A Text book of

DAIRY CHEMISTRY AND TECHNOLOGY

First Edition
2063

By
Pushpa Prasad Acharya
Lecturer
Central Campus of Technology

Institute of Science and Technology

Central Campus of Technology
Dahran, Hattisar
A Textbook of DAIRY CHEMISTRY AND TECHNOLOGY
Pushpa Prasad Acharya

First Edition 2063/2006

© Author

Rs.

Institute of Science and Technology

Central Campus of Technology
Dharan-14, Hattisar
 PREFACE

Milk has been the subject of scientific study for about 150 years. In context of our country
still we have not separate courses on Dairy Science and Technology. Though it has
included in the courses of Food Technology since twenty seven years. This book is written
on the basis of courses included in Bachelors level. Even though, I hope this book will be
useful to master level student also. Till now we have lack of consolidated book on Dairy
Technology. Without this students are facing many problems.
This book is intended to fill the gap and should be as useful to undergraduate and post
graduate level and who are working in the dairy industry. This book assumes knowledge of
chemistry of the principle milk constituents of milk, i.e. water, lactose, lipids, proteins
(including enzymes), salts and vitamins. As well as this book also includes technology of
different milk products i.e. market(liquid) milk, cream products, ice cream, fermented milks,
cheeses, butter, ghee, powder milk, concentrated milk, condensed milk. In spite of this
book also included basic operation of milk processing, hygienic milk handling, quality
assurance of milk, milk microbiology, testing of milk and milk products etc.
I hope that the book will answer some of your questions on the chemistry and technology of
milk and milk products and encourage to the reader to under take more extensive study of
these topics.
Lastly, I humbly request to the reader for their healthy, creative and constructive
suggestions from which it could make further more attractive and informative.

Pushpa P Acharya
Contents
1 History of dairy industry in Nepal 1-10
1.1 Introduction 1
1.2 Dairy development commission 1
1.3 Establishment of the Central Dairy at Lainchour 2
1.4 Kharipati Milk collection centre 2
1.5 Dairy Development Board 2
1.6 Foreign Assistance 3
1.7 HRD in Nepalese dairy development 3
1.8 The period of 1960-64 4
1.9 The Period of 1964-69 4
1.9.1 New approach in milk collection and marketing 4
1.9.2 Expansion of the central dairy 4
1.9.3 Handover to Nepalese technicians 4
1.9.4 Dairy Development Corporation 5
1.9.5 Biratnagar milk plant 5
1.10 Entry of DANIDA in the dairy sector 6
1.11 Hetauda milk plant 6
1.12 Pokhara milk supply scheme 6
1.13 The period of 1975-85 6
1.13.1 Milk supply scheme 6
1.13.2 Cheese and butter factories 6
1.13.3 Growth of the private sector dairies 7
1.13.4 Cooperative in dairy sector 7
1.13.5 Nepal cheese producers cooperative society 7
1.14 Formation of Dairy Development Board 8
1.15 Status of market milk industries of Nepal 8
1.16 Milk and milk distribution system 10
2 Biosynthesis and secretion of milk 13-20
2.1 The mammary gland 13
2.2 The physiology of milk secretion 14
2.2.1 Cell cytology and milk secretion 14
2.2.2 Mitochondria 15
2.2.3 Lysosomes 15
2.3 Secretion of milk (Milk ejection) 17
3 General aspects of milk 21-28
3.1 Milk definition 21
3.2 Milk composition 21
3.3 General introduction to proximate composition of milk 21
3.3.1 Water 22
3.3.2 Milk proteins 22
3.3.2.1 Definitions 22
3.3.2.2 Casein 22
3.3.2.3 General composition of milk 25
3.3.3 Lactose 25
3.3.4 Milk fat 27
3.3.5 Gases present in milk 28
3.3.6 Enzymes in milk 28
4 Physicochemical properties of milk 29-34
5 Milk components 35-84
 5.1 Water 35
5.2 Carbohydrates 35
5.3 Milk lipids 48
5.4 Milk Proteins 57
5.5 Enzymes in milk 64
5.6 Salts in milk 72
5.7 Other components 78
5.8 Flavor components 83
5.9 Pigments 84
6 Colloidal particles of milk 85-108
6.1 Fat globules 85
6.1.1 Properties of fat globules 85
6.1.2 Emulsion stability 87
6.1.3 Interaction with air bubbles 92
6.1.4 Creaming 93
6.1.5 Lipolysis 96
6.2 Casein micelles 97
6.2.1 Description 97
6.2.2 Changes 99
6.2.3 Colloidal stability 100
6.3 Physical properties 103
6.3.1 Optical properties 103
6.3.2 Viscosity 104
6.4 Whey proteins 105
6.4.1 Definition 105
6.4.2 Classification of whey 105
6.4.3 Proximate composition 105
6.4.4 Composition of whey 105
6.4.5 Whey proteins 106
7 Microbiology of milk 109-122
7.1 Introduction 109
7.2 Milk as a substrate for bacteria 113
7.3 Undesirable bacteria 114
7.3.1 Pathogenic microorganisms 114
7.3.2 Spoilage microorganisms 115
7.4 Pathogenic microorganisms in milk 117
7.5 Sources of contaminations 118
7.6 Hygienic measures 121
7.7 Protection of the consumers against pathogenic microorganisms 121
7.8 Measures against spoilage microorganisms 122
8 Milk processing 123-181
8.1 Introduction 123
8.2 Objectives 124
8.3 Quality assurance 125
8.3.1 Concepts 125
8.3.2 HACCP 127
8.4 Quality assurance of raw milk 132
8.5 Milk storage and transport 133
8.6 Preservatives 135
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 8.7 Collection and reception of milk 136
8.8 Milk chilling and storage 137
8.9 Transport of milk to the dairy 140
8.10 Filtration and clarification 141
8.10.1 Types 142
8.11 Membrane filtration process 144
8.12 Standardizing 146
8.13 Heat treatment 150
8.13.1 Changes caused by heating 151
8.13.2 Heating intensity 153
8.14 Inactivation of enzymes 155
8.15 Method of heating 157
8.16 Cream separation 165
8.17 Bactofugation 170
8.18 Homogenization 172
8.18.1 Objectives of homogenization 172
8.18.2 Operation of the homogenizer 173
8.18.3 Factors affecting homogenization 174
8.18.4 Theory of homogenization 175
8.18.5 Factors affecting the fat globule size 177
8.18.6 Effect of homogenization 178
8.18.7 Homogenization clusters 179
8.18.8 Other effect of homogenization 180
8.19 Creaming 181
9 Milk for liquid consumption 184-195
9.1 Pasteurized milk 182
9.1.1 Manufacture 182
9.1.2 Shelf life 186
9.1.3 Use of microfiltration 188
9.2 Sterilized milk 189
9.2.1 Description 189
9.2.2 Method of manufacture 190
9.2.3 Shelf life 192
9.2.4 Flavor 193
9.2.5 Nutritive value 195
10 Cream products 196-202
10.1 Sterilized milk 196
10.1.1 Manufacture 297
10.1.2 Heat stability 298
10.1.3 Stability in coffee 298
10.1.4 Clustering 298
10.2 Whipping Cream 299
10.2.1 Desirable properties 299
10.2.2 Manufacture 200
10.2.3 The whipping process 201
10.2.4 Variables 201
11 Ice cream production 203-217
11.1 Introduction 203
11.2 Classification of ice cream 203
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 11.3 Chemical composition 204
11.4 Specification of ice cream 204
11.5 Role of various components in ice cream 204
11.6 Manufacture 205
11.7 Physical structure: Formation and stability 209
11.8 Over-run 212
11.9 Hardening of ice cream 213
11.10 Ice cream defects 213
11.11 Sofety ice cream 214
12 Concentrated milk 215-227
12.1 Evaporated milk 215
12.1.1 Description 215
12.1.2 Manufacture 215
12.1.3 Organoleptic properties 219
12.1.4 Heat stability 219
12.1.5 Creaming 220
12.1.6 Age thickening and gelation 220
12.2 Sweetened condensed milk 222
12.2.1 Description 222
12.2.2 Manufacture 223
12.2.3 Homogenization 223
12.2.4 Cooling and seeding 224
12.2.5 Microbial spoilage 225
12.2.6 Chemical deterioration 229
12.3 Lactose crystals 227
13 Milk powder 228-246
13.1 Objectives 228
13.2 Composition 229
13.3 Manufacture 229
13.4 Hygienic aspects 232
13.5 Physical properties 236
13.6 Ease of dispersing (instant powder) 236
13.7 Influence of process variables on product properties 237
13.8 Deterioration 240
13.9 Other types of milk products 243
13.10 Reconstituted products 243
14 Butter production 244-261
14.1 Description 244
14.2 Definition 245
14.3 Description of types 245
14.4 Composition of butter 246
14.5 Manufacturing schemes 246
14.6 The churning process 248
14.7 Structure and properties 251
14.8 Cold storage defects 254
14.9 Culture butter from sweet cream 255
14.10 High fat products 256
14.10.1 Anhydrous milk fat 257
14.10.2 Modification of milk fat 258
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 14.10.3 Recombined butter 259
14.10.4 Butter products with a laow fat contents 261
15 Ghee manufacturing 262-269
15.1 Chemical composition 262
15.2 Physico-chemical constants 262
15.3 Food and nutritive value 263
15.4 Method of manufacture 263
15.4.1 Cooling and granulation 266
15.4.2 Packaging and storage of ghee 266
15.4.3 Renovation of ghee 268
15.4.4 Neutralization of high acid ghee 268
15.4.5 AGMARK 268
15.4.6 Grading of ghee 268
15.4.7 Physiochemical quality of ghee 269
15.4.8 Nutritive value 269
16 Fermented milk 270-284
16.1 General aspects 270
16.1.1 Preservation 270
16.1.2 Nutritive value 271
16.1.3 Composition 271
16.1.4 Nutritive aspects 272
16.2 Various types 274
16.2.1 Types of fermentation 274
16.2.2 Fat content 275
16.2.3 Concentration of milk 276
16.2.4 Withdrawal of whey 276
16.2.5 Milk of various animal species 276
16.3 Yoghurt 278
16.3.1 Introduction 278
16.3.2 Yoghurt bacteria 279
16.3.3 Manufacture of set and stirred yoghurt 280
16.3.4 Physical properties 281
16.3.5 Flavor defects: Shelf life 282
16.3.6 Dahi or curd 282
17 Cheese production 285-321
17.1 Definition 285
17.2 Brief history of cheese 285
17.3 Basic principles of cheese making 286
17.4 Retention of constituent of milk 286
17.5 Classification of cheese 288
17.6 Chemical composition of different varieties of cheese 289
17.7 Nepalese cheeses 290
17.7.1 Yak cheese 291
17.7.2 Kanchan cheese 293
17.7.3 Mozzarella cheese 294
17.7.4 Danish mozzarella 295
17.7.5 Soft mozzarella 295
17.8 Manufacturing process of hard and soft varieties of cheese 296
17.8.1 Cheese milk 296
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 17.8.2 Additives to cheese milk 300
17.8.3 Curd treatment 308
17.8.4 Moulding 312
17.8.5 Salting 312
17.8.6 Ripening of cheese 313
17.8.7 Cleaning and packaging 314
17.8.8 Grading of cheese 316
17.8.9 Cheese faults and their removal 317
17.8.10 Process and spread cheese 320
18 Dairy plant sanitation and hygiene 322-335
18.1 Introduction 322
18.2 Cleaning 322
18.2.1 Importance of cleaning and sanitizing 323
18.3 Detergents 323
18.3.1 Classification of Dairy detergents 324
18.3.2 Sanitizers 324
18.4 Cleaning and sanitization procedure 325
18.5 Selection of detergents and sanitizers 326
18.6 Method of cleaning 327
18.6.1 Hand washing 327
18.6.2 Mechanical washing 328
18.6.3 Cleaning-in-place (CIP) 329
19 Testing of milk and milk products 336-356
19.1 Platform testing of milk and milk products 336
19.2 Chemical and enzyme test 343
19.3 Freezing point test of milk 347
19.4 Microbiological tests 348
19.5 Detection of adulterants in milk 348
19.6 Determination of preservatives 352
19.7 Determination of neutralizers 354
20 Organization in dairy 356
20.1 Organizational chart of Kathmandu Milk supply scheme 356
20.1 Organizational chart of Biratnagar Milk supply scheme 356

vi
List of figures

S.N Contents Page

2.1 Diagramatic sketch of the precursors for milk synthesis of the ruminant mamals 17
2.2 Diagramatic sketch of the duct system of one quarter of the bovine udder 19
3.1 Chart shows the general composition of milk 25
5.1 Chemical structure of lactose and lactulose 38
5.2 Mutarotation of lactose solution 41
5.3 The different forms of lactose 42
5.4 Common shapes of the α- lactose hydrate crystals 44
5.5 Solubilities of α- lactose and β-lactose hydrate crystal. 45
5.6 Relative concentration of antioxidants, hydroperoxides and free carbonyls during
53
autoxidation of a fat
5.7 Melting curve of milk fat 54
5.8 Examples of the proportion of fat being solid after 24 h cooling to temperature T 55
5.10 Time(t) and temperature(T) of heating milk needed to inactivate some enzymes 72
6.1 Types of physical changes in oil-in-water emulsion 87
6.2 Adsorption of aggregation (black dots) onto milk fat globules and the ensuring
94
flocculation of the globules
6.3 Acidity of milk fat of milk susceptible to lipolysis 97
6.4 A casein micelle 98
6.5 Aggregation of casein micelles 103
6.6 Preparation of lactoglobumin 107
6.7 Flow chart for the preparation of different whey products 108
7.1 Growth curve of a bacterial culture 110
7.2 Change of the colony count during the keeping of milk of two initial counts, at two
111
temperatures
7.3 Time needed to reach a count of 106 bacteria per ml when keeping raw milk at
112
various temperatures.
8.1 Shows the some possible hazards and CCPs applied in the pasteurized milk
132
production chains
8.2 The principle of Ultrafiltration 145
8.3 Shows the RO membrane, where salt are separated 146
8.4 Diagram of the continuous standardization process 148
8.5 Simplified diagram of a heat exchanger for heating and cooling of liquids, showing
159
principles of regeneration
8.6 Shows the different section of Plate heat exchanger (a continuous pasteurizer) 161
8.7 The flow diagram of a heat exchanger in which the milk is heated and cooled by
162
water
8.8 Temperatures of milk versus time during heat treatment(direct &indirect) 162
8.9 Diagram of continuous bottle sterilizer 164
8.10 Detail of cross section of a modern homogenizing valve with a cone type 174
8.11 Diagram showing the principle of the two stage homogenizer 175
8.12 Showing deformation of the fat globule in the homogenizer slit 175
8.13 Diagrammatic representation of the homogenization process 176
8.14 New surface layer of fat globules 178
9.1 Flow diagram for the manufacture if standardized, homogenized, pasteurized milk 183
9.2 A manufacturing process for pasteurized beverage milk by using microfiltration 188
9.3 Manufacture of in-bottle-sterilized milk 190
9.4 The manufacture of UHT-sterilized milk(indirect &direct heat) with aseptic packaging 191
vii
10.1 The manucature of coffee cream (top) and dessert cream(bottom) 197
10.2 Manufacturing flow diagram of whipping cream 200
11.1 Flow diagram for the manufacture of ice-cream 212
11.2 Freezing of the ice cream mix 211
11.3 Shows the structure of the ice cream at about -5oC 211
11.4 The extent of clumping of the fat globules on the shape retention of ice cream 212
12.1 Manufacture of in-bottle (left) & UHT(right) sterilized whole milk 217
12.2 Manufacture of recombined evaporated milk 218
12.3 The changes obserbed in the casein micelles of evaporated milk during storage 221
12.4 The manufacture of sweetened condensed milk (SCM) 223
13.1 Flow chart for the manufacture of whole milk powder 229
14.1 Flow diagram for the manufacture of butter from (sour) cream 246
14.2 Effect of temperature and time of storage on the firmness of butter 253
14.3 The manufacture of recombined butter (or margarine) 260
15.1 Flow diagram for the manufacture of ghee by creamery method 263
15.2 Flow diagram for the manufacture of ghee by creamery butter method 264
15.3 Simplified flow diagram for the manufacture of butter oil 265
15.4 Shows the pre-stratification method using in ghee manufacture 266
16.1 Traditional production of yoghurt 280
16.2 Flow diagram for the production of improved method of set and stirred yoghurt 280
16.3 Flow diagram for the production of dahi 284
17.1 Interrelationship between different classes of cheeses 289
17.2 Flow diagram for yak cheese manufacture 292
17.3 Flow diagram for kanchan cheese manufacture from cow milk 293
17.4 Manufacture of Nepalese mozzarella 294
17.5 Flow diagram for normal mozzarella manufacture 295
17.6 Flow diagram of soft mozzarella manufacture 295
17.7 Different products formed during ripening of cheese 313
17.8 Flow diagram for Nepalese processed cheese 320
17.9 Flow diagram for the preparation of processed cheese 321
18.1 Flow chart for general cleaning and sanitizing procedure 325
18.2 Flow chart for hand washing procedure 327
18.3 Mechanical can and bottle washing procedure 328
18.4 Mechanical bottle washing procedure 328
18.5 Elements of a Cleaning-In-Place (CIP) system 334

viii
List of Tables

1.1 List of milk processing industries under DDC 9

1.2 Existing market milk processing industries owned by private sectors 9
1.3 Factory reception cost price 10
1.4 Price paid according to chilling centre 11
2.1 Composition of blood plasma and milk 20
2.2 Shows the precursors in blood responsible for the constituents of milk 20
3.1 General composition of milk 21
3.2 Differences of milk composition due to species 21
5.1 Example of the rate of growth of some faces of an α-lactose hydrate crystals as
44
affected by liquid composition
5.2 Fatty acid profile of milk fats of different species (mole %)
5.3 Some enzymes in milk 64
5.4 The most important salts in milk and their distribution between serum and casein
73
micelles
5.5 Shows the vitamins content in fresh milk 79
5.6 Most important radionucleides that can occur in milk 81
6.1 Essential amino acids in cow milk products (g/100g) 108
7.1 Generation time(h) of some groups of bacteria in milk 111
7.2 Effect of the keeping temperature of milk on its count after 24 h, and on its keeping
111
quality
8.1 Examples of standards for (pooled) milk before processing 139
8.2 Loss of % of some nutrients i.e., available lysine and various vitamins, due to some
153
heat treatment in milk
8.3 Heat inactivation of some milk enzymes in milk 156
11.1 Approximate chemical composition of ice cream and its specification 204
13.1 The approximate composition of come types of powder 229
15.1 Difference between different methods of ghee making 265
17.1 Composition of different types of cheeses 290
17.2 Some chemical composition of Nepalese cheese varieties 291
17.3 Proximate composition of milk from milch animals of Nepal 291
19.1 Correction to be applied to lactometer readings st temperature other than 27oC to
342
obtain corrected lactometer reading of milk at 27oC

ix
Chapter 1

History of dairy industry development in Nepal

1.1 Introduction
History of dairy development showed that, the organized market milk industry was started
at Langtang by producing the experimental cheese during the year 1953-56. A cheese store
was also built at Kyangjin Ghyang in 1955. At this place a cheese factory also built during
the period 1956-60. Those establishments still exist in their original form with only some
repair and maintenance.
The period 1958-60 was spent in development of infrastructure for establishment of another
factory at Thodung with cheese store, staff quarter, office rooms, and several mobile
cheese units to cover the up and down movement of chauri herds, during the cheese
manufacturing season usually from March/April to November/December.
Mr. Warner Schulthess, a foreign expert, played a pioneering role by starting the
pasteurization of milk at Tusal village near Banepa. The success of the cheese production
and pasteurization convinced the Department of Agriculture to generate reimbursement
funds from New Zealand grant in 1955, which Mr. Schulthess had taken to finance to Tusal
dairy.
1.2 Dairy development commission
In the development agenda of Nepal, the dairy industry received a very high priority.
Government of Nepal requested and received the financial grant assistance from
government of New Zealand under the Colombo plan in 1953-54 for the development of
dairy industry. A high powered commission was made to facilitate dairy industry
development. In the year 1954 the late king Mahendra was inaugurated first milk collection
and processing unit at Nala Tusal showed the importance given to the dairy sector by the
Government.
The beginning of milk supply scheme for Kathmandu was established during the period of
1953-55. The success of first milk collection, processing and marketing of pasteurized milk
was received very enthusiastically by the producers, consumers and the government
authority. Regular and enhanced income attracted the consumers and producers. Both of
these factors encouraged the government to embark upon an ambitious plan for dairy
development.
1.3 Establishment of the central dairy at Lainchour
In the first decade of democratic Nepal, dairy industry development had placed in first
priority. It was evidenced by laying down the foundation stone of Central Dairy at Lainchour
by late His Majesty King Mahendra in Februrary/March 1956 (where the Dairy Development
Corporation has its head office now). In the same year (june 1956) two candidates were
selected and sent to NDRI, Banglore, India for higher studies. The need of human resource
development (HRD) in dairy sector was realized and from 1956 onward 2-3 candidates
were selected every year and sent for studies in Dairy science and Technology that
continues for many years. Central dairy at Lainchour was planned very modestly, a 500 L
per hour high temperature short time (HTST) pasteurizer, cream separator to standardized
milk at 3.0 % fat and 8.0 % solid-not-fat, a manual bottle filling and capping machine, a
manual bottle and can washing machines, and wire milk crates were the main equipments
for milk processing. One lb (half litre) and half lb (¼ litre) bottles were used for milk
distribution. For milk distribution tri-cycle was introduced in Kathmandu to door to door in
the morning and in the evening.
Surplus cream separated was purchase by International hotel named as Hotel Royal
managed by legendary Mr. Boris. Hotel Royal was single largest customer of milk, cream,
butter, cheese. Bir hospital was another recognized large customer of milk products. The
rest were small buyers. The product become so popular, in course of time that people
(particularly students, teachers and professors) used to drink milk straight from the bottle at
the central dairy shop located within the premises of the Central dairy at Lainchour. The
milk was safe and healthy.
1.4 Kharipati milk collection centre
In order to meet the demand of milk and milk products, another potential milk pocket was
located at Kharipati, north-east of Bhaktapur. It was developed during 1956-58, on the
same lines as at Tusal. However, by this time the quality of milk became doubtful as the
farmers had learnt to take the advantage of the faulty quality control and payment system.
1.5 Dairy Development Board
The dairy industry had taken a firm root during a period 1950-60. In 1954-55 dairy
development section was created, dairy development commission was dissolved and a
dairy development board (DDB) was constituted to oversee and facilitate dairy development

2
in Nepal under the Chairmanship of minister of agriculture. The DDB was responsible to
coordinate between the related agencies including HMG/N, New Zealand Government,
FAO and SATA (Swiss Association of Technical Assistance).
1.6 Foreign Assistance
In the dairy sector Switzerland mobilized SATA for technical assistance. The entry of
Switzerland, FAO and New Zealand in Nepal was through dairy sector development. FAO
provided the technical leadership, New Zealand provided financial grant and SATA
provided technical assistance in terms of required qualified manpower.
1.7 HRD in Nepalese dairy development
The first candidate Mr. Gauri Prasad Sharma had received degree in dairy science and
technology in 1940, sponsored by government in 1938. After returning back, he headed the
livestock and dairy development section within the department of agriculture. However, in
1954-55 an independent dairy development section was created under his leadership. This
significantly contributed for the establishment and development of dairy industry in Nepal.
After that for a long time (about 18 years) no any manpower was trained in dairy
technology.
But after the beginning of the organized dairy sector development with financial grant
provided by the New Zealand government and technical support from FAO and SATA, it
was started to train manpower in this sector since 1956. It was continued through nineteen-
seventies up to which time more than 20 persons received dairy education from diploma to
post-graduate and doctoral level. But, after 1980 very little attention has been paid for the
development of manpower in the dairy sector. After 1990, apart from short training, no
formal education has been provided in this field.
In the early sixties, the requirement of specialized manpower in special field of dairy
development was so great that many more people were invited from SATA to fill the gap. In
this way by 1960 a big contingent of Swiss technicians were working in this sector.
However, very recently the NDDB/DSP had identified the need for HRD in the Dairy sector,
including the need of establishing a national dairy training facility in the country as well as
efforts are underway to train and educate the numbers of people at every level.

3
1.8 The period of 1960-64
The successful introduction of organized cheese industry and the milk supply in Kathmandu
created very positive environment for consolidation of the dairy industry. The cheese factory
at Thodung was completed in 1962 and production of cheese was started. It was also used
as training centre for Nepalese technicians in milk reception, quality control and cheese
production. In the same year, near Jiri a cheese store, which was latter developed into a
full-fledged cheese factory, was completed. In 1963, another cheese factory was also
initiated at Semila Pike in Solukhumbu district. A cheese factory was also developed at
Chordhum north-east of Jiri. This factory came in production during 1966-67.
1.9 The period of 1964-69
1.9.1 New approach in milk collection and marketing
The first two milk collecting centers established in Tusal and Kharipati were in fact complete
processing plants. The new approach in milk collection was to establish small milk
collecting centers with minimum facilities for receiving, sampling, testing, weighing and
pooling milk in 40 L Al-alloy milk cans. Milk was collected in the morning as well as in the
evening immediately dispatched both in the morning and evening to a centrally located milk
chilling center easily accessible to 5 to 7 collecting centers. First the chilling center was
established in Sallaghari, Bhaktapur with a capacity of 3000 L. It was a ice bank type
chilling vat. Milk was chilled and transported in insulated milk tankers to Central Dairy
Lainchour.
1.9.2 Expansion of the central dairy
The Lainchour milk plant was expanded with adding automatic bottle washing, filling, and
sealing machines. The pasteurized standardized milk was sold in milk bottles in Kathmandu
and Lalitpur. Cream, curd, butter, ice cream, cheese, processed cheese were marketed
with good success.
1.9.3 Handover to Nepalese technicians
During this period number of dairy technicians having graduated in dairy science and
technology slowly took over the industry in their own hands. The dairy expert from FAO had
left Nepal in 1964 (after 12 years of involvement in dairy development). All Swiss
technicians and experts also went back. Dairy industry was fully managed and supervised
by Nepalese experts.

4
1.9.4 Dairy development corporation
Dairy development board and the Dairy development section were dissolved and a Dairy
development corporation was constituted under the Corporation’s act of 2021 B.S.(1964
AD). HMG/N announced the constitution of DDC through Nepal Gazette dated 2026, Kartic
18th (November 1969). DDC started functioning from 1st of shrawn 2026 B.S. (16th July
1969) with its head office at Lainchour. DDC was responsible both for development and
expansion of dairy industry and at the same time to operate commercially with the principle
of no profit no loss. It is quite successful in this regard. The DDC begin milk processing
plant at Lainchour had a capacity of 1,000 L/h. Three Yak cheese factories in the alpine belt
under a separate scheme called Cheese Production and Supply Scheme (CPSS). The
other cheese factories were also established at Gosainkunda (Chandanbari) Rasuwa
district in 1970; Kyamawalding (North of Thodung) in Ramechhap district in 1972; at
Kyangsing in Sindhupalchouk district in 1974 and at Taksindhu in Solokhumbu district in
1975.
A temperature and humidity controlled cheese store with the capacity of 30,000 kg of
cheese was constructed within the premises of Dairy at Lainchour.Several milk collection
center were added DDC in different centers of Kathmandu town.
1.9.5 Biratnagar milk plant
During a visit of late His Majesty King Mahendra in the town of Utrecht, the Netherlands,
citizens of the town made a gift of a dairy plant for the children of Nepal, The people had
donated certain funds, which was supplemented by the Government of Netherlands. This
fund was used by DDC to build a milk plant with 2000 L/h capacity at Kanchanbari,
Biratnagar. The mayor of town Utrecht laid the foundation stone. The plant came into
operation in June 1973. This plant was named as Biratnagar Milk supply Scheme (BMSS).
FAO had organized an Inter Governmental meeting at its headquarters in Rome in 1972 to
foster an International Scheme of coodination for Dairy Development (ISCDD). Nepal had
presented its country paper on dairy development and requested for assistance from the
member countries attending that meeting. A generous offer was made by Danish
International Development Agency (DANIDA), Government of New Zealand and WFP.
Following the commitment made at the ISCCD meeting at Rome, the Government of New
Zealand provided grant assistance to build a modern full-fledged dairy plant at alaju with

5
5,000 L/h capacity with provisions for production of by-products. The foundation atone of
this plant was laid by H.E. the Ambassador of New Zealand to Nepal in May, 1973. The
milk plant is presently known as Kathmandu Milk Supply Scheme (KMSS).
1.10 Entry of DANIDA in the dairy sector
On June 26,1973, an agreement was signed between HMG/N and Royal Danish
Government for a Danish assistance of DKr. 13 million for the development of dairy industry
in Nepal. These funds were used for the establishment of milk plants and chilling centers.
1.11 Hetauda milk plant
Establishment of a milk processing plant at Hetauda with a capacity of 3,000 L/h with a
provisions to make milk products including yoghurt and ice cream was accomplished out of
the above Danida assistance. It came into operation in 1975. This plant is called Hetauda
Milk Supply Scheme(HMSS).
1.12 Pokhara Milk Supply Scheme
Out of the above DANIDA fund, a milk plant was also established in Pokhara with a modest
capacity of 2,000 L/h with facilities for manufacturing other dairy products such as, cream,
butter, yoghurt and ice cream. This plant was named as Pokhara Milk Supply Scheme
(PMSS) and came into operation in 1980. Now the plant has been privatized.
1.13 The period of 1975-85
1.13.1 Milk supply scheme
Milk schemes had been established and started milk collection, processing and marketing
of milk and milk products in Kathmandu, Biratnagar, Hetauda and Pokhara. The collection
of raw milk was however, much less than the total market demand. Heavy reconstitution
was done from imported skim milk powder (SMP). Share of local milk varied from a
maximum of 56 % to a minimum of 36 % averaging about 46 %.
1.13.2 Cheese and butter factories
Yak/Chauri cheese production activities were expanded to new pockets. Buffalo cheese
production also initiated at Dhulikhel, at Pauwa in Kavre district (1975), latter in Nagarkot,
Kalinchok 1877 and Deurali 1978, all in Kavre district but these are closed now. These
factory was used for paneer production. Two yak cheese factories were established at
Gatlang 1977 in Rasuwa district and at Chinchu 1981 in Dolakha district. A milk collection
and chilling center was also established at Trisuli with assistance from Rasuwa Nuwakot

6
Integrated Rural Development Project funded by the World bank. The beginning of the
production of cow milk cheese called Kanchan cheese at Pashupatinagar, Ilam was started
in 1985. In this period many activities concentrated towards reforms and structural
adjustment in the field of livestock and dairy development.
1.13.3 Growth of the private sector dairies
In the latter part of 1970s with a very small scale operations private sector started dairy
processing in Kaathmandu. The sector grew slowly during 1980s. However, after the
adoption of economic liberalization policy by the Government after the restoration of
democracy in 1990, the private sector grew rapidly. New dairies were emerged and the
existing ones expanded their capacities. Presently the private sector has been able to
capture the market share of about 46 % in pasteurized milk sale, which were merely about
2 percent in 1981. The growth of milk sale of the private sector is and has exceeded the
amount being produced by DDC. The available information shows that presently 131 milk
processing plants of varying capacities (mostly in the central region), 21 yak cheese
production centers and 5 cow cheese production centers are operating in the private sector.
The private dairies have not only increased in number but they have also been successful
in showing their capabilities of diversifying their products. Himalaya dairy has diversified its
product line to UHT drinking yoghurt in three flavors and is soon producing UHT milk.
1.13.4 Cooperative in dairy sector
After the enforcement of the cooperative act 1992, Milk Producers Co-operative Unions
(MPCU) were at secondary level organized in order to guide the MPC’s on their managerial,
promotional and business affairs. On April 2, 1993 a Central milk procedures cooperative
union ltd. (CMPCU) at national level was voluntarily and jointly organized by the MPC’s and
MPCU’s and registered under the Act.
1.13.4.1 Nepal cheese producers cooperative society
FAO/NDDB organized a workshop on cheese production in Nepal held from March 6-10,
2000 in Pokhara which recommended developing common cooperative cheese storage and
handling and marketing facilities of cheese to help solve the participants to immediately
bring this recommendation into action. Hence, the cheese producers from mid hills and high
Himalayan region producing Kanchan and Yak cheese respectively formed Nepal cheese

7
producers cooperative society ltd. (NCPCS) in 2055 (1998-99). It was registered under the
cooperative act of 1992 with head office at Kathmandu.
1.14 Formation of dairy development board (DDB)
An workshop cum-seminar on Dairy development management in Nepal jointly organized
by FAO and DDC was held from 8th to 11th in Katmandu. Among other recommendations,
this workshop recommended formation of Dairy development board (DDB) to facilitate and
promote the development and management of dairy industry of Nepal. It was formed in
September 1989 under the chairmanship of the minister of agriculture. Members
represented the Board from private dairies, MPA’s, the Nepal peasant organization, the
banking sector, the development of livestock services (DSL) and Minister of agriculture,
Finance and industries. The secretariat of the Board was kept at DDC central office and the
General Manager of DDC was appointed as its appointed as its member secretary.
1.15 Status of market milk industries of Nepal
Nepal is an agricultural country. More than 80 per cent of the total population of Nepal is
based on agriculture. At present Livestock contributes to about 31 per cent of the gross
domestic production (GDP), whereas agriculture contributes to about 60 per cent of the
same.
Dairying is an integral part of Nepalese way of life. It has contributed to alleviate the
poverty of Nepalese to a great extent. Rearing of cattle and buffaloes contribute about 77 %
of the total live stock sector. Of the total live stock about 6 % is contributed by milk and milk
products. Of these 70 % of the milk is produced by buffaloes and 30 % by cows, in this
country. Annual growth rate of milk production has been 1-2 %.
Average per capita annual milk consumption amounts to about 45 liters. Annual milk
production is about 1 028 000 tons, which is above 6 % of GDP at current market price.
Dairy Development has multiplier effect in the growth of other sector too. Thus the growth of
Dairy sector has a vital bearing on the overall development of the country. Around 1 00 000
dairy farmers’ households produce and sell milk to the organized sector, and about 10 000
people are engaged in milk processing industry including rural enterprises, ancillary
workers, porters, venders, booth men, etc.
Dairy Development Corporation is the pioneer and major sector in the Nepalese dairy
industry with four market milk plant, one skimmed milk powder plant, one product plant and

8
small scale plants. The government is putting emphasis on free market economy, thus the
private sector participation is also increase. Number of small, medium and large sized
modern dairy plants are owned and managed by the private sector people.
Table 1.1 List of milk processing industries under Dairy Development Corporation.
MPA/ Prodn Sales Distribution
MPCS Chilling center booth
Name District covered centers
capacity
(L/shift)
Kathmandu Katmandu, Bhaktapur,
milk supply Lalitpur, Sindupalchok,
475 14 75,000 - 750
scheme Kavre,Gorkha, Dhading,
Nuwakot and Chitwan
Biratnagar Morang,
milk supply Saptari,Sunsari, 25,000 +
116 8 2 87
scheme Jhapa,Ilam,Dhankuta 35,000
and Terhathum.
Hetauda milk Makawanpur, Bara,
supply Rautahat, Sarlahi and 187 8 15000 2 76
scheme mahottari
Pokhara Gorkha, Tanahu, Kaski,
milk supply Sangiya, Palpa, Parbat 100 6 10,000 2 135
scheme and Baglung
Lumbani Palpa, Nawalparasi,
milk supply Dang, Rupandehi,
90 7 - 1 -
scheme Kapilbastu, Banke,
Bardiya and Surkhet.
Milk pdts Ilam, Panchthar, Kavre,
pro-duction Ramechhap, Dolakha,
25 - - 2 -
and Solukhumbu and
distribution Rasuwa
Central Lamahi, Kohalpur,
Collected the milk from different areas and
Western Chinchu
supply to Pokhara and Kathmandu through
milk supply
National Milk Grid.
scheme
Table 1.2 Existing market milk processing industries owned by private sectors.
Name of some private dairy Daily milk collection Daily milk sales
industries (l/day) (l/day)
1. Sandesh dairy 100 100
2. Kishan dairy 200 200
3. Swait dairy 80 80
4. Nepal dairy 3000 3000
5. Kathmandu dairy 3000 3000
6. Bhaktapur dairy 1900 1900
7. Sainju dairy 1200 2000
8. Narayani dairy 1200 1500
9
9. Modern dairy 3000 3000
10. Pushpa dairy 1000 1000
11. Laxmi dairy 200 200
12. Sitaram Gokul Dairy Pvt. Ltd. 18000 ?
13. Baneswor milk bar 400 400
14. Koshi dairy 80 80
15. Quality dairy 300 300
16. Shree Chamunda dairy 300 300
17. Bhagawati dairy 150 150
18. Anmol dairy 4500 5500
19. Shree Ganesh Dairy 80 80
20. Everest dairy 200 200
21. Ugrachandi dairy 500 500
22. Silwal dairy 600 600
23. Cottage dairy 1500 1500
24. Pushpa dairy 160 160
25. Himalayan dairy(Today) 17000 32000
26. Panjee dairy (pokhara) 500 500
27. Krishna Dairy(Pokhara) 150 150
28. Nilayam Himal Cheese 400 Cheese
29. Yeti Hard Cheese, Ilam 1400 Cheese
30. Gurudev dairy, Banke 600 600
31. Godar dairy, Sinduli 100 100
32. Lila dairy, Dang 250 250
33. Nobel dairy Pvt. Ltd.
34. Ram Janaki Dairy (sunsari)
35.Kailash Kamdhenu Dairy (sunsari)
36. Dharan Dairy 500 500

1.16 Milk and milk distribution system

Dairy development corporation have altogether 1828 sales booth and 8 sales center,
from which the milk and milk products are sales to the consumers. The DDC sales their
products through the city areas of Kathmandu, Lalitpur, Bhaktapur, Biratnagar, Dharan,
Narayanghat, Hetauda, Birgung, Pokhara, Butwal, Bhairahawa etc.
The private dairy industries also concentrated in city areas and nearby and distributed
their products to the consumers of that areas.
Milk procurement and sales price
Implemented from 2057-04-05
Table 1.3 Factory reception cost price
Milk supply Fat per Solid not Total solid
schemes kg fat per kg per kg
10
 Kathmandu, Balaju 160 115 16
Hetauda 146 104 13
Biratnagar 152 103 12
Pokhara 148 106 16.50
Butwal 143 104 12.50

Table 1.4 Price paid according to chilling center

S. Particulars Fat/ Kg SNF/ Kg TS/Kg*
N
Kathmandu milk supply scheme(KMSS)
Bhaktapur, Sankhu, Banepa,
1 157 113 15
Panauti, Dhungre paini
Tika Bhairab* ,Belefi*, Panchkhal**,
2 153 111 18* 15**
Banepa**, Mahadev benshi**
3 Sifaghat*, Chauradi** 152 109 17* 16**
4 Bhaktapur, Jayamire 147 105 12
5 Chanauli( Gunjanagar) 147 105 11.50
Hetauda milk supply scheme (HMSS)
1 In industry 146 109 13
2 Chaughoda, Bhimfedi 141 104 11.50
3 Chandrani gohapur, Garuda 137 104 13
4 Nawalpur, Barathawa, Malangawa,
135 104 11.50
Gausala
Biratnagar Milk supply scheme (BMSS)
1 In industry 152 103 12
2 Inaruwa, Kanchanpur, Budhabare,
147 103 10
Salakpur, Surunga
3 Phikal, Tinghare, Bahundagi,
147 98 10
Kutidada
4 Biblayate, Puakhola 142 98 12
5 Hile,Chitre 142 98 12
Pokhara milk supply scheme(PMSS)
1 In industry 148 106 16.50
2 Damauli, Galayang, Dumbre 142 103 14
Lumbini milk supply scheme(LMSS)
1 Manigram, Butwal 143 104 12.25
2 Chaunata, Sunwal 141 102 12.65
Mid -Western milk supply scheme
1 Lamahi 139 102 13.85
2 Kohalpur 138 101 11.85
3 Chinchu 138 101 11.75
Milk product production, sales and distribution scheme
1 Pashupatinagar Cheese Factory,
147 98 10
Ilam
2 Nagarkot Cheese factory 157 113 15

11
3 Rakshe, Ranke 142 98 12
4 Jaubari 141 97 10
Note: * TS/Kg means TS commission per Kg.
Minimum standard fixed by DDC for procurement
Cow milk - fat 3.5 % and SNF 8.2 %
Buffalo milk- fat 5.1 % and SNF 8.4 %
Sales price of standard pasteurized milk and whole milk in all sales booth of different
milk supply schemes.
Standard pasteurized milk Rs. 22/= per litre and Whole milk Rs. 26/= per litre.

12
Chapter 2

Biosynthesis of milk and its secretion

2.1 The Mammary Gland

Mammary gland is made up of secretary and connective tissue. Secretary units are
small grape like objects called "alveoli". A group of alveoli are joined together in a cluster
with a common duct, so that they resemble grapes attached to a stem.
Each lobule has a small duct that leads into larger ducts and all of the alveoli in a cluster
(lobule) are drained by a common duct. Each lobule has between (150 to 220) alveoli and
measures roughly 0.75 mm3.
Each lobule is surrounded by a connective tissue capsule. A number of lobules together
form a lobe, which is drained by a larger duct and surrounded by a connective tissue
capsule. The udder is made up of lobes that are attached to the duct system.
The basic component of an alveolus is a single layer of epithelial cells surrounding a
central cavity, the lumen. Each alveolus is supplied with small capillaries and venuoles and
drain away unused blood. The capillaries and venuoles connect with the larger arteries and
veins in the udder. Each alveolus are also surrounded with a series of specialized cells (
the myoepithelial cells) that are responsible for milk ejection.
Milk is a unique product secreted by the mammary gland under complex hormonal
control. Milk is synthesized in distinctive secretory cells, which comprise the secretory
epithelium. There is a sequential changes in the hormonal environment, so that these cells
proliferate and develop during pregnancy to be brought into full synthetic activity near the
time of parturition.The major hormones are estrogen, progesterone , and growth hormone,
basically involved in the development of the mammary gland. Insulin is a hormone
promotes cell division of the mammary epithelium, corticosteriods are involved for the
developments of the cellular organelles for the synthesis of milk constituents, and prolactin
is essential for the initiation and maintenance of lactation. Each secretory cell secretes
complete milk. Most of the constituents of the milk are synthesized in these cells, and
others are transported through them.
The mammary secretory cells, which are arranged around alveoli. In general, precursors
enter the secretory cells from the blood circulation on the basal side, and milk is secreted
from the apical side into the lumen of the alveolus.Myoepithelial cells surround the alveolus.
The hormone oxytocin stimulates contraction of these cells, thus squeezing milk out of the
alveoli into the ducts. The alveoli are interconnected by an intricate network of the ducts
that lead to the milk cistern. A considerable portion of the milk secreted between milking
accumulates in the cistern.
The biosynthetic and physiological processes involved in the milk secretion have been
elucidated by studies of differences in concentrations of certain constituents between
arterial blood supplying the mammary gland and venous blood leaving it, by electron
microscopy of mammary tissue at various stages, and by enzyme assays and radioactive
tracer studies of biosynthetic processes in intact animals, cultures of mammary cells, and
cell fractions. The biosynthetic processes occur at various sites in the cell. Intermediate
compounds required for synthesis of protein, lactose, and fat are produced in cytosol and
mitochondria; protein and lipid are synthesized in the endoplasmic reticulum; lactose
synthesis, post-translational modification of proteins, and assembly of casein micelles occur
in the Golgi apparatus. The mitochondria play an important role for to transfer the energy
essential for synthesis processes. Citrate is synthesized in the mitochondria and can diffuse
through their membranes into the cytosol. The mitochondria also supply carbon for the
synthesis of nonessential amino acids.
2.2 The Physiology of milk secretion
2.2.1 Cell cytology and milk secretion
The major function of the mammary gland is the secretion of milk, which takes place in
the epithelial cells of the alveolus. The cells have relatively large nuclei with one or more
nucleoli. The nucleus has two membranes, an outer one that is continuous with the
endoplasmic reticulum and the golgi apparatus and an inner one that has pores for the
movement of material from the nucleus to the cytoplasm. Chromatin material in the nucleus
contains in formation for cell duplication and the synthesis of cell milk protein including the
production of enzymes required for synthesis of other milk constituents.Each of the
secretory cells of lactating animal has an abundant network of endoplasmic reticulum and
an enlarged golgi apparatus. There are numerous protein droplets in the golgibodies and
the lumina of the alveoli, and a large number of fat droplets exist in the cell apices and the
lumina. A marked increase in the number of mitochondria occurs preparatory to secretion.

14
2.2.2 Mitochondria
It is located throughout the cell. It is responsible for breaking down various molecules
especially glucose and acetate. It captures the released energy in the form of ATP. The
energy is then utilized in the synthetic reactions that are necessary to the secretion of milk.
The endoplasmic reticulum and the golgi apparatus are continuous with the cell
membrane and the lumen of the alveolus. Part of the endoplasmic reticulum is lined with
dense particles known as ribosomes which are attached to the outer surface of the
membrane and are responsible for most of the protein synthesis of the cell. Some
ribosomes are not attached to the endoplasmic reticulum but are scattered throughout the
cytoplasm.
Protein synthesis
Three types of RNA molecules are involved in the protein synthesis. All of these are formed
in the nucleus and are transported through the cytoplasm to the sites in which protein
synthesis occurs. The protein molecules moves through the lumen of the endoplasmic
reticulum to the smooth endoplasmic reticulum and then to the golgi apparatus, in which the
milk proteins are condensed into vacuoles to be exposed to the alveolar lumina.
2.2.2 Lysosomes
The lysosomes in the cell contain enzymes for the breakdown of larger molecules into
smaller molecules. The lysosomes also contain degradative enzymes from being released
into the cell. Which prevents degradative enzymes from being released into the cell. The
lysosomes probably play a major role in the involution of mammary gland.
Milk fat secretion
Almost all milk fat is made up of triglycerides. The fatty acids have different carbon
chain length. Ruminant milk fat contains higher proportion of short chain fatty acids than
does milk fat from non-ruminants and also contains a smaller amount of unsaturated fatty
acids (oleic & linoleic acids).
The fatty acids that are esterified to glycerol arise from two major sources.Most of the
short chain fatty acids up through the C14 acids are synthesized within the mammary
gland. They are built up from acetate units, which contain 2 carbon atoms and beta-
hydroxybutyric acid molecules, which contain 4 carbon atoms.These short carbon units

15
come from the rumen fermentation process. Much more acetate than beta-hydroxy butyric
acid is present in the milk fat synthesis.
Almost all of the C18 fatty acids and many of the C16 fatty acids, come from the fatty
acids that are absorbed as such from the blood stream. The mammary gland can remove
H2 ions from the C18 fatty acid to unsaturated them. The mammary gland absorbs smaller,
amounts of oleic acids and linoleic acids from the blood stream than are found in the ; most
of the glycerol molecules are synthesized from glucose, but some of them comes from the
glycerol portion of the triglycerides that are absorbed from the blood secretion.
Lactose Secretion
Lactose formation apparently occurs in the golgi-apparatus. α – lactalbumin is synthesized
at the ribosomes and moves through the lumen of endoplasmic reticulum to golgi-apparatus
where the second subunit (galactosyl transferase) of lactose synthetase is present.
Because lactose synthetase is formed in the golgi-apparatus. It is believed that lactose
formation takes place these and the lactose is exported with the protein particles through
the vacuoles.
The lactose content of milk is relatively constant, lactose is the major component
responsible for the osmotic pressure of milk, but chloride, K and Na ions also play a role in
this respect. Water is transferred to the lumen of the alveolus until the osmotic pressure of
the milk is similar to that of the blood. Most of the water passes through the epithelial cells
into the milk by filtration.
Other Secretion processes
The minerals and vitamins content of milk’s governed by the filtration process. However
epithelial cells act as a membrane barrier or carrier of the particles from the blood to the
lumen of the alveolus. The epithelial cells combine some of the minerals with organic
compounds; for example 75 % of the calcium in milk is in chemical or physical combination
with casein, phosphate and citrate, & more than one-half of the phosphorus in milk is
combined with casein. Vitamin molecules in milk are transferred unchanged from the blood.
Concentrations of some vitamins, especially the fat soluble vitamins, can be increased by
increasing the vitamin content of the blood plasma.
The filtration process account for the secretion of water, vitamins and minerals and the true
cell metabolism process accounts for the production of fat, lactose and most of the protein.

16
 Fatty
acids Intestine Body fat
Liver
Rumen C2,C3,
C4 Glucose

Fatty acids Amino Fatty

Minerals Vitamins Glucose
C18, C6 acids acids
Water C18
Blood

Energy

Energy
Proteins Lactose
Fat
Milk

Fig.2.1 Diagrammatic sketch of the precursors for milk synthesis of the ruminant mammal.
2.3 Secretion of milk (Milk ejection)
The dairy cow’s has the best developed mammary glands of any animal. A good cow will
produce milk in one year about ten times its own weight, and exceptional cows may
produce 25 times of their weight. The cow’s udder consists of 4 distinctly separate glands,
known as the quarters. Each quarter is provided with a teat, the hollow, interior portion of
which is known as the teat cistern. The teat cistern extends upwards into the body of the
gland and is connected with the gland cistern, which may vary in capacity from less than ½
to 1 liter. Radiating from the walls of the cistern are numerous tubes or ducts which branch
out or divide innumerable times. The ducts are very small in that portion of the quarter
removed from the cistern, but become larger as they approach and enter the gland cistern.
Some secretion of milk occurs in the lining of the ducts, but the principal secretion takes
place in the alveoli, the enlargements at the very end of each of the smallest branches of
the duct system. Milk is secreted more or less continuously between milking periods. It may
be slowed or stopped completely by the pressure of the milk accumulated in the alveoli.
Frequent milking decreases this pressure and favors milk production. At the milking time
the cow ejects or lets down her milk. This is a reflex or involuntary action, controlled by

17
sensory nerves which carry the message to the pituitary body in the brain. This message
may be originated by the suckling calf or by the washing and manipulating of the udder by
the milker. The pituitary gland then furnishes to the blood stream a hormone known as
prolactin, which is an important factor in the growth of the udder and milk production.
Another hormone, oxytocin, causes the alveoli to contract and thereby squeeze the milk
from them into the ducts through which it travels to the cistern and teats.
Conditions that frighten or anger the cow at milking time interfere with the function of the
pituitary body so that its hormones are not released into the blood stream and the result is
that she does not yield her milk. Oxytocin gradually is destroyed in the blood stream, and
therefore the milking operation must be completed before its influence is lost, which usually
is within ten minutes.
Most of the milk obtained at a single milking is present in the udder at the time of milking.
About half of this milk is stored between milking in the milk cistern and ducts, while the
balance is stored by stretching the udder. As the udder is stretched, the pressure within it
increased, then restricting its blood supply and so reducing the rate of secretion.
Milk is a secretion and in its composition differs greatly from that of the blood from which it
is derived. For e.g. milk fat, casein and lactose, which are synthesized in the udder, are not
found elsewhere in the body. Very large amounts of blood must be passed through the
udder in the production of milk; it is estimated that a cow producing six gallons of milk will
pass the equivalent of over ten times of blood through the udder.
A number of substances that are not normal constituents of milk may be eliminated from
the body by the milk. In this category are many drugs and certain flavor producing
substances may be present, such as onions, turnips and silage that the cow may consume.
Of recent interest is the elimination in the milk of radioactive fall-out material such as
Strontium-90 and pesticide chemical, such as DDT, which might be present in the cow
feed.
A usual occurrence, possibly associated with the transfer through milk of an injurious
substance, was reported from Tasmania School Children there showed an increase
incidence of goiter, even though their diet was supplemented with iodine. This was
attributed to the use of milk from cows on pasture that contained narrow-stem kale. This
plant, with others such as cabbage and turnips, contains an antithyroid or goiter producing

18
substance, capable of being transmitted through the milk. Different workers regarding the
secretion of milk have proposed numerous theories. But it is now believed that most of the
milk is formed within the milk gland and the udder contains all the milk, which is yielded in
the milking.
The process of milk formation (Fig.2.1, above) in the udder can be divided into two steps:
a. The supply of the precursors of milk to the udder by the blood stream.
b. The conversion of the raw material into the constituents of milk by the gland cells.
The blood enters the mammary gland through the mammary artery, which branches into
numerous small arteries, and capillaries, which run throughout the tissues of the mammary
glands and supply

Alveoli

Large milk ducts

Gland cistern Myoepithelial cell

Annular fold
Milk
Teat cistern
cavity
Sphincter muscle

Streak canal

Alveolus
Fig.2.2 Diagrammatic sketch of the duct system of one quarter of the bovine udder. Each
quarter is composed of many lobes that contain lobules. The lobules contain alveoli,
the secretary tissue. Only one lobe is shown.
nutrients to each secreting cell, i.e., alveoli. After supplying the necessary nutrient to
alveoli, the blood returns to the heart through the milk vein. If the size of the milk vein is
good, this is a good indication of good yielding animal. For every liter of milk secreted,
about 400 litres of blood are required to circulate through the udder.
Thus, blood is carrier of nutrients and waste product of the body. Maynard gave the
following analysis of the blood plasma and milk, which shows similarly between the two:

19
Table 2.1 Composition of blood plasma and milk.
Blood plasma Milk
Constituents % Constituents %
Water 91.0 Water 87.0
Glucose 0.05 Lactose 4.8
Serum albumin 3.20 Lactalbumin 0.52
Serum globulin 4.40 Globulin 0.05
Amino acids 0.003 Casein 2.9
Neutral fats 0.09 Neutral fats 3.8
Phospholipids 0.20 Phospholipids 0.04
Cholesterol ester 0.17 Cholesterol ester Traces
Calcium 0.009 Calcium 0.12
Phosphorus 0.011 Phosphorus 0.10
Sodium 0.34 Sodium 0.15
Potassium 0.03 Potassium 0.15
Chlorine 0.35 Chlorine 0.11
Citric acid Trace Citric acid 0.20

Table 2.2 Shows the precursors in blood responsible for the constituents of milk.

Constituents of milk Precursor in Blood

Water Water
Lipids
Fat Fatty acids (from food fat or from
Cholesterol the catabolism of proteins and
Ergosterol carbohydrates).
Lecithin
Cephalin
Pigments Per se from blood.
Proteins
Casein Globulin, amino acid and other
Albumin non-protein nitrogenous
Globulin compounds.
Non Protein nitrogen
creatinine From the filtration products from
urea the blood and metabolic products
Ammonia etc. from glandular activity.
Carbohydrates Glucose, lactic acid.
Lactose
Inorganic constituents Salts (selective filtration and
Ca, K, Na, Mg, Mn, PO4, molecular rearrangement).
Cl, SO4, CO3 etc
Vitamins Per se from blood.
Enzymes
Dissolved gases Not definite.
O2 Metabolic products from glandular
N2 activity, diffusion through tissues.
CO2

20
Chapter 3
General aspects of milk

3.1 Milk definition

Milk is an important article of food for mankind and young ones of all mammals. It has a complex
nature and possesses extraordinary properties.
Different workers have defined it in a number of ways. The important definitions are:
Milk has been defined as the entire product of the complete and uninterrupted milking of milch
cows, which are properly cared for and are in good health.
Federal definition of U.S.A. is," Milk is the fresh, clean lacteal secretion obtained by the
complete milking of one or more healthy cows, properly fed and kept, excluding that obtained
within 15 days before and 10 days after calving and containing not less than 8.5 percent solids-not-
fat and not less than 3.25 percent milk fat."
It has also been defined as the, “Physiological secretion from the mammary gland of mammals”.
Since the dairy cow is most commonly and extensively used source of milk, the term milk, denotes
to mean milk of the dairy cows unless otherwise specified.
India’s legal definition of milk reads, “Milk is the secretion derived from the complete milking of
healthy milch animals. It shall be free from colostrums”.
3.2 Composition of milk
Table 3.1 General composition of milk.
Constituents %
Water 87.54
Fat 3.71
Casein 2.63
Whey protein 0.42 Protein 3.18
Protease-peptone 0.13
Other nitrogeneous substances 0.11
Lactose 4.70
Ash 0.76
Total 100.0

Table 3.2 Differences in milk composition due to species

Species Water (%) Fat(%) Sugar(%) Protein (%) Ash (%)
Cow 87.54 3.71 4.70 3.31 0.76
Goat 85.58 4.93 4.78 4.11 0.89
Buffalo 82.90 7.50 4.70 4.10 0.80
Human 88.50 3.30 6.80 1.30 0.20
3.3 General introduction to proximate composition of milk
3.3.1 Water
It is found in milk in three forms. Such as free, bound and crystallized water. Water act
as a suspending, dispersing and dissolving media for the components of milk. It occupies
about 87 % in fresh milk.
3.3.2 Milk Proteins
3.3.2.1 Definition
Proteins are high molecular weight polymers, generally over 10000 MW,of amino acids
covalently linked by the peptide bond. Most of the proteins of milk contain more than 150
amino acids. The properties of the various proteins are dictated by the aminoacids in the
molecule and by their sequence in the polypeptide chain(s), which regulates the molecular
configuration of the protein molecule and the surface electrical charge. Proteins can exist in
helical coils, random coils, pleated sheets or in a combination of these forms. These
properties relate to the stability of the proteins in the food system and influence processing
of the products. Milk proteins generally possess several of these forms in a single protein
molecule.
Protein is one of the most essential nutrients of milk present in about 3.5 %. Milk protein
contains almost all of the essential amino acids and hence high nutritive value. Carbon,
Hydrogen, Nitrogen, Oxygen, Sulphur, and phosphorus are the elements present in protein.
In milk among the total protein, casein contributed 2.9 % and whey protein 0.6 %.Milk
protein may be divided into two main groups: Casein and whey proteins (lactalbumin,
lactoglobulin).
3.3.2.2 Casein
Their relatively high phosphorus content characterizes the casein. Casein is a generic
term for a class of proteins that are synthesized in the mammary gland and make up about
80-85 % of the total milk protein content. It is present in the form of micelles or particles of
macromolecular sizes.
Casein is composed of four recognized components called α-s,β,κ,γ on the basis of
differences in their electrical charge. Chemically the casein system is defined as a
glycophospho-protein, since it contains both carbohydrate (glycol group) and phosphorus
as integral parts of the protein. In milk, casein exists as its calcium salts, namely, calcium

22
caseinate, in distinct globular particles, ranging from 40-300 mµ in diameter. These
particles are called micelles; each micelle contains all the component casein held together
in part by calcium phosphate.
Commercial casein is obtained from fat-free skim milk by precipitation either by addition
of an acid, or by the addition of rennet extract (containing the enzyme rennin).
Caseins are high molecular weight (89 000) compounds. They are the charged. They
are the charged particles having IEP 4.6. The elementary composition of casein indicates
carbon (53%), hydrogen (7.07%), sulfur (0.76%) and phosphorus (0.85%). The factor used
for determination of protein is 6.38. This is because N2 content in milk is taken as 15.65
instead of normal value (16 %). Milk and milk products provide food proteins of excellent
quality for the nutrition of man and animals. Casein, the dominant protein of milk, is a good
source of amino acids, which are the indispensable for human nutrition.
The milk protein differs considerably from each other in amino acid composition however
they all contain considerable amount of dietary essential amino acid. The α-lactalbumin
contains very high amount of tryptophan, while all milk proteins are rich in aspartic acid and
glutamic acid. The essential amino acids such as tryptophan, threonine, isoleucine, leucine,
lysine, methionine, cystine, phenylalanine, tyrosine and valine, all are found in milk.
The peptide chain obtained after enzymatic hydrolysis found to have phosphorus bound
to the peptide. Hence, they are also called phosphopeptides or phosphopeptone.
The casein in normal milk accounts for close to 80 % of the nitrogen present. Casein is
present in milk in solution and as micelles containing calcium, inorganic phosphate,
magnesium, and citrate. The micelles are in colloidal suspension.
The diameters of micelles are between 80 nm & 250 nm in buffalo and between 50 nm
200 nm in cow casein. Casein is precipitated with or without those mineral components
depending upon the coagulating agent used. The casein of normal fresh milk only partially
precipitates on prolong heating at high temperatures. Good quality milk may be preheated
to render it more stable during condensing or drying, although there might be some
reduction of the ability or power to reconstitute if made from such milk. Preheating helps to
prevent age thickening. Adding small concentrations of sodium & potassium chloride
increases the heat stability of milk. Browning may result from the reaction of the amino
acids and carbohydrates to prolong heating at temperatures in excess of 100oC. Heat
23
aggravates the affect of acid in milk. Casein readily precipitates with acid or rennet. Acid
frees the casein by reacting with the calcium with which this protein is associated in milk.
The casein is then no longer able to remain in suspension, so it separates out. Another
means of coagulating casein is by the action of proteolytic enzymes such as rennin. Curd
strength is measured in grams of force required to cause a standard knife to pass through
the surface of a standard curd. This value may range from 30 to 80 g for fresh normal milk
from individual cows. Boiling reduces the curd tension nearly 30 %. The curd tention of raw
buffalo milk is reported to be 44.54g. This value is reduced to about 32.5 % by
pasteurization by the holding method & nearly 73 % by homogenization. Colostrum and
mastitis milk is unstable to heat.
Globulin & albumin are soluble proteins of milk. They are called whey proteins because
they do not usually precipitate with acid & only partially precipitate with rennet.High
temperatures and strong salt solutions will precipitates the whey proteins, which account for
about 12 % of the nitrogen of milk.
Casein also precipitated by alcohol is calcium caseinate. The casein precipitated with
weak acids is free of calcium. When casein is precipitated with rennin, paracasein is
formed. It contains more calcium than Ca-caseinate. Pure casein is precipitated by heat,
but in fresh milk prolong heating at high temperature (100oC) for 12 Hrs or more Hrs or
(120oC) under pressure will cause the precipitation of casein. On boiling fresh milk, a thin
layer of finely precipitated casein, together with other milk constituents including fat, forms a
thin layer over the surface of the milk. The application of heat to milk that is slightly acid will
cause the precipitation of casein.
One to two molecule of serine combines with one phosphoric acid molecule a
diphosphoric ester is formed. Therefore phosphoric acid may form a cross link between
polypeptide chain. Casein has acidic nature, as compared to other proteins, is quite
distinctive as it has considerable base binding properties and can even liberate CO2 from
carbonates. As lactic acid develops, calcium from colloidal solution of protein moves to
soluble condition and this process continuous till the milk curdling. The calcium remove
from calcium hydrogen carbonate to form calcium lactate. The protein is precipitated (or
progressible removal of calcium starts) only after pH 4.6. The concentrated and dry form of
milk, when left for a long time gradually develops a brown color. The browning in milk is

24
caused by a maillard type of reaction between amino group of milk protein and aldehyde
group of lactose. Such products are unfit for human consumption.
Casein is the macromolecule in the form of micelles and each micelle composed of
about 1000 casein molecules. Casein is a mixture of different fractions and is
heterogeneous in nature. The different fractions are differing in composition, solubility and
rennet coagulation.
It has been found that in electric field casein separates into three separate components
moving at different speeds. Those components are α, β and γ casein which are arranged in
descending order of mobility. The α and β casein are phosphoprotein of high quality where
γ-casein contain very little phosphorus. On the basis of electrophoretic analysis, whole
casein contains about 75 % of α-casein, 22 % of β-casein and 3 % of γ-casein. Alpha
casein is responsible for the stabilization of the micelle in the milk and it is the component
where readily attack. It is composed of two sub-fractions of α-s casein (or calcium sensitive
casein) and κ-casein (or calcium insensitive casein).
3.2.2.3 General composition of milk

Milk

Fat Water Non fatty substances

Associated Lactose Nitrogenous Mineral Other

Total fat matter constituents
substances substances

Protein Non-protein
Several
glycerides Casein Albumin Phosphates, citrates,
Globulin
chlorides of K, Na,
Lecithin Cholesterol Carotene Vitamin Ca, Mg. Traces of
ADEK Fe, Cu, I etc

Enzyme, Vitamin B1,B2, C Dissolved

Bacteria gases
Fig.3.1 Chart shows the general composition of milk.

The average composition of milk are as follows

3.3.3 Lactose
It is sugar found in mammal milk. In cow's milk it is found about 4.7 %, whereas in human
milk it contains about 6.3 %. Lactose is present in milk as true solution. When it crystallizes
from water, it forms hard gritty crystals, which have one molecule of water of crystallization
25
(C12H22O11.H2O). These gritty crystals sometimes appear in certain milk products under
certain circumstances under which these crystals are allowed to grow in size. These
crystals appear in ice cream when the mix contains a high proportion of milk solids. Lactose
is faintly sweet, about 1/6th the sweetness of cane sugar. That is why milk is only faintly
sweet. Whey appears to be sweeter due to removal of casein, which masks the flavor of
lactose.
Lactose belong to disaccharide group of sugar and is made by the combination of one
molecule of glucose and a molecule of galactose with elimination of one molecule of water
in such a manner that only one reactive aldehyde group is left in lactose. Thus, the
hydrolysis of lactose by the enzyme lactase found in intestines splits lactose molecule into
the component hexose sugar, e.g., glucose and galactose.
lactase enzyme
C12H22O11 + H2O C6H12O6 + C6H12O6
Lactose Galactose Glucose
The mineral acids also have the hydrolysing capacity.The fermented milk contains about
0.7 % alcohol , CO2 and lactic acid. Casein of such fermented milk is very digestible.
Lactose solution in water has reducing properties and reduces fehling solution and
ammonical silver nitrate. The reducing power of lactose is doubled by hydrolysis as both
the sugars formed as a result of hydrolysis have one active aldehyde group each. Lactose
is insoluble in alcohol and ether but is soluble in hot acetic acid. Oxidation of lactose in
slightly acid condition yields formic acid and finally laevulinic acid, but oxidation by
concentrated nitric acid breaks lactose into oxalic acid and carbonic acid. When lactose is
heated between 110-1300C, the lactose hydrate crystals lose the water
of crystallization and above 150oC they turn yellow and at at 175oC they turn brown & form
caramel. The slight burning and characteristic colour of cooked milk is due to the formation
of caramel, though the milk and milk products are never treated to such high temperatures
under ordinary processes. Lactose, when gently warmed with nitrogeneous bases like
ammonia, certain amines, amino acid, any of which may be present in heated milk, forms
brown coloured complexes. Thus, the extent of these changes will depend upon the
severity of the heat treatment and the time of holding. Pasteurized milk is only slightly
effected, while sterilized milk or evaporated milk have more effect.

26
Lactose is a valuable product and is used for the preparation of infant,weaned food
products & invalids. This is the main source of galactose, present in brain & nerve tissues
of human body. Lactose helps in assimilation of calcium and phosphate from intestines has
a tonning effect on the alimentary canal. Whey is the main sourse for its manufacture.
3.3.4 Milk fat
If a drop of milk is observed under microscope, the presence of fat fat globules is revealed.
The size of these fat globules vary widely.The size of the fat globules increases with
advancing lactation and also depend upon the breed.Usually the breeds giving more
percentage of fat, have higher proportion of smaller globules.
If cool raw milk is kept for some time without much mixing, the fat globules cluster round
each other in many hundreds of individuals.This tendency of clustering is important in the
separation cream either by gravitational or centrifugal means.In cream, where the
percentage of fat is higher than in milk, the fat globules are tightely/ closely packed, but
neither in milk nor in cream, globules form a continuous phase, though they may be in
actual contact with their immediate neighbours.Thus, each individual globule is dependent
due to the presence of a protctive layer around them.
About the protective layer, the earlier theories explain that the fat globule is enclosed in an
enveloping membrane. It is known that the surface area of the fat globules is very large,
due to the small size of the fat globules. Such large surface areas associated with small
particles exibit considerable surface forces which is responsible for the phenomenon of
adsorption. In milk, casein and fat are present in colloidal state and hence they are
responsible for adsorption and concentration of certain milk constituents which form a very
thin layer which prevent the merging of the fat globules and the formation of a continuous
fat phase.
In a protective layer, a variety of substances are found such as protein, phospholipid,
phosphatase, an unidentified ether soluble substances, and a complex of riboflavin-
phosphoric acid-protein. When metals like Cupper come in contact with the protective layer
and modifies with the protective layer, they stimulate oxidative changes in the fat phase.
The dissolved metal forms metallic proteinates either by combining with protein of the layer
or protein of the plasma and then the metallic proteinates are absorbed by the globules.

27
Thus, the water soluble metal is brought in contact with the fat globules. In fact, the
adsorbed layer is the center of vigorous chemical action.
The melting and setting points of fat varies from 28-330C and 24-290C, respectively. It
means fat is a mixture of various kinds of fat having different properties.
The physical properties of the milk fat is determined by the nature and the proportion of its
constituents. It is a mixture of true fats which dissolve or absorb a number of substances,
including lecithin, cholesterol, carotene and the fat soluble vitamins : The vitamins present
in milk are :
Fat soluble A,D,E,K and water soluble B-complex (B1 such as aneurin or thiamine, B2 such
as riboflavin, nicotinic acid, pyrodoxine, pantothenic acid, biotin, vitamin B12, folic acid) and
vitamin C (Ascorbic acid). Milk is a good source of vitamin A and B2 as far as human
nutrition is concerned.
3.3.5 Gases present in milk
When milk is drawn from the udder of the cow, it contains dissolved gases to about 8 % by
volume amongest which carbon dioxide predominates.But when milk is exposed to
atmosphere, the volume decreases to about 6 percent. This change in volume is not merely
quantitative but is qualitative as well. Carbon dioxide is lost and oxygen and nitrogen are
gained. Dissolved oxygen is responsible for oxidation of fat and ascorbic acid. It can be
reduced by a combination of heat and vacuum treatment.
3.3.6 Enzymes in milk
These are organic catalysts bring many complex reactions both anabolic and catabolic. It do not
enter into chemical change. They are colloidal, proteineous and are classified according to the
chemical change which they bring about, such as hydrolases (hydrolysing enzymes), oxidases
(oxidising enzymes), reductases (reducing enzymes). They are also classified on the basis of the
substrate which they act upon as protease (protein splitting enzymes); lipase (fat splitting enzyme),
amylases (starch splitting enzymes), etc.
Enzymes are susceptible to heat, light and pH changes. Substances accelerating the activity of
enzymes are known as co-enzymes and those inhibit their activity are known as antienzymes.
A number of enzymes occur in milk as it is drawn from udder itself, while some gain entrance as a
result of bacterial contamination.
Peroxidase, catalase, reductase, phosphatase, lipase are present in fresh milk, while other
enzymes enter as a result of bacterial contamination.
28
Chapter 4

Physico- chemical properties of milk

4.1 Physical state of milk

In milk water is the continuous phase in which other constituents are either dissolved or
suspended. Lactose and a portion of the mineral salts are found in solution, proteins and
the remainders of the minerals are in colloidal suspension and fat as emulsion.
4.2 Acidity and pH of milk
4.2.1 Acidity
Freshly drawn milk is amphoteric to litmus that is it turns red litmus blue and blue litmus
red. However, it shows acidity during titration and is due to presence of casein, acid
phosphates, citrates etc, which is known as natural or apparent acidity. Titratable acidity of
cows’ milk (0.13 to 0.14%) and buffalo milk (0.14 to 0.15 %). During storage, lactic acid
formed as the result of bacterial action on lactose and hence titratable acidity of stored milk
is equal to some of the natural acidity and developed acidity. Generally, acidity is
expressed as lactic acid.
C12H22O11 + H2O C6H12O6 + C6H12O6
Lactose glucose galactose
2C6H12O6 4CH3CHOH.COOH (Lactic acid)
4.2.2 pH
The pH of normal, fresh, sweet milk usually varies from 6.4 to 6.6 for cow milk and 6.7 to
6.8 for buffalo milk. A higher pH value for fresh milk indicates udder infection (mastitis) and
lower values due to bacterial action.
4.3 Density and specific gravity
Milk is heavier than water. The average specific gravity ranges (at 600F) from 1.028 to
1.030 for cow milk, 1.030 to 1.032 for buffalo milk and 1.035 to1.037 for skim milk. The
proportion of its constituents greatly influenced the specific gravity of milk. The specific
gravity of water 1.00, fat 0.93, protein 1.346, lactose 1.666 and salts 4.12 (SNF
1.616).Although, buffalo milk contains more fat than cow milk, its specific gravity is higher
due to excess of SNF.
 The specific gravity of milk is lowered by addition of water and cream and increased by
addition of skim milk or removal of fat. Specific gravity can be determined by lactometer
and is equal to:
Lactometer reading (at 600 F )
Specific gravity = 1 +
1000
The percent total solid and solid-not-fat in the milk can be calculated by the following
formula.
% T .S = 0.25 D + 1.22 F + 0.72
% SNF = 0.25 D + 0.22 F + 0.72
Where, D = 1000(d − 1)
d = density of milk at 20 0 C.
F = Fat % of the milk sample.

Recknagel phenomenon
The specific gravity of milk increases gradually after milking up to certain time. During
milking O2 or other gas mixed and this phenomenon proceed until the gas comes out. This
process continuous at 15oC for 1 to 2 days. He found this phenomenon during 1883. So
this phenomenon is called Recknagel’s Phenomenon. The causes of increase in specific
gravity are due to casein hydration and slow solidification of fat.
4.4 Freezing point of milk
Milk freezes at temperatures slightly lower than water due to the presence of soluble
constituents such as lactose, soluble salts etc. The freezing point of water is 0oC but due to
the solutes, cow milk becomes lower (- 0.555oC), and buffalo milk -0.560oC and addition of
water will raise the value (below -0.53oC). The instrument measuring the freezing point is
Cryoscope. By the help of this instrument, it is easier to determine the adulteration. The
freezing point test of milk is a highly sensitive one and even up to 3 % of watering can be
detected. Boiling and sterilization increase the value of freezing point depression, but
pasteurization has no effect on it. The fat and protein content of milk have no direct effect
on the freezing point of milk. The major factors affecting the freezing point of milk are:
lactose, soluble salts.
4.5 Boiling point
Milk is heavier than water. Milk boils at some higher temperature due to the reason that the
boiling point depends on the specific gravity. The water boils at 100oC at sea level but milk
30
boils at 100.17oC. When water is added to milk, the boiling point also lowers due to
decrease in the concentration of solutes.
4.6 Specific heat
Specific heat is the ratio of heat necessary to raise the temperature of some substances
and the heat to raise the temperature up to some extent of water having same weight. The
unit is ‘calorie’. The specific heat of milk in average is 0.9457, whereas pure water is 1.00 at
STP. It depends on the physical and chemical condition. Milk boils faster or temperature
rises faster than water but the boiling point of milk is higher than water. It is because milk
dissolves more soluble solids but water do not dissolves it.
4.7 Color of milk
Milk seems as white nontransparent color. The white color is due to “ calcium-casienate” in
the form of colloidal. Generally the color is a blend of the individual effects produced by
• the colloidal casein particles and the dispersed fat globules, both of which scatter light,
and,
• the carotene ( to some extent xanthophylls) which imparts the yellowish tint.
Milk ranges in color from yellowish creamery white (cow milk) to creamery white( buffalo
milk). The greater the intakes of green feed, the deeper yellow the color of cow milk. The
larger the fat globules and higher the fat percentage, the greater the intensity of the yellow
color. Skim milk has a bluish and when a greenish yellow color. The greenish of whey is
due to riboflavin (Lactoflavin).
The color of foods is an important aspect of their marketability. Color has 3 aspects that is
tint, intensity and uniformity.
4.8 Flavor
This is composed of smell (odor) and taste. The flavor of milk is a blend of the sweet taste
of lactose and salty taste of minerals, both of which are damped down by proteins. The
phospholipids, fatty acids and fats of milk also contribute the flavor.
The sulfydryl compounds significantly contribute to the cooked flavor of heated milk and
milk products. Sometimes the cowy flavor appears. In late lactation milk and milk of mastitis
animal, the increase amount of chloride cause salty flavor. Late lactation milk has been
found bitterness due to the action of lipase enzyme splitting the fat to fatty acids, so

31
bitterness causes. Microorganisms may enter and cause sourness due to lactic acids.
Similarly various chemical changes causes oxidized, metallic or fishy odor.
4.9 Viscosity / Plasticity
It is measured by Ostwald Viscometer. It is the measurements of resistant flowing the
liquid. Water flows simply than milk. A 20oC water has viscosity 1.005 cP but milk has 1.5 to
2.0 cP, skim milk 1.5, whey 1.007, Whole milk 2.0 cP. The value decreases both with an
increase of temperature and with the removal of fat. Pasteurization does not appreciably
alter the viscosity of milk. Both sterilization and boiling reportedly increase milk viscosity.
Plasticity refers to a high degree of viscosity. Once a quantity of plastic or semi-solid
material is formed into a particular shape, it will tend to retain that shape. For such
materials, outside force is required to cause the material to flow most liquids flow by the
force of their own mass.
Pump design, the application of external force in printing butter, and the mechanical power
requirements of a processing operation are affected by the viscosity of the products.
4.10 Surface tension
When a droplet of liquid is free flowing in a gas or in another liquid with which it doesn’t mix,
the droplet tends to pull itself into a spherical shape, or into that shape which has the
smallest surface area per unit of volume. The force pulling the droplet into this shape is the
surface tension of the liquid. The surface tension of milk is about 45 to 50 dynes/cm but
water has 72.5 dynes/cm. It is mainly affected by fat, temperature and ageing also.
Homogenization increases the surface tension. Du Nuoy Tensinometer is used for its
measurement.
4.11 Action of milk on metals
Milk acts on certain metals, so that a small amount of the metal is dissolved in it. The
metallic salts thus formed may give rise to a metallic taste in the milk. Some salts acts as
catalysts, thus hastening the oxidation of fat and producing an oxidized flavor. These
metals are said to taint milks.
The factors which influence the degree of action by milk in the metal are:
1. Temperature of the milk.
2. Period of contact.
3. Cleanliness and polish of metals.

32
 4. Amount of free air in milk, and
5. Acidity of the milk.
The metals used for the milk contact surfaces must, as far as possible, meet the
following requirements.
1. Non toxic,
2. Non tainting,
3. Insoluble in milk and milk products,
4. Highly resistant to corrosion,
5. Easy to clean and keep bright
6. Light and strong,
7. Good agents of heat transfer
8. Good in appearance throughout life,
9. Low in cost.
10. Non absorbent.
11. Durable.
No single metal or alloy meets all these requirements. However, 18:8 stainless steel alloy
are the most satisfactory at present. Corrosion can’t be entirely prevented in dairy
equipments but its rate can be controlled to a large extent. To prevent corrosion of
stainless steel surface the following measures should be taken.
1. The surface should be clean.
2. Surface air dry, whenever possible.
3. Cleaners and sanitizers use in the lowest concentration and for shortest duration.
An invisible film of chromium oxide forms on stainless steel surface when it is dry and
exposed to the atmosphere. This film protects it from being corrosion.
Chloride and its compounds are very corrosive. Equipments should be sanitized with
chloride solutions. Preferably just before it is to be used, so as to avoid prolonged contact
and thus corrosion (pitting).
4.12 Refractive index
sin i
µ=
sin r

33
When light passes through dense to rare medium, it does not go straight, but deflects from
its path. The measurement of this deflection is refractive index. At 20oC, water has
refractive index of 1.333 but normal milk has 1.344 to 1.3445. The instrument measuring
the refractive index is Abbe refractometer.
4.13 Oxidation-reduction potential
Oxidation means loss of electron and reduction means gain of electron. Redox potential
means the potential of oxidation and reduction. The redox potential of milk is Eh +0.2 to 0.3
volts. Two methods of measuring the redox potential is electrometric and colorimetric. This
potential decreases when boiling the milk. The growth of microorganisms also decreases
the redox potential.
4.14 Adhesiveness of milk
The adhesiveness is due to casein so casein is used to make gums in the industry.
4.15 Effect of heat on milk
The objectives of heating the milk are
1. The destruction of pathogenic and other microorganisms.
2. For the concentration of the milk.
3. For inactivation’s of milk enzymes.
4. To mix other constituents simply.
In factory the milk is heated during pasteurization, fore warming, condensation, drying etc.
The effect may cause caramelization, cooked flavors and loss of some nutrients. This
depends on the temperature of heating.
4.16 Significance of physical properties of milk
1. Helps in detection of the adulteration.
2. Helps in manufacture of dairy products (fermented and unfermented).
3. Helps in the fabrication of dairy equipments.
Assessment of physical and chemical changes in milk and milk products during
manufacture

34
Chapter 5

Milk components

5.1 Water: Free, Bound and Crystallized water

Water act as a suspending, dispersing and dissolving media for the other components of
milk. It occupies about 87 per cent in fresh milk. It exists in three forms such as free water,
bound water and crystallized water. Any variations in the amount of other constituents also
affected the water percentage.
5.1.1 Free Water
Largest portion of water exists in this form; this is the one which act as dispersing and
dissolving media for other constituents. It freezes at 0oC and evaporates at 100oC. It can
dissolve soluble substances. It can be removed easily while processing of milk. It is also
supportive to microbial growth.
5.1.2 Bound water
It exists in milk as binding with protein (through hydrogen bond), fat globules and
hydrophobic radical of milk constituents. In fresh milk, 3.18 % of the total water exists in this
form. It is very hard to remove while processing milk. This can be neither freeze at 0oC nor
can evaporate at 100oC and it cannot act as a solvent. It cannot support microbial growth.
5.1.3 Crystallized water
This water exists within chemical structure of any of the milk constituents eg α-lactose
hydrate (C12H22O11.H2O). This is most stable and hence hardest to remove. It also cannot
support microbial growth.
In various milk products, high extent of water makes it liable to microbial growth. Hence, it
is essential to remove water up to required level. But it is not advisable to remove water
below required level. For e.g. 2-3 per cent of water content is normal for milk powder but if
water is below 1.89 per cent it has been observed that milk powder oxidizes, and which will
results bad smell, solubility reduces and protein denatures.
5.2 Milk Carbohydrates - Lactose
It is sugar found in mammal milk. In cow's milk it is found from 4.4 to 5.2 per cent in an
average of 4.8 per cent anhydrous lactose, whereas in human milk it contains about 6.3 per
cent. Lactose is present in milk as true solution. This usually amounts to 50-52 per cent of

35
the total solids in skim milk. When it crystallizes from water, it forms hard gritty crystals,
which have one molecule of water of crystallization (C12H22O11.H2O). These gritty crystals
sometimes appear in certain milk products under certain circumstances under which these
crystals are allowed to grow in size. These crystals appear in ice cream when the mix
contains a high proportion of milk solids. The lactose content of milk is increased slightly by
the over feeding of carbohydrates, especially soluble carbohydrates or decreased by
mastitis infection of the udder.
Lactose is the major carbohydrate in milk. Milk contains traces of other carbohydrates but
no polysaccharides. The glucide compounds like hexosamines and N-acetylneuraminic
acid occurs in milk, but these are largely associated with proteins and cerebrocides.
Lactose solution in water has reducing properties and reduces fehling solution and
ammonical silver nitrate.The reducing power of lactose is doubled by hydrolysis as both the
sugars formed as a result of hydrolysis have one active aldehyde group each. Lactose is
insoluble in alcohol and ether but is soluble in hot acetic acid. Oxidation of lactose in slightly
acid condition yields formic acid and finally laevulinic acid, but oxidation by concentrated
nitric acid breaks lactose into oxalic acid and carbonic acid. When lactose is heated
between 110-130oC, the lactose hydrate crystals loose the water of crystallization and
above 150oC they turn yellow and at at 175oC they turn brown and form caramel. The
slight burning and characteristic colour of cooked milk is due to the formation of caramel,
though the milk and milk products are never treated to such high temperatures under
ordinary processes. Lactose, when gently warmed with nitrogeneous bases like ammo-nia,
certain amines, amino acid, any of which may be present in heated milk, forms brown
coloured complexes. Thus, the extent of these changes will depend upon the severity of the
heat treatment and the time of holding. Pasteurized milk is only slightly effected, while steri-
lized milk or evaporated milk have more effect. Lactose is a valuable product and is used
for the preparation of infant, weaned food products & invalids. This is the main source of
galac-tose, present in brain & nerve tissues of human body. Lactose helps in assimilation of
calci-um and phosphate from intestines has a tonning effect on the aliment-ary canal. Whey
is the main sourse for its manufacture.
5.2.1 Physical properties of lactose
Lactose is normally found in two crystalline forms i.e. α-hydrate and β-anhydrous.

36
1. α-hydrate
Commercial lactose is α-lactose monohydrate (C12H22O11.H2O) or α-hydrate. It is prepared by
concentrating an aqueous lactose solution to super saturation and allowing crystallization to take
place at a moderate rate below 93.5oC. It is stable solid form at ordinary temperature because it
forms hydrate below 93.5oC.
20

Specific rotation in water [α ] = 89.4 melting point = 201.6oC.

D

α-may form a number of crystal shapes, depending on the conditions of crystallization, the most
familiar forms being prism and tomahawk shapes. As the crystals are hard and not very soluble,
they feel gritty in mouth as sand particles may arise problems in ice cream, condensed milk or at
cheese (depends on size and number of crystals).
Size -10 µ or small -undetectable.
Above 16 µ or below 30 µ - tolerated without effect.
30 µ or large - sandiness’
Other physical properties
a. Density : Various lactose crystals differ slightly.
α-hydrate = 1.540 α- anhydride = 1.544 ( from dehydration)
β-anhydrate = 1.589 α- anhydride = 1.575 ( from crystallization by alcohol)
Densities of lactose solutions are not straight line function of concentrations.
b. Relative sweetness: β-lactose is sweeter than α and β is appreciably sweeter than the
equilibrium mixture except when the concentration of lactose solution equals or greater than 7 %.
Since there is approximately 63 % β in the equilibrium mixture, a β-lactose solution differs less in
sweetness from a solution in equilibrium than does α-lactose solution. However, for practical
purposes there is little advantage in using β for sweetness in preference to an equilibrium solution
at these concentrations, since the small difference is quickly eliminated by mutarotation.
5.2.2 Chemical properties of lactose

O
CH2O CH2O

OO O
O O
O O
(galactose) (glucose) α-form
β-LACTOSE

37
 O O
CH2O CH2O O
CH2O
O O CH2O
O O
O
Pyranose form furanose form
(galactose) (fructose)
LACTULOSE
Fig. 5.1 Chemical structure of lactose and lactulose.
Hydrolysis of lactose by acid does not occur easily. If it occurs at high temperature, low pH,
many reactions takes place as well. Lactose can, however, simply be hydrolysed using the
enzyme lactase (β-D-galactosidase). This enzyme is highly specific for the β-1,4 linkage of
a galactopyranose residue. Besides glucose and galcatose the enzyme also produces
some di- and oligosaccharides up to a few percent of the hydrolyzed lactose.
Several reactions of lactose occur when milk is heated. Lactose may isomerize into
lactulose. That means the glucose moiety converts to a fructose moiety (Fig.5.1).
Isomerization of the glucose moiety into mannose may occur as well, yielding epilactose (in
a trace amounts). The quantity of lactulose in heated milk products can be used as an
indica-tor for the intensity of the heat treatment . On heating, caramelization also can occur.
A mix-ture of reaction products is formed, including:
Hydroxymethylfurfural Furfuryl alcohol

H2CO CO H2COH

Acetol CH3.CH.CH2OH Formic acid HCOOH Acetic acid CH3.COOH

Methylglyoxal CH3.CO.CHO Pyruvic acid CH3.CO.COOH
Formaldehyde HCOH Levulinic acid CH3.CO.CH2.CH2.COO
The proportion of the products formed depends on concentration of sugar, pH, heating
time, and temperature. The very important maillard reaction occurs in the presence of
amino compounds in milk ( mainly concerns the ε-amino groups of the lysine residue in milk
proteins). It involves formation of a Schiff base between the amino group and the aldehyde
group of lactose. The initial reaction product undergoes a series of rearrangements,
yielding nitrogeneous reaction products in addition to such products as mentioned above

38
for caramelization. Further reactions lead to brown color, loss of nutritive value, and off
flavors. All these changes occur on prolonged storage, and especially during heating.
Lactose is ≈ 0.3 times as sweet as sucrose. The sweet taste in milk is somewhat masked
by the protein, primarily the casein. Whey has a sweeter than milk. The mixture of glucose
and galactose formed by hydrolysis tastes much sweeter than lactose.
5.2.3 Physiochemical aspects of lactose
5.2.3.1 Equilibrium in solution (Mutarotation)
Lactose exists in two forms, α and β. By definition, α is the form having the greater optical
rotation in the dextro direction. The specific rotation of a substance is characteristic of that
substance and is defined as the rotation in angular degrees produced by a length of 1 dm
of a solution containing 1 g of substances per ml. Therefore, the specific rotation may be
20
a
represented by the formula [α ] = 100
D
lc
Where, α = Specific rotation. c = concentration of substances in g/100 mil solution.
a = degrees of angular rotation.l = length of the tube in dm.
Also, important besides the variables of the equation, are temperature of the solution,
wave-length of the light source, and concentration of the solution. The standard light source
used to measure optical rotation has been the bright yellow D lines of the sodium spectrum,
but the single mercury lines, λ 5461 Ao is now used frequently for precision measurements.

Generally the specific rotation is reported at 20oC and expressed as: [α ]20D or [α ]20Hg .
The following formulas express variations in specific rotation in terms of these variables:

[α ] tD = 56.75 – 0.017C- 0.058 T (D line, λ = 589 ηm)

For pure α-lactose : αD = 91.1 at 20oC and For pure β-lactose : αD = 33.2 at 20oC.
(Where, c = is gm anhydrous lactose /100 ml solution)
T = is degree centigrade.

[α ] tHg = 66.25 – 0.007 C – 0.054 T. (Hg line, λ = 546 ηm)

Where, C = is gm of lactose monohydrate /100 mil solution.

When either of α and β lactose is dissolve in water, however there is a gradual change over
of one form to other until equilibrium is established. Regardless of one form used in prepa-

39
ring a solution, the rotation will change (mutarotation) until [α ]20D = 55.3, at equilibrium
(anh-ydrous weight basis). This is equivalent to 37.3 % in α-form and 62.7 % in the β-form;
since the equilibrium rotation is the sum of the individual rotations of the α and β forms. The
62.7
equi-librium ratio of β to α at 20oC, therefore, is =1.68.
37.3
Mutarotation has been shown to be a first order reaction, the velocity constant being
indepe-ndent of reaction time and concentration of the reactant.
In solution, conversion of α- to β- lactose and vice versa occurs.
α-lactose β- lactose, reaction constant k1
β-lactose α-lactose, reaction constant k2
k1 [β ]
Mutarotation equilibrium ratio, R = R=
k2
;
[α ]
The rate of mutarotation reaction has the constant K= k1+ k2
[α ]
If we dissolve, for instance, α lactose and if we define x =
[α ] equilibrium
 β 

α
  β αe   R 
ln   = kt ln  × =kt ln   = kt
 ( x −1) 
or, or, We have,
 α −1  α α − α e 
α 
 e 

(
β = α 1 − e −k t )
if time is very large the term e-kt becomes too small so it is neglected, then it should be,
βt = αo e-k t or, αt = βoe-k t
In other words, the proportion of the mutarotation reaction that has been completed at time
t is given by 1-e-kt
The same holds for conversion of β-lactose. The changes may be observed by using
polarimeter. The rotation of plane of polarization then is found to change (to mutate) with
time because α- & β-lactose differ in specific rotation. Hence the term called “mutarotation”.

40
 %
A k B
100 5

80 K=4 4
(T=40oC)
60 3

40 K=0.2 2
(T=15oC)
20 1

0 0
0 0.5 1 1.5 2 0 2 4 6 8
T(h) pH

Fig. 5.2 Mutarotation of lactose solution. (A) Course of the reaction (% finished) against
time t. (B)Mutarotation reaction constant k (h-1) as a function of pH (approx. 25 oC).
Mutarotation rate k depends closely on temperature. At 20oC and pH 6.7, k ≈ 0.37 h-1, it
increases by a factor of 3 or more per 10oC rise in temperature. At room temperature it
takes many hours before mutarotation equilibrium is reached; at 70oC a few minutes. (Fig.
5.2 B)
Several substances may affect the mutarotation rate. For example the salt in milk increases
the reaction rate by a factor of almost two as compared to the rate in water.
The mutarotation equilibrium likewise depends on temperature: R = 1.64─0.0027 T, where,
T is degrees Celsius. Thus the change in temperature causes mutarotation.
Mutarotation depends on lactose concentration. With increasing concentration, k decreases
R changes as well. If other sugar such as sucrose present in high concentration, k
decreases considerably. At very high lactose concentration, i.e., in amorphous lactose as,
for example, occurs in spray-dried milk powder, after equilibration R ≈ 1.25, independent of
temperature; mutarotation may occur, but extremely slowly.
Table 5.1 Some properties of lactose
1. Solubility in water ( q, in g /100 g water) as a function of temperature (T, oC);
α-lactose : log q ≈ 0.613 + 0.0128 T β-lactose : log q ≈ 1.64 + 0.003 T
Equlibrium solution; q ≈12.48 + 0.2807 T + 5.067(10-3T2 + 4.168(10-6 T3 + 1.147(10-6 T4
2. Density, viscosity, and refractive index as a function of concentration.

Conc.of Density of Apparent density of Viscosity of Ref. index of

lactose(g solution at lactose dissolved in solution(m.Pa.s) soln at 25oC,
/100g water) 20oC kg/m3 water (kg/m3) 20o 60oC λ = 589 ηm

41
 0 998.2 - 1.00 0.47 1.3325
10 1043 1750 1.38 0.70 1.3484
20 1082 1629 2.04 0.90 1.3659
30 1124 1592 3.42 1.29 -
40 1173 1591 7.01 2.19 -
c. Specific rotation α
α is expressed in degrees of arc of rotation per cm path length in a hypothetical solution of
1 g anhydrous lactose solution. The rotation depends on temperature (T, oC), on the wave
length of light (λ), and somewhat on concentration(C,g /100 mL solution), etc.The equations
are given above.

Lactose in solution α β
β/ α = 1.64 – 0.0027 T

Supersaturation
Supersaturation

T>93.5oC
T<93.5oC

Amorphous lactose
β/ α = 1.25
Water uptake

Water uptake
T<93.5oC

Dissolve

T>93.5oC

T≈100, presence
of water vapor β - anhydrous
Dissolve, T<93.5
T≈100, in vacuo
water uptake Anhydrous
Α-hydrate
T<93.5 α- unstable
(1 mol H2O)
T≈150, presence
of water vapor
Anhydrous
Dissolve, T<93.5 α- stable
CH3OH

+HCl

Supersaturation
Compound
crystal α 5 β3 in ethanol

Fig. 5.3 The different forms of lactose. T = Temperature (oC)

42
5.2.4 Crystallization of α-lactose hydrate

Crystallization is of great practical importance. Because α-hydrate is poorely soluble, it may

crystallize in some milk products, especially ice cream and sweetened condensed milk.
Large crystals can easily be formed because both nucleation and crystal growth are slow.
We usually have to add numerous tiny seed crystals to ensure the rapid formation of
sufficient, hence small sized crystals.To prevent segregation and development of
“sandiness” in milk products, the largest crystals formed should be no more than 10 µg in
size. This implies that at least 1010 crystals per gram of crystalline lactose should be
present. The α-lactose hydrate crystals can have many geometrical form ( but the crystal
lattice is always the same). The commonest shape is the “ tomahawk” (Fig.5.4 below).
Usually, the crystals does not grow in the direction of the ‘b’ axis, i.e., the crystal faces oīo
and k or 150. Likewise, lateral faces do not grow at all. Consequently, the “apex” of the
crystal is also the point where the crystal started to grow. Furthermore, crystal growth is
slow, far slower than may be accounted for by the combined effect of mutarotation and
diffusion of α-lactose to the crystal.
Presumably, there is some difficulty of fitting molecules into the crystal lattice.But otherwise
the observations of the preceeding paragraph are largely explained by the inhibition of β-
lactose. It appears as if β-lactose fits well into the oīo and oī ı faces of the crystal lattice but
then prevents any further uptake of α-lactose. Growth of other faces is inhibited as well (
Fig. 5.6 and Table 5.2 below). If very little β-lactose is present ( difficult to accomplish), the
oīı faces grow fast, causing formation of needle crystals.
Several other substances can retard crystal growth; the individual crystal faces are inhibited
in different ways, which lead to variation in crystal habit. Some inhibitors, such as riboflavin,
are present in milk. Some additions can speed up the growth rate of certain crystal faces.
If α-lactose hydrate is purified by recrystallization, its rate of crystal growth decreases
(Table 5.2, below). Moreover, the pH of the lactose solutions falls upon further
recrystallization.

43
 -b 010

K (~150) 110

011
Fig. 5.4
110 011 100 Common shapes of the α-
lactose hydrate crystal. The
main axes (a,b,c) and the
100
110 indices of the various faces
-a are given.
110
010
70
-c

It appears that a crystal growth inhibitor is present; it has a stronger affinity for the lactose
crystal than α-lactose itself. The inhibitor is a mixture of lactose monophosphates; its
concentration in milk is about 15 mg.L-1. It particularly inhibits at low supersaturation and
causes inhibition of nucleation in lactose solutions. The substance can be removed by ion
exchange.
Table 5.1 Examples of the rate of growth of some faces of an α-lactose hydrate crystal as
affected by liquid composition.
Growth (µm.h-1) of face
Supersaturation (%) Remarks oıo ııo ıoo ıīo

55 -
55 +10 ppm gelatin 3.8 3.3 1.3 0.3
55 +100 ppm riboflavin 1.2 1.0 1.0 0.4
55 + 10 ppm TMODAC* 2.7 0.0 0.0 0.0
120 - 43 34 21 12
55 Own pH 3.2 2.7 1.6 0.4
55 pH 7 6.6 5.0 2.7 1.2
55 3 × recrystallized 0.2 0.7 1.3 0.5
55 Nonionic* * 19.1 9.1 3.1 1.2
55 Nonionic + Inhibitor* * * 0.0 0.0 0.9 0.5
*
Trimethyloctadecylamonium chloride.
**
Solution passed through an anion exchanger.
***
Lactose Monohydrate.

44
5.2.5 Lactic Acid Fermentation

Lactose is the main source of of energy for most of the bacteria growing in milk. Commonly,
the organisms attack lactose by hydrolyzing it to form galactose and glucose. The latter
molecules are each fermented into lactic acid (CH3CHOHCOOH), but part of the galactose
may not metabolized. Lactic acid bacteria are of two types. The one which can produced
only lactic acid is called homofermentative and those produce lactic acid as well as CO2,
acetic acid and ethanol.
Glucose + 2ADP + 2H3PO4 2 lactate + 2ATP + 2H2O
Lactic acid is an essential component of many milk products. Most bacteria produce about
1 % lactic acid in milk, but some of these can reach as much as 2 %. Such high lactic acid
concentration inhibit growth of most microorganisms.
5.2.6 Solubility of lactose
The solubility of α and β lactose differ considerably and it depends on the temperature. If α-
lactose is brought in water, much less dissolves at the beginning than later. This is because
of mutarotation : α-lactose is converted to β, hence the α- concentration diminishes and
more α can dissolve. If β-lactose is brought in water, more dissolves at the beginning than
later (at least below 70oC): On mutarotation more α-lactose forms that can stay dissolved,
and α-lactose starts to crystallize.
200

100
6
1

1.
2.

β
ble at
e
40 lia di d
e le te α
a
rm ab ur
Solubility

te st at
in et
a ts
M No
20

10
5
0 20 40 60 80 100
Temperature (oC)

Fig. 5.5 Solubilities of α- and β- lactose, final solubility of lactose (curve1), and
supersaturation by a factor of 1.6 and 2.1 as a function of temperature.
45
The solubility thus depends partly on the mutarotation equilibrium, the rate of dissolution on
the mutarotation rate. The so-called final solubility is identical whether we dissolve α- or β-
lactose. It is R + 1 times the solubility of α-lactose. This applies below 93.5oC because
above this temperature β-lactose determines the final solubility. At lower temperature, it
takes a long time to reach equilibrium.
When α-lactose hydrate is added in excess to water, with agitation, a definite amount
dissolves rapidly, after which an additional amount dissolves slowly until final solubility is
attained. Its solubility in water is only 17.8% at 77oF (25oC).
Β-lactose at room temperature is about 7 times more soluble than α-form. The initial
solubility is the true solubility of α-form. The increasing solubility with time is due to muta-
rotation. Since the solution was already saturated with α, α formed by mutarotation will
crystallize to reestablish equilibrium. Since β-lactose is much more soluble and
mutarotation is slow, it is possible to form more highly concentrated solutions by dissolving
β-rather than α-lactose hydrate. The increase solubility makes it appear sweet to taste and
favors the use of it in dietary and baby foods.
The overall rate of lactose crystallization can be summarized by the reaction.
Β-lactose α-lactose α-lactose hydrate crystals.
Lactose solution can be supersaturated easily and to a considerable extent. At
concentrations over 2.1 times the saturation concentration, spontaneous crystallization
occurs rapidly, probably because of homogeneous nucleation (i.e., formation of nuclei in a
pure liquid). At less than 1.6 times the saturation concentration, seeding with crystals
usually is needed to induce crystallization, unless we wait a very long time, the solution is
thus metastable. In the intermediate region, crystallization depends on several factors, such
as time.
5.2.7 Uses of lactose
The large amount of lactose is consumed in the form of milk products but the sugar itself
has only few industrial applications.
• as a constituents of infant foods and medical products.
• in the early stages of brain formation, galactose is needed , which is found in lactose.
• for the manufacture of tablets and capsules and general fillers.
• use in fondants and tablets like candies.

46
 • use in baking industries to produce a desirable brown color in pie, crusts, cookies and
other baked goods.
• to improve the flavor and body of the product like chocolate milk, butter milk, and
modified SMP
• used as substitute for cream in coffee as a dry mixtures.
• used for the formation of penicillin due to its slow rate of fermentation properties.
• lactobacillus spp. of bacteria only utilized lactose sugar and produce lactic acid which
is used as substitute for acetic acid and citric acid in food stuffs.
• used as an ingredients of the medicine in which mold is grown.
• used as a reducing agent in silvering of mirrors.
• minor use of lactose include use of it military technology for making of smoke screen
and signal and target candles.
For the manufacture of several dairy products like sweetened condensed milk, instant milk
powder, stabilized whey powders and lactose ice cream etc., the process of crystallization
is important. The control of lactose crystallization become important for various products
like ie cream etc.
5.2.8 Nutritional and physiological effect of lactose – Lactose intolerance
Milk and its derivatives contribute lactose to the diet. In the digestive tract, lactose may be
fermented by bacteria; in the intestine it may be absorbed directly or hydrolysed by β-D-
galactosidase (Lactose) and its components absorbed. Β-D-galactosidase is an intracellular
enzyme which in man is found within the cells of the intestinal mucous membrane
hydrolysis, therefore, occurs during transport through the intestinal wall.
A great deal of interest has recently centered in the enzyme lactose because it brings about
certain diarrheal syndromes in infants, but also because of its wide spread deficiency in
adults who normally consume little or no milk after weaning. “Lactose intolerance” causing
abdominal cramps, gaseous distention or diarrhoea in severe cases, is atributed to a
deficiency of lactase in the intestinal mucosa. Lactose deficiency is detected by a biopsy of
the mucosa or by feeding lactose to the subject after a period of fasting and measuring the
rate of increase in “ blood sugar” level.

47
The significance of this subject is obvious because of its implications on the suitability of
milk as a food after weaning in countries whose there is a high incidence of lactase
deficiency in the population.
Lactase may also be transferred to the blood or urine (lactosuria) without being hydrolysed,
particularly after consumption of a large quantity of lactose or during lactation.
5.3 Milk Lipids

Lipids are esters of fatty acids and related components that are soluble in non polar organic
solvents and insoluble or nearly so, in water. Alternatively, the term fat is used. But “fat” is
usually considered to consist largely of a mixture of triglycerides, especially when the
mixture is partly solid at room temperature.
Nearly all of the fat in milk is in fat globules. It can therefore be concentrated readily by
means of gravity creaming, possibly followed by churning. Products rich in fat, such as
cream and butter, have a specific and often desired flavor and texture. On the other hand,
milk fat is prone to deterioration, leading to serious off flavors. The consistency of high fat
products greatly depends on the crystallization of the fat. In turn, crystallization behavior of
milk fat depends on such factors as the widely varying fat composition.
5.3.1 Composition
About 98 % of milk fat is a mixture of triglycerides. Various other lipids, even present in
trace amounts, are also dissolved in the fat. Most of the more polar lipids are in the fat
globule membrane.
The chemical and physical properties of a lipid primarily depend on the kind of molecule.
For example, triglycerides are different from lecithin’s or sterols. But each lipid class
consists of many different kinds of molecules since it contains various fatty acid residues.
Such a fatty acid pattern is an important factor in determining lipid properties, such as
melting range, chemical reactivity, and nutritional value.
The main variable among fatty acids in milk fat.
a. Chain length : Most fatty acids contain 4-18 carbon atoms; even numbered acids are
predominant.
b. Number of double bonds (the degree of unsaturation): It mainly determines chemical
activity, including proneness to autoxidation.
c. Position of double bonds, e.g., conjugated ( -CH=CH-C=CH- ) or non-conjugated.
48
d. Configuration of double bond: Each double bond can be either in the cis or trans
position. The cis form is the common one in nature. Milk fat contains about 3 mol %
trans acids, predominantly monosaturated.
e. Branching: Nearly all the fatty acids have an unbranched carbon chain. But some have a
terminal –CH(CH3)-CH3 group.
f. On hydrolysis, the glycerides may yield some fatty alcohols and fatty aldehydes, in
addition to fatty acids.
On heating, some of the fatty acids show chemical reactions. Residues of 3-ketoacids give
rise to free methylketones, 4- and 5- hydroxyl fatty acids residues give γ- and δ- lactones,
respectively. These compounds are present in fresh milk and are partly responsible for the
characteristic flavor of milk fat. Heat treatment or during long storage of dried milk, cause a
typical flavor. At still higher heating temperatures (e.g., 150oC)the position of double bonds
changes and some are transformed from cis to trans. Intersterification or randomizing (i.e.,
interchange of fatty acid residues among their position in the triglyceride molecule) also
occurs.
The fatty acid pattern on the triglycerides is highly variable and depends on several factors.
Milk fat can be characterized as follows.
a. Its fatty acid composition is very wide. Including fatty acids with keto or hydroxyl
groups, with an uneven number of C atoms, with branched carbon chain, the total
number is some 250 different acid residues.
b. It contains a relatively high proportion (15-20 mol %) of short chain fatty acid residues,
with 4-10 carbon atoms. This is typical of milk fat of ruminant species.
c. The proportion of saturated fatty acid residues is high, e.g., 70 mol % (approx. 63 %
w/w).
d. Oleic acid is the most abundant of the unsaturated fatty acid residues (about 70 %).
e. The other unsaturated fatty acid residues are present in a wide variety of chain length
as well as number, position, and configuration of the double bonds.
Various lipids are unevenly distributed among the physical fractions of milk, the fat
composition of different milk products varies. The largest differences originate from
variations in the amount of material from the fat globule membranes. Anhydrous milk fat is

49
prepared from butter by melting it, and by separating and drying the oil layer obtained; it
composition is virtually equal to the fat in the core of the milk fat globules.
Milk fat can be modified chemically by intersterification (which occurs also somewhat
obtained can no longer be called milk fat). Milk fat can be fractionated into high and low
melting portions by letting it partly crystallize. In milk, fat changes can occur by
autoxidation, lipolysis and intense heating.
5.3.2 Properties of individual lipid classes
1. Tryglycerides
Milk fat consists almost entirely of triacylglycerols or triglycerides, which constitute nearly
97-98.5 per cent of the total lipid. Small quantities of di- and monoacylglycerols are also
present. These glycerides are formed by the esterification of the hydroxyl groups of
glycerols with the fatty acids varies vastly and has a significant influence on the chemical,
physical and organoleptic properties of the fat.
There are as many as 437 different fatty acids from bovine milk has been compiled. Among
there these only five fatty acids are present in significant quantities, nearly 80 %, and are of
nutritional, chemical and physical importance. These are oleic, palmitic, butyric, stearic and
myristic acids. The remaining fatty acids are present in small quantities. Nearly 50 of the 80
commonly encountered fatty acids together make up hardly 1 % of the total.
The ways of classifying the myriad types of fatty acids encountered in milk fat based on
saturation (saturated, mono-unsaturaed, polyunsaturated fatty acids), geometric isomerism
(straight chain, branched chain); chain length ( short, medium and long), etc.
Polyunsaturated fatty acids are further classified as either cis or trans (geometric
isomerism), or conjugated- non conjugated (positional isomerism). Not very often, fatty
acids are also termed as either even or odd numbered fatty acids are rare and not
commonly found in milk fat. A convenient method of notation of fatty acids involves two
numbers designating the carbon chain length and the unsaturation (number of double
bonds). Thus, C4: 0 (butyric acid) which have 4 carbon atoms and is saturated without any
double bond. Similarly C18: 1 or C18: 2 denotes oleic acids and linoleic acid having a carbon
chain length of 18 with 1 and 2 double bonds, respectively. Milk fat contains a wide range of
fatty acids varying in chain length from C4 to C26.

50
Table 5.2 Fatty acid profile of milk fats of different species (mole %).
Component mole % Fatty acid cow buffalo Goat Sheep
Butyric 4:0 8.8 11.4 7.6 8.4
Capric 6:0 3.5 3.1 4.5 5.4
Caprylic 8:0 2.0 1.0 6.2 5.8
Capric 10:0 3.0 1.6 11.1 10.1
Lauric 12:0 3.8 2.6 5.1 6.0
Myristic 14:0 9.9 10.6 11.2 11.8
Palmitic 16:0 26.1 30.3 21.5 20.4
Stearic 18:0 9.1 10.5 7.5 5.4
Higher saturation 20-26 1.0 0.7 1.3 1.3
Unsaturated (mono) 10:1-14:1 1.8-3.6 1.0-3.6 - -
Lower 16:1 26.2 21.6 24.2 22.2
Unsaturated (poly) 18:1 3.5 2.0 1.2 3.2
2. Di- and Monoglycerides
Some of these occur in fresh milk fat. Lipolysis increases their quantities. Diglycerides are
apolar and do not differ much from triglycerides in properties. Monoglycerides present in far
smaller quantities, are fairly polar; they are surface active and thus accumulate at an oil
water interface.
3. Free fatty acids
These already occur in fresh milk and lipolysis increases their amount. The shorter acids
are somewhat soluble in water. Fatty acids can dissociate into ions; their pK is about 4.8. In
the milk plasma, they are thus predominantly in the ionized form ( i.e., as soaps), and these
are much more soluble in pure water than the pure fatty acids.
Fatty acids dissolve well in oil, though only in the nonionized form; moreover, they tend to
associate into dimmers. Obviously, the partition of the acids over the oil and water fractions
is rather intricate.
The shorter acids (C4 and C6) are predominately in the plasma, the longer ones (C14) in the
fat. The other acids are distributed between both fractions, though more go into the fat with
decreasing pH (i.e., with ionization becoming weaker). This shorter acids are responsible

51
for the soapy rancid flavor perceived after lipolysis. The fatty acids especially long chain
ones, are surface active and tend to accumulate in the oil water interface.
4. Compound lipids
Most abundant in milk are the phospholipids or phosphatides. Most of these have two
charged groups (an acid and a basic one) and therefore are fairly polar. They do not
dissolve well in water or oil, but form micelles in either micelles in either liquid. They are
highly surface-active and tend to associate with several proteins to yield lipoproteins.
In milk, the component lipids are largely in the fat globule membrane; in plasma, they are
present in lipoprotein particales or “milk membrane”.
5. Unsaponifiable matter
It consists largely of cholesterol, which is fairly apolar and associates easily with phospho-
lipids, accordingly, part of cholesterol is in the fat globule membrane. A fraction of the
cholesterol is esterified to a fatty acids (i.e., actually being saponifiable). Carotenoids are
responsible for the yellow color of milk fat.
5.3.3 Autoxidation
The double bonds in a fatty acids or a fatty acid residue can oxidize. Several
components can be formed from oxidation. Some of these can be perceived in
exceptionally low concentrations and thereby cause off-flavors, including tallowy, fatty,
fishy, metallic, and cardboard-like.
Off-flavor development can cause problems in beverage milk, sour-cream butter milk,
cream and especially long keeping high fat products like butter and whole milk powder. The
complex of reactions involved is highly intricate, and there are several complicating factors.
The formed hydro peroxides have no flavor. But they are fairly unstable and can break
down in various ways to form unsaturated ketones and aldehydes, some (especially those
with the group –CH=CH-CH=CH-COOH) having a very strong flavor i.e., they may have a
threshold concentration as low concentration as low as 10-3 ppm.
The reaction rates depends significantly on such conditions as temperature. The off-
flavors develops under certain conditions e.g., high temperature do not always correlate
satisfatorily with low temperature.
The rate at which off-flavors develop especially depends on the extent of un-saturation.
Antioxidants can block the chain reaction or prevent the initiation. Natural antioxidants are

52
tocopherol (which reacts with radicals) and β-carotene, which can react with singlet oxygen.
Synthetic antioxidant also called synergists (e.g., phospholipids dissolved in fat) inhance the action
of antioxidants. Citrate can react with the catalyzing metal ions.
Autoxidation usually takes some time to set in. Initially, the antioxidants are consumed; after this
has been achieved, peroxides are first liberated and subsequently are broken down to form
perceptible amounts of flavor products.
Autoxidation of milk fat in milk and milk products usually starts with the phospholipids of the fat
globule membrane. These lipids have highly unsaturated fatty acid residues. Moreover, cupper, the
main catalyst, can be in the membrane of the fat globules but not in the core, which contains the
triglycerides. Copper is particularly active as a catalyst if phospholipids are present; moreover, at
least in milk and buttermilk a little ascorbic acid is needed. Copper entering the milk by
contamination with as little as 5 µg.kg-1 suffices for an oxidized flavor to develop, whereas in others
200 µg.kg-1 is sufficient. Iron can also be active as a catalyst, but not in the presence of proteins,
i.e., not in milk.
Autoxidation of fat in milk can also be enhanced by exposure to light of short wavelengths.
Riboflavin is an essential factor in light-induced autoxidation.
Early lactation milk on the average is more prone to develop flavors caused by fat oxidation.

ANTIOXIDANT
A NTIOXIDANT PEROXIDES

CARBONYLS

TIME

Fig. 5.6 Relative concentration of antioxidats, hydroperoxides,

and free carbonyl components during autoxidation of a fat.
5.3.4 Crystallization of fat
Crystallization of fat (tryglycerides) is a complicated phenomenon, especially for milk fat with its
very broad composition.

53
 1. Melting range
The fats are widely varying melting points. This is reflected in the melting range of the fats in which
the acids have been esterified. The shorter the chain length and the greater the number of double
bonds, the lower the melting point. The other variables are as follows:
a. Odd number of carbon atoms: Odd numbered ones melt ~5 k lower than fits the series with
an even number of carbon atoms.
b. Branching: May cause a decrease by 1-40 k as compared to unbranched carbon chains.
c. Position of double bond: May make a difference up to about 20 k.
d. Trans: 20-30 k higher than cis.
e. Conjugated: ~25 k higher than nonconjugated.
Melting point of a triglycerides molecule also depends on the distribution of the fatty acid residues
over the three positions. A strongly asymmetric triglycride e.g., palmitic, butyric, has a lower
melting point than a symmetric one with the fatty acid residues.
The melting point range varies from -30 to +40oC, ilk fat usually consists of liquid as well as solid
fat, i.e., oil with various crystals.
Every individual triglyceride is tri-stearic; its melting point is 72oC, i.e., about 35 k above the final
melting point of milk fat. The “solid” triglycerides are thus dissolved in the liquid fat. The solubility of
a single triglyceride in oil in consistent with the thermodynamic theory of perfect solutions. But if
several triglycerides crystallize, they may interface with each other solubility. In turn, this effect on
the solubility depends on external conditions, as will become clear below. That explains why the
melting curve of milk fat cannot be simply derived from its composition. Naturally, a higher content
of high melting triglycerides causes a higher content of solid fat at, say, room temperature.

FIG. 5.7 Melting curve of milk fat.

2 3 1. Summer fat slowly cooled.
% SOLID FAT

80 2. Summer fat rapidly cooled.

1
3. Winter fat rapidly cooled.
40
80

0
-4 -2 0 0 20 40
o
Temperature ( C)
Solidification curves depend even more closely on external conditions than melting curve
do. This is because of super cooling may occur.

54
 2. Nucleation
A substance cannot crystallize unless nuclei have formed, i.e., embryonic crystals, just large
enough to escape immediately dissolving again. (The solubility of a small particles increases as its
radius of curvature decreases). Homogeneous nucleation (i.e., the formation of nuclei in a pure
liquid) often requires considerable super cooling to form nuclei within, say, a few hours. In facts, a
super cooling of about 35 k below the final melting point is needed. But nucleation is normally
heterogeneous, i.e., it takes place at the surfaces of very small “contaminating particles” such
particles are called catalytic impurities. As a rule, the number of impurities that catalyze, nucleation
(N) significantly increases with decreasing temperature.
In milk fat, if present in bulk (i,e., as a continuous mass), a super cooling of a few, say, 5 k, causes
significant catalytic impurities to induce crystallization. As soon as fat crystals have been formed,
they can, in turn, act as catalytic impurities for other triglycerides. For this to happen, a very small
super cooling suffices. This is because of epitoxy, i.e., the crystal lattice of the molecules to
crystallize almost fit on that of crystals already present. Milk fat in bulk may indeed show little
hysteresis between solidification and melting curves (Fig.5.8,A)
The situation may be very different if the fat has been emulsified. In bulk fat, some 103 catalytic
impurities per g would suffice to ensure rapid crystallization. But in milk, 1 g of fat is divided over
about 1011 globules, in homogenized milk even over > 1014 globules. In each of these at least one
nucleus must be formed. Consequently, considerable super cooling may be necessary, and
significant hysteresis between solidification and melting curves occurs (Fig.5.8, B)
× ×
A B

0.4
0.4

0 10 20 30 0 10 20 30
× proportion of fat being solid (×) after 24 h cooling to temperature T, and after warming
again.
Fig.5.8, A & B : Examples of the proportion of fat being solid (×) after 24 h cooling to
temperature T, and after warming again ( after keeping it at 0oC). (A) fat in bulk (B)
Same fat in recombined cream.

55
 3. Crystal growth
In milk fat, growth of nuclei or crystals is very slow. The large, elongated, and flexible
triglyceride molecules takes a long time before obtaining the correct position and
conformation to fit into the crystal lattice. Before a molecule actually fits in, it often diffuses
away again. Moreover, there are many competing molecules, which are so similar to those
in the crystal that they almost fit into its lattice. They have to diffuse out again before a
properly fitting molecule can occupy a site in the crystal lattice.
For isothermal crystallization of milk fat at 25oC, it may take 1-2 h before half of the final
amount is crystallized. A lower temperature implies a greater supersaturation. Crystallizat-
ion rates depend on conditions such as crystal size and the rate of removal of the heat of
crystallization.
4. Polymorphism
Triglycerides, like most molecules with long aliphatic chains, can crystallize in three
different modifications can be identified by ×-ray diffraction, denoted as α, β’, and β. Each
modifications is characterized by its crystal lattice type i.e., the mode of packing of the
molecules.) including the corresponding distances between molecules; therefore, the
modifications can be identified by X-ray diffraction. In the order α, β’, and β, the melting
points increase (for tristearate, e.g., 55, 64, and 72oC), as do the enthalpy of fusion and
density of the crystals. This implies that closeness and intricacy of fit of the molecules in the
lattice increase and their freedom of motion decreases in the same order. The α and β’
modifications are metastable. Transitions can only take place according to the scheme.
α
Liquid β’
β
other transitions cannot occur.
Nucleation usually occurs in the α modification. The α modification has a very short lifetime,
whereas β’ may persist for a longer time. But in milk fat, α crystals can be very persistent
and especially at lower temperature both α and β’ modification are found. The final melting
point in milk fat are approx. 22oC, 30oC, and 36oC for α, β’ and β, respectively.
The importance of the polymorphism is that a partially crystallized fat essentially is never in
equilibrium.
56
 5. Compound crystals
In fats, compound crystals readily occur. Milk fat exhibits extensive mixed crystallization,
probably, because of the very great number of distinct, though similar, triglycerides
molecules. Compound crystals are formed easily and abundantly in the α, and usually not
in the β, modification. Cooling the fat more rapidly to lower temperature causes more
compound crystals to be formed.
6. Size and shape of crystals
The geometrical form (habit) of a crystal should be distinguished from each other. Each
modification may include widely varying habits. In most cases, rapid cooling of milk fat
leads to formation of platelets. The ratio of the platelet sizes approximates 4:2:1; platelet
length ≈ 0.1- 3 µm; platelet concentration may be on the order of 1012 per gram of fat.
7. Rheological aspects
Fat crystals suspended in oil attract each other because of Van der Waals forces and
only repeal each other because of hard core repulsion ( i.e., when they touch). Accordingly,
they always flocculate and this causes formation of a continuous crystal network, in which
the oil is held if sufficient crystals are present (a few per cent by weight). The network
causes the fat to have a certain firmness, which increases with increasing fraction of solid
fat and depends also on size and shape of the crystals.
5.4 Proteins in milk
5.4.1 Chemistry of proteins
5.4.1.1 Definition
Proteins are high molecular weight polymers, generally over 10000 MW, of amino acids
covalently linked by the peptide bond. Most of the milk proteins contain more than150
Peptide bond
H H O H
R1 C COOH + H2 N C R2 R1 C C N C R2
NH2 COOH NH2 COOH
Amino acid 1 Amino acid 2 Dipeptide
amino acids. The properties of the various proteins are dictated by the amino acids in the
molecule and by their sequence in the polypeptide chain(s), which regulates the molecular
configuration of the protein molecule and the surface electrical charge. Proteins can exist in
helical coils, random coils, pleated sheets or in a combination of these forms. These
57
properties relate to the stability of the proteins in the food system and influence processing
of the products. Milk proteins generally possess several of these forms in a single protein
molecule.
Protein is one of the most essential nutrients of milk present in about 3.5 %. Milk protein
contains almost all of the essential amino acids and hence high nutritive value. Carbon,
Hydrogen, Nitrogen, Oxygen, Sulphur, and phosphorus are the elements present in protein.
In milk among the total protein, casein contributed 2.9 % and whey protein 0.6 %. Milk
protein may be divided into two main groups: Casein and whey proteins (lactalbumin,
lactoglobulin).
5.4.2 Casein
The caseins are characterized by their relatively high phosphorus content. Casein is a
generic term for a class of proteins that are synthesized in the mammary gland and make
up about 80-85 % of the total milk protein. It is present in the form of micelles or particles of
macromolecular sizes.
Casein is composed of four recognized components called α-s, β, κ, γ on the basis of
differences in their electrical charge. Chemically the casein system is defined as a
glucophosphoprotein, since it contains both carbohydrate (glycol group) and phosphorus as
integral parts of the protein. In milk, casein exists as its calcium salts, namely, calcium
caseinate, in distinct globular particles, ranging from 40-300 mµ in diameter. These
particles are called micelles; each micelle contains all the component casein held together
in part by calcium phosphate.
Commercial casein is obtained from fat-free skim milk by precipitation either by addition
of an acid, or by the addition of rennet extract (containing the enzyme rennin).
Caseins are high molecular weight (89,000) compounds. They are the charged. They are
the charged particles having IEP 4.6. The elementary composition of casein indicates
carbon (53 %), hydrogen (7.07 %), sulfur (0.76 %) and phosphorus (0.85 %). The factor
used for determination of protein is 6.38. This is because N2 content in milk is taken as
15.65 instead of normal value (16 %). Milk and milk products provide food proteins of
excellent quality for the nutrition of man and animals. Casein, the dominant protein of milk,
is a good source of amino acids, which are the indispensable for human nutrition.

58
 The milk protein differs considerably from each other in amino acid composition however
they all contain considerable amount of dietary essential amino acid. The α-lactalbumin
contains very high amount of tryptophan, while all milk proteins are rich in aspartic acid and
glutamic acid. The essential amino acids such as tryptophan, threonine, isoleucine, leucine,
lysine, methionine, cystine, phenylalanine, tyrosine and valine, all are found in milk.
The peptide chain obtained after enzymatic hydrolysis found to have phosphorus bound
to the peptide. Hence, they are also called phosphopeptides or phosphopeptone. Protein is
present as a phosphoric ester of serine in which the hydroxyl group reacts with one of the
3- hydrogen atom of phosphoric acid.
HO P O CH2H
H
CHNH2CO
O OH Serine
Phosphoric acids

The casein in normal milk accounts for close to 80 % of the nitrogen present. Casein is
present in milk in solution and as micelles containing calcium, inorganic phosphate,
magnesium, and citrate. The micelles are in colloidal suspension.
The diameters of micelles are between 80 nm & 250 nm in buffalo and between 50 nm
200 nm in cow casein. Casein is precipitated with or without those mineral components
depending upon the coagulating agent used. The casein of normal fresh milk only partially
precipitates on prolong heating at high temperatures. Good quality milk may be preheated
to render it more stable during condensing or drying, although there might be some
reduction of the ability or power to reconstitute if made from such milk. Preheating helps to
prevent age thickening. Adding small concentrations of sodium & potassium chloride
increases the heat stability of milk. Browning may result from the reaction of the amino
acids and carbohydrates to prolong heating at temperatures in excess of 100oC. Heat
aggravates the affect of acid in milk. Casein readily precipitates with acid or rennet. Acid
frees the casein by reacting with the calcium with which this protein is associated in milk.
The casein is then no longer able to remain in suspension, so it separates out. Another
means of coagulating casein is by the action of proteolytic enzymes such as rennin. Curd
strength is measured in grams of force required to cause a standard knife to pass through
the surface of a standard curd. This value may range from 30 to 80 g for fresh normal milk
from individual cows. Boiling reduces the curd tension nearly 30 %. The curd tension of raw
59
buffalo milk is reported to be 44.54g. This value is reduced to about 32.5 % by
pasteurization by the holding method & nearly 73 % by homogenization. Colostrums and
mastitis milk are unstable to heat during processing.
Globulin & albumin are soluble proteins of milk. They are called whey proteins because
they do not usually precipitate with acid and only partially precipitate with rennet. High
temperatures and strong salt solutions will precipitate the whey proteins, which account for
about 12 % of the nitrogen of milk. Casein also precipitated by alcohol is calcium caseinate.
The casein precipitated with weak acids is free of calcium.
When casein is precipitated with rennin, para-casein is formed. It contains more calcium
than Ca-caseinate. Pure casein is precipitated by heat, but in fresh milk prolongs heating at
high temperature (100oC) for 12 hrs or more hrs or (120oC) under pressure will cause the
precipitation of casein. On boiling fresh milk, a thin layer of finely precipitated casein,
together with other milk constituents including fat, forms a thin layer over the surface of the
milk. The application of heat to milk that is slightly acid will cause the precipitation of casein.
One to two molecules of serine combines with 1 phosphoric acid molecule a
diphosphoric ester is formed. Therefore, phosphoric acid may form a cross link between
polypeptide chain. Casein has acidic nature, as compared to other proteins, is quite
distinctive as it has considerable base binding properties and can even liberate CO2 from
carbonates.
Calcium–hydrogen-caseinate + Lactic acid Acid casein precipitates + Calcium lactate
(solution)
The protein precipitated only after pH 4.6. The progressible removal of Ca-starts at pH 4.6,
when lactic acid develops.
Ca-caeinate + Ca3(PO4)2 + 4CH3CHOH Ca(H2PO4)2 + 2(CH3CHOHCOO)2Ca+Casein↓
Concentration and dry form of milk when left for a long time gradually developed a brown
color. The browning in milk is caused by a maillard type of reaction between amino group of
milk protein and aldehyde group of lactose. Such products are unfit for human
consumption.
As lactic acid develops, calcium from colloidal solution of protein moves to soluble
condition and this process continuous till the milk curdling. The calcium removes from
calcium hydrogen carbonate to form calcium lactate. The protein is precipitated (or
progressible removal of calcium starts) only after pH 4.6. The concentrated and dry form of
60
milk, when left for a long time gradually develops a brown color. The browning in milk is
caused by a maillard type of reaction between amino group of milk protein and aldehyde
group of lactose. Such products are unfit for human consumption.
It is known that casein is the macromolecule in the form of micelles and each micelle
composed of about 1000 casein molecules. Casein is a mixture of different fractions and is
heterogeneous in nature. The different fractions are differing in composition, solubility and
rennet coagulation.
It has been found that in electric field, casein separates into three separate components
moving at different speeds. Those components are α, β and γ casein which are arranged in
descending order of mobility. The α and β casein are phosphoprotein of high quality where
γ-casein contain very little phosphorus. On the basis of electrophoretic analysis, whole
casein contains about 75 % of α-casein, 22 % of β-casein and 3 % of γ-casein. Alpha
casein is responsible for the stabilization of the micelle in the milk and it is the component
where readily attack. It is composed of two sub-fractions of α-s casein (or calcium sensitive
casein) and κ-casein (or calcium insensitive casein).
5.4.2.1 Precipitation of casein
Generally, casein can be precipitated by the following methods.
1. Acid coagulation of casein
The pH value of normal milk often ranges from 6.6 to 6.7. When acid is added, the pH will
be lowered and pH 4.6, i.e., IEP will be obtained in which casein will be precipitated. The
casein precipitated with weak acids is free of calcium or free casein at pH 4. Depending
upon the acid used, the product is termed sulphuric, muriatic(hydrochloric), or lactic acid
casein.
After the addition of acid, first of all reaction will be taken place between Ca++ / Mg++ salts
and acid. The added acid should be in excess amount to remove calcium ion (ca++) from
calcium caseinate. The Ca++creates metallic odor in processed product like cheese.
Ca-caseinate + HCl Casein + CaCl2
Calcium phosphate [Ca3(PO4)2] Calcium hydrogen phosphate Ca(H2PO4)2
Before precipitation micelle is defined as Calcium-caseinate and calcium phosphate.
Generally, hydrochloric acid is used for the prcipitation. The use of sulphuric acid will be

61
adversely affected on the quality of precipitated casein. Due to weakened of κ-casein and
weak absorb layer, permanent precipitation takes place at 4.5 pH.
Natural precipitation of casein
Naturally present microorganisms produce lactic acid which can cause coagulation.

Lactic acid
Lactose Lactic acid (pH 4.5)
Coagulation will be takes place
.
If a excess of acid is added to a certain coagulum the casein will redissolve, forming a salt
with the acid.
2. Alcoholic precipitation of casein
When the fresh milk is acted upon by microorganisms and the pH value is lowered than 6.6,
due to the formation of lactic acid, at this condition, if we add alcohol the precipitation will
takes place. It is called alcoholic precipitation of caasein.
The principle of alcoholic precipitation is the dehydration of adsorbed layer of micelle and
exposure of κ-casein and ultimately action of developed acidity due to microbial
contamination upon the milk.
Hence alcoholic precipitation of milk will be only possible if the milk is contaminated by
microbes and acidity is developed. This method is used to quick check for freshness of
milk. If the milk is immediately precipitated out on adding alcohol, it is not fresh and vice
versa.
3. Enzyme precipitation of casein
The dynamic stability of micelle is due to κ-casein but it is most susceptible to enzymes. It
responces positively to enzyme. When enzyme like renin and pepsin are added in milk,
precipitation of casein takes place with the formation of para-casein or para-casein. The
reaction is as follows.
Rennet
κ-casein of micelle para-κ-caseinate + Glycomacropeptide
Hydrophobic Hydrophillic
(insoluble in water) (soluble in water)
Actually κ-casein has 169 amino acids molecules. In the position of 105th , there is
phenylalanine and on the position of 106th there is methionine. The link between these
amino acids (105----106 ) is the most weakest for enzyme action, thus it is more favourable
for proteolysis. Thus almost all the proteolytic enzyme acts upon this link.
Chain 1……….to……105 para-κ-caseinate (insoluble)
62
Chain 106…….to…….169 Glycomacropeptide (soluble)
In the above reaction, para-κ-caseinate remains in micelle with αs and β casein. Being
insoluble, it causes precipitation of casein which is known as enzymatic precipitation of
casein. The soluble glycomacropeptide part which is soluble drawn with whey during the
precipitation especially on cheese manufacturing.
Phases of enzyme precipitation reaction
Actually the above reaction takes place by two phases.
a. Phase 1 / Primary phase:
In this phase, splitting κ-casein into para-κ-caseinate and macro-peptide take place.

ΑS Β Rennet Glycomacropeptide
ΑS Β
+

Para-κ-caseinate
b. Phase 2/ Secondary phase/ Precipitation phase
In this phase, calcium bridge form where para-κ-caseinate is not present and their
synthesis occurs and finally precipitation occurs.
Ca
ΑS Β

ΑS Β

Ca
ΑΒ + Ca++ Ca Ca
ΑS Β

Ca ΑS Β

Para-κ-caseinate Calcium-caseinate (ppted)

4 Heat precipitation of casein

Pure casein is not precipitated by heat, but in fresh milk prolonged heating at high
temperatures 100oC for 12 or more hours or 120oC under pressure will cause the
precipitation of casein. On boiling fresh milk, a thin layer of finely precipitated casein,
together with other milk constituents including fat, forms a thin layer over the surface of the
milk. The application of heat to milk that slightly acid will cause the precipitation of casein.
63
When milk is heated at 130oC for several hours (1 hrs) hydrogen bond rupture and β-
dimensional structure of casein disruption causes ultimately governing the protein
denaturation and finally precipitation occurs. The cooked curd casein is less soluble and
contain more ash than the other caseins.
5.5 Enzymes in milk
An enzyme is a biological catalyst elaborated by a living cell, milk enzymes being
elaborated by the cells of the mammary tissue. The enzymes produced by bacteria in milk
can not be controlled as inactivated by high temperatures, they posses a pH of optimum
activity, and they exhibit specificity for certain substrates.
Milk contains numbers of enzymes. The native enzymes, i.e., those known to be excreted
by mammary gland, may include several present in the leukocytes, e.g., catalase. In
addition, enzymes of microbial origin may be involved. The latter may be present in
microorganisms, secreted by the organisms (e.g., proteinases and lipases), or released
after lysis. The native enzymes can be present at different locations in the milk. Many of
them are associated with the fat globule membrane. Most of the membrane originates from
the apical cell membrane, which contains several enzymes. Other enzymes are in solution,
i.e., dispersed in the serum, but some of these (e.g., lipoprotein lipase) are partly
associated with the casein micelles.
Table 5.3 Some enzymes in milk
Optimum Activity1
Name Temp Poten Where in milk? Inactivation2
pH Actual
(oC) -tial
Xanthine oxidase ~8 37 >>40 40 Fat globule membrane 7min 73oC
Sulfhydryl oxidase ~7 45 ? ? Plasma 3 min 73oC
Catalase 7 37? ? 300 Leukocytes 2 min 73oC
Lactoperoxidase 6.5 20 ? 22000 Serum 10 min 73oC
Superoxide dismutase ? ? ~2000 ? Plasma 70min 76oC
Casein micelles
Lipoprotein lipase ~9 33 3000 0.3 30 s 73oC
Fat globule membrane
Alkaline phosphatase ~9 37 500 <<500 Serum 20 s 73oC
3
Ribonuclease 7.5 37 () ? ?
Casein micelles
Plasmin 8 37 3 0.05 40 min 73oC
1
µmol.min-1.L-1
2
Heat treatment needed to reduce activity to approximately 1 %.
3
11-25 mg enzyme/kg milk.
Most of the milk enzymes seem to have no biological function in milk, even if they are
present in high concentrations (e.g., ribonuclease). Often, they do not significantly alter the

64
milk. Some milk enzymes have an antimicrobial function or play other beneficial roles. A
few of the enzymes may facilitate resorption of milk constituents into the blood if and when
milking is stopped. It presumably concerns plasma and lipoprotein lipase, which are not
very active in fresh milk through they are present in high concentrations (Table 1).
These, as well as some other enzymes, can cause spoilage of milk during storage. Some
enzymes are used for analytical purposes. Formerly, catalase was applied to detect
mastitis, but the correlation is too weak. Furthermore, particular enzymes are used to
monitor pasteurization.
5.5.1 Enzyme Activity
The properties of a solution are governed by activities rather than by concentrations, and
this certainly holds for enzymes. The maximum rate of catalysis is expressed as kcat or
turnover number for the enzyme, i.e., the number of molecules of substrate converted per
enzyme molecule per second if the substrate is in excess and the conditions for the
enzyme (pH, temperature, ionic strength, and other factors affecting enzyme activity) are
ideal. The total turnover rate is defined by Vmax = kcat [E], where [E] = enzyme
concentration. According to Michaelis and Menten, the velocity of reaction as a function of
the substrate concentration [S] is
vi = vmax [S] / (Km + [S]) 1
The Michaelis constant Km is a measure of the affinity of an enzyme for its substrate (the
lower Km, the greater the affinity). It depends on the type of substrate used and is thus a
variable next to kcat, Km equals the substrate concentration when vi = Vmax / 2; only applies
to the initial velocity of reaction vi because reaction products may inhibit the reaction
(product inhibition); moreover, the substrate concentration decreases during the reaction.
The following are the additional factors that can affect enzyme activity.
f. In addition are the above-mentioned reaction products, several substances may inhibit
the reaction products, several substances may inhibit enzyme activity, e.g., because
these substances also bind to the enzyme (competitive inhibition) or because they
affect the conformation of the enzyme molecule.
g. The above equation does not apply to so-called allosteric enzymes.
h. Many enzymes need a “cofactor”. An example is apoprotein C2, which is essential for
lipoprotein lipase action. The concentration of cofactors in milk varies.

65
 i. There may be other stimulators that inactivate an inhibitor.
j. The substrate can be inaccessible. AN example is the triglycerides, which are
screened from enzymes in milk by the fat globule membrane.
k. The enzyme can be adsorbed onto particles, thereby becoming less active. Some
enzymes (lipases, proteinases) are adsorbed onto casein micelles.
l. The enzyme may be present in a nonactive form, a so-called zymogen, and slowly be
activated. An example is plasmin, largely occurring in milk as the inactive
plasminogen.

Vmax 3

Approximate
2 zeroth-order
kinetics
Vmax/2
vi

1
Approximate first
order kinetics
0
0 2 4 [S]

Fig. 5.5.2 Effect of substrate concentration [S] on initial reaction rate vi of an enzyme
reaction, according to equation 1. Arbitrary scale.
m. The enzyme may be (slowly) inactivated. For instance, lipoprotein lipase in milk loses
activity, presumably caused by an oxidative reaction.
5.5.2 Some Milk Enzymes
1. Antibacterial Enzymes The main is lactoperoxidase. It catalyses the reaction:
H2O2 + 2HA 2H2O + 2A
Where the substrate HA can include several compounds:aromatic amines, phenols,
vitamin C, and so on. The enzyme can also catalyze oxidation of thiocynate [CNS-] by
H2O2 to an unidentified product that inhibits most bacteria. If the bacteria themselves
produce H2O2, as most lactic acid bacteria do, they are inhibited. ( In milk, H2O2
decomposition by catalase, is too to prevent this. But the thiocyanate concentration in
milk varies widely because it depends on the cyanoglucoside content of the feed.
Sometimes thiocyanate plus a little H2O2 is added to raw milk to prevent spoilage. In

66
 this way, even in a warm climate spoilage of milk can be delayed for many hours. In
milk, the action of the lactoperoxidase-hydrogen peroxide-thiocyanate system can be
enhanced by xanthine oxidase, which can form some substrates.
Lysosome is another bactericidal enzyme, it hydrolyses polysaccharides of bacterial
cell walls, eventually causing lysis of the bacteria. In cows’ milk the lysosome activity
is weak; in human milk it is much stronger.
2. Oxidoreductases
g. Xanthine oxidase Milk is the best known source of the enzyme xanthine oxidase.
An oxidase is an enzyme which catalyzes the addition of oxygen to a substance or
the removal of hydrogen from it. For the removal of hydrogen, the dehydrogenase
is sometimes used; for the donor of oxygen, the term reductance is applied. The
enzyme can reduce nitrate(trace amount in milk) to nitrite. This property is use in
the manufacture of cheeses, where nitrate is added to milk to prevent the
proliferation of the detrimental butyric acid bacteria. Nitrate inhibits these bacteria. If
nitrate has been added to the milk, nitrite is present in sufficient amounts in cheese,
though it is fairly rapidly decomposed. Cows’ milk has a relatively high xanthine
oxidase content. Most of the enzyme is associated with the fat globule membrane.
Because of this it is only partly active, but the activity is increased by such
treatments as cooling and homogenization, which may release enzyme from the
membrane.The enzyme has a molecular weight 300 000, and IEP is at pH 5.3-5.4.
It contain in milk about 160 mg/l.
h. Superoxide dismutase catalyses the dismutation ( it mean oxdation of one molecule
and simultaneous reduction of another) of superoxide anion O2- to hydrogen
peroxide and triplet oxygen according to: 2O2.-+2H+ H2O2+3O2 . Its biological
function is to protect cells from oxidative damage. In milk, the superoxide anion can
be generated by oxidations catalysed by xanthine oxidase and lactoperoxidase,
and by photoxidation of riboflavin. This enzyme may inhibit oxidation of milk
constituents. The enzyme also counteract the oxidation of lipids from this off-flavors
will be prevented. It is not inactivated by low pasteurization.
i. Sulphydryl oxidase catalyses oxidation of sulphydryl groups to disulfides, using O2
as electron acceptor: 2RSH +O2 RSSR + H2O2 - SH groups of both high-

67
 and low- molar-mass compounds are decomposed. Most of the enzyme is bound to
lipoprotein particles. Pasteurization inactivates enzymes only partially. In
pasteurized milk, the enzyme may be essential for reducing the cooked flavor
caused by –SH compounds.
3 Phosphatases Several phosphatases occur in milk. Best known phosphatase in milk
is alkaline phosphate (Alkaline phosphomonoesterase). It catalyses the hydrolysis of
phosphoric monoesters. Generally, determination of the activity of the enzyme by the ‘
phosphatase activity test’ is applied as a check low pasteurization of milk. Inactivation
of the enzyme ensures that all of the pathogenic microorganisms present in the milk
during heating have been killed; most but not all of the lactic acid bacteria and Gram-
negetive rods have also been killed. Most of the enzyme is in the membranes of the fat
globules. Accordingly, the phosphatase test is less sensitive when applied to skim milk.
The pH of the optimum activity is about 9.6. Average alkaline activity of milk to be 160
IU for normal milk and 30 units for colostrum.
Acid phosphatase (Acid phosphomonoesterate): This phosphatase catalyze the
hydrolysis of certain phosphoric esters in milk, but slowly. It is reported in milk in low
concentration. It is found in the skim milk(serum portion), and its optimum pH is about
4.0. It is quite heat resistant. It is unstable when exposed to sunlight or UV radiation
but is very heat resistant, requiring 5 min at 96oC for complete destruction. The activity
of acid phosphatase to be 72 IU for normal cow’s milk and 14 units for
colostrum.Another phosphatase can release phosphoric acid groups esterified to
casein.
4. Lipolytic Enzymes Several esterases, which can hydrolyze fatty acid esters, occurs in
milk. Some of these attack esters in solution, but the principal lipolytic enzyme of cows’
milk, i.e., lipoprotein lipase liberates fatty acids from triglycerides and diglycerides and
is only active at the oil-water-interface. It is bound largely to casein micelles. In milk,
lipolysis causes a soapy-rancid off-flavor.
A lipase is an enzyme which catalyzes the hydrolysis of fats to glycerol and fatty acids.
It cause increased acidity in canned butter. Though bitter, rancid flavors were observed
earlier in products made from homogenized milk. Raw milk became rancid as a result

68
 of homogenization and and that the lipase causing the development of objectionable
flavours could be inactivated by holding the milk at 55oC for 30 min.
Milk lipase is a mixture of several enzymes and is capable of hydrolyzing many types
of fats. Lipases are present in the plasma of fresh milk, but differentiate between
‘plasma lipase’ which remains in the plasma when the milk is cooled, and “membrane
lipase”which is irreversely adsorbed on the fat globule membrane as fresh milk is
cooled. The “plasma lipase” requires an activation treatment (homogenization or
agitation) before it produces rancidity. Whereas the ‘membrane lipase’ requires cooling
to place it in contact with the substrate. Cows late in lactation and on dry feed contain
more of this ‘membrane lipase’, and the milk is subject to spontaneous lipolysis; but
many investigators do not agree that lipolytic activity increases toward the end of
lactation.
Lipase has not yet been crystllized or obtained in pure form, and the percentage in
milk is still in doubt. The effect of heat treatment on lipase activity in milk and noted a
decrease as the temperature was increased. However, complete inactivation did not
occur at 73.9 or 85oC for 30 min.
5. Proteinases In milk at least two trypsin-like endopeptidases occurs. One of these is
the so called alkaline milk proteinase, which is identical to the plasmin of blood. Most of the
alkaline proteinase in milk is present as the inactive plasminogen. The enzyme is largely
associated with the casein micelles. Its activity in milk varies widely, partly because of a
variable ratio of plasmin to plasminogen. Usually, the activity increases with time as well as
by heating, e.g., pasteurization. Milk contains one or more promoters that catalyze the
hydrolysis of plasminogen to yield plasmin. Moreover, milk contains at least one substance
that inhibits the promoter(s). The inhibitor is inactivated by heat treatment. It also appears
that leucocytes contain a promotor, and milk by heat treatment. It also appears that
leukocytes contain a promotor, and milk of a high somatic cell count generally shows
enhanced plasmin activity.
Plasmin can hydrolyze proteins to yield large degredation products and is responsible for
production of γ-casein and protease-peptone from β-casein. The enzyme causes
proteolysis in some products, e.g., in cheese. In UHT milk products its proteolytic action
causes a bitter flavor and eventually can solubilize the casein micelles; in some cases,

69
gelatin has been obserbed. This is because the enzyme is very heat resistant. Accordingly,
appropriate UHT treatment (e.g., 140oC for 15 s) should be applied to prevent this
problems. In milk an acid milk proteinase also occurs, though with a lower activity. This
enzyme is less heat resistant than plasmin.
For whole raw milk, proteolytic activity showed a range of 4-126 µg/ 5ml with an average
of 57.0. The protease activity of milk was not serious affect in milk processing.
6. Lactoperoxidase: It catalyses the transfer of oxygen from peroxides, especially H2O2,
other substances. All milk contains peroxidase. Milk peroxidase is known as
lactoperoxidase , it is a heme protein with an iron content of about 0.07 %. Molecular
weight of about 82000. Its optimum pH is 6.8, and pasteurization temperature do not
inactivate it.
The average peroxidase activity of milk as 22,000 IU, colostrum 29200 IU.
7. Amylases: It catalyses the hydrolysis of starch to dextrin or maltose. An amylases that
catalyses the hydrolysis of central glucosidic linkases in the starch molecule, thus
producing dextrinization, is designated as α-amylase, one that catalyses saccharafication is
called β-amylase.
Cows milk contain both α, and β-amylases. Α-amylases is inactivated by heating milk at
55oC for 30 min; β-amylase withstands 65oC for 30 min without loss of activity.
α-amylase, a normal , native constituent of cow’s milk. The amylase activity of the milk
was 1029 IU (µM/min/1000 ml /37oC). Some milk contain low percentage of β-amylase.
8. Aldolase: The enzyme aldolase reversely splits fructose 1,6-diphosphate into
dihydroxyacetate phosphate and phosphoglyceric aldehyde. Present in fresh milk in the
same concentation range as in blood serum. It is present in greater concentration in cream
than in milk. It is unstable when in milk, stability is enhanced by purification. It is completely
inactivated in milk by heating at 45oC for 20 mins.
9. Ribonuclease: This enzyme hydrolyzes nucleic acid to its component nucleotides. It is
present in milk in relatively large amounts. The enzyme is quite heat stable, there being
little loss when heated to 90oC for 20 min at pH 3.5; however, under the same conditions at
pH 7, all the activity is lost. The optimum activity at pH 6.9 and 60oC. Its activity is not
reduced by pasteurization. It content in milk about 1100 µ/100ml.

70
10. Lysozome: It content of milk 13 µg/100ml. For human milk it is 40 000 µg/100ml. The
average lysosome content in brown Swiss, Guernsey, Holstein and Jersey milks were 21,
15 and 5 µg/100 ml, respectively. The stage of lactation or milk yield did not affect
lysosome secretion.
10. Carbonic anhydrase: The enzyme catalyses the hydration of CO2 and the reverse reaction,
the dehydration of carbonic acid. It is a Zink-containing protein associated with the RBC’s and is an
important enzyme in the animal body. Its value in milk is unknown.
11. Rhodonase: It catalyzes the conversion of cyanide into thiocyanate. Its activity has been
reported in cow’s milk and some what higher activity in goats and sheep milks.
12. β-galactosidase (Lactase): It catalyses the hydrolysis of lactose to glucose and galactose.
13. Other enzymes : These are transaminase and sorbitol anhydrase, phosphohexose isomerase,
sulphydryl oxidase, lactase dehydrogenase, β-glucuroxidase, thromboplastin and p-diamine
oxidase.
5.5.3 Inactivation
To inactivate enzymes, heat treatment is mostly applied. Inactivation is generally due to
denaturation (unfolding) of the enzyme molecule. The heating time and temperature relationship
for the inactivation of various enzymes in milk is given below. The inactivation by heat treatment is
of great importance because
a. Enzymes that cause spoilage can be inactivated.
b. System that inhibit bactrial growth can be inactivated. This can be either desirable or
undesirable.
c. Spoilage inhibiting enzymes (e.g., superoxide dismutase) can be inactivated.
d. The intensity of milk pasteurization can be checked, low pasteurization through alkaline
phosphatase and high pasteurization through lactoperoxidase.
The rate of inactivation by heat treatment may be closely depend on conditions like pH and the
presence or absence of substrate. Moreover, different isozyms (i.e., genetic variants of one
enzyme) that differ in heat stability may be involved. Often, they cause curved plots for the
relationship between the log of the required heating time and the temperature.

71
 t’
60
40 Ac Fig.5.10
id
ph Time(t’) and

Cat
20 Pla os
temperature (T)
Minutes

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10 in a ta
se 3 of heating milk

e
5 needed to

Cold
inactivate some

Log t’(s)
Xa
2
enzymes (i.e.,

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2
1 reduce the

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Re

40
Alk

u
activity by about

ox
tinat
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ali

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ne
era

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se
Seconds

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prevent cold
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10
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ha

5 xida op
top

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60 70 80 90 100
T (oC)

5.6 Salts in milk

Milk contains organic and inorganic salts. The concept of ‘salts’ thus is not equivalent to
‘mineral substances’. Salts are by no means equivalent to “ash” because ashing of milk
causes loss of organic acids including citrate and acetate, and because organic
phosphorus and sulfur are transferred to inorganic salts during ashing. Mass concentration
of salt can be expressed in various ways, e.g., as element (e.g., P), acid residue (PO43-),
oxide (P2O5).
5.6.1 Composition and distribution of salt among the phases
The salt composition varies, but the various components do not vary independently of
each other.
All of the salts are not dissolved, and all of the dissolved salts are not ionized. The
casein micelles contain the undissolved salts. In addition to counterions of the negatively
charged casein, the micelle contains the so called colloidal calcium phosphate, which also
contains Mg, citrate, Na, and K, as well as presumably small quantities of other ions. The
colloidal phosphate is amorphous, can vary in composition, and may have ion exchange
properties.
The distribution of phosphorus among the fractions is even more intricate. The phospha-
tases present in milk may hydrolyze phosphoric acid esters, causing the content of organic
72
(esterified) phosphate to decrease and that of inorganic phosphate to increase. Milk
contains sulfur, again in several forms. Not more than 10 % of the sulfur in milk, amounting
to about 36 mg/100g milk, is present as inorganic sulphate, whereas the remainder is
present in protein.
The dissolved salts affect various milk properties, e.g., protein stability. These salts are
only present in the serum. Note that the solute content in the serum ~ 1.09 times the
content in milk; in the plasma it ~ 1.04 time the content in milk.
The composition of the salt solution of milk is best determined from a dialyzate of milk.
The solution can be obtained by dialysis of water against a large excess of milk, as it is in
equilibrium with the colloidal particles and dissolved proteins in milk. It does not reflect
precisely the concentration of the various dissolved salts in the water. To begin with, part of
the water in milk is not available as a solvent. Second, the proteins have a “diffuse double
layer,” so that they are accomplished by counterions. Of the cations associated with the
casein micelles, all of the Na and K, most of Mg, and as far smaller portion of the Ca are
present as counterions. The serum proteins also take along some counterions.
Table 5.4 The most important salts in milk and their distribution between serum and casein
micelles.
Molar Fraction In micelles
Range Average
Compound mass present (mmol/g
mmol/kg (mg/100kg)
Da in serum dry casein
Cations
Na 23 17-28 48 0.95 0.04
K 39.1 31-43 143 0.94 0.08
Ca 40.1 26-32 117 0.32 0.77
Mg 24.3 4-6 11 0.66 0.06
Amines - ~ 1.3 - ~1 -
Anions
Cl 35.5 22-34 110 1 -
CO3 60.0 ~2 10 ~1? -
SO4 96.1 ~1 10 1 -
PO4* 95.0 19-23 203 0.53 0.39
Citrate 189 7-11 173 0.92 0.03
Carboxylic acids - 1-4 - ~1? -
Phosphoric esters** - 2-4 - 1 -
*Inorganic only
**Soluble

73
5.6.2 Trace elements
Among the trace elements found in milk the Zink is found in the highest concentration ~ 3
mg/kg of milk, while others are present in lower concentrations. The others are Al, As, Ba,
B, Br, Cd, Cs, Cr, Co, F, I, Fe, Pb, Li, Mn, Hg, Mo, Ni, Rb, Se, Si, Ag, Sr, Sn, Ti, V.
These are natural components in milk. Concentrations of some of these elements in milk
can be increased by increasing their level in the feeding ration of the cow. Consequently,
their concentration in milk can vary widely. For instance, Se can range from 4-1200 µg/kg
of milk. Concentration of other metals are not affected by cattle feed, except on shortage
(e.g., Cu, Fe), or if extreme levels in the ration cause poisoning of the cow (e.g., Pb).
Finally, some elements can enter in milk by contamination following milking which can
considerably increase the concentration. For instance, the natural Cu content of milk is
about 20 µg/kg (colostrums contains more); contact of milk with bronze parts in milking
utensils or with copper pipes can increase its Cu content easily to 1 mg.kg-1. Iron is
already entering the milk due to contamination.
Parts of the elements are associated with protein, e.g., some heavy metals with lacto-
ferrin, whereas most of other elements are dissolved. About 10 % of cupper and nearly half
of iron is associated with the fat globules membrane. Zink is predominantly in the colloidal
phosphate.
Some of trace elements are important from a nutritional point of view, whereas other
elements (e.g., Cd, Hg, and Pb) are toxic, though they are hardly ever detected in milk in
too high a concentration.
For dairy manufacturer, copper is of great importance due to its catalytic action on fat
autoxidation. “ Natural copper” in milk does not promote oxidation, or does so hardly at all,
but “added copper” often does, even when added in minute amounts.
Manganese is of importance in the metabolism of some lactic acid bacteria, especially
for fermentation of citrate, and in some milks the Mn content too low for production of
diacetyl by leuconostocs.
5.6.3 Properties of salt solution
Only the dissolved salts are considered here, i.e., roughly the salts in the milk serum.
There is a extensive association of ions so the composition given in (Table 5.6.1) above
does not follow simply.

74
 Some of the acid and bases in milk (phosphoric acid, carbonic acid, secondary amines,
etc) are not fully ionized at milk pH. But some of the salts may also be partly associated.
This primarily concerns binding of Ca++ and Mg++ to citrate3 - and to HPO32-.
Consequently, the concentration of Ca++ ions is much lower than that of dissolved calcium,
because Ca-citrate- and CaHPO4 are present in appreciable amounts.
Also other salts, e.g., the chlorides of Na, K, Ca, and Mg, are not completely ionized.
The appropriate ion composition can be calculated from the atomic composition of the milk
salt solution and the association constants.
The total ionic strength in milk is about 0.5 A.V-1.M-1 (variation~0.4- 0.55) at 25oC. This
roughly corresponds to the conductivity of 0.25 % (w/w) NaCl in water.
5.6.4 Colligative properties
Freezing point depression, osmotic pressure, (1-aW), etc., are called the colligative
properties. Nonionic solutes as well as ions determine the magnitude of these
properties.The freezing point depression of 0.53 k (equivalents to ~ 0.85 % of NaCl
solution) is calculated from the total concentration of dissolved substances (0.28 mol/L
solution), the molar freezing point depression of water (1.86 k for 1 mol in 1 kg of water),
average osmotic coefficient (~0.93 for ionic species and 1.00 for natural molecules), and
from the fact that 1 L of milk serum contains about 950 g of water. The calculated value
corresponds satisfactorily to the observed freezing point depression of about 0.53 k, which
varies among milk samples from ~0.515 to 0.55 k.
The freezing point depression of milk is very constant (relative standard deviation between
individual milking about 1 %) in as much as it is proportional to its osmotic pressure, which
is essentially equal to that of blood, which in turn is kept almost constant. The osmotic
press-ure of milk is about 700 kPa (7 bar ) at 20oC.
Lactose accounts for over half of that value, complemented by K+, Na+,and Cl-.
Furthermore, a boiling point elevation of milk 0.15 k calculated from the molar
concentration, and a water activity of 0.993.
5.6.5 Colloidal calcium phosphate
Part of the salts in milk is present in or on the casein micelles, i.e., in colloidal particles.
This undissolved salt is called the colloidal or micellar, calcium phosphate, though it
includes other components, i.e., K, Na, Mg and citrate. The total amount is about 7 g / 100

75
g dry casein. Part of it is to be considered as counterions. This is because the casein is
negatively charged at the milk pH and is thus associated with positive counterions. This
presumably involves the K, the Na, most or all of the Mg, and part of the calcium in the
micelles. The rest, which is mainly calcium and phosphate together with a little Mg and
citrate, is present in a different state. Milk is supersaturated with respect to calcium
phosphate and accordingly, a large part of it is undissolved. The calcium phosphate in the
caseins micelles consists of small non-crystalline regions and is, moreover, bound to the
protein.
The molar ratio Ca/inorganic phosphorus in the micelles is high. Even if the part of the
calcium present as counterions is subtracted, a ratio of about 1.5 remains for the calcium
phosphate, i.e., as in tricalcium phosphate. That seems astonizing because we would
except a diphosphate (Ca/P =1) at most, due to the pK2 of phosphoric acid being about 7.
Therefore, the phosphate esterified to serine residues of casein, i.e., the organic
phosphate, is believed to participate in the colloidal phosphate as a result of which a ratio
of ~ 1 would be found. However, the colloidal phosphate should not be seen as one of
many known types of calcium phosphate. Moreover, it has a variable composition that
depends on the ion atmosphere. For instance, as stated above, it contains Mg and citrate
as well as traces of several ions, primarily Zn. In other words, the colloidal phosphate can
be considered to have ion exchange properties.
5.6.7 Changes in salts
The salts of milk are in dynamic equilibrium: among themselves in solution, between solut-
ion and colloidal phosphate, between solution and proteins. Changing external conditions
of milk may cause alterations in equilibria. To be sure, there is no true equilibrium,
especially not of calcium phosphates, but local or pseudo equilibria exist.
Milk is saturated with respect to CaHPO4 (solubility in milk is about 1.8 mmol/L, that of
Ca3(PO4)2 about 0.06). Furthermore, a small part of the citrate in milk is undissolved, as in
table 1. Milk is not saturated with respect to other salts (e.g., solubility of MgHPO4 in water
is ~12 mmol/L). Milk has a buffering capacity for some ions, primarily Ca2+. This is mainly
caused by the presence of undissolved colloidal phosphate, which can vary not only in
quantity but also in composition, as a result of ion exchange. Imposing changes in ionic

76
composition by temperature, pH, etc., has therefore a different effect on the salt solution of
milk, where exchange with the micellar salts may occur, and that of whey or ultr-afiltrate.
During storage of milk, some changes may occur. The milk loses CO2; the original content
in the udder is roughly twice. Enzyme action on dissolved phosphoric esters causes a
decrease in pH and in [Ca2+], and an increase in the amount of colloidal phosphate.
Lipolysis yields free fatty acids that decrease the pH and bind some Ca2+ ions. The calcium
salts of fatty acids are poorly soluble.
1. Acidity The pH may change as a result of additions, by concentrating or heating the
milk, and so forth. Microbial fermentation of lactose to lactic acid is of great importance.
The ensuring drop in pH causes a partial dissolution of the colloidal phosphate, and a
decrease of the negative charge of the proteins, which goes along with a decrease in the
association with counterions.
A decrease in pH reduces the dissociation of weak acids, increases the [Ca2+], and
increases the ionic strength, presumably by more than corresponding to the amount of
lactic acid formed. The acid production causes a decrease of the freezing point by about 2
m K per mmol lactic acid produced, and an increase of the electrical conductivity by~ 4
mAV-1.m-1 per mmol lactic acid. Several lactic acid bacteria also breakdown citrate, and
this would enhance the increase in [Ca2+]. On the other hand, lactate associates to some
extent with Ca2+, and increase in [Ca2+] on lactic fermentation will thus be less than would
follow from the drop in pH formed.
2. Temperature treatment
The dissociation constants are temperature dependent. During heating, the most important
change is that dissolved calcium and phosphate become partly insoluble and largely
associate with the calcium micelles. The additional colloidal phosphate formed has a molar
ratio Ca / P ≈ 1. The reaction is roughly as follows:
Ca2++ H2PO4-- CaHPO4 + H+
This implies that the milk becomes more acidic. (The drop in pH partly counterbalances the
insolubilization of calcium and phosphates). The reactions are slow and occur especially in
a fairly narrow temperature range. Below 60oC, changes are small, whereas above 80oC
any further increase in temperature has little effect. The reactions reverse at room
temperature, though very slowly. At low temperature, the reverse occurs, after 24 h at 3oC,

77
dissolved Ca is increased by about 7 %, dissolved phosphate by about 4 %, and Ca++
concentration is also increased. The magnitude of all of these changes may vary.
3. Concentration
Concentration of milk by evaporation of water causes several changes, but it should be
taken into account that heating is usually involved as well. The pH drops by about 0.3 unit
for 2:1 concentration (i.e., concentrating to twice the original dry matter content) and by
about 0.5 unit for 3:1 concentration. Again, the main cause of the changes is formation of
additional calcium phosphate in the casein in the casein micelle. The Ca2+concentration
does not contain appreciably.
The fractions of dissolve citrate, phosphate, and calcium decrease, but less than
proportiona-lly to concentration; for instance, dissolved calcium decreases from 40 % to 30
% for 2.5:1 concentration. This can be attributed partly to the pH decrease, partly to the
increase of the ionic strength, but otherwise the changes are poorely understood.
4. Calcium ion activity
The Ca2+ activity (aCa2+) in milk is an important variable, especially for the stability of
casein micelles. It differs markedly from the content of dissolved calcium. Because of this,
direct determination of aCa2+ is essential. This can be achieved by using a calcium ion-
selective electrode, in much the same way as the pH is measured.
Addition of sugar, e.g., sucrose, to milk (as is applied in the manufacture of sweetened
condensed milk, ice cream mix, and other milk products) significantly increases aCa2+
expressed in mmol/kg water). For example, the activity coefficient of Ca2+ increases from
0.40 to 0.46 by the addition of 150 g sucrose to 1 L milk. Different milks vary widely in
aCa2+, from 0.6 to 1.6 mmol.L-1. The variation usually correlates significantly with pH, i.e.,
the higher the aH+ (the lower the pH), the higher the aCa2+.
5.7 Other components
Several minor components are also present, which are equally important in all respects.
The no of compounds detected in milk is large and will certainly increase with further
research. The other components are partially by enzymatic and microbial changes, or by
changes caused by manufacturing processes. The natural components may considerably
increase in concentration due to contamination, etc.

78
5.7.1 Natural components of milk
These are entered in milk directly from the blood or are intermediate products of the
metabolic processes in the secretory cell.
a. Organic acids
These are citric acids, low molar mass fatty acids, small quantities of organic acids (e.g.,
trace amounts of lactic and pyruvic acids) occurs in milk serum. Bacterial action may
greatly increase the concentration of such acids.
b. Carbohydrates
In addition of lactose, milk contains traces of glucose, galactose and oligosaccharides. It
contains several sugar derivatives, again in trace quantities: hexose monophosphates,
hexos-amines, and many others. A large part of these is associated with other compounds,
e.g., proteins (κ-casein, fat globule membrane proteins) and cerebrosides.
c. Nitrogeneous compounds
About 5 % of total nitrogen in milk is non-protein nitrogen (NPN). The compounds are partly
intermediate products of the protein metabolism of the animal (e.g., ammonia, urea,
creatine, creatinine, uric acid). Most of the amino acids as well as their derivatives (amines,
serine, phosphoric acid) are also found in trace amounts free in solution. Milk also contains
small peptides. These compounds may be essential nutrients for some bacteria.
d. Vitamins
Table 5.5 Shows the vitamins content in fresh milk.
Vitamins mg per Range
100 ml
Vitamin A 159.000 136-176
Carotenoids 0.030 0.025-0.060
Vitamin D 2.210 0-10.9
Vitamin E 0.100 0.02-0.18
Vitamin K 0.00467 0.0-0.0160
Vitamin C 2.09 1.57-2.75
Biotin 0.003 0.0012-0.0060
Choline 13.7 4.3-28.5
Folacin 0.0059 0.0038-0.0090
Myo-inositol(total) 11.0 6.0-18.0
Niacin 0.09 0.03-0.20
Pantothenic acid 0.34 0.26-0.49
Riboflavin 0.17 0.08-0.26
Thiamine 0.04 0.02-0.08
79
 Vitamin B-6 0.06 0.02-0.08
Vitamin B-12 0.00042 0.00024-0.00074
p-aminobenzoic acid 0.01 0.004-0.015

e. Phosphate esters For examples hexose monophosphates and glycerol phosphate.

f. Ribonucleic acids and their degredation products, e.g., phosphate esters and organic
bases. Furhermore, orotic acid typically occurs in milk of ruminant animals; it is a growth
factor for Lactobacillus delbrueckii ssp. bulgaricus.
g. Sulphuric acid esters only indoxyl sulphate has been found in milk.
h. Carbonyl compounds An example is acetone; more of it occurs if the cow suffers from
ketosis. Fat soluble aldehydes and ketones.
i. Gases
In milk the amount of nitrogen is about 16 mg.kg-1, that of oxygen about 6 mg.kg-1, or
about 1.3% and 0.4 % by volume, respectively. Milk is almost saturated with respect to air;
however, it contains relatively far more carbon dioxide though in the form of bicarbonate.
The O2 content of milk while in the udder is lower, i.e., about 1.5 mg. kg-1.
j. Enzymes (in separate chapter).
k. Hormones In trace quantities in milk. Examples are prolactin, somatotropin, and steroids.
5.7.2 Contaminants
They have potential toxicity or mutagenicity. Some of them may be allergic compounds
e.g., antibiotics used to treat several diseases of cow. Several pathways to enter into the
milk.
a. Illness of the cow
If the cow will suffer from sever mastitis there is a chance of entering blood compounds and
somatic cell to the milk.
b. Pharmaceuticals (drugs)
The drugs used for the treatment of mastitis, and also from blood may pass to milk can be
taken as contaminants.
c. Feed Many compounds can gain entrance into the milk through the feed, through the
cow may act as a filter, sometimes, substances are partly broken down first.
• Chlorinated hydrocarbons such as several pesticides (DDT, aldrin, dieldrin); PCOS
(polychlorinated biphenyls), which are widely used in materials; dioxins, which are
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 potentially harmful to the consumer even in extremely low concentrations. Some of
these components are toxic or carcinogenic, and occasionally too high a level (i.e.,
higher than the standard, which normally has a safety factor of, say, 100) has been
detected, for instance, in milk from cows fed large quantities of vegetables sprayed
with pesticides. These substances are lipophillic and hence tend to accumulate in the
fat.
• Other pesticides, herbicides, and fungicides- like phosphoric esters and carbamates.
Most of these components are broken down by the cow.
• Mycotoxins – It may originate from molds growing on concentrates fed to cows.
• Heavy metals – Pb, Hg, and Cd, are especially suspect, but toxic levels have never
been found in milk. Most heavy metals do not gain entrance into the milk because the
cow acts as a filter, unless extremely high quantities are fed.
5.7.3 The following substances / compounds that may enter the milk during
milking and milk handling.
• Pesticides It may gain entrance through air e.g., when aerosols with insecticides are
used.
• Plasticizers It may gain entrance from plastics or antioxidants from rubber (teat cup
linings).
• Metal ions- e.g., Cu. It may cause off-flavors via autoxidation.
• Cleaning agents and disinfectants They may cause off-flavors and decreased activity
of starters.
• Substances added on purpose – Sometimes disinfectants are added to milk to arrive at
a low colony count. This is, of course, adulteration. Active chlorine may be determined
to detect such adulteration. Addition of water is best checked by determining the
freezing point.
5.7.4 Radionuclides
Radioactive isotopes of several elements are always present in milk but in minute
quantities. It especially concern 40K. It feed or drinking water contaminated after
radioactive fallout is ingested by the cow, part of the radionuclides will be secreted in the
milk, despite the fact that cow acts as a filter. For example, of the radioactive Sr ingested
only a small part enters the milk, of 131I much more.
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Table 5.6 Most important radionuclides that can occur in milk.
Radio- Physical Biological
Location in milk
nuclides half life* half life* *
89Sr 52 days ~ 50 years > 80% in casein micelles, the rest in
serum.
90Sr 28 years ~ 50 years > 80% in casein micelles, the rest in
serum.
131I 8 days ~ 100 Serum (~2 % in the fat)
days
137Cs 33 years ~30 days Serum

* The physical half life refers to the period needed to reduce the radioactive emission by
an isotope to half of its original level.
* * The biological half life refers to the time it takes until half of the amounts of a compound
injested by the body has been excreted.
Strontium is distributed in milk is much the same way as Ca, but because SrHPO4 is very
poorly soluble, by far the greater part of the Sr in milk is the colloidal phosphate. Cs behave
like K+ and Na+. Most iodine is found as dissolved iodine.
5.8 Flavor compounds
The main flavor compounds in milk are lactose and the dissolved salts, which cause a
sweet and salty taste, respectively. The sweet taste prevails, whereas the salty taste, is
prevalent if the Cl/lactose ratio is high, as in mastitis milk. The fat globules must also
contains flavor compounds since skim milk and milk differ considerably in flavor, these are
responsible for the creamy or “rich” flavor, though milk with an increased fat-free dry matter
content also has enhanced “richness”. Other compounds including dimethyl sulphide,
diacetyl, 2-methyl butanol-1, and some aldehydes, are responsible for the characteristic
flavor of fresh milk.
Fresh milk may have off-flavors originating from the feed. The compounds causing the off-
flavors enter the milk through the cow, the air, and sometimes through both pathways.
Examples are clover and garlic flavors. If the cow suffers from ketosis, such as that caused
by feed deficient in protein, increased concentrations of ketones (including acetone) are
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found in the milk. Consequently, the milk exhibits a typical cowy flavor. Vacuum heating
may remove part of such flavor compounds if they are hydrophilic. Removal of the many
fat-soluble compounds is much more difficult. Hay feeding leads to cumarin in milk; this
flavor compound is not always undesirable.
Spoilage of milk, especially microbial spoilage, may produce flavor defects; the defects are
referred to as acid; unclean, fruity, ester, malty/burnt, phenolic, bitter, rancid flavors etc.
Enzymatic spoilage includes lipolysis. Autoxidation of fat, as caused by catalytic action of
cupper, usually leads to a tallowy flavor. In milk, a cardboard flavor also occurs, which
results from autoxidation of phospholipids; it can be observed in skim milk as well. The
phospholipids in the plasma appear to oxidize more readily. In sour-cream buttermilk, this
may readily lead to a metallic flavor if the defect is weak and to a sharp (pungent) flavor if
the defect is strong.
Flavor defects in milk can also be included by light. The tallowy flavor developed slowly
also result from autoxidation catalyzed by light. But on exposure to light, additional flavor
compounds can also develop in milk. Development of this “sunlight flavor” requires ribo-
flavin (vitamin B2). Oxidation of free methionine yields methional (CH3.S.CH2.CH2. CHO)
whereas free -SH compounds are formed from protein-associated sulfur-containing amino
acids. Presumably, all of these compounds together cause the sunlight flavor.
Moderate heat treatment of milk (say, 75oC for 20s) causes the characteristic raw milk
flavor to disappear so that a fairly flat flavor results. More intense heat treatment, e.g., 80-
100oC for 20 s result in ‘cooked flavor’, caused mainly by H2S. The preponderant
“sterilization flavor” of high treated milk (e.g., 115oC for 10 min) mainly results from maltol,
furanone compounds (formed from lactose), and aliphatic methyl ketones and lactones
(formed from lipids).
5.9 Pigments
Milk contains fat soluble and water soluble pigments. Which are largly the carotenes and
riboflavin, called earlier lactoflavin. The yellow appearance of milk fat is caaused by the fat
soluble carotene . Water soluble riboflavin produces a slight yellow or greenish tint in skim
milk which becomes a distinct green fluoroscence in whey after removal of the light
reflecting components in cheese making.

83
The white or “milky” appearance of cow’s milk is caused by the scattering of reflected light
by the fat globules, the colloidal calcium caseinate ,and the colloidal calcium phosphate in
the milk. Dispersions of each of these to that occuring in milk , and , in the case the fat, with
the particles the size of the globules in milk.

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Chapter 6

Colloidal particles of milk

6.1 Fat globules

All of the fat globules in milk are in separate small globules. Milk is an oil-in-water emulsion.
6.1.1 Properties of fat globules
i. Distribution of size: The milk fat globules vary in diameter varying from 0.1 to 15 µm.
Several dispersion properties of fat are affected by its size distribution. One gram fat
contains ~ 4×1011globules, of which 1011globules are > 1 µm in diameter.
ii. Fat surface layers: Each fat globules of milk is surrounded by a surface layer or
membrane. The functions of layers are to prevent the fat globules from being
coalescence. Its composition is completely different from that of milk fat or milk plasma,
and is like that of a cell membrane, from which the fat globule membrane largely
derives.
The composition of the membrane proteins is complicated; as there are at least 10
major species and several minor compounds are also found. They are predominantly
glycoproteins, specific for membranes, and include butyrophillin, which appears to be
specific for milk fat globules.
Several of the membrane proteins are enzymes. Alkaline phosphatase and xanthine
oxidase are of special importance; these enzymes make up a substantial part of the
membrane proteins. The membrane is often said to contain many high melting
triglycerides, but that is poor concept. The presence of monoglycerides and free fatty
acids in the membrane has been rather firmly established. The membrane structure is
poorly established so difficult to understand.
The original structure of the membrane is presumably a lipid monolayer adsorbed from
the cytoplasm surrounded by a layer of proteins and, on top of this, a lipid bilayer
interspersed with proteins, some of which protrude into the milk plasma. However, much
of this structure is lost during with and after milk secretion and the membrane shows
considerable variation from place to place.
The average layer thickness is some 15 nm but varies from about 10 to 20 nm. The
globules have a negative charge. The interfacial tension is 1-1.5 mN. m-1.
85
 The fat globule membrane is also found in skim milk, where it often amounts to about one –
third of the total amount of membrane material present in milk. Not all of this originates from the
fat globules but a part does, because treatment of milk causes the fat globules causes a
decrease of their surface area, which leads to release of membrane if they came into contact
with air. Cooling leads to a migration of membrane material.
Coalescence of fat globules causes a decrease of their surface area, which leads to release
of membrane material. Fat globules also lose part of their membrane if they come into contact
with air. Cooling leads to a migration of membrane material to milk plasma (the change is
irreversible) about 20 % of the phospholipids as well as some protein, xanthine oxidase, and
cupper are released. By contrast, cooling causes adsorption of other proteins (cryoglobulins)
onto the fat globules; but this process is reversible.
Releasing part of the membrane from the surface area of a fat globule causes surface active
substances (mainly protein) to adsorb from the plasma onto the denuded fat-water interface.
This may happen when air is beaten in. Alternatively, increasing the fat surface area by
reducing average globule size creates an uncovered interface, which subsequently acquires a
coat of plasma proteins. This especially happens during homogenization.
iii. Crystallization: Crystallization of fat in fat globules differs from that of fat in bulk. Supercooling
must be deeper to induce crystallization. The crystals in a fat globule cannot grow larger than
the globule diameter. The arrangement of the crystals may also be different from that of fat in
bulk. If there are sufficient crystals in a globules they can flocculate into a network that provides
a certain rigidity to the globule. Sometimes, especially after churning, crystals tend to be sited
in the fat water interface and orient tangentially; this causes a bright layer in polarized light. As
crystallization proceeds (especially by cooling), the tangentially oriented crystals may grow into
a solid layer. Crystallization of fat in the globules is of great importance in their coalescence
stability.
iv. Differences among fat globules: Differences among fat globules refer especially to their size.
Differences in size associated with variations in composition. For eg., the phospholipids content
of the fat globules will be almost inversely proportional to volume surface average diameter.
However, globules of the same size also show variations in compositions, especially with
respect to triglycerides. There are considerable variations in composition among globules in
one milking of the cow; for eg; the final melting point of the globules in such milk can vary up to
10 k. Membrane composition of individual fat globules can also vary, but quantitative data are
not available.

86
6.1.2 Emulsion stability
In many products a stable emulsion should be needed. The emulsion should not be changed on
standing &/or during treatment. But, in some cases moderate instability is desired during some
treatments (e.g., in the whipping of cream and the freezing of ice cream). In churning operation the
emulsion should be break.
6.1.2.1 Types of Instability
The various physical changes may occur simultaneously (Fig 6.1). Some changes (e.g.,
creaming) always occur, though possibly slowly. In principle, flocculation and coalescence
can occur spontaneously, but often the activation energy is high, so that these processes
can be retarded or virtually prevented.

Breaking or
deemulsification

Rapid creaming

Coalescence
COARSER

Rapid creaming

Flocculation Coalescence
enhanced
Slow creaming
Milk

disruption
FINE

SEGREGATION

FIG. 6.1 Types of physical changes in oil-in-water emulsions.

Fat globules can aggregate in various ways. The three types of aggregates are as follows.
a. In floccules, attractive forces between globules are weak, so stirring disrupts the

87
floccules. Flocculation does not happen normally with milk fat globules because
electrostatic and steric repulsion prevents it. Milk fat globules do not flocculate even at their
IEPH (the iso-electric pH of “washed” milk fat globules is 3.7). Some of the glycoproteins in
the membrane cause sufficient steric repulsion.
Flocule
b. In clusters, two globules share part of the membrane material, generally micellar casein.
Examples are so called homogenization clusters and heat coagulated fat globules. Clusters
usually cannot be disrupted by stirring.

cluster
c.In granules, fat touches fat. Aggregation to granules can only occur if the fat globules
contain a network of fat crystals, giving the globules a certain rigidity. Granules usually
cannot be disrupted.
granules

6.1.2.2 Coalescence
Coalescence of fat globules may have several consequences:
I. It may lead to rapid creaming and to the formation of fat lenses or butter granules in milk
products.
II. It may cause cream to become more viscous, or a fairly solid cream plug to be formed in
a bottle of milk.
III. It permits the isolation of the fat from milk products, as in churning.
IV. It is desirable, when controlled, during the whipping of cream and the freezing of ice
cream for the acquirement of desirable texture and stability.
V. It leads to transfer of fat globule membrane substances (e.g. phospholipids,
glycoproteins, xanthine oxidase) to the milk plasma because of the decrease in total
surface area of the globules.
Milk fat globules should thus be stable to coalescence during handling and storage, but
unstable under other conditions. These requirements can usually be met. Coalescence is
the flowing together of two emulsion droplets into one. Its ultimate cause is the rupture of
the rupture of the film of the continuous phase (in this case milk plasma) between two

88
droplets that are close to each other; the thickness of a film that can rupture is generally of
the order of 10 nm. Coalescence stability should thus be treated in terms of the stability of
such a film on the basis of the interfacial tension, γ, between the oil water phases and of the
interaction energy between the droplets as a function of film thickness. These factors
determine whether any disturbance of the film (i.e. a local fluctuation in thickness) will be
damped-out or enhanced. In the latter case, the film ruptures. It turn out that the probability
of rupture is higher as the size of the film is larger (hence globule diameter larger), γ is
smaller and the droplets can come closer together (hence net repulsion is lower). The
theory is, however, insufficient. Generally, if it predicts instability the emulsion is unstable,
but if it predicts stability the emulsion may still show coalescence.
The presence of crystals (solid fat) in the globules strongly affects coalescence. If all of the
fat is solid, coalescence is definitely impossible; solid particles can flocculate but not
coalesce. If part of the fat is solid, the globules may show partial coalescence; irregular
clumps or granules are formed that keep their shape because of the crystals (that form a
network), while liquid fat act as a sticking agent – subsequent melting of the crystals causes
complete coalescence. The presence of crystals actually promotes coalescence if some
crystals are located in the oil-water interface; presumably such crystals may pierce the film
between globules. Streaming in the liquid (e.g. caused by agitation) may promote (partial)
coalescence, but can also cause disruption of globules, particularly of large globules.
Consequently, coalescence during agitation may never lead to visible oil separation, since
the larger droplets formed are disrupted again. But if part of the fat is solid, partial
coalescence is possible and disruption of the granules is mostly impossible; consequently, if
the fat is partly solid, rapid granule formation may be observed in an agitated cream, while
no visible change occurs when the fat is either fully solid or fully liquid.
Partial coalescence
When two emulsion droplets are close together and the film between them has been
decreased to a few nm, the film can be ruptured. Consequently, the globules can fuse or
coalesce into one droplet. But the globules usually have a surface layer that causes
sufficient repulsion between them to prohibit close approach, hence coalescence. The
globules may contain fat crystals of which some may stick somewhat out of the globule. If
so, such a protruding crystal may pierce the film between two approaching globules,

89
usually leading to a commencing coalescence. A complete coalescence cannot occur
because the crystals in the globules present this, and a granule is formed. The process
therefore is called partial coalescence.
True coalescence is rarely an important process in milk products. This is because of the
high stability of milk fat globules, though coalescence may occur in unhomogenized cream
at sterilization temperature. Partial coalescence, however, can easily occur, at least if part
of the fat is solid and especially during vigorous flow of the product.
Partial coalescence (formation of granules, clumping) differs definitely from true
coalescence in its effects. In the latter case, small fat globules transform into larger ones, in
the former into large irregularly shaped granules (“butter grains”) or even a continuous
network as in whipped cream. We also may envision a granule of two fat globules as a
collection of separate, rigid globules held together by a “neck” of liquid fat between them.
Often crystals will be present in these necks, and sometimes the original fat globules can
no longer be distinguished.
Increasing the temperature and thereby melting the fat crystals will cause coalescence of
the formed granules into (large) droplets; consequently, the oil-water interface decreases
and substances of the fat globule membrane (e.g., phospholipids) are released into the
plasma. During partial coalescence release of membrane components may also occur,
although to a lesser degree. Often the terms “free fat“, or even “uncovered fat” are used to
describe occurrence of visible or invisible clumping or coalescence. Uncovered fat as such
cannot exist in milk because there is a large excess of surface active material present in
milk plasma. Fat globules or granules will immediately (i.e., within 0.1 s) acquire this
material at their damaged spots. Incipient granule formation can only be detected by
examining whether the average particle diameter has increased.
The main factors affecting the rate of (partial) coalescence are as follows.
a. Streaming or agitation
The main variable is the velocity gradient or shear rate, S, although the type of flow is also
important. For liquid milk fat globules (at 40oC), laminar flow at S = 102 to 103 s-1 causes
partial coalescence at such a rate that ~10-7of the calculated number of encounters
between globules lead to coalescence; the initial coalescence rate first order kinetics with
time, but soon levels off, presumably because of simultaneous disruption. Liquid fat

90
globules show no significant coalescence in milk or cream at rest. If part of the fat is solid,
coalescence during agitation is more pronounced. Turbulence causes even faster
coalescence. The presence of birefringent outer layers in the globules appears to be
necessary for liquid coalescence, and the deformation of droplets, even if very slight, may
enhance the genesis of such globules.
b. Beating: of air is again a way of stirring; moreover enhances the fat clumping in another
way (see section 6.1.3).
c. Fat content The higher the fat content, the higher the rate of coalescence during
agitation. The dependence of the coalescence rate on fat content is very strong, much
stronger than the quadratic relation that would be expected from the effect on encounter
frequency. Presumably, the fat globules are pressed closer together during agitation in a
high-fat cream.
d. Proportion of solid fat: It is crucial. If there are no crystals, partial coalescence cannot
occur. If the globules contain too much solid fat (e.g., after keeping them for same hours
at < 5oC), the residual liquid fat is retained in the pores of the crystal network, leaving no
sticking agent to hold the globules together.
e. Freezing: Freezing of cream causes considerable partial coalescence, presumably
because the ice crystals damage the fat globules.
f. Creaming: Creaming, as such, causes little or no coalescence of liquid globules.
However, if the fat globules are closely packed in the cream layer (thus in the absence
of agglutination) changes in crystallization may cause partial coalescence and thus the
formation of a cream plug.
g. Fat globule size: The smaller the globules, the better their stability to coalescence. A
milk or cream with small globules is more stable than systems with large globules
follows from the greater number of coalescence events that must take place for a visible
change to occur, but the rate constant for coalescence is also smaller, often much
smaller for smaller globules. Consequently, homogenized milk products are usually very
stable to coalescence, despite their modified surface layers.
h. Surface layer or membrane of the fat globules: Undoubtedly, the properties of the
surface layer considerably affect the coalescence stability. The composition of the
surface layer determines the interaction energy, and thereby affects stability. Moreover,

91
 clear correlations between surface-rheological properties and coalescence stability of
emulsion droplets with widely varying surface layers have been established, but not
explained. Partial degradation of membrane phospholipids by added or bacterial
enzymes leads to coalescence.
i. Temperature: It primarily affects the proportion of solid fat and thereby partial
coalescence. Pre-cooling may be needed to obtain solid fat in the globules and thus
may affect coalescence but it also promotes coalescence by some other mechanism,
perhaps via changes induced in the globule membrane. A special case is the
‘rebodying’ of cream; cream is cooled to ~4oC, kept for ~2 h, warmed to ~30oC and
cooled again. This causes the (apparent) viscosity of the cream to increase greatly; the
main cause is a limited partial coalescence.
If the fat is liquid, coalescence rate (during agitation), is faster at higher temperatures. In
some types of heat exchanger, considerable coalescence may occur in cream.
Size, orientation, and network formation of the crystals in the globules will have a
significant effect on the tendency to form granules, e.g., is re-bodying.
6.1.2.3 Disruption
The breaking up of fat globules into smaller ones does not happen spontaneously. Beating
in of air, intense turbulence as applied in a high-pressure homogenizer or a very high
velocity gradient all may cause disruption. The smaller the fat globules, the more difficult
the disruption.
6.1.3 Interaction with air bubbles
Skim milk foams readily, especially at low temperature. The foam is most stable at a
somewhat higher temperature, say, 40oC. Proteins, above all casein, stabilize the foam
lamellae. Milk fat depresses foaming tendency of separated milk to less than half.
Apparently, the milk fat globules affect the foam lamellae.
When a milk fat globules makes contact with a newly formed air-water interface, membrane
material of the droplet and thereafter part of its contents will spread over the interface. This
happens as long as no other surface active substances especially proteins, have been
adsorbed onto the interface, globules can become attached to the air-water interface, and
thus to an air bubble. In this way, collection of globules in foam can occur; this process may
be called flotation.

92
A fat globule can make contact with an air bubble having an adsorption layer or it can be
caught between two such bubbles i.e., in a foam lamella. In these cases, electrostatic and
steric repulsion between globule and air –water surface layer usually prevents the globule
making contact with the air in the bubble. However, globules containing fat crystals may
pierce this layer and make contact with air, causing membrane material and fat to spread
over the interface. The spreading usually causes rupture of the foam lamella. Small fat
globules as well as fully liquid globules show fewer tendencies to make foam unstable
because these pierce the adsorption layer of the air bubble less readily.
Rapid beating in of air in milk or cream (as in churning or whipping) causes new air –water
interface to be continually formed, and fat may spread over the interface. If the fat is fully
liquid, the subsequent breaking up of air bubble covered with fat causes disruption of the
fat. Churning warm milk or cream thus yields smaller fat globules. If the globules also
contain solid fat, they become attached the air bubbles.
As the air surface area diminishes (because air bubbles coalesce), the attached fat
globules are driven nearer to each other; the liquid fat spread over the air bubble surface
readily causes the globules to form granules. Furthermore, the liquid fat makes a foam less
stable. Further aggregation of granules yields butter grains, in which a phase inversion has
apparently take place, i.e., oil and moisture droplets, concentrating and then working the
grains removes excessive moisture and reduces the moisture droplets in size. In this way,
butter is formed.
If there is very little liquid fat during beating in of air and if, moreover, the fat content is fairly
high, structures of fat clumps are formed. However, churning does not occur; the structures
entrap the air bubbles, so that whipped cream is obtained. Similar processes occur during
freezing and whipping of ice cream.
6.1.4 Creaming
Because of the differences in density between milk plasma and fat globules, the globules
tend to rise. This property is of great importance because it causes it causes (undesirable)
creaming during keeping and enables milk to be separated into cream and skim milk.
Creaming is much enhanced if the fat globules have been aggregated into floccules or
clusters.

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6.1.4.1 High pasteurized milk
In this milk, creaming of single fat globules may occur. The velocity versus of a rinsing
globules is usually obtained from Stoke’s equation.

− a( ρ p _ ρ f )d 2 Where,
Vs = ρp = density of plasma, ρf = density of the fat.
18η p ηp = Viscosity of plasma, a = acceleration, i.e. g
if creaming is due to gravity.
(ρ p _ ρ f )
= Temperature affects factor.
ηp
Homogenization is the main variable. For Stoke’s law to hold several conditions must be
fulfilled, but the equations is quite useful to predict trends. In practice, globule size and
temperature mainly determine the extent of creaming in high pasteurized milk.
Cream fat globules clusters are formed much faster than single globules. Clustering may
be caused by homogenization or by intense heating (sterilization). The latter clustering can
occur in evaporated milk where it causes undesirable creaming.
6.1.4.2 Raw milk
The creaming in the cold is usually determined by the flocculation of the globules by
“agglutinin,” i.e., a complex of cryoglobulins (predominantly immunoglobulin M) and
lipoproteins.

Low
temperature

Fig.6.2 Adsorption of “agglutinin” (black dots) onto milk fat globules and the ensuing
flocculation of the globules.
a. In the cold, cryglobulins precipitate onto all kinds of particles, especially fat globules (at
a rate about 10-3 times that predicted by the theory for fast flocculation).
b. The so “covered” fat globules aggregate into fairly large floccules.
c. Large floccules rise rapidly.
d. Large floccules overtake smaller ones and single fat globules, thereby enhancing
flocculation and rinsing still faster. In this way, a cream layer forms rapidly, even in a
deep vessel or a large mlk tank.
94
Main variables affecting natural creaming in raw milk.
a. Temperature: No creaming occurs at 37oC. The colder the milk, the quicker the
creaming is.
b. Concentration of agglutinin: it varies among cows and with lactation stage, i.e., high in
colostrums, negligible in late lactation milk.
c. Fat globule size, partly because smaller globules have a relatively larger surface area
and, consequently, need more agglutinin.
d. Fat content the higher the fat content of milk, the quicker the creaming is because
formation of floccules is quicker.
e. Agitating milk for a while at low temperature seriously impairs creaming. A possible
explanation is that aggregates of agglutinin are formed causing a decrease of the
number of active particles.
f. Warming such milk largely restores the ability to flocculate because it dissociates
agglutinin from the aggregates.
g. Heat treatment of milk can inactivate agglutinin. The effect precisely parallels the
insolubilization (by denaturation) of the immunoglobulins. Heating for 20 s at 71oC has
no effect, 73oC causes 25 % less creaming, and 78oC often leads to complete
inactivation.
h. Homogenization inactivates agglutinin, also if takes place without the fat globules being
present.
6.1.4.3 Cream and skim milk
Cold agglutination in raw or low-pasteurized milk leads to a deep cream layer of loosely
packed floccules, containing much plasma, the layer may contain 20 % to 25 % fat. The
cream can readily be redispersed throughout the milk. It contains many of the bacteria from
the milk. In low-pasteurized cream of 20% fat or higher, creaming will hardly occur; the
globules aggregate into one large floccule that fills the whole volume. However, gravity
tends to compact this flocculated system and a thin layer of plasma may form at the bottom
slowly increasing in height.
A different cream layer forms on high-heated milk. This is a thin layer of a high fat content
(40 -50 %), but the closest possible packing of globules (about 70 % fat) is not attained.
When the fat starts crystallizing in the cream layer, this may promote partial coalescence of

95
the fat globules, and so does stirring the layer. Consequently, it may be impossible to
redispose the cream throughout the milk. By centrifugal action, cream with a much higher
fat content, i.e., 80 % fat or more (plastic cream), can be obtained so that globules are
deformed.
Skim milk obtained after natural creaming rarely has a fat content of less than 0.5 %; the
milk still contains fairly large globules. Centrifugally separated milk has a far lower fat
content, made up by the smallest fat globules and about 0.025 % “non globular fat”.
Most of the agglutinin is found in the cream if raw or low-pasteurized milk is separated
when cold, e.g., 5oC. This can cause considerable agglutination in the cream. Most of the
agglutinin is in the skim milk if the milk is separated when warm, say, >35oC. Consequently,
a mixture of cold-separated skim milk and of cream obtained by warm separation displays
hardly any cold agglutination.
6.1.5 Lipolysis
In milk, lipolysis (i.e., enzymatic hydrolysis of triacyl glycerides) causes free fatty acids to be
formed, and this may give the milk as soapy-rancid taste. Several enzymes can be
responsible for lipolysis, but here we consider the main lipolytic enzyme of milk, i.e.,
lipoprotein lipase.
Optimum temperature of the enzyme is about 33oC, optimum pH about 8.5. Milk contains
10-20 mmol of the enzyme per liter. The natural fat globule membrane protects the inside of
fat globules against enzyme attack.
Increasing the number of milking also leads to increased lipolysis. Increased fat acidity
especially occurs when milk yield becomes low (<3 kg / milking), i.e., at the very end of
lactation. All of these conditions enhance “leakage” of lipoproteins from the blood into the
milk.
Lipolysis can, however, readily be induced in milk. Removing the natural membrane from
the fat globules by intense beating in of air, or increasing the fat plasma proteins, this
causes interfacial tension up to 15 mN.m-1. As a result, lipoprotein lipase can penetrate the
membrane. Homogenization of raw milk thus causes very rapid lipolysis. Lipolysis can also
be induced by cooling raw milk to 5oC, warming it to 30oC, and cooling it to again, although
there is wide variation among milk samples. Inactivation of the enzyme by heating can
prevent lipolysis.

96
 Fat acidity
3

+
0
0 50 100
% susceptible milk
Fig.6.3 Acidity of milk fat (as mmol/100 g ) of milk “ susceptible” to
lipolysis, “normal” milk and mixtures thereof. The milk were kept for 24 h at 4oC.
6.2 Casein micelles
6.2.1 Description
Almost all casein in fresh uncooled milk is present in roughly spherical particles, mostly 40-
300 nm in diameter. On average, the particles comprise approximately 104 casein
molecules. These casein micelles also contain inorganic matter, mainly calcium phosphate,
about 8 g/100 g casein. They also contain small quantities of some other proteins, such as
part of the protease-peptone and certain enzymes. The micelles are voluminous, holding
more water than dry matter. They have a negative charge.
6.2.1.1 Submicelles
When casein is in a solution comparable with milk serum, but at low calcium ion activity, small and
roughly spherical aggregates form. These are about 12-15nm in diameter, and each of them
contains 20-25 molecules, called submicelles. The hydrophobic bonds and salt bridges keep the
molecules together.
Each micelles contains different casein molecules, but not all submicelles have the same
composition. There are two major types of submicelles with or without κ-casein. The submicelles
contain at the most 25 protein molecules. The molar ratio being αs1: αs2 : (β+γ) : κ = 4:1:4:1.6; κ-
casein exists in milk as a polymer, on average consisting of six molecules held together by –S-S-
linkages. Presumably, the C-terminal hydrophilic part of κ-casein, which may contain sugar
residues, is sticking out from the submicellar surface.

97
Adding an excess of calcium and phosphate, as occurs in the mammary secretary cells, results in
aggregation of the submicelles into larger units, i.e., the casein micelles. The deposition of the
calcium phosphate in the submicelles lowers their electric charge and also make them more
compact. Two submicelles aggregates each other protruding hairs of κ-casein of at least one of
them are in between will not be able to bound each other. The aggregation will goes on until a
roughly spherical aggregate is formed, whereas the surface of the micelle is covered with a more
or less continuous layer of hairs of κ-casein.
6.2.1.2 Model

Submicelle

Protruding peptide chain

Calcium phosphate

Fig.6.4 A casein micelle

A model of the casein micelle derived from the above reasoning. The micelle is a fairly
dense aggregate of submicelles. The latter contain small regions (often called
nanoclusters) of calcium phosphate in which the serine phosphate residues of the casein
are involved. The major portions of the peptide chains of casein in loose submicelles have
great freedom of motion. The calcium phosphate present in micelle almost fully lost the
flexibility of the peptide chains. Which are essential in providing the stability to the micelles.
6.2.1.3 Variability
The micelles are not the same in size. The great numbers of very small particles are loose
submicelles, which make up only a small part of the total casein. Authors are not fully
agreed about the size distribution. Some have found a small number of very large micelles,
up to 600 nm diameter. The volume surface average diameter (dvs), excluding the loose
submicelles, to be at least 100 nm. The variation in size may occur in milking intervals of
one cows and different lots of milk. Different cows produce milk with a different size
distribution.
The proportion of κ-casein varies it largely determines the average casein micelle size.
Because κ-casein may be at the surface of the micelles. Moreover, a small part of the κ-

98
casein at the interior of the micelles. The voluminosity of the micelles also varies. It will
markedly increase with decreasing micelle size because of the constant thickness (~ 7 nm)
of the hairy layer and the low protein content in that layer.The content and composition of
the colloidal calcium phosphate may vary, too. Large micelles probably have a higher
colloidal calcium phosphate content.
6.2.2 Changes
6.2.2.1 Dynamic equilibria
The casein micelle and its surroundings keep exchanging components. The exchanges can
be considered dynamic equilibria, though they may be pseudo- rather than true equilibria.
6.2.2.2 High temperature
On increasing the temperature the micelles shrink somewhat and the amount of colloidal
phosphate increases. The additional colloidal phosphate may not have the same properties
as the natural phosphate.
At temperatures above 70oC the casein molecules become more flexible, as if part of the
submicelle structure melts. At still higher temperature (above 100oC), dissolution of part of
the κ-casein occurs. The extent closely depends on pH; no dissolution occurs below pH 6.2
(measured at room temperature), but there is almost complete dissolution at pH 7.2.
Serum proteins become largely become largely become associated with the casein
micelles during their heat denaturation, and they largely become bound to the micelle
surface. The association should at least partly be ascribed to formation of -S-S- linkages.
The association of β-lactoglobulin with κ-casein is an example. Most of the associations are
irreversible on cooling.
6.2.2.3 Acidity
The colloidal phosphate goes into solution, with the dissolution being completed at pH 5.25.
Removal of all of the calcium requires a still lower pH, i.e., until below the isoelectric pH of
casein. The net charge of the micelles decreases by decreasing the pH because of
increasing association of hydrogen ions with the acid and basic groups. In other words, Ca
substitutes calcium phosphate to a certain extent. On further decrease of the pH, the
negative charge of casein increases again, due to dissociation of the calcium ions, and
eventually decreases again, due to association with H+ ions. At still lower pH casein
becomes positively charged. Furthermore, lowering the pH leads at first to swelling of the

99
micelles and eventually to considerable shrinkage. When the pH is lowered to, say, 5.3, a
large part of the casein goes into “solution”, more so with increasing hydrophobicity of the
casein concerned. At higher temperature the effect is smaller, and vice versa.
When the pH is lowered, the phosphate dissolves, resulting in increasingly weaker bonds.
Consequently, swelling of the micelles occurs, along with dissolution of part of the casein.
At low pH, internal salt bridges between positive and negative groups on the protein keep
the molecules together. Obviously, the total attraction is strongest near the isoelectric pH of
casein, i.e., near pH 4.6. It may conclude that the number and/or the strength of the sum of
all kinds of bonds is weakest near pH 5.25; this optimum pH depends somewhat on
temperature. Increasing the pH of milk also occurs swelling of the micelles and their
eventual disintegration.
6.2.2.5 Disintegration
Weakening of the bonds between the submicelles or those between protein molecules in
the submicelles can lead to disintegration. The former may be due to dissolution of the
colloidal phosphate at constant pH, e.g., by adding an excess of a Ca binder like citrate,
EDTA, or oxalate. The second type of disintegration occurs by addition of reagents like
sodium dodecyl sulphate or large quantities of urea, which break hydrogen bonds &/or
hydrophobic interactions.
6.2.3 Colloidal stability
Casein micelles are colloidal particles, large enough to flocculate as a result of mutial
attraction caused by van der Waals forces. The micelles do not flocculate under
physiological conditions. There is counteracting repulsive forces that prevent aggregation.
The micelles can be flocculate if the following conditions are met, e.g., by adding large
quantities of calcium ions or ethanol, or by applying very high temperature. Casein micelles
are much less stable than dissolved casein because of the much higher entropy of the free
molecules.
6.2.3.1 Causes of stability
The electrostatic repulsion and attractive van der Waals forces are responsible for colloidal
stability of particles. The free activation energy would prevent flocculation. Steric repulsion
caused by the hairy layer around the micelles must be responsible. The hair consists of a
flexible peptide chain, which keeps exhibiting conformational changes because of Brownian

100
motion. If the presence of another particle restricts the freedom of motion of hair, this
causes loss of entropy and thus repulsion. This is called volume restriction. It yields one of
the terms in the formula for steric repulsion. The other term is so-called mixing term, which
becomes important if the hairy layers of two micelles overlap. If the solvent quality of the
liquid for the hair is poor, mixing will lead to attraction of the micelles; if it is good, there will
be repulsion. The parts of the κ-casein sticking out from the micelles readily dissolve in milk
serum. This means that the solvent quality is good. Thus there is repulsion, which will
monotonously increase with closer approach of the surfaces of the micelles, because of
increasing interpretation of the hairy layers.
Variables affecting the micelle stability are:
- An increase in calcium ion activity
- Decrease of the pH
- Decrease of the dielectric constant
All of which would cause a decrease of electrostatic repulsion and are observed to
enhance flocculation.
6.2.3.2 Causes of instability
A change in environment may lead to flocculation (aggregation of the micelles). The
aggregation mostly seems irreversible. Age thickening and gelation mainly occur in
evaporated and sweetened condensed milk. Electron microscopy reveals that the micelles
in these products become much less smooth and increasingly show protrusions. This
change causes an increase of the viscosity of the product and eventual formation of a
continuous network, hence a gel.
Beating of the air in milk causes adsorption of casein micelles onto the air bubbles formed.
The micelles can partly spread over the bubble surface. The partly spread micelles can
conceivably touch each other at sites devoid of hairs, leading to their fusion.
Aggregation as caused by acidification can simply be explained; casein becomes insoluble
near its isoelectric point. Again lowering the pH results in a significantly decreased charge
which diminishes colloidal stability. Addition of ethanol lowers the solvent quality for the
hairs of κ-casein. This causes the hairy layer to collapse, and the steric repulsion to
diminish or even to change in attraction. The latter effect is enhanced by decrease
repulsion. Moreover, the colloidal phosphate passes into another, unknown state which

101
causes aggregation of the “micelles” to be irreversible. The lower the pH of milk, the
smaller the ethanol concentration needed to cause coagulation. This principle may be
applied to quickly detect slight sourness in milk.
As excess of Ca2+ ions enhances the possibilities of Ca bridge formation. Moreover, it
decreases the charge of the micelles and increases the super saturation with to calcium
phosphate in the milk serum. The latter would cause formation of additional colloidal
phosphate, which would cause fusion of micelles.
Freezing of milk leads to highly increased salt concentration in the remaining non frozen
solution.
6.2.3.3 Effect of temperature
The lower the temperature, the higher the colloidal stability of the casein micelles. There is
no strong bonds between micelles can exist at low temperature. After their aggregation at,
for instance, 30oC by renneting or acidification of milk, the formed gel is not dispersed
again on lowering the temperature to, say, 5oC; it even becomes firmer, which implies that
the number or the strength of the bonds between the micelles increases. In other words,
the casein micelles do not flocculate at low temperature because of high activation free
energy is required for flocculation.
6.2.3.4 Consequences of aggregation
Casein micelles encounter each other frequently because of Brownian motion. After
making contact they will draw apart again, but sometimes they can keep together due to
formed bonds and become aggregated. Subsequently, fusion of micelles may occur.
Usually fusion will be a slow process, but it is quicker if most of the hairs have been
removed. Essentially, fusion is the same reaction as the formation of casein micelles from
submicelles.
Most aggregations need a much longer time for fusion (reaction 2) than for flocculation
(reaction 1), and run as follows: Small flocs first flocculate with each other and with single
micelles, and then increasingly larger flocs flocculate with each other. Due to the chance
factor in flocculation, the flocs formed have a quite open structure.

102
1. +

2.

3.

Fig. 6.5 Aggregation of casein micelles. 1. Flocculation reaction. 2. Fusion. 3. Examples of

a floc of micelles formed during ongoing flocculation.
6.3 Physical properties
Some of the physical properties of milk are to a considerable degree determined by its
being a dispersion of colloidal particles. This is obvious for optical properties because milk
is turbid. Rheological properties are also strongly dependent on concentration and
properties of the particles in milk.
6.3.1 Optical properties
The refractive index, n, of a transparent liquid is defined as the ratio of the velocity of light
in air to that in this liquid. It depends on the wavelength of the light and decreases with
increasing temperature. It is generally determined at a wavelength of 589.3 nm and at
20oC.
The refractive index of milk (about 1.338) is determined by that of water (1.333) and the
dissolved substances. Particles larger than about 0.1 µm do not contribute to n.
Consequently, fat globules, air bubbles, or lactose crystals do not contribute to the
refractive index of milk and milk products, though they may hamper the determination of n,
by the turbidity they cause. Casein micelles, though many are larger than 0.1 µm. Do
contribute to n because they are inhomogenous and have no sharp boundary.
The difference between n of water and a aqueous solution is given by
n (solution) – n(water) = ∆ n ≈ ρ ∑ mi ri

103
Where, ρ is the mass density of the solution, m mass fraction of a solute, and r is its
specific refraction increment. The equation is not precise because the contribution of the
various components (i) is not always precisely additive. Values for r ( at 589.3 nm and
20oC) are, in ml/g (which implies that ρ has to be in g/ml).
Casein micelles, 0.207 per g casein
Serum proteins, 0.187
Lactose, 0.140
Other dissolved milk components,~ 0.170
Sucrose, 0.141
With these data, the refractive index of milk products can be calculated. When
concentrating milk or another liquid by evaporation, ∆ n/ ρ increases proportionally with the
concentration factor. Since n can be determined easily, rapidly, and accurately (standard
deviation 10-4 or better), it is a useful parameter to check changes in composition, such as
solids-not-fat content. Light scattering is caused by particles whose refractive index differs
from that of the surrounding medium. For instance, nfat globule/ nplasma ≈ 1.084.
6.3.2 Viscosity
Milk behaves as a Newtonian liquid; this means that the shear stress is proportional to the
shear rate (dv/dx). The viscosity of milk is about twice that of water. The difference is
caused by the dissolved substances and the dispersed particles. For a UF-concentrated
skim milk, casein micelles make up a far greater part of the mass for a given percentage
dry matter than is the case for evaporated skim milk. At low temperature, the voluminosity
of the micelles is markedly increased and part of the β-casein becomes dissociated from
the micelles. Consequently, the viscosity increases steeply.
Heat treatment of skim milk to such a degree that the serum proteins become insoluble
causes an increase in viscosity by about 10 %. This is because of increasing the the
voluminosity of the serum proteins.
Increases the pH of milk also increases its viscosity, presumably by additional swelling of
casein micelles. Slightly decreasing the pH usually leads to a small decrease of η. A more
drastic pH decrease causes η to increase, which is due to aggregation of casein.
Homogenization of milk has little effect on η, but homogenization of cream may
considerably enhance apparent viscosity.
104
6.4 Whey (serum) Proteins
6.4.1 Definition of whey
Broadly, whey may be defined as the serum or watery part of milk remaining after
separation of the curd that results from the coagulation of milk by acid or proteolytic
enzymes. Its composition will vary substantially, depending on the variety of the cheese
produced or the method of casein manufacture emplyed. On average, whey contains about
65 kg-1 of solids, comprising about 50 g lactose, 6 g protein, 6 g ash, 2 g non-protein
nitrogen and 0.5 g fat.
The protein fraction contains about 50 % β-lactoglobulin, 25 % other protein fractions
including, immunoglobulins. However, there will be wide variations in composition
depending on milk supply, and the process involved in the production of the whey.
6.4.2 Classification of whey
Sweet wheys : titratable acidity 0.10-0.20 %, pH = 5.8 to 6.6.
Medium acid wheys : titratable acidity 0.20-0.40 %, pH= 5.0 to 5.8.
Acid wheys : titratable acidity >0.40 %, pH < 5.0
In general, wheys produced from rennet-coagulated cheeses develop low levels of acidity,
while the production of fresh, acid cheeses, such as ricotta or cottage cheese, yields
medium acids or acid wheys. Whey from caseins produced by acid addition is classed as
high acid whey, whereas whey from rennet casein is sweet whey.
6.4.3 Proximate composition
Content (per kg powder)
Parameter
Sweet whey Acid whey
Water (g) 31.9 35.1
Food energy (kJ) 14760 14190
Protein (N×6.38) g 129.3 117.3
Total lipid (fat) (g) 10.7 5.4
Carbohydrate(total) (g) 83.5 107.7
6.4.4 Composition of whey
Whey variety
Parameters
Rennet sulphuric lactic HCl casein Cheddarcheese
T.S*** (g solids/kg whey) 64-67 60-64 62-65 58-61 61-66
True protein(g /kg solids) 110-140 85-120 84-110 96-124 99-110
NPN* : TN** ratio 0.24-0.28 0.24-0.29 0.30-0.35 0.18-0.24 0.22-0.30
Ash (g/kg solids) 74-78 120-132 117-123 116-194 76-91
Lactose ,, 750-810 680-760 620-690 - 744-810
Calcium ,, 6-8 21-24 24-27 - 6-7
105
Inorg. phosphate ,, 10-16 29-34 30-34 - 8-30
Potassium ,, 18-26 8-24 20-24 - 19-25
Sodium ,, 6-16 4-14 6-12 - 6-11
Chlorine ,, 14-21 12-18 13-19 - 14-20

Note: *NPN = non protein nitrogen; **TN = Total nitrogen ; ***TS = Total solids.
Whey is the liquid remaining after removal of casein and fat from milk in the process of
cheese making. It contains most of the salts, lactose, and water soluble proteins of the milk.
Casein may be defined most simply as the protein precipitated by acidifying skim milk to a
pH value near 4.6 at 20oC. The protein remaining after casein has been removed from skim
milk are known as whey proteins or milk serum proteins. The whey protein consists of 20 %
of the total milk protein.
Whey proteins are high molecular proteins. They have higher affinity with water and
remains highly dispersed in milk. Due to highly dispersed nature in milk, even in their IEP,
they remain unchanged.
6.4.5 Whey proteins
About 20 % of the total protein of bovine milk belongs to a group of proteins generally
referred to as whey or serum proteins or non-casein nitrogen. These whey proteins are
further divided into two groups.
6.4.5.1 Heat liable (Thermounstable)
when whey protein is boiled to 20-30 min at pH 4.6, a part of proteins are precipitated and
are called thermostable protein occupies 81 % of whey protein. They are still further divided
into two types.
1. Lactalbumin
α -Lactalbumin consists (2-5 %) of skim milk protein. Lactalbumin is that portion of heat
liable milk serum protein that is soluble in saturated MgSO4 solution or saturated
(NH4)2SO4 solution. The lactalbumin fraction of bovine milk contains three main proteins, β-
lactoglobulin, α-lactalbumin and blood serum albumin which represent approximately 50, 20
and 10 % of total whey protein, respectively, and trace amounts of several other proteins,
notably lactotransferrin, serotransferrin and several enzymes.
2. β -Lactoglobulin
It consists of 7-12% of skim milk proteins. The lactoglobulin fraction consists mainly of
immunoglobulins (Ig), especially IgG1, with lesser amounts of IgG2, IgM. Lactoglobulin is
106
that portion of heat liable milk serum protein that insoluble in half saturated ammonium
sulphate or MgSO4 and are precipitated. Among 81 % of thermoliable whey protein, 68% is
lactoalbumin and 13 % lactoglobumin. Lactoglobulin constitutes by α-lactalbumin, 19.7 %;
β-lactalbumin, 43.6 %; serum albumin 4.7 % of whey protein. Whereas lactoglobumin
constitutes by euglobulin (true globulins), pseudoglobulin (false globulin).
Euglobulins are insoluble in water but soluble in 0.6% of NaCl solution. Pseudoglobulins
are soluble in water but insoluble in ethyl alcohol. Mostly colostrum is pseudoglobulins are
also called immunoglobulins because they fight with the infectants or contaminants.
6.4.5.2 Heat stable (Thermostable) protein
When milk serum (whey protein) is treated to boiling point for 20-30 mins at pH 4.6 to 4.7,
the protein which remains stable and would not precipitate out are called heat stable whey
proteins e.g. protease and peptones. These heat stable proteins are 19% of whey protein.
Preparation of soluble lactalbumin
Among α, β, lactalbumin, α-is the most heat stable. Lactalbumin is heat coagulated and
hence about small amount is coagulated when milk is pasteurized and nearly ⅓rd of the
lactalbumin is coagulated when milk is heated at 160oF for 30 min. Lactalbumin is not
coagulated by rennins.
Skim milk

Acidification to pH 4.6
or rennet treatment

Drain

Casein or Cheese curd

Optional pretreatment
pH 7-8

Heat 90-120oC

Acid, pH 4.5

Centrifuge

Lactalbumin

Fig.6.6 Preparation of lactalbumin

107
 Table 6.1 Essential amonoacids in cow’s milk products (g/100g)
Amino acids Casein Lactalbumin Dried non fat milk Dried whey
Tryptophan 1.3 2.2 0.5 0.15
Threonine 4.3 5.2 1.6 0.64
Isoleucine 6.6 6.2 2.3 0.73
Leucine 10.0 12.3 3.5 1.04
Lysine 8.0 9.1 2.8 0.77
Methionine 3.1 2.3 0.87 0.19
Cystine 0.38 3.4 0.32 0.25
Phenylalanine 5.4 4.4 1.7 0.32
Tyrosine 5.8 3.8 1.8 0.13
Valine 7.4 5.7 2.4 0.64

6.5.4.3 Manufacture of whey powder

Whey

Cream separation

Whey protein Ultrafiltration Separated whey

concentrate
Permeate Evaporate

Demineralize Crystallize Whey powder

Demineralized Centrifugation Mother liquor

whey powder
Roller/Spray dried Delactosed whey powder
Crude lactose
Redissolve Refine Refined lactose
Dry
Unrefined lactose

Fig.6.7 Flow chart for the preparation of different whey products.

108
Chapter 7
Microbiology of milk

7.1 Introduction
Milk is a good source of nutrients and edible energy, not only for mammals but also for
numerous microorganisms, which thus can grow in milk. It primarily concerns bacteria, but
some molds and yeasts can also grow in milk.
Growth of organisms
Bacteria can multiply by division. Every cell division yields two new bacterial cells. The
multiplication is a geometrical progression.
2o 21 22 23 2n
If a growing bacterial culture contains No cell ml-1, the bacterial count N after n divisions is
N = No . 2n (i)
Or, log N = log No + n log 2 (ia)
= log No + 0.3 n
The time needed for a full cell division thus determines the growth rate. It is called the
generation time g; it can be derived from the number of divisions occurring during a certain
time t;
t
g= (ii)
n
Consequently, in well-defined conditions the count N after a storage time t can be
calculated from equations (ia) and (ii), if No and g are known.
log N = log No + 0.3 t/g (iii)
Generation time g depends on several factors. In milk, the bacterium species (or strain) and
the temperature are of special importance. Other factors involved are pH, oxygen pressure,
and concentration of inhibitors and nutrients, which are fairly constant in raw milk.
Growth of the bacteria means an increase in the number present. It can be determine in
colony-forming units (CFU) per ml or g. Since several bacteria tend to remain attached to
each other after division, forming shorter or longer chains of individual cells (up to about
100, but generally less), the colony count may be much smaller than the actual numbers of
living cells present. This is especially true for lactococcus and streptoccous species, as well
as for some species of lactobacillus.
109
The equation given above should thus be interpreted with care. In some cases,
determination of the bio-mass of bacteria present would be preferable. It should further be
realized that yeast cells are far bigger than bacteria. The same holds true for molds, where
the differences between CFU and individual cell number also may differ greatly.
The above equations apply to the exponential growth phase of the bacteria (sometimes
called logarithmic or log phase) (fig. below). During the lag phase the bacteria do not
multiply; primarily because their enzyme system needs adaptation, enabling the bacteria to
metabolize the nutrients in the medium. The duration of the lag phase closely depends on
the physiological state of the bacteria, the temperature, and the properties of the medium.
Log count

Stationary
phase Dying-of
phase
Exponential
phase
Lag phase

Time

Fig. 7.1 Growth curve of a bacterial culture.

During the exponential phase, the growth is at a maximum rate until the stationary phase is
reached. In the latter phase, some growth still occurs, together with dying off. The decrease
of the growth rate is usually caused by action of inhibitors formed by the bacteria
themselves and/or by a lack of available nutrients. Eventually, the stationary phase turns
into the dying-off phase, during which the count decreases. The former two phases are of
special importance for the quality and the keeping quality of milk. In fermented milk
products the latter two phases also are essential.
Temperature has a great effect on bacterial growth. Lowering of the temperature retards
the rate of nearly all processes in the cell, thereby slowing down and decreasing
fermentation rate (e.g., acid production). Moreover, it extends the duration of effectiveness
of some of the natural bacterial inhibitors in milk.
Furthermore, many bacteria coming from a medium such as dung or teat surface and
entering a substrate like milk must adapt themselves to the new medium, hence the lag
110
phase. At a lower temperature the lag phase thus will last longer. The extent to which a lowering of
the temperature affects bacterial growth depends on the type of organisms present.
Table 7.1 gives some examples of the effect of temperature on generation time. It shows that lactic
acid bacteria will not spoil cold stored milk and that at 30oC pseudomonad’s grow more slowly than
other bacteria. The temperature dependence of the growth rate has consequences for the keeping
quality of milk, as is shown in Table 7.2 below. At high storage temperature, the milk has a poor
keeping quality, even if its initial count is low; it should be processed within a few hours after
production.
Temperature (oC) 5 15 30
Lactic acid bacteria >20 2.1 0.5
Pseudomonad’s 4 1.9 0.7
Coliforms 8 1.7 0.45
Heat resistant Streptococci >20 3.5 0.5
Aerobic sporeformers 18 1.9 0.45
Table 7.1 Generation time (h) of some groups of bacteria in milk
(not including the lag phase).
Milk held at (oC) Count after 24 h (ml-1) Keeping quality (h)*
4 2.5 × 103 >75
10 1.2 × 104 30
16 1.8 × 105 19
20 4.5 × 106 11
30 1.4 × 109 5
*
Keeping quality is defined as the storage time during which the milk remains suitable for
processing (count not exceeding 0.5 – 1.0 × 105 ml-1).
Table 7.2 Effect of the keeping temperature of milk on its count after 24 h, and on its
keeping quality (Initial count 2.3×103ml-1).
15OC 15OC
109
4OC
Count (ml
-1
)

107

105 4OC

103
24 48 72
Incubation time (h)

Fig. 7.2 Change of the colony count during the keeping of milk of two
initial counts, at two temperatures.
Broken lines mark the region where spoilage of milk usually becomes observable.
111
 15

Storage periods (days)

1 Legend:
4 1 : Streptococcus faecium 104
10 2 2 : Aeromonas spp. 100
3 : Bacillus cereus 0.1
3 4 : Lactic acid bacteria 100
5 : Pseudomonas spp. 100
5 5

0
0 5 10 15 20
Temperature (oC)

Fig. 7.3 Time needed to reach a count of 106 bacteria per ml when keeping raw milk
at various temperatures.The Bacterium considered and its initial count (in ml-1) are
indicated.

Fig. 7.2 shows that a low initial count and a low storage temperature are essential. Whether
the milk is kept at a low or at a higher temperature, a lower initial count always means that
it takes more time for the milk to spoil. Naturally, the combination of low initial count and low
storage temperature is to be preferred. It is important to not that for raw milk to be of
greater importance than the total count. For example, contamination by 105 mastitis
bacteria per ml of milk has less effect on the keeping quality at low temperature than 103
psychrotrophs per ml (Fig.7.3).
Growth of microorganisms in raw milk is generally undesirable. Of all treatments known to
reduce the growth of microorganisms, for raw milk only lowering of the temperature is
generally feasible. Heat treatment kills bacteria. Milk contains some natural growth
inhibitors, but addition of a bacterial inhibitor to milk generally is not allowed by the public
health authorities. Such inhibitors may pose a health hazard or cause off-flavor
development. But in some tropical countries high temperatures and poor hygienic
standards prevail; in order to bring the milk in good condition to where and when it is
needed for processing, addition of a bacterial inhibitor seems unavoidable. The use of H2O2
can under certain conditions be tolerated. A better preservation may result from activation
of the lactoperoxidase-thiocyanate-H2O2 system.

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7.2 Milk as a substrate for bacteria
Milk products can be taken as an ecosystem. The interaction between bacteria and
environment determines what will happen; bacterial action affects the environment, and the
latter determines which bacteria can proliferate. The environment includes the properties of
the substrate (i.e., milk or a derivative) and outside conditions, of which temperature is by
far the most important variable. The bacterial population can vary greatly. Raw milk left
contact with the outside world is essentially an open ecosystem; almost any bacterium can
be present, and the properties of the milk and the temperature largely determine which
bacteria will outmatch the others. Many milk products are closed or controlled ecosystems,
and the microbial changes occurring depend greatly on the particular contamination by
bacteria important, but so are their physiological conditions, such as stage of growth, and
the possible presence of bacteriophages.
The effect of the environment is usually different for growth, fermentation and in the case of
certain species, sporulation. Generally, conditions permitting growth are more restricted
than those for fermentation. For instance, several lactic acid bacteria do not grow to any
extent near 5oC but, other conditions being favorable, they may go on producing lactic acid
from lactose.
Milk contains a wide range of nutrients including all of the vitamins, that can grow in milk
may have very different properties. For some bacteria lactose is not a suitable energy
source. Another rely on free amino acids as a nitrogen source, and fresh milk contains only
tiny amounts of amino acids. Consequently, such bacteria often starts to grow after other
bacteria have hydrolyzed proteins, thus providing suitable nutrients.
Another such example is the production of CO2 by some lactic streptococci, which
stimulates growth of some lactobacilli.
Milk contains natural inhibitors. Some bacteria do not grow in milk despite the presence of
sufficient nutrients and suitable conditions. The important inhibitor found in milk is the
immunoglobulins.
Lysozyme is an enzyme that hydrolyzes polysaccharides of the bacterial wall, particularly
splitting off N-acetylmuraminic acid; this may cause lysis of the bacteria.

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7.3 Undesirable bacteria
Most microorganisms are undesirable in milk because they can be pathogenic or produce
enzymes that cause undesirable transformations in the milk.
7.3.1 Pathogenic microorganisms
Those enter in milk can be pathogenic for humans or animals. Human pathogens are
usually classified into those causing food infections and those causing food poisoning.
Food infection implies that the food, e.g., milk, acts as a carrier for the number of
microorganisms; which enters the human body through milk. So a person can become ill,
often not until a day or so after drinking the milk. In food infection, fairly small numbers of
microorganisms may suffice to cause illness, but it it does not grow in milk, it is very unlikely
to cause illness.
In food poisoning, the microorganisms form a toxin in the food (or such a toxin
contaminates the food by another route). The consumer rapidly falls ill. Large numbers of
the pathogenic microorganism are usually needed to cause food poisoning. The amount of
toxin produced should be large enough to give symptoms. Unlike food infection, food
poisoning does not imply that the pathogenic organisms are still in the food. Some toxins
are more heat resistant than the toxin producing microorganisms itself, e.g.,
Staphylococcus spp.
Non-pathogenic microorganisms: by themselves would not impair milk quality. It is that the
organisms require nutrients, which are achieved by producing enzymes that hydrolyze
lactose, protein, fat or other substances in the milk, in order to yield compounds suitable for
their growth. These conversions cause the milk to develop off-flavors and to be less
suitable for processing into retail milk and milk products, e.g., because of a decreased heat
stability of the milk. Most heat processing applied in dairy processing does not destroy all
microorganisms or all microbial enzymes.
7.3.2 Spoilage microorganisms
Milk is an excellent culture medium for many kinds of microorganisms, being high in
moisture nearly neutral in pH, and rich in microbial foods.
A plentiful supply of food for energy is present in the form of milk sugar (lactose), butter fat,
citrate and nitrogenous compounds in many forms (proteins, amino acids, ammonia, urea,
and other compounds), and the accessory foods and minerals required by microorganisms

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are available. Some inhibitory substances (lactoperoxidase and agglutinins) are present in
freshly drawn milk but soon become comparatively ineffective. Because of the fermentable
sugar, an acid fermentation by bacteria is most likely under ordinary conditions in raw milk,
but other changes may take place if conditions are unfavorable to the acid formers or if they
are absent. With a trend toward higher pasteurization temperatures, the spoilage flora of
pasteurized milk is more frequently becoming heat resistant, spore forming bacilli which can
be psychotropic.
When milk sours, it usually is considered spoiled, especially if it curdles, although the lactic
acid fermentation of milk is utilized in the manufacture of fermented milks and cheese. The
evidences of acid formation are first a sour flavor and then coagulation of the milk to give a
solid jellylike curd or a weaker curd that releases clear whey. The lactic acid fermentation is
most likely to take place in raw milk held at room temperatures.
In raw milk at temperatures from 10 to 37oC, Streptococcus lactis is cause souring. There is
some growth of Coliform bacteria, Enterococci, lactobacilli, and Micrococci. At higher
temperatures, e.g., 37 to 50oC, Str. thermophillus and Str. faecalis may produce about 1
percent acid and be followed by lactobacilli, such as Lactobacillus bulgaricus, which will
produce more acid. Some of the lactobacilli can grow at temperatures above 50oC but
produce less acid there. Thermophillic bacteria can grow at still higher temperatures, e.g.,
Lact. thermophillus. Little formation of acid takes place in milk held at temperatures near
freezing, but proteolysis may take place.
The pasteurization of milk kills the more active acid-forming bacteria but may permit the
survival of heat resistant lactis (e.g., Enterococci, Streptococcus thermophillus, and
lactobacilli), which will cause a lactic acid fermentation if the subsequent storage
temperature is high enough.
Many bacteria other than those termed lactics can cause an acid fermentation in milk,
especially if conditions are unfavorable for the lactic acid bacteria. The coliform bacteria
produce some lactic acid and considerable amounts of volatile products, such as hydrogen,
carbon dioxide, acetic acid, formic acid, alcohol, etc. Species of Micrococcus,
Micobacterium and Bacillus can produce acid in milk, mostly lactic, but ordinarily cannot
compete with the lactics.

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Butyric acid may be produced in milk by the action of Clostridium spp. under conditions that
prevent or inhibit the normal lactic acid formation. Thus after a heat treatment which
destroys all vegetative cells of bacteria but allows the survival of spores of clostridium, milk
may undergo the butyric acid fermentation with the production of hydrogen and carbon
dioxide gas.
At any given temperature most samples of raw milk undergo a typical series of changes
caused by a succession of microorganisms. At refrigeration temperatures, proteolysis may
be initiated by psychrotrophic bacteria such as Pseudomonas, and molds may then appear.
At room temperatures, an acid fermentation is most preferable, first by lactic streptococci
and coliform bacteria and then by the acid tolerant lactobacilli. Then molds or film yeasts on
the surface lower the acidity, permitting the formation of more acid. Eventually, when most
of the acid has been destroyed, proteolytic or putrefactive bacteria complete the
decomposition.
Pasteurization as applied commercially in HTST systems kills yeasts, molds, most
psychrotrophic bacteria, the coliforms, and rapid acid procedures like Streptococcus lactis.
The spoilage of pasteurized milk then depends upon
1. the bacteria that survive pasteurization, the “thermodurics” and sporeformers;
2. the bacteria that enter the milk following pasteurization, postpasteurization
contamination from equipment, filling , operation and the package itself;
3. the possible presence of heat-resistant residual microbial enzymes; and
4. the temperature of storage.
Of course, once the milk is pasteurized and sealed in the cartoon, the microbial flora
present is established unless the carton losses its integrity. Therefore, the storage
temperature dictates which organism will predominate and with this fixed flora and number
per millimeter the temperature determines the rate of spoilage for that particular carton.
Likewise, the initial number and types present following pasteurization affect the spoilage
rate if the temperature of storage is fixed. Since milk is refrigerated, spoilage usually results
from organisms with psychotropic tendencies. Growth at low temperatures is slow but
significant. Assuming a generation time of 8 hr at 7oC, one cell could become 1 million in 20
generations, or about 6 to 7 days, or 2 million in 7 to 8 days, or 1 billion in 10 days.

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7.4 Pathogenic organisms in milk
The organisms that may gain entry to milk may be placed into two groups.
a. Those transmitted in the milk by a diseased cow’s udder.
b. Those that may contaminate the milk though some human agency after it has left the
cow’s udder.
(a) Milk from diseased udder
(i) Tuberculosis: It is widespread disease in cattle. Particles of infected dust or manure
may contaminate the milk, or it may be infected directly from a tubercular udder. The
causative organisms; in cattle, called Mycobacterum bovis (one of the heat-resistant
of the non-spore forming pathogenic bacteria). It is destroys by pasteurization.
Infected raw milk is carrier by which milk-borne tuberculosis is transmitted to man.
The feeding of skim milk and cheese whey derived from infected raw milk has been
a source of tuberculosis in animals. Some strains of mycobacteria, have been found
to survive pasteurization.
(ii) Brucellosis: It is the name applied to the disease caused by members of the group
of organisms known as the Brucella. In man, brucellosis is also called Malta fever,
meditarian fever, and undulant fever. In cattle the disease is often called Bang’s
disease, or contagious Abortion.
Brucella abortus, the organism that causes the disease in cows, is a common
source of brucellosis in man, but brucella suis, the organisms associated with the
disease in hogs, also has been found associated with milk borne epidemics of the
disease. Brucella suis is more infective to man than Brucella abortus. The causative
organisms are destroyed easily when milk is pasteurized.
(iii) Leptospirosis:
It is a spirochetal infection which is found throughout the world. In man it is known
as Weil’s disease, or infectious or hemorrhagic jaundice. Mild attacks resemble
influenza and the illness also has been confused with typhoid fever. The Leptospira
have been found in the milk of diseased cows.
Coxiellosis – Q-fever- It is a pneumonia- like disease of rickettsial origin. The rate of
mortality is negligible. The causative organism is Coxiella burnetii. It is one of the
organisms most resistant to destruction by pasteurization and proper relationships of

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 time and temperature must be carefully observed to ensure it destruction.
Pasteurization at 143oF for 30 min was found to be inadequate, but 145oF for 30 min
or 161oF for 1 sec destroys the organisms.
b. Diseases from contamination of milk by an external source :
The contamination of milk, milk products, or milk-handling equipment by persons
recovering from an infectious disease, or acting as “carriers” of the disease, it
perhaps the most frequent cause of milk-borne disease. In such cases infection by
Salmonella and Staphylococci is most common.
7.5 Sources of Contamination
1. Microorganisms present in the udder
a. Healthy cows
In most cows, no microorganisms are present in the milk in the alveolus, duct, and teat
cistern, but they are in the teat canal and the sphincter of the teat, mainly heat-resistant
Micrococcus and Staphylococcus spp. and Cornybacterium bovis. Sometimes other
bacteria are also involved. During milking these bacteria enter the milk. Directly after
milking, their number varies widely among cows, from hardly any to about 15000 ml-1; the
colony count of aseptically drawn milk of healthy cows is usually low, e.g., <100 ml-1. At 5oC
the bacteria hardly grow and after low pasteurization these organisms can often not be
detected. Obviously, microbially high grade milk can be collected from the healthy cows.
The cows has several defense mechanisms to keep microorganisms away from the udder.
• The sphincter of the teat.
• Bacteriostatic and bactericidal agents present in the keratin material of the teat canal
and in the milk itself, and the leukocytes in the milk.
• The “rinsing effect” due to discharge of the milk.
b. Unhealthy cows
When a cow is ill due to microbial infection, the organisms involved can enter the milk. In
the case of mastitis, pathogenic organisms are already present in the udder and thereby in
the milk. Because of this, mastitis milk usually has a high count. Some of these mastitis
organisms, including Mycobacterium tuberculosis, certain Streptococci, Staphylococcus
aureus, and certain strains of Escherichia coli are also pathogenic to humans.

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If organs other than the udder are inflamed, pathogens may directly enter the milk through
the body, especially if the cow is also mastitic. Naturally, the organisms can also enter the
milk through, for instance , dung or urine. Among such organisms that are pathogenic to
humans are : Leptospora serotype hardjo, Micobacterium tuberculosis, Campylobacter
jejuni, Listeria monocytogenes, Bacillus anthracis ( cause anthrax), Brucella abortus
(causes an illness resembling Malta fever in humans), and the foot-and-mouth disease
virus. Obviously, it is essential to exclude milk of diseased animals from processing and to
heat the milk in order to kill any pathogens. The drinking of raw milk is highly inadvisable.
2. Contamination during and after milking
The hygienic measures taken during and after (mechanical) milking essentially determine
what foreign microorganisms enter the milk, including human pathogens. This applies also
to their numbers. The count of properly drawn mixed milk from healthy cows is about
10,000 ml-1, sometimes even less. If, however, the hygienic standards during milking are
poor, freshly drawn mixed milk can have a much higher count, up to 1 million ml-1. Potential
sources of contamination of milk together with the characteristic microorganisms involved
will now be discussed.
a. The cow
During milking, microorganisms can enter the milk from the skin of the teats, which often
are contaminated by dung, soil, or dust, flakes of skin, hairs, and dirt from the feet and
flanks can also enter the milk. Several types of microorganisms can contaminate the milk,
including coliforms, fecal streptococci, other intestinal bacteria, bacterial spores (mostly
clostridium spp.), yeasts, and molds, Some of the microorganisms are human pathogens.
Appropriate housing and care of the cows is an essential measure to promote clean
udders. As a result, dry treatment, including removal of loose dirt, suffices at milking. Such
a dry treatment, moreover, causes less leakage of milk against the teats. Fewer bacteria
then because detached from the teat skin. Dirty udders have to be cleaned thoroughly
before milking. However, the complete removal of bacteria is impossible.
b. Soil, Dung, Dust
All of these contaminants can reach the milk and thereby increase counts. Moreover,
spores of bacteria, yeasts and molds also occur in air. Well known is B.subtilis, originating

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from hay dust. The spores can enter milk through air sucked in during mechanical milking,
or fall directly into the milk during milking in open milking pails.
c. The feed
Feed also contains large numbers of microorganisms. Feed can sometimes fall directly into
the milk but, more significantly , certain microorganisms in the feed survive passage
through the digestive tract and subsequently enter milk through dung; it includes some
human pathogens. Spore forming bacteria Bacillus cereus, B. subtilis, and Clostridium
tyrobutyricum, which can spoil milk and milk products, are especially involved. Large
numbers of C. tyrobutyricum occur in silage of inferior quality. The bacterial spores survive
low pasteurization of cheese milk, to which a more intense heat treatment cannot be
applied, and may cause “late blowing” in some types of cheese.
d. Milking unit
Contact infection poses the largest threat of contamination to almost all foods, including
milk. Poorly cleaned and disinfected milking equipment can contain large numbers of
microorganisms. Since these organisms generally originate from milk, they will grow rapidly
and can decrease quality. Residual milk often contains about 109 bacteria ml-1, and even 1
ml of such milk entering 100 L of milk during the next milking would increase the count by
10,000 ml-1.
The methods of cleaning and disinfection applied largely determine the species of the
contaminating organisms. If high temperatures are used and cleaning and disinfection of
milking utensils are unsatisfactory, the main species will be heat-resistant, including
Micrococci, Micobacterium lacticum, some Streptococci and spore forming bacteria. If, on
the other hand, low temperature are used, lactic acid bacteria, e.g., Lactococcus lactis,
Pseudomonads, and coliforms, will mainly be involved. Use of milking equipment that can
be adequately cleaned and disinfected is thus paramount. Small cracks in worn-out rubber
units and “dead ends” in the equipment that are insufficiently rinsed should be avoided.
e. Water used
Tap water may be of good quality. Any private water supply must be examined at intervals.
Surface water can contain many microorganisms, including human pathogens, and it must
therefore on no account be used for cleaning and rinsing. Gram-negetive rods like
Pseudomonas, Achromobacter, Flavobacterium and Alcaligenes spp., most of which are

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psychrotrophic, often occur in contaminated water (also in dung, soil, and poorely cleaned
utensils). Especially in tropical countries, the water may have very high counts.
f. The milker
The milker affects many of the factors mentioned and thereby the microbiological quality of
the milk. He can also contaminate the milk directly, e.g., from his hands. If the milker suffers
from a microbial infection, he might directly contaminate the milk with pathogens.
7.6 Hygienic measures
In discussing measures that would result in a satisfactory bacteriological milk quality,
contamination by undesirable bacteria should be distinguished from growth of the bacteria
in milk. Butyric acid bacteria, for example, can’t grow in milk, but the presence of more than
1 spore /ml of milk is undesirable in the production of some types of cheese. Psychotropic
bacteria, however, grow rapidly in milk, and contaminated by 102-103 ml-1 during milking is
hard to avoid. Counts lower than 105 ml-1 do not harm. The distinction should be made in
relation to food infection and food poisoning. Hygienic measures should aim at suppressing
pathogens and inhibiting spoilage organisms.
7.7 Protection of the consumer against pathogenic microorganisms
The following are the main reasons why contamination of raw milk by pathogens and
growth of these organisms in milk during storage should be avoided as much as possible.
a. During microbial growth in raw milk, toxins may be formed. Some toxins are fairly heat-
resistant.
b. Some pathogens survive heat treatments such as pasteurization. Fortunately, this is
exceptional. The higher the content in raw milk, the more organisms may survive heat
treatment. This is of importance if the heat treatment applied leaves only a small
margin.
c. The heavier the contamination of raw milks by pathogens, the greater the risk of
recontamination of the heated milk.
Contamination of raw milk by pathogens can never be ruled out. Milk intended for liquid
consumption or for transformation into milk products is therefore often required by law to be
heated to such an extent that the common pathogens are killed; this implies at least low
pasteurization.

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The drinking of raw milk is highly inadvisable. However, semi-hard cheese made from raw
milk is harmless for the consumer. The lactic acid bacteria rapidly hydrolyze the lactose to
yield lactic acid, which is not a suitable carbon source for most pathogens. As a result, pH
decreases rapidly to below, say, 5.5, which is unfavorable for many pathogens. The redox-
potential drops to a low value, about –150 mV, which prevents aerobic microorganisms
from growing. Moreover, the lactic acid bacteria form compounds that are antagonistic to
some pathogens. Most pathogens, if present, die within a few weeks. However, there is a
real danger that pathogens are present in soft cheeses made from raw milk.
Measures taken to prevent growth of spoilage organisms also stop growth of pathogenic
bacteria that can produce heat-resistant toxins. Pasteurized milk is therefore among safest
food products of animal origin.
7.8 Measures against spoilage organisms
A low contamination by microorganisms is the first aim. To achieve this, the sources of
contamination should be known. Some are found before milking, especially in housing
(clean cows) and fodder production (butyric acid bacteria).
Cleaning and disinfection of the milking equipment is essential. It is specifically meant to
remove and kill bacteria. Bacteria originating from inadequately cleaned equipment usually
have no lag phase and can grow rapidity in milk.
Cooling is the main means of slowing down the growth of bacteria in milk. The maximum
storage time of milk closely depends on storage temperature. A satisfactory operation of
refrigerated milk tanks on the farm is essential. However, cooling of milk kills no bacteria
and it can’t remedy unsatisfactory hygiene.
In dairy factories, the raw milk received often is not simply stored before processing, but is
thermalized and then cooled to below 4oC. Thermalization is a mild heat treatment, e.g., 15
s at 65oC. It kills nearly all pshychrotrophic bacteria, which are not at all heat-resistant. In
this way growth of these bacteria to harmful numbers during cold storage of the milk in the
factory is prevented, as is the formation of heat-resistant enzymes ( lipases and
proteinases). Thermalization kills part of the other bacteria, including many lactic acid
bacteria.

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Chapter 8
Milk processing : general aspects

8.1 Introduction
Milk is the raw material for the manufacture of different food products. These are primarily
made in dairies (dairy factories). Some typical characteristics of the dairy industries are as
follows.
a. Milk is liquid and homogeneous (or be made homogeneous). This implies that transport
and storage are relatively simple and greatly facilitates the application of continuous
processes.
b. Milk properties vary according to source, season, and storage conditions, and during
keeping. This may imply that processes have to be adapted to the variation in
properties.
c. Milk is highly perishable and the same holds true for many intermediates between raw
milk and final product. This requires strict control of hygiene and storage conditions.
d. Raw milk may contain pathogenic bacteria and some of these can thrive in milk. This
also requires strict control of hygiene and stabilization processes.
e. Raw milk generally is delivered to the dairy throughout the year, but in varying quantities
(in some regions there is even no delivery during part of the year). Because the milk
must be processed within at most a few days, this implies that the processing capacity
of a dairy can generally not be fully used during most of the year.
f. Milk contains several components and it can be separated in fractions in various ways,
e.g., in cream and skim milk, in powder and water or in curd and whey. Moreover,
several physical transformations and fermentations can be applied. This means that a
wide variety of products can be made.
g. Besides milk, fairly small amounts of raw materials are needed for the manufacture of
most milk products but consumption of water and energy may be high.
h. One and the same unit operation can be applied in the manufacture of a range of
products this includes heat treatment, cooling, cream separation, and homogenization.
Nearly all processes steps, or unit operations, that are applied in food industry are also
applied in the dairy. They can be grouped as follows:
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 a. Transfer of momentum: Pumping, flow.
b. Heat transfer: Heating and cooling.
c. Mixing / Communition: stirring, atomization, homogenization, recombination. Last two
are physical transformation.
d. Phase separation: Skimming, separating milk powder from drying air, part of the
churning process.
e. Molecular separation: Evaporation, drying, membrane processes, crystallization (of
water, lactose, milk fat).
f. Physical transformation: gel formation (as due to renneting or acidification of the
milk), important elements of butter making, making of ice cream, etc.
g. Microbial and enzymatic transformation: production of fermented products, cheese
ripening.
h. Stabilization: Pasteurize, sterilize, cool, freeze. At least one of these operations is
virtually always applied. Most stabilization are also, or even primarily, aimed at
ensuring food safety.
The process affects the material interactions are intricate, specific, and of practical
importance. A through knowledge of the physics, chemistry, and microbiology of milk and
its components is needed to understand the changes, both intended and undesired,
occurring in the material during processing.
8.2 Objectives
In the development of processes for the manufacture of food products, several constraints
have to be taken into account. This includes availability of skilled staffs, material,
machinery, and specific knowledge, as well as legal conditions. However, the objectives of
the production process are paramount. The ensuring requirements can be grouped as
follows:
a. Safety of the product for the consumer: The health of the consumer can be threatened
by pathogenic bacteria (or their toxins) and by toxic or carcinogenic substances.
b. Quality of the product: This generally involves:
• Nutritional value
• Eating quality: taste, odor, mouth feel.
• Appearance: Color, texture

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 • Keeping quality and shelf life: i.e., the time a product can be kept before it
significantly decreases in quality.
• Usage properties, e.g., spreadability of butter, whippability of cream, dispersibility of
milk powder; and, in general, ease of handling.
• Emotional value: a wide range of aspects, greatly varying among consumers.
c. Quality of the process: The process should be safe and convenient for the staff
involved as well as for other people around. It should not cause environmental problems,
such as pollution or excessive use of exhaustible resources (e.g., energy and water).
d. Expenses: Often, the necessity to maintain the processing costs within limits is
overriding. It may concerns the concerns the price of raw materials (including packing), the
use of energy, the equipment expenditure, the labor intensity etc. Also the flexibility and the
complexity of the process, with the ensuring probability of making mistakes (poor quality or
even discarding of products), may affect production costs. The same holds true for the
costs of storage. The objective are manifold and often mutually conflicting. This means that
process optimization may be far from easy to achieve.
8.3 Quality assurance
Quality assurance (QA) is paramount importance in all manufacturing and handling. It
involves a coherent system of activities that assures (guarantees) that the products made
meet a set of defined quality marks.
8.3.1 Concepts: Quality: Definition: A well definition (by J.M.Juran) is: “Quality is fitness
for use.”
A product or a service is fit for use if it meets the expiations of the user. However, it is far
from easy to establish what these expectations are. This is because the expectations vary
among consumers. Often widely so, and generally depend on conditions under which a
product is purchased or used. Moreover, several quality marks are highly subjective and it
is difficult to translate these into measurable product attributes. A high quality does not
merely mean that the product complies with legal requirements or preconceived ideas of
the manufacturer. Marketing specialists and technologists should cooperate in establishing
the desired quality marks. Food technologists then play a key role in translating quality
aspects into defined criteria and in developing methods for determining whether and to
what extent a criterion is meeting.

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The value can be estimated by more or less objective methods (e.g., safety, shelf life,
dispersibility), and others can only be assessed by consumer panels (e.g., flavor). It is often
tried to establish correlations between objective criteria and consumer opinions, e.g.,
between acidity or diacetyl content of fermented milks and their flavor, or between a
rheological parameter and the subjectively observed spread ability of butter.
Quality is a management function. It must be controlled (enforced). The current approach is
a system of integrated or total quality management. It involves integration at three levels.
a. Throughout the production chain i.e., from the farm to the consumer. It may even have
to start before the farm, for instance in the design of milking machines or in the
specifications for concentrates fed to the cows. Distribution of the products made also
involves several steps that bear on product quality.
b. For the product in the widest sense, including service. This would involve the way in
which the product reaches the consumer and the information given about the product.
c. Throughout the organization, i.e., at all hierarchical levels and in all departments.
Quality begins with the design: can good products be made by the planned procedures?
The next question is whether the desired quality can be reproduced: does every item
produced comply with the set quality criteria? For the latter, a control system has to be
installed. However, the general rule should be that “prevention is better than cure”.
The product should be safety to the consumer. Milk may contain several types of
pathogenic bacteria. Because their presence is largely determined by chance, and because
a single bacterial cell can in principle be dangerous (since same pathogens can grow in
milk), safety cannot be assured by selecting and inspecting samples. It is almost never
possible to check every unit of the product.
Other measures are-
• Treating the raw milk in such a way that all of the pathogenic bacteria that can be
present and harmful are killed.
• Prevention of recontamination of intermediates and end product. This requires strictly
enforced hygienic measures and packing.
• Transformation of the material into the product in which pathogens can not grow; a good
example is fermented milk. Preferably, any pathogens present will die off.

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 • Combination of these three treatments methods will give at least chance for “accidents”
to happen. However, all of these measures, especially the third one, cannot always be
taken. Consequently, a rigorous inspection and control system must be established.
Health hazards due to toxic or carcinogenic concentrations of substances in the product are
very rare in milk products.
8.3.2 Hazard analysis and critical control points (HACCP)
It is method to establish for an existing production process what control measures are
essential to assure the safety of the products made. The same method can be applied for
other quality characteristics, but the emphasis generally is on safety. HACCP should be
applied separately to every manufacturing process actually in operation; this means a
separate system for every product or group of closely related products. The main features
of the method are:
• Make an analysis of the potential hazards,
• Identify critical points in the process, and
• Established criteria for control.
HACCP is also a control system applied after the analysis has been made. It involves
corrective measures where needed, e.g., via feedback or control loops that adjust process
variables if needed; a single example is adjustment of heating temperature. A HACCP
study may reveal that the process should be changed to allow efficient control.
The manufacturing process is carefully described in a flow diagram, including the control
points for adjusting the process (e.g., temperature, flow rate, mixing intensity, rate at which
a component is added). Each step in the process is analyzed what measures can be taken
to minimize the hazard, and it is finally decided, on the basis of systematic criteria, whether
this is made critical control point. If so, a monitoring scheme is devised an essential point of
which is the monitoring frequency. Monitoring too often is unnecessary expensive and
tends to demotivate the operators, monitoring too seldom may lead to an unacceptable
hazard. This procedure is applied to every process step, leading to a complete HACCP
system. Furthermore, a corrective action plan should be developed, i.e., what measures
should be taken when some critical parameter is observed to be outside the limits set. The
system should regularly be evaluated and verified during its application, and modified were
needed.

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An essential step is that HACCP system can not be copied. Every manufacturer has his
own particulars in the process applied and in outside conditions and constraints. Moreover,
the development and repeated evaluation of conditions and constrains. Moreover, the
development and repeated evaluation of the system by the people involved in its
application is a prerequisite for its success.
8.3.2.1 Principles
HACCP is a system which identifies specific hazard(s) (i.e., any biological, chemical or
physical property that adversely affects the safety of the food) and specifies measures for
their control. The system consists of the following 7 basic principles.
Principle 1:
Conduct a hazard analysis. Prepare a flow diagram of the steps in the process. Identify and
list the hazards and specify the control measures.
Principle 2:
Identify the critical control points (CCPs) in the process.
Principle 3:
Establish critical limits which must be met to ensure each CCP in under control.
Principle 4:
Establish a monitoring system to ensure control of the CCP by schedule testing or
observations.
Principle 5:
Establish the corrective action to be taken when monitoring indicates that a particular CCP
is moving out of control.
Principle 6:
Establish documentation concerning all procedures and records appropriate to these
principles and their application.
Principle 7:
Establish verification procedures which includes appropriate supplementary tests, together
with a review which confirms that HACCP is working efficiently.
Successful application of these principles requires a well defined and consistent
methodology. A logical sequence of 14 stages (HACCP procedure) is recommended.

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HACCP Procedure for developing a control system for the manufacture of a food product,
according to the European Hygienic Design Group.
1. Define terms of reference
2. Select the HACCP team
3. Describe the product
4. Identify intended use of product
5. Construct a flow diagram
6. On-site verification of flow diagram.
7. List all hazards with each process step and list all measures which will control the
hazards.
8. Apply HACCP decision tree to each process step in order to identify CCPs.
9. Establish target level(s) and tolerance for each CCP.
10. Establish a monitoring system for each CCP.
11. Establish a corrective action plan.
12. Establish record keeping and documentation.
13. Verification.
14. Review the HACCP plan.

HACCP-form (scheme)1.
Procedure description Risks Monitoring system CCP

HACCP-form (scheme)1.
Monitoring Corrective
CCP Critical limits Responsible
Method Frequency Responsible action plan

Article name: Page:

Article no:
Process step Identifying risk-factors Preventive measure(s)

HACCP documentation of risk factors.

Carry out: Approval: Date: Version: Page off Doc.no:
CCP no:
Dept :
Process:
Hazard: Description:

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 Chemical
Physical
Microbiological
Fixed by law
Others
Assessment of gravity and probability of occurrence:
Result of any use of decision tree:
Conclusion, determination of objectives and tolerances:
Remarks:
HACCP examination of raw material and packaging material
Carry out: Approval: Date: Version: Page off Doc.no:
Product/Product category: (Chemical, Physical and Microbial characteristics).
Origin:
Description of delivery, packing and store condition:
Description of ready making before use:
Assessment of risk factors
Conclusion
Remarks
Example of HACCP analysis:
Target level and tolerance Monitoring procedures Corrective action
o
• < 6 C ex. Farm • Temperature check • Investigate farm supply.
• Scheduled cleaning adhered • Supervision during production • Retraining
to: and site audits
• < 5oC in silo. • Temp. chart recorder. • Investigate milk supply temp. and silo
temp. control
• Maximum holding time of 48 • Time recorded on process log • Review production schedule
hrs. sheets.
• Barrier hygiene guidelines • Supervision during production • Retraining or building uprate.
adhere to. and site audits.

• Scheduled cleaning adhere • Site audits. • Retraining

to.
• Barrier hygiene guide lines • Supervision during • Retraining or building uprate.
adhere to. production and site audits.
• Scheduled cleaning adhered • Site audits. • Retraining.
to.
o
• 80± 2 C. Diversion valve. • Chart record/plant thermo- • Automatic flow diversion if low
Set at 78oC. meter. Operation of diversion temperature.
valve check daily.
• Time 15±1 sec. • Chart record. Holding tube • Adjust flow rate as necessary.
check
• To agree with plant master • Plant calibration • Adjust as necessary.
Thermometer.
• No raw cream or chilled • Annual checks for plate leaks. • Repair or replace as required.
water leak to product.
• No product/water leakage. • Press gauge check during • Repair as required.
production.
• Chill water TPC • Daily water sample laboratory. • Result outside target level initiate
< 1000/ml. urgent review.
• Coliform absent in 1 ml • First product through and • Result outside target level initiate
cream; TPC < 1000/m2 swabs – laboratory. urgent review.

130
 equipment surface. • Site audits. • Retraining.
• Conformance to CIP
schedule • Chart record/ plant • Result outside target level initiate
o o
• 4 ±1 C. thermometer. urgent review.
• Visual check on valve. • Machine adjustment.
• System on valve open.
• Scheduled cleaning • Site audits. • Retraining.
adhered to
• Coliform absent in 1 ml • First product through and • Result outside target level initiate
cream; TPC < 1000 /m2 swabs – laboratory. urgent review.
equipment surfaces.
• Air filtered to specified • Scheduled inspection of filters. • Replace or repair as required.
microbiological standards.
• Intact tank wall. • Annual checks for jacket leak. • Replace or repair as required.
• Chill water TPC <1000/ml. • Daily water sample – Lab. • Result outside target level initiate
urgent review.
• 4o ±1oC. • Chart record/ plant • Result outside target level initiate
thermometer. urgent review.
• System on valve open. • Visual check on valve. • Machine adjustment.
• Held<5oC, 24-48 h (max) • Temperature indicator and • Machine adjustment.
operator check. Discard product if outside
Time process log sheets. tolerance.
• Scheduled cleaning • Site audits. • Retraining.
adhered to
• Coliform absent in 1 ml • First product through and • Result outside target level initiate
cream; TPC < 1000 swabs – laboratory. urgent review.
/m2equipment surfaces.
• No chilled water leaks to • Schedule checks for water • Repair as required.
product. leaks.
• 4o ±1oC. • Chart record thermometer. • Result outside target level initiate
urgent review. adjust chill water as
required.
• System on valve open. • Process log checked during • Retraining of operatives.
production.
• Scheduled cleaning • Site audits. • Retraining.
adhered to
• Coliform absent in 1 ml • First product through and • Result outside target level initiate
cream; TPC < 1000 /m2 swabs – laboratory. urgent review.
equipment surfaces.
• Lid in place. • Inspection during production • Retraining or design review.
• Air TPC < 50 CFU/m3. • Air sampled for microbial • Review environmental cleaning if
level weekly. advance trend is observed.
• +ive press maintained • Air pressure check. • Repair or adjust.
• Air filtered to specified • .Scheduled inspection of • Replace or repair as required.
microbiological standards. filters.
• No condensation. • Observation for condensation • Review of unit &/or production
conditions if condensation observed.
• Barrier hygiene guideline • Supervision during production • Retraining or building up rate.
adhere to and site audits.
• Filtered to microbiological • Scheduled inspection of • Replace as required.
standards. filters. Site audits.
• Pneumatic exhaust directed • Inspection during production. • Retraining or design review.
away from product.
• Maximum 4 hrs. between • Temp. chart recorder. • Adjust air conditioning system
clean.
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Some source of contamination with possible CCPs in a simple milk flow diagram has shown below.
Milking

Milk handling ∅

Morning milk Evening milk (CCP)

⊗ ⊗∇ Hold overnight
Collection of farmer’s milk
∇
Mixing individual farmer’s milk CCP

Milk producer associations ⊕CCP

Can filled and
transported Chilling center
Water ∅
⊕∅
⊗ Milk storage Vat
Water ∇ CCP
⊗ Tanker at chilling center
Transportation
∇ Tanker at Factory
Pumped to plant
⊕ Raw milk storage tank ⊗∇◊

Pre-warming ⊗∇◊
Legends
Filtration / Clarification ⊗∇◊ ⊗ Possibility that milk or
water initially contaminated.
Cream separation ⊗∇◊ ⊕ Possibility of contamination
Skim milk /SMP / Water from surface equipment.
Standardization ∅ Possibility of contamination
from handlers
After pasteurization ♣ Destruction of most vegetative
Pasteurized at 74
o
C for 15 sec. cells, but spore of thermoduric
Before packing and thermoduric and
Circulated in CCP thermophiles can survive.
⊗ pipe line
After packing ∇ possibility of multiplication
⊕ Laminated plastic bacteria
Cold storage CCP ◊ Growth unlikely.
⊕ Consumer ∅

FIG.8.1 Shows the some possible hazards and critical control points applied in the
pasteurized milk production chains.
8.4 Quality assurance of raw milk
Total quality assurance involves the full chain from the production of raw milk to the
consumption of dairy products. Obtaining high-quality raw milk is a matter of particular and
lasting concern for a dairy. This is because in milk production so many steps and aspects
play a role and because so many individual producers are involved. Extensive measures for
quality assurance also have been taken in the distribution of dairy products. Raw milk
quality has several aspects, the most important being gross composition and hygienic
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quality. Gross composition means fat and protein contents in random samples; the price of
the raw milk is determined by its composition.
Assurance of the hygienic quality poses more problems, mainly because mistakes leading
to poor quality can readily be made and because sampling and analysis of every lot
delivered would often be too expensive.
The actual measures to be taken will vary greatly with local conditions, problems, and
regulations, but in any case milk must be sampled and analyzed on a regular basis.
The success of a quality assurance system would depend on a number of conditions:
a. Training and information to the farmers/procedures.
b. The procedure should be knowledgeable about hazards and remedies, and should be
committed to deliver high-quality milk.
c. Provisions of rewards and punishment :
The producer should be financially rewarded for producing milk of good hygienic quality,
and should be penalized for delivering milk that adulterated or potentially harmful.
d. Providing help in finding remedies of problem in high quality milk production.
e. Financial penalty for poor-quality milk should be restricted: If the farmer suspects that his
milk accidentally is of poor quality, e.g., because of cooling system have failed, he
should have the option of reporting this to the dairy. The milk can be collected separately
and not give any financial penalty to the procedures. The problem is that raw materials
for feed concentrate are sometimes contaminated with substances that may reach the
milk, e.g., aflatoxin.
Milk unacceptable hygienic quality should always be rejected. Quality criteria for raw milk
should not be more stringent than necessary to make safe and good quality milk products.
8.5 Milk Transport and Storage
8.5.1 Transportation of milk
Milk can is used to transport small quantities of milk, whereas tanker is used to handle
large quantities of milk. Tanker has certain advantages:
• Quicker mode of transport,
• Better temperature control,
• Less risk of contamination,
• More time and labor saving;

133
 • Lower investment in cans;
• Overall saving in detergents, etc.
Types of containers and equipments used.
• Baked earth
• Wood or bamboo
• Metal (generally brass)
• Galvanized-iron(GI)
• Second hand tins (mainly vegetable oil/ghee)
• Tinned iron or Aluminum-alloy ( used by organized dairies)
Problems in relation to collection and transportation of milk are
• Milk is liquid, perishable and bulky;
• Small and scattered production of milk;
• Tropical climate;
• Lack of transport facilities;
• Lack of countrywide organizations for milk collection and transport;
• Vested interests among local milk merchants.
8.5.2 Milk storage
Milk storage and transport operations are aimed at having good quality milk available where
and when needed for processing. The milk should not be contaminated by microorganisms,
chemicals, water, or any other substance. The costs involved in storage and transport
should be kept low, which implies that, for e.g., loss of milk should be minimized. Simple
and effective cleaning of all the equipment involved should be possible. A satisfactory
record of actual losses is desirable; most manufacturers determine a mass and fat balance
on a daily basis.
Transport and storage refer to raw milk as well as to intermediate products.
Almost all developed countries dairying also developed and production of milk is basically
confined to rural areas, while demand is mostly in urban areas. Therefore, milk has to be
collected and transported from production areas in the milk shed areas to processing
distribution points in cities.
Milk collection-cum-chilling centers
- Normally attached to city dairies.

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 Objectives: These are
• to preserve the quality of raw milk supplies, and
• to provide easy transport to the processing dairy.
Location: The chilling center should be located if the following requirements are fulfilled.
• Adequate milk production.
• Adequate water supply.
• Proximity to good road facilities.
• Electric supply and
• Sewage disposal facilities.
8.5.3 Equipments
• Milk weighing tank/pan and weighing scale;
• Drop(dump) tank with cover;
• Can washer;
• Milk pump (sanitary type);
• Surface/ plate cooler;
• Refrigeration unit ( of suitable capacity);
• Cold room ( of suitable capacity);
• Milk testing unit.
8.5.4 Operational procedure
On arrival, the milk is graded for acceptance/rejection, weighed, sampled for testing,
cooled and stored at a low temperature until dispatched to the dairy industry.
8.6 Preservatives
No preservatives are added to milk except soda (sodium carbonate) which is added for
neutralization. Sometimes factory neutralizes acidic milk by the addition of soda just before
processing. But, if neutralizer is detected in milk when receiving it is considered as an
adulteration, such milk is not accepted.
Milk seasons
1. Falgun – Shrawan (lean season)
2. Bhadra – Magh (flush season)

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8.7 Collection and reception of milk
Milk may be supplied to the dairy in milk cans or by a tanker after it has been cold-stored at
the firm (tank milk).
During transport, milk in cans usually has a temperature of >10oC, up to 20-30oC according
to the climate. Consequently, bacterial growth can occur between milking and milk arrival at
the dairy, as this interval may take as long as a day. The extent of bacterial growth depends
primarily on the quality of hygiene during milking, the temperature, and the storage period.
Spoilage of the milk is mainly by mesophilic bacteria and usually involves lactic acid
fermentation, however, heavy contamination with polluted water (mainly pseudomonad’s)
may cause a non-souring spoilage. On reception at the dairy plant, milk is cooled to <6oC,
which helps to more or less stabilize its bacteriological quality for at least 2 days.
Tank milk has been kept at low temperature for a longer time. It mainly contains
psychrotrophs and consequently requires another treatment than milk in cans. Among the
advantages of tank over milk in cans are the cheaper transport costs (if the collection routes
are not too long) and a regular supply of good quality milk, provided that the temperature of
the milk at the farm and during transport is satisfactorily controlled.
On reception, the quantity of milk is recorded first. At the dairy, milk in can is weighed by a
platform balance. The quantity of tank is determined by metering the intake line of the milk
tanker. Milk volume is then converted to weight.
Collected milk ought to be routinely to identify poor quality milk supplies. A simple, rapid
examination of the sensory properties would include odor, appearance, and temperature. In
addition, the intake pipe of the milk tanker can be equipped with a continuously recording
thermometer and a pH meter that may switch off the intake pump if the valves recorded
exceed a predetermined level. Incidentally, an off-flavor is more easily detected in the
warmer milk in cans than in tank milk, and souring of milk can be detected easier than the
growth of psychrotrophs. In addition to this simple inspection, the milk can be tested for the
presence of antibiotics, as well as freezing point depression, acidity and bacterial count.
It is advisable that the reception of milk in cans at the dairy occurs as soon as possible after
milking. This implies twice a day milk collection. Often this is not practical and the evening
milking is cooled by mains or well water. Once a day collection may, however, seriously
impair the milk quality in cans.

136
Tank milk should be refrigerated to <4oC. After 4 or 5 days storage substantial growth of
psychrotrophs may have occurred. Consequently, tank milk can normally be kept on the
farm for 3 days, i.e., 6 milkings, and stored for another day at the dairy before processing.
Milk can be contaminated during transport if the tanker had been inadequately cleaned.
Milk tankers can contaminate milk with high numbers of psychrotrophs. This means that
rigorous cleaning of the tanker and routine monitoring are essential. Furthermore, the
temperature of the milk during transport must be kept low, i.e., <5oC. Satisfactory quality of
raw material supplied. Milk supplies of poor quality should preferentially be eliminated.
8.8 Milk chilling and storage
8.8.1 Importance of chilling

Milk contains some of the micro-organisms when drawn from the udder. Their numbers
increases during the subsequent handling. The common micro-organisms grow best
between 20-40oC.Bacterial growth is invariably accompanied by deterioration in market
quality due to development of off-flavour, acidity etc. The method of preserving milk is by
prompt cooling to a low temperature i.e.,below 5oC. Effect of storage temperature on
bacterial growth in milk:

Milk held for 18 hrs. at Bacterial growth factors*

temperature (oC)
0 1.00
5 1.05
10 1.80
15 10.00
20 200.00
25 1,20,000.00
*
The initial bacterial count will be multiplied with this factor to get the total final count

8.8.2 Storage
The objectives are:
To maintain milk at a low temperature so as to prevent any deterioration in quality prior
to processing and/or product manufacture.

• To facilitate the bulking of the raw milk supply which will ensure uniform composition.
• To allow for uninterrupted operation during processing and bottling.
137
 • To facilitate standardization of the milk.
Methods of chilling:

a. surface cooler.
b. Plate coolers
c. Internal tubular cooler
d. Jacketted vat/ tank
Methods of storage:

a. Insulated / refrigerated.
b. Horizontal and vertical.
c. Rectangular, cylindrical or Oval.
d. Built for gravity flow, air pressure or vacuum operation.
Parts of the storage tank:

Sight glass, Light glass & lamp, ladder, Manhole, Agitator, Outlet valve, Inlet, Air vent,
Safety valve, Legs, Indicating thermometer, volume meter.

Variations in composition, properties, and quality of the raw milk directly affect the
manufacturing process as well as the composition and quality of the final products, and are
therefore undesirable. Some variation is inevitable, but mixing many deliveries in large
storage tanks, containing for example, 300 000 kg of milk, results in only a small variation
among lots of milk within 1 or 2 days.
1. Bacterial growth
The maximum keeping quality of raw milk in storage tanks is mainly determined by the
growth of psychrotrophs. Prior to processing, bacterial numbers greater than 5 × 105 ml-1 in
milk imply a risk that psychrotrophs have produced heat stable enzymes, i.e., bacterial
lipases and proteinases, which may impair the quality of the final product. It is important to
note that a high count originating from mixing a similar count resulting from limited growth in
the whole lot. This is because extra cellular enzymes are predominantly produced at the
end of the exponential growth phase. Initially, during storage on the farm, the total count
remains almost constant and only starts to increase after 4 and 5 days. The delay in the
growth of psychrotrophs to high number is often thought to be due to an extended lag phase

138
at low temperatures. However, a very low initial contamination with fairly fast growing bacteria
i.e.,<10 ml-1.
Depending on the age of the milk supplied to the dairy, it can be stored for further 1 or 2 days
without further treatment. All milk supplies should, however, be cooled to <4oC because the
temperature of the farm to the dairy and the generation time of bacteria is markedly shorter at high
temperatures.
Often, the dairy is unable to process all milk supplies within 4 days of milking. Consequently,
measures must be taken to keep the raw milk for a longer time. Pasteurization (72oC for 15s) is not
desirable because it will be done later on, and pasteurizing twice may impair the quality of the
finished products. A more moderate heat treatment (e.g., 65 for 15s, called Thermalization)
reduces the number of psychrotrophs considerably while leaving most enzymes and agglutinins
intact. After Thermalization, the milk can be kept for another 3 or 4 days at 6-7oC without
substantial increase in the bacterial count, provided that there is no recontamination by
psychrotrophs. Milk should be thermalized as soon as possible after arrival at the dairy.
Thermalization is a far better method for controlling the quality of dairy products than merely
cooling the raw milk, but it also more expensive. Since many bacteria survive thermalization,
considerable bacterial growth can occur at 30-40oC in the regeneration section of the heat
exchanger. Therefore, it may be necessary to clean it after operating for 4 to 6 h. The quality of
thermalized milk may still be threatened by the presence of any psychrotrophs that are fairly heat
resistant e.g., Alcaligenes tolerans.
Usually, the quality of milk is examined after it arrives at the dairy. It is advisable to test the milk
again just before processing. Standards for the milk quality before processing are given in table
below.
Table 8.1 Example of standards for (pooled) milk before processing.
Quality mark Standard Absolute unit
Acidity 17 ≤18 oN (mmol/L)
Count (Raw) 100 ≤250 <500 µL-1
Count (thermalized) 50 <100 <250 µL-1
Heat-resistant bacteria 5 <10 <25 µL-1
Bacillus cereus 0.1 <0.2 ≤1 ml-1
Fat acidity 0.6 <0.8 ≤0.9 mmol/100g
Freezing point depression 520-525 >515 mk
Antibiotics Not detectable
Disinfectants Not detectable

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2. Enzyme activity
Lipase activity is normally the main problem in fresh milk, although other milk enzymes,
e.g., proteases and phosphatases, also cause changes. Therefore, extensive temperature
fluctuations, in the range of 5oC to 30oC, and damage to fat globules should be avoided.
3. Chemical changes
Exposure to light should be avoided because it results in off flavors. Contamination with
rinsing water (which causes dilution) disinfectants (oxidation) and especially with cu
(catalyses lipid oxidation) all should be avoided.
4. Physical changes
The main physical changes during storage are:
a. Raw or thermalized milk stored at low temperature creams rapidly. Formation of a
cream layer can be avoided by regular stirring of the milk, e.g., stirring for 2 min every
hour.
This is often done by aeration; the air supplied should be sterile, for obvious reasons,
and the air bubbles fairly large, since otherwise too many fat globules would adsorb onto
the bubbles.
b. Damage to fat globules is mainly caused by air incorporation and by temperature
fluctuations that allow some fat to melt and crystallize. These events can lead to
increased lipolysis, to disruption of fat globules if the fat is liquid, and to clumping of fat
globules if the fat is partly solid (10oC to 30oC).
c. At low temperatures, part of the casein, primarily β-casein, dissolves from the micelles
to end up in the serum. This dissolution is a slow process and reaches equilibrium after
approx 24 h. The dissolution of some casein increases the viscosity of the plasma by
approx 10 % and reaches the rennetability of the milk. The reduced rennetability may be
partly due to a changed calcium ion activity. Temporarily heating milk to ~50oC or higher
almost fully restores the original rennetability of the milk.
8.9 Transport of milk to the dairy
To move the milk about, a dairy needs an intricate system of pipelines, pumps and valves,
as well as controlling units. The system should be flexible while excluding such errors as
milk running off or unintentional mixing of different products. To save on pumping costs,
gravity is often used. Pumping of viscous products requires much energy and the common

140
centrifugal pumps are unsuitable. These pumps are preferentially used for milk because
they keep turning without great problems if the milk cannot be discharged.
Some specific problems are:
8.9.1 Milk losses
The residues in pipes and equipment after processing, spillage, mixing of milk with different
products or with water when valves are switched over. Ensuring a satisfactory discharge of
the milk, avoiding “dead ends” in pipes, and minimizing the surface area wetted by milk all
are obvious measures to reduce losses. Minimizing the diameter (D) of pipes can reduce
the amount of mixing that occurs between milk and water and water; the volume of the
mixing region is proportional to D. Milk diluted with water may be evaporated or mixed with
skim milk powder or used as cattle feed. Proper operation reduces the cost due to milk
losses to approximately 1 % of the total coat of the raw material.
8.9.2 Damage to milk
Air incorporation may damage milk fat globules. Excessive shear rates and intense turbulence
during transport may cause clumping, i.e., formation of visible lumps of fat, especially in cream. In
transporting cream it is thus advisable to avoid narrow and long pipes, as well as obstacles (e.g.,
sharp bends) in the pipeline system; the cream should not be transported at temperatures between
10oC and 40oC. Furthermore, the viscosity of products like yoghurt and custard can be markedly
reduced by high deformation rates occurring during transport (irreversible breakdown of structure).
8.9.3 Bacterial growth
During transport contamination of milk by bacteria can readily occur. Balance tanks are
often situated before various kinds of processing equipment to ensure a constant milk flow
rate.
If the temperature is such a balance tank is high enough for bacterial growth, the tank tends
to act as a continuous fermenter, allowing growth of bacteria in the milk to be processed
leaving raw milk for same time in non-insulated pipelines favors bacterial growth. All such
situations should be avoided.
8.10 Filtration and Clarification
The purpose of filtration is to remove visible particles and dirt that may have entered the
milk. A new, dry filter is used for each lot of milk, because a used filter may harbor vast
number of microorganisms (bacteria) that would be washed into the milk if the old filter
were re-used. Usually the milk is heated to about 90 to 110oC before filtration.
141
 Instead of filtration, most milk plants use a mechanical clarifier to remove foreign matter
from the milk. The clarifier operates on the principle of the centrifugal cream separator. It is
so designed that the cream is not removed from the milk, but dirt, body cells, leucocytes
and some bacteria (so called clarifier slime), are caught in the bowl of the apparatus. Nearly
less than one half of the leucocytes present are removed during clarification.
In recent years, milk filtering devices which are provided with clearly woven cotton cloths
or cellulose pads have been used. The mesh of these filters is very fine and provides a
much more efficient filtering capacity.
In most cases, it is intended that these filter pads should be used but once and then
discarded, since their efficiency is impaired or destroyed by washing. They are found to be
effective in removing visible sediment from milk but do not filter out the leukocytes,
epithelial cells, cell fragments or microorganisms unless these are trapped on the large
particles which make up the visible sediment.
Clarification is an efficient way to remove dirt from milk, but it does not materially reduce
its bacterial content. It sometimes has the effect apparently of increasing the bacterial count
because its mechanical action tends to breaks up lumps of bacteria and release the
individual members throughout the milk. Clarification is done preferably before the milk is
pasteurized.
Clarification carried out at temperature below 32.2oC has little or no effect on the
creaming ability of the milk.
The object is to improve the aesthetic quality of milk by removing visible foreign matter
which is unsightly and unacceptable and may therefore cause consumer complaints. While
filtration removes suspended, foreign particles by the straining process, clarification
removes the same by centrifugal sedimentation.
8.10.1 Types
There are two types of filters/clarifiers.Those that operate with cold milk and those
operating with warm milk.
1. Filters
The important features are
i. A filter cloth or pad of the desired pore size which can retain the smallest particle.

142
ii. A frame or support to compress and hold the margins of the cloth or pad; so that milk
can pass only through the pores.
iii. A perforated metal or other support for the cloth or pad which will not tear or break
under the pressure of the milk.
iv. An enclosure to confine both the unfiltered and filtered milks in a close system fitted
suitably with inlet and outlet connections for sanitary piping.
v. A means of distributing the incoming stream of milk so that it does not damage or tear
any part of the cloth or pad by vigorous washing.
vi. A design so planned that filter cloths or pads can be changed quickly and all parts are
easily accessible for washing.
Where continuous operation is essential or where large volumes of milk are handled, two or
more filters are used so that operations needn’t be interrupted when it becomes necessary
to change the filter cloth. The frequency of cloth damage depends on the temperature of
milk, the amount of foreign matter etc.
2. Clarifiers
In general appearance and construction clarifiers are quite similar to centrifugal cream
separator. The main differences are
a. In clarifier, there is only one outlet.
b. The discs in the clarifier bowl smallest in diameter.
c. The milk distributing holes are at the outer edge of the disc in clarifiers, but near the
center in separators.
Relative Efficiency
Clarification removes sediment much more efficiently than filtration; clarifier removes still
finer particles that escape from filters.
General Remarks
a. Both filtration and clarification tend to decrease the depth of the cream layer that will
form on milk and this effect becomes more pronounced as the processing temperature
increases.
b. Neither filtration nor clarification improves the keeping quality of milk.
c. Milk should neither be filtered nor clarified after pasteurization, as this might
contaminate it.

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8.11 Membrane filtration process
8.11.1 Ultrafiltration (UF)
The colloidal and high molecular particles in solution are concentrated by means of
pressure drop(fig. below). The larger molecules such as protein, retained by the membrane,
whereas smaller molecules such as salts pass through it.
The liquid separated after UF called high molecular material known as concentrate
(retenate) and the liquid passes through the membrane is known as ultrafiltrate (permeate)
The pore sizes of the membranes ranges from 0.0005 to 0.005 µm. For normal operation
the maximum product pressure is 105Pa. The first membrane is made of cellulose acetate.
But it has poor mechanical and thermal strength as well as not resistant to lye and acid
during cleaning. The other membrane were developed later, such as synthetic polymers
and inorganic colloids, which overcomes the limitations of the former one. The suitable
polymers are cellulose acetate, polyamide, polysulphone and polyacrylic nitrile / polyvinyl
chloride, Z-f-99 and zerochromic monoxide etc.
Characteristics of the membranes
• They must manage high permeate flows.
• They must have a high relativity.
• They must have good chemical and bacteriological resistance.
• They must be resistant to detergents and disinfectants.
• They must be cheap.
The separation accuracy for a membrane is determined by pore size and pore
distribution. The molecular weight determine the separation limit, likewise the shape of the
separate particles also has an influence. A spherical particle is easier to separate than a
chain formed particle.
The different factors that affect the separation capacity are:
The membrane resistance: It is due to
• The thickness of the membranes.
• The surface area which consists of pores.
• The pore diameter.
The transport resistance: It is due to
When the separated material is deposited on the filter surface forming a layer. The layer
causes the actual filter resistance and it is often described as a secondary membrane. This
144
membrane causes reduced flow through the filter, changes the separation ability and the
bacteriological standard. The flow velocity, the length of the membrane and the product
concentration are the factors which affect the result.
SALTS

Macromo
Lecules &
Salts
Macromolecules
salts
Fig.8.2 The principle of Ultrafiltration (UF).
Only salts can pass through the membrane, whilst protein and other molecules are
retained by the membrane. The productivity of a UF plant is mainly determined by the
specific load on the membrane, expressed in lit/m2 per hr. This affects the density and
viscosity of the product. The specific load on the membrane is in the range of 25-50 lit/m2
per hr. Larger particles retains on the filter.
100 (C f − C p )
% retention ( R ) = J .
Cf

Where,
Cf = molecules concentration in feed.
Cp= molecules concentration in permeate.
J = flux ( L/ hr. m2 )
The flux (filtration flux) initially high and latter on during filtration goes on decreasing
down. Ultra Filtration mainly used for concentration of whey proteins from whey,
concentration of cheese milk and pre-concentration of milk for yoghurt manufacture.
8.11.2 Reverse osmosis (RO)
When a counter pressure (hydrostatic pressure) is greater than the osmotic pressure, the
solvent will flow from the concentrated product, through the membrane to the low viscosity
product. This is known as reverse osmosis. The pressure designated to be 70×105 Pa.
The membrane is cellulose acetate or synthetic materials- polymers such as polyamide.
In dairy industry mainly, the synthetic membranes are used because natural membranes
145
have narrow pH units (3-8) & low temperature resistance (50oC). Low molecular weight
molecules will be separated and the active filter layer must therefore be designed for the
purpose. The membranes used are called diffusion membranes. They are unsymmetrical or
have extremely thin, symmetrical layers. There is no membrane that can separate all
materials and only allow the solvent to pass. In dairy industry reverse osmosis is used for
the concentration of milk and whey. It is also used to produce fresh water from sea water.
R O is often used for pre-concentration of the product and the final concentration takes
place in an evaporator.

MACROMOLECULES AND SALTS

WATER

Fig.8.3 Shows the Reverse osmosis (RO) membrane, where salts are also separated.
8.12 Standardization of milk
It refers to the adjustment of the fat and solid not fat content in milk at a desired value so as
to conform or meet the prescribed legal standard. It refers to the adjustment of fat and
solid-not-fat percentage in standard milk & milk products by raising or lowering these
components is known as standardization. It should be remembered that all the
measurement are done by weight basis not by volume.
Standardization of the composition of a milk product is needed because it is legally required
or balance the manufacturer sets a standard for his product. It mostly concerns the fat
content, often also the dry matter content (or the degree of concentration), sometimes the
protein content or still another component. From the economic point of view, continuous
standardization is desirable; turbidity or density measurements can be applied for fat
content, density, or refractive index for dry matter content. Measuring infrared reflection is
also used, e.g., to determine the water content of the milk powder.
In continuous standardization, the mostly amplified measuring signal may control the
position of a regulating valve, e.g., a valve in a cream line or in a steam supply pipe; in this
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way the desired content can be adjusted. To achieve this, the relationship between turbidity
and fat content, between density and dry matter content, etc; in the original milk must be
known. This is because these relationships are not always the same. The adjustment is
often difficult because great fluctuation can easily occur when the adjustments are not
always the same. The adjustment is often difficult because great fluctuations can easily
occur when the adjustments are not always the same. Therefore, a double adjustment is
often employed, based on measurement of the volume flows as well as a concentration
dependent variable.
After the standardization, performed tentatively or by means of continuous determination,
the desirable content will have to be checked. This implies that it may be necessary to
make an adjustment by addition of cream, skim milk, water etc. Any bacterial or other
contamination should be rigorously avoided. The added component should have been
treated (especially with respect of heating) in a way similar to that of the product itself.
Standardization is always subject to inaccuracy because the results of the methods of
determination and the measuring or weighing of the components have a certain inaccuracy.
The same holds for the determination by the supervising authority. Therefore, a certain
margin should be left, e.g., twice the standard deviation. In same cases, for example with
respect to the fat content of beverage milk, a deviation of ±0.05 % fat may be permitted,
whereas the average value over a prolonged period should deviate by no more than 0.01 %
fat from the accepted standard value. Standardization of products (e.g., beverage milk) with
respect to protein content is generally not allowed. All the same, the nutritive value and the
cost price of the milk greatly depend on the (variable) protein content. Technically,
standardization is possible by applying ultra filtration.
The operation is carried out in two stages:
1. The cream and skim milk are separated by means of a disc centrifuge. At the same
time, the centrifugal force purifies and clarifies the milk
2. Thus, after separation, we have one circuit carrying cream, and one carrying skim milk,
which will be subsequently re-blended in proportions calculated by a microprocessor
according to the desired fat content in the standardized milk.

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The process of standardization
Initially, the fat content of skim milk, and the desired fat content for the standardized milk,
are entered into the control system which collects data from the viscous peripherals. Since
the fat content of cream is inversely proportional to the flow rate, this fat content is
controlled by a flow meter. By establishing a ratio between the flow rate of standardized
milk and that of the added cream, the control systems microprocessor is able to maintain a
constant fat content in the standardized milk.
Flow meter
Skim milk

Milk Standardized milk

Control system Disply of fat content

Separator
Flow
Flow meter
Flowmeter
meter

Cream

Surplus cream
Control valve Control valve
Flow meter

Fig. 8.4 Diagram of the continuous standardization process.

Thus, if we use M1to represent the mass flow of added cream containing a G1 % of fat and
M2 to represent the mass fle of skim milk containing G2 % of fat, and Ms to represent the
flow rate of standardized milk containing Gs % of fat,
We have : Ms = M1 + M2
MsGs = M1G1 + M2G2 i.e., (M1 + M2) Gs = M1G1 + M2G2
Therefore,
M 1 G s − G2
=
M 2 G1 − Gs

Ex. If G1 = 30 % and G2 = 1 %, and the standardized milk must have a fat content of 3 %.
M1 3 −1 2
= = = 0.074074
M 2 30 − 3 27

If M1, the flow rate of skim milk, is 30,000 kg/h. Then, M1, the flow rate of 30 % cream will
be 2222.22 kg/h and the flow rate of standardized milk will be 3222.22 kg/hr

148
Solving the problems on standardization
A D D = B − C or, C − B
E = A − C or, C − A
A = Fat % in first raw material such as whole milk.
C B = Fat % in second raw material such as skim milk.
C = Fat % required in standardized milk.
D = Parts of raw material A.
B E E = parts of raw material B will be needed to produce. F
parts ( D+E) of standard milk of correct composition.
F

To calculate the amount of each type of raw material required per batch of a 1000 litre of
standard milk;
B−C or C − B A−C or C − A
A= × 1000 B= × 1000
F F
Problems on standardization
1. How much cream 32 % fat will be separated from 10,000 L of raw milk with 4.8 % fat?
The skim milk separated has a fat content of 0.1 % and a loss of fat during separation is
0.15 %.
2. Calculate the required quantity of milk having 3.8 % fat and 8.6 % solid-not-fat; cream
having 35 % fat and 5.9 % solid-not-fat, and skim milk powder contains 1.2 % fat and
95.8 % solid-not-fat for the preparation of 5000 kg of super lacto milk with composition 4
% fat and 10.5 solid not fat in the finished product.
3. You have given 1000 Kg of whole buffalo milk testing 7.5 % fat and 9.8 % SNF; SMP
testing 0.5 % fat and 96.5 % SNF. You are required to produce single tonned milk
containing 3.0% fat and 8.5 % solid-not-fat and double tonned milk having 1.5 % fat and
9.0 % solid-not-fat. How much water and skim milk powder should be added to produce
these products?
4. You are required to produce 30 000 L of standard homogenized milk having 3.2 % fat,
8.2 % SNF, from 10 000 L of whole milk having 5.6 % fat, 9.0 % SNF and calculate the
amount of skim milk powder, butter oil and water required.
5. How much Kg of salt required for making salted table butter 80 % fat and 1.5 % salt in
the final product through 5000 kg of cream with 32 % fat and fat in butter milk contains
0.8 %.

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 6. You have given 10 000 Kg of raw milk testing 7.2 % fat, 8.6 % SNF; skim milk powder testing
0.6 % fat, 96.3 % TS. You are preparing tonned milk to having 3.7 % fat and 8.2 % SNF.
Calculate the amount of required quantity of SMP, water and raw milk. SNF/Fat = 2.21
8.13 Heat treatment
In the manufacture of almost all milk and dairy products involves heat treatment. The main
aims of it are killing of microorganisms and inactivation of enzymes. The result greatly
depends on the intensity of the heat treatment, i.e., the combination of temperature and
duration of heating. It is also useful to distinguished between irreversible and reversible
changes. The later are often involved when milk is brought to a high temperature to
facilitate some reaction or process, such as renneting of cheese milk, growth of starter
organisms, efficiency of water evaporation or centrifugal separation, etc.
Heat treatment may also cause undesirable changes, although desirability may depend on
the product involved or its intended use. Examples are browning, development of a cooked
flavor, loss of nutritional quality, inactivation of bacterial inhibitors, and impairment of rennet
ability. Heat treatment should be carefully optimized.
Objectives
The main objectives of heat treatments are as follows:
• Warranting the safety of the consumer
It is mainly concerns with the killing of pathogens like Mycobacterium tuberculosis, Coxiella
burnetii, Staphylococcus aureus, Salmonella species, Lisreria monocytosis and
Campylobacter jejuni. It also concerns potentially pathogenic bacteria that may unintentially
enter the milk. A fairly moderate heat treatment kills all of these organisms. Highly heat-
resistant pathogens either do not occur in milk (e.g., Bacillus anthracis), or they become
readily overgrown with other bacteria (e.g., Clostridium perfringens), or they are pathogenic
only at such high numbers (e.g., Bacillus cereus) that an approaching spoilage of the milk is
detected long before these counts are reached. Some toxins (especially from
Staphylococci) a withstand moderate heat treatment.
• Increasing the keeping quality
It primarily concerns killing of spoilage organisms and of their spores if present. Inactivation
of enzymes, native in milk or excreted by microorganisms, also is essential. Chemical

150
deterioration by autoxidation of lipids can be limited by intense heat treatment. Rapid
creaming can be avoided by inactivating ‘agglutinin’.
• Establishing specific product properties
The examples are
a. Heating the milk before its evaporation to increase the coagulation stability of the
evaporated milk during its sterilization.
b. Inactivating bacterial inhibitors like immunoglobulins and the lactoperoxidase-CNS-
H2O2 system to enhance the growth starter bacteria.
c. Obtaining a satisfactory consistency of yoghurt; and during acidification of milk.
8.13.1 Changes caused by heating
Overview of changes
• Changes may be reversible or irreversible.
• Interest mainly in irreversible or slowly reversible reactions; such changes scarcely
occur at heat treatments of lower intensity than low pasteurization.
• Reversible reactions must be taken into account because they determine the state at
increased temperature, that is, the conditions in the milk at which the irreversible
changes take place.
• Reversible changes include the mutarotation equilibrium of lactose and ionic equilibria,
including pH.
8.13.1.1 Chemical and physical changes
Changes caused by heat treatment are as follows:
1. CO2 and other gases are removed during heating. Loss of O2 is important for the rate of
oxidation reactions during heating and for growth rate of some bacteria afterward. The
loss of gases is reversible, but uptake of air may take a long time.
2. The amount of colloidal phosphate increases and the [Ca++] decreases. Again, the
changes are reversible, though slowly (~24 h).
3. Lactose isomerizes and partly degrades to yield, for instance, lactulose and organic
acids.
4. Phosphoric esters, those of casein in particular, are hydrolyzed. Phospholipids and
some dissolved esters are also split. Consequently, the amount of organic phosphate
increases.

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5. The pH of the decreases, and the titratable acidity increases, mainly due to changes
depend on conditions.
6. Most of the serum proteins are denatured and thereby are rendered insoluble.
7. Enzymes are inactivated.
8. Reactions between protein and lactose occur (Maillard reaction).This involve loss of
available lysine.
9. Free sulphydryl group are formed. This causes a drop of the redox potential.
10. Other reactions of proteins occur.
11. Casein micelles become aggregated. Aggregation may eventually lead to coagulation.
12. Several changes occur in the fat globules membrane, e.g., in its Cu content.
13. Glycerides are hydrolyzed and intersterified.
14. Lactones and methyl ketones are formed from the fat
15. Some vitamins are degraded.
8.13.1.2 Consequences
Usually the main effect of heat treatment is the far slower rate of deterioration due to
microbial and enzymatic action. The most important other effects are as follows:
a. Color: Heating milk at first makes it a little whiter ( no. 2 above).At increasing heating
intensity the color becomes brown; due to (8).
b. Viscosity: May increase slightly due to (6), and much more due to (11). The latter
change especially occurs when concentrated milk is sterilized.
c. Flavor: changes appreciably, mainly due to changes 8,9,10 and 14.
d. Nutritive value: decrease, at least for some nutrients, due to 15 and 8 and may be 10.
e. Bacterial growth: Several bacteria can grow faster in heat treated milk because bacterial
inhibitors like lactoperoxidase-H2O2-CNS and immunoglobulins are inactivated (nos. 6
&7). Furthermore, heat treatment may lead to formation of stimulants for some bacteria
or inhibitors for still other bacteria. All of these changes greatly depend on heating
intensity.
f. Tendency of age thickening and for heat coagulation of concentrated milk may be
decreased.
g. The rennetability decreases.
h. Creaming tendency of the milk decreases mainly caused by (6).

152
i. The proneness to autoxidation is affected in several ways mainly due to 12, 9 and 7.
j. The composition of the surface layers of the fat globules formed during homogenization
or recombination is affected by the intensity of heating before homogenization, mainly
because of change no 6. This affects some properties of products. For example, the
tendency to form homogenization clusters increased.
Heat Available Vitamins
treatment lysine B1 B6 B9 B12 C
o
15 s at 75 C 0 5-10 0-5 3-5 3-10 5-20
15 s at 0 5-15 5-15 5-10 10-20 10-
o
140 C 20
20 min at 5-10 20-40 10-20 20-50 30-80 30-
115oC 60
Note: Loss of vitamins B1, B2 and C depends on the O2 concentration during heat treatment.
Table 8.2 Loss in % of some nutrients, i.e., available lysine and various
vitamins, due to some heat treatments of milk.

8.13.2 Heating Intensity

The intensity of heating follows from the duration of heating and the temperature. The effect
of certain combination of duration and temperature will differ because they depend on the
reaction considered, e.g., inactivation of a certain enzyme or formation of Maillard products.
After all, certain reactions occur fairly quickly at relatively low temperature, whereas others
need a much higher temperature before having an appreciable effect. The dependence of
the reaction rate on temperature varies widely among reactions.
Process of different intensity
Classifying heating processes is the basis of their intensity, special attention is usually paid
to the killing of microorganisms and to the inactivation of enzymes.
The following are customary processes:
a. Thermalization: This heat treatment of lower intensity than low pasteurization, usually 20
s at 60-69oC. The purpose is to kill bacteria, especially psychrotrophs, as several of
these produce heat resistant lipases and proteinases that may eventually cause
deterioration of milk products. Except for the killing of many vegetative microorganisms,
Thermalization causes almost no irreversible changes in the milk.
b. Low pasteurization: This is a heat treatment of such intensity that the enzyme alkaline
phosphate of milk is inactivated. It may be realized by heating for 30 min at 63oC or for
15 s at 72oC. Almost all pathogens that can be present in milk are killed; it specifically
153
 concerns Mycobacterium tuberculosis, a relatively heat- resistant organism that formerly
was among the most, but not all, vegetative bacteria are killed. Furthermore, some
enzymes are inactivated but by no means all of them. Flavored of milk is hardly altered,
little or no serum protein is denatured, and cold agglutination and bacteristatic
properties remain virtually intact. A more heat treatment is, however, often applied (e.g.,
20 s at 75oC). This causes for instance, denaturation of immunoglobulins (hence
decrease in cold agglutination and in bacteristatic activity) and sometimes a perceptible
change in the flavor of milk.
c. High pasteurization: This is a treatment such activity of the enzyme lactoperoxidase is
destroyed, for which 20 s at 85oC suffices. However, higher temperatures, up to 100oC,
are sometimes applied. Virtually all vegetative microorganisms are killed but not
bacterial spores. Most enzymes are inactivated but milk proteinase (plasmin) and some
bacterial proteinases and lipases are not or fully. Most bacteriostatic properties of the
milk are destroyed. Denaturation of part of the serum proteins occurs. A distinct cooked
flavor develops; a gassy flavor if it concerns cream. There are no significant changes in
nutritive value, with the exception of a loss of vitamin C. The stability of the product
toward autoxidation of fat is increased. All the same, only irreversible chemical reactions
occur.
d. This heat treatment is meant to kill all microorganisms, including the bacterial spores.
To that end, 30 min at 110oC, or 1 s at 145oC usually suffices.
The latter two are examples of so called UHT (Ultra-High-Temperature, short time)
treatment. The effect of all such heat treatments are different. Heating for 30 min at 110oC
inactivates all milk enzymes, but not all bacterial lipases or proteinases fully; causes
extensive Maillard reactions, leading to browning formation of a sterilized milk flavor, and
some loss of available lysine; reduces the content of some vitamins, causes considerable
changes in the proteins including casein; and decreases the pH of the milk by about 0.2
unit. Heating for 1 s at 145oC does not inactivate all enzymes, i.e., plasma hardly and some
bacterial lipases and proteinases not all, and therefore such heat treatment is rarely
applied; chemical reactions hardly occur, most serum proteins remain unchanged, and only
a weak cooked flavor develops.

154
e. Preheating: This may mean anything from very mild to quite intense heating. It mostly
concerns heating intensities anywhere low pasteurization and sterilization.
The changes caused by heating the milk
a. Inactivation of inhibitors
1. Immunoglobulin – 1 s 80oC inactivate this class of inhibitors which is largely coincide
with those for the inactivation of cold agglutination. Bacillus cereus is fairly sensitive to
immunoglobulins because IgM causes the bacterial cells to agglutinate and to sediment
to the bottom of the vat.
2. Lactoproxidase system – Most of the lactic acid bacteria are fairly sensitive; most gram
-ive bacteria are not. The system is inactivated by denaturation of the enzyme, but its
activity should be reduced to at least 0.001 because peroxidase is in excess in milk.
The effects of inactivations (1) and (2) are seen, at about 60oC and 70oC, respectively.
3. Bacillus stearothermophillus is sensitive to lactoferrin (the bacterium grows faster in
milk when ferro salts have been added), which appears to be inactivated as a bacterial
inhibitor at a much more intense heating than is needed for its denaturation.
B. sterothermophiphillus hardly grows, in UHT milk, but it does grow in traditionally
sterilized milk. The effect of lactoferin can considerably affect the analysis of the
kinetics of the killing of bacteria.
b. Formation of stimulants: Some lactic acid bacteria, especially the thermophillic ones,
are enhanced by the presence of formic acid, which is formed during intense heating.
This may explains the stronger acid production (more rapid growth) at heating
temperatures above 100oC.
c. Formation of inhibitors or inactivation of stimulants:
For example, Bacillus stearothermophillus has been found to grow more slowly in
moderately heated milk than in raw milk.
d. Killing of bacteriophages: This will rarely play a role because raw milk contains at most
a very low number of phases (e.g., 1 or less per liter).
8.14 Inactivation of enzymes
Heat inactivation of most enzymes follows firsts order kinetics as occurs during
denaturation of globular proteins. The inactivation is strongly temperature dependent, Q10
mostly being at least 50. A D-value of 1 min is usually reached between 60oC and 90oC.

155
But the milk contains some enzymes that can cause spoilage and that show a strongly
differing heat inactivation. After the actual denaturation of the enzyme molecule, at least
one reaction is needed to prevent renaturation of the enzyme from occurring on cooling.
[Renaturaion would mean preserving the enzyme activity]. Various enzymes are
inactivated at far higher heating intensities than mentioned above, and they shows a lower
Q10. The high heating intensity used for the ensuring reactions to proceed because these
enzymes generally are in a denatured state at a temperature of, say, 80oC.
Table 8.3 Heat inactivation of some enzymes in milk.
Enzymes Temp (oC) D (s) Q10
Milk enzymes
Alkaline phosphatase 70 33 60
Lipoprotein lipase 70 20 13
Xanthin Oxidase 80 17 46
Lactoperoxidase 80 4 230
Superoxide dismutase 80 345 150
Catalase 80 2 180
Plasmin 80 360 33
Plasmin 120 30 (1.5)
Acid phosphatase 100 45 10.5
Extracellular bacterial enzymea
Lipase Pseudomonas fluorescens 130 500 (1.3)
Lipase Pseudomonas sp. 130 700 2.4
Lipase Alkaligenes viscolactis 70 30 2.6
Proteinase Pseudomonas flurorescens 130 630 2.1
Proteinase Pseudomonas sp. 130 160 1.9
Proteinase Achromobacter sp. 130 510 2.1
Chymosin 60 25 70
a
The results may vary widely among strains and may also depend on conditions during
growth.
8.14.1 Some enzymes
Lipoprotein lipase of milk is inactivated, to about Q10 at 75oC.
Plasmin is very heat-resistant. Above 110oC, the inactivation rate increases only slightly
with increasing temperature. Even at 140oC, the milk should be heated for at least 15 s to
prevent the occurrence of proteolysis during keeping.
The bulk of the enzyme occur in milk as an inactive zymogen, the plasminogen, which can
be slowly transformed into the active form by the enzyme urokinase, plasminogen also is
very heat-resistant.
156
Bacterial lipases, especially lipases exreated in the milk by some gram negative rods, can
also be very heat resistant.
Bacterial proteinases – especially extracellular endoproteinases of gram negative rods, can
also be very heat resistant.
The incomplete inactivation of lipases, lipolysis may cause a rancid flavor. Residual milk
proteinase especially attacks β- and αs2- caseins. As a result, a bitter flavor may develop,
and skim milk may finally become more or less transparent. Residual bacterial proteinases
mainly attack κ-casein. Consequences may be bitter flavor development, gel formation, and
wheing off.
Most of the bacterial enzymes are insufficiently inactivated by heat treatment because of
their great heat resistance.
8.15 Method of heating
Different ways of heating and/or cooling of liquids can be done with various kinds of
machinery.
Prerequisites for a heating process may be defined as follows:
a. The desirable time-temperature relationship should be practicable. It also involves such
aspects as controllability and reliability, and uniformity of heating. In establishing the
result of the heat treatment (e.g., the sterilizing effect), the times needed for warming
and cooling should be accounted for.
b. No undesirable changes should occur in the product, such as absorption of extraneous
matter (including Cu, Sn, and plasticizers), loss of compounds (e.g., water), disruption or
coalescence of fat globules, coagulation of protein, etc. Sometimes excessive growth of
thermophillic bacteria can occur in plasticizers.
c. The expenses should be low. They partly depend on the price, the life time, and the
maintenance and operating costs of the machinery. Of much concern is the amount of
energy needed for heating and cooling, which may be kept low by regeneration of heat
and cold, respectively. Furthermore, the extent of fouling plays a role. Rapid fouling
causes the heat transfer and the rate of flow to diminish. As a result, consumption of
energy increases significantly. This necessities frequent cleaning, hence brief operating
times.

157
 d. The way of working should fit into the planning. For example, insertion of operations like
configuration or homogenization in the process line may desirable and so may be good
possibilities to adjust heating temperature, heating time, and flow capacity.
A particular apparatus or processing scheme is selected on the basis of:
a. The desirable combination of time and temperature. Heating for 30 min at 68oC requires
other machinery than 1 s at 145oC.
b. Properties of the liquid. The main factor involved is the heat transfer rate which, in turn,
depends on the thermal conductivity and especially on the viscosity. A part from that, the
tendency to exhibit fouling is of importance. Generally, highly viscous products show
poor heat exchange.
c. Requirements for the prevention of recontamination and for ensuring process steps,
especially packing.
An additional factor in the selection may be the effect of the method of heating on the air
content, especially the O2 content of the milk. The O2 content affects the possibilities for
growth of several bacteria. For example, Bacillus types need some O2, lactic acid bacteria
are slowed down at high O2 pressure. The O2 content of long-life milk products may affect
the development of off-flavor by fat autooxidation.
Holder pasteurization causes significant deaeration, but air can be reabsorbed during
cooling. Heating in a heat exchanger does not affect the O2 content, unless a special
deaerator (e.g., a flash cooler) is connected, as in direct UHT treatment. The extent of
deaeration during autoclaving depends on the type of sealing applied. For e.g., bottles fitted
with crown corks lose most of the air, but sealed cans lose most of the air, but sealed cans
lose nothing.
8.15.1 Equipment
Liquids can be heated and cooled in a batch process, in a heat exchanger, or in a packed
form. Originally, batch processing was in general use for pasteurizing beverage milk. It is
the so-called holder pasteurization, e.g., 30 min at 63oC.
The process still use in the manufacture of starters, whipping cream and other small scale
products. A jacketed vats involved are fitted with an agitator; through the double jacket,
steam or hot water circulate followed by cold water.

158
Advantages
• Simple
• Flexible
• Satisfactory temperature control (little fluctuation in temperature unless highly viscous
liquids are used).
Disadvantages
• Warming and cooling times are too long (excessive for large containers).
• Regeneration of heat is not well possible and
• Connection to continuous process is awkward (clumpsy).
Currently, flow through heaters or heat exchangers are commonly used. Heat water or
condensing steam constitutes the heating medium. Sometimes, vacuum steam heating is
applied to minimize the difference in temperature with the liquid to be heated.
In plate heat exchangers, a large heating surface is assembled in a confined space and on
a small floor area. Heating agent and incoming liquid are present in thin layers and are
separated by a thin wall, i.e., a plate. Because of the large heating surface per unit volume
of liquid that is to be heated can be small, e.g., 2oC when milk is heated from 65oC and
75oC. This may be an advantage for heat sensitive products, where fouling of the heat
exchanger is greater for a higher wall temperature. Furthermore, warming and cooling
processed rapidly in plate heaters. Another advantage is that the energy consumption (for
heating and cooling) can be relatively small because heat can be regenerated. The
principle is shown in Fig. 8.4 below.
Holder
Product out
Regeneration
35 section

20 85 85

10 70 95
25 20
80
Hot water
Cooling water Product in

Fig 8.5 Simplified diagram of a heat exchanger for heating and cooling of liquids, showing
the principle of regeneration. The numbers are temperatures (oC) and merely give an

159
 example. Thick walls denote insulating walls; thin lines are walls allowing rapid heat
transfer.
When the milk comes in the heat exchanger, it is heated by milk that has already been
heated and that is at the some time being cooled by the milk coming in. The later is
subsequently being heated further by hot water (or steam). It may then flow through a
holder section, to achieve a sufficient heating time. After being cooled by the incoming milk,
it is cooled further by means of cold water (or another cooling agent). Note that the liquids being
heated and cooled are always in counterflow and and that the temperature difference between the
two remains constant.
Drawbacks
• Vulnerability
• Rubber gasket can become leaky, they do not resist high temperatures and should occasionally
be changed.
• Cracks can be formed in the plates. As a result, small amounts of raw milk might leak into the
milk already pasteurized, e.g., in the regeneration section, possibly with detrimental results on
bacterial quality.
• Plate heaters are only fit for heating liquid products.
• To attain a high enough speed through the machinery, highly viscous liquids require such a high
operating that leakage may occur. Moreover, heating to above 100oC requires special
construction because it involves pressures over 1 bar.
A plate heat exchanger is made up of various sections connected in series, including regeneration
section, heating section, holding section (may also be tube), and cooling sections. Each section
consists of great number of plates, being partly parallel and partly connected in series. In this way
the liquid is properly distributed among the plates and arrives at a high enough speed to reduce
fouling. The plates are shaped in such a way as to greatly enhance turbulence in the liquid. This
enhances heat transfer and diminishes fouling.

160
 Cooling sections Regeneration sections Pasteurizer

72
25 25 45 45 Flow diversion valve
Pasteurized milk 5 15 15 72
40 40 67 67
20

e.c.

Cooling agent Cold water Holder

Centrifuge
Heating agent
e.c. = extra cream
Temperature in oC Homogenizer
Insufficiently
heated milk

Fig 8.6 Shows the different section of plate heat exchanger (a continuous pasteurizer).
Tubular heat exchangers generally have a smaller heating surface per unit volume of liquid
than plate heat exchangers. Accordingly, the difference in temperature between the heating
agent and the incoming liquid is greater. To restrain fouling and entrance heat transfer, high
flow rates are used, which necessitates high pressures. But this causes no problems
because tubes are much stronger than plates; after all, tubular heat exchangers have no
sealing gaskets but mostly have (spirally bent) concentric tubes. It can readily be applied to
obtain very high temperatures (e.g., 150oC). Accordingly, they are excellently fit for indirect
UHT treatment. Like a plate heat exchanger, a tubular heat exchanger can be built of
regeneration, heating, holding, and cooling sections.
In modern heat exchangers, the milk may be in counter flow with water throughout the
apparatus. The water is kept circulated and is heated by means of indirect steam heating,
immediately before it should heat the milk to the maximum temperature desired (Fig. 8.6)
Often up to about 90 % heat regeneration is achieved.
Advantages:
• Temperature control,
• Rapid and even heat transfer, and
• Energy saving.

161
 water Cold water

Steam
Heat exchanger
Holder Aseptic
Raw milk 25 5 packaging
5 115 140 70

Fig. 8.7 The flow diagram of a heat exchanger in which the milk is heated and cooled by
water.The milk temperature in oC.
UHT treatment with direct heating there is no wall between the heating agent and the liquid
to be heated, but the heating agent (steam) is injected in to the desired temperature (e.g.,
in 0.1 s from 80oC to 145oC) occurs, provided that the steam have immediately condense.
For this to happen, finely disperse steam and a significant back pressure in the liquid are
needed. The incoming liquid becomes diluted with the condensing steam. Of course, the
steam has to be of high purity. After maintaining the liquid at the desirable temperature for
few seconds, it is discharged in a vessel of reduced pressure. Here almost instantaneous
evaporation of water occurs, causing very rapid cooling. The amount of water evaporating
should equal the amount of steam have been absorbed before.
150
2
Temperature (oC)

100

50

0
0 100 200 300 400 500
Time (s)

Fig. 8.8 Temperature of milk versus time during heat treatment. 1= direct
heating; 2 = indirect heating
Steam injection heating causes disruption of fat globules and some coagulation of protein.
Homogenization (aseptic) at high pressure redisperses the coagulum. Without homogeni-

162
zation, the product gives the inhomogeneous astringent or non-smooth sensation in the
mouth, and a sediment tends to form in storage.
The heating section (upto e.g., 80oC) that precedes the steam injection, and the cooling
section connected after the “ flash cooling” (starting from, e.g., 80oC), may be plate heat
exchangers. In Fig.8.8, the hold up time of the milk above say, 80oC is very short, i.e.,
insufficient to inactivate milk proteinase.
The so-called vacreator is quite similar to a direct UHT heater, but in this apparatus the
liquid is heated to pasteurization temperatures. The evaporative cooling in vacuum, aimed
at removal of volatile flavor components. Such apparatus is occasionally used for
pasteurizing cream for butter manufacture.
Disadvantage
• The considerable damage (coalescence and disruption) to fat globules.
• Limited heat generation.
8.15.2 Heat regeneration
Regeneration is the regaining of heat and the saving of cooling energy in the combined
processes of heating and cooling. The regenerated effect is the heat absorbed in the
regeneration section as a percentage of the total heat absorption. If the specific heat is the
same for all liquids at all temperatures, the regenerating effect can simply be calculated
from:
Temp after heating up in regeneration sec tion − inlet temp
Heat regeneration = × 100 %
Pasteuriza tion or sterilizat ion temp. − inlet temp
Theoretically, as much as 100 % regeneration might be achieved, but this would need an
infinite heating surface and absence of heat loss to the surroundings. Loss of heat does,
however, occur, and the surface area is by no means infinite. It is often amounts to about
90 % regeneration.

163
8.16 A continuous bottle sterilizer
b

40
70

80
110

Bottle supply 30 c
Bottle discharge a

Fig.8.9 Diagram of continuous bottle sterilizer. a = water seal in the bottle intrance and
preheating section; b = steam space with adjustable pressure; c = water seal in the
bottle discharge and first cooling section; d = second cooling section. Indicated
temperature(oC) refer to that in the bottles. The water is partly circulatd.
mostly concerns Streptococcus thermophillus (maximum growth temperature about 53oC).
But S. faecium (= S. durans, maximum temperature ~ 52oC) and S. faecalis (maximum
~47oC)
8.16.1 Control measures during heat treatment
1. Heating intensity: it may be insufficient because steam supply, heating temperature,
may fluctuate or because of a sudden increase in fouling. A pasteurizing plant has flow
diversion valve. The milk flow back to the supply pipe if the pasteurization temperature
decreases to below a preset limiting valve. Alternatively, an automatic “pump stop” may
be applied. Moreover, the heating temperature should be recorded continuously. The
risk of too brief a heating time will be slight, the volumes in the heat exchanger

164
 (especially the holder) are fixed, and it is almost impossible that the milk pump would
suddenly run faster.
2. Recontamination: Raw or insufficiently heat treated milk may gain entrance in to the
heated milk, e.g., because of the leaky heat exchanger or because a mistake is made
in connecting pipes. Naturally, contamination occurs when milk passes through a
machine or pipe that is not absolutely clean. Recontamination should be rigorously
avoided in UHT treatment, because it is usually combined with aseptic packing; it
specifically concerns the homogenizer. One bacterium per 1000 L of milk may cause
unacceptable spoilage.
3. Bacterial growth: Growth of bacteria may occur in heating equipment, e.g., in a batch
pasteurizer. Especially in a vessel such as balance tank through which milk flows while
it maintains a relatively high temperature for some time, organisms like Bacillus
sterothermophillus ( maximum growth temperature ranging from 65 to 75oC) and B.
coagulans, maximum temperature 55 to 60oC) can grow. As a result the counts of
these bacteria in raw milk are very low. Accordingly, the contamination will become
perceptible only after many hours. Obviously, occasional cleaning and disinfection of
the machinery can overcome these problems.
After having been in use for hours, a pasteurizer may contain growing bacteria in that
part of the regeneration section where the milk is being cooled. The bacteria involved
survive the milk pasteurization and may colonize on the metal surface of plates or
tubes that show some fouling by milk components; such a layer of bacteria and milk
components is called a biofilm. Bacteria in a biofilm can grow rapidly, so that
significantly increased counts in the pasteurized milk may be found after, say, 10 h of
continuous use of the apparatus. It may also cause problems. Timely cleaning is the
obvious remedy.
8.17 Cream Separation
8.17.1 Principle
The basic principle of cream separation, whether by gravity or centrifugal method, is
based on the fact that milk fat is lighter than skim milk portion. At 16oC, the average density
of milk fat is 0.93 and skim milk is 1.036. Hence milk is subjected to either gravity or a

165
centrifugal force, the two components viz. cream and skim milk by virtue of their differing
densities, separate from one another.
Cream is concentrated form of milk fat. As a rule cream as sold for household use
contains from 20 to 25 % fat. Cream having higher fat percentages are called “whipping
cream”. Cream as received at butter factories usually contains from 25 to 40 % fat.
8.17.2 Method of separation
There are two methods of separation of fat from milk. These methods are as follows.
8.17.2.1 Gravity Methods
When the milk is allowed to stand for sometime, there is a tendency for the fat
globule to rise up. The velocity or rate, at which the globules rise is given by the following
equation which is known as stokes law.

2G (d s − d f ).r
2

V=
9 n
Where,
V = Velocity or rate at which a single fat globule rises.
G = acceleration due to gravity.
ds = density of milk.
df = density of fat.
r = radius of fat globule.
η = viscosity of the skim milk.
In practice the important factors affecting are
1. Size of fat globules: As the size of the fat globule increases, the rate at which cream
rises also increases.
2. Temperature: As the temperature increases, viscosity of milk decreases and hence
velocity of fat globule rise increases.
3. Clumping: A clump or cluster acts like a single globule, radius of fat globules increases,
velocity of fat globule rise also increases.
4. Addition of adhesives: Ultimately helps in increasing the rate at which fat globules rise.

166
8.17.2.2 Centrifugal Methods
When the milk enters the rapid revolving bowl of the cream separator, it immediately
subjected to a tremendous force acting outwards (centrifugal force), which is 3000-6000
times greater than the gravitational force. While both fat and skim milk are subjected to the
centrifugal force, the difference in density affects the heavier portion (skim milk), more
intensely than the lighter fat portion (cream). Thereby the skim milk is forced to the
periphery while the fat portion moves towards the centre. The skim milk and cream both
form vertical walls within the bowl and are separated by being lead through separate
outlets.
The equation given by the Stoke’s law, which is applied in centrifugal separation is follows.
(d s − d f )
V = r2 .N 2 .R.K
n
Where, V = velocity of moment of a single fat globule.
r = radius of the fat globule.
ds = density of the skim milk.
df = density of the fat.
R = distance of the fat globule from the axis of rotation.
N = speed of bowl (rpm).
K = constant.
η = viscosity of the skim milk.

167
 SKIM MILK CREAM
CREAM

WHOLE MILK

SKIM MILK A

WHOLE MILK

B
Fig 8.5 A. Flow of cream and
skim milk in the space between
disces in a centrifugal separator.
B. A disc stack.
C. Photograph of a separator
disc showing holes for
chanelling of milk and spacers
(caulks).
C

Comparison of gravitational and Centrifugal method

S.N. Particulars Gravity Method Centrifugal Method
1 Nature of force causing separation Gravitational Centrifugal
2 Speed of separation Extremely slow Partially instantaneous

3 Direction of movement of fat and

Vertical Horizontal
skim milk particles
4 Microbial quality of cream and skim milk Low High
5 Fat % of cream 10-25 18-85
6 Skim milk 0.2 or above 0.1 or less
7 Scale of operation Small Large
8 Fat % recovered in cream Not more than 90 99.0-99.5

168
Factors influencing the fat percentage of cream
• Position of the cream screw or skim • Mechanical condition of machine (
milk screw. vibration of the disc, amount of slime).
• Fat percentage in milk • Size of the fat globules.
• Speed of the bowl. • Degree and temperature of agitation.
• Rate of inflow of milk. • Presence of air in milk.
• Temperature of the milk. • Acidity of the milk.
• Amount of water or skim milk added
to flush the bowl.
Skimming Efficiency
It refers to the % total fat from milk recovered in the cream. The higher the fat % in milk
and greater the fat loss in skim milk, the lower the skimming efficiency and vice versa.
Example 1 : 100 kg milk testing 7.5 % fat, cream produced 14.1 Kg having 52.5 %.
Calculate the SE.
52.5
14.1 × ( )
S .E. = 100 × 100 = 98.7
7.5
100 × ( )
100
Yield of Cream
fm − fs
THIS IS GIVEN AS C=M
fc − fs
Where, C = weight of cream (Kg)
M = weight of milk (Kg)
fm = fat % of milk,
fs = fat % of skim milk.
fc = fat % of cream.

169
Yield of skim milk
fc − fm
S=M
fc − fs
This is given as S = Weight of skim milk (Kg)
Fat recovery in cream
% fat in cream = (kg fat in cream / kg fat in milk) ×100.
% fat in skim milk = (kg fat in skim milk/ kg fat in milk) × 100
Working Principle of cream separator
The centrifugal force (Fc) acting on an object of mass m, rotating in a circular path of radius
R, at an angular velocity of ω is
π N
FC = M R Ω2 AND, ω =
30
where, N = rotational speed in rpm.
Thus the magnitude of the centrifugal force depends on the radius of rotation, speed of the
rotation and mass of the body (density × volume). For unit volume of material, density
becomes the factor affecting centrifugal force,
FC = (V × R × ω2) ρ
If volume, radius and rotational speed of cream separator are constant ( during the
operation), the milk fat having low density experience less force where as skim milk having
high density experience greater force. Therefore, the fat displaced towards the center of
rotation and skim milk displaced towards the outer wall of the bowl, these two forming inner
and outer ring.
8.17 Bactofugation
Bactofugation is a process in which a specially designed centrifuge called a Bactofuge is
used to separate microorganisms from milk. Originally the Bactofuge was developed to
improve the keeping quality of market milk. At the present time bactofugation is also used
to improve the bacteriological quality of milk intended for other products like cheese, milk
powder and whey baby food. Bacteria, especially heat resistant spores, have a significantly
higher density than the milk. A Bactofuge is therefore a particularly efficient means of
ridding milk of bacteria spores are also resistant to heat treatment, the Bactofuge makes a
useful complement to thermization, pasteurization and sterilization.
170
Bacteria, and especially bacterial spores, can be separated at very high centrifugal force
and high temperature in a specifically designed centrifuge, called a bactofuge. A double
treatment at approximately 73oC leads to three decimal reductions, slightly more for
bacterial spores. The method is expensive, partly because a small percentage of milk solids
becomes incorporated in the sludge. The sludge is removed continuously and often is
readded to the milk after sterilization. It has been suggested that the method be used for
the removal of spores of Bacillus cereus from pasteurized beverage milk. It is employed to
reduce the number of spores of Clostridium tyrobutyricum, which can cause late blowing of
cheese even if present in small numbers. That is why cheese milk is bactofugated.
The original Bactofugate was a solid bowl centrifuge with nozzles in the periphery of the
bowl. It was long considered necessary to have a continuous flow of the heavy phase,
either through a peripheral nozzle or over the heavy phase outlet of the Batofuge, to
achieve efficient separation. This was possibly true of the old solid bowl centrifuges with
vertical cylindrical walls, but in modern self-cleaning separators with a sludge space outside
the stack, bacteria and spores can be collected over a period of time and intermittently
discharged at preset intervals.
There are two types of modern Bactofuge:
1. The two phase Bactofuge has two outlets at the top: one for continuous discharge of
bacteria concentrate (bactofuge) via a special top disc, and one for the bacteria-
reduced phase.
2. The one phase Bactofuge has only one outlet at the top of the bowl for the bacteria
reduced milk. The Bactofugate is collected in the sludge space of the bowl and
discharged at present intervals.
The amount of bactofugate from the two-phase Bactofuge is about 3% of the feed, while
from one phase can be as low as 0.15 % of the feed.
Bactofugate always has a higher dry matter content than the milk from which it originates.
This is because some of the larger casein micelles are separated out together with the
bacteria and spores. Higher bactofugation temperature increases the amount of protein in
the bactofugate. Optimal bactofugation temperature is 55-60oC.

171
8.18 Homogenization
It refers to the process of forcing the milk through a homogenizer with the object of
subdividing the fat globules. In efficiently homogenized milk, the fat globules are subdivided
to 2 microns or less in diameter.
Homogenized milk is a milk which has been treated in such a manner as to insure breakup
of the fat globules to such an extend that after 48 hrs of quiescent storage at 45oF; no
visible cream separation occurs on the milk.
8.18.1 Objectives of homogenization
a. Counteracting segregation, for the most part creaming.
To achieve this, the size of the fat globules should be greatly reduced. The
sedimentation of cocoa particles; the homogenizer can reduce these particles.
b. Improving stability towards partial coalescence.
The increased stability of homogenized fat globules is caused by the reduced diameter
and by the acquired surface layer of the fat globules. Moreover, partial coalescence
especially occurs in a cream layer, and such a layer forms much more slowly in
homogenized products. Prevention of partial coalescence usually is the most important
purpose of homogenization, a cream layer per se is not very inconvenient because it
can really be re-dispersed in the milk.
c. Creating desirable rheological properties.
Formation of the homogenization clusters can greatly increase the viscosity of a
product, e.g., cream. Homogenized and subsequently soured milk (e.g., yoghurt) has a
higher viscosity than un-homogenized milk. This is because the fat globules that are
now partly covered with casein participate in the aggregation of the casein micelles.
d. Recombining milk products.
A homogenizer, however, is not emulsifying machine. Therefore, the mixture
concerned should first be pre-emulsified, e.g., by vigorous stirring, the formed coarse
emulsion is subsequently homogenized.
Merits and Demerits
Merits
• No formation of cream layer.
• Fat in milk does not churn due to rough handling or excessive agitation.

172
 • More palatable, heavier body and richer flavor.
• Produces soft curd and is better digested.
• Less susceptible to oxidized flavor development.
Demerits
• Increased cost of production.
• Returned homogenized milk difficult to salvage fat recovery
• Sediment appears to a greater degree.
• Curdling in cookery.
• More susceptible to production of activated or sunshine flavour defects.
• Greater tendency for milk “seepage” through bottle cap.
8.18.2 Operation of homogenizer
The homogenizer is a piston type of pump. It consists of high pressure pump which forces the milk
at high pressures and velocity through a narrow opening between the homogenizing valve and its
seat; the fat globules in the milk are thereby subdivided into smaller particles of uniform size.
Homogenizers are either single stage or double stage. This is a machine which causes the sub-
division of fat globules. It is designed to subject a liquid milk product to a high pressure then allow
the product to pass through a special valve. This process breaks up the fat globules, increases the
ability of the protein to hold water hydration, and slightly softens the curd.
The diameter of the globules in unprocessed milk varies with breed and stage of lactation of the
producing animal, and other factors, averaging about 3 µ, those in homogenized milk might
average only 1 or 2 µ depending, among other things, upon the pressure used. Such division
greatly increases the number of globules and total globules surface area; it reduces the average
volume of fat per globule. One hundred kg of milk testing 5.0 % fat in globules averaging 3 µ in
diameter might contain approximately 3.78×1014 globules. If homogenization reduces the average
globule diameter to 2 µ, the number of globules will increase about 3.0 times, to about 1.14 × 1015
globules. While their total surface area before homogenization is nearly 1.070 m2, after
homogenization this area becomes nearly 15.750 m2, an increase of about 15 times.
The milk fat must be liquid for the effective homogenization. The temperature should be 30oC and
40oC; the exact temperature will vary with the composition of the fat. Ice cream mix must be
homogenized. This assures uniform distribution of the fat throughout the product, prevents
churning in the freezer, and gives the product a smooth and creamy body and texture. The
improvements in texture are partly a result of the affect on proteins. They are then able to hold
more water as bound water. This contributes to the formation of smaller ice-crystals during
freezing.
173
Whole milk must be homogenized for fluid milk sale. Homogenization divides the fat
globules and distributes them throughout the milk; cream does not rise in such milk. The
flavor is fuller and creamier. The curd is slightly softer.
Pressure used in homogenization varies with the product or the purpose of the process.
Single stage homogenization aggregates the clumping of the fragmented fat globules.
Homogenizing in two stages breaks up these clumps to a great degree. The two stage
process is advised for both fluid milk and ice cream mix. Whole milk for fluid sale is
homogenized at pressure of 106 to 212 kg/cm2 on the first stage plus another 35-70 kg/cm2
on the second stage. Ice-cream mix homogenized at 140 kg/cm2 on first stage plus 70
kg/cm2 on the second stage.
8.18.3 Factors affecting homogenization
j. Temperature of homogenization: The milk should be at a temperature above the melting
point of fat (33oC) at the time of homogenization. This is because fat should be in the
liquid state for proper subdivision. The enzyme lipase should be inactivated prior to
homogenization or immediately afterwards. This can be achieved by heating the milk
55oC or above (65-70oC) for homogenization. The temperature of 38-49oC is the danger
zone of lipase activity and should be avoided during or after homogenization.
Homogenizing valve

Breaker ring

Homogenized
product out

Valve sheet

Homogenizing valve
Unhomogenized
product in

Fig. 8.10 Detail of cross section of a modern homogenizing valve with cone type.
k. Pressure of homogenization: In a single stage up to 6 % fat milk, usually 2000-2500
psi pressure is sufficient. Higher pressures may increase the tendency for milk to curdle
when cooked, due to the increased destabilizing effect on milk proteins. For liquid

174
products with more than 6 % fat, two stage homogenizer is needed to prevent fat
clumping; 2000 psi at the first stage and 500 psi at the second.

Handle
Adjustable valve second
stage
Second stage valve

500 lbs homogenized

product out
first stage
adjustable valve
Unhomogeni
zed product Handle
in with high 2500 lbs
pressure First stage
valve

Fig. 8.11 Diagram showing the principle of the two stage homogenizer.

Homogenized fat globule –

0.1-3 µ /Average = 2 µ

o
F rise = psi / 318 for milk
Time = about 1/10,000 sec.

Globule deformation
V= 300-800 ft/sec
Valve

Unhomogenized
fat globule size
=Ф1-10 µ Average
= Ф3.5 µ
Velocity = 1-10 ft/sec

FIG. 8.12 Showing deformation of the fat globule in the homogenizer slit.

175
 Membrane 2-3 % Homogenized fat
Normal
globule, 25% of
Homogenization casein
Fat membrane protein
globule Partial transfer of
phospholipids 10-20 % fat interacts with
Partial transfer of casein to give a high density
casein from skim products.
Skim milk Partially hydrophobized fat globule
increases in frequency with an
increase in fat content, rarely found in
well homogenized milk.

FIG 8.13 Diagrammatic representation of the homogenization process.

8.18.4 Theory of homogenization
The disrupting action takes place as a result of a shearing action between the globules as
they flow through a passage at high velocity, much as rocks are sheared and worn by the
action of a fast moving water. The solid particles nearest the edge of the stream are
retarded somewhat by the friction of the fluid on the banks of the stream, and the center or
fast moving part of the stream therefore carries the particles nearest in the center at a more
rapid place than those nearer the edge. This difference in speed causes the solid particles
to grind against each other with a shearing effect, resulting in the reduction in size of the
particles. The faster the flow and narrower the stream, the greater is the shearing action.
The principal action in the homogenizer is the same nature; because of the extremely high
velocity and the minute orifice the shearing action is very great. In addition to the above,
there is also a certain disrupting action due to impact which takes place when the high-
velocity stream strike a solid surface as the breaker ring, for example, in same types of
valves. Then there is undoubtedly drop in pressure or explosive effect as the fluid leaves
the valve. Studies by Loo indicate that forces caused by collapse of bubbles due to
cavitations may also be an important factor in homogenization. In actual practice, most
valves employ a combination of the three principles;
• The size of the orifices;
• The shape are determined by the volume to be handled in a given time, and
• Also by the viscosity of the product.

176
The best results are obtained from a homogenizer valve when the fluid is forced through it
under steady pressure. This is because the shearing effect of the valve changes with the
velocity of the fluid passing through it and for a certain valve can be correct at only a given
velocity. With fluctuating pressure the velocity will fluctuate, thus causing irregular results.
8.18.5 Factor affecting the fat globule size
Type of homogenizer
It depends on the construction of homogenizer valve. For the same pressure the passage
time will differ. The spread in conditions will differ.
a. Homogenizing pressure
The size distribution of the fat globules depends only a little on pressure.
b. Two-stage homogenization
The milk first passes the “usual” homogenizer valve by which the pressure is reduced from
20 to 5 MPa (the minimum pressure inside the valve equals zero). By the second
homogenizer valve the pressure is reduced to about 1 bar (0.1 MPa). There is no significant
homogenization in the second valve slit. Accordingly, the second stage influence on the fat
globule size is small. In other words, one stage homogenization at 20 MPa leads to a result
similar to that in two stages at 20 MPa and 5 MPa, respectively. The result is worse if the
pressure drop in the second stage increases to more than about 30 % of the total pressure
drop.
c. Fat content and ratio of amount of surfactants (usually protein) to amount of fat
If insufficient protein is available to cover the nearly formed fat surface, the average
diameter of the fat globules and the relative distribution width will be larger. In cream, the
time needed for formation of adsorption of adsorption layers is longer than in milk; on the
other hand, the average time between encounters of one droplet with another is much
shorter. As a result, in cream far more recoalescence of newly formed droplets can occur.
d. Temperature
Homogenization is usually done at temperatures between 40oC and 75oC. Homogenization
is poor if the temperature is so low that part of fat is crystalline. Further increased in
temperature in temperature still has a small effect, presumably because the viscosity of the
dispersed faction decreases somewhat.

177
e. Proper operation of the homogenizer.
A varying pressure (caused by leaking valves, etc.), a worn homogenizer valve, and air inclusion all
may work out badly. Air inclusion and wear of the homogenizer valve should be rigorously be
avoided. If the liquid contains solid particles like dust or cocoa, the valve may quickly wear out,
resulting in unsatisfactory homogenization.
8.18.6 Effect of homogenization
The average fat globule size may be derived from specific turbidity measurements at long
wavelength (e.g., 1 µ) after the milk has been diluted and the casein micelles dissolved. In this way
the homogenization effect can be evaluated rapidly and simply. In principle, continuous
determination is possible. In actual practice, however, an accelerated creaming test is usually
done. A certain quantity of milk is centrifuged and the fat content of the resulting skim milk
determined.
8.18.6.1 Surface layers
The newly formed membranes are predominantly composed of micellar casein and serum protein.
Some of the casein micelles in the layer are present as such, but most are more or less spread out
into micelle fragments or a layer of submicelles. The spreading occurs if a micelle touches a
denuded oil-water interface. (It is sometimes assumed that the micelles are disrupted by the
homogenizer and that the resulting micelle fragments are subsequently adsorbed onto the fat
globules, but this hardly occurs; compare the size of the micelles with the scale of the smallest
eddies. The spreading occurs in a time scale of the order of 0.1 µs.

Submicelle
Serum protein
Casein micelle
FAT
100 nm

Fig. 8.14 New surface layer of fat globules.

178
8.18.6.2 Stability
Plasma protein, predominantly casein, covers a large part (up to about 90 %, in recombined
milks 100 %) of the surface area of homogenized fat globules. This makes the globules
behave to some extent like large casein micelles. Any reaction that causes casein micelles
to aggregate, such as renneting, souring , or heating at very high temperatures, will also
cause the homoge-nized fat globule to aggregate. Moreover, aggregation will take place
more quickly because homogenization has increased the apparent casein concentration
(i.e., the casein concentration effective in aggregating).
To altered surface layer due to:
• The addition of emulsifiers before homogenization may increase stability to heat
coagulation.
• Homogenized fat globules generally show a high stability toward partial coalescence
because they are so small. Moreover, surface layers that consist fully of plasma protein
(as in recombined milk) yield a very high stability to the globules. The addition of
surfactants (emulsifier) displace protein at the oil-water interface can considerably
decrease the stability to coalescence. This plays a part in a product such as ice cream.
• Sour products (yoghurt, sour cream) and cheese made of homogenized milk (or cream)
have different rheological properties from those of unhomogenized milk; this is caused
by the fat globules becoming part of the casein network.
8.18.7 Homogenization clusters
On microscopic examination one sees large agglomerates of fat globules rather than single
globules in homogenized cream. These so-called homogenization clusters contain very
many fat globules, up to about 105. Because the cluster contain interstitial liquid, the
effective volume fraction of particles in the cream is increased, and hence also its(apparent)
viscosity. Adding casein micelle-dissolving agents can disperse the clusters. The fat
globules in cluster are interconnected by casein micelles.
8.18.7.1 Formation of homogenization clusters
During homogenization, when a partly denuded fat globule collides with another globule
that has been covered with casein micelles, such a casein micelle can also reach the
surface area of the former globule. As a result, both fat globules are connected by a
“bridge” and form a homogenization cluster. The cluster will immediately be broken up

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again by turbulent eddies. If however, too little protein is available to fully cover the newly
formed fat surface, clusters are formed from the partly denuded fat globules just outside the
valve slit of the homogenizer, where the power density is too low to disrupt the cluster
again.
The conditions promote for the formation of homogenization clusters:
• High fat content
• Low protein content.
• High homogenization pressure
• A relatively high surface excess of protein, this is promoted by a low homogenization
temperature (less rapid spreading of casein micelles), intense preheating (little serum
protein available for adsorption) and, again, a high homogenizing pressure.
Clustering due to homogenization does not occur in a cream with less than 9 % fat,
whereas it always does in a cream with more than 18 % fat. At intermediate fat contents,
clustering closely depends on pressure and temperature of homogenization.
Clusters can be disrupted again to a large extent (but not fully) in a two stage homogenizer.
In the second stage the turbulent intensity is too low to disrupt fat globules, and hence to
form new clusters, whereas clusters are disrupted; this goes along with some coalescence.
Two stage homogenization of high fat cream (e.g., 30 % fat) insufficiently breaks up
homogenization clusters.
8.18.8 Other effect of homogenization
Homogenizing milk that contains lipase strongly enhances lipolysis. Raw milk turns rancid
within a few minutes after homogenization. This should be explained by lipoprotein lipase
being capable of penetrating the membrane formed by homogenization, but not the natural
membrane. Accordingly, raw milk homogenization should be avoided, or the milk should be
pasteurized immediately after homogenization in such a way that the lipase is inactivated.
Homogenization is often done before pasteurization, since in the homogenizer the milk may
readily be contaminated by bacteria. Furthermore, mixing of homogenized milk with raw
milk should be prevented again to avoid lipolysis.
Homogenization of milk has several other effects:
• The color becomes whiter.
• The tendency to foam increases somewhat.

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• The proneness to fat autoxidation, and hence to the formation of ensuing off-flavors, is
reduced.
• The fat globules lose their ability to be agglutinated upon cooling. This is caused by
inactivation of the cold agglutinin rather than by changing the fat globules;
homogenization at very low pressure (1 MPa) can be inactivated also; to achieve this, a
higher pressure is required, e.g., 10 MPa.
8.19 Creaming
An important purpose of homogenization usually is to slow down creaming and thereby to
prevent partial coalescence. The purpose is primarily achieved by reducing the fat globules
in size. The creaming will be much faster when the fat globules are somehow aggregated.
The following are causes of aggregation:
• Cold agglutination: Fat globules in most homogenized products cannot flocculate in the
cold.
• Homogenization clusters: These are rarely formed, except during homogenizing of
cream.
• Heating at high temperature (sterilization) can cause small clusters of homogenized fat
globule to be formed, especially in evaporated milk; this is the beginning of heat
coagulation.

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Chapter 9

Milk for liquid consumption

Liquid milk can be delivered to the consumer after various heat treatment: none (raw milk),
pasteurized or sterilized, and either packed or not (although sterilized milk is, of course,
always packed). The properties of liquid milk that require the most attention are safety to
the consumer, shelf life, and flavor. Safety is, of course, essential and consumption of raw
milk cannot be considered safe. Consequently, the delivery of raw milk is prohibited or
severely curtailed in many countries. Likewise, delivering milk that is not packed may
involve a health hazard.

9.1 Pasteurized milk

The pasteurization process involves heating milk or other fluid products to a predetermined
tempera-tures, holding the product for a definite period of time and cooling the same
immediately to sufficiently low temperature 4.4oC and storing at low temperature to arrest
bacterial growth till relea-sed for sale. When additional solids are present (chocolate milk,
ice-cream mix), higher temperatures are required for an equvalent microbial destruction. It
is also necessary to cool immediately to less than 10oC.
Pasteurization is practiced in the modern dairy plant to make milk and dairy products safe
for human consumption by destroying any pathogenic organisms which may be present. It
improves the preservation quality of the dairy products so that they they will reach the
consumer in better condition. It also helps to retain the good flavour over a longer period of
time.
The pasteuring process as develoved by Louis Pasteur consists, in principle, of heating the
product to a predetermined temperature temperature and holding it until all or nearly all
objectionable organi-sms, which may be present, are killed. The original works was done
with wine.For pasteurising milk, apparatus must be provided which will heat each and every
particle of milk to the proper temp-erature, hold it for the required length of time, and do it
accurately, for overheating will impair the flavor and affect other physical characteristics.
United States Public Health Service defines pasteurization for milk milk products as the
process of heating every particle of the product to 63oC, and holding it continuously at that

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temperature for at least 30 minutes; or to at least 72oC and holding at that temperaturefor at least
15 seconds.
Type of pasteurizers: Pasteurizing units are classified broadly as follows:
1. Long hold or Vat with single or multipurpose operations – LTLT.
2. High temperature short time (HTST) continuous plate type for single or combined operations.
Common time temperature relationship for milk, cream, ice-cream mix etc are given below.
Milk 63oC for 30 minutes.
Batch pasteurization Cream 66oC for 30 minutes
Ice-cream mix 69oC for 30 minutes.
Milk 72oC for 15 seconds.
High Temperature Short
Cream 75oC for 15 seconds
Time Pasteurization
Icecream mix 80oC for 25 seconds
UHT heat treatment milk above or equal 130oC at least 2 seconds
9.1.1 Manufacture

Raw milk ~ 4 % fat

Preheating to 30-35oC

Filtering/Clarifying

Thermalization Cooling to 4oC

65 C, 20s, cool to 6oC
o

Centrifuge

Cream 10-12 % fat Skim milk

Standardize , fat
3 % and SNF 8 %

Homogenize 10 MPa

Standardize milk

Pasteurize 75oC, 20 s
Bottles
cool to 4oC

Packing material Packing Cleaning

Storage (Dark, 4oC) Inspecting

Fig.9.1 Flow diagram for the manufacture of standardized, homogenized,

pasteurized milk.

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Details of Production

The importance of thermalization to prevent fat and protein breakdown by heat resistant
enzymes of psychrotrophic bacteria. But as rule, the keeping time of pasteurized milk is too
short to cause noticeable decomposition by these enzymes, unless the original milk had a
high count of psychrotrophs. Furthermore, thermalization at a rather high temperature (say
20 s at 67.5oC) causes a considerable inactivation of milk lipase (about 50 %) and permits
a somewhat lower pasteurization temperature in the manufacture of homogenized milk.
Despite these obvious advantages of thermalization, dairy plants often only cool the milk
(mainly because of the lower costs), taking the risk of some growth of psychrotrophs.
Separation is needed to adjust to the desired fat and solid-not-fat content. If
homogenization is omitted, only a part of the milk will be skimmed, while the skim milk
volume obtained should suffice to standardize the milk.
Homogenization serves to prevent the formation of a cream layer in the package during
storage. Many users dislike such a layer. In low-pasteurized milk (alkaline phosphatase just
inactivated) a loose cream layer of agglutinated fat globules forms that can be easily
redispersed throughout the milk. In high pasteurized milk the cold agglutinin has been
inactivated and a cream layer of forms far more slowly, but then it is a compact, hardly
dispersible layer, a solid cream plug may even result from partial coalescence of the fat
globules. Therefore, this milk is usually homogenized. To reduce the cost not all milk is
homogenized, only its cream fraction is partially homogenized. Obviously, all milk should
then be separated. Homogenization clusters should be absent after the homogenization;
therefore, the fat content of the cream should be rather low (10 to 12%) and the
homogenizer temperature not too low (≥55oC). Usually the homogenization precedes the
pasteurization, to minimize the risk of recontamination. Because milk lipase is still present,
the milk should immediately be pasteurized.
After partial homogenization the milk may still cream due to cold agglutination. This results
from the agglutinin being in the skim milk after separation and being not fully inactivated by
the subsequent pasteurization. In spite of low agglutinin to the surface area, the fat globules
can agglutinate if the raw milk contained much agglutinin.
Homogenized milk has a increased tendency to foam, especially at low temperature.

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Standardization with respect to fat content can be done by adding cream and/or skim milk
to the milk in the storage tank or by continuous standardization. See in detail in the
previous section.
Pasteurization ensures the safety and greatly enhances the shelf life of the product. A mild
heat treatment, e.g., 15 s at 72oC, kills all pathogens that may be present (especially
Mycobacterium tuberculosis, salmonella spp. enteropathogenic Coli spp., Campylobacter
jejuni, and Listeria monocytogenes), to such an extent that no health hazard is left. Some
cells of some strains of Staphylococcus aureus can survive the heat treatment, but they do
not grow to the extent as to form hazardous amounts of toxins. Such low pasteurization
inactivates alkaline phosphatase to the extent as to be no longer detectable (the enzyme
may, however, regenerate after keeping the product for some days, but this especially
holds for pasteurized cream). Most of the spoilage microorganisms in raw milk, such as
coliforms, Mesophilic lactic acid bacteria, and psychrotrophs, are also killed by
pasteurization. Among those not killed are heat-resistant Micrococci (Micobacterium spp.),
some Thermophillic streptococci, and bacterial spores. But these microorganisms do not
grow too quickly in milk, except Bacillus cereus.
The natural milk enzyme lipoprotein lipase may cause lipolysis; undesirable changes in fat
will results. Homogenized milk is more susceptible to lypolysis because of its readily
accessible “substrate”, hence it should be rather intensely heated (e.g., 20 s at 75oC) to
reduce its lipase activity to 10-3 to 10-4. A decrease to 10-2 suffices for nonhomogenized
milk, which implies a heating of, say, 15 s 72.5oC. Milk proteinase is not inactivated by
pasteurization; but the keeping time of pasteurized beverage milk generally is too short to
cause problems.
In high pasteurized milk (e.g., 15 s 85oC) the bacterial growth inhibitors are eliminated and,
despite its lower initial bacterial count, the milk may have a shorter shelf life than has low-
pasteurized milk. High-pasteurized milk is often heated in the bottle, and this improves its
keeping quality because recontamination does not occur; however it also causes a distinct
cooked flavor.
In the manufacture of low-pasteurized beverage milk flow through heating is commonly
applied, as a rule in a plate heat exchanger. The time-temperature combination selected is
a compromise between sufficient inactivation of milk lipase and conservation of the activity

185
inhibiting bacterial growth. Usually temperature is adjusted, but adjusting the length of time
at a constant temperature may give better results. On pasteurizing homogenized milk the
agglutinins should be inactivated to such an extent as to prevent creaming of the milk. A
cooked flavor sometimes be observed. High-pasteurized milk has a somewhat whiter color
due to homogenization. A more intense heating causes browning due to Maillard reactions.
Sometimes heating to over 100oC is applied, to kill spores of Bacillus cereus, thereby
enhancing shelf life.
Packing of low-pasteurized beverage milk is generally done in single service containers,
such as cartons. A certain quantity of milk is still filled in glass bottles. Great care should be
taken to ensure hygiene during packing in terms of the shelf life of the product, but
especially because of the effect of recontamination on the shelf life of the product; aseptic
packing is desirable. The temperature of the milk may increase by about 1 K during packing
due to the transportation in pipelines and on conveyor belts, and due to the use of sealing
machinery.
9.1.2 Shelf life
It is the time during which the pasteurized product can be kept under certain conditions
(e.g., at a given temperature) without apparent undesirable changes. Changes in beverage
milk during storage can be distinguished in:
• Decomposition by bacteria growing in the milk, like acid production, protein
breakdown, and fat hydrolysis.
• Decomposition by milk enzymes or by extracellular bacterial enzymes like fat and
protein breakdown.
• Chemical reactions causing oxidized or sunlight flavor.
• Physicochemical changes like creaming, flocculation, and gel formation which may in
turn be caused by changes mentioned above.
Pasteurized beverage milk should kept for several days after purchase, provided it is kept
refrigerated (below 7oC). Sometimes, a “day of ultimate sale” is given with the product; in
other cases an “ultimate day of consumption” (or minimum guaranteed shelf life).
Chemical changes especially concern the high susceptibility of low pasteurized milk to light
induced off-flavor. Deterioration of pasteurized milk is especially caused by growth of
microorganisms. It is determined by:

186
 • Storage temperature.
• Extent of contamination.
• Growth rate (generation time, g) of the bacteria involved.
• Number of spores of B. cereus in the original milk.
• Activity of substances inhibiting bacterial growth.
The storage temperature of the milk is important because the generation time of the
microorganisms is highly temperature-dependent. The growth rate of bacteria depends on
the temperature and the bacterium species involved. Starting from a count in the milk of 10
per liter, and with g amounting to 4, 7, and 10 h, a shelf life of 5, 8, and 13 days,
respectively, is calculated.
After pasteurization of the milk its count normally amounts to 500-1000 ml-1, unless many
heat resistant bacteria are present in the original milk. As a rule, the milk is spoiled by
“sweet curdling”, caused by B. cereus (g ≥ 10 h at 7oC), unless it is recontaminated. B.
cereus, forming lecithinase, is also responsible for the “bitty cream” defect in
nonhomogenized milk, i.e., the enzyme coagulates the fat globules in the cream layer that
in the vicinity of a “colony” of these bacteria. At a storage temperature below 6oC, B. cereus
cannot grow; deterioration may then be caused by B. circulans. High-pasteurized milk,
made by heating at about 100oC, is mainly spoiled by B. licheniformis, or by B. subtilis if the
keeping temperature is relatively high. Milk contains, say, 10 spores of B. cereus per 100
ml; its shelf life for normal storage conditions amounts to 12-14 days if it is not
recontaminated. Shelf life can be extended by decreasing the count of B. cereus spores by
means of bactofugation. Provision should then be made against enzymic deterioration while
aseptic packaging has to be applied.
If the pasteurized milk is recontaminated, deterioration is generally faster and of a different
nature. Thorough cleaning and disinfection of the filling and lidding machine is needed to
avoid recontamination after flow through pasteurization. In determining of the day of
ultimate sale, one usually assumes that some recontamination of the milk occurs.
Frequent and thorough inspections are needed during processing to limit recontamination
and to meet the requirements at the day of ultimate sale.

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9.1.3 Use of micro filtration
By using micro filtration, bacteria and bacterial spores can almost fully be removed from a
solution; this can be advantageous in the manufacture of milk for liquid consumption. The
method can readily be used in milk processing tanks to ceramic membranes and to further
technological developments. Pressures applied are below 1 bar. A high flux and long
operating periods can be achieved. The fat globules are also retained, considering that the
membrane has a pore size of about 1 µm; therefore, the milk should first be separated. An
outline of manufacturing process by using micro filtration is given below. Some 0.1 % to 1
% of the total number of bacterial cells passes to the permeate, of B. cereus < 0.05 %.
Stronger reductions, even up to sterility, can be obtained by using membranes with smaller
pore size, but that is at the expense of the flux and of the maximum operating time. The
amount of retenate is only a small percentage of the initial volume; the protein content is
slightly increased, by about 0.5 percentage unit. The retenate is sterilized along with the
cream.
The principal effect of the method is the enhanced shelf life of the product. On the other
hand, part of the product (about 12 %) is sterilized; the ensuing cooked flavor is restricted
by applying a brief UHT treatment, but it is important to note that the fat globules (which
generate the greater part of the sulphydryl compounds on intense heat treatment) are in the
most intensely heated fraction.

Raw milk
100
Cream 40 % Centrifuge Skim milk
2.8 10 90
Mixing Retenate Microfiltration
11.7 4.5 85. 5
UHT treatment Mixing Permeate
4 s 130oC 97.2
Low pasteurize Homogenize

Beverage milk

Fig. 9.2 A manufacturing process for pasteurized beverage

milk by using microfiltration.

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9.2 Sterilized milk
9.2.1 Description
Sterilization of milk is aimed at killing all microorganisms present, including bacterial
spores, so that the packed product can be stored for a long period at ambient temperature,
without spoilage by microorganisms. Since molds and yeasts are readily killed, we are only
concerned about bacteria. The undesirable secondary effects of in-bottle sterilization like
browning, sterilization flavor and losses of vitamins can be diminished by UHT sterilization.
During packing of UHT-sterilized milk, contamination by bacteria has to be rigorously
prevented. After UHT sterilization, certain enzymatic reactions and physicochemical
changes still may occur.
To achieve the objectives it is necessary that:
• The count of microorganisms, including spores, is reduced to less than 10-5 per liter.
• The original milk does not contain enzymes of bacterial origin that cannot be fully
inactivated by the heat treatment.
• Enzymes naturally present in milk are sufficiently inactivated.
• Chemical reactions during storage are minimal.
• Physical properties of the milk change as little as possible during treatment and
storage.
• The flavor of the milk remains acceptable.
• The nutritive value of the milk decreases only slightly.
These are difficult to achieve, so attention should be given to processing costs,
complexicity of the machinery and processing, and the consumer’s wishes.
Oxidation causes off-flavors and decomposition of vitamins. This can be overcome by
deaeration, by excluding air from the package, and by using the package i.e., impermeable
to light and oxygen. Furthermore, Millard reactions can occur, both during the heat
treatment (in-bottle sterilization) and during storage (UHT milk). The latter reactions are
responsible for browning, off-flavor, and decreased nutritive value.
Sterilized milk is kept for a long time so that it will show strong gravity creaming, if
unhomogenized. Most intense creaming would occur in in-bottle sterilized milk because
during the sterilization fat globules can coalesce. Creaming as such is undesirable.
Besides, partial coalescence of closely packed fat globules will lead to formation of a cream
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plug, which is hard to mix throughout the remaining milk; oiling off may even occurs at
somewhat elevated temperatures. Therefore, sterilized liquid milk is always homogenized.
If the milk is in-bottle sterilized, little variation in process conditions is possible; the product
obtained can be clearly recognized by the user because of its inevitable sterilized flavor. If
the milk is UHT-heated, a sufficient sterilizing effect can readily be achieved, which implies
that the appropriate process conditions can be selected on the basis of additional
considerations. The flavor can vary from a mild (at, say, 0.6 s at 145oC) to a marked
cooked flavor (UHT heating of, e.g., 16 s at 142oC in a heat exchanger with a warming and
cooling profile that can be scarcely distinguished from the flavor of in-bottle sterilized milk.
The UHT milk can be characterized by means of chemical change, for which the formation
of lactulose is generally used. A standard for UHT milk then would be that it contains less
than 600 mg lactulose per liter.
9.2.2 Method of manufacture
Raw milk ~ 4 % fat

Preheating to 30-35oC

Filtering/Clarifying

Thermalization
65oC, 20s, cool to 6oC

Centrifuge 40oC
Excess cream
Cream 35 % fat Skim milk
Standardize , fat
3 % and SNF 8 %

Standardized milk

Bottles Pasteurize 85oC, 20 s Homogenize 65oC

20 MPa
Bottle cleaning Bottle filling

Sterilize 12 min 118oC

Cool to 20oC

Fig.9.3 Manufacture of in-bottle sterilized milk.

The proteinases and lipases of psychrotrophs, especially of the genus Pseudomonas, can
be very heat-resistant and even in-bottle sterilization does not suffice to fully inactivate
190
these enzymes. Therefore they should be absent in the raw milk. In particular, the addition
of some milk psychrotrophs may have grown extensively. These bacteria especially
produce heat-resistant enzymes in an (almost) full-grown culture (stationary phase).
The various types of heating processes are:
• In-bottle sterilization
• Flow-through preheating and a mild in-bottle sterilization
• Flow-through sterilization and aseptic packing.
The sterilizing effect required determines the lower limit of the time-temperature relationship
to be selected. The sterilization intensity also has an upper limit, which is reached when the
milk protein starts to coagulate. Nearly all good quality raw milk is stable enough to
withstand sterilization. The heating step in the UHT process with direct heating causes
formation of aggregates of casein micelles, which lead to an astringent mouth-feel and to
some sediment on storage of the milk. Heat coagulation is responsible for the aggregates.
High pressure homogenization (40 MPa) is needed to disrupt them; since homogenization
must be done aseptically, this needs a specifically designed homogenizer. Browning of in-
bottle sterilized milk is inevitable because at the usual temperature of 118oC the curves of
sufficient sterilizing effect and significant browning intersect.
Standardized milk

Pasteurize at 80oC,20 s

Homogenize 20 MPa Steam injection

at 150o, 2 s
De-aeration
Direct expansion
Cooling to 80oC
Sterilize at 142oC
Cool to 20oC
Aseptically
Packaging Homogenize
material 40 MPa

Aseptic packaging Sterilize Centrifuge 40oC

Fig.9.4 The manufacture of UHT- sterilized milk (indirect or direct heating)

with aseptic packaging.

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UHT sterilization is mostly performed at temperatures above 140oC. Accordingly, the
sterilizing effect required is readily attained. But a sufficiently long shelf life at ambient
temperature is only obtained if the residual activity of the milk proteinase (plasmin) is at
most 1 %.
600 mg of lactulose represents the upper limit of UHT sterilization, but at that limit a
significant cooked flavor is already formed. For short sterilization times, however, both the
selected time-temperature combination and the full ‘thermal load” of the product, including
heating and re-cooling, are important.
When indirect UHT heating is applied, oxygen should first be removed from the product by
means of deaeration, preferably up to less than 1 mg per kg milk. In direct heating this is
already achieved during the evaporative cooling of the product. If small amount of O2 is
present, it can lead to removal of a slight cooked flavor within a few days, but high O2
contents cause development of an oxidized flavor and partial loss of some vitamins during
keeping. Since the intense heat treatment during in-bottle sterilization forms sufficient
antioxidants, deaeration is not necessary in that case (bottles with a crown cork become
deaerated during sterilization).
The package for sterilized milk should be impermeable to O2; on aseptic packing complete
filling should be aimed at (no head space). UHT milk is, moreover, highly susceptible to off
flavors caused by light, so that a package impervious to light is to be preferred.
9.2.3 Shelf life
Spoilage of in-bottle sterilized milk can be caused by insufficient heat treatment, due to
which spores of, for instance, Bacillus subtilis, B. circulans, B. coagulans or B.
stearothermophillus have survived sterilization. B. subtilis has relatively heat-resistant
spores and this bacterium may cause deterioration of in-bottle sterilized milk. If the milk is
stored under tropical climate, it may spoil due to B. stearothermophillus, which is very heat
resistant spores. Both a low count of these spores in the original milk and a UHT preheating
step can help B. stearothermophillus does not grow below about 35oC. A mild in-bottle
sterilization after a UHT presterilization is only possible if during filling not more than a very
slight contamination by bacterial spores occurs. If the package is not completely tight (e.g.,
due to an ill fitting crown cork), then the milk can also be recontaminated and thus become
spoiled.

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Deterioration of UHT milk by bacterial growth is usually caused by recontamination. The
type of determination is determined by the species of the re-contaminating bacteria.
Recontamination by pathogens may even occur, possibly without marked deterioration. Up
to now (rare) cases of food poisoning due to UHT milk contaminated by Staphylococci have
been reported.
Enzymatic deterioration of UHT milk due to the presence of heat-resistant bacterial
enzymes, such as gelation or development of bitter, rancid, or putrid flavors, can only be
prevented by good quality raw material. Deterioration by milk proteinase, causing, say,
bitter flavor, will mainly, occur in those cases where it is desirable to store UHT milk for a
longer time (e.g., up to 6 months) and at higher temperature, as in tropical countries. A
more intense heating can partially prevent this.
Due to plasmin activity at an ambient temperature off flavors like “gluey” and “bitter”
develop, primarily and milk will be only keep for 2-3 weeks. The period can be extended up
to 6 weeks if product is kept refrigerated. Non-enzymatic deterioration of UHT milk during
storage may concern: Oxidation, influence of light, and Maillard reactions.
The keeping quality of in-bottle sterilized milk is checked by incubation of samples at
various temperatures, mostly 30o and 55oC. After a few days, one can, for instance,
determine: smell, flavor, appearance, acidity, colony count, and oxygen pressure. The
sterility of UHT milk can, in principle, be verified in much the same way. O2 pressure can be
done rapidly, it is only suitable to the product just after packing, still contain some oxygen,
reduction of O2 pressure then points to microbial growth. The increase in bacterial ATP via
bioluminescence is also possible. The sterilized milk should preferably be sold only after
the result of the shelf life test has become known and is satisfactory.
9.2.4 Flavor
Good flavor is, of course, an essential quality mark of beverage milk. Fresh milk has a fairly
bland flavor, whereas full-cream milk has a “richer” taste than (partly) skimmed milk. The
fresh milk may already have off-flavors. These can mostly not be removed, although flavor
compounds formed by heating may to some extent mask off-flavors; the first effect of heat
treatment mostly is that the typical “cowy” flavor of fresh milk is reduced (masked), so that
the flavor becomes even more bland. Flash boiling of milk, as occurs in the cooling section
of a direct UHT heater or in a “vacreator,” may reduce some off-flavors.

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Microbial growth, either before or after processing, may cause various off-flavors. Unclean
or even putrid flavors are mainly caused by some Psychrotophs, whereas other (e.g.,
Pseudomonas fragii) cause fruity flavors. Lactic acid bacteria turns milk to sour and malty
flavor ( caused by Lactobacillus lactis var. maltigenes), may also occur. Bacillus circulans
occasionally causes a phenolic flavor in in-bottle sterilized milk. Growth of B. cereus in
pasteurized milk readily leads to very unclean flavor; might contain sufficient toxin to be
hazardous. Milk enzymes may cause a bitter flavor due to proteolysis by plasmin, as may
occur in UHT milk; and a soapy rancid flavor, due to lipolysis by lipoprotein lipase in low
pasteurized milk.
From lipid oxidation produces off-flavor called “tallowy” or just oxidized flavor, but in some
cases cardboard-like flavor develops. The later may be caused by oxidation of
phospholipids, and it can also develop in skim milk.
Exposure to sunlight can be highly detrimental to milk flavor, and 10 min of direct sunlight
on milk in a glass bottle or 10 h of “fluorescent” light on milk in a carton (not provided a Al-
foil layer) be sufficient to produce defects. The off-flavor is formed not immediately but
rather several hours after illumination. It may concern oxidized flavors, but also a quite
different “sunlight” flavor. The latter is mainly due to oxidation of free methionine to
methional (CH3.S.CH2.CH2. CHO) and to free thiols formed from sulphur containing amino
acids residues; the presence of riboflavin is needed for the sunlight flavor to develop.
Heat treatment leads to a change in flavor, the appreciation or dislike of which varies
greatly among consumers. The main flavor profile compounds are cooked flavor, UHT
ketone flavor, and sterilized-caramelized flavor. Cooked flavor mainly caused by the
presence of H2S liberated after denaturation of protein high pasteurization and boiling. UHT
milk also has cooked flavor but, in addition, it has a ketone flavor that predominantly
originates in the lipid fraction and is due to methyl ketones and, to a lesser extent, to
lactones and sulphur compounds. The cooked flavor and ketone flavor greatly depend on
the type of UHT process used. In-bottle sterilized milk has no real cooked flavor (-SH
compounds); it has a UHT ketone flavor, but this is largely masked by the sterilized-
caramelized flavor formed form certain Maillard and caramelized products.

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9.2.5 Nutritive value
The nutritive value of pasteurized and UHT-sterilized milk changes little by the heat treatment and
during storage. In-bottle sterilized milk shows a somewhat greater loss of nutritive value. The
decrease of available lysine and the total or partial loss of some vitamins are of special concerns.
Maillard reactions are responsible for the partial loss of lysine. They occur to some extent in UHT-
sterilized milk during storage and in in-bottle sterilized milk during heating.
The losses of vitamins mainly concern vitamin C and some five vitamins of B-group. Vitamins A
and E are sensitive to light and/or oxidation, but mostly their concentrations do not decrease in
sterilized milk. Losses of vitamins in milk should be evaluated relative to the contribution of
beverage milk to be supply of these vitamins in the total diet. Especially losses of vitamin B1, B2,
and B6 should be considered undesirable. The loss of vitamin C is generally less importance, but it
may affect the nutritive value in other ways. The breakdown of Vitamin C is connected with that of
vitamin B12; moreover, vitamin C protects folic acid from oxidation.
Loss of vitamins during storage can largely be avoided if O2 is excluded. Vitamin C and B9 may
completely disappear within a few days if much O2 is present. The loss is accelerated by exposure
to light, with riboflavin (vitamin B1) being a catalyst. Most of the riboflavin disappears on long-term
exposure to light.

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Chapter 10

Cream products

Cream is sold in many varieties. The fat content may range from 10 % (half- and –half”) to
48 % (“double cream”). Although it may be used for several purposes, mostly it is
something of a luxury and therefore an excellent flavor is paramount. Because of the high
fat content, any off-flavor of the fat becomes concentrated. For instance, milk with a fat
acidity of 1 mmol per 100 g fat will not be perceived to have a soapy-rancid flavor by most
people, but a whipping cream made from it will definitely taste rancid. Hence, the milk
should be impeccable with regard to lipolysis and fat oxidation.
Sometimes anhydrous milk fat is used in cream products and recontamination is applied.
This enhances the danger of an oxidation flavor and also, if impeccable in this respect, the
test of the product may be different because of the absence of components from the milk
fat globule membrane. One may improve on this by using a limited quantity of good quality
(dried) sweet cream buttermilk. Besides plain cream, some derived products are made,
such as sour cream and ice cream.

10.1 Sterilized cream

The cream contains about 20 % fat (light cream). A good keeping quality is essential
because many consumers use it a little at a time or want to have it stored for special
occasions. The cream is usually sterilized to guarantee microbial shelf life. Chemical
stability is generally not a problem, although ongoing Maillard reactions can occur during
long-term storage. Due to the intense heat treatment oxidative deterioration scarcely occurs
and neither does lipolysis. Physical deterioration may be considerable; gravity creaming
and fat clumping or oiling off. Therefore, the cream must be homogenized, if stored for a
long time it may thicken with age, form a gel, or become lumpy.
Most of the cream is used in coffee; hence the name coffee cream. Thus it is important that
the cream not feather in coffee and that it cause sufficient whiteness (i.e., turbidity) after
dilution with coffee. Sterilization flavor mostly is not too objectionable since this is largely
masked in the coffee.

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Cream is also used in desserts, e.g., on fruit. A pure flavor, then, is paramount, as are a
white color and a relatively high viscosity. Sometimes a very thick, almost pudding-like
cream is made.
10.1.1 Manufacture
In-bottle sterilized coffee cream prepared traditionally has been given below. Alternatively,
raw or thermalized milk may be skimmed and the cream obtained may be standardized,
pasteurized, and homogenized at the pasteurization temperature. The manufacture of ultra-
high temperature short time heated (UHT) cream. In this case the cream should be
homogenized after; sterilizing; otherwise, UHT heating will cause coagulation of protein and
fat globules, and coalescence of fat globules.
If a highly viscous cream is desired, the cream will be homogenized at a lower temperature
and in one stage in order to produce a maximum of homogenization clusters.
Milk Pasteurize Centrifuge Cream
Phosphatase 40-50oC
negetive
Skim milk
Homogenize Heating to 70-75oC Standardize
11 + 3 MPa 20 % fat, pH

Bottle filling Sterilize to Cool to 25oC

115oC,20 min Stabilizing salt
Bottles
crown corks
Milk Pasteurize Centrifuge Cream
Phosphatase 40-50oC
negetive
Skim milk
Cool to 50oC UHT treatment Standardize
140oC for 10 s 20 % fat, pH
Aseptically
homogenize Aseptic Cool to 10oC
10 MPa packing Stabilizing salt

Packing material

Fig.10.1 The manufacture of coffee cream (top) and dessert cream

(bottom). Added stabilizing salt, e.g., is 0.15 % Na3C6H5O7.5H2O.

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10.1.2 Heat stability

In making sterilized cream it is hard to avoid coagulation during sterilization while at the
same time the product is sufficiently homogenized to prevent rapid creaming and (partial)
coalescence of fat globules. Homogenization is largely responsible for the poor stability
toward heat coagulation. Although the heat stability of cream (like that of evaporated milk)
can be improved by adjusting the pH and by adding stabilizing salts (e.g., citrate), the main
variables are the conditions during the homogenization. It appears that as the surface area
of fat globules that is covered with casein increases, the cream becomes less stable.
Because of this, preheating at a high temperature does not help; it causes the serum
proteins to precipitate so that a larger part of the oil-water interface is covered by casein.
Furthermore, the presence of homogenization clusters will shorten the heat coagulation
time.
The higher the homogenization pressure, the lower the heat stability. However, creaming
and (partial) coalescence will cause problems at lower homogenization pressures. It is
advantageous to make the fat globule size distribution as narrow as possible.
10.1.3 Stability in coffee
Feathering of the cream in coffee is due to coagulation of the fat globules and runs largely
parallel to the heat stability. UHT cream is rather susceptible to feathering. In its
manufacture no problems do arise with heat coagulation, but feathering occurs readily if the
homogenization pressure is too high. Moreover, UHT cream is liable liable to thicken with
age or to show aggregation during storage. The latter phenomenon starts with the
aggregation of fat globules. Soon this also leads to feathering in coffee. Feathering
obviously depends on temperature, pH, and Ca++ activity of the coffee, also. Stability in the
coffee may be improved by increasing the solids-not-fat content of the cream, presumably
as it buffers for H+ and Ca++ in the coffee.
10.1.4 Clustering
Dessert cream should be somewhat viscous. It can be achieved by the formation of
homogenization clusters, though thickening agents (carrageenan, alginate) can be added
successfully. Clustering increases the viscosity because the effective volume fraction of the
fat globules increases; first, because of the plasma entrapped between the fat globules (this
part of the plasma is essentially immobilized), and second, because of the irregular shape
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of the clusters (causing them to occupy an efficiently enlarged volume when rotating due to
shear). As the fat content of the cream is higher, the increase in viscosity due to a given
extent of clustering of the fat globules is stronger. Moreover, the clustering itself is more
extensive for a higher fat content.
The viscosity can be reduced considerably by a second homogenization at a much lower
pressure: the homogenization clusters then are partly disrupted again (hence reduced in
size); moreover, the remaining clusters are more rounded. The same is achieved by
exposing the “clustered” cream to shear, as, for instance in a rotating viscometer. The
decrease of the (apparent) viscosity with increasing shear rate; the greater the rate, the
further the clusters are disrupted, and the latter do not reform on release of the shear; as
the hysteresis loop shows. High shear rates should therefore be avoided during pumping
and packing if homogenized cream is to retain its high viscosity.
10.2 Whipping cream
This concerns 35 to 40 % fat cream. It is primarily designed to be beaten into foam, often
with sugar added. It is mostly available as a pasteurized product in small bottles, plastic
cups, or large cans. It is also sold as in-can sterilized cream, and even supplied with sugar
and a driving gas in an aerosol can that delivers a ready-made whipped cream.
10.2.1 Desirable properties
The most important specific requirements are:
1. Flavor: The product is eaten for its flavor, which obviously must be perfect. Rancid and
tallowy flavors in the original milk should be rigorously avoided; this requirement is even
more essential than for coffee cream. Not everybody appreciates a sterilization flavor or
even a pronounced cooked flavor, and partly because of the cream usually is
pasteurized.
2. Keeping quality: Many kinds of spoilage can occur, but it is often desirable to store the
cream for a prolonged time. The original milk should contain not more than a few heat
resistant bacteria; above all, Bacillus cereus is a disastrous microorganism in whipping
cream (it causes a fat emulsion to become unstable). No growth of Psychrotrophs can
occur in original milk because they form heat resistant lipases. To allow for a fairly long
shelf life, the pasteurized cream should be packed under strictly hygienic or even

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 aseptic conditions. Recontamination by bacteria may occur, therefore, whipping cream
is often heated by in-can or in-bottle pasteurization.
3. Whippability: The cream should quickly (i.e., in a few min) and easily whip up to form a
firm and homogeneous product, containing about 50 % v/v of air (=100 % over run).
4. Stability after whipping: The whipped cream should be firm enough to retain its shape,
remain stable during deformation (as in “decoration”), not exhibit coarsening of the air
cells, and show negligible leakage of liquid.
Sometimes carrageenan is added as a thickening agent.
10.2.2 Manufacture
The classical manufacture of whipping cream is fairly simple. The pasteurization of the
cream should at least be sufficient to fully inactivate milk lipase. Usually, the heat treatment
is far more intense in order to improve the bacterial keeping quality. The way of heating, as
well as the heating intensity, varies widely, holder pasteurization (e.g., 30 min at 85oC),
heating in a heat exchanger (possibly over 100oC), and in can (bottle) heating (e.g., 20 min
103oC) are used. Likewise the manufacturing sequence, separation temperature, and so
forth vary widely. Sometimes the cream is stirred in an open vat at rather high temperature
in order to deodorize it; vacreation is not suited because it damages the fat globules.
Such damage, especially (partial) coalescence of the fat globules, should be avoided. The
milk, and especially the cream, should be handled gently. The cream should not be
processed or pumped unless the fat is completely liquid or largely solid, i.e., only at
temperatures below 5oC or above 40oC. Hence, but it is rather uneconomical. Sterilization
of whipping cream may cause problems. In-bottle or in-can sterilization often causes
coalescence, unless the cream is first homogenized. However, most homogenized cream
can not be whipped. UHT heating is to be preferred, also because of the flavor; the cream
should then be homogenized aseptically at low pressure and the composition should then
be homogenized aseptically at low pressure and the composition should be adjusted
(emulsifier added). A disadvantage of UHT whipping cream is that the temperature
fluctuations to which it may be subject can cause “rebodying”. To prevent creaming during
storage, a thickening agent is generally added (e.g., 0.01 % κ-carrageenan).

200
 Cream

Milk Pasteurize Centrifuge Thickening Standardize

at 72oC,15 s 50oC agent 36 % fat
Packing material
Skim milk
Cold storage Packing Pasteurize at
Cool to 5oC
85oC for 30 min

Fig.10.2 Manufacturing flow diagram of whipped cream.

10.2.3 The Whipping process
When skim milk is beaten, foam very rich in air is rapidly formed on top of the liquid. This
proceeds more slowly when cream is beaten and the air bubbles stay in the liquid for a
longer time. This is partly because of the higher viscosity but also because the fat globules
directly penetrate the air-water interface, attaching them to the air bubble and spreading
some liquid fat onto the bubble surface. Because of this the film between closely
approached air bubbles are rather unstable and initially the bubbles coalesce readily. The
fat globules are so highly concentrated that they readily show partial coalescence
(clumping). In this way a structure of clumped fat globules forms, enclosing the air bubbles
and giving a rigid and stable form. To achieve this, air cells and fat clumps should be of
similar size, preferably 10-100 µm. The foam increases in firmness during whipping, but it
also becomes coarser. On prolongs beating, the clumps become so large and few that they
cannot stabilize but a few large air cells; the whipping becomes churning and the clumps
become butter grains; the air bubbles coalesce and disappear again.
The balance between foaming and churning partly depends on the way of beating. If this is
too slow, the cream may churn prematurely. Vigorous beating causes a high overrun and
finely structured and smooth foam. The smaller the air cells, the less clumping are needed
to enclose the bubbles and to produce firm foam.
Proteins and other surfactants may cause some foam stability. But since encapsulation of
air bubbles with fat globules does not occur, the foam is mostly unstable to manipulation.
These products often have a high over run over 200 %, instead of around 100 % for
ordinary whipped cream.
10.2.4 Variables
Several properties of the cream affect the whipping process.
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a. Fat content: It has considerable effect. But the influence depends on the conditions
during whipping. The more intensive the beating, the lower fat content of the cream
allowing a stable foam to form, and the higher the overrun.
b. Crystallization of fat: It is essential for clumping. If the amount of liquid fat is high,
clumping is too rapid and the foam becomes unstable. Hence, deep cooling and a
sufficient cooling time of the cream are essential, as is a low temperature during
storage and at whipping. The composition of the fat has an effect: more problems in
summer than in winter.
c. Protein content in cream: When beating starts protein is needed to form foam cells.
Addition of thickening agents hardly affects whipping, but leakage of liquid is
considerably reduced.
d. Homogenization of cream: Homogenization considerably impairs whippability; the
globules become too small to clump rapidly. This may, however, be better than
expected if the fat globules have formed homogenization clusters because far less
clumping is needed in that case. Homogenization at low pressure (1-4 MPa), preferably
in two stages (e.g., 2 and 0.7 MPa at 35oC), can give clusters of some 15-20 µm in
diameter.
e. Surface layer with surface active agents: Supplying these decreases the formation of
clusters and increases the tendency to clumping; then homogenization at higher
pressure may be applied. The surfactant added may be a monoglycerides or a Tween;
the latter drastically affects the whipping properties.

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Chapter 11

Production of ice cream

11.1 Introduction

Ice cream may be defined as a frozen dairy product made by suitable blending and
processing of cream and other milk products, together with sugar and flavor, with or without
stabilizer or color, and with the incorporation of air during the freezing process.

According to the PFA rules(1976), ice cream is the frozen product obtained from cow or
buffalo milk or a combination thereof or from cream and/or other milk products, with without
the addition of cane sugar, eggs, fruits, fruit juices, preserved fruits, nuts, chocolate, edible
flavors and permitted food colors. It may contain permitted stabilizers and emulsifiers not
exceeding 0.5 % by weight. The mixture must be suitably heated before freezing. The
product should contain not less than10% milk fat, 3.5 % protein, and 36.0 % total solids.
However, when any of the aforesaid preparations contain fruits or nuts or both, the content
of milk fat may be proportionately reduced but may not less than 8 % by weight. Starch may
be added to a maximum extent of 5 %, with a declaration to that effect on the label.

11.2 Classification of ice-cream

No standard classification of ice cream has yet been adopted even in developed
countries. However, some of the important frozen desserts can be classified as follows:

1. Pain ice cream: In this ice cream the color and flavoring ingredients together amount to
less than 5 % of the volume of the unfrozen ice cream. For examples – Vanilla & coffee
ice creams.
2. Chocolate ice cream: Flavored with cocoa or chocolate.
3. Fruit ice cream: It contains fruits, with or without additional fruit flavoring or color. Fruits
such as strawberry, apricot, pineapple, mango, banana etc. Fruits should be fresh,
frozen, packed, canned or preserved.
4. Nut ice cream: Ice cream containing nuts, such as almonds, walnuts, cashew nut, etc
with or without additional flavoring or color.
5. Milk ices or milk lollies: It should contain not greater than 2% milk fat, not less than 3.5
% proteins, not less than 20 % total solids and not greater than 0.5 % stabilizers. It is
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 produced by mixing milk, skim milk or milk products, with or without the addition of can
sugar, eggs, fruits, fruits juices, nuts, chocolates, edible flavors and permitted colors.
6. Soft ice cream: It is sold directly from the freezer without being hardening.
11.3 Chemical composition
Table 11.1 Approximate chemical composition of ice cream (%).
Economy ice cream Good average ice
Particulars cream
Milk fat 10 12
Milk solid not fat (MSNF) 10-11 11
Sugar 13-15 15
Stabilizer & Emulsifier 0.30-0.50 0.30
Total solids 35-37 37.5-39.0
11.4 Specification of ice cream
Characteristics Requirements
Weight (g/L) - minm 525
Total solids (%) ,, 36
Milk fat (%) ,, 10
Acidity (%Lactic acid) -maxm 0.25
Sucrose( % wt.) - ,, 15
Stabilizer/Emulsifier (%wt.)- ,, 0.5
Standard plate count (SPC) per gm 2,50,000
Coliform count ( per gm) 90
Phosphatase test - ive

11.5 Role of different ingredients in ice cream making

a. Milk fat
Increases the food value, but it is expensive.
It enriches & mellows the ice cream, giving a full, rich, creamery flavor.
Also contributes to the body & melting resistance of ice cream while producing a
smoothness of texture.
Gives stability to the ice cream but impairs whipping ability.
b. Milk solid not fat
It is known as serum solids, they consists of milk proteins, milk sugar and mineral matter.
• High in food value, but it is expensive.
• Improves its body and texture, but little to the smell.
• Milk sugar adds to the sweet taste.
• Milk protein helps to make the ice cream more compact and smooth.
• It can be added as large as possible without being sandiness to the product.
c. Sugar
• To increase the stability of the ice cream.
• Desired sweetening effect is only produced by adding sucrose.
• Cheapest source of total solid in the ice cream mix.
• Lowers the freezing point of the mix so that it does not solidify in the freezer.
d. Stabilizers
• To prevent the formation of objectionably large ice crystals in ice cream, especially during
storage.
• Added very small quantities, they have a negligible influence on food value and flavor
e. Emulsifiers
• To provide a uniform whipping quality to the mixture, and
• To produce a drier ice cream with smoother body and texture.

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f. Flavor and colors
• Increases the acceptability of the ice cream product.
• Improves appearance.
• Aids in identifying flavors.
11.6 Manufacture of ice cream
Milk

Centrifuge 40-50oC

Skim Milk Cream

Sugar Standardize Additives

Mix

Pasteurize at 80oC 25 s

Homogenize15+3 MPa
Hardened
Cool 4 C o ice cream

Hardening
Additions Ripening 4-24 h
-25oC

Air Freezing -5oC Packing

Ice cream Packing

material

Fig11.1 Flow diagram for the manufacture of ice cream

11.6.1 Mix calculation

Example of the procedure for calculation of an ice cream mix:

Carry out a calculation for 100 kg of ice cream mix, with the following composition:
12% fat
12% sugar
2 % glucose syrup solids
0.5% emulsifier/stabilizer
17 parts m.s.n.f. per 100 parts of water.
The following raw materials are available:
Cream with 30 % fat
Skim milk with 9 % m.s.n.f.
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 Skimmed milk powder with 97 % m.s.n.f.
Sugar 100% solids
Glucose syrup with 80% solids
Emulsifier / stabilizer 100% solids.
Calculation of cream
kg mix 100
kg cream = fat % in the cream × = 12 × = 40 kg
fat % in cream 30
Calculation of the skimmed milk in cream:
Kg skimmed milk in cream = kg cream – kg fat in cream
Kg skimmed milk = 40 – 12 = 28.0 kg.
Calculation of m.s.n.f. in ice cream:
Kg m.s.n.f. in cream = Kg skim milk in cream x % m.s.n.f. in skim milk/ 100
Kg m.s.n.f. in cream = 28.0 x 9.0 / 100 = 2.52 kg
Calculation of % m.s.n.f.
M.s.n.f. % may be calculated first, because it depends of the amount of water in the ice
cream mix and therefore it can not be specified directly.
100
The m.s.n.f. factor is defined as follows: % m.s.n. f . × =A
% water
Since, “% water” can be expressed as “100 less total solids”, and “total solids” as other
solids plus m.s.n.f., the formula can be condensed as follows:
(100 − other solids )
% m.s.n. f = A ×
(100 + A)
(100 − 26.5)
= 17 × = 10.68 %
(100 + 17)
Calculation of glucose syrup
Kg glucose syrup = percentage glucose syrup x kg mix / % of solids in glucose syrup
100
= 2× = 2.50 kg
80
To get 100 kg mix we are missing:
100 – ( 40.0 +12.0 + 2.50 + 0.5 ) = 45.00 kg (skimmed milk + skimmed milk powder)

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To get 10.68 kg m.s.n.f. we are missing: 10.68 – 2.52 = 8.16 kg (comes from skimmed milk
and skimmed milk powder).
Equation
kg skimmed milk = x ; kg skimmed milk powder = 45.00 - x
x×9 97
+ (45.0 − x) = 8.16
100 100
or, 0.09 x + 43.65 – 0.97 x = 8.16
or, 0.88 x = 35.49
kg skimmed milk (x) = 40.33
Skimmed milk powder = 45.00 – 40.33 = 4.67.
11.6.2 Preparation of Ice cream mix
The first step in making ice cream is to mix the ingredients in a tank in the proportions
calculated on the basis of the formula used. This may lead to two different situations,
depending upon whether the ingredients used are powders and other liquids, or only
liquids.

In the first case, the liquid ingredients are put into the tank where the mix is prepared.
This tank, which may rest on a balance, is equipped with devices which can heat the
contents. The liquid ingredients are agitated and heated, and the powders are added before
the temperature goes above 50oC. It is possible to obtain better dispersion of the dry
ingredients (powder milk, sugar, stabilizer, etc.), if they are mixed before pouring them into
the tank. Nevertheless, certain stabilizers disperse better at lower temperatures, while other
require higher temperatures, such as alginates, which must be mixed at 65oC. If frozen
cream or butter is used, it is essential to ensure that they are melted before reaching the
pasteurization temperature.
In the second case, where all ingredients are liquid, they are mixed by simply controlling
the valves and pumps to introduce into the tank the required quantities of each ingredient.
In this case, it is possible to use more advanced automatic processes. The precision of the
system must be in order of 0.1 % to 1.15 %.

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11.6.3 Pasteurization of the mix

It is very essential steps because of sanitary view point. Heating is required to dissolve
the ingredients and homogenize the mix, pasteurization adds very little to the cost. This
can be carried out in three different ways:
• Batch process, at 68o to 70oC for 30 minutes. This process is not widely used. It is
used mainly in small operations.
• High temperature pasteurization (HTST), at 700 to 850C, for 2 to 20 seconds. This is
most widely used process. It gives better results, produces ice cream with better
rheological and organoleptic qualities, is more economical, and is well suited for
automatic processing.
• Ultra high temperature process (U.H.T.), at 1000 to 1300C, for 1 to 40 seconds. This
process modifies the structure and properties of the proteins and improves the body
and texture of the products. The increased water retention capacity makes it possible
to use smaller amounts of stabilizers.

11.6.4 Homogenization the mix

In order to produce a quality product, it is essential to obtain proper homogenization. The
main purpose of homogenizing the mix is to obtain a more uniform and stable emulsion.
The diameter of fat globules varies between 0.5 and 4µm, with an average of 1 µm, and
few globules measure more than 2µm. The size of the fat globules affects the physico-
chemical properties of the mix, so that viscosity, whipping ability, body, texture, and melting
properties, are improved.

11.6.5 Aging the mix

It consists of keeping the mix at -2o to -4oC for a period of 4 to 24 hrs, before freezing.
This holding period allows the hydration and crystallization phenomena to take place, and
produce a mix with better physical properties. The length of the aging period may vary
depending upon the ingredients used. Some authors believe that an aging of four hours is
generally enough, except in the case of batch freezing.

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 When the mix contains a fast-acting stabilizer and consists only of liquid ingredients,
where proteins are hydrated, it is possible to obtain good quality ice cream without an aging
period.

11.6.6 Freezing the mix

During the freezing process, part of the water in the mix is converted into ice, while, at the
same time, air is added to obtain the desired overrun. The mix & the air are introduced into
a cylinder containing a dasher equipped with scrapper blades which scrape the refrigerated
surface where the mix freezes. A jacket surrounds the cylinder where the liquid coolant
circulates.

Two conditions are essential to obtain a quality product:

• Ice must be in the form of many small crystals, with a diameter of less than 35 µm.
• Air must be evenly distributed, in the form of cells, small and numerous according to the
overrun.
These two physical conditions produce the microstructure which determines the body and
texture of the product.
During the freezing period, the temperature of the mix drops to about –5oC to –6oC.
Under these conditions, and in the case of a mix of average composition, about 50% of the
water is transformed into ice. More than 70% of the water becomes ice at –10oC. The
remaining liquid water is either associated with the proteins and stabilizers, or forms parts
of a "syrup" rich in sugars and minerals.
The speed with which this operation takes place, which is only a few seconds in a
continuous process, and the violent agitation of the product by the blades, facilitate the
formation of many small ice crystals. In fact, the thickness of the layer of ice cream whicsh
freezes on the surface of the drum must be greater than the diameter of the ice crystals,
that is, between 50 – 75 µm.
With conventional freezers, a lower temperature will cause certain mechanical problems
and substantially decrease equipment performance.
11.7 Physical structure: Formation and stability
The chemical composition of an ice cream mix with air on top is exactly equal to that of the
corresponding ice cream. The differences in appearance, consistency (mouth-feel), and

209
flavor are considerable, and these are caused by the difference in physical structure (Fig.11.2).
When half of the water is frozen (amount -5oC) the following structural elements can be
distinguished ( d = diameter, ∅ = volume fraction):
Ice crystals : d = 7 – 170 µm, on average about 50 µm, ∅ = 0.3
Lactose crystals : length ≈ 20 µm, ∅ = 0.005; not always present.
Air cells: d = 60- 150 µm, ∅ = 0.5
Thickness of foam lamellae: 10-20 µm
Fat globules: d < 2 µm , ∅ ≈ 0.06 (including globules in clumps)
Fat globules clumps : up to 10 µm, in size
The size of the ice crystals depends on the stirring intensity and on the cooling rate during freezing,
the quicker the freezing; the smaller the crystals. Immediately after freezing, no lactose crystals are
present. To be sure, the temperature is below that for saturation of lactose, as is seen in Fig.11.1
below, but it is still above that for its homogeneous nucleation. Only after deep cooling can lactose
crystals form.
Microscopically, many air cells can be observed to be somewhat deformed by the ice crystals,
which is not surprising considering the system to be more or less completely “filled”, i.e., the
combined volume fraction of the structural elements is about 0.8. Furthermore, the air cells are
almost entirely covered with fat globules and their clumps. The clumped fat globules, together with
the air cells to which they are attached, form a continuous network throughout the liquid. This has
important effects:
The air cells become stabilized by the fat globules.
After the ice crystals have melted (e.g., in the mouth) the mass retains some firmness (“stand-up”);
in which the extent of fat clumping is expressed as a “churned fat index”(which may be determined
by examining which proportion of the fat creams rapidly after complete melting of ice crystals). The
stand-up is a valued organoleptic property.
The clumping (partial coalescence) of the fat globules changes the texture, i.e., the ice cream looks
less glossy and thereby appears more attractive for most people. This property is called “dryness,”
and it correlates very well with the experimentally obtained churned fat index.
Ice cream having insufficient dryness sticks to the processing equipment, which may interface with
the packing operation, etc.
The network of clumped of fat globules is formed during freezing. Although the air bubbles become
almost completely covered with fat globules, floatation churning presumably does not occur
because too little liquid fat is available to spread over the air bubbles. Clumping is dominantly

210
caused by mechanical forces, i.e., the fat globules are pushed together during beating due to the
presence of ice crystals, and are damaged by them.The lower the temperature (more ice), the
faster the clumping.

100
g ice
100 g (ice + water)
80

g dry matter
60 100 g solution

40 Homogeneous nucleation

20
Lactose saturated

0
0 -10 -20 -30
Temperature (oC)

Fig.11.2 Freezing of ice cream mix. Approximate amount of frozen water and
concentration of the remaining solution, assuming that the ice is in
equilibrium with the liquid and that no other constituents crystallize. The
estimated temperatures for saturation of lactose and for its homogeneous
nucleation are also indicated.

Casein micelle

Fat globule.
Lactose
crystal.
Ice crystal.
Plasma
Air cell.
200 µm

Fig.11.3 Shows the structure of the ice cream at about -5oC.

211
 100

Shape retention (%)

50

0
0 20 40 60
CF
Fig. 11.4 The extent of clumping of the fat globules (expressed as churned fat
index, CF) on the shape retention of ice cream. The retention is the
height of a cube of ice cream after keeping it for some time at room
temperature, expressed as a percentage of its initial height.

11.8 Overrun in ice cream

1. 1800 liters of ice cream were made using 1000 liters of the ice cream mix. Calculate
the overrun (OR)? (volume basis).
(Volume of icecream) − (Volume of mix )
OR = × 100
(volume of mix )

1800 − 1000
OR = × 100 = 80 %
1000
Overrun can also be calculated on the basis of weight.
Example: 1800 liters of ice cream at 0.5 kg/L have been made with 900 kg of mix weighing
1.07 per liter.

Solution : weight of the same mix = 1800 x 0.5 = 900 kg.

900 × 0.5
Weight of the same volume of ice cream = = 420.5
1.07
(900 − 420.5
OR = × 100 = 114 %
420.5
(1.07 − 0.5)
or , OR = × 100 = 114 %
0 .5
2. Calculate the weight of one volume of ice cream for a given overrun:
Weight of one litre of mix = 1.07 kg
Desired overrun = 110 %

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Calculate the weight of one litre of ice cream (x).
(1.07 − x) (1.07 − x)
OR = × 100 or , 110 = × 100 = 0.509 kg (0.51) //
x x
11.9 Hardening the ice cream
As ice cream comes out of the freezer, it is put in containers, and the freezing period
continues. Thus, on the average, the percentage of water converted into ice will go from 50
% to about 80-85 %. This new ice is formed without agitation and over a longer period of
time. The crystallization nuclei formed in the freezer are very important. Ideally, hardening
should take place as quickly as possible. In a cold chamber kept at between –200 and –
300C, without force circulation of cold air among the containers, hardening will take several
hours; while in a tunnel specially designed to provide good circulation of air at temperatures
between –350 and –400C, it is possible to harden the product in less than two hours in one
liter containers.

11.10 Ice-cream defects

Ice cream defects affect mainly the flavor, texture, and body of the product. Appearance
or color defects are relatively rare, and, in this case, the solutions are obvious. However, it
is important to point out that color is an important factor, which is greatly valued by
consumers.

11.10.1 Flavor

This defect are mainly due to the use of poor quality ingredients, the improper use of
these ingredients, or serious deficiencies in the manufacturing and conservation processes.
Abnormal flavors in the raw material used can be detected in the finished product, and
can even develop there, under favorable conditions. This is the case particularly with the
development of oxidized flavors and hydrolytic rancidity associated with fat. The same is
true for the "cooked" and "stale" flavors that are mainly caused by over-heating the milk
proteins. Products where acidity has developed will produce a typical "acid" flavor.
Flavor defects may be due to misuse of ingredients, such as too much or too little sugar
or flavorings: in which case the flavor may be too sweet or too bland, too strong or too
weak.

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11.10.2 Texture and body
The structure of the product can be affected by the air cells ; the ice crystals; the condition of the
fat, proteins and minerals contained in the milk or added to the ice cream; the interface between
the fat globules and the mix; the interface between air and the mix, etc.

Texture is a function of the number and size of the particles, their organization, and their
distribution. It must be smooth and produce a pleasant sensation in the mouth. In this case, the
most important defect is a coarse or sandy texture.
The use of a mix with an unbalanced composition, inadequate manufacturing processes, or
improper conservation conditions, such as excessively high temperatures, or significant
temperature variations, can all lead to the development of this type of defect.
11.10.3 Body defects
This defect is more numerous and more frequent. Good ice cream is firm, keeps well under
adequate melting conditions, and does not produce an unpleasant cold sensation when eaten. On
the other hand, the body of defective ice cream can be flaky, greasy, soft, sticky, heavy, brittle,
soggy etc. It may tend to melt too quickly and form foam. These defects may be the result of using
an improperly balanced mix, or a mix which contains ingredients whose functional properties have
been modified; or they may be the result of inadequate manufacturing conditions.
11.11 Softy ice cream
The composition of the mix used to make soft ice cream is different from that used for regular
ice cream. Freezing point, whipping ability, and the stability of the emulsion are critical factors,
which must be taken into account in the composition of the mix, and the choice of ingredients.

In order to prevent an excessive drop in the freezing point, the sugar content must be kept
around 12-15 %, and the use of corn sweeteners (glucose syrup) must be limited to 25 % of the
total sugar content. The utilization conditions of this product require a certain amount of firmness
and consistency, and good melting resistance.
In order to obtain good consistency and a smooth texture, the stabilizers and emulsifiers used
must be suitable for this particular product and the manufacturing processes used. Proper whipping
requires violent agitation and this increases the likelihood of churning of the fat. The amounts of
stabilizers and emulsifiers used are higher, in the order of 0.4 to 0.6 %. Often, a higher proportion
of solids non-fat is also used, so that the content may be in the order of 14 %.
The fat content must meet legal requirements. When the fat content is too high, this may
facilitate churning, or the separation of the fat in the form of clumps. It may also give the product
too rich a flavor which consumers do not appreciate.

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Chapter 12
Concentrated milks

12.1 Evaporated milk

12.1.1 Description

Evaporated milk is sterilized, concentrated, homogenized milk. The product has a long
shelf- life (it can be kept for several months, even at tropical temperatures), is completely
safe for the user, and can be kept without refrigeration. After dilution, flavor and nutritive
value of the product are not greatly different from that of fresh milk. Traditionally,
sterilization occurs in cans or bottles. Currently, ultra-high temperature short time heated
(UHT) heating is also applied, followed by aseptic packing in cardboard containers. A major
problem with sterilization is the heat stability; the higher the concentration of the milk, the
lower its stability. Therefore, concentrating cannot be by more than about 2.6 times, which
implies about 22 % solid-not-fat in the evaporated milk.
The traditional product shows browning due to Maillard reactions, and it also has a
“sterilized” flavor. The sterilization can destroy up to 10 % of the available lysine, about half
of vitamins B1, B12, and C, and smaller proportions of vitamin B6 and the folic acid. The
product is quite viscous; its viscosity amounts to 40 mPa.s or about 20 times that of fresh
milk. These disadvantages do not seem in UHT evaporated milk, wherein the loss of
nutrients is far smaller and there is a whiter color, a lower viscosity, and a better flavor.
Evaporated milk is mainly used in countries with little or no milk production, especially in
the tropics; it is generally diluted with water before use. An alternative is to make
recombined milk from skim milk powder, anhydrous milk fat, and water. Currently,
evaporated milk is used in coffee in certain countries. After the bottle has been opened the
milk can be kept in the refrigerator for up to 10 days because it initially contains no bacteria
at all and because contaminating bacteria grow somewhat more slowly due to the reduced
water activity, which is about 0.98. In order to control the heat stability of the milk, CaCl2 or
sodium carbonates, phosphates, or citrates may be added. Sometimes, a thickening agent
(e.g., 0.015 % carrageenan) is added to slow down creaming.
12.1.2 Manufacture
The outlines of manufacturing processes of in-bottle and UHT sterilized whole evaporated
milk has given below. Some process steps are discussed in more detail.
215
Preheating serves to enhance the heat stability of the evaporated milk, to inactivate
enzymes, and to kill microorganisms as well as many bacterial spores. The heating
temperature- time relationship is usually selected on the basis of heat stability. Formerly, a
long heat treatment (e.g., 20 min) at a temperature below 100oC was often applied.
Currently, UHT treatment is generally preferred. It reduces the number of spores in the milk
considerably and therefore a less intensive sterilization suffices.
Concentrating: The milk is usually concentrated by evaporation. Standardization to desired
dry matter content is of much concern. A higher concentration causes a lower yield and
poorer heat stability. Continuous standardization is generally applied by determining the
mass density. Based on that parameter either the raw milk supply or the steam supply is
adjusted; it is obvious that density and dry matter content of the raw milk must be
determined. Alternatively, standardization can be based on refractive index determination.
The milk can also be concentrated by reverse osmosis.
After concentration, the manufacturing processes for in-bottle sterilized and UHT sterilized
evaporated milk differ. The process for in-bottle sterilized milk has discussed first.
Homogenization serves to prevent creaming and coalescence. It should not be too
intensive because the heat stability becomes too low.
Stabilization: To ensure that the evaporated, homogenized milk does not coagulate during
sterilization and at the same time does acquire a desirable viscosity, a series of sterilization
tests is often done on small quantities of the evaporated milk to which varying amounts of
stabilizing salts (for the most part Na2HPO4) are added. The tests are needed because
variation occurs among batches of milk. Essentially, the addition of the salt means
adjusting the pH. Because further processing must be postponed until the test results are
available, this necessitates cooling the evaporated milk after its homogenization and
keeping it for a while. Long term storage should be avoided to prevent bacterial growth;
moreover, cold storage of the milk increases the tendency of age thickening. The stabilizing
salt is added as an aqueous solution, which dilutes the evaporated milk slightly. Therefore,
the milk is often concentrated somewhat too far and re-standardized to the correct dry
matter content during “stabilization”.
Packing in cans is common. The tin plate of the cans is coated (provided with a protective
layer of a suitable polymer) to prevent iron and tin from dissolving in the product. After

216
filling, the cans may be soldered up, but mechanical sealing is currently preferred.
Evaporated milk intended for use in coffee is usually packed in bottles that are closed with
a crown cork.
Sterilization In-bottle or in-can sterilization can be applied batch wise (in an autoclave) or
continuously. Machines that have rotary air locks (to maintain the pressure) may be applied
for cans and hydrostatic sterilizers for bottles.
The sterilization is primarily aimed at killing all bacterial spores–reduction to, say, 10-
8
spores per ml- and inactivating plasmin, i.e., milk proteinase. Lipases and proteinases
from Psychrotrophs should be absent from the raw milk because these enzymes would be
sufficiently inactivated. The most heat resistant spores are those from Bacillus
stearothermophillus. This bacterium does not grow at moderate temperatures but may do
so in the tropics. D121 of the spores is some 4-7 min. If the sterilization effect is adequate
for B. stearothermophillus, then B. subtilis, Clostridium perfringens, Clostridium botulinum,
are also absent.

Milk

Standardize

Preheat 30 s 130oC

Evaporation or reverse osmosis

Homogenize 65oC 22 + 5 MPa (Stabilize with Na2HPO4)

Cool to 10oC Sterilize 15 s 140oC cool to 60oC

Stabilize with Na2HPO4 Homogenize 45MPa

Packing Cool to 10oC

Sterilize 15 s 120oC Aseptic packaging

Fig.12.1 Manufacture of in-bottle (left) and UHT (right) sterilized whole milk.

UHT sterilization kills bacterial spores more effectively than in-bottle sterilization. The pre-
heating is also required to prevent excessive heat coagulation in and fouling of the UHT
sterilizer. Some heat coagulation nearly always occurs, and the subsequent
homogenization is also meant to reduce the size of the protein aggregates formed. Aseptic
homogenization must be applied. Indirect UHT sterilization in a tubular heat exchanger
217
allows the pump of the homogenizer to be fitted before the heater and the homogenizing
valve behind it. Thereby the risk of recontamination is diminished. The addition of
stabilizing salt can often be omitted if UHT sterilization is applied or if the amount to be
added is not so critical that sterilization tests must be carried out. It implies that the whole
process from preheating up to and including aseptic packing can proceed without
interruption.
Recombination: Manufacture of evaporated milk by means of reconstitution is given below.
The skim milk powder used has to comply with strict requirements. The powder must have
been made from skim milk that is heated so intensely (e.g., 1 min at 130oC) that the
recombined concentrated milk after its homogenization is sufficiently heat stable. Spores of
B. Stearothermophillus should largely be absent, so that a somewhat more moderate
sterilization of the evaporated milk suffices. Sometimes, up to 10 % of the skim milk powder
is displaced by sweet cream buttermilk powder to improve the flavor of the product. The
copper and peroxide contents of the anhydrous milk fat should be low to avoid flavor
deterioration. A high level of calcium in the water used can cause problems with heat
stability. “Filled evaporated milk” is also made. A fat different from milk fat is used.
Water
Skim milk powder Milk fat
o
Heating to 40 C
Heating 40oC
Dissolution/emulsifying

Filtration

Heating to 62oC

Homogenize 20 + 4 MPa
Na2HPO4, carrageenan Cool to 10oC
Cans Packing

Sterilize at
120oC for 15 s

Fig.12.2 Manufacture of recombined evaporated milk

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12.1.3 Organoleptic properties
A Maillard reaction is responsible for flavor and color of evaporated milk. The temperature
and the degree of heat treatment during manufacture determine the initial concentration of
the reaction products, but ongoing Maillard reactions occurs during storage, especially at a
high temperature. The milk eventually develops a stale flavor, also due to Maillard
reactions. The flavor after a long storage time differs considerably from that directly after
intense heating. This is because the complicated set of reactions involved leads to different
reaction products at different temperatures. A “sterilized flavor “may be appreciated when
the milk is used in coffee. Off-flavors due to autoxidation need not occur.
When the milk is used in coffee the brown color is often desirable, to prevent the coffee’s
acquiring a grayish hue. The brown color depends greatly on the Maillard reactions, though
the color of the fat is also involved.
The viscosity of evaporated milk is an important quality criterion. Many consumers prefer
the milk viscous.
If the bacterial lipases and proteinases may remain active in evaporated milk can cause
deterioration and will result soapy rancid and bitter flavors, and to age thickening.
12.1.4 Heat Stability
Concentrated milk is far less stable during sterilization than non-evaporated milk and the
fatty intensive homogenization applied decreases the heat stability further. Evaporated milk
should increase in viscosity during sterilization. Essentially, the viscosity increases by
incipient coagulation. A subtle process optimalization is needed to meet these
requirements.
The milk must be preheated before evaporation in such a way that most serum proteins are
denatured. Otherwise the evaporated milk forms a gel during sterilization due to its high
concentration of serum proteins. Pre-heating to about 120oC for 3 min is done.
The pH should always be adjusted. Preheating and evaporation have lowered the pH to
about 6.2 and that is clearly below the optimum pH. In practice, NaHPO4.12H2O is usually
added, but NaOH can also be used.
The influence of some variables like the homogenization temperature differs from that in
sterilized cream. Homogenization of evaporated milk does not lead to formation of

219
homogenization clusters. It is often observed that a slight homogenization increases HCT
which cannot be easily explained.
The heat stability can be improved by lowering the calcium content of the milk before
evaporation by means of ion exchange. Addition of 15 mmol H2O2 (0.05 %) or of about 15
µmol Cu++ (0.5-1 mg.kg-1) after preheating but before evaporation increases the heat
stability.
12.1.5 Creaming
Creaming of evaporated milk eventually leads to formation of a solid cream plug that
cannot be re-dispersed. Partial coalescence or bringing of adjacent fat globules due to
“fusion” of the fragments of casein micelles in their surface layers may be responsible.
Accordingly, intense homogenization is necessary.
The newly formed surface layers during homogenization can be fairly thick. pre-heating has
left hardly any dissolved serum proteins and the evaporation and sterilization steps have
increased the average diameter of the casein micelles. Especially after homogenization at
high pressure and low temperature, the layers may be thick enough for the globules to
have a higher density than the plasma; consequently, they sediment rather than cream. As
a result, the fat content of evaporated milk in booth the top and bottom layers of a can than
that in the middle.
A higher viscosity of the evaporated milk often involves a slower creaming, but the relations
are not straightforward. The viscosity of the plasma phase, not that of the product, that
determines creaming rate. Generally, a high viscosity is due to approaching heat
coagulation. The homogenized fat globules tend to participate in this coagulation. The
homogenized fat globules tend to participate in this coagulation, hence to form clusters that
would cream rapidly. Addition of traces of cupper, usually lead to a decreased creaming.
UHT evaporated milk can be homogenized far more intensely because the sterilization
precedes the homogenization. Intense homogenization is also required to prevent
excessive creaming because the viscosity of the plasma phase is much lower than in
conventional evaporated milk.
12.1.6 Age thickening and gelation

Age thickening is a most serious problem associated in condensed/concentrated or

evaporated milk. This property is thixotrophic and so as the high initial viscosity can be
220
broken down by stirring to give a much lower viscosity, which, however, increases with age.
When evaporated milk is kept, its viscosity may initially decrease slightly. This may be
explained in terms of casein micelle aggregates changing from an irregular to a spherical
shape, as a result of which the effective volume fraction decreases (Fig. below).
Subsequently, the viscosity tends to increase, and it becomes dependent on the shear rate
(hence, it is an apparent viscosity). Soon the milk displays a yield stress and a gel is
formed that firms rapidly. The mechanism is not quite clear. In most cases it is not caused
by proteolytic enzymes neither are Maillard reactions responsible, through the latter parallel
gelation. Moreover, gelation is not related to heat coagulation. For intense, it does not
depend significantly on the pH and its rate increases rather than decreases after lowering
of the calcium content. Electron microscopy reveals that thread-like protrusions appear on
the casein micelles, which eventually from a network (C & D).

A B C D
Fig.12.3 The change observed in the casein micelles of evaporated milk during
storage. The apparent viscosity is at a minimum in stage B.
It is observed that a slow change in the micellar Ca(PO4)2 is at partly responsible for the
changes observed.
Age thickening and gelation tend to occur fast in UHT evaporated milk. It may then be due
to proteolysis caused by enzymes released by psychrotrophs, but also if such enzymes are
absent, fast gelation occurs. A more intense sterilization after evaporation delays gelation.
Gelation is faster in more concentrated milk and at a higher storage temperature. Addition
of sodium polyphosphate (about 0.4 % in the dry matter) delays gelation considerably; the
higher the molar mass of the phosphate, the more effective it is. Addition of citrate or
orthophosphate often accelerates gelation, presumably because of binding of calcium.
Polyphosphates may be hydrolyzed to yield orthophosphate, especially during heating.
Consequently, addition of polyphosphate does not counteract gelation of in-bottle sterilized
evaporated milk, to the contrary.

221
Conventional evaporated milk only gels if kept for a long time at a high temperature (as in
the tropics). Rapid gelation can occur, however, if the evaporated milk before its
sterilization is kept refrigerated at, say 4oC for a few days.
Adequate measures can be taken to delays gelation of the evaporated milk for a
considerable time. Gelation can be examined by suspending a can of evaporated milk on a
torsion wire and checking whether the milk has elastic properties. If so, the can will keep
oscillating for a while when it is given a turn and then released.
12.2 Sweetened Condensed Milk
12.2.1 Description
Usually, sweetened condensed milk contains 8-9 % fat and 28-31 % total solids.
Furthermore, 43-46 % sugar (can sugar) is added. The high sugar content gives the
product good keeping qualities as it causes the osmotic pressure in the aqueous phase to
become so high that most micro-organisms are unable to develop. To obtain this effect, the
sugar content in the aqueous phase must be at least 62.5 %, which value may be
calculated according to the following equation:
% sugar in the finished product × 100
% sugar in aqueous phase =
100 − % milk solids
The sugar content in the aqueous phase should not exceed 65.5 % as this increase the risk
of sugar crystallization. In addition to the standardization of the desired ratio of MSNF to fat,
the ratio of MSNF to sugar must also be adjusted. The latter should be approx. 1:2.
After standardization of the fat content and the addition of sugar, the milk is pasteurized at
115-120oC for 0.1-2 min. The product is evaporated to a TS content of approx. 72 %.
When evaporation has been completed, the concentrate will be supersaturated with
lactose, which has a strong tendency to crystallize. To prevent too widespread formation of
large crystals (max. 10 µm), which causes the defect sandiness and increases the risk of
sedimentation, cooling must be rapid. Such crystals formation can also be inhibited by
seeding the product with small lactose crystals after the temperature has fallen to 30-34oC
followed by rapid cooling to 20oC and crystallization in a tank – during which the product
must be constantly stirred. The product is then packaged in tins or large drums.

222
Sweetened condensed milk will keep for 2-3 months and is used in coffee, for desserts, ice
cream, chocolate and caramel confectionery, and in diluted form for drinking, cooking and
other purposes.
12.2.2 Manufacture
A flow diagram of manufacturing process for sweetened condensed milk has been shown
in figure below. Some of the important steps have been discussed here.
Heating
Pathogens and potential spoilage organisms must be killed. Among the enzymes, milk
lipase should primarily be inactivated; bacterial lipases are not inactivated and, if present,
can cause severe deterioration. Deterioration caused by proteinases has not been
reported. The heating intensity considerably affects viscosity, age thickening, and gelation
of the product, so the actual heat treatment must be adjusted to these properties. UHT
treatment at about 130-140oC is commonly applied.
Milk

Cleaning and standardizing

Heating to 135oC for 5 s

Sugar
Homogenizing 70oC, 4 MPa

Dissolve hot Evaporation

Water Cool to 50oC

Seed lactose 0.05 % Seeding

Cans Cool to 18oC

Packing

Fig.12.4 The manufacture of sweetened condensed milk (SCM).

12.2.3 Homogenization
Creaming is not a big problem, so homogenization is not always applied. Now a day SCM
is made less viscous. If homogenization is not done, the creaming rate would be about 1 %
of the fat per day. This is too high, so that homogenization is often applied, though at low
pressure, i.e., 2-6 MPa. Homogenization also increases the viscosity of the product slightly.

223
 Sugar can simply be added to the original milk. The amount added can be adjusted
readily and accurately, and the sugar is pasteurized along with the milk. This will cause
faster Maillard reaction during heating and evaporation, and above all a faster age
thickening. Likewise, evaporation is more difficult (especially when a multiple effect
evaporator is used) because of a further increase in boiling point and a decreased
coefficient of heat transmission (due to the higher viscosity). It is better to use a
concentrated refined sugar solution which is sufficiently heat treated to kill osmophillic
yeasts, at the end of evaporation step. To prevent excessive Maillard reactions avoid of
invert sugar.
Concentration is usually done by evaporation, but reverse osmosis can also be used. A
falling film evaporator is usually used to remove the bulk of the water and another
evaporator is usually used to remove remainder. The latter is usually a conventional rising
film evaporator with separate steam supply, in which part of the product recirculates to
ensure that the highly viscous SCM keeps the heating surface covered. To achieve that,
somewhat higher temperature (up to 80oC) is often applied, which implies a lower viscosity
in the evaporator but a higher initial viscosity of the final product.
The low water content of the SCM implies high viscosity and boiling point. Evaporation
is continuously operating equipment with many effects is therefore not easy(the stem
management is thus less economical). Fouling thus readily occurs. It is hard to accurately
adjust the desired water content, which is mostly monitored by means of refractive index.
This method can be effectively used because SCM is hardly turbid.
12.2.4 Cooling and Seeding
The formation of large lactose crystals must be avoided. Consequently, seed lactose is
added. Before that, the condensed milk must be cooled to a temperature at which lactose is
supersaturated so that the seed lactose does not dissolve. However, the temperature must
not be so low that spontaneous nucleation can occur before the seed crystals are mixed in.
After seeding, cooling should be continued to crystallize the lactose. In a continuous
process, the cooling should be fast, but that is by no means easy for such a highly viscous
product. A vacuum cooler is often used. In which a thin layer of the milk passes the wall of
a vat that is under vacuum. Cooling from 50oC to 18oC causes some 3 kg of water to

224
evaporate from 100 kg of SCM, which needs to be taken account of during evaporation.
Alternatively, the SCM can be cooled in a scraped-surface heat exchanger.
Packing in cans is common. The cans are then covered with a lid and the seams sealed.
Cans and lids are first sterilized, e.g., by flaming. The packing section is supplied with air
purified through bacterial filters. In this section, rigorous hygienic standards are paramount.
12.2.5 Microbial Spoilage
SCM is not sterile. It contains living bacteria and spores. The low water activity (about 0.83)
or, rather, the high sugar content prohibits growth of most but not all microorganisms.
a. Deterioration usually occurs by osmophillic yeasts, most of which belong to the genus
Torulopsis. The yeasts often cause gas formation (bulging cans), a fruity flavor, and
coagulation of protein. Coagulation may result from ethanol production. As a result, the
product becomes unacceptable. The yeasts do not start easily, especially is the sugar
concentration is high. It may thus take several weeks for incipient growth to be
perceptible.
b. Some Micrococci may grow in SCM, though slowly, especially if water activity and
temperature are high. Presumably, the presence of oxygen is required. It may happen
that they grow to reach a colony count of, say, 105 ml-1 and then stop growing,
aggregates eventually form and several off-flavors develop.
c. Some molds, especially strains of Aspergillus repens and A. glaucus, can grow as long
as oxygen is present. If so, fairly firm colored lumps are formed and an off-flavors
develops. One spore in one air bubble can cause such a lump.
Remedies
Killing of all saprophytes and mold spores in the milk and in the sugar. No single bacterial
spore can germinate in SCM. Growth of harmful microorganisms in the dairy plant should
be rigorously avoided. No sugar and residues of the milk should be left around.
Satisfactory hygienic standards must therefore be maintained, especially in the packing
section. The machinery used for evaporation must be thoroughly cleaned immediately after
processing. Any mold spores can be removed by air filtration. No air is left in the cans
during packing. The self life evaluation of cans have been made at elevated temperature,
say, 35oC.

225
12.2.6 Chemical deterioration
The main change in SCM during storage is presumably age thickening and, finally,
gelation. SCM is far more concentrated than evaporated milk. Nevertheless, it does not
thicken markedly faster with age. It is usually assumed that added sucrose inhibits age
thickening; other sugars or hexitols have a similar effect. An important difference from
evaporated milk is the far higher viscosity of the continuous phase of SCM. Sucrose
increases the Ca++ activity. Another difference with evaporated milk is that an initial
decrease in viscosity before age thickening is not observed but that need not be surprising
because SCM would contain no flocculated casein micelles. The viscosity η’ increases
almost linearly with time. The thickened sweetened condensed milk after age gelation
shows considerable shear rate thinning, and the effective shear rate during the viscosity
measurement is often not reported.
The main factors affecting the age thickening are:
a. Kinds of milk: Variation occurs among batches of milk, often with an effect of season.
Milk of cows in early lactation may be more sensitive.
b. Preheating of milk: Longer heating times at UHT temperatures lead to little age
thickening. The heating affects the initial viscosity considerably and that will be the main
effect, i.e., the lower the initial viscosity, the lower the volume fraction of casein
particles, and the longer the elapsed time before a gel is formed (assuming the volume
fraction of the casein particles increases at the same relative rate.)
c. Stage at which sugar is added: The latter in the evaporating process, the less the age
thickening.
d. The concentration factor: The higher the concentration factor, the more the age
thickening.
e. Stabilizing salts: Salts are added up to, say, 0.2 %. Adding a small amount of sodium
tetra polyphosphate (0.03 %) mostly delays age thickening, whereas adding more often
has the opposite effect.
f. Storage temperature: Age thickening considerably increases with storage temperature:
Q10 = 3.4. At tropical temperatures, gelation inevitably occurs within about 1 year.

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12.3 Lactose crystals
SCM contains around 38-45 g lactose per 100g water. The solubility of lactose at room
temperature to be about 20 g per 100 g water, but in SCM the solubility is about half as
large due to the presence of sucrose. It implies that 75 % of the lactose tends to crystallize,
meaning about 7.5 g per 100g SCM. Without special measures, SCM would obtain a
relatively large quantity of lactose in the shape of large crystals. These crystals sediment
(after sedimentation an even distribution throughout the viscous liquid is hard to achieve)
and are responsible for a sandy mouth-feel. Although the crystals are not so to be left singly
in the mouth, they can be large enough to cause a non-smooth impression. To avoid this,
they should be smaller than about 8 µm in length.
Preventing crystallization is not possible and, accordingly, a large number of crystals should
be the goal. This may be achieved by using an ingenious cooling schedule, but fully
satisfactory results can only be obtained by using seed lactose. Adding 0.03 % seed
lactose represents 0.004 time the amount of lactose to be crystallized. The final size of the
crystals in the product should not exceed 8 µm. Consequently, the seed lactose would
contain enough seed crystals (one per crystal to be formed) if its crystal size does not
exceed about (0.004 × 83)1/3 = 1.25 µm. Such tiny crystals can be made by intensive
grinding of lactose, which is crystallized as usual. Another method is as follows: A lactose
solution is spray dried. Then the amorphous lactose powder is left to absorb just enough
water (at aW ≈ 0.4) to allow crystallization, which gives extremely small crystals (<1 µm
length) embedded in amorphous lactose. Grinding readily pulverizes the powder into
separate crystals.
In addition to lactose crystals, sucrose crystals can also be formed if too much sucrose has
been added or the milk has been concentrated too far, and the sweetened condensed milk
is kept at a low temperature. Since the super saturation of sucrose would be slight, large
crystals are formed that make the product definitely sandy.

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Chapter 13
Production of powder milk

13.1 Objectives

a. To convert the liquid perishable raw material to a product that can be stored without
substantial loss in quality, preferably for some years. Decrease in quality mainly
concerns formation of gluey and tallowy flavors (due to Maillard reactions and
autoxidation, respectively) and decreasing nutritive value (decrease in available lysine).
b. The powder should be easy to handle. It should not dust too much or be overly
voluminous. It should be free-flowing, i.e., flow readily from an opening, and not stick to
the walls of vessels and machinery. The latter requirement is of special importance for
powder used in coffee machines, etc.
c. After adding water the powder should be reconstituted completely and readily to a
homogeneous mixture, similar in composition to the original product. Complete
reconstitution means that no undissolved pieces or flakes are left and that neither butter
grains nor oil droplets appear on top of the solution. “Readily reconstituted” means that
during mixing of powder and water no lumps are formed because these are hard to
dissolve. In the ideal situation the powder will disperse rapidly when scattered on cold
water; this is called “instant powder”. If the powder is instantizing to attain this property.
d. The reconstituted product should meet specific requirements to its intended use. If the
use is beverage milk, the absence of a cooked flavor is of importance. If it use for
cheese making it should have good clotting properties. It should have a satisfactory heat
stability if used to make recombined evaporated milk. So there are several widely
divergent requirements that can not be reconciled in one powder. It is not possible to
make whole milk powder that has no cooked flavor and at the same time develops no
oxidized flavor during storage. For the first only a mild heat treatment is allowed,
whereas for the second intensive heating is needed
e. The product must be free of health hazards, be it toxic substances or pathogenic
organisms.

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13.2 Composition
Table 13.1 The approximate composition of some types of powder.
Powder from
Constituents
Whole milk Skim milk Whey Sweet cream butter milk
Fat 26 1 1 5
Lactose 38 51 72 48
Casein 19.5 27 0.6 26
Serum protein 4.8 6.6 8.5 6.2
Ash 6.3 8.5 8 8
Lactic acid - - 0.2-2 -
Water 2.5 3 3 3

13.3 Manufacture
Milk

Separating

Skim milk
Cream
Standardizing
Closing
Preheating to 95oC, 1 min Gas flushing

Evaporating 45 % DM Tins Packing

Homogenizing 15 MPa Bags Waiting (4 days)
Cooling/Storing Bagging Gas fluishing
Heating to 78oC
Storing in silo
Spray drying (8 % water)

Lechithin Separating powder Fines Cool to 25oC

Fluid bed drying/Agglomerating Separating Agglomerated

powder

Fig.13.1 Flow chart for the manufacture of whole milk powder.

Intense pasteurization is needed to obtain resistance to autoxidation. Vapor from the last
stage of the evaporator is often used to supply a part of the heat needed for pasteurization.
The pasteurizing step and the heating to the required temperature before evaporation are
combined. If a nozzle is used for atomization it may effectively disrupted the fat globules so
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homogenization is not needed. Homogenization of highly concentrated milk will increases
the viscosity (since the transfer of large casein micelles to the fat globules gives the latter
such an irregular shape as to increase the effective volume fraction of fat globules plus
casein micelles).This will cause the coarser droplets during atomization. If the concentrated
milk is not homogenized, evaporation can be up to a higher dry matter content. Storage
(buffering) of the concentrate before atomization is not always applied, it is done partly to
overcome differences in capacity between evaporator and drier. The concentrate should
not be kept warm for more than a short time to prevent the growth of microorganisms. A
refrigerated concentrate is too viscous to be atomized readily and it is therefore heated.
The later must be done just prior to atomization because otherwise the viscosity increases
again (age thickening). The heating can at the same time serve to kill bacteria that may
have recontaminated the concentrate.
Lecithinizing during the drying in the fluid bed is not always applied; it is meant to obtain
instant properties. The so-called gas flushing, essentially displacing air by N2 or a mixture
of N2 and CO2, is to remove a considerable part of the oxygen and thereby to improve the
stability toward autoxidation; it can be done once or twice. If it is not done, the powder may
be packed in to multilayer paper bags with a polyethylene inner layer. While milk powder,
however, is often packed in tins or plastic containers to minimize oxygen uptake.
The pasteurization can be less intense (at least phosphatase negative) in case of
production of skim milk powder. Homogenization is not necessary. The milk can be
concentrated to some what higher solid content. Nor are lecithinizing and gas flushing
carried out. Sometimes vitamins preparations are added, especially vitamin A.
The manufacture of whey powder is largely similar to that of skim milk powder. At first, curd
fines should be removed from the whey by filtration or by means of a hydrocyclone, and the
whey should be separated. Sour whey can cause fouling of the machinery so problems
arise during processing. Sour whey or skim milk) may be neutralized with alkali.
Whey can be evaporated to more than 60 % dry matter, but then crystallization of lactose
readily occurs. The lactose in the evaporated whey to crystallize as completely as possible,
e.g., by keeping it for 3 h at 25oC while stirring. If the dry matter content is over 60 %,
seeding with lactose crystals is not necessary. Atomization should be with a disk, as a
spray nozzle would be blocked. The precrystallized whey powder then obtained has some

230
attractive properties, especially as regards caking. The conventional methods for
determining the water content of powders do not remove the bulk of the water of
crystallization of α- lactose monohydrate, hence, crystallization of 80 % of the lactose yields
up to maximum of 3 % “more” whey powder. In this way crystallized skim milk powder also
can be made, but then a longer crystallization time and seeding with lactose crystals are needed.
It is done by spray-drying or by roller-drying (drum-drying). In spray-drying, the water content of full
fat milk or skimmed milk is decreased by evaporation to 50-65% of the original content. All milk
components, i.e., proteins, lactose, minerals, and fat are retained in evaporated milk. It is then
atomized (dispersed) into minute droplets and these are passed through hot air in a drying tower.
The inlet air is quite hot (~220°C) but its temperature is
decreased as its energy is used to evaporate water and at the
outlet it is less than 100°C. The droplets of evaporated milk turn
into powder particles as the water present in the milk evaporates.
During this process, lactose dries in an amorphous form which
holds no water. Although this form is very hygroscopic (which
means that is readily takes up water), the primary milk powder dissolves in water with difficulty - it
foams and forms lumps. Because of its hygroscopicity, the powder easily bakes into blocks. The
powder may appear is various forms. The form reminiscent of a dried apple (up at right) is the most
common. Fine superficial wrinkles on the powder particles are believed to be associated with the
presence of casein micelles in the powder.

A small particle Dents in the large particle Roller-dried milk particles

(surrounded by a 'crater' have been caused by small are produced as
rim) has been captured by hard dry particles. fragments of a dried milk
a large particle. sheet.
Atomization of evaporated milk produces
minute droplets of varying dimensions.
Small droplets dry and harden more
rapidly than larger droplets. When such
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hard particles collide with larger particles which are still sticky, they lodge in the sticky
surfaces and eject some material from them.
Instantization results in agglomeration of powder particles
Exposure of such milk powder to excessively humid air leads to an extensive crystallization
of lactose as is evident in the micrograph at left. However, a controlled crystallization of
lactose is used to alleviate problems encountered with the primary milk powder when
reconstituting it, i.e., dissolving it in water. Instantization has been introduced into the
production of spray-dried milk powders. It is based on controlled crystallization of lactose in
the form of a monohydrate, which means that in the crystals there is one water molecule
per each molecule of lactose. Instantization is achieved by wetting the milk powder by
exposing it to a humid atmosphere before it settles at the bottom of the drying chamber.
Water mist, mist of 10% skim milk, or steam is used to humidify the atmosphere. Under
these conditions, lactose crystallizes and the powder particles agglomerate (micrograph at
right). The agglomerated powder is transported to a vibrating fluid drier where it is redried
and then cooled.
13.4 Hygienic aspects

The requirements for bacteriological quality of milk powder partly depend on its intended
use and also on manufacturing process. Whether the powder is used for direct
consumption or it is used for heat treatment after reconstitution ( e.g., for recombined milk).
The heat treatment during the manufacture of (skim) milk powder, classified as “low heat”
usually is not more intense than the heat treatment during low pasteurization (say, 75oC for
20 s). Consequently, many bacteria may survive the manufacturing process.
The causes for milk powder to be bacteriologically unacceptable or even unsafe can be of
three kinds:
a. In the fresh milk bacteria are present that are not killed by the heat treatments to which
the milk is subjected before and during drying.
b. Conditions during the various process steps until the product is dry do allow growth of
some bacteria.
c. During manufacture, incidental contamination may occur. The level of contamination is
generally low and remains low if the bacteria involved cannot grow.

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 1. Bacteria in the original milk
In deep-cooled milk psychrotrophic gram-negative rods can develop during prolonged
storage (e.g., Pseudomonas spp.). As is well known, these bacteria do not survive even a
mild heat treatment. Proteases and lipases formed by these rods may survive and become
incorporated into the powder. Prevention of the growth of these bacteria (refrigeration,
limiting storage time, Thermalization) is discussed previously. Contamination and growth
during storage of the thermalized milk should be avoided.
The heat resistance bacteria and bacterial spores are taken of great importance. They
survive low pasteurization (72oC for 15 s), and most are not killed during evaporation and
drying. Due to concentrating, the powder contains about 10 times as many bacteria per
gram as the milk immediately after preheating. A more intense pasteurization will kill the
heat-resistant streptococci (e.g., S. faecalis, S. thermophillus ) and in a high-quality
medium-heat or high-heat milk powder only bacterial spores and Microbacterium lacticum
can originate from the original milk.
Among the aerobic and anaerobic spore-forming bacteria, especially Bacillus cereus and
Clostridium perfringenes, are important to the powder quality the reconstituted milk is to be
used for cheese making, a very low content of gas forming anaerobic spore forming
bacteria ( C. tyrobutyricum and C. butyricum may be essential. All of these bacteria are
likely to originate mainly from contamination during milking (dung, soil). A low count of
anaerobic spore forming bacteria points to a good quality silage (silage being the good
source of most of the Clostridia : feed → cow→dung → milk). But the pathogenic C.
perfringens usually does not originate from the silage, though from the dung. Hence, a low
content of anaerobic spore forming bacteria need not be an indication of the absence of C.
perfringens. Likewise, the total count of aerobic spore forming bacteria is not always an
indication for the spore number of B. cereus. Usually, the total count is higher during winter,
but the count of B. cereus may be highest in summer and autumn. This probably is
because at higher environmental temperatures B. cereus can develop and sporulate in
imperfectly cleaned and disinfected machinery, outside the operating periods. Obviously,
during the first few hours of processing, slightly higher counts of B. cereus are found than
latter on. To kill bacterial spores, heat treatment at 90-110oC for 10-20 s is insufficient, a
UHT treatment should be applied. The D-value is about 4 s at 125oC for B. cereus as well

233
as for C. perfringens, so that heating for 15-20 s at that temperature would cause a
sufficient reduction of these spores.
2. Growth during manufacture

Temperature and water activity during the successive steps in the manufacture are such
that some Thermophillic bacteria can readily grow, while they are not or insufficiently killed
during drying. The type of bacterium often is characteristic of the cause of the
contamination.
In the regeneration section of pasteurizers and thermalizers (and possibly in the part of the
evaporator plant where the milk is heated in counter flow), S. thermophillus can develop in
particular. The bacterium grows fastest at 45oC but scarcely at temperatures over 50oC. It
does not grow in the evaporator because the temperature is too high in the first effects,
whereas in the later effects aW mostly is too low. Since S. thermophillus in the milk just
before the preheater may give the good indication of the fouling of the heating section of
the evaporator and of the moment at which it should be cleaned. In some drying plants that
employ a wet washer to recover powder fines, the outlet air is brought into contact with a
film of milk rather than water, thereby preheating the milk and saving energy; this implies
that the preheated milk acquires the wet bulb temperature (about 45oC), which leads to
ideal growth conditions for S. thermophillus.
The conditions in the second half of the evaporator and in the balance tank are not optimal
for S. thermophillus. Especially D streptococci (S.faecium being an important
representative) will start to grow. If the milk is properly preheated and the plant is
satisfactorily cleaned and sterilized, it will take a rather long time, however, before
substantial counts of S. faecium often is the predominant species.
Likewise, the conditions in the second part of the evaporator and in the balance tank are
favorable for growth of Staphylococcus aureus. The bacterium generally killed by
pasteurization, and strains of Staphylococcus aureus in the milk powder have been shown
to have phage characteristics different from the strains in raw milk. They originate from
direct and indirect human contamination. Heat stable enterotoxins can be formed at counts
of 10-7-10-8 per ml. The amounts formed may cause the powder to be a health hazard.
Although S. aureus is not heat resistant, the conditions during drying appear to be such that
complete killing is not achieved. Roughly 10-5 to 10-1 of the initial count of these bacteria
234
have been found to survive under various practical conditions. This means that S. aureus
can at least be found in 1 g of fresh powder if its counts before the drying was so high that
production of enterotoxins could have occurred.
Bacillus sterothermophillus can readily grow at high temperatures. Its growth range is from
45oC to 70oC, with an optimum near 60oC. It can also grow in concentrated milk and
therefore throughout the equipment between preheater and drier. Moreover, the bacterium
can form spores under these conditions, which further limits its killing during drying. Some
growth of Bacillus sterothermophillus will always occur during manufacture even in a
cleaned and disinfected plant. However, under normal conditions this will not cause
problems.
The concentrated milk may be pasteurized just before it enters the drier. The temperature
of 78oC for 45 s causes a considerable reduction of S. faecalis and S. faecium.
3. Incidental Contamination
A distinction can be made between contamination before the drying (wet part)and after the
drying (dry part). Bacteria involved in these types of contamination generally do not grow
during the process and contribute little to the count of the powder.
Contamination after preheating and before drying can readily occur if the equipment has
been insufficiently cleaned. This is only important if the bacteria involved can survive the
drying (and the pasteurization before drying, if carried out).
About 70 % of the S. faecalis and S. faecium survive during drying whereas survival of S.
aureus varies widely. About 10-4 to 10-5 of the initial count of Salmonella spp. and E.coli will
survive. Due to the relatively low level of contamination, the powder may be expected to
contain no appreciable counts of enterobacteriaceae, immediately after the drier.
Contamination of powder occur at many places- in the spray dryer, during fluid bed drying,
and during packing. Contamination via (in)direct human contacts should also be considered
(e.g., S. aureus). Bacteria can easily survive in dry powder, and undesirable bacteria can
start to grow if the water content increases to over 20 %. The supply of cooling air into the
drier and into the fluid bed drier can be a source of direct contamination. It may also be
responsible for indirect contamination because it gives, at certain sites, better conditions for
survival and growth of bacteria in remnants of not fully dried powder.

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4. Sampling and checking
Bacteria originating from contamination or growth prior to drying will usually be
homogeneously distributed throughout the powder and can cause no problems with
sampling. This is different for bacteria originating from incidental contamination of the
powder, which may be distributed quite inhomogeneously. It is not possible to devise
sampling schemes that guarantee detection of incidental contamination. To ensure that a
product is bacteriologically safe, not only the powder should be sampled; that a product is
bacteriologically safe, not only the powder should be sampled; indeed, samples must also
be taken at sites that are potential sources of contamination.
13.5 Physical properties
The milk powder particle generally consists of a continuous mass of amorphous lactose
and other low-molar mass components in which fat globules, casein micelles, and serum
protein molecules are embedded. The lactose generally remains amorphous, the time
available for its crystallization being too short. If, however, pre-crystallization has taken
place, large lactose crystals may be present (some tens of micrometers in size). If lactose
has been allowed to crystallize afterward due to water absorption, its crystals are generally
small (about 1 µm). Most fat globules are less than 2 µm, but a small proportion of the fat
(e.g., 2 % of it) is to be found as a thin layer on parts of the surface of the powder particles.
The vacuole volume v of most powders amounts to 50-400 cm3.kg-1. When pre-crytallized
whey powder is examined microscopically, most of the particles look more like lactose
crystals of the tomahawk shape to which other material adheres.
Free flowingness refers to the ability of a powder to be poured. The property may be
determined by pouring out a heap of powder under standardized conditions and measuring
its angle of repose α. α = r/h.

13.6 Ease of dispersing (Instant powder)

An important property of milk powder is its ease of dissolution in water or, more precisely,
of dispersing it. With hot water and a high-speed agitator, dissolution causes few problems,
but dissolving the powder in cold water under household conditions may be far from easy.
Instant powder disperses rapidly in cold water with gentle stirring. The dispersibility is not
related to the solubility of the powder, but to the rapid penetration of water in a mass of

236
powder. Due to this penetration the powder particles disperse separately in the water,
where they can subsequently dissolve.
The instant properties depend on the phenomena occurring when a quantity of powder is
brought in to or upon water.
The powder should be wettable by the water. Consider two powder particles close together
on a water surface. The wetting depends on the contact angle (θ, as measured in the
aqueous phase) of the system consisting of dried milk, water, and air. The contact angle
depends on the three interfacial tensions: solid-water, solid-air, air-water. For a hydrophobic
solid θ is large, for a hydrophilic solid it is small. If θ is less than 90o, the particles are
wetted. For dried skim milk θ is about 20o, for dried milk about 50o. This would imply that
water is always sucked into the pores between the particles by capillary forces: If the water
surface is curved, the capillary force acts in the direction of the concave side, i.e., upward.
13.7 Influence of process variables on product properties
13.7.1 Flavor
Milk powder often has a cooked flavor, which results from the flavor components formed
during pre-heating and possibly during evaporation. During drying, conditions are mostly
not such that off-flavors are induced. On the contrary, a considerable part of the volatile
sulphydryl compounds (esp. H2S) is removed. Obviously, a cooked flavor in milk powder
mainly results from methyl ketones and lactones formed by heating of the fat (they thus are
almost absent in skim milk powder) and from Maillard products.
13.7.2 WPN index
Heat treatment of the original product, concentrate, and/or drying droplets can cause
denaturation of serum proteins, though the conditions during drying are rarely such as to
cause extensive heat denaturation. The extent of denaturation is an important quality mask
in connection with the use of milk powder. If the dissolved powder is to be used for cheese
making, practically no serum protein should have been denatured in view of the
renneability, in infant formulas, on the other hand, the rennetability should be poor.
The extent of the denaturation of serum protein can be used as a measure for the heating
intensity applied. This also holds where denaturation by itself may be of no importance but
other changes, associated with intense heat treatment, are. An example is the flavor of
powder to be used in beverage milk, which require a mild heat treatment. A good stability
against heat coagulation in the manufacture of recombined evaporated milk, or a high
237
viscosity of the final product in the manufacture of yoghurt from reconstituted milk, requires
intensive heat treatment. The latter is also desired if the powder is used in milk chocolate;
presumably, Maillard products contribute to its flavor.
The whey protein nitrogen (WPN) index is a generally used to classify milk powders
according to the intensity of the heat treatment(s) applied during manufacture. To the end,
the amount of denaturable protein left in the reconstituted product is determined, usually by
making acid whey and determining the quality of protein that precipitates on heating the
whey. This can be done by Kjeldahl analysis of protein nitrogen or by means of a much
easier turbidity test that is calculated on the Kjeldahl method. The result is expressed as the
quantity of undenatured serum protein per g skim milk powder. The classification is as
follows:
WPN ≥ 6 mg per g : “low heat”
6 > WPN > 1.5 mg N per g : “medium heat”
WPN ≤ 1.5 mg N per g : “high fat”
13.7.3 Insolubility
It can be determined in various ways. In all tests, powder is dissolved under standardized
conditions (concentration, temperature, and duration and intensity of stirring), then the
fraction that has not been dissolved is determined (e.g., volumetrically after centrifugation
or by determination of dry matter). Often one refers to this as a “solubility index”, but this is
a confusing expression because it concerns an insolubility figure or insolubility index. The
insoluble fraction-essentially the material that sediments during centrifugation-will
predominantly consists of protein. In whole milk powder, flocks of coagulated protein with
entrapped milk fat globules (the so-called flecks) may float to the surface; the quantity
involved usually is more than the sediment. Hence, the insolubility found closely depends
on the method used.
The insolubilization of a fraction of the milk powder is related to heat coagulation.
Consequently, the extent to which it occurs greatly depends on the time during which the
drying material is at high temperature and on the degree of concentration during drying.
The preheating has an effect: high preheating → higher viscosity → larger droplets on
atomization → more intense heat treatment during drying → increased insolubility. But
evaporation to a certain viscosity would imply a lower degree of concentration for highly

238
preheated milk, so that heat coagulation and insolubilization during drying would be less.
Finally, homogenization of the concentrate may increase the insolubility, but this may hardly
be noticeable because considerable homogenization occurs during evaporation and
atomization.(The insolubility of whole milk powder is indeed increased most readily than
that of skim milk powder). Homogenization of the concentrate may also lead to greater
insolubility if it considerably raises its viscosity (thus at a high degree of concentration),
increasing the droplet size and thereby the drying time.

13.7.4 Specific volume

Specific packed or bulk volume is equal to 1/ρb.
ρb = bulk density or packing density of the whole powder. Sometimes a distribution is made
between specific bulk volume (if the powder is lightly packed) and specific packed volume
(if the powder is allowed to set, e.g., by tapping). For determination of the latter, a tapping
apparatus may be used, which repeatedly brings a graduated cylinder with powder to fall on
a solid base. Obviously, the packed volume is of great importance in connection with the
mass of powder that can be stored in a certain package. The volume should not be greatly
reducible by tapping because otherwise a can, being full of powder when filling, may
appear to be partly empty later on when used.
The particle density ρb includes the vacuoles and is therefore given by ( v in m3.Kg-1)
ρt
ρp =
1 + vρ t
In skim milk powder ρp usually is 900 – 1400 Kg.m-3. The bulk density or packing density of
the whole powder, ρb, is given by
1− ε
ρb = ρp (1 - ε) = ρ t
1 + vρ t
where, ε is the void volume fraction or porosity, i.e., the volume fraction of voids (pores)
between the particles. Generally, ε = 0.4 – 0.75, but it depends on the way of handling the
powder. It decreases considerably when a lightly bulked mass of powder is set by means of
tapping or shaking, e.g., from 0.75 to 0.45 for whole milk powder and from 0.55 to 0.40 for
skim milk powder. ε depends also on other factors. All in all, ρb can widely vary, for
instance, from 300 to 800 kg.m-3.

239
Generally, factors causing a higher viscosity of the concentrate during atomization, and a
lower average drying temperature will lead to a lower v and thus to a higher ρb. The drying
temperature, however, may also affect ε, and not all of those are easy to explain. In
general, ε will be higher if the powder particles are more irregular in shape differ less in
size. Accordingly, agglomeration and removal of small powder particles clearly raise ε,
hence decrease ρb. ε may decrease slightly with increasing water content of the powder;
presumably, it causes the particle surface to become smoother. Precrystallization of lactose
makes the particles more angular and raises ε slightly.
Obviously, ρb increases due to shaking, tapping, or vibrating. In general, the effect
produced is reversible. But the agglomerated powder the agglomerates can be disrupted;
consequently ρb is increased irreversibly and the instant properties are impaired.
13.7.5 Free flowingness
The higher the ε, the poorer the free-flowingness.
13.7.6 Free fat content
The term “free fat content” is, properly speaking, incorrect because it suggests that some
fat entrapped in the powder particles without a surrounding membrane. So-called free fat is
determined by extraction with an organic solvent. In fact, the fat is then extracted from all
fat globules that are in contact with the surface of the particle, or with a vacuole, with pores
or cracks, etc. The amount of extractible is higher as the powder particles are smaller, a
greater number of vacuoles have been entrapped, and the drying has been performed at a
higher temperature (giving more cracks). Increasing the water content of the powder
decreases the amount of extractible fat since cracks are closed due to swelling.
The amount of extractible fat can be reduced by intensive homogenization. Only in milk
powder with less than 20 % fat does this affect the dispersibility, and then only slightly.
13.7.7 Dispersibility
Discussed under the ease of dispersing.
13.7.8 Stability
Changes in properties during storage are discussed below.
13.8 Deterioration
The rate of undesirable changes in milk powder is the water content. When comparing
different types of powder, it is probably easiest to consider water activity (aw). The higher aw

240
of whole milk powder as compared to skim milk powder as the same water content is
caused by the fat not affecting aw. Whey powder has an aw slightly different from that of
skim milk powder because in a dry product the soluble constituents (especially sugar and
salts) decrease aw somewhat less than casein. This only holds, however, as long as all
lactose is amorphous, which often does not apply to whey powder. aw is considerably
reduced if lactose crystallizes without the powder absorbing water, at least at aw less than
0.5. Crystalline lactose thus binds water very strongly and that is also why the usual
methods to determine the water content do not include the bulk of the water of
crystallization. If the water content excluding the water of crystallization is taken as a basis,
then aw, is even higher for the powder with crystallized lactose.
It thus is advisable to make milk powder sufficiently dry and to keep it sufficiently dry. If it is
not hermetically sealed from the outside air, it will attract water in most climates. The higher
the temperature, the higher the water activity. Several reactions are faster at a higher aw,
this implies that a temperature increase often causes an extra acceleration of deterioration.
Microbial and enzymic deterioration are rare in milk powder. For microbial deterioration to
occur, aw should increase to over 0.6 (and for the majority of microorganisms much higher),
such a high aw is only reached if the powder is exposed to fairly moist air. Deterioration
then often caused by molds. Enzymatic hydrolysis of fat has been observed at aw ≥ 0.1, be
it extremely slow. Accordingly, whole milk powder must be free of lipase. Milk lipase always
be inactivated by the intense pasteurization of the milk as applied in the manufacture of
whole milk powder. This is by no means ensured, however, for bacterial lipases. Hence, not
too many lipase-forming bacteria should occur in the milk. Proteolysis in milk powder
appears highly improbable and it has never been reported.
Of course, enzymic deterioration of liquid products made from the milk powder can occur if
enzymes are present before the drying, since drying usually does not cause substantial
inactivation of enzymes.
Caking: When milk powder or whey powder absorb water from the air is that lumps are
formed, eventually, the whole mass of powder turns into a solid mass (cake). Crystallization
of lactose is responsible, as it causes the powder particles, largely consisting of lactose, to
grow together. Since water is needed for crystallization of α – lactose, caking does not
occur at low aw, say, below 0.4. At a higher temperature crystallization can occur for more

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readily, aw being higher, moreover, the viscosity of the highly concentrated lactose solution
(essentially the continuous phase of the powder particles) is lower, causing nucleation,
hence crystallization, to be faster.
The susceptibility to caking, essentially high in whey powder, is considerably reduced if
most of the lactose is crystallized before the drying (in the concentrate). Such pre-
crystallized powder is usually called “nonhygroscopic,” which may be a misnomer because
the powder concerned does not attract less water (this is determined by it’s aw in relation to
that of the air), but the effects differ.
Maillard reactions increase considerably with water content and with temperature. They
lead to browning and to an off-flavor. The gluey flavor that always develops during storage
of dry milk products with a too high water content is usually ascribed to Maillard reactions,
the main component appears to be Ο-aminoacetophenone. If extensive Maillard reactions
occur, this is always accompanied by insolubilization of the protein. Accordingly, the
insolubility index increases if milk powder is stored for a long at a high water content and
temperature; at a normal water content the ADMI ( ml sediment per 50 ml reconstituted
milk; dissolving at 24oC, intensive stirring; centrifuging) number may increase to 0.5 in 3
years.
Autoxidation of the fat and the ensuring tallowy off-flavor pose a difficult problem when
storing whole milk powder. The rate of autoxidation strongly increases with decreasing aw;
however, to prevent other types of deterioration (esp. Maillard reactions) aw should as low
as possible. The effective Q10 of the autoxidation reaction in milk powder is relatively low
(about 1.5) since a higher temperature also causes higher aw.
A toffee flavor can develop during storage of whole milk powder with a high water content at high
temperature, due to the formation of δ-decalactone and related components in the fat.
Loss of nutritive value during storage primarily concerns loss of available lysine due to
Maillard reactions. Storage at 20oC at normal water content does not cause an appreciable
loss; at 30oC a loss of 12 % after storing for 3 years has been reported. Maillard reactions
cause a decrease in protein digestibility and formation of weak mutagens.
Extensive autoxidation results in formation of reaction products between hydro peroxides
and amino acid residues (this partly gives methionine sulfoxide), and between carbonyl
compounds and ε-amino groups; this may cause the biological value of the protein to

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decrease slightly. Of greater concerns is the loss of vitamin A in vitamin fortified skim milk
powder due to its oxidation. This especially occurs if the vitamin preparation is dissolved in
oil and then emulsified into the skim milk before atomization. Usually, dry added
preparation are stabler. It is, however, very difficult to homogeneously distribute a minute
amount of a powder throughout a bulk mass.

13.9 Other types of milk powder

Roller dried milk looks completely different from spray powder in the microscope. It consists
of fair-sized flakes. Due to intense heat treatment during the drying it has a brownish color,
a strong cooked flavor, and the availability of lysine has been considerably reduced by 20
% to 50 %.
Freeze dried milk consists of coarse, irregularly shaped and very voluminous powder
particles, which dissolve readily and completely. However, the fat globules show
considerable coalescence, unless intense homogenization has been applied. In most
cases, damage due to heat treatment is minimal.
13.10 Reconstituted products
Milk powder can be used to make a variety of liquid milk products. Some common types are
the following:
Reconstituted milk is simply made by dissolving whole milk powder in water to obtain a
liquid that is similar in composition to whole milk. Likewise, reconstituted skim milk can be
made.
Recombined milk is made by dissolving skim milk powder in water, generally at 40-50oC,
then adding liquid milk fat (preferably “anhydrous milk fat” of good quality, making a coarse
emulsion by vigorous stirring or with a static mixer, and then homogenizing the liquid. The
product is similar to homogenized whole milk, except that it lacks most of the material of the
natural fat globule membrane, such as phospholipids.
Filled milk is like recombined milk, except that instead of a vegetable oil is used to provide
the desired fat content.
Toned milk is a mixture of buffaloes’ milk and reconstituted skim milk. The high fat content
of buffaloes milk (e.g., 7.5 %) is there by toned down.

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Chapter 14

Butter production

14.1 Description

Butter is generally made from cream by means of churning and working. It contains
somewhat over 80 % fat which is partly crystallized. The churning proceeds most easily
with sour cream, at a temperature of about 15-20oC. Therefore, butter typically is a product
originating from regions of temperate climate.
The most important specific requirements for the product and its manufacture.
a. Flavor: Off-flavors of the fat are to be avoided, especially those caused by lipolysis, but
also those due to volatile contaminants. The latter mostly dissolve readily in fat;
notorious examples are off-flavors caused by feeds like silage and Allium species. If the
cream is heated too intensely the butter gets a cooked flavor. Moreover, much attention
has to be paid to the souring. The formation of so called aroma by heterofermentative
lactic acid bacteria is essential. The most important aroma substance is diacetyl, not
only is its formation important but also its persistence, since some starter bacteria can
reduce the diacetyl again. The homofermentative Lactococcus lactis ssp. lactis biovar.
diacetylactis also form diacetyl, but some strains produce so much acetaldehyde that
the butter develops a yoghurt flavor.
b. Shelf life: Spoilage by microorganisms may cause several off-flavors (putrid, volatile
acid, yeasty, cheesy, rancid); in culture cream butter it usually involves molds and
yeasts, the pH of the moisture being too low (~ 4.6) for bacterial growth. Lipolysis
causes a soapy-rancid flavor; no lipases formed by psychrotrophs should be present in
the milk. Furthermore, autoxidation of the fat can also occur, especially at prolonged
storage, even at a low temperature (~20oC), leading to a fatty or even a fishy flavor.
c. Consistency: Butter derives its firmness largely from fat crystals that are aggregated into
a network. Butter should be sufficiently firm to retain its shape, likewise, oiling off (i.e.,
separation of liquid fat) should not occur. On the other hand, the butter should be
sufficiently soft as to be spreadable on bread. This causes great problems because the
firmness ad the spreadability closely depend on the composition of the fat and on the
temperature.
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 d. Color and homogeneity are mostly easy to get right.
e. Yield: Losses of fat occur at skimming (in the skim milk) and at churning (in the butter
milk). If the water content is below the legal limit (e.g., 16 %), this also means a loss of
yield.
f. The product buttermilk is sometimes desirable, but it is often undesirable because of
insufficient demand. Sour cream buttermilk is only applicable as a beverage (or is used
for cattle feeding purposes), but it will keep poorly due to rapid development of an
oxidized flavor. Sweet cream buttermilk can more readily be incorporated in certain
products.
14.2 Definition

Butter may be defined as a fat concentrate which is obtained by churning cream, gathering
the fat into a compact mass and then working it .
According to the PFA Rules (1976), table (creamery) butter is the product obtained from
cow or buffalo milk or a combination thereof, with or without the addition of common salt
and annatto or carotene as coloring matter. It should be free from other animal fats, wax
and mineral oils, vegetable oils and fats. No preservatives except common salt and no
coloring matter except annatto or carotene may be added. It must contain not less than 80
% by weight of milk fat, not greater than 1.5 % by weight of curd and not greater than 3 %
by weight of common salt. Diacetyl may be added as a flavoring agent but, if so used, the
total diacetyl content should be not greater than 4 ppm . Calcium hydroxide, sodium
bicarbonate, sodium carbonate, sodium polyphosphates may be added, but must not
exceed the weight of butter as a whole by more than 0.2 % .

14.3 Description of types

Sweet cream butter: It is made from sweet cream, no starter has been added. Cream is
generally pasteurized for its production.
Ripened cream butter: Starter has been added to the cream before production.
Creamery butter: It is a butter made from cream produced on a number of farms.
Sweet butter: Butter to which salt is not added.
Salted butter: Butter to which salt is added.
Whey butter: Butter made from cream separated from whey which results in the
manufacture of cheese made from whole milk.
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Farm or Dairy butter: Butter made on the farm from a single herd, generally from
unpasteurized cream.
Renovated or Process butter: Butter which is melted, refined and renovated.
14.4 Composition of butter
Constituents %
Butter fat 80.2
Moisture 16.3
Salt 2.5
Curd 1.0

Butter is very high in fat and fat soluble vitamins A,D,E, & K.

14.5 Manufacturing scheme

Milk

Skimming 50oC
Skim milk
Cream 35 % fat
Pasteurizing at 90oC
for30 min Pasteurizing at 85oC for 15 s

Starter, at 20oC 5% Inoculating

for 20 h
0.5 % Ripening at 14oC for 20 h
Culture
Churning 14oC

Buttermilk Packing
Separating

Butter grains Homogenizing

Water 12oC Washing Cold storing 10oC, 7 d

Working, Butter
Water, salt Standardizing
Packing
Buttermilk
Fig.14.1 Flow diagram for the manufacture of butter from ripened (sour) cream.

The skimming is mainly done for economical reasons: reduction of fat loss (e.g., the fat
content of buttermilk is 0.4 %, that of skim milk is 0.05 %, this means that removal of 1 kg of
skim milk from the liquid to be churned will result in an additional yield of about 4 kg butter);
reduction of the size of the machinery (especially the churn); reduction of the volume of
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buttermilk. Hence, a high fat content of the cream (e.g., 40 %) has advantages, also
because of counteracts development of off-flavors. If a continuous butter making is used,
the fat content of the cream is often taken even higher, i.e., up to 50 %.
In some countries the farmers skim the milk and deliver the cream to the dairy. Such cream
is often in a poor condition, i.e., more or less turned sour and having off-flavors. Generally,
such a cream to be neutralized, e.g., with NaOH or NaHCO3, to allow its pasteurization.
The cream may also be washed, i.e., diluted with water and reseparated, to remove
undesirable flavors, however, thus is not very effective.
Pasteurization serves to kill microorganisms, to inactivate enzymes, to make the cream a
better substrate for the starter bacteria, and to render the butter more resistant to oxidative
deterioration. Overly intense heating causes a cooked or gassy flavor. Sometimes the
cream is pasteurized in a vacreator, which implies that the hot cream is put under vacuum
to cool, due to which some off-flavors are (partly) removed.
The starter should be “aromatic,” i.e., produce aroma substances and retain these. To that
end it can be necessary to aerate the starter for a while at the end of the incubation period,
e.g., by stirring. The starter bacteria should be able to grow fairly fast at low temperature.
The purpose of ripening is to sour the cream and to crystallize the fat. Without solid fat,
churning is impossible, and too little solid fat goes along with excessive fat loss in the
buttermilk. The way of cooling (temperature sequence) affects the butter consistency.
The churning is in most cases achieved by beating in of air. It can be done in a churn,
mostly consisting of a large vessel (tub, cylinder, cube, double ended cone) with so called
dashboards, which is partly (at most half) filled with cream, and which is rotated at several
revolutions per minute(rpm). The churning is then takes, say, 20 min. There are also churns
with a rotary agitator (e.g., 20 min). The latter principle is also applied in the frequently used
continuous butter making machine according to Fritz. Here the paddle turns very quickly
(500-3000 rpm) and the cream stays in it for less than 1 min. For that purpose high fat
cream has to be used, i.e., about 50% fat. These machines can have a very great capacity.
The churning should proceed rapidly and completely (low fat content in buttermilk) and the
formed butter grains should have the correct firmness to allow for efficient working. The
size of the butter grains can be varied by continuing the churning for various lengths of time

247
after grains have formed. Very fine butter grains (say, 1 mm) are hard to separate from the
buttermilk, especially in continuous machines.
If the butter grains are not too large their firmness can to some extent be affected by
washing, i.e., via the wash water temperature. The washing consists of mixing the butter
grains with water, after which this again is drained off. It reduces the dry matter content of
the butter moisture. Formerly, washing was applied to improve the keeping quality of the
butter, but nowadays, it is only done to control the temperature, if needed.
The working (kneading) is meant to render the butter grains into a continuous mass, to
finely disperse the moisture in the butter, to regulate the water content, and, if desired, to
incorporate salt. Washing consists of deforming the butter. This can, for instance, be
achieved by squeezing the butter through rollers, by allowing it to fall from a height (in the
modern churn- and –workers), or by squeezing the butter through perforated plates (in the
continuous machines). During the working the water content is regularly checked and, if
need be, additional water is added to arrive at the accepted standard value.
The butter can now be immediately packed, e.g., in retail package. Often one wants the
butter after the working to be soft enough to be pumped from the churn-and-worker by
means of a suitable positive pump. Sometimes, the butter is allowed to set or it is for
another reason kept for some time before packing. It is then too firm to pass through the
packing machine, and it must be passed through a butter homogenizer to soften it, this may
also prevent the moisture dispersion from becoming too coarse during packing.
14.6 The churning process
Churning means agitation of cream at a suitable temperature until the fat globules are adhere or
coalesce each other forming a compact solid mass and release the serum occurs. There is a
complete separation of fat and serum occurs.

Air bubble

Liquid fat
1 2 3

Fig. 14.2 Shows the iteractions between fat globules and air bubbles during
churning. The fat is liquid and it is disrupted by beating in of air (top).
The fat globules contain solid fat and form fat clumps (bottom).

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The object of churning cream is to produce butter. In milk/cream, the fat exists in the form
of an emulsion, i.e. a continuous phase. This emulsion is fairly stable. As long as it remains
intact, there is no formation of butter.
A greater concentration of fat globules in cream promotes a more profuse & rapid
coalescence and aggregation than in milk. Above melting point of (31-36oC), agitation of
cream results in subdivision of fat globules. At lower temperatures (maximum effect at 7-
8oC), agitation causes coalescence of fat globules.
Factors influencing churnability of cream and body of butter
Chemical composition of fat : Milk fat is a mixture of numerous fats (glycerides) of widely
varying melting points and solidifying points.
Low melting point fats : e.g. tri-butyrin, olein etc – soft fats.
High melting fats : e.g. sterin, palmitin etc. – hard fats.
An increase in the proportion of soft fats shortens the churning period, diminishes the
firmness of butter and increases the fat losses in buttermilk and vice versa.
Size of fat globules: Smaller the size of fat globules, longer the churning period and greater
the fat loss in butter milk, and vice versa.
Viscosity of cream: The greater the viscosity of cream, the greater the churning period, and
vice versa.
Temperature of the cream at churning: A higher churning temperature causes a shorter
churning time, higher fat loss and a weak body in butter. A lower churning temperature
prolongs the churning period. The optimum churning temperature ranges from 9-11oC for
about 30-60 min, a butter of satisfactory firmness and exhaustive churning.
Fat % of cream: The higher the fat % in cream the lower the churning period and vice
versa.
Acidity of cream: Cream churns more rapidly and exhaustively than sweet cream.
Load of churn: The optimum load of cream which the churn can take more efficiently is to
be one-half and one third of its total capacity . Overloading in churn prolongs the churning
period, while under loading reduces the normal capacity of the churn.
Nature of the agitation: It influenced by the size, type and r.p.m. of the churn, and affects
the churning period.
Speed of the churn: More speed shortens the churning period.

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Details of production
The butter of good quality can only be produced by using good raw material. So, the milk
and cream received for butter production should be tested and graded.
Cream acidity should be reduced to 0.06 to 0.08 % before churning. The acidity of cream
should be 0.25 to 0.30 % during churning. Therefore neutralization of cream should be
needed.
The objectives of neutralization are:
• To avoid excessive fat loss in butter milk (When pasteurizing sour cream, the casein
curdles, thereby entrapping fat globules, as the bulk of the curd goes to butter milk, this
causes high fat loss.)
• To guard against the production of an undesirable off-flavors in cream.
• To improve the keeping quality of butter made from high acid cream.
Salted acid butter develops a fishy flavor during commercial storage at –23 to –29oC.
The neutralizers are, Calcium hydroxide (Ca(OH)2, Magnesium hydroxide, Sodium
hydroxide, Sodium carbonate, Sodium bicarbonate, Sodium sesquicarbonate.
Standardization of cream reduced the fat loss in butter milk during churning.
Pasteurization of cream is necessary to kill pathogenic organisms and enzymes. The time
and temperature used for pasteurization of cream are:
71oC for 20 min – Promptly cooled- holder pasteurization.
95-100oC for 15-16 seconds –high temperature short time pasteurization.
Vacuum or Vacretion pasteurization.
For the ripening of butter starter culture containing lactic acid producers for examples
Streptococcus lactis, Streptococcus cremoris, together with diacetyl producers e.g.
Streptococcus diacetilactis, Leuconostoc dextranium at the rate of 0.5 – 2.0 % and
incubated at 21oC for 15-16 h.
Objectives of ripening
To produce butter with pleasing, pronounced characteristics flavor and aroma.
To obtain exhaustive churning i.e. a low fat loss in butter milk.
The flavor is mainly due to the effect of diacetyl, biacetyl and smaller extent of acetic
and propionic acids. In a sweet cream there is no diacetyl found. The ripened cream

250
contains diacetyl in an average of 2.5 ppm and rarely over 4 ppm. The diacetyl is produced
from its mother substance acetyl methyl carbinol.
Ageing of cream is necessary for crystallization of fat in the fat globules. Optimum
temperature for cooling and ageing cream depends on :
Composition of fat, size of fat globules, fat % in cream, period of ageing , temperature of
churning & acidity of cream.
The optimum temperature of cooling and ageing and churning provides a degree of
solidification of fat in cream that may yield a normal churning period, reasonably exhaustive
churning, satisfactory washing and a satisfactory firmness in the body of butter.
Overrun in butter
B−F
% Over run = × 100
F
where, B = Kg. butter made.
F = Kg. fat in churn
14.7 Structure and properties
14.7.1 Microstructure

Striking (and an important difference with, for example, margarine) is the presence of many,
partly intact fat globules. Their number depends on the way of manufacture and it
decreases, for instance, during intense working. Note that most crystals in the fat globules
are tangentially arranged.
The continuous phase is liquid fat. Sometimes a continuous aqueous phase persists,
especially in insufficiently worked butter. This aqueous phase partly passes through the
surface layers of the fat globules. The fat that displacement of water through butter can
occur generally has another cause: approximately 0.2 % (v/v) water can dissolve in liquid
fat. This implies that water can diffuse through the continuous oil phase.
Moisture droplets are not always equal in composition. Differences are found due to water
addition, washing, and the working in of starter, salt, or brine. Differences in osmotic
pressure then cause a slow water transport to the most concentrated droplets. Hence,
moisture droplets in the vicinity of a salt crystal mostly disappear, and the salt crystals leads
to a large droplet; accordingly, the butter turns ‘wet’.

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The number and size of the fat crystals greatly depend on the temperature and on the
temperature history. A considerable part of the crystalline fat may be inside the fat globules
because during churning liquid fat is extruded from the globules, mainly by spreading over
the air bubbles. But there are also crystals outside the globules and these aggregate to a
continuous network and may grow together to form a solid structure, which is mainly
responsible for the butter firmness. The crystals inside the globules do not precipitate in this
network and, therefore, they hardly make the butter firmer. Because of this, butter generally
contains more solid fat than margarine, when both products are equally firm; this results, in
turn, in the butter feeling cooler in the mouth (due to the greater heat of fusion).
The crystals outside the fat globules thus make up a continuous network in which part of
the water droplets (often with crystals attached to their surface) and damaged fat globules
may precipitate. This network retains the liquid fat as a sponge. Note that butter thus has at
least two continuous phases (oil and fat crystals) and possibly a third one (aqueous). If the
temperature increases many crystals melt and the network becomes less dense and
coarser. Because of this, destabilization can eventually occur, i.e., the butter separates oil.
Oiling-off occurs more readily (at equal solid fat content) if the crystals are coarser.
Air cells always occur in butter, unless the working is done in vacuum (which is possible in
some continuous machines). Moreover, butter contains up to about 4 % (v/v) of dissolved
air.
14.7.2 Consistency
Butter should be sufficiently firm against sagging. It should be readily spreadable and
thereby not too “long” (extensible) or too “short” (crumbly, flaky). It should be easily
deformable in the mouth without being greasy, should melt rather quickly, and should
thereby feel cool. A full rheological description of butter would be very complicated and
cannot yet be given. Generally, we use the term firmness (hardness) to denote the main
property. Firmness is a poorly defined quantity and depends on the type of deformation
applied, on the time scale of the experiment etc. In most cases, spread ability correlates
rather well with it.
The main factors affecting the firmness are
a. The fat crystals aggregate into a network with meshes of the order of 1 to several µm in
size. This gives the fat certain elasticity at very small deformation. At greater

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 deformation, bonds have to be broken; this involves the most important contribution to
the firmness of a freshly crystallized fat.
b. As soon as aggregated crystals grow further they sinter i.e., grow locally together to
form a solid structure, thereby very much increasing the firmness. At (large) deformation
the solid structure has to be broken.
c. The amount of solid fat: A very strong influence of the temperature.
d. Size and shape of the crystals: Milk fat with 20 % crystals in the form of large spherulites
is still liquid, but with 10 % of the fat in small crystals it behaves like a solid of
considerable firmness.
e. The inhomogeniety of the network of crystals has a very great effect because the fat will
start to flow where the bonds are weakest. In a quantitative sense, however, practically
nothing is known about this aspect.
As soon as are starts to work a fat its firmness decreases sharply; this is called work
softening. On keeping, the fat sets again, but not up to only after a few hours; a solid
structure also forms again, but that takes a longer time. Hence, in a sense, the fat is
thixotrophic.
The fat globules in butter contain a considerable part of the crystalline fat. These crystals
therefore cannot or hardly particulate in forming a continuous network or a solid structure.
In other words, butter is much softer than butter fat (or margarine) with the same amount of
solid fat. Butter is also a water-in-oil emulsion, but the water droplets have a small effect on
the consistency, unless their volume fraction is very great, as in low fat spreads.
17 days
Firmness at 16oC

4 days

1 days

fresh

0 5 10 15
o
Temperature ( C)

Fig. 14.3 Effect of temperature and time of storage on the firmness of butter.
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14.7.3 Cold storage defects
To keep the butter for a long time, it should be stored at a temperature, say, -20oC, if the
butter has been well made, and if the original milk did not contain too many bacteria with
thermo resistant lipases, it can keep for a very long time in cold storage. It now deteriorates
by autoxidation of the fat, leading to flavor defects after 1 month to 2 years.
The keeping quality in cold storage greatly depends on the method of manufacture.
Processing variables having an effect are as follows.
a. Contamination with even minute quantities of copper should be strictly prevented.
b. By cooling the milk for some time before its use (e.g., for at least 2 h at 5oC) a part of
the copper on the fat globules moves to the plasma; this may restrict the autoxidation.
Moreover, the cooling causes a migration of protein to the plasma, and this is precisely
the protein that liberates H2S during the heat treatment. Hence, in this way, a cooked
flavor after heat treatment can be limited.
c. Heating of milk or cream causes migration of copper from the plasma to the fat
globules. Pasteurization of the milk should therefore be avoided because more copper
is available to migrate in milk than in cream.
d. Due to souring of the cream (or the milk) a considerable part (30 % to 40 %) of the
added copper (i.e., copper entered by contamination) moves to the fat globules.
Because of this, butter from sour cream is much more affected by autoxidation than that
from sweet cream. But sweet cream butter is not aromatic.
e. It is important to adjust the fat content of the cream to a high level because this causes
a lower copper content in the butter as discussed in c and d .
f. Heating of the cream prevents the migration during souring mentioned in d. In all
probability, copper becomes bound to low-molar mass sulfides, especially H2S, formed
by the heat treatment. This causes a strong reduction of the autoxidation in sour cream
butter. Therefore, it would be desirable to pasteurize the cream very intensely, but then
much H2S is formed, yielding butter with a gassy or cooked flavor. Although this flavor
defect decreases slightly during storage, it is objectionable. The pasteurizing conditions
should thus be optimized (not too high, not too low); of course, the smaller the spread in
holding time, the better.

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 g. Adding salt to sour cream butter considerably accelerates the autoxidation. In sweet-
cream butter a high salt content has a oxidation diminishing effect.
h. The lower the storage temperature, the higher the keeping quality.
14.9 Culture butter from sweet cream

It is often a problem to dispose of sour cream buttermilk because it has very short shelf-life
and the demand as a beverage often is small. Moreover, it cannot be pasteurized. Sweet
cream buttermilk can be processed much more easily. On the other hand, several markets
prefer an aromatic butter, which has to contain acid (lactic acid) and aroma substances
(mainly diacetyl). It has indeed been tired to churn sweet cream and to add starter to the
butter granules afterward to work it into the butter, but the result is disappointing, i.e., the
flavor remains almost equal to that of sweet butter. This is not surprising because the
souring and the diacetyl production in the butter can hardly occur if the moisture has been
well dispersed, the pH also remains high.
NIZO has developed an alternative manufacturing process. Sweet butter grains are worked
together with a very aromatic starter and with a concentrated starter permeate, essentially a
lactic acid solution. Churning and working can proceed in a churn-and-worker or in a
continuous butter making machine. The initial water content (i.e., after the first working)
should be low so as not to exceed the 16 % limit afterward. In preparing the starter
permeate a partly delactosed whey is soured by means of Lactobacillus helveticus; then the
liquid is purified by ultrafiltration, and the permeate concentrated by evaporation. The lactic
acid content of the permeate then amounts to about 16 % (standard acidity degree
1800oN).
At first a normal aromatic starter was worked in, apart from the starter permeate. This had
the disadvantage that the aroma must develop after the butter making and that the aroma
formation depends too much on the conditions; hardly any aroma is produced if the butter is
immediately transferred to cold storage, whereas at room temperature the aroma the aroma
production can be excessive if the butter has a not very fine moisture dispersion. Therefore,
nowadays a very aromatic starter grown in evaporated skim milk (starter I); it is mixed with
the starter permeate and subsequently aerated. In this way sufficient diacetyl is can occur.
Starter II contributes some other flavor components and can cause a continued formation of
diacetyl; the concentration of diacetyl in the butter eventually is 1.5-2.5 mg/kg.
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The moisture droplets will occur in the butter that more or less differ in composition: sweet
buttermilk, starter, starter permeate with starter. Some compounds can migrate, e.g., water
and lactic acid (both slowly), and especially diacetyl, which is fairly fat-soluble (the partition
coefficient water/oil roughly equals unity)
14.10 High fat products
Besides butter there are other high fat products in which fat is the continuous phase. Such
products are made for various reasons.
Traditionally, butter was “melted down” to increase its keeping quality, i.e., after heating the
butter, the formed butter oil was separated. The rendered butter thus obtained had a long
shelf life. Nowadays, this product is designated anhydrous milk fat; it may be used as such
in the kitchen because, contrary to butter, it allows heating at a high temperature. In the
country like Nepal where the temperature may be too high for butter making, ghee is made
from buffalo’s milk. This milk has fairly large-sized fat globules, which at a somewhat higher
temperature cream sufficiently fast (without agglutinating) to yield a cream layer. The cream
obtained is subsequently heated over an open fire until the water has boiled off.
Precipitated dry matter substances are removed by decanting.
The separated fat can be modified and/or fractioned in various ways. The main aim may be
to change the crystallization behavior of the fat. The modified fat can be used in
recombined butter, in chocolate (as a partial substitute for cocoa butter), in bakery
products(such processes as kneading of dough and paste, and beating in of air require a
constant melting behavior and often a high final melting point), in recombined cheese
(restricting oiling-off), or in instant milk powder (liquid fat yields better instant properties).
From milk fat and skim milk recombined butter can be made. The purpose may be to
enhance the value of poor-quality cream or to obtain a product with other properties, e.g.,
firmness (spreadibility), a higher water content, or a higher content of polyun-saturated fatty
acids. For example, several kinds of spread are made, especially low-fat-spreads. To
achieve this, blends of milk fat and vegetable oil can be used. Alternatively, a vegetable oil
(e.g., 25 % to 30 % refined soybean oil) can be worked into butter to obtain a spreadable
mixture, such as the Swedish Bregott. The products and processes are as follows:

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14.10.1 Anhydrous milk fat

The most important general requirement is that the fat be very pure and stable to
autoxidation. To secure this, good quality fresh milk should be used. Contamination by
traces of copper is highly detrimental. The water content should not exceed 0.1 %, because
otherwise moisture droplets may form at low temperature. If the water content is higher (up
to 0.4 %), the product is usually designed “butter oil”.
Manufacturing process
One starts from butter. Alternatively, one can make high fat cream (by centrifuging twice)
and accomplish a phase conversion in it: If a very concentrated o/w emulsion is
destabilized, a w/o emulsion is usually formed. To achieve this, fat cream can be passed
through an agitator, a special pump, or even a homogenizer, often, the phase inversion
occurs easier if the cream first is subjected to “washing”, i.e., diluting it with water and re-
separating. If cream with 82 % fat is passed through a scraped-surface heat exchanger
while being cooled sufficiently for fat crystallization to occur, then butter is formed.
Examples are the Alpha process and the Meleshin process. Sweet cream Alpha butter can
be very durable.
In the manufacture of anhydrous milk fat, the cream destabilization usually occurs at high
temperature and butter oil containing plasma droplets then is obtained; often, a more or
less complete separation into two layers immediately sets in. This can also be achieved by
melting of butter. Subsequently, a separation has to take place by decanting or, in common
practice, by centrifuging, for this purpose a suitable separator is needed. The fat obtained in
this way is very pure; if the temperature during the melting and separating has not been too
high, it is almost free from polar lipids.
Another method of working is based on evaporation of water by heat treatment; from butter,
from fat cream, or from an intermediate product, e.g., butter grains; alternatively, washed
cream can be used. When the evaporation is done in an open vat, i.e., at atmospheric
pressure, the temperature becomes somewhat high, up to 120oC; the product obtained is
ghee. Furthermore, cream or butter can be “dried” in a vacuum evaporator or in a spray
drier. In all cases, the non fat solids are left dispersed in the fat. These can be removed by
decanting, filtering, or centrifugation. Due to the higher temperature and the strong lowering
of the water activity, the polar lipids, especially the phospholipids have an antoxidant
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activity, which is beneficial for the keeping quality. Genuine ghee, especially ghee made
from sour cream, contains numerous flavor compounds originating from phospholipids,
proteins, and triglycerides (e.g., methyl ketones from keto fatty acids).
Milk fat made by one of the above processes usually contains about 0.4 % water. On
cooling, droplets are formed (the solubility of water in milk fat is 0.1, 0.2, and 0.4 % at 10,
40, and 90oC, respectively) and the product can spoil rapidly. Therefore, vacuum drying is
generally applied e.g., at 40oC and 2 KPa (= 0.02 bar). It causes a decrease of the water
content to below 0.1 %. Also the oxygen content decreases significantly. A product made in
this way may keep for some years if made from milk without any beginning autoxidation, if
stored in isolation from air and light, and if copper contamination has been rigorously
prevented.
14.10.2 Modification of milk fat

The most widely used modification is fractionation by means of crystallization. After

solidification of milk fat at a certain temperature in such a way as to form fairly large
crystals, the fat can mechanically be separated into a solid and a liquid portion. The
purpose is to obtain fractions with different melting behavior. The composition is also
altered in another respect since fat-soluble components, like carotenoids, vitamins, and
flavors, become concentrated in the liquid fraction. Accordingly, it remains to be seen as to
whether a “solid” fraction may still be considered milk fat.
The success of a single fractionation is less than expected. The separation is incomplete
because the network of fairly small crystals readily retains liquid fat. To form large crystals
by cooling the fat very slowly. This may cause formation of spherulites. Spherulites are
sphere-shaped crystals, but they are made up of a great number of ramified radial needles,
between which, again, liquid fat is held. A far better fractionation can be achieved by
crystallization of the fat from acetone, but this is an expensive method; moreover, use of
the product obtained in foods may not be allowed by the public health authorities.
Fractionation of milk fat generally obtained more spreadable butter. It has also been tried to
make recombined butter with a firmness that is not greatly temperature dependent.
Milk fat can be modified chemically, but the products can no longer be called milk fat.
Hydrogenation decreases the number of double bonds and thereby increases the high
melting proportion of a fat; this is why the process is often called hardening. It also causes
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several other changes, such as displacement of remaining double bonds and cis- trans
isomerization. Hydrogenation can be applied to make cocoa butter replacers from milk fat.
Intersterification causes the distribution of the fatty acid residues over the positions of the
triglyceride molecules to become increasingly random. It can be achieved by heating the fat
in the presence of catalyst, like sodium methoxide. (A very high temperature –say, 150oC- it
also occurs without a catalyst being present.) After intersterification, the melting range of
milk fat is sifted to higher temperature. Similar effects occur on cis-trans isomerization.
None of these chemical modifications is applied to milk fat on a commercial scale.
14.10.3 Recombined butter

Milk fat and skim milk can be recombined to yield a butter-like product. Manufacturing
process and physical structure of the product then are practically identical to those of
margarine. The difference is in the composition.
Recombined butter is firmer than natural butter of the same fat composition; the texture is
also slightly different. This is largely due to the product not containing fat globules, so that
all fat crystals can participate in networks and solid structures. Accordingly, the firmness
has to be adjusted in a different way, mainly via the fat composition. The final melting point
of the fat should be below body temperature because the butter should fully melt in the
mouth. If temperature fluctuates widely a course texture readily develops, due to the fat
crystals becoming very large.
Manufacture
In the production of margarine a good deal precedes in the way of treatment of the fat,
including degumming, alkali refining, bleaching, deodorizing, and partial hydrogenation.
The treatment of the aqueous phase mainly serves to attain a good a good flavor and to
improve the keeping quality (preventing spoilage by microorganisms, lipase, etc.). Usually,
in the production of recombined butter, hardly any additives are used, whereas margarine
may contain an antioxidant with or without synergist, coloring matter, added vitamins A and
D, fat soluble flavoring agents, and an emulsifier.

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 Skim milk Milk fat

o Additives
Culture Pasteurizing Melting at 50 C

Souring
Starter
Emulsifier
Salt Emulsifying

Rapid cooling 10oC

Crystallizing for 10 min

Working
Packaging
material Packing

Fig.14.3 The manufacture of recombined butter (or margarine)

Emulsification serves to obtain a fairly homogeneous mixture of constant composition. The
water-in-oil (w/o) emulsion is not stable and the liquid must be agitated at all times, until fat
crystals have been formed. The crystals stabilize the moisture droplets by adsorbing onto
the interface. The emulsifiers (in margarine, e.g., monoglycerides seem to prevent the fat
crystals from flocculating so strongly into a network that they can insufficiently adsorb onto
the moisture droplets. Moreover, the emulsifier plays a twofold role during the heating of the
product in the frying pan. Due to melting of the fat the moisture droplets become unstable,
and if they now quickly flow together into large droplets these can start to splash very
annoyingly when reaching the boiling point. The emulsifier slows down such coalescence.
Furthermore, lecithin contributes to the typical aroma during frying. Therefore, in the case of
recombined butter it has its advantages to displace a part of the skim milk by sweet cream
buttermilk; in this way the composition also is closer to that of natural butter.
The cooling is usually performed in a scraped-surface heat exchanger; otherwise the heat
transfer would proceed far too slowly, causing the formation of overly large crystals. Due to
the scraping and stirring the moisture droplets, which during emulsification may still be up
to 1 mm in size, are divided in to much smaller ones. A further reduction occurs by the
working that consists of forcing the mass through a small opening (valve) or perforated
plate.
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The working also serves to destroy solid structures should then indeed already have been
formed, which implies that by far most of the crystallization should have taken place. This is
achieved by the passage through a “ crystallization tube” before the working. However, milk
fat crystallizes very slowly and, accordingly, it is mostly impossible to achieve a fairly
complete crystallization in one processing step (as is common in margarine manufacture).
Consequently, the butter can still set considerably after manufacture. During crystallization
one often has to cool again. If, for instance, 20 % of the fat crystallizes, then the heat of
crystallization suffices to increase the blend temperature by 8oC.
14.10.5 Butter products with a low fat contents

Low-fat butter products have long been in existence (from the early 1940s), but they have
received renewed interest following low-fat margarine. These “spreads” have a water
content of, say, 40 %. There are also spreads that partly consist of fat extraneous to milk fat
and that are held to be better for health than butter by some people.
Production of low-fat butter cannot be achieved by churning because the formation of a
product with a discontinuous aqueous phase fails. Most surface active substances in cream
are water soluble, and this largely prevents the formation of a w/o emulsion. The same
manufacturing process given above can be applied; a suitable (i.e., oil-soluble) emulsifier
should be added. It may be a problem to make the moisture droplets sufficiently small and
to prevent them from coalescing in the product, especially at a somewhat higher
temperature. Therefore a gelling agent (e.g., gelatin) may be added. It causes the droplets
to become more or less solid, unable to coalesce. At the least, a thickening agent should be
added to the aqueous phase. This may be a protein mixture, like serum protein that has been
coagulated to yield aggregates of about 1µm in size. The agent improves flavor and mouth-feel of
the product, which tends to be similar to butter. Flavor substances as well as aroma forming starter
bacteria may be added. All the same, there is no common appreciation of the flavor of such
“butter”. One of the aspects involved may be that the product contains less crystalline fat than
butter does and therefore feels less cool in the mouth.
Spreadability is generally not a problem. However, if the butter product contains excess liquid fat it
may become too soft and at room temperature it may not retain its shape and show oiling-off. A
product with a low number of fat crystals might have large moisture droplets, which can easily
cause microbial spoilage. A preservative can then be added.

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Chapter 15
Ghee Manufacturing

Ghee may be defined as clarified butter fat prepared chiefly from cow or buffalo milk. Sheep
and goat milk can also be used rarely for the production of ghee. The unwanted solid
matter and impurities should be removed while purifying ghee.
According to the PFA Rules (1976), ghee is the pure clarified fat derived solely from milk or
nauni (cooking) or also called desi (in India) butter or from cream to which no coloring
matter is added.
15.1 Chemical composition

Parameters Quantity
Milk fat 99.5 %
Moisture > 0.5 %
Carotene 3.2-7.4 µg/g
Vitamin A 19-34 IU/g
Tocopherol 26-48 µg/g
FFA (As oleic acid) Max. 2.8
Charred casein, salts of Cu, Fe etc. Traces

15.2 Physico-chemical constants

Ghee, as in the case of other fats and oils, is characterized by certain physico-chemical
properties, Which have been found to be the basis for the fixation of physicochemical
constants for defining the quality of the product. These properties, however, show some
natural variations depending on such factors as: Method of manufacture, age condition of
the sample, species, breed, individuality, animal’s stage of lactation, the season of the year,
region of the country, feed of the animal etc.
The important analytical constants or standards of mixed ghee produced under standard
conditions are given below.

Melting* point of ghee 28-44oC (≈ 33oC)

Solidifying* point 28-15oC
Specific gravity 0.93-0.94
Refractive index 40-45 at 40oC
Reichert-Meissel (RM) value** Not less than 28
Polenske value*** Not more than 2
Saponification value 220
Iodine value 26-38
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 *
• Being a mixture of glycerides of short and long chain fatty acids, ghee does not have
sharp melting and/or solidifying points.
**
• Ghee from cotton seed feeding areas, Where the limit is 20.
***
• Cotton seed feeding areas, where the limit is 1.5.
15.3 Food and nutritive value
Ghee is the richest source of milk fat. It contains fat soluble vitamins A, D, E, and K.

15.4 Methods of manufacture

1. Deshi method.
2. Creamery method.
3. Creamery butter method.
4. Pre-stratification method.

1. Deshi(Nauni) method
The ghee is prepared on small scale in the household or in cottage scale industry only.
The fresh and clean nauni (in India called mukkhan) collected for some days is heated
and stirred on a metallic open pot/vessel with a low fire to drive out moisture. When all
the moisture has been removed, as it is judged by experience, further heating is stopped
and the pot/vessel removed from the fire. On cooling, when the residue has settled
down, the clear fat is decanted into suitable containers.
2. Creamery method
Milk reception

Cream separation

Cream of 50 % 70 % fat
L. cremoris, left for 10-12 hrs.
for ripening the cream

Boiling for 115-120oC

effervescence
Ghee
(Good flavor ghee produced)
Fig.15.1 Flow diagram for the manufacture of ghee by creamery method.

Cephalin acts as a antioxidant and preserves the ghee.

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Stages of fat heating process
1. Butter fat melts at 40-60oC.
2. Heating above 60oC a foam forms on the surface which increases on volume until a
temperature of 94oC is reached.
3. The liquid butter simmers at 94-96oC and the temperature remains constant till all the
water evaporates out.
4. By the time the last drop of water evaporates out it temperature remains constant till all
water evaporates out.
5. By the time the last drop of water evaporates out it temperature rises 96oC the liquid
becomes thicker and bubbles begins break with sufficient violence to cause some
spattering.
6. As the temperature approaches 105oC the curd begins to form the lumps and these
lumps are carried to the surface to form a scum.
7. As the temperature approaches 120oC the curd particles combined and sink to the
bottom of the pan. The scum disappears from the surface and the particles of white curd
can be seen near the bottom of pan; floating it in the clear yellow liquid. As the
temperature rises slightly above 120oC the larger bubbles are changed to smaller ones,
until foams of smaller bubbles suddenly rises up which indicates the end point of the
operation. As this stage steam should be closed the color of the curd particles at the end
point should be brownish.
Filtration of ghee
Ghee and ghee residue are separated through the filtration process. The ghee residue
contains lactose, whey protein and charred casein. In ghee residue 8-12 % ghee is lost.
Some of the ghee from ghee residue can be separated by putting it in boiled water. When
boiled water is put in ghee residue and agitates the ghee float on the top and it can be
separated out. This minimizes the loss of ghee in ghee residue.
If we take high % fat cream say 70 % instead of 50 %, there is less loss of fat in ghee
residues, because high % fat means less % of other residues.

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3. Creamery butter method
Receiving milk

Cream separation

White butter without salt added

Boiling for 115-120oC

Crude ghee

Filtration

Ghee Ghee residue

Fig.15.2 Flow diagram for the manufacture of ghee by creamery-butter method.

In a flush season, when plenty of milk produced then the creamery butter method is used to
prepared ghee making.
Butter-oil Butter oil
Heating at Centrifuge
Butter
100oC
SNF
Fig. 15.3 Simplified flow diagram of butter oil-manufacturing
Table 15.1 Differences between different methods of ghee making
Characteristics Deshi method Direct creamery method Creamery butter method
Adaptability Small scale industry Medium scale industry Large scale industry
% output of fat 85-88 % 93-95 % 96-98 %
Quality of ghee Fair Good Good
Flavor Excellent Good Good
Keeping quality Poor Excellent Good

4. Pre-stratification method

The lower or bottom portion which contain butter milk or skim should be separated out and
heated to 110-115oC. The keeping quality of ghee by this method is poor. It has produced
low flavor ghee so now a days not used by the most of the producers of India but in Nepal
Dairy development corporation using this method for ghee manucature.

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 Steam in Steam out
Scum
Cream heated at 80-
85oC to 30-40 min.
Fat

Skim or butter milk 60-70

% SNF and Water 80 %.

Fig.15.4 Shows the pre-stratification method using in ghee manufacture.

15.4.2 Cooling and granulation

Granulation of ghee is important parameter, which prefer by consumer. The high molecular
weight fatty acid such as stearic and palmitic acid contain in ghee may affect the
granulation of ghee. These are saturated fatty acids having high melting point. These
caused good crystallization of ghee. In this regard buffalo ghee contains more amount of
these fatty acid so crystallizes more effectively than cow ghee.
If the ghee from 60-100oC, rapidly cooled to room temperature small fine crystals are
formed. If the ghee crystallization is kept at 2-3oC, very good grain formation will results.
Good crystallization of ghee will takes place if the temperature reduces from 60-80oC to 32-
34oC. Cold storage of granulated ghee should be avoided which leads to pasty, waxy and
greasy consistency of ghee.
15.4.3 Packaging and storage of ghee
Since milk fat is susceptible to deterioration due to exposure to light, air and metals. Ghee
should be properly packed after production so as to retain its initial flavor and nutritive
value.
1. Selection of container
- nontoxic, non tainting character
- easily available
266
 - economical
- resistance to rough handling.
2. Filling and storage
- store at room temperature 30-32oC.
- filling at melted condition.
- Air space should not be more than 2 % of total volume of fill in a container.
- Storage temperature of ghee should be around 21oC.
3. Keeping quality: It depends on the-
- method of production.
- Sedimentation.
- Packaging and storage.
Shelf life of ghee is about 6-8 months at 21oC.
Keeping quality is affected by development of acidity and off-flavor etc.
Shelf life of ghee is affected by-
• temperature of storage.
• Initial moisture content of ghee.
• Initial acidity content of ghee (acid value-oleic –not greater than 2.5 %)
• Cupper and iron content in ghee i.e., responsible for oxidation of ghee.
• Heating at high temperature i.e., 120-125oC affect the shelf life, so centrifugation is
done to reduces.
• Methods of packaging.
Extending the shelf life of ghee:
- addition of BHA @ 0.02 %.
- Addition of phospholipids – solvent extraction method @ 0.1 % phospholipids
(cephalin) in ghee will extend the shelf life 2 times.
Accelerating shelf life testing
There is no reliable test for measuring the developed rancidity and correlating the same
with the keeping quality of ghee. Determination of Induction period (IP) for oxygen
absorption at an elevated temperature of 79oC for ghee has been observed to be helpful in
predicting its self life. An induction period of 20 h or more will corresponds 6 months storage

267
life or self life. The induction period is the period measured in hours, after which the rate of
increase in the peroxide value of a ghee sample shoots up.
15.4.3 Renovation of ghee
1. Reheating inferior ghee with curd, betel, curry leaves- heat for 10-15 min. Then filter the ghee.
2. Adding a yellow substances such as saffron, annatto color, Rattanjot, turmeric juice etc, to
make it appear that it is cow ghee. The ghee mass is boiled and filtrate.
3. Blending an inferior quality ghee with a superior quality product.
15.4.4 Neutralization of high acid ghee
High acid ghee produces harmful effects in body due to high content of oleic acid. In this method
ghee is heated to 60-70oC and good quality lime produced to 60 mesh is sprinkled on the surface
@ 3 % of ghee. The temperature is quickly raised to 108oC with gentle stirring and the mass is
cooled and filtered at 60oC by the above neutralization method, the flavor, granularity and keeping
quality of high-acid ghee are all improved.
15.4.5 AGMARK: Ag = Agricultural, Mark = marking
Taking care of ghee and vegetable oil in India.
Collect the ghee samples and melted, kept in container. The samples send to different places like
I II III IV
Test for melting point, free fatty acids, butyrorefractometer reading.
If all the above parameters are correct and within the standards, then AGMARK certify the product.
AGMARK ghee is packed under two grades, viz., ‘special’ and ‘General’, which are represented by
two differently colored labels. The only difference in the gradesis in the maximum limit of free fatty
acids (oleic acid), which in special grade (red label) ghee is limited to 1.4 % and in general grade
(green labels) to 2.5 %.
15.4.6 Grading of ghee
Score card of ghee
Grading parameters Perfect score
Flavor 45
Texture 10
Acidity 25
Color 10
Freedom from suspended particles 5
Package 5

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15.4.7 Physiochemical quality of ghee
Ghee made from buffalo milk is whitish with greenish tinge in comparison to the cow milk
ghee. Which is due to tetrapyrazole pigments-biliverdin and bilirubin. These pigments will
present or interact in conjugated linoleic acid along with protein and gives greenish color.
The flavor of pigment is not only by one compound.
Flavor of ghee develop during heating process due to interaction between protein (casein)
and reducing sugar (lactose) and minerals. Some of the flavoring compounds are produced
during fermentation of cream which is natural flavor. When fermentation is omitted the
flavor of ghee tends to be flat oily. A wide range of chemical compounds has been identified
is a possible contribution to typical aroma of ghee. Which are propanone-ethanol, η-
butanol, η-butanone, 2-ketohydrocarbons of 3 and 9 carbon chain length, keto glycerides,
δ-lactones.
15.4.8 Nutritive values
Conjugated linoleic acid (CLA) present in milk fat has been recognized as anticarcinogenic
properties. The CLA content of milk fat increases substantially during manufacture of ghee.
The natural microbial action during fermentation of dahi in traditional process of ghee
making also increases the CLA content to 1 %.

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Chapter 16

Fermented milks

16.1 General aspects

16.1.1 Preservation
The fermenting of milk is a fairly simple, cheap, and safe way to preserve milk. The lactic
acid bacteria alter the conditions in the milk in such a way that most undesirable organisms,
including pathogens, cannot grow or even die. These conditions include a low pH (4.6-4.0),
which also help in maintaining a low pH in the stomach after consuming the milk; growth
inhibition by un-dissociated acids (e.g., lactic acid) and by other metabolites such as H2O2
and compounds an antibiotic activity; a low redox potential; consumption by the lactic acid
bacteria of compounds that are vital for the growth of other organisms. Appropriate
pasteurization of raw milk kills any pathogens that may survive the fermentation.
Microbial, chemical, and physical spoilage of fermented milk products can occur. Yeasts
and molds may keep growing at a pH below 3.8. Accordingly, these are by far the most
important sources for microbial defects. Pasteurizing the milk and preventing its
recontamination are thus required. The growth of aerobic yeasts and molds in
contaminated products is determined by the extent of contamination, the temperature and
time of storage, the amount of air in the package, and the air permeability of the packing
material. Incidentally, in certain types of sour milk products yeasts, and sometimes also
molds, are part of the essential flora. When the products are kept for a long time or when
they are stored at a high temperature, the ongoing action of enzymes of the starter bacteria
can also cause defects, such as bitter, acid, and cheesy flavors.
An excessive acid production in the milk leads to an objectionable acid flavor. At equal
titratable acidities, the pH of concentrated milk products is higher, which causes the effect
of an excessive acid production of the flavor to be less readily perceptible. Contamination
by metals like copper and iron and exposure to light may have adverse effects on the flavor
of the fermented products. The packing material can also cause off-flavors. The different
factors of physical deterioration (such as wheying off) are:
a. Pretreatment of the milk: The viscosity of the fermented products will increase by
pasteurization and homogenization and decrease the risk of wheying off.
b. Composition of the milk: Lower casein content increases the risk of the defect.
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 c. Inoculation temperature: The higher the incubation temperature, the greater the
tendency for whey separation.
d. The pH of the product: The high as well as a low pH enhances wheying off.
e. Mechanical treatments of the fermented milk, such as stirring and pumping, should be
performed continuously, especially before pH 5 is reached.
f. Presence of air cells and gas cells: Beating in of air during manufacture of the product
enhances wheying off, and so does gas production by coliforms, yeasts, etc. Excessive
production of CO2 by starter bacteria may also cause it and it may follow from a change
of CO2 from the dissolved into the gaseous state caused by an increase in temperature.
Therefore, for instance, buttermilk is designed to some extent before packing.
16.1.2 Nutritive value
The nutritive values of fermented milks predominantly deal with yogurt and are mainly
carried out on animals. The most important aspects when a fermented milk product is
compared with plain milk.
16.1.3 Composition
a. Lactose content: Fermentation decreases the lactose content but is not continued to
such a low pH that any further sugar breakdown is impossible because the resulting
product would become too acidic. At a lactic acid content of, say, 0.9 % the
fermentation is often slowed down by cooling. About 20 % of the lactose in the milk has
then been split, if both glucose and galactose are fermented. In yogurt twice as much
lactose is split since most of the yogurt bacteria do not decompose galactose.
b. Vitamin content: Lactic acid bacteria often require certain B-vitamins for growth, and
can produce other vitamins. Accordingly, the properties of the culture involved largely
determine the extent to which the concentrations of vitamins in the fermented milk differ
from those in the original milk. In yogurt the folic acid content may be increased.
The vitamins contents in fermented products are also affected by the storage
conditions and especially by the pretreatment of the milk. For instance, heat treatment
of milk results in a decrease of vitamins B1, B12, C and folic acid.
c. Other changes due to bacterial action are nutritionally insignificant.

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 d. Composition can be changed by much process steps as standardization and ultra
filtration, and by addition of skim milk powder, caseinates, stabilizers, flavorings, or fruit
pulp.
16.1.4 Nutritional aspects
a. Edible energy: The fermentation process per se does not cause of substantial change of
energy content of milk. The conversion of lactose to lactic acid reduces the energy value by
only a small percentage.
b. Digestibility:
Protein and fat: The digestibility may be improved by a slight predigestion of the
compounds by enzymes of the lactic acid bacteria. People with a weakened intestinal
function may take advantage of the predigestion, but healthy people digest these
compounds efficiently. In the stomach, the protein in fermented milks coagulates into finer
particles than in plain milk, which may increase digestibility. The gastric juice of babies
contains little acid and accordingly, sometimes (dextrorotatory) lactic acid is added to baby
formulas.
Lactose: Lactose intolerant users digest a sour milk product like yogurt much better than
plain milk. The lower lactose content plays a part.
In addition factors must exist that cause easier digestion of lactose. The lactose activity of
the yoghurt bacteria as well as the stimulation of the lactase activity of the intestinal
mucosa by yoghurt have been held responsible. Alternatively, the depletion of the stomach
contents into the duodenum may be retarded when fermented milks are consumed; the
contact time of lactose hydrolyzing enzymes with the substrate in the stomach would be
extended, resulting in a better digestion of lactose.
c. pH adjustment: The fermented milks consumption will cause a smaller increase of the
pH of the stomach contents and thereby diminishes the risk of passage of pathogens. This
is of particular importance for people suffering from a weakened secretion of gastric juice,
e.g., many elderly people and babies.
d. Antimicrobial action: Lactic acid bacteria can form antibiotic compounds that injure
pathogens in vitro. The in vivo significance of these compounds in suppressing
gastroenteritis is not quite clear.

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e. Absorption of minerals: Due to low pH of fermented milk; some minerals are better
soluble than in plain milk; it is sometimes assumed that a better absorption of minerals is
thus to be expected. However, the absorption of various elements, especially that of Mg
and Zn, is enhanced by lactose.
The lactose content decreases by fermentation, causing the net absorption from sour from
sour milk to be lower. The absorption of phosphorus, is less affected by lactose. There has
no uptake of minerals in the fermented milk.
f. Some additional positive and negative effects of fermented milk
a. Intestinal flora: The consumption of living lactic acid bacteria through fermented milk is
supposed to result in the implantation of a favorable flora of lactic acid bacteria in the
large intestine, the flora might repress pathogens. The most probable effects result from
bacteria that do not only survive the action of gastric juice in the gastrointestinal tract but
can also colonize in the intestine, such as strains of the intestinal bacteria Lactobacillus
acidophilus, L. salivarius, and Bifidobacterium bifidum.
b. Cholesterol level: The consumption of fermented milk might contribute to a decreased
cholesterol content of the blood and, accordingly, could reduce the risk of approaching
heart and vascular diseases. However, even if this is true, the effect would be small.
Consumption of fermented milk could further contribute to an increased resistance to
pathogens by activating the immune system and to a decreased risk of colon cancer.
c. Dental carries: Fermented milks have not been shown to cause caries due to damage to
the enamel at low pH. The lactic acid bacteria of the mouth flora from no sticky dextrin’s
from lactose (they form these from sucrose) and they consequently cause no dental
plague. Obviously, the saliva has adequate counteracting activity to prevent dental
caries.
d. Cataract: The consumption of yogurt can cause this eye disorder. Rats exclusively fed
with yogurt (made of concentrated milk) went blind because of the accumulation of
galactitol in the eye lens.
However, unlike the rat, humans can readily convert galactose to glucose; therefore, the
galactose content of the blood does not increase and no galactitol is formed.
e. Lactic acid type: The type of lactic acid formed has physiological significance. Two
sterio-isomers of lactic acid exist: dextrorotatory L(+) lactic acid and levorotatory D(-)

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 lactic acid. L (+) lactic acid can readily be metabolized in the body, but D(-) at a slower
rate. The latter acid is partly removed from the body through the urine. In traditional
yogurt some 40 % to 60 % of the lactic acid is levorotatory and is formed by
Lactobacillus delbrueckii ssp. bulgaricus. Ingesting excessive quantities of D(-) lactic
acid may cause acidosis, resulting in some tissue injury.
16.2 Various types
Fermented milks can be subdivided according to a number of variables, such as type of
fermentative process, fat content, concentration of the milk, withdrawal of the whey, and
use of milk from various species.
16.2.1 Types of fermentation

1. The fermented products are purely lactic fermentation. The fermentation is of two types:
a. Mesophilic starters, consisting of Lactococcus lactis, Leuconostoc cremoris/lactis and/or
Lactococcus lactis ssp. lactis biovar. Diacetylactis. Examples are sour milk, butter milk
and related products, sour cream etc.
b. Thermophillic starters, composed of:
• The flora of Streptococcus thermophillus and Lactobacillus delbruekii Ssp. bulgaricus,
used in yoghurt .
• A pure culture of Lactobacillus acidophilus, used for the manufacture of acidophilus milk;
or a flora of this organism and/or Bifidobacterium bifidum combined with the yogurt
bacteria, used for the manufacture of yogurt-like products.
Acidophillus milk owes its existence to its supposed therapeutic value. L. acidophilus is not
a natural representative of the milk flora. It grows slowly in milk and, hence, contamination
during the manufacture of acidophilus milk, must be avoided. Sterilized milk is inoculated
with a large amount of starter (2 % to 5 %) and inoculated at about 38oC for 18-24 h. The
product is cooled to 4oC and distributed soon. This is because L. acidophilus is fairly acid
tolerant, due to which the lactic acid content of the milk can become high, i.e., some 1 % to
2 % if stored at an insufficiently low temperature; the flavor of the milk becomes sharp and
the number of living bacterial cells decreases quickly. The latter problem can be overcome
be blending plain milk with a deep-frozen concentrated culture of the starter, and by
keeping the mixture at a low temperature (say, 4oC), which prevents the milk from souring.

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Yoghurt-like products may contain L. acidophilus and/or B. bifidum in addition to S.
thermophillus and L. delbruekii ssp. bulgaricus. Alternatively, S. thermophillus can be
combined with L. acidophilus, or with B. bifidum, or with both types of bacteria. The
resulting products are designated as Bioghurt, Bifighurt, and biogarde, respectively.
2. Fermented milk combining a lactic fermentation with alcohol production, e.g., kefir and
kumiss.
16.2.2 Fat content
The fat content will vary in different product like sour milk, buttermilk, and related products,
and sour cream.
Sour milk is prepared by acid production in whole or separated milk at 20oC by using a D
starter. The standards for the fat content vary widely and this holds also for the lactic acid
content (0.5 % to 1.5 %).
Butter milk derives from the churning of cultured cream in the manufacture of butter.
Related products (cultured buttermilk or cultured skim milk) are made by souring skim milk
at 20oC aromatic L or DL starter. Sometimes, the milk used for the manufacture of cultured
buttermilk is required to have a minimum fat content of, say, 0.4%. This is because of flavor
becomes too acid at lower fat contents. The milk is preheated (e.g., 20 s at 80-85oC) to
increase the viscosity of the cultured buttermilk. After having attained the desirable acidity
for the viscosity and the flavor, the milk is stirred until it is smooth, degassed, cooled, and
stored at about 4oC.
Real or churned buttermilk has a more characteristic flavor than cultured skim milk,
probably due to the higher level of membrane compounds of fat globules, especially
phospholipids. The higher the fat contents of the cream, the higher the phospholipids
content of the resulting buttermilk. The difference in composition renders cultured skim milk
far less susceptible to oxidized flavor. Churned buttermilk, especially if derived from high-fat
cream, readily shows flavor defects and thereby becomes unacceptable. Vitamin C can be
added to retard lipid oxidation. At present buttermilk is sometimes manufactured by
churning sour milk; the phospholipids content will be only a little higher than the cultured
skim milk, causing the product to have shelf life than traditional buttermilk.
Sour cream is high-pasteurized, 18 % to 20 % fat cream, homogenized (preferably at a low
temperature, to promote formation of homogenization clusters), and is inoculated with an

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aromatic starter and incubated at 20oC. During the acid production the homogenization
clusters flocculate, resulting in a highly viscous cream. To increase the firmness some
rennet and/or a thickening agent are sometimes added to the sweet cream. When the pH
has reached 4.5 the cream is cooled during gentle stirring and packed. Alternatively,
souring in the package may be applied.
16.2.2 Concentration of the milk
The milk used for making concentrated yogurt is reported. Excessive acid production then
is less detrimental to the flavor due to the higher buffering capacity of a concentrated
product.
16.2.3 Withdrawal of whey
This is another way to prepare concentrated milks. An example is the manufacture of ymer.
Ymer is a Danish soured milk drink. High-pasteurized milk is acidified to pH 4.6 by using an
aromatic starter; the sour milk is gradually heated to 35oC and some syneresis occurs. The
CO2 formed causes the curd to float. The whey is drawn off, homogenized cream is added
to the curd, and the mixture is stirred, cooled, and packed. Ymer contains 11.5 % fat-free
dry matter, 6.5 % protein, and 3 % fat. It is a high-protein and relatively low-calorie product,
with a fairly thick but pourable consistency. Alternatively, ymer is made by direct
fermentation of ultrafiltered milk; the product is stirred and packed after
fermentation.Various other concentrated fermented milks are made by applying
ultrafiltration processes.
16.2.4 Milks of various animal species
Most fermented milk are made of cows’ milk. For some products milk of other species is
applied, especially ewes’, goats’, and mares’ milk. Well-known products are kefir and
kumiss. There is also Greek style yogurt, i.e., a type of concentrated yogurt. It is made of
ewes’ milk and therefore has a high fat content.
KEFIR is made of ewes’, goats’ or cows’ milk. During the fermentation lactic acid and
alcohol are produced. Origionally, the milk drink was made in Russia and Southwestern
Asia. Origionally, the milk drink was made in various countries on an industrial scale by
using cows’ milk.
The microflora used in kefir are Lactococci (Lactococcus lactis ssp. lactis, ssp. cremoris
and biovar. Diacetylactis), leuconostocs (L. lactis, L.cremoris, and Lactobacilli (L. brevis, L.

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kefir, somewhat also L.delbruekii ssp. bulgaricus, L. acidophilus) can have lactic acid, while
yeasts, including Candida, Kluveromyces, and Saccharomyces species, produce alcohol.
Kefir of a satisfactory quality is believed to contain acetic acid bacteria also. Typically, the
organisms involved in the cultured product are present in structures (grains). During
fermentation of the milk the grains grow due to coagulation of protein while they become
connected by means of a formed polysaccharide (“kefiran”).
Kefir is a cream like, sparkling, acid milk drink. Its lactic acid content is 0.7 % to 1 % and its
alcohol content ranges from 0.05 to 1 %, but it rarely over 0.5 %. These levels depend on
the incubation and storage conditions. Metabolites should be formed in certain proportions
to obtain a good flavor. Some conversions are detrimental to quality; an example is the
formation of acetic acid from alcohol by the yeasts after uptake of oxygen from the air.
In the traditional manufacture of kefir, milk with added active grains is first kept for some
time at a temperature of 20-25oC to enhance the lactic fermentation. Subsequently, the
grains are sieved out of the milk and the milk and the milk is further ripened at a
temperature of 8-10oC, which stimulates the alcoholic fermentation.
Modern ways of processing use homogenized, pasteurized whole or standardized milk.
The milk is not inoculated with the grains as such but with soured milk obtained by sieving
a previously fermented culture of grains. A certain amount of L starter may be added. The
inoculated milk is put in well-closed packages and incubated. In this way,“ firm kefir” is
obtained. A considerable amount of gas forms during the fermentation. Inoculation time
and temperature determine the properties of the final product, i.e., amount of lactic acid,
alcohol and CO2, and aroma. In the manufacture of “stirred kefir,” the milk is fermented at a
fairly high temperature, slowly cooled while stirred, further ripened at low temperature, and
packed. Modern packing material, e.g., Al-foil-capped plastic cups, cannot resist a high
CO2 pressure and ballooning can readily occur. Accordingly, a hole is made in the foil, or
the fermentation is stopped at an earlier stage, at the expense of the traditional
characteristics. Continuous production of kefir is also possible. A substitute of kefir can be
obtained by adding sucrose to butter-milk (e.g., 20 g/L) together with the yeast
Saccharomyces cerevisiae and incubating for 3-4 days at 18-21oC in a closed firm
package.

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Kumiss is a well-known milk drink in Russia and western Asia. The cultured milk formerly
was valued because of its supposed control of tuberculosis and typhus. The product is
traditionally made of mares’ milk. The fermenting flora is variable, as in kefir.
Kumiss is a sparkling drink. It contains 0.7 % to 1 % lactic acid, 0.7 % to 2.5 % alcohol, 1.8
% fat, and 2 % protein, it has a grayish color. During its manufacture protein is substantially
degraded. Together with the fermentation compounds formed, the proteolysis is
responsible for a specific flavor. The fermentative processes must proceed in such away
that the metabolites are formed in certain proportions.
Traditional kumiss is not manufactured on an industrial scale. To raw mares’ milk,
temperature 26-28oC, 40 % starter is added, which increases the acidity to 50oN. (The
starter is propagated as a kind of continuous culture in mares’ milk.) The mixture is
intensely stirred and subsequently left undisturbed , which rises the acidity to 60oN. The
milk is stirred for an additional hour, to aerate it and to obtain dispersed protein particles,
and it is bottled. The bottles are kept for a few hours at 18-20oC and then for a certain time
at 4-6oC, which temperature sequence enhances the lactic acid and alcoholic fermentation.
An imitation product of kumiss is now being made on an industrial scale, starting from
cows’ milk. Compares to mares’ milk, cows’ milk has a high ratio of casein to serum
proteins and a low lactose content. The composition of mares’ milk is therefore simulated
by mixing cows’ milk and ultrafiltered, heated whey; heating of the whey is necessary to
inactivate rennet. The starter contains Lactobacillus delbrueckii ssp. bulgaricus and
Candida kefir.
16.3 Yoghurt
16.3.1 Introduction

The name yoghurt have known by different names in countries to countries.Turkish yoghurt,
Egyptian leben, Armenian matzoon and Indian and Nepalese dahi are all similar products.
Originally, yoghurt was made from boiled concentrated whole milk, but most modern
methods of manufacture use whole or partly defatted milk containing small amounts of
skim milk powder or concentrate. The fat content in youghurt may vray from 0-5 percent
and solids contents from 9-20 percent.

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16.3.2 The yogurt bacteria
Two microorganisms, Lactobacillous bulgaricus and Streptococcus thermophillus growing
together symbiotically, are responsible for the lactic fermentation of yoghurt. In some
countries, it contains lactose fermenting yeast, Leuconostoc strains, Str. diacetilactis and L.
acidophillus are also added to improve the flavour of the product.
The method of culture control is very important in yoghurt manufacture, and for this reason
stock (mother) cultures are best maintained individually, rather than mixed. The optimum
pH and temperature for growth of Str. Thermophillus is 6.8 and 38oC. This organisms
normally attain acidities of 0.85 – 0.95 % , whereas Lactobacillous bulgaricus reaches
acidities of 1.20- 1.5 %. The extreme sensivity of both these micro-organisms to penicillin
makes it essential to select antibiotic-free milk for yoghurt manufacture and all starter
culture propagation.
Mother culture is preepared as follows: Fresh or reconstituted skim milk is autoclaved at 15
psi for 10-15 minutes, cooled to 41oC and inoculated with 0.2–1 % inoculum. Cultures of S.
thermophillus are incubated at 38oC and L. bulgaricus at 43oC. Normal coagulation occurs
in 12-18 Hrs, at which time the cultures are cooled to 5oC. Transfer should preferably be
made daily. A commercial yoghurt culture, which contains both microorganisms should be
incubated between 41- 43oC, or according to the supplier's instructions.
Bulk starters are carried in stainless steel vessels or vats with sufficient capacity to contain
2 parts of starter for every 100 parts of yoghurt made. The necessary quantity of skim milk
is heated to 85-88oC for 30-45 minutes, cooled to 43oC and inoculated preferably with 1 %
each of L. bulgaricus and Streptococcus thermophillus, which in a starter or product have a
marked effect on its flavour and odour. Starters are mixed in the milk and the latter
incubated at 41-43oC until coagulation occurs. The bulk starter is cooled to 10oC and stored
at 5oC if not required for immediate use.
16.3.3 Methods of yoghurt production
16.3.3.1 Traditional process

The cow or buffalo milk is received. It is filtrated with fine wire net or muslin cloth to remove
visible dust, dirt and other contaminants. It is then subjected to boiling to 2/3rd of the
origional volume which may cause partial concentration. It helps to increase the total milk
solids and prevent the syneresis of water from the curd after fermentation. The boiled milk
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is then cooled to a incubation temperature it is roughly to room temperature. It is then
adding the starter culture i.e. previous days yoghurt culture.It is incubate in bulk until
coagulum is formed i.e. overnight at room temperature. The yoghurt formed is cool in home
refrigerator and dispatch in the market.

Milk reception Adding starter culture

Filtration Incubation

Cooling
Boiling
Dahi
Cooling
Dispatch

Fig.16.1 Traditional production of yoghurt.

16.3.3.2 Manufacture of set and stirred yoghurt
Receiving milk Heat treatment
Preliminary treatment Cooling to incubation Starter propagation
temperature
Pre-warming to 50-60oC
Incubate with starter culture
Homogenization
Produced set or stirred yoghurt

Set yoghurt Stirred yoghurt

Packed in retail container Incubate in bulk

Incubate Mixed Mixed with Cool

with fruit synthetic flavor
Cool Package Mixed Mixed with synthetic
Incubate Incubate with fruit flavor and color
Dispatch Dispatch
Cool Cool Package Package
Natural/ Dispatch Dispatch Natural/plain
plain Dispatch Dispatch
yoghurt
yoghurt Fruit Flavored Fruit Flavored
yoghurt yoghurt yoghurt yoghurt

Fig.16.2 Flow diagram for the production of improved method of set and stirred
yoghurt.

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16.3.4 Physical properties
The physical structure of yoghurt is a network of aggregated casein particles onto which
part of the serum proteins have been deposited due to their heat denaturation. The network
encloses fat globules and serum. The largest pores of the network are of the order of 10
µm. The existence of a continuous network implies that the yoghurt is a gel, a viscoelastic
material characterized by a fairly small yield stress (say, 100 Pa). If the gel is broken up, as
in the making of stirred yoghurt a fairly viscous non-Newtonian liquid can be formed; it is
strongly shear rate thinning and has an apparent viscosity. Set and stirred yoghurt have
markedly different textures.
a. Firmness of Set yoghurt: Firmness of set yoghurt is often estimated by lowering a probe
of given weight and dimensions into the product during a given time. The reciprocal of
the penetration depth than is a measure of firmness. Firmness is not closely related to
an elastic modulus, but rather to a yield stress. Its value depends on the method of
measurement, especially the time scale, and on several product and process variables.
i. Casein content of the milk: firmness is about proportional to casein content cubed.
Natural variation in casein content can thus have a marked effect. Evaporating the
milk, adding akim milk powder, or partial ultrafiltration increase firmness.
ii. Fat content: The higher the fat content, the weaker the gel because the fat globules
interrupt the network.
iii. Homogenizing the milk: It leads to a much enhanced firmness because the fat
globules then contain fragments of casein micelles in their surface coat, by which
they can participate in the network upon acidification. The volume fraction of casein
is thus effectively increased. (homogenizatin of skim milk is no difference.)
iv. Heat treatment of the milk: It considerably enhances firmness. The deposition of
denatured serum proteins increases the volume fraction of aggregating protein; it
also may alter the number and the nature of the bonds beteween protein particles.
Milk is generally heated for 5-10 min at 85-90oC.
v. Yoghurt cultures vary with firmness they produce ( at a given acidity), but as rule the
differences are small.
vi. Incubation temperature. The lower it is, the longer it takes before a certain pH, and
thereby a certain firmness, is reached, but the finished product is much firmer.

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 vii. Temperature of the yogurt. For the same incubation temperature a lower measuring
temperature gives a greater firmness.
16.3.5 Flavor defects and shelf-life
A main quality problem with yogurt is that souring tends to go on after delivery to the
retailer, and the product may be too acidic when consumed. Moreover, the yogurt may
become bitter due to excessive proteolysis, this would also depend on the starter strains
used. The development of these defects generally determines the shelf life. Of course, the
product is cooled to slow down acidification, but it is difficult to cool fast enough. Set yogurt
is present in a package and cannot be stirred, stirred yogurt should not be stirred too
vigorously because it would then become too thin. And even at refrigerator temperatures,
acidification and other changes caused by the enzyme systems go on, albeit(even though)
slowly.
Other defects may be caused by contaminating organisms, mainly yeasts, molds. The off-
flavors may be characterized as yeasty, fruity, musty, cheasy, or bitter, and occasionally,
sopy-rancid. A flavor threshhold is generally reached at a count of about 104 yeasts +
molds per ml. The growth of these microbes is largely determined by the amount of oxygen
available, hence by the head space volume and the air permeability of the container.
Another defect is the flavor. It may be due to a low incubation temperature, an excessive
growth of the streptococci, or by the lactobacilli being weak aroma producers. Insufficient
acidification e.g., because the milk is contaminated with penicillin, also leads to a blend
product.
16.3.6 Dahi or curd
It is well known fermented milk product consumed by large sections of the population
throughout the country, either as a part of the daily diet or as a refreshing beverage. The
conversion of milk into dahi is an intermediary step in the manufacture of indigenous butter
(Nauni) and ghee.
Definition: Dahi or curd is the product obtained from pasteurized or boiled milk by souring,
natural or by a harmless lactic acid or other bacterial culture. Dahi may contain additional
cane sugar. It should contain same percentage of fat and solid-not-fat as the milk from
which it is prepared. Sweet dahi can be prepared by using Str. Lactis, Str. Diacetilactis, Str.

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cremoris ; single or in combination with or without Leuconostoc species. Sour dahi can be
prepared by using the above organisms along with L.bulgaricus or S. thermophillus or both.
Requrements for fermented milk products:

Requirements
Characteristics
Sweet dahi Sour dahi
Acidity, as lactic acid (%) Max 0.70 1.0
Yeast & mould count per g. (Max) 100 100
Coliform count per g.(Max.) 10 10
Phosphatase test -ve -ve

Food and nutritive value: It has been observed that fermented milk products including dahi
has good food and nutritive value as compared to the original milk.
i. It is more palatable, as compared to milk.
ii. It is easily digested and assimilated than milk.
iii. It has more therapeutic value in the stomach and during intestinal disorders.
16.3.6.1 Composition of whole milk dahi
Water 85 – 88
Fat 5–8
Protein 3.2– 3.4
Lactose 4.6– 5.2
Ash 0.70– 0.72
Lactic acid 0.5 – 1.1
16.3.6.2 Method of production

Sweet/Sour dahi
Traditional method
This mainly involves production on a small scale , either in the consumer's or in the sweet-
meat-maker's shop in urban areas. In the household , the milk is boiled, cooled to a body
temperature , inoculated with 0.5 – 1 percent starter (previous day's dahi or buttermilk) and
then allowed to set undisturbed overnight. In the cooler weather, the dahi-setting vessel is
usually wrapped up with woollen cloth to maintain warmth.

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 Milk receiving Homogenization-2500 psi
Packaging
Pre-heating (35-40oC)
Pasteurization (80-90oC
Incubation (22-25oC
Filtration/clarification for 15-30 min
for 16-18 hrs
Standardization Cooling(22-25oC)
Dahi
Pre-heating(60oC) Inoculation Cooling and storage

Fig 16.3 Flow diagram for the production of dahi.

Details of production
Fresh, sweet, good-quality milk (cow, buffalo or mixed) is received, preheated to 35 – 40oC,
and subjected to filtration/ clarification. It is then standardized to 2.5 – 3.0 % fat and 10 %
SNF ( in order to improve the body), pre-heated to 60oC and homogenized in single stage
at a pressure of 2500 psi. The milk is pasteurized at 80-90oC for 15-30 minutes, cooled to
22-25oC and inoculated with 1-3 % of specific starter culture. It is then filled in suitable
containers (glass bottles/ plastic cups, etc) of the required capacity and incubated at 22-
25oC for 16-18 Hrs., during which period the acidity reaches 0.6-0.7 % and a firm curd is
formed. The curd is cooled to less than 12oC in about 1 hour ( by circulating chilled water or
air around the containers) and then stored at about 5oC in a cold room.
Sweetened dahi
It has a characteristic brown colour, a cooked and caramelized flavour and a firm body. It is
prepared commercially by adding 6.25 % cane sugar to milk either before boiling or at the
time of setting. The pronounced and intence heating causes the milk to brown and get
partially concentrated (the volume gets reduced to about ¾th of the origional). Artificial
colour, sugar caramel and jaggery are also added during production. After heat treatment,
the milk is cooled to room temperature and then seeded with the previous day's product. It
is usually set in wooden pot called (Theki), and earthenware basins and finished product
obtained after 15-16 hours.

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Chapter 17
Production of cheese

17.1 Definition
Simply cheese may be defined as “the curd of milk separated from the whey and pressed into solid
mass”.
Davis (1965) has defined cheese as “a product made from the curd obtained from milk by
coagulating the casein with the help of rennet or similar enzymes in the presence of lactic acid
produced by added or adventitious micro-organisms, from which part of the moisture has been
removed by cutting, cooking and/or pressing, which has been shaped in a mould, and then ripened
by holding it suitable temperatures and humidities”.
According to the description of hard cheese in PFA (1955), cheese (hard) means the product
obtained by draining after coagulation of milk with a harmless milk coagulating agent under the
influence of harmless bacterial cultures. It shall not contain any ingredients not found in milk,
except coagulating agent, sodium chloride, calcium chloride (anhydrous salt) not exceeding 0.02 %
by weight, annatto or carotene color, and may contain certain emulsifiers and/or stabilizers, viz.,
citric acid, sodium citrate or sodium salts of orthophosphoric acid and polyphosphoric acid, not
exceeding 0.2 % by weight; wax used for covering the outer surface contain anything harmful to
the health. In case the wax is colored, only permitted colors should be used. Hard cheese should
contain not more than 43 % moisture and not less than 42 % milk fat of the dry matter. Hard
cheese may contain 0.1 % of the sorbic acid or its sodium, potassium or calcium salt; or o.1 % of
nisin.
According to the definition of FAO/WHO Standard No. A-6 (1978), cheese is the fresh or
matured solid or semi-solid product obtained by coagulating milk, skimmed milk, partially skimmed
milk, cream, whey cream, or buttermilk, or any combination of these materials, through the action
of rennet or other suitable coagulating agents, and by partially draining the whey resulting from
such coagulation.

17.2 Brief history of cheese

Cheese has been known to mankind for thousands of years. Cheese is an art that predates the
biblical era. The origin of cheese has been dated to 6000 to 7000 BC. The worldwide annual
production is more than 12 million tons and is growing at a rate of about 4 %.

Through out the ages numerous varieties of cheese have evolved. The development of cheese
closely depended on local conditions, climate, type of milk, and other economical and geographical
285
factors. Furthermore, it was a matter of technological experience and of finding, by trial and error,
the manufacturing conditions that led to a product being tasty, easy to handle and durable. Along
with this, the general level of technological and hygienic know-how was of great importance.
Various process steps that now are very common are relatively new, say 100 years old: use of
starter, use of industrially made rennet, washing of curd, pasteurization of cheese milk. Of more
recent date are the uses of inocula for the surface flora, application of latex emulsions to the
cheese rind, use of enzyme preparations to accelerate ripening, and so forth.

17.3 Basic principles of cheese making

17.3.1 Concentration
Milk can be coagulated either by acidification or renneting. Both methods entail that the casein of
milk becomes insoluble and forms an unbroken network throughout the milk. The precipitation
takes place in different ways in the two methods. The rennet coagulum is relatively firm and elastic;
an acid coagulum is less cohesive. A common feature of the two types of coagulum is that they
have a tendency to contract and release whey, a feature upon which all cheese making is based.
The contraction and whey exudation of the coagulum can be accelerated by cutting, by heating
and, in the case of rennet coagulum, by acidification. Thereby, the casein and the constituents
enclosed in the casein network are concentrated and the result is a cheese curd which can be
shaped in various ways.
If the protein free whey or permeate is removed from the milk before coagulation by using
ultrafiltration, the whey drainage after coagulation must be correspondingly smaller.
17.3.2 Preservation
The cheese curd contains valuable nutrients and it is exposed to destruction by bacteria, yeasts
and moulds. It is therefore necessary to keep the number of harmful microorganisms in the cheese
as low as possible and to inhibit the growth of the harmful microorganisms that do exist. It means
the cheese must be preserved.
The number of harmful microorganisms can be limited by good milking hygiene, by pasteurizing of
the milk, and by good production hygiene. The factors which are especially useful for inhibiting the
growth of unwanted microorganisms are: low moisture content (concentration), acidification,
salting, addition of saltpetre, and various kinds of cheese surface treatment. Storage at lower
temperatures will increase the keeping qualities.
17.3.3 Ripening
Cheese curd may be eaten fresh, but it may also be subjected to a ripening process, i.e., a process
in which the milk solids are changed so as to bring about the characteristic flavor, consistency and

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appearance of different varieties of cheese. These changes are caused by enzymes and
microorganisms in the cheese curd and, in many cases, also by microorganisms on the surface of
the cheese. The ripening process may last several months.
There are thus three basic processes:
1. Concentration [coagulation, whey exudation (in cheese vat, during pressing and during
salting), evaporation during storage]
2. Preservation (hygiene, pasteurization, concentration, acidification, salting, addition of saltpeter,
surface treatment, cooling).
3. Ripening [changes in total solids(protein, lactose, fat)]
These three processes are used for the manufacture of all cheese varieties. By controlling
these processes in different ways – more or less whey drainage, stronger or weaker
acidification, different moulding, different surface treatment, addition of different
microorganisms, storage at different temperatures, etc. It is possible to manufacture a large
number of very different cheese varieties from the same raw material i.e., milk.
17.4 Retention of constituents of milk
At the beginning, the three-dimensional casein network which is formed during coagulation
encloses all the other milk constituents. When the coagulum contracts, water and the
constituents dissolved in the water are squeezed out, whereas fat globules and bacteria are
retained in the fine-meshed casein network.
17.4.1 Protein
As casein is the principal constituent inn the coagulum, the retention figure for rennet
casein will be close to 100. A small loss of casein may occur through very small cheese
particles being lost with the whey. Rennet casein makes up some 74 % of the milk protein,
and as some of the whey proteins also remain to the cheese, the retention figure for protein
about 75.
The high temperature pasteurized cheese milk the retention figure for protein will be higher
because heat-denatured whey proteins will precipitate together with the casein. The
retention figure will not be higher than approx. 88 because low molecular weight nitrogen
compounds and peptides, as well as the peptide which is cleaved from the casein by the
rennet, will always follow the whey.
When the milk protein is concentrated by ultrafiltration prior to rennet coagulation, the whey
drainage, and thus the loss of protein through the whey, can be reduced. This method
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increases the protein retention to about 93, because low molecular weight nitrogen
compounds and small peptides will follow the permeate.
17.4.2 Fat
The fat globules are retained by the fine-meshed casein network. The usual retention for fat is
around 92 (88-95). The rest of the fat is released during cutting of the curd and enters the whey
from the surface of the curd grains.
17.4.3 Lactose
The retention figure for lactose is low, usually in the range of 3-5 %, equivalent to the amount of
whey remaining in the cheese.
17.4.4 Ash
The ash constituents of milk are found in the true solution, whereas others are present as colloidal
particles linked to the casein particles. It is especially parts of calcium and phosphate that is found
linked to the casein, and these constituents will therefore go mostly into the cheese. The rest of the
ash constituents, which are found mainly as ions in true solution, will follow the whey out of the
cheese. In rennet cheese about 30-40 % of the ash goes into the cheese, but in acid curd cheese
the fat retention is lower because calcium and phosphate are released from the casein during
acidification.
When ultrafiltration is used, the extent to which ash goes into the same cheese can be reduced
slightly by means of diafiltration. The ash especially the calcium and the phosphate ash goes into
the cheese to perform a pH adjustment by means of pre-ripening prior to ultrafiltration. The lower
the pH value is during ultrafiltration, the less calcium and phosphste will remain linked to the
casein, and the lower the ash retention will be.
17.4.5 Citric acid
It is found in milk in true solution just like lactose. Approx. 10 % of the citric acid is linked to the
casein together with the calcium in the same way as phosphate. The retention will depend partly on
the whey content of the cheese, partly on the acid development during whey exudation or prior to
ultrafiltration.
17.5 Classification of cheese
There are hundreds of different varieties of cheese. The classification of cheeses is based on a
number of factors like raw material, type of consistency, appearance (interior and exterior), fat
content, moisture content and ripening methods. The most commonly used criteria are the
moisture content of the finished product and mode of ripening. A classification of these cheese
varieties can be made in several ways.

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17.5.1 Types based on moisture content
a. Very hard (maximum 34 % moisture)
b. Hard ( maximum 39 % moisture)
c. Semi-hard / semi soft ( 39 – 50 %)
d. Soft ( 50 – 80 % moisture)
17.5.2 Types based on mode of ripening
a. Bacteria ripened: Ripening is brought about by different bacteria like
streptococci, lactobacilli, pediococci, leuconostocs,
propionibacteria, Brevibacterium etc.
b. Mold ripened: Mold spp. like Penicillium spp.
c. Unripened: Ripening not involved.
17.6 Composition of different varieties of cheeses
Cheese constituents, viz., fat, protein, moisture, minerals and vitamins differ greatly with the variety of the
product. The composition of some important cheeses has been shown below.
Different classes of cheeses
cheeses

Natural cheeses Miscellaneous cheeses

Bacteria Mold Unripened White Whey Processed

ripened ripened brined cheese (or process
cheese e.g. cheese)
e.g.Feta ricotta
(whey)
Very hard Hard Semi Soft e.g., Mysost
(grating) hard Cambridge,
e.g., (semisoft) Limburger
Parmesan
, With Without
Ramano, eyes e.g., eyes e.g.,
Asiago swiss Cheddar, Semi-hard/ semi soft Soft (surface-ripened)
(old), (Emment Chesire, (internally ripened) e.g., Camembert, Neufchatel
Sbrinz al), Provolone, Blue, Roquefort,
Gruyere, Stilton Gorgonzola, Stilton
Herrgard

Internally- ripened e.g., Surface ripened

Edam, Gouda, e.g., Brick
Fig.17.1 Interrelationship between different classes of cheeses.

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 Vitamins Energy
Mois content
Types of Vit.A Thiamin Riboflavin
ture Fat Protein Calcium
( Kcal/
cheeses % % (µg/1 (µg/100 (µg/100 g)
% % 100g)
00 g) g)
Hard
(Cheddar) 35 33 26 0.83 380 50 0.50 400
Semi-hard
(Edam) 43 24 26 0.76 250 60 0.35 320
Blue-vined
(Roquefor) 40 31 21 0.32 300 30 0.70 360
Soft
(camembe 51 23 19 0.38 240 50 0.45 280
rt)
Unripened 79 0.4 16.9 0.09 3 30 0.28 82
(cottage)

Table 17.1 Composition of different types of cheeses.

17.7 Nepalese cheeses

In Nepal, cheese production started in 1953 . Despite its history of about 47 years
cheese production has not substantially improved, qualitatively as well as quantitatively. In
the recent years, DANIDA has been playing a material role in promoting cheese industry in
Nepal.
Nepal produced some 350 MT cheese in 2000. In 1994, Nepalese cheese generated a
foreign exchange of $ 525,000. The statistic may not be very impressive but is nevertheless
encouraging. The demand for cheese is steadily increasing. At a very conservative
estimate, the annual requirement of cheese in Nepal is around 800 MT. As of now, Nepal
has been meeting the demand for cheese by importing them from foreign countries. Such
cheeses are usually sold at prices about 5-10 times higher than the price for Nepalese
cheese. Cheese making can thus be a very important economic activity for Nepal.

The important varieties of cheese produced in Nepal are Yak cheese, Kanchan cheese,
Mozzarella–like cheese and Processed cheese. Yak- and cow-milk cheeses are produced
by both Dairy Development Corporation (DDC) and private sectors. A substantial
percentage of private yak cheese is of lower quality than DDC yak cheese in terms of taste,
storage qualities, texture and health safety. This is due to private producers’ inexperience,
shortcuts in production that save resources, difficulty in obtaining quality inputs and poor

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credit facilities that make investment cost high (Colavito,1994). From this and other facts
(personal communication) it can be said that the situations for other varieties of cheese are
also much the same.

17.1.1 Yak cheese

Cheese from yak - or chauri milk has been collectively termed yak cheese. Yak and
Chauri are reared in the alpine regions of Nepal. Yak cheese is a hard cheese with unique
taste and characteristics. The uniqueness, for the most part, is due to the chemical
composition of the milk (Table 3). Tourists prize yak cheese as a souvenir from Nepal. The
credit of introducing yak cheese production in Nepal goes to Dr Warner Schulthess (a
senior specialist from FAO) and Gauri P. Sharma. According to a recent report, Dairy
Development Corporation (DDC), Nepal runs six yak cheese factories. There are some 10
private yak cheese factories operating alongside DDC plants. The annual production of yak
cheese in the fiscal year1999/2000 was about 150 metric ton. Composition of yak and other
cheeses of Nepal are given in Table 2 below.

Parameter Yak Processed Kanchan Mozzarella

Moisture (%) 38 41 38 NA
Fat (%, dry basis) 45 45 47 NA
Salt (%, dry basis) 1.37 1.87 1.12 NA
pH 5.75 5.78 NA NA
Table 17.2 Some chemical compositions of Nepalese cheese varieties
NA : Data not available.
Table 17.3 Proximate composition of milk from milch animals of Nepal
Parameters Cow Chauri/ Yak Buffalo
Fat (%) 3.5 – 5.0 7.3 7.3
Protein (%) 3.4 5.5 3.5
Lactose (%) 4.6 4.7 4.8
Ash (%) 0.75 0.85 0.78
Total solids (%) 12.25 – 13.75 18.35 16.38
Milk Solis Not Fat (MSNF) 8.75 11.05 9.08
(%)
Water (%) 83.25 – 87.75 81.65 83.62
Source: Cheese production in Nepal (2001).

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 Steps followed Processing parameters
Quality tests: Organoleptic, COB, alcohol test (in some cases),
Milk
fat test, Acidity test etc
Standardization 3.5 % fat
Pasteurization 65oC for 30 min
Cooling 33oC
1% starter culture( Streptococcus thermophillus and
Pre-ripening
Lactobacillus helveticus in the ration 1:1); 33-37OC for 5-12 min
2.5.g rennet powder / 100 L; temp ≈ 25-30oC; Stirring for 4-5
Renneting
min; Setting for 30-35 min
Cutting 3×3×3 mm cubes
Pre-stirring 30 min
Whey drainage 30%
33oC →35oC→45oC→50oC in 30 min (i.e., 10 min for each
Cooking
stage)
Final stirring 30 min
Molding Wooden mould
Pre-pressing 1-2 times the weight of cheese; duration ≈3 h
Final pressing 5-6 times the weight of cheese; duration ≈16-17 h
Brining 24% salt; temperature ≈ 11-13oC
Ripening 4 months; temp ≈ 11-16oC; 80-85 % RH
Final preparation Washing with cold water and paraffining; 5-6 kg block

Fig. 17.2 Flow diagram for Yak cheese manufacture.

17.1.2 Kanchan cheese

Kanchan cheese making process closely resembles Swiss Gruyere process. Due to the
process developed overtime, kanchan cheese has now emerged as a variety in itself.

Kanchan cheese is prepared from cow’s milk. It is a variety on its own. It closely
resembles Swiss Gruyere cheese (Kanchan cheese does not have eyeholes, though. Dairy
Development Corporation has established a standard method but the process seems to
vary somewhat based on experience of cheese makers. For hard cheese production,
ideally, milk should be renneted at 31-40ºC. However, Kanchan cheese makers of Nepal,
rennet the cheese milk at a relatively high temperature (38-40ºC). The main reason behind
it is the lack of facilities for maintaining uniform temperature.

Production of kanchan cheese

Steps followed Processing parameters used

Milk Quality tests: Organoleptic, COB, alcohol test (in some
cases), fat test, Acidity test etc
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 Remove dust, dirt, hair, nail and suspended visible and
Filtration/Clarification
invisible matter.
Standardization 3 % fat
Pasteurization 63oC for 30 min
Cooling 36oC
Saltpeter addition ≈25 g per 100L milk
Starter culture: 1-2 % at 30oC; inoculation temp: 36-38oC;
Ripening
Duration: 10-30 min (depending on the acidity)
2.5 g rennet / 100L milk; Temp.: 37-38oC; Setting for 30-35
Renneting
min at 37-38oC
Cutting Small cubes (2×2×2 cm3)
Stirring Gently and slowly for 5-10 min
With constant stirring up to 48-50oC within 30 min or until the
Cooking
curd pieces appear firm enough.
Draining ⅓ of the whey
Cheese grains scooped off in cheese cloth to obtain cheese
Scooping off
block of 10 kg
Hooping The mass kept on mould for pressing
10 times the weight of cheese; Turning at intervals of 0.5, 1.0,
Pre-pressing 2.0, 5.0, and 8.0 h with change of cheese cloth at each
turning
Final pressing Final pressing for about 12-14 h
During the pre-pressing intervals the irregular edges removed
Trimming
with a sharp knife
Brining 23-25% salt solution for 48 h
Storing/Ageing Temp.: 12-14oC ; Duration: 3-4 months
Wiping the cheese surface with salt solution and turning each
Final preparation
day for ripening until dispatched
Fig.17.3 Flow diagram for Kanchan cheese manufacture from cow milk.

17.1.3 Mozzarella cheese

True mozzarella is made from the milk of the water buffalo. Today, what is called
mozzarella is a cow’s milk adaptation of the original process. Distinction between true
mozzarella and that from cow’s milk is rapidly fading. In Nepal, mozzarella is produced from
cow’s milk. It is also prepared from a mixture of cow and buffalo milk. It is produced by

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some of the private cheese makers. It is mainly consumed by hotels for making pizzas.
Although the cheese is very popular, due to various reasons, the quality of cheese does not
remain consistent. The main problem encountered in making mozzarella has been found to
be lack of access to good culture.
Production of Nepalese mozzarella
Steps involved Processing parameters
Quality tests: Organoleptic, COB, alcohol test (in some cases),
Milk reception
fat test, Acidity test etc
Standardization Separation of excess cream; Fat adjusted to 3 %
Pasteurization HTST pasteurization; Cooling to 36oC
1 % starter culture(Streptococcus diacetylactis and
Leuconostoc cremoris); 0.5% yoghurt culture (thermophillic
Pre-ripening
lactic culture); if needed, 20 ml CaCl2/ 100 L milk; stirred well
and left for about 15-30 min
2.5 g rennet powder/ 100 L milk (25 ml equivalent of rennet
Renneting
solution) ; Setting for 30 min
Cutting of the curd 1 cm3 size; Duration of cutting ≈ 20 min
Heating Slow heating to raise the temperature to 42oC within 0.5 h
All the whey removed when the temperature reaches 42oC ; pH
Draining
of the whey should be 5.8
Temperature raised to 42oC ; The mat of cheese curd cut in to
Cutting
large pieces
The powder cheese kept in water having temp. of 80oC and
Cooking double the amount; Stirred and cooked until the whole curd
melts to form a plastic mass
Molding The plastic mass is kept on moulds
Cooling The moulded mass is chilled immediately to 4-6o C in water
Storage / Marketing Sold fresh
Fig. 17.4 Manufacture of Nepalese mozzarella.
17.1.4 Danish mozzarella
In Denmark, two types of mozzarella are made. They are normal mozzarella and soft
mozzarella. Of the two types, soft mozzarella has more similarities with Nepalese
mozzarella. A brief flow diagram for both have been shown below. Danish mozzarella is
rindless, whitish cheese, found on a number of export markets around the world. Danish
Mozarella is specially designed for use in warm dishes. Through heating, the cheese’s fine,
mild, and slightly acidulous flavor is brought out and it is this piquant taste which
characterizes a good result.

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Production of Danish Normal Mozzarella
Steps involved Processing parameters
Cheese milk
Pasteurization 72oC for 20 min
Pre-ripening 33-35oC for 20 min
Renneting 30 ml per 100 L milk, left for 25 min
Cutting 5 mm cubes; resting for 3 min
Pre-stirring Very gentle, 15 min
Whey drainage ⅓ of milk
Intermediate stirring For 10 min
Scalding By addition of water at 70oC; Final Temperature : 39oC
Final stirring At pH~ 6.0
Drainage + Pressing
Cheddaring Mixing with dry salt ~ 2 % of the estimated cheese
Stretching In hot salt water at 95oC, 6 % NaCl, Final temperature in
cheese ~ 60oC
Filling To give forms
Cooling In cold water ~ 6oC for 1 h
Vacuum packing
Marketing
Fig.17.5 Flow diagram for normal mozzarella manufacture.
17.1.5 Soft mozzarella
Production of Danish soft mozzarella
Steps Processing parameters
Milk
Standardization
Pasteurization Temp.: 72oC; Time : 15 sec
Pre-ripening 30-45 min ; 38oC streptococcus thermophillus (DVS = 20
g/ 100 ml)
Renneting 38oC, 35-40 min till the firm curd is formed
Cutting 30 mm cubes
Stirring As little and gentle as possible, by hand
Resting on the whey Until pH = 5.0- 5.1
Molding and stretching Water temperature 75-80oC, Cheese temp.: 55-60oC
Molding in a balls
Chilling in ice water
Packing in cups
Storing At 4-7oC in salt brine ( 5% NaCl + 1% CaCl2

Fig. 17.6 Flow diagram of soft mozzarella manufacture.

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17.7.1 Cheese milk
To obtain a satisfactory result in cheese making, a requirement is that the milk used is of
the highest quality both bacteriologically and chemically.
Standards requirements for high-quality milk
• Few bacteria.
• The milk must come from healthy cows. There must be no disease related bacteria.
• Good ability to starter culture.
• Good ability to rennet.
• Milk must be of natural composition.
• Good smell and taste.
• Few butyric acid bacteria.
Few bacteria
When bacteria grow, they convert fat, protein and lactose into acids or other foul testing
substances, which can even be toxic. These substances are not broken down by
pasteurization, and they harm products taste. Many bacteria are not inactivated through the
pasteurization process used on milk destined for cheese production.
Milk must come from healthy cows
Milk from diseased cows has an unsatisfactory culturing and renneting ability. There is also
the risk of spreading contagious diseases.
Milk ability to culture
It is an obsolete necessity for obtaining the desired taste, consistency, and shelf life that the
cultivated lactic acid bacteria should be able to grow and form lactic acid and flavoring.
The milk stored in refrigeration cultures a little slower than fresh milk, but the difference in
that respect is so small that in by far the majority of cases, this will not be perceived in
practice. Because with small changes in starter culture technology and possibly with a
maturing of milk one can completely compensate for any possible decrease in acidification.
If the milk titer rises 3-4 degrees, an over-ripening of the milk has occurred, and the starter
ability is decreased. Storing the milk too long can also encourage the formation of
bacteriophages which also reduces the milk’s ability to culture.

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The starter ability is, to a greater extent, depended on the cow’s diet, cell composition, and
the milk’s dry matter content. Other important factors are the milk’s cleaning and
disinfecting substances, antibiotics, and very important bacteriophages.
• Cleaning and disinfecting substances: chlorine concentration may affected the starter
ability appreciably. The quaternary ammonium compounds (0.0002-0.005 %) presents
in milk creates initial inhibition of starter ability.
• Bacteriophages: Dairies must focus on a significant increase in hygienic standards,
especially aimed at lowering bacteriophage infection. Whey should under all
circumstances be pasteurized in order to kill the bacteria and, therefore,
bacteriophages’ host organisms also killed. Whey and milk should not be handled in the
same skimming and pasteurizing system.
Whey spills on the floor, etc should be prevented to the greatest extent possible, and
the floors should be rinsed with chlorinated water several times for day as well as being
carefully disinfected at the end of each work day. All joints should be kept in perfect
condition.
Waste water ejection and handling of waste water should be done as far way from the
cheese production facility as possible. There should be quick and easy asses for all the
personnel to disinfect their hands, arms etc, in chlorinated water; similarly, the work
place should be arranged so that the personnel perspire as little as possible. For the
culturing of cream and returned milk, the culture stock should be different from that
used for the cheese. The cheese starter culture should be cultivated in a special room
where all precautions are taken to prevent infection.
Similarly, one can use Ca++ free milk, as bacteriophage can’t grow in this milk. In
regards to preventing bacteriophages, closed cheese tanks and vats are a step forward.
Good renneting ability
In cheese production, it is necessary that the milk is able to form a firm curd and later
syneresis. Milk can have very different abilities to rennet, and this is caused in part by
differences in the amount of Ca++ in the milk.
Various factors influence calcium content as well as phosphate content, which has a
special significance for renneting. Late lactation milk and milk from mastitis cows contains
especially small amount of the two substances.

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If the milk is stored at a low temperature, β-casein is released from the protein-micelles
together with Ca and PO4. (CaPO4 is more easily dissolved at low temperature). Its
changes involve a use of H+, as well as an increase of pH, which results in slower rennet
coagulation.
Pasteurization of milk precipitates Ca and PO4, but a slow re-dissolving can occur, so that
the renneting ability is regarded the longer the milk stands after pasteurization. To
compensate for the precipitation of Ca++, one can add CaCl2 to the milk.
If the milk’s salt content (NaCl) is too high, there will occur an ion exchange with calcium,
so that calcium is driven out of the casein by the salt, which to a certain extent decreases
the milk’s renneting ability.
Milk must be of natural composition
Colostrum, late lactation and mastitis milk do not have natural composition and the cheese
process will not be normal. The ratio between fat and protein must be normal.
• Blending in of colostrums milk
The milk contain colostrums may coagulate easily in pasteurization temperature, because
of containing high albumin and globulin content. So no blending of such milk. The cheese
grains is completely abnormal and less ability to press water, from colostrums milk. Such
milk is unfit because almost completely curdling during heating. Lactic acid bacteria also
grow very poorly in this milk. The acid production is too weak and regular spoiling of the
cheese can occur.
All of these things happen because the milk is not of natural composition. The Ca-content is
nearly double that of regular milk. The ratio of Ca to Na2O + K2O is nearly 4 times greater
than the regular milk.
• Late lactation milk
Late lactated milk is poor for cheese making, because of its large alkali salt content (K and
Na salts). This produces a dissolving of colloids, in which the protein is significantly more
hydrated than in normal milk, and coagulation and water-processing from the cheese grains
will be consequently worse. If the milk has a salty or bitter taste, it should not be used for
cheese. Such milk also has a poorer starter ability.
• Mastitis milk
Such milk has a lower casein, Ca and phosphorus content than normal milk. It has:

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 - a lower yield,
- a poor renneting ability.
• Milk’s dry matter content
It would be natural to choose milk which has the highest protein content and therefore
produces the greatest cheese yield. In selecting milk based upon fat and protein content,
one must be sure to understand that with a previously selected cheese making technique,
there must be a very specific ratio between milk’s content of fat and protein in order to
achieve a satisfactory curdling.
• Addition of water
When the milk’s content of protein is high during the autumn season water is added. In this
way, one achieves a prolonged renneting time and a lower coagulant, which has lost some
of its ability to squeeze out whey. As a result, the water content in the finished cheese
becomes higher, but when the whey is thinned with water, the lactose content falls and the
cheese can contain more whey than normal without becoming sour. The result: the cheese
is softer and less sour. The water content in the cheese increases 0.2 % and increases pH
by 0.01 for every 2 % addition of water to the volume of the milk.
The milk must have good smell and taste
Substances causing bad smell and taste found in milk cannot be destroyed through
pasteurization. Milk with a bad smell or taste will normally also impart a bad smell or taste
to the cheese.
Milk can have a poor smell and taste, even at the time of milking, as a result of poor diet.
The smell and taste lowers the cheese quality, because it can not be removed.
There are great problems with beet taste caused by feeding the animals large amounts of
beet or beet-tops. Turnips contain mustard containing glucosides.
Few butyric acid bacteria
-“late blowing cheese” with bad smell and taste.
Low butyric acid bacteria would be possible in the following ways:
• Cheese dairies accept only milk from non-silage fed livestock.
• Cheese dairies sort the milk at pick-up and use spore rich milk for other purposes than
cheese production.
• Milk dairies produce spore-poor silage.

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 • Udder hygiene is considerably improved.
17.7.1.2 Additives to the cheese milk
Additives to prevent fermentation faults
Early blowing (gassy curd)
Coliform bacteria can ferment lactose into different kinds of organic acids, etc., and large
amounts of carbon dioxide and hydrogen. Hydrogen is insoluble in water and will therefore
immediately cause eyeholes to be formed in the curd. Hydrogen gas produces strongest
effects, which tears apart the curds and completely destroy the structure. Carbon dioxide
can also cause difficulties if it is produce in large amounts and at an early stage. The
coliforms also create an off-flavor. Besides gas, some of these microorganisms contain
volatile and unpleasant tasting acids. Some are rich in proteases and cause a through and
unfortunate breakdown of proteins.
Lactose is usually fermented by the lactic acid bacteria in the course of 24-48 hours and
when the process is complete, the coliforms can no longer produce gas in the cheese. If the
growth of the lactic acid bacteria is inhibited, the fermentation of lactose will be retarded,
and this increases the possibility of even small amounts of coliform bacteria causing a
gassy curd.
Saltpetre prevents the coliforms from producing hydrogen and, thus, blowing is prevented.
However, saltpetre does not inhibit the growth of the metabolism of the coliform bacteria.
Effective pasteurization and good hygiene will help to make the less number of such
coliform bacteria or rapid growth of starter culture also helps to make unfavorable
environment for the growth of such bacteria. Coliform bacteria may also cause the type of
gassy curd characterized by closely-spaced eyeholes with thin, broken membranes
between them.
It is therefore important in the production of the above-mentioned cheeses to avoid these
fermentation faults. First and foremost, infection of the milk by these microorganisms
should be prevented as far as possible through better hygiene at the point of production,
as well as during processing with various provisions not only against infection but against
reinfection after pasteurization as well.

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Late blowing
Butyric acid bacteria are spore-forming and the spores are not killed by pasteurization.
Butyric acid bacteria are found in soil and, especially, in bad silage, and they may find their
way into milk through manure particles. Butyric acid bacteria grow relatively slowly in
cheese but, as they ferment lactate (lactic acid salts), they can grow in the cheese during
ripening. Butyric acid bacteria can ferment lactate into butyric acid, CO2, H2, etc., and
thereby cause blowing of the cheese and development of off-flavors (sickly, bitter-sweet
taste). Butyric acid fermentation usually starts at a relatively late point in the in the ripening
process, ie., butyric acid bacteria cause late blowing.
Butyric acid bacteria are not inhibited by nitrate, nor can nitrate inhibit their production of
hydrogen. However, the nitrate in cheese can be reduced to nitrate which is toxic to butyric
acid bacteria. This effect is increased by salting and acidification.
Butyric acid bacteria can cause a significant decrease in the cheese’s quality by causing a
strong generation of gas and an unpleasant flavor. They are found in more or less all
cheese making milk. The lactate fermenting Clostridia (Cl. tyrobutyricum, etc), which are
the spore formers that are normally considered to be able to spoil cheese. The vegetative
cells of butyric acid bacteria can be killed or inactivated by heat treatment of the milk, but
spore forms will survive.
Spores remaining in the cheese will change into a vegetative form and will therefore, cause
serious damage in the stored cheese. The growth of butyric acid bacteria can be somewhat
inhibited by using a lower storage temperature, by decreasing the cheeses water content,
and by increasing the level of pH and salt content.
It is necessary to use saltpetre (KNO3 and NaNO3) in some of the Danish cheeses but not
necessary for the production of Cheddar and Cheddar types as well as fresh cheese and
molded cheese.
Nitrates
Saltpetre (KNO3 and NaNO3) –are good preservatives (esp. for meat and pork). It is also
used for a long time as an agent against fermentation faults in cheese (especially in round-
eyed cheeses), and have been of significant use.
The addition of saltpetre as chlorate results in a change in the length of fermentation and
that one could encounter an enzyme blockade. For e.g., in both cases, there is a

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considerable accumulation of formic acid. Addition of nitrites and nitrates also causes the
amount of lactic acid to decrease and the amount of acetic acid to increase. These
additives inhibit the further decomposition of the formic acid.
Further decomposition of formic acid that gas production occurs (CO2 and H2). The addition
of saltpetre to the milk usually takes place before the addition of rennet, but it can be added
to the whey during, for e.g., scalding. The most usual amount is 5-10 g /100 kg of milk.
Lysozome
• Found in chicken egg white.
• Also found in number of other plant and animal tissues, in tears and in milk.
• Present in human milk in a concentration of 0.03 – 0.04 % ( it is 300 times to that of the
cows milk).
• Has proven to have an antimicrobial effect, esp. for gram positive bacteria, including
anaerobic spore formers.
• It chemically breaks down the cell walls and therefore destroys the cells.
• 1-2 g lysozyme per 100 litres of milk was has effective as 10 g of saltpetre per 100 of
milk.
• 20 g per 1000 liters of milk should be used.
Advantages over saltpetre
Lysozyme follows the casein, and that < 1 % of the added amount is lost in the whey.
Saltpetre on the other hand follows the aqueous phase, and therefore the majority of it is
lost in the whey where it is not wanted if it is later to be concentrated and processed
further.
Lysozyme can be found again complete in the cheese, and that it is not inactivated during
the ripening of the cheese, but saltpetre, is slowly reduced during the ripening of the
cheese.
Lysozyme has no effect on coliform bacteria and therefore a high standard of hygiene can
be required.
Calcium chloride ( CaCl2)
A Ca++ is necessary in milk in order to be able to precipitate the para-casein produced by
renneting, and the more Ca++, the stronger the rennet effect, the more solid the
coagulation, and the faster the curdling.

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There must be the fitting ratio of calcium salts and sodium potassium salts in the milk. If
there is too much calcium, the coagulum will be too solid, and if there is too little, it will be
too loose and soft. The addition of CaCl2, is therefore used when the milk originally
contains too little calcium ions, or in order to replace the calcium which is precipitated
during the milk’s pasteurization, or to improve the rennet ability that was weakened by
cooling and/or homogenizing the milk (e.g., in feta). In this way, one gets both more
calcium ions in solution and a higher titre (lower pH which improves renneting ability). The
various other advantageous changes of calcium chloride additions are:
• larger grain size.
• Lower temperature, and
• Stronger stirring by which reduced fat loss in the whey.
• Dissolved CaCl2 in a little boiling water before addition kills any possibly harmful
bacteria.
• It is possible to produce a saturated solution (about 33 %) by boiling CaCl2 in water,
and this selection more than 20 ml/100 kg milk added.
• CaCl2 is added to milk before addition of rennet.
Calcium chloride usually 5-20 g/100 kg milk is used.
Stabilizers and emulsifiers
• To improve the cheeses’ consistency and make than more cohesive and pliant (pliable).
• Fat content in fatter cheeses gives these a softer and more cohesive consistency.
Fatty cheeses are therefore not so sensitive to small changes in starter culture and calcium
content as the leaner cheeses which easily become either tough (too weak a starter
culture) or short in consistency (too strong a starter culture).
Buttermilk can be added and is used as an emulsifier, esp. sweet buttermilk. The content of
fat, copper and phospholipids are highest in sweet buttermilk. These are about 0.63 % fat,
89 µg Cu per kg and 0.90 mg phospholipids. Phospholipids are good emulsifiers, and
together with proteins they form a lipoprotein complex which emulsifies better than
phospholipids alone. Sweet cream buttermilk is more sensitive to oxidation because of
higher content of phospholipids and copper. The renneting ability in milk with added sweet
buttermilk is poor, and therefore quite a lot of CaCl2 must be added. Sweet butter milk in

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amounts up to 30 % of the total amount of milk can substitute for ordinary milk during
manufacture of 30+ cheeses.
Color and Bleaching agents
Color
The color of cheese varies, just as it does in butter, according to the seasons and related
changes in animal feeding.
Fatty cheeses are colored more strongly than lean cheeses because most coloring agents
follow fat. To keep the color constant all the year round and to keep a specific hue in a
specific kind of cheese, the cheese curd is colored by color to the milk before renneting.
The most common coloring agent for cheese is, as in butter, annatto, only in cheese it is
dissolved in a base-most often potash lye- which binds itself to the casein, while in butter
the color is dissolved in plant oil.
Saffron pigment (extracted from the dried parts of a crocus flavor), produces a light yellow
color and has been used to some extent in special kinds of cheese such as Emmentaler
and Parmesan. Carotene is now an excellent color which gives the cheese a yellowish
color.
Bleaching agent
Green-blue chlorophyll stain which gives the cheese a lighter shade can be used as a
bleaching agent because it is a complementary color to the milk’s yellow pigment. Other
bleaching agents are: Aniline stain, Titanium dioxide, Benzyl peroxide etc.
Acidification of cheese milk
Rennet
Many enzymes from both plants, bacteria and animals can cause casein particles to
“polymerize”- creating a network that encapsulates the milk’s other parts in a firm, elastic
coagulum. Usually the enzyme chymosin (chymase) or rennin a sulphur-containing protein,
which is extracted from the fourth stomach abomasums of calves, has been used as the
sole source for producing the coagulation that is a necessary beginning to the usual
cheesing process. The enzyme rennet is added to cheese milk to coagulate the casein
particles which is necessary for whey exudation. The enzymes also play a part role in
breaking down the casein during storing and maturing the cheeses. Chymosin is a
proteolytic digestion enzyme.

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Rennet consists of mainly rennin and pepsin, principally the former being responsible for
milk clotting and the latter for proteolysis (although both possess the two characteristics).
Rennin is liable to heat and alkali besides other physical and chemical agents and gets
inactivated at 70oC in 14 min at pH 6.8-7.0.
Calf rennet
For centuries, rennet has been made from calf stomachs (abomasums, the fourth stomach
or the vell). The stomachs are sliced into strips which are extracted in a slightly acid salt
solution. As rennet enzymes are destroyed by heating to 55-60oC, the rennet extract cannot
be preserved by heating. To obtain the best possible preservation, the pH value of the
extract is adjusted to approx. 5.5, the salt solution to 15-20 %, and benzoic acid is added.
The rennet must be protected from light and stored cold. Under these conditions, it will only
lose about 1 % of its activity (strength) per month. If it is stored at higher temperatures, or is
subjected to light, its activity may decrease much faster. The enzymes are destroyed by
alkali and strong acid.
Most of the coagulation activity of calf rennet is caused by the enzyme chymosin (rennin).
Part of the coagulation activity, however, is caused by another enzyme: bovine pepsin. As
the calf matures the amount of pepsin increases and chymosin (rennin) decreases.
Other types of rennet
The increased demand and production of cheese the world over, coupled with the
decreased culling and slaughtering of calves due to improved management practices has
resulted in a substantial shortage of calf rennet. The resulting high cost combined with
certain sentimental and religious reservations of vegetarian population in consuming the
cheese made with animal rennet substitutes from non-animal source like plants and micro-
organisms. Among the microbial milk clotting enzymes investigated so far, a few have been
introduced in the market commercially by some foreign firms.
Commercial microbial rennets
Trade name Micro-organisms Trade name Micro-organisms
Suparen Endothia parasitica Rennilase Mucor miehei
Sure-curd Endothia parasitica Emporase Mucor pusillus var. Lindt
Fromase Mucor miehei Meito rennet Mucor pusillus var. Lindt
Hennilase Mucor miehei Noury rennet Mucor pusillus var. Lindt
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 modilase Mucor miehei Milcozyme Bacillus polymyxa
Marzyme Mucor miehei Mikrozym Bacillus subtilis
The different milk coagulants (rennet substitutes) hydrolyze κ-casein at or near to chymosin
(calf rennet) sensitive Phe.-Met. bond.
Limitation of microbial rennet as compared to calf rennet:
• Low milk clotting to proteolytic activity ratio,
• High retention of the enzyme in the curd.
These properties tend to cause the hydrolysis of caseins to the point of forming bitter
peptides during cheese ripening.
Rennet activity
The activity of rennet is stated as the quantity (ml) of fresh mixed milk that can be
coagulated by 1 ml rennet in the course of 40 minutes at 35oC. The activity of a liquid
rennet is usually 12,000- 15,000. The activity of powder rennet is usually ten times higher
(calculated per gramme). One part of rennet liquid (about 2 % proteins) is used to clot
about 5,000 parts of milk during cheese-making.
The coagulation process
The coagulation of milk by means of rennet takes place in two phases.
a. First phase, a negatively charged part of the κ-casein (≈ ⅓ of the κ-casein molecule)
is released. Thereby the casein particles lose part of the negative charge which
otherwise prevents the particles from coagulating. The casein which is formed in this
manner is known as para-casein. It is insoluble in the presence of Ca++ (free calcium
ions).
b. Second phase, is the precipitation of the para-casein formed in the first phase. The
assumption is that precipitation occurs because the casein particles-having lost part
of their negative charge- can amalgamate through so-called hydrophobic (water
repellant) bonds between amino acids side chains of the protein. Ca++ is necessary
for the precipitation because the linkage of the positively charged calcium ions to the
negatively charged phosphate groups of the caseins neutralizes a great part of the
surplus of negative charges of the casein.
At 30oC and using 30 ml ordinary cheese rennet per 100 L fresh mixed milk, the first
phase lasts about 10 minutes and the second phase about 1 min. The speed of the
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 first phase depends on the quantity of enzymes, whereas the speed of the second
phase is independent of the quantity.
At first, the coagulum which is formed is very soft and weak, but, gradually, it
becomes firmer and more cohesive. In the case of fresh milk at 30oC, it will usually
be about 20 minutes from the onset of coagulation until the coagulum is sufficiently
firm for cutting.
Factors affecting the coagulation process
The activity of enzymes usually depends on temperature and pH. This is also the case for
rennet enzymes, which are also influenced by the content of Ca++ in the milk.
1. pH
The activity of rennet enzymes is highest in acid media. In connection with the splitting of
the κ-casein in milk (the primary phase), the chymosin optimum is at a pH of 5.4. Even
small changes in the activity of the milk greatly influence the activity of rennet enzymes. In
the case of chymosin, the enzyme acts twice as fast when the pH is lowered from 6.7 to
6.4. The effect of pepsins-both bovine and porcine-increases even more when the pH value
is lowered from 6.7 to 6.4.
Changes in the amount of starter culture added or in the point of time at which it is added
(pre-ripening) will therefore cause changes in the coagulation time of the milk.
2. Temperature
The activity of rennet enzymes is highest at a temperature of approx. 42oC (optimum
temperature) and is lower at both higher and lower temperatures. At 55-60oC, the enzyme
is destroyed. Changes in milk temperature influence the coagulation time considerably. At
30oC, coagulation takes two to three times longer than at 42oC. In cheese making, the
normal renneting temperature is around 30oC (30-33oC).
3. Ca++ concentration
The casein which is converted by the rennet enzyme (para-casein) can only precipitate if
the milk contains free calcium ions. Variations in the content of free calcium ions in the milk
will cause changes in coagulation time, firmness of the curd, and whey exudation.

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Amount of rennet
The usual amount of rennet is 20-40 ml (standard rennet) per 100 kg cheese milk. The
coagulation time is roughly inversely proportional to the amount of rennet, i.e., the
coagulation time is halved if the amount of rennet is doubled.
The whey exudation is unaffected by variations in the amount of rennet. The splitting of
proteins during ripening of the cheese increases somewhat with increasing amounts of
rennet, but the use of larger amounts of rennet also increases the risk of a better flavor
because of the development of a relatively larger amount of bitter-tasting peptides. It would
seem that there are no advantages in using more than 20 ml rennet per 100 kg milk.
17.7.1.3 Curd treatment
The coagulum that is produced when the para-casein formed by the action of the rennet
enzyme is precipitated has a natural ability to contract and squeeze out whey (i.e., water
and the constituents dissolved in it: lactose, whey proteins and ash).
Purpose
The purpose of curd treatment in the cheese vat after coagulation is to promote the
contraction of the casein network and the resulting whey exudation.
Contraction is promoted by cutting the coagulum into small pieces because, thereby, the
distance the whey has travel is shortened, and the cutting provides a much larger surface
area from which whey can escape. Contraction is also promoted by the acid development
during treatment in the cheese vat; finally, it can be increased by increasing the
temperature (scalding). The water binding capacity of casein decreases with increasing the
temperatures. While these processes are going on, it is usually necessary to stir the curd to
make sure that the curd grain surfaces are exposed so as to allow the whey to escape.
Stirring is also necessary to secure a constant temperature throughout the mixture, which is
especially important during scalding. If scalding takes place by means of hot water addition,
stirring also serves to distribute the water among the curd grains.
The whey exudation is the visible result of the curd treatment, a profuse development of
bacteria and acid is taking place inside the curd grains.

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Cutting
After a few minutes of rennet addition the precipitation of para-casein will be taken place. At
the beginning, the coagulum is very soft but it gradually becomes firmer. The coagulum
hardens faster at higher temperature and lower pH values.
Cutting begins when the coagulum has reached a suitable degree of firmness. Before
cutting, the linking of the casein particles must be progressed so far that all the casein
particles and most of the fat globules will remain in the curd grains, i.e., the whey which is
released must be clear and greenish yellow tint but not white.
The rennet coagulum is cut (horizontal and vertical) with cheese knives into pieces,
thereby, increasing the surface area of the curd for easy expulsion of whey. The mode of
cutting the coagulum varies with the variety of cheeses, as it directly determines the rate
and extent of moisture removal and the final cheese texture. The smaller the size of the
curd piece, the greater is the moisture expulsion and harder is the texture (lower moisture)
of cheese obtained. Besides the size of curd pieces, the initial firmness of the rennet
coagulum is also important. Softer coagulum will lead to poor moisture removal and poor
yield due to losses of fat and protein in whey.
Healing
After cutting stages, the curd is kept undisturbed for sometime during which the whey is
expelled out and the curd pieces get immersed and settle towards the bottom of the cheese
vat. The firming up of the curd (or formation of a semi permeable membrane) at this stage
is called healing. Meanwhile, the cheese-maker may wipe or “squeeze” the curd particles
sticking on the sides of the vat. This helps in two ways. Firstly, it reduces ‘cook-on’ during
subsequent stages involving heating. Secondly, clostridial spores, which may be present in
the sticking curd particles are not activated and hence the defect of “stink spots” caused by
the activation of these spores in high cooked out varieties (e.g., Swiss), is avoided.
Fore-working
The curd is subsequently (after 10-15 min) subjected to fore-working (manual agitation with
a paddle or shovel) to avoid its matting or balling.
Pre-drawing
This step involves withdrawing of a portion (about one-third) of the whey from the cheese
vat before subjecting to cooking. Pre-drawing of whey has three-fold advantage:

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• The quality of whey withdrawal is more suitable for processing as its acidity is low
(about 0.11 %) at this stage.
• Withdrawing of a sizeable portion of whey leads to lesser requirements of energy during
cooking.
• The amount of whey to be removed after cooking is lesser and hence there is time
saving during cooking.
Cooking
The cooking temperature ranges from 38-45oC or more depending on the final moisture
required and more depending on the final moisture required and tolerance of starter
bacteria.
• Further contracting the curd particles
• Expulsion of whey.
These effects are controlled by temperature, acidity and agitation during cooking.
Low cooking temperature leads to softer cheese due to higher moisture retention which
facilitates faster acid development by starter bacteria during ripening. Higher temperature
will result harder and more dry cheese curd which ripens slowly. The rate of increasing the
temperature during cooking is also important as a gradual heating rate (0.2o to 0.5oC/ 5 min
initially followed by 1.5o to 2oC/min) prevents “cheese hardening” (the formation of dense
surface layer on curd particles) which retards the expulsion of whey.
Acidity development in and around the curd pieces due to starter activity helps in further
controlling the curd casein leading to greater moisture removal.
Inadequate agitation hinders whey removal as the curd particles lump together and
decrease the effective surface area, thereby, causing poor expulsion of whey.
Draining or dipping the whey
The process is performed to separate whey from curd and to aggregate or coalesce the
curd particles. It takes about 15 minutes. The dipping is a traditional approach in which all
the curd mass was collected in a cheese cloth or scooped into perforated moulds to
separate it from whey. Later, draining practice was adopted in which the curd is pushed
aside in the vat to allow the whey to drain through the outlet.

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Curd trenching / Furrowing
It takes about 10 min. The curd is pushed aside in the vat to allow the whey to drain
through the outlet. The separation of the whey from the curds in a horizontal vat can be
facilitated by raking the curd into long piles along the sides leaving a trench or ditch down
the center. This method of draining is termed as ‘ditching’.
Washing
Washing of the curd with cold water during and after draining is practiced only in certain
cheeses like cottage and Colby but not in other like cheddar. It helps in a following ways:
• Cools the curd.
• Prevents matting the curd particles.
• Reduces acidity
• Leaches out lactose, the available fermentable sugar.
Cheddaring process
Expulsion of whey and to form body and texture. Casein will change to para-caseinate.
“Cheddaring” refers to a combined event of matting (packing), turning, piling and repiling of
curd and further promotes and controls the expulsion of whey.
a. Matting/packing After draining, the curd particles are placed as two long slabs one on
either side of the vat, for allowing the residual whey removal to continue.
b. Turning The small strips/blocks are turned or inverted frequently for about 30 to 45 min
to promote acid production by starter bacteria and whey expulsion.
c. Piling and repiling of curd strips The strips are piled two high or three high (piling)
followed by changing of postion (repiling) within 15 min.
“Cheddaring” is carried out in certain cheeses, viz., cheddar cheese and related types and
usually takes two hours. During this processes, the curd gets transformed from a course
rubber-like mass to a plastic-like flexible mass resembling white meat of chicken. Starter
activity continuous during cheddaring and the acidity produced controls the whey expulsion
and final and final moisture content of the cheese.
Hot iron test – If a hot iron is placed on a curd and make up elastic mass produced because
of converting casein into para-casein. It shows cheddaring completes.

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Milling process
In this stage the cheese blocks are cut into small pieces of sizes 2” × 3”. The reduction in
size of curd increases the surface area and helps it the following ways.
• Promotes further whey expulsion.
• Allow to quicker and more uniform salt distribution,
• Contributes to further cooling of curd before hoping and
• Enables the curd to be pressed into desired forms.
17.7.1.4 Salting
Food grade salt is used. Dry salting can be used for the expulsion of whey. To check the
growth of undesirable microorganisms @ 0.75-1 % or (1.0- 1.5 %) salt on the basis of
amount of curd is used. For example 200 g (First part, second part, third part).
First part should be mixed well; it may penetrate inside the curd and moisture expulsion.
Second part mix which is effective for microorganisms (e.g.,proteolytic and lipolytic types).
However, E.coli, an unwanted coliform, gets stipulated by 3 % salt concentration and
requires about 12 % concentration for its inhibition.
17.7.1.5 Moulding / Hooping The placing of the milled and salted curd into forms, moulds
or heaps to give it the desired shape is called moulding and hooping.
- vertical and horizontal hoops are used.
- rectangular, circular, square, ball, cone, pear, pipe shaped etc and are usually made up of
stainless steel or plastic.
The salted curd is filled in mould. The temperature is about 30-32oC. It gives the desired
shape.
Pressing This refers to an operation of compressing the milled curd particles in the hoops to
the final compact shape. It is carried out in two stages by gradual application of pressure so
that the entrapped whey and air are expelled out properly. Sudden application of pressure
leads to openness, to entrapped air and pockets of free whey. For pressing at about 70 psi
pressure is applied. There should not be any mechanical hole in between the curd particles,
if there is holes aerobic bacteria will grow. It takes about 14-15 hours for complete pressing.
Drying
surface drying – keep it for 4-5 hours at 4-5oC.
Paraffining
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Melted crystalline wax.
17.7.1.6 Ripening of cheese
keeping it in cool temperature 7-8oC.
it also reformed as curing, maturing or ageing.
Process of storage of fresh cheese curd at suitable temperatures till it transforms into a
finished into a finished product of desirable body, texture and flavor. The transformation
involves the activity of ripening agents, viz. microorganisms (starter or non-starter flora)
and enzymes (from milk, rennet and microorganisms) on various cheese constituents
(mainly lactose, protein and fat) to cause physical, chemical (biochemical) and
microbiological changes in the product.
Raw cheese

Milk
Coagulant Microorganisms
(Normally present)

Fats Citrate
Proteins Carbohydrates Fatty acids
Proteoses Sugars Alcohols
Esters
Acetic acids
peptones
Aldehydes Ketones Butyric acids
Peptides
Caproic acids
Amino Flavor
To
acids Stearic acid
Texture Aroma
Body Oleic acid
Ammonia
Hydrogen sulphide

Fig.17.7 Different products formed during ripening of cheese

17.7.1.7 Cleaning and packaging of cheese
Cheeses can be cleaned and packaged before sold. Only a few cheese varieties such as
Camembert are not cleaned before packaging. Semi-hard cheeses are usually paraffin
waxed after cleaning and before further packaging.
Cleaning
Cheeses may be cleaned by scraping or washing. Before washing, the cheeses are
softened in water for 15-20 minutes. To promote the softening of the proteins in the smear

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coat, small amounts of slaked lime or NaCl may be added to the softening water. The
cheeses should not be soaked for too long because this may weaken the rind.
Washing
For washing, macines with rotating brushes or spray nozzles are used. Clean water must
be used for washing and then rinsed with clean water. Instead of it, the chill water may be
used both washing and rinsing. The temperature should probably not exceed 20-25oC
during washing.
Drying
After washing, the cheeses must be dried because paraffin will not adhere to a damp rind.
The cheeses passes through a drying tunnel on a conveyor belt. The temperature of the
drying air should not exceed approx 60oC as higher temperature may cause fat to run off
the cheese surface, which make it difficult for the paraffin coat to adhere to the cheese rind.
Drying may also take place in a dry room with vigorous air circulation. The cheeses are
placed in this room for 24-48 hrs. after washing.
Waxing (Paraffin coating)
Purpose
• To give the cheese a clean and attractive appearance.
• To reduce the water evaporation (weight loss) during further storage and shipment.
• To prevent further development of microorganisms as the cheese surface.
Function
• To form a cohesive, closely adhering membrane around the cheese.
• Paraffin coat is strong and elastic that will tolerate the changes in shape that the cheese
undergoes during the subsequent ripening and shipment.
Waxing: The dipping time is 4-5 seconds and a paraffin temperature of about 150oC is
usually suitable. Lower temperature will give a thicker coat and vice versa. Longer dipping
time will give a thinner coat and vice versa. The cheese should be cooled to ≈ 12oC before
waxing, because it will then be so rigid that it will not be deformed and the paraffin coat will
not break during removal after immersion. After waxing, most cheeses should be stored at
approx 12oC. At higher temperature the cheese may change shape and break the paraffin
coat, and there is also a risk that the CO2 production will be too rapid and, thus, cause
blisters and rind cracking.

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Time for waxing
Waxing must not be performed too early because a sufficiently hard rind must have formed
before waxing, and because the diffusion of CO2 must be more or less complete before the
cheese is enclosed in a tight paraffin coat. Waxing must not be performed too late because
then evaporation of water from the cheese will have been too high, causing too thick a rind
to form (less edible cheese) and too large a weight loss.
Re-waxing
When cheeses have been stored for some weeks in the ripening store or in cold store, it will
often be necessary to re-wax them before shipment.
Cheeses which have stored at a temperature of ≈ 12oC (ripening store) can be re-waxed
immediately, but cheeses which have been cold stored must be warmed at 12oC for 24-48
hrs. to prevent condensation of vapor on the cheese surface prior to re-waxing. It should be
performed at a slightly higher temperature and/or with a slightly longer dipping time than the
first waxing.
Packaging
Inner packaging material
The packaging material which comes into contact with the surface of the cheese. This
material is usually rather thin and its primary function is to protect the cheese from dirt. The
cheese rind and the paraffin coat must also be regarded as packaging materials.
For packaging of ordinary semi-hard cheese varieties, parchment substitutes are used. For
some markets, cellophane is used. Aluminium foil is also used.
Packaging of cheese portions
The best packaging material for these purposes is a synthetic film which, on the one hand,
protects the cheese against drying out(weight loss) and oxygen, and on the other, allows
the consumer to assess the appearance of the cheese. It is generally done to avoid
infection.
Outer packaging material
The purpose of the outer packaging material is to protect the cheese from mechanical
damage during its journey from the place of production to the retailer. The outer packaging
material may be wooden crates or vats, corrugated paper or fiber boxes. The outer

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packaging material must be both practical and easy to handle and it must be packed tightly
around the cheeses to avoid loose cheeses.
17.7.1.8 Grading of cheese
Cheese grading is a fairly complex matter in the sense that, in principle, the evaluation of
cheese should be performed in such a way that it assesses not only the extent to which the
cheese satisfies the general demands of the cheese variety, but also the extent to which
the cheese satisfies the specific demands of the consumers (the market) for whom it is
intended.
Cheese may be graded by making use of the human senses and by making use of
chemical, physical and microbiological methods. The following are the parameters analyzed
for grading the cheeses.
Fat content, Moisture content, Salt content, Acidity and pH, Dirt content, Extent and depth
of ripening, Total bacterial count, Special groups of microorganisms in cheese e.g., coliform
bacteria, Different compounds e.g., various peptides, amino-acids, free fatty acids, metals.
Organoleptic evaluation
Outer appearance of cheese (shape, size, marking, possible packaging, paraffin coat and
rind) including color, cleanness, thickness and strength of rind.
Color, texture, consistency, aroma & taste of cheeses have been evaluated. At the end of
the evaluation, a total number of points is awarded.
The Danish grading scheme operates with a 15-point scale.
The current Danish scale is used as follows:
11-15 points: satisfactory to good quality.
9 -10 points: Average quality (minor, significant fault).
7 – 8 points: Poor quality (one or more significant faults).
0 - 6 points: very poor to defective quality (major faults, putrified).
17.7.1.9 Cheese faults and their removal
Faults in outer appearance
Faults in shape
Atypical. The cheese is manufactured in a different shape &/or size than the one
prescribed.
Deformed. Uneven pressing. Insufficient turning in fermentation store.

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Rounded corners and edges. Too tight cheese cloth during pressing.
Collapsed. Too high moisture and too weak acidification. Too high temp. and humidity in
the fermentation store. Keeping qualities are reduced and faulty fermentation and
unclean flavors may easily develop.
Step sides and sharp edges. Too acid and too salt which delays the ripening.
Concave planes. Cheese has been blown and greatly expanded, but the gas has since
escaped and thereby the cheese has shrunk.
Faults in paraffin coat
Thick paraffin. Too low waxing temperature &/or too short dipping time.
Thin paraffin. Too high waxing temperature &/or too long dipping time.
Cracked paraffin. Weak rind and soft cheese.
Loose paraffin. Poor cleaning &/or drying before waxing. Too early waxing (before sufficient
rind formation.
Blistered paraffin. Too tight paraffin coat &/or too early waxing so that the CO2 cannot
escape through the paraffin coat at the rate at which it diffuses from the center of the
cheese.
Uneven paraffin. Too high waxing temperature and too long dipping time.
Muddy paraffin. Contaminated paraffin mixture.
Marking faults
Smudged. Insufficient care during stamping.
Poor marking. Insufficient care during and application of casein marks.
Rind faults
Weak rind. Insufficient drying and possibly too strong smear formation whereby micro-
organisms in the smear coat have decomposed tha rind.
Thick rind. Too strong drying.
Cracked rind. Too weak rind. Incorrect cutting of pressing edges. Mechanical damage. Too
weak pressing. Excessive kneading and too high pasteurization temperature (kneaded
cheese).
Injured. Mechanical damage. Attacks by rats or mice.

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Attacked. The rind is dissolved in places by microorganisms, e.g., bacteria or yeasts
underneath paraffin blisters or by moulds. The rind may also be broken to a dry powder
by cheese mites.
Smeary, slimy, greasy. Too early waxing of badly cleaned &/or dried cheeses.
Cloth folds. Incorrect application of cloths during pressing.
Discoloration. Colored patches (brown, yellow, black, etc may be pigment from colonies of
pigment-forming microorganisms or they may steam from rust particles from piping,
racks, etc.
Dirty. Insufficient cleaning before waxing. Rust patches. Cheese placed on dirty floors,
trucks, etc.
Faults in inner body
Color faults
Saltpeter color. The whole of the has a slightly reddish color . Too much saltpeter addition.
Saltpeter rim, red-rimmed. The outer layers of cheese are red or brown.
Brown rim. Too strong smear formation. Pigments from the smear coat may be diffuse into
the cheese.
Salt rim. A pale area underneath the rind. Found immediately salting and should disappear
when salt is distributed in the cheese.
Acid rim. A pale area underneath the rind. Too strong acidification in the outer layer of the
cheese which, in turn, may be caused by excessive cooling during moulding and
pressing.
Discoloration. Red, yellow, brown patches in the cheese. Colonies of the pigment-forming
microorganisms. Rust particles.
Blue or brown blotches. May be caused by copper particles or rust particles. Brown or black
patches may also be caused by oil from agitators or stirrers.
Faults in texture
Slits in cheese. Too strong acidification &/or salting (whereby the curd becomes inelastic
and slits rather than round eyeholes are formed)
Close blind. Few or no eyeholes. Too few gas producing bacteria, i.e., too few aroma
bacteria in the starter culture. Too much saltpeter added to cheesemilk. Too low
temperature in the fermentation store.

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Blown. Too strong gas production. Early blowing: Coli fermentation, a large number of relatively
small holes); Late blowing: usually relatively few, but very large holes) or, in some cases, by
too strong propionic acid fermentation. Also caused by certain rod shaped bacteria which
produce gas by converting free amino acids.
Gassy curd. Irregular, closely-shaped eyeholes with thin broken membranes between them. May
be caused by lump formation during stirring or coli fermentation in connection with too slow
acidification. Starter culture with too many Sc .diacetylactis can also cause too early and
irregular formation of eye-holes.
“Rind fermentation”. Too early penetration of salt between curd grains which have not fused
completely. This is prevented by cooling and drying before salting. It may also be caused by
cooling of the cheese before moulding or by loose, cold curd grains being pressed into the
cheese surface. Too weak pressing.
Partial rind fermentation. Lump formation during stirring. Piercing by fingers during transfer into
moulds. Holes in cheese cloths.
Rind cracking. Slits underneath the rind. Secondary fermentation in connection with too tight
paraffin coat.
Mould growth, too much, too little or to regular. Too good or too poor growth conditions throughout
or in parts of the cheese.
Faults in consistency
Rubbery. Too weak acidification (too many Ca-bonds between casein molecules in the casein
network).
Brittle. Too strong acidification &/or salting (too few Ca-bonds between casein molecules in the
casein network). Old cheese.
Hard, dry. Too low moisture content, possibly in connection with low iodine values in the cheese
fat.
Soft. Too high moisture content, possibly in connection with high iodine values in the cheese fat.
Flabby. Soft cheese with too weak acidification.
Pasty. Soft cheese-probably with normal or too strong acidification.
Soft rind. Too high moisture content in the outer layers of the cheese immediately underneath the
rind.
Spongy. Usually kneaded or dipped cheeses with too strong gas production.
Faults in aroma and taste
Acid. Too strong acidification.

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Wheyey, yeasty. Too high moisture content and too slow and too weak acidification, and with a
strong growth of coliform bacteria or yeasts.
Bitter. Too much rennet.
Bitter –sweet. Often in connection with butyric acid fermentation.
Malty. Infection by Sc. Lactis var. maltigenes in the starter culture.
Smeary (possibly putrid). Too strong smear formation. Unsuitable smear flora. Wrapping or waxing
of badly cleaned cheese.
Bland. Too weak ripening. Too low salt content.
Salty. Too high salt content.
Feed flavors. Milk with off-flavors from feed.
Off-flavors. Undesirable flavor which cannot be characterized more precisely.
Unclean. Undesirable flavor which cannot be characterized more precisely. However, it is usually
caused by growth of unwanted microorganisms in the milk or cheese.
17.7.1.10 Processed and spread cheese

Processed cheese
Processed cheese may normally be defined as a modified form of natural cheese prepared with
the aid of heat, by comminuting and blending one or more lots of cheese with water, salt, color and
emulsifier in to homogeneous plastic mass.

Processed cheese is very popular in Nepal. A typical method used for the preparation of Nepalese
processed cheese is as follows:
Ingredients: natural cheese, butter, trisodium citrate, Na2HPO4, common salt and water.
The production flow chart is given below.

Addition of water 18.6 L

Hard cheese per 100 kg of cheese; Na2HPO4: 1.39 kg
Tri-sodium citrate: 1.39 kg
Milling Salt : 0.39 kg
Butter: 7.56 kg
Cheese kettle
Vacuum packaging
Vacuum generation

Cooking at 80oC to melt Molding Cutting

Fig. 17.8 Flow diagram for Nepalese processed cheese.

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 Natural cheese
Additives (water, color,
Analysis and selection of salt and emulsifier)
cheese of proper age groups
Heating and stirring

Tempering and cleaning Packaging

Quartering and grinding Processed cheese

Fig. 17.9 Flow diagram for the preparation of processed cheese.

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Chapter 18

Dairy Plant Sanitation and hygiene

18.1 Introduction
In a dairy plant, the type of space is characterized by the degree of the microbiological risk
involved. The sources of contamination are as follows: human: Offices, washrooms,
cloakrooms, milk receiving areas; process : Milk powder, storage of raw materials, and
finished products, milk reception; Low risk areas : either because the product is not
exposed to the air : pasteurization, separation, evaporation rooms etc; or because the
product is relatively stable at this stage: packaging of butter, cheese etc. The Critical areas
are: Starter culture room, curd room, room used for bottling fresh milk. Obviously,
communication between 1 and 3 should be minimized. In general, access to the critical
areas must be restricted to a small number of employees. These must follow certain
decontamination procedures before entering the room, such as scrubbing their hands with
sanitizer, or walking or an antiseptic material or bath.
Overloaded storage areas, lift trucks, and pallets are often sources of contamination.
Generally, preventive measures can be summarized as follows: proper maintenance of
storage areas, restricting the circulation of personnel and mobile equipment, disposing of
waste as quickly as possible, limiting access to critical zones, keeping the laboratories
properly cleaned etc.
There are many regulations concerning the types of construction materials which may be
used, the protection of electrical circuits the construction of drains and sewers, etc.
18.2 Cleaning
Cleaning or washing of dairy equipment implies the removal of 'soil ' from the surface of
each machine.
Sanitization (Sterilization) implies the destruction of all pathogenic and almost all non-
pathogenic microorganisms from the equipment surface.
(Soil: It consists milk and milk products residues which may be more or less modified by
processing treatment , or by interaction with water or cleaning materials previously used, or
by dust , dirt or other foreign matter.

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 Soil may be one or more of the following: liquid milk films, air-dried films, heat
precipitated films, heat hardened films, milk-stone and miscellaneous foreign matter.)
Milk- stone: It is an accumulation of dried milk solids and salts from hard water and washing
solutions. It consists largely of calcium phosphate, milk protein precipitated, coagulated and
baked on by heat, and insoluble calcium salts from water and washing solutions. It has the
following approximate composition: Moisture (2.7 to 8.7 %), Fat (3.6 to 17.7 %), Protein (4.4
to 43.8 %), Ash (42.0 to 67.3 %).
Detergents or cleaning / washing compounds are substances capable of assisting cleaning.
Sanitizers are substances capable of destroying all pathogenic and almost all non-
pathogenic microorganisms.
18.2.1 Importance of cleaning and sanitizing
All dairy equipment should be properly cleaned and sanitized as milk provides an excellent
medium for the growth of microorganisms. At the same time, detergents and sanitizers
used for cleaning and sanitization should be so selected as not to affect the material of the
equipment.
Cleaning and sanitization are complementary processes; either of them alone will not
achieve the desired result, which is to leave the surfaces as free as possible from milk
residues and viable organisms.
18.3 Detergents
The detergents should have the following desirable properties:
i. wetting and penetrating power;
ii. emulsifying power;
iii. saponifying power;
iv. deflocculating power;
v. sequestering and chelating power
vi. quick and complete solubility
vii.should be non-corrosive to metal container.
viii. free rinsing.
ix. economical,
x. stability during storage,
xi. should be mild in hands,

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xii. should possess germicidal action,
(Note: No single detergents are possesses all the above properties. Hence two or more
detergents are mixed for different cleaning operations so as to efficient cleaning with
safety.)
18.3.1 Classification of dairy detergents
Dairy detergents can be classified into four groups
1. Alkalies: Sodium hydroxide, sodium carbonate, sodium phosphate, sodium bicarbonate/
sesquicarbonate, sodium silicate, sodium sulphite (as inhibitors), etc.
Strong alkalies: to saponify fat.
Weak alkalies: to dissolve protein.
2. Acids (mild): Phosphoric acid, tartaric acid, citric acid, gluconic acid, hydro acetylic acid;
Strong : Nitric acid.
Mild acids are used for milk stone removal; nitric acid may be used in not greater than 1 %,
for stainless steel surfaces.
3. Polyphosphates and chelating chemicals: These are used together with acid or alkalis,
e.g., Tetra phosphate, hexametaphosphate, tripolyphosphate, pyrophosphate etc.
4. Surface active / wetting agents : These are either used alone or in combination with
acids or alkalies. e.g. Teepol, Acinol-N, Idet-10, common soaps etc.
18.3.2 Sanitizers
Desirable properties of sanitizers are:
i. non-toxic,
ii. quick acting,
iii. relatively inexpensive,
iv. non-corrosive to hands and equipments,
v. easily and quickly applied.
Commonly used dairy sanitizers are
- steam, hot water and chemicals (chlorine compounds, iodophor and quaternary ammonium
compounds).
The methods of chemical sanitization
i. Flushing ii. Spraying iii. Brushing iv. Fogging v. Submersion
Consideration for detergent selection
For the selection of any particular detergent, consideration should be given to :

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- type of soil,
- quantity of water supply,
- material of surface and the equipment to be cleaned, and
- method of cleaning, eg. Soaking, brushing, spraying, and / or recirculation.
Detergents are used as an aqueous solution.
In the selection of dairy sanitizers, the following considerations are kept in mind.
a. High temperature sanitizing
It has good penetrating ability and quick drying of the equipment. Heat is the most reliable
sanitizer, especially when both temperature and time are controlled. Thus effective
sanitization can be done by steam ( 15 psi for 5 min. or 0 psi for 15 min) or scalding water (
90-95oC for 10 min).
b. Low temperature sanitizing
i. It permits sanitizing immediately before the equipment is used (when hot equipment will
be injurious to the quality of milk and milk products).
ii. It avoids excessive strain on equipment (such as ice-cream freezers); and
vi. It permits flushing out of equipment immediately before use, thereby removing any
possible dust that may have entered. Generally, chlorine solution at 15-20oC containing
150-200 ppm available chlorine, is used for a contact time of 1-2 min.
18.4 Cleaning and sanitization procedure

Draining - To remove any residual loose

milk and any other matter
Pre-rinsing - With cold water, to remove as much
milk residue and other materials
Warm water - To hot detergent washing with solution
rinsing of 0.15 to 0.60 % alkalinity, to remove
the remaining milk-solids
Hot water rinsing - To remove traces of detergents

Sanitizing - To destroy all pathogens and almost all

non-pathogens (usually also just before
using the equipments).
Draining and rinsing - Help to prevent bacterial
growth and corrosion

Fig 18.1 Flow chart for general cleaning and sanitizing procedure.

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18.5 Selection of detergents and sanitizers
Material Cleaning Sanitization

All alkalies may be used. Care should be All sanitizers may be

Stainless steel
taken with acids. used.

All alkalies may be used, acids should be

Mild steel -do-
used together with inhibitors.

Tinned Weak alkalies, together with sodium All sanitizers may be

steel/Copper sulphite as inhibitor, should be used. used.

Bronze -do- -do-

Galvanized -do- -do-

Weak alkalies, together with sodium

Aluminium alloy -do-
silicate as inhibitor, should be used.

Glass All alkalies and acids may be used. -do-

Weak alkalies, together with sodium

Vitrous enamel -do-
silicate as inhibitor, should be used.

Cleaning temperatures should not be Onlychemical sanitizers

Plastics
above the softening point of plastic. should be used.

Strong alkalies should be used to remove

Rubber -do-
any fatty material stuck to the surface.

Note:
i. Chlorine sanitizers, if left in contact with metal surfaces, cause corrosion. Hence they
should preferably be just before processing.
ii. Inhibitors are substances which minimize the corrosive action of acids or alkalies on
metal surfaces.For eg. Sodium sulphite for a tinned surface and silicate for an aluminium /
Al-alloy surface.
Tri-sodium silicate is usually replaced by sodium carbonate, mainly due to the lower cost
and easy availability. However, sodium carbonate has a lower cleaning efficiency than tri-
sodium silicate.
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The sanitizers chosen may be steam, scalding water (90-95oC) or chlorine solution (150 to
200 ppm available chlorine).
18.6 Methods of cleaning
The different methods are
1. Hand Washing, 2. Mechanical washing and 3. CIP (Cleaning In Place).
18.6.1 Hand washing
Prepare detergent mixture – 0.8 to 1.0 % in tap water. So as to give a minimum alkalinity
of 0.5% (pH over 11.0) in a wash up tank and maintain the temperature at about 50oC.

Rinsing with clean cold water

Introduce the detergent solution into the equipment. Thoroughly brush the equipment
surface, inside and outside, with a clean can-brush.

Washing the utensil with enough fresh cold water, using a clean brush again if needed; to
remove all traces of detergents.

Draining and dry for 1-2 hrs.

Sanitize by steam or hot water after cleaning or by rinsing with chlorine solution (200
ppm, available chlorine) just before using.

Fig. 18.2 Flow chart for the hand washing procedure.

Bottle hand washing Hand brush or a motor driven brush and a mild alkaline solution.
18.6.2 Mechanical washing For can and bottle washing.
Rotary or straight-through / Tunnel type machines are used for washing.
Rotary washing machine is used in small dairy plants. It occupies small space. It can be
operated by a single worker. It can wash 2-6 cans and lids per minutes.
Straight through washer is used in bigger plants. It has a greater capacity, 4 –12 cans & lids
per minutes.

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Procedure

Draining -liquid milk residues

Pre-rinsing -pump fed cold or luke warm water

Draining

Pumping the -not less than 70oC

hot detergents
Draining

Fresh water Rinsing -pump-fed or, by steam and water injector

at 88-93oC
Live Steam injection

Drying – at 95 to 115oC.

Fig. 18.3 Mechanical can and bottle washing procedure

- not greater than 0.5 % alkalinity is desirable for can metal.
- Sanitization with chlorine is not recommended for tinned milk cans.
Bottle Washing
The mechanical bottle-washer may either be a soaker (soaking), or Hydro (jetting) or
Soaker-Hydro (part soaking and part jetting). Further it may be of the come-back or
Straight-through type; in the former, loading and unloading take place at the same end,
while in the latter they are done in opposite ends. Generally, soaker-hydro-come-back
types are used for small capacities, and straight-through-hydro types are used for larger
capacities. In all machines, bottles are loaded manually or semi-manually and are
discharged automatically onto one or more conveyors.
Procedure for mechanical bottle washer
Pre-rinse Using water at 32-38oC

Washing with detergents 1-3 % NaOH together with chelating

and washing agents
Rinsing with warm water Remove all traces of detergents 25-
30oC water is usually recirculated.

Rinsing with cold water Manually recirculated ahlorinated water

(35-50 ppm available chlorine) is used to
prevent recontamination of bottles.
Draining Visual inspection.

Fig. 18.4 Mechanical bottle washing procedure

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18.6.3 Cleaning In place (CIP) or In place cleaning
CIP means that rinsing water and detergent solutions are circulated through tanks, pipes
and process lines without the equipment having to be dismantled. CIP can be defined as
circulation of cleaning liquids through machines and other equipment in a cleaning circuit.
This technique is used for permanent installations with many pipes and tanks, which are
practically impossible to clean by other means. Cleaning conditions vary widely from one
installation to the other. Tanks, pipes, conveyors, etc, can all be cleaned in place.
The passage of the high-velocity flow of liquids over the equipment surfaces generates a
mechanical scouring effect which discharges dirt deposits. This only applies to the flow in
pipes, heat exchangers, pumps, valves, separators etc.
For cleaning large tanks is to spray the detergent on the upper surfaces and then allow it to
run down the walls. The mechanical scouring effect is then often insufficient, but the effect
can to some extent be improved by the use of specifically designed spray devices.
CIP circuits
The type of equipment that can be cleaned in the same circuit is determined according to
the following factors:
• The product residue deposits must be of the same type, so that the same detergents
and disinfectants can be used.
• The surfaces of the equipment to be cleaned must be of the same material or, at
least of material compatible with the same detergent and disinfectant.
• All components in the circuit must be available for cleaning at the same time.
Dairy installations are therefore divided for cleaning purposes into a number of circuits
which can be cleaned at different times.
18.6.3.1 Compatible materials and system design
For effective CIP
• The equipment must be designed to fit into a cleaning circuit and must also be easy
to learn.
• All surfaces must be accessible to the detergent cannot reach or through which it
cannot flow.
• Machines and pipes efficiently drained.

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 • Residual water cannot drain will provide sites for rapid multiplication of bacteria and
cause a serious risk of infecting the product.
Materials in process equipment, such as stainless steel, plastics and elastomers, must be
of such quality that they do not transmit any odor or taste to the product. They must also be
capable of withstanding contact with detergents and disinfectants at the cleaning
temperatures.
In some cases the surfaces of pipes and equipment may be chemically attacked and
contaminate the product. Copper, brass and tin are sensitive to strong alkalis and strong
acids. Even small traces of copper in milk result in an oxidized flavor (oily, train-oil taste).
Stainless steel is the universal material for product wetted surfaces in modern dairies.
Metallic contamination is therefore normally no problem. Stainless steel can however be
attacked by chlorine solutions.
Electrolytic corrosion is common when components made of copper or brass are built into
systems of stainless steel. In such conditions the risk of contamination is great. Electrolytic
corrosion may also occur if a system with steels of different grades is cleaned with cation
active agents.
Elastomers (e.g., rubber gaskets) can be attacked by chlorine and oxidizing agents, which
cause them to blacken or crack and release rubber particles into the milk.
Various types of plastic in process equipment may present a contamination hazard. Some
of the constituents of some types of plastics can be dissolved by the fat in milk. Detergent
solutions can have the same effect. Plastic materials for use in dairies must therefore
satisfy certain criteria regarding composition and stability.
CIP programs
Dairy CIP programs differ according to whether the circuit to be cleaned contains heated
surfaces or not. We distinguish between:
• CIP programs for circuits with pasteurizers and other equipment with heated surfaces
(UHT etc).
• CIP programs for circuits with pipe systems, tanks and other process equipment with no
heated surfaces.

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The main difference between the two types is that acid circulation must always be included
in the first type to remove encrusted protein and salts from the surfaces of heat treatment
equipment.
A CIP program for a pasteurizer, “hot components”, circuit can consist of the following
stages:
1. Rinsing with warm water for about 10 minutes.
2. Circulation of an alkaline detergent solution (0.5 – 1.5 %) for about 30 minutes at 75oC.
3. Rinsing out alkaline detergent with warm water for about 5 minutes.
4. Circulation of (nitric) acid solution (0.5 -1.0 %) for about 20 minutes at 70oC.
5. Post rinsing with cold water.
6. Gradual cooling with cold water for about 8 minutes.
The pasteurizer is usually disinfected in the morning; before production starts. This is
typically done by circulating hot water at 90-95oC for 10-15 minutes after the returning
temperature is at least 85oC.
In some plants, after pre-rinsing with water, the CIP system is programmed to start with the
acid detergent to first remove precipitated salts and thus break up the dirt layer to facilitate
dissolving of proteins by the subsequent alkaline detergent. If disinfection is going to be
done with chlorinated chemicals; there is an imminent risk of fast corrosion problems if any
residues of the acid detergent remain. Therefore, when starting with alkaline cleaning and
ending with acid cleaning after an intermediate water rinse, the plant should be flushed with
a weak alkaline solution to neutralize the acid before disinfection with a chlorinated
chemical can start.
A CIP programme for a circuit with pipes, tank and other “cold components” can comprise
the following stages.
1. Rinsing with warm water for 3 minutes.
2. Circulation of a 0.5 – 1.5 % alkaline detergent at 75oC for about 10 minutes.
3. Rinsing with warm water for about 3 minutes.
4. Disinfection with hot water 90-95oC for 5 minutes.
5. Gradual cooling with cold tap water for about 10 minutes (normally no cooling for tanks).
18.6.3.3 Design of CIP systems

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In practice there is no limitation to satisfy stringent individual demands as to the size and
complexicity of CIP plants.
The CIP station in a dairy consists of all necessary equipment for storage, monitoring and
distribution of cleaning fluids to the various CIP circuits.
The exact design of the station is determined by many factors, such as:
• How many individual CIP circuits are to be served from the station? How many are “hot”
and how many are “cold” ?
• Are the milk rinses to be collected ? Are they to be processed (evaporated) ?
• What method of disinfection is to be used ? Chemical, steam or hot water ?
• Will the detergent solutions be used only once or recovered for reuse ?
• What is the estimated steam demand, momentary and total, for cleaning and sterilization ?
Looking back over the history of CIP, we find two schools of thought:
1. Centralized cleaning.
2. Decentralized cleaning.
Until the end of the 50’s, cleaning was decentralized. The cleaning equipment was located
in the dairy, close to the process equipment. Detergents were mixed by hands to the
required concentration- an unpleasant and hazardous –procedure for the personal involved.
Detergent consumption was high, which made cleaning expensive.
The centralized CIP system was developed during the 60’s and 70’s. A central CIP station
was installed in the dairy. Rinsing water, heated detergents solutions and hot water were
supplied from this unit by a network of pipes to all the CIP circuits in the dairy. The used
solutions were then pumped back to the central station, and from there to the respective
collecting tanks. Detergents recovered in this way could be topped up to the correct
concentration and reused until they were too dirty and had to be discarded.
Centralized CIP works well in many dairies, but in large dairies the communication lines
between the central CIP station and the peripheral CIP circuits have grown excessively
long. The CIP pipe systems contain large volume of liquids, even when they are “drained”.
The water remaining in the pipes after pre-rinsing dilutes the detergent solution, which
means that large amounts of concentrated detergent must be added to maintain the correct
concentration. The greater the distance, the greater the cleaning cost. A move back

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towards decentralized CIP stations therefore began in large dairies at the end of the
seventies. Each department had its own CIP station.
18.6.3.3.1 Centralized CIP
Centralized CIP systems are used mainly in small dairy plants with relatively short
communication lines. The detergent solutions and hot water are kept hot in insulated tanks.
The required temperatures are maintained by heat exchangers. The final rinse water is
collected in the rinse water tank and used as pre-rinsing water in the next cleaning
program. The milk/water mixture from the first rinsing water is collected in the rinse-milk
tank.
The detergent solutions must be discharged when they become dirty after repeated use.
The storage tank must then be cleaned and refilled with fresh solutions. It is also important
to empty and clean the water tanks, especially the rinse water tank, at regular intervals to
avoid the risk of infecting an otherwise clean process line.
A station of this type is usually highly automated. The tanks have electrodes for high and
low level monitoring. Returning of the cleaning solutions is controlled by conductivity
transmitters. The conductivity is proportional to the concentrations normally used at dairy
cleaning. At the phase of flushing with water the concentration of detergent solution
becomes lower. At a preset value a change over valve routes the liquid into the drain
instead of the relevant detergent tank. CIP programs are controlled from a computerized
sequence controller. Large CIP stations can be equipped with multiple tanks to provide the
necessary capacity.
18.6.3.3.2 Decentralized CIP ( satellite CIP system)
Decentralized CIP is an attractive alternative for large dairies where the distance between a
centrally located CIP station and peripheral CIP circuits would be extremely long. The large
CIP station is replaced by a number of smaller units located close to the various groups of
process equipment in the dairy.
This still has a central station for storage of the alkaline and acid detergents, which are
individually distributed to the individual CIP units in main lines. Supply and heating of
rinsing water (and acid detergent when required) are arranged locally at the satellite
stations.

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These stations operate on the principle that the various stages of the cleaning program are
carried out with a carefully measured minimum volume of liquid just enough to fill the circuit
to be cleaned. A powerful circulation pump is used to force the detergent through the circuit
at a high flow rate.
The principle of circulating small batches of cleaning solutions has many advantages.
Water and steam consumption, both momentary and total, can be greatly reduced. Milk
residues from the first rinse are obtained in a more concentrated form and are therefore
easier to handle and cheaper to evaporate. Decentralized CIP reduces the load on sewage
systems as compared to centralized CIP, which uses large volume of liquids.
The concept of single use detergents has been introduced in conjunction with decentralized
CIP, as opposed to the standard practice of detergent recycling in centralized systems. The
one-time concept is based on the assumption that the composition of the detergent solution
can be optimized for a certain circuit. The solution is considered spent after having been
used once. In some cases it may however be used for pre-rinsing in a subsequent program
A simple sketch of cleaning-in-place

Tank Spray balls

Filter
Return pump

Clean water
Sanitizer at 60oC
Detergent

Thermometer

Outlet pump

Fig.18.5 Elements of a cleaning-in-place system (CIP)

The flow rate of the pump must produce a speed of circulation of 1.5 m/s (5 ft./sec.). Higher
speed may result in dead spots, while lower speeds, will not generate enough cleaning
action.
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In this system, which involves the use of detergents and sanitizers, it is possible to reuse
the detergent. The cleaning tank can be either fixed or movable, and pressure may vary
between 300 to 800 KPa (40 to 120 psi), when choosing the spray ball, it is important to
ensure that the surface is coated with the detergent, and that there is adequate cleaning
action. The outlet pump must operate at a high enough flow rate to prevent overfilling the
tanks during the course of the cleaning cycle.

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Chapter 19

Testing of milk and milk products

19.1 Platform testing of milk

The market milk quality mainly depends on the efficiency of platform testing of milk at the
collection points. The collection points are usually milk collection centers, milk chilling
centers, and the milk processing plants. Milk is an ideal substrate of microorganisms, if
there is any carelessness in testing the keeping quality of milks at these collection points,
there is possibility of spoilage of the whole lot only of one particular point.
It has therefore been an integral part of quality control exercised by dairy plant operator
almost all over the world. Although the Dairy Development Corporation has revealed to
have contained minimum quality control measures to be adopted at various points of
receiving milk. The different platform tests and laboratory quality control tests are:
Organoleptic tests, Sediment tests, Lactometer test, Alcohol (ethanol) tests, Fat tests,
Acidity tests, Clot-on-boiling test, MBRT, Direct microscopic test and Standard plate count
etc.
19.1.1 Organoleptic test
The acidic smell and taste tend to develop in milk by the microbial action which is
perceptible to senses of smell and taste. Good quality fresh milk has a mild sweet smell
and taste. A clear perceptible acidic smell and taste indicate that milk quality is doubtful and
be further tested before accepting and bulking it together.
a. Smell of the milk in the container immediately on removing the lid presence of any foul
or abnormal smell, other than the faintly sweet agreeable odor of good quality natural
milk, the sample should be kept aside for subjecting it to confirmatory tests.
b. Color of the milk- either the milk white color with faint golden yellow tinge for cow’s milk
or the same with vary pale blue tinge for buffalo’s milk are desirable.
The performance of the evaluations depends on the skill and experience of the person
doing the tastes.
19.1.2 Sediment test
This test indicates the presence of insoluble extraneous matter in milk. For this test filtering
the certain volume of milk through a filter paper disk and observe the kinds and amount of
such matter. Sediment test provides a simple, rapid and fairly quantitative measure to
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indicate the cleanliness of the milk with respect to visible dirt, which gained access to the
milk and has not been removed by single service strainers. However, the test has its
limitations as to the interpretation of the result and therefore should be cautiously applied.
The test is based on the principle of trapping visible dirt on a fixed area of a special filter
disc mounted to a suction type tester when a measured volume of milk has passed through
it.
19.1.3 Clot-on-boiling (COB) test
Introduction
This test gives an indisputable end point and the fact that it indicates not only the life of milk
due to souring but also the end of the life of milk when it is due to sweet curdling. Sweet
curdling means some organisms have the property of producing a rennin-like enzyme
which will curdle milk without the production of any acid, at least in the initial stage of
coagulation. The causative organisms may be spore formers such as Bacillus subtilis and
Bacillus cereus or non-spore forming rods belonging to the genus proteus. Various species
of cocci also sweet curdle the milk, the most organisms being Streptococcus liquifaciens.
Procedure
i. Pipette 5 ml of milk sample in to a clean, dry test tube.
ii. Heat the tube in a boiling water bath for 5 min or boil the milk over an open flame
(bunsen burner for a few seconds).
iii. Remove the tube from the bath after 5 min, gently tilt it and observe the inner sides of
the tube.
Observation
Observe the formation of floccules (small precipitated particles) on the inner side of the test
tube. The formation of the floccules (precipitated particles) is due the developed acidity.
Result interpretation
The precipitated particles (or clotting or coagulation) attached in the inner side of the test
tube will be a positive test and the milk should be rejected.
19.1.4 Alcohol test
The fresh milk is not precipitated by the addition of equal volume of 68 % ethyl alcohol
whereas souring milk, colostrums milk, bad mastitis contaminated milk (due to the diseases
of the udder) and sweet curdling milk, or when calcium and magnesium compounds are

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present in greater than normal amounts it will coagulate on the addition of alcohol. This fact
is the basis of the alcohol test which is provided a means of judging the quality of milk.
If the acidity of the milk is lower than 0.21 per cent, coagulation by alcohol is taken to
indicate a salt balance in the milk, which is unfavorable to processing.
Chemicals required
Ethanol- 68 % by weight or 75 % by volume having density 0.8675 g/ml at 27oC.
Procedure
i. Pipette 5 ml of milk sample in a clean, dry test tube.
ii. Add 5 ml of ethanol to the milk.
iii. The test tube mouth should be closed with suitable stopper and/or with the tip of the
thumb. Mix the contents of the test tube by inverting several times after closing.
iv. Note the presence of any flakes or clots, which denotes a positive test.
Interpretation of result
A negative test (do not form the clots) indicates a good heat stability of the milk sample. But
a positive test with a sample denotes poor heat stability of the milk sample which might be
unsuitable for processing in to condensed or sterilized milks.
19.1.5 Fat test (Volumetric)
The method used for rapid estimation of fat in fluid milk is known as “Gerber method”. It is
based on the principle of measuring the volume of fat released from the known volume of
the milk sample in a specially devised and accurately calibrated modified form of cylinder,
called “butyrometer” after adding to the milk requisite quantity of protein dissolving reagent
(Sulphuric acid) and a surfactant like amyl alcohol.
Principle
Digestion of the protein, particularly of the fat globule membranes, by H2SO4 separation of
the released fat by centrifugation (to ensure complete separation of the fat), and addition of
amyl alcohol or suitable substitute; results are in weight percentage.
Apparatus
a. Butyrometers with stoppers
b. Tilt measure (10 ml capacity) or automatic measure for Conc. H2SO4.
c. Tilt measure (1 ml capacity) for amyl alcohol.
d. Gerber centrifuge.

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 e. Water bath maintained at 65±2oC, provided for stand for butyrometers.
f. Pipette (10.75 ml) for milk at 27oC.
Reagents
a. Gerber Sulphuric acid: Conc. H2SO4 to be suitably diluted to give an acid with density of
1.807-1.812 g/ml at 27oC which corresponds to a concentration of H2SO4 90-91 % by
weight.
Preparation of Gerber H2SO4
Mix 910-900 ml of commercial conc. H2SO4 having sp.gr. of 1.84 at 20oC with 90-100 ml of
distilled water and cool in ice cold water bath. The sp.gr. of acid prepared should be
between 1.807-1.812 at 27oC. (Always add conc.acid to distilled water)
b.Amyl alcohol: The iso-amyl alcohol shall have a density of 0.803-0.805 g/ml at 27oC and
shall be clear, colorless, free from water. Furfural, acids and fatty matters.
Procedure
Transfer the 10 ml of H2SO4 in to the butyrometer, using 10 ml pipette or automatic
measure, keeping care not to wet the neck of the butyrometer with acid. Take out 10.75 ml
with specific pipette the properly mixed homogeneous milk sample of 27oC. Pour the milk to
the butyrometer containing H2SO4 carefully and slowly by touching the tip of the pipette jet
to the base of the neck of the butyrometer and slanting the pipette so that its delivery tube
rests on the lower end of the neck of the butyrometer. In this position the pipette stands at
45o angle to the verticle axis of the butyrometer. Holding the pipette thus, gently release the
index finger closing the tip of the pipette so that the milk flows gently along the inner wall of
the butyrometer and rests on top of the layer of H2SO4 without mixing with the acid, as far
as possible. Add 1 ml of amyl alcohol using 1 ml pipette or automatic measure and
following the procedure for addition of acid and milk so that the neck of the butyrometer
remains dry. Close the neck of the butyrometer firmly by inserting the stopper carefully so
as not to disturb the content inside. Carefully shake the contents of the butyrometer by
applying gentle swirling motion. When the contents are thoroughly mixed without any
charing, the curd is completely dissolved without leaving any suspended white particles.
Finally mix the contents again by gently inverting the butyrometer a few times. Transfer the
butyrometers to a water bath maintained at 65±2oC and allow it to stand for 5 min. Remove
the butyrometer from the water bath, dry it with a cloth and transfer it to the Gerber

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centrifuge. Centrifuge for 4 min at a maximum speed of 1100 rpm. After a centrifuge has
stopped gradually transfer the butyrometers from centrifuge with stopper downwards and
place again in to water bath at 65±2oC. Remove the butyrometer from the water bath in
between 3-10 min. Hold the butyrometer vertically with the graduated end upward and at
eye level. Take the reading of the butyrometer from the two markings on the scale which
correspond to lowest point of the clear fat meniscus and to the fat-acid interphase. Before
taking the butyrometer reading adjust the position of the fat column by manipulating the
stopper (either pushing in or drawing out) so that the lower end of the fat column is brought
within the graduation mark. The difference between the two scale readings gives the
percentage (by weight) of fat in the milk. Read the butyrometers to the nearest half of the
scale division, i.e., with a precision of 0.05 % fat.
19.1.6 Lactometer test (Total solids and solid-not-fat (SNF) test)
In routine analysis density of fluid milk is expressed in terms of sp.gr. compared with
density of water as unity at 15.56oC. The specific gravity of milk usually is determined with
a lactometer. It consists of a long, slender glass stem of uniform diameter connected to a
larger glass air chamber. This chamber causes the instrument to float. The lowest end of
the body is a glass bulb which is filled with lead shots or mercury which causes the
lactometer to sink to a proper level and also to float in an upright position in milk. The
lactometer is a hydrometer with a scale adapted to the limits of the specific gravity of milk.
To determine the specific gravity, bring the sample of milk to a temperature of very nearly
60oF (15.56o). For each degree Fahrenheit above 60oF (or 1.8oC above 15.56oC), 0.1 must
be added to the lactometer reading while for each degree below 60oF (1.8o below 15.56oC),
0.1 must be subtracted. The temperature correction is accurate only when applied within a
few degrees of the standard temperature. The reading corrected for temperature, is usually
designated as “corrected lactometer reading” (Corr. L) and is used for calculating either
specific gravity or total solids.
Corr . L
Specific gravity ( Sp. gr.) = +1
1000
No calculation is necessary if 1.0 is placed before the corrected lactometer reading.

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Calculation of total solids
In skim milk milk, the percentage of total solids is approximately one-fourth of the corrected
lactometer reading. However, in whole milk the fat causes and decreases in the lactometer
reading but an increase in the total solids. Furthermore, the percentage of solid-not-fat
increases about 0.2 for each 1 percent fat. Therefore, the “Babcock formula” has been
found to yield approximately correct results for estimating the percentage of total solids:
Corr. L
T.S.(%)= + (1.25× % fat) + 0.5
4
SNF (%) = Total solids(%) − Fat (%)
Corr . L
or, S .N .F . = + (0.25 × % fat ) + 0.5
4
Of the various types of lactometers in use the Quevene type is mostly followed. The
lactometer may or may not contain a built thermometer. The lactometer is usually
graduated at equal intervals from 15 at the top to 40 at the bottom, each division is being
known as lactometer degree.
Procedure
i. Warm the milk sample to about 40oC for 5 mins by dipping the container in hot water.
ii. Remove the container and mix the milk gently but thoroughly by inverting and rotating
the container (bottle), taking care to avoid frothing.
iii. Cool the samples to a temperature close to that of calibration of the lactometer; in any
case see that the sample is at a temperature within the range shown in the correction
table when the actual reading is taken.
iv. Poor gently the milk in to the cylinder avoiding formation of air bubbles; adjust the
quantity of milk in the cylinder so as to cause a slight overflow when the lactometer is
inserted.
v. Lower the lactometer into the milk as far as the graduation 28-29; it is of utmost
importance that the lactometer should float freely and should not touch the walls of the
cylinder.
vi. Take the lactometer reading carefully avoiding parallax; the reading should correspond
to the graduation in contact with the horizontal portion of the milk surface. (Wait for
some time for taking reading).
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vii. Record the temperature of milk by gently flapping the top of the stem, allow it to come
to rest and take a second reading; take the average of two readings.
The correction factor from variation in temperature and obtain the corrected reading.

Temp. Fat percent of sample

o
C 0 2 4 6 8
19.0 -2.2 -2.4 -2.6 -2.7 -2.9
19.5 -2.1 -2.3 -2.4 -2.6 -2.7
20.0 -2.0 -2.1 -2.2 -2.4 -2.5
20.5 -1.8 -2.0 -2.1 -2.2 -2.3
21.0 -1.7 -1.8 -1.9 -2.0 -2.2
21.5 -1.5 -1.7 -1.7 -1.9 -2.0
22.0 -1.4 -1.5 -1.6 -1.7 -1.8
22.5 -1.3 -1.4 -1.4 -1.5 -1.6
23.0 -1.1 -1.2 -1.3 -1.4 -1.4
23.5 -1.0 -1.1 -1.1 -1.2 -1.3
24.0 -0.8 -0.9 -1.0 -1.0 -1.1
24.5 -0.7 -0.8 -0.8 -0.9 -0.9
25.0 -0.6 -0.6 -0.6 -0.7 -0.7
25.5 -0.4 -0.5 -0.5 -0.5 -0.5
26.0 -0.3 -0.3 -0.3 -0.3 -0.4
26.5 -0.1 -0.2 -0.2 -0.2 -0.2
27.0 0 0 0 0 0
27.5 +0.1 +0.2 +0.2 +0.2 +0.2
28.0 +0.3 +0.3 +0.3 +0.3 +0.4
28.5 +0.4 +0.5 +0.5 +0.5 +0.5
29.0 +0.6 +0.6 +0.6 +0.7 +0.7
29.5 +0.7 +0.8 +0.8 +0.9 +0.9
30.0 +0.8 +0.9 +0.9 +0.0 +0.1
30.5 +1.0 +1.1 +1.1 +1.2 +1.3
31.0 +1.1 +1.2 +1.2 +1.4 +1.4
31.5 +1.3 +1.4 +1.4 +1.5 +1.6
32.0 +1.4 +1.5 +1.5 +1.7 +1.8
32.5 +1.5 +1.7 +1.7 +1.9 +2.0
33.0 +1.7 +1.8 +1.8 +2.0 +2.2
33.5 +1.8 +2.0 +2.0 +2.2 +2.3
34.0 +2.0 +2.1 +2.1 +2.4 +2.5
34.5 +2.1 +2.3 +2.3 +2.6 +2.7
35.0 +2.2 +2.4 +2.6 +2.7 +2.9
Table19.1 Correction to be applied to lactometer readings at temperature other
than 27oC to obtain Lactometer reading of milk at 27oC.

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19.2 Chemical and enzyme test
19.2.1 Acidity test in milk
The acidity test is of great value in grading milk and cream. Many of the microorganisms
common to milk produce lactic acid from lactose. Many cream and milk buyers use the
acidity test as one of the indices of quality in the products they purchase. The acidity test is
not only index of quality in milk and cream. The odors and flavors are not detected by the
acidity test, but are readily noted by the senses of taste and smell. Acidity is expressed as
% lactic acid (1 ml of 0.1 N NaOH = 0.009 g of lactic acid). It can be put in formula as
follows.
ml of NaOH × N NaOH × 9
% of lactic acid =
Weight of the sample
To determine the total acidity (natural + developed) in milk.
Aim To determine the total acidity in milk.
Apparatus Burette, porcelain dish-white flat bottomed of approximately 100 ml capacity,
stirring rod, Pipette-10 ml and 1 ml.
Reagents N/9 sodium hydroxide solution, 0.5 % phenolphthalein indicator, 0.005 %
rosaniline acetate solution.
Principle
Fresh milk on keeping at room temperature for same time develops acidity due to bacterial
action. This along with the natural acidity could be measured by titrating a known volume of
milk with a standard alkali to the end point of an indicator like phenolphthalein.
Lactic acid reacts with sodium hydroxide as shown below:
CH3CH(OH)COOH + NaOH = CH3CH(OH)COONa + H2O
According to the reaction one molecular weight of sodium hydroxide neutralizes one
molecular weight of lactic acid.
Procedure
I. Mix thoroughly the sample of milk by gently pouring from one container to another,
avoiding incorporation of air bubbles.
II. Transfer 10 ml of milk sample with the help of bulb pipette to each of the two porcelain
basins (50-100 ml).
III. Dilute the samples with equal volumes of freshly boiled and cooled water.
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IV.Prepare a blank by adding 1 ml of rosaniline acetate solution to 10 ml milk in one of the
porcelain basins. Add 1 ml of phenolphthalein indicator to the other 10 ml milk sample.
V. Rapidly add 1 ml of 0.1N NaOH solution to milk containing phenolphthalein and continue
to add drop by drop until by comparison the color matches the pink tint of the blank.
Repeat the experiment to get concordant values.
Observation
Take the total volume of alkali delivered in ml.
Calculation
The titratable acidity is expressed as lactic acid equivalent per 100 ml of milk.
TA = 0.9 X V1 X N
Where, V1 = Volume in ml of the standard NaOH solution required for titration.
N = Actual normality of the NaOH solution.
TA = Titratable acidity.
Result
Acidity of the given sample expressed as percentage of lactic acid.
Note:
Through mixing should be done during titration. Titration should be carried out within 20 s.
Titration shall be made under diffused day light or under illumination from a daylight lamp.
19.2.2 Methylene blue reduction test (MBRT)
Introduction
It is known by several names, such as the “reductase”, “methylene blue”, and “MBRT”. The
test shows the comparative activity of the bacteria in milk, therefore, it is a rough indication
of the number of bacteria per milliliter. The MBRT is based on the fact that the color
imported to milk by the addition of a dye such as methylene blue will disappear. The
agencies responsible for the oxygen consumption are the bacteria. The greater the number
of bacteria in milk, the quicker will the oxygen be consumed, and in turn the sooner will the
color disappear. This test has lost much of its popularity because of its low correlation with
other bacterial procedures. This test is true particularly in those samples which show
extensive multiplication of the psychotropic species.
For carry out the test, definite quantities of MB solution are added to 10 ml of milk and the
samples held at a uniform temperature (37oC) until the blue color has disappeared. The

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milk which remains blue for the longest time is considered to be the best quality. For sweet
cream more concentrated methylene blue is used.
Principle
When a methylene blue solution is added to milk. The time taken for decolor-ization will
serve as an indirect and approximate indicator of the bacterial quality of the milk. The
sources of error involved by admixture of air and cream formation in the mixture are
eliminated by inverting the tubes every half hour.
Reagent
Sterile methylene blue solution prepared by dissolving 1 standard methylene blue tablet in
sterile water to a volume of 400 ml.
Apparatus
1. Sterile test tubes holding 20 or 40 ml
2. Sterile rubber stoppers
3. Sterile pipette for measuring 0.5 or 1.0 ml.
4. Rack for the test tubes.
5. Water bath for maintaining a constant temperature of 37oC ±0.5oC
Procedure
i. The milk to be tested is thoroughly mixed and 10 or 20 ml are measured in to the sterile
tube should hold twice the volume of milk employed for the test.
ii. To 10 ml of milk is added 0.5 ml (to 20 ml of milk 1.0 ml of the sterile methylene blue
solution.
iii. The tube containing the mixtures of methylene blue and milk are then stoppered,
inverted to complete mixture, and placed in a water bath maintained at 37oC.
iv. The tubes should be inspected every half hour. At each inspection every tube should be
inverted and the numbers of the tubes showing decolorization noted.
vi. The time required from immersion of tubes in water bath to decolorization indicates the
reduction time for the sample concerned. The decolorization is considered complete when
the entire column of milk except an upper layer of approx. 5 mm has been decolorized.

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Reading of test
Time for reduction of color Class Remarks
0 -1/2 h IV Very unsatisfactory
½-2h III Unsatisfactory
2 - 5.5 h II Less satisfactory
> 5.5 h I Satisfactory

19.2.3 Phosphatase test

Introduction
This test was introduced by Kay and Graham in 1933. This test is applied to dairy product
to determine whether pasteurization is done properly or not and also to detect the possible
addition of raw milk. A negative phosphatase reaction cannot be interpreted as an absolute
index of proper pasteurization.
The principle behind this test is that the alkaline phosphatase enzyme is raw milk liberates
phenol from phosphoric esters (disodium phenylphosphate) or phenolphthalein from a
phenolphthalein monophosphate substrate when the test are conducted at suitable
temperature and pH. The amount of phenol liberated from the substrate is proportional to
the activity of enzyme. The phosphatase test is now being applied to chocolate milk, butter,
ice cream, and cheese to determine if these products have been manufactured from
adequately pasteurized milk or cream.
19.2.3.1 Sharers method
Reagents
1. Phenyl-phosphate solution (2 tablets Ewos I in 50 ml carbonate buffer (Ewos solution I)
2. Phenol reagent [1 tablet di-brome-kinon-chlorimide (Ewos tablet II in 3 ml 93 % alcohol
(Ewos solution II)].
3. Chloroform saturated, distilled water (Ewos solution III).
The tablet tablet-solutions must be freshly prepared.
Principle Phosphatase
Phenyl phosphatase Phenol + Phosphate
+ Phenol reagent
Procedure Blue color

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1. 1 ml milk is pipetted with a clean pipette in a clean test tube. For comparison make also
the test on a milk heated to 80-85oC.
2. Add 5 ml chloroform saturated water.
3. Add 5 ml phenyl-phosphate solution.
4. Place the tubes in a water bath at 38oC for 1 h.
5. After incubation add 6 drops phenol reagent, shake and incubate again 15 min. at 38oC.
6. If a blue color develops, phosphatase enzyme is present.
19.2.3.2 “Phosphatest” paper strip method
1. Cut the plastbag in two.
2. The paper strip is marked with a pencil and half immersed in the milk and again placed
in the bag.
3. Hold the bag in tweezers and close it by melting near a flame.
4. Incubate at 37oC (e.g., in a pocket) for two hours.
5. If phosphatase enzyme is present in the milk a yellow color develops. With raw milk the
color develops in a few minutes.
The “phosphatest” strips shall be stored in a freezer.

Principle
Para-nitro phosphate Para-nitro-phenol + phosphate

Colorless Yellow

19.3 Freezing point test of milk

Milk has a quite constant freezing point nearly -0.55oC. It mainly depends on lactose and
chlorides, which are soluble in milk. The constancy is due to the fact that these corresponds
have inverse relation, it mean, when one rises in amount the other lowers and vice versa.
Whole milk (-0.544), cream (-0.545) and skim milk have their differences in composition,
even though the freezing point is very close i.e., -0.544. Therefore, freezing points depends
on the dissolved substances, mainly lactose and salts, but not on the composition. The milk
fat exists as coarse globules and the casein as colloids have no effect ion the freezing
point.

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Freezing point of milk may vary daily, seasonally, as well as according to bread, feed, and
other many factors. The freezing point of milk is related to that of the blood of the cow,
requiring two or three hours to approach equilibrium.
19.4 Microbiological tests
19.4.1 Direct microscopic count method
The method is developed by Dr. R.S. Breed so it is also called “Breed count Method”.
This method of counting bacteria in milk consists of examining a stained film of milk under a
compound microscope. It is much quicker and less expensive than the plate method and
individual organisms and leucocytes can be observed and studied. The apparatus required
consists of a microscope, pipettes, slides and stains.
A measured volume of milk (0.01 ml) is spread over an area of 1 sq.cm on a glass slide,
dried and stained. Bacteria and body cells (leucocytes) are stained selectively whereas the
milk solids are relatively unstained.
The examination of a few fields can indicate the quality of the milk-poor milk will show
many clumps of bacteria per field whereas high quality milk shows few or no bacteria per
field.
19.4.2 Standard plate count (SPC)
It consists in mixing a definite volume of milk with melted agar, allowing the agar to solidify
and then counting the colonies that appear upon incubation. A colony consists of the
progeny of one cell or group of cells, and the results are reported as so many colonies per
ml of milk. Agar is available commercially in dehydrated form. The composition of agar is as
follows.
Standard medium agar
Pancreatic digest of casein (USP) – 5 g
Yeast extract – 25 g
Glucose – 1 g
Agar bacteriological grade – 15 g
Distilled water – 1 g
Final pH – 7.0 at 25oC
19.5 Detection of adulterants in milk
Some of common adulterants in milks are:

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i. Partial removal of fat by skimming.
ii. Increasing the bulk of whole milk by addition of skim milk.
iii. Dilution by addition of water.
iv. Increasing the density of water milk by addition of starch, can sugar, glucose, gelatin
etc.
19.5.1 Detection of partial removal of fat by skimming
An indication of the removal of excess fat from milk is given by the following observations:
a. Lower percentage of fat.
b. Higher density reading of the sample at 27oC.
c. Higher ratio of solid-not-fat to fat.
19.5.2 Detection of addition of skim milk
When fresh separated milk or skim milk has been added to whole milk, it could be inferred
from the following observations:
a. Lower percentage of fat;
b. Higher density reading of the sample at 27oC.
c. Higher ratio of solids-not-fat to fat.
19.5.3 Dilution by adding water
Presence of extraneous water in milk is detected by the following observations:
a. Lower percentage of fat;
b. Lower density of milk at 27oC;
c. Lower percentage of solids-not-fat; and
d. Depression of freezing point of the milk sample.
19.5.4 Detection of compounds which increase density
Substances commonly misused to increase the density of normal milk through
incorporation in to milk are starch or other cereal products, cane sugar, glucose, gelatin,
urea, etc. Tests for detection of these extraneous compounds in milk are given.
19.5.4.1 Starch and cereal flours
Reagent
Iodine solution- 1 %. ( Dissolve 2.5 g KI in 100 ml water, add to it 1 g iodine crystals;
shake well to give a clear solution.

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Procedure
i. Take about 3 ml well mixed milk sample in a test tube.
ii. Heat the milk to just boiling by holding the tube over flame, and thereafter cool to room
temperature.
iii. Add 1-2 drops of 1 % iodine solution.
iv. Observe the development of color.
Interpretation
Formation of blue-violet color indicates presence of starch or cereal flours. This color
disappears on further heating and reappears on cooling.
19.5.4.2 Can sugar (Selivanoff’s Test)
Reagent
Selivanoff’s reagent- is prepared by dissolving 0.05 g resorcinol in 100 ml dil. (1:2) HCl
Procedure
i. Take about 3 ml of well mixed milk sample in a test tube.
ii. Add 5 ml of Selivanoff’s reagent.
iii. Place the tube in a boiling water bath for 5 mins (or gently heat over flame to boiling)
and observe the development of color.
Interpratation
Development of a red color with or without the separation of a brown red precipitate
indicates the presence of cane sugar in milk.
19.5.4.3 Glucose
Reagents
a. Barfoed’s reagent (Tauber and Keleiner modification)- Dissolve 24 g cupric acetate in
450 ml boiling water and immediately add 25 ml of 8.5 % lactic acid to the hot
solution. Shake to dissolve almost all precipitate, cool and dilute with water to 500 ml.
If necessary decant or filter to get a clear solution.
b. Phosphomolybdic acid reagent – take 35 g ammonium molybdate and 5 g sodium
tungstate in a large beaker; add 200 ml of 10 % NaOH solution and 200 ml water.
Boil vigorously (20-60 mins) so as to remove nearly whole of ammonia. Cool, dilute
with water to about 350 ml. Add 125 ml conc. H3PO4(85 %) and dilute further to 500
ml.

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 Procedure
e. Take 1 ml of milk sample in a test tube.
i. Add 1 ml of modified Barfoed’s reagent.
ii. Heat the mixture for exact 3 mins in a boiling water bath and then rapidly cool under
tap water.
iii. Add 1 ml of Phosphomolybdic acid reagent to the turbid solution and observe the
color.
Interpretation Immediate formation of deep blue color indicates the presence of added
glucose. In case of pure milk only faint bluish color is formed due to dilution of Barfoed’s
reagent.
19.5.4.4 Gelatin
Reagents
a. Acid mercuric nitrate solution (or Millon’s reagent)- is prepared by dissolving 1 part of
metallic mercury in 2 parts of conc. HNO3 (sp.gr. 1.42) and diluting solution with 2
volumes of water.
Procedure
i. Take 10 ml of the milk sample in an Erlenmeyer and add 10 ml acid Hg(NO3)2 solution.
ii. Shake the mixture, add 20 ml water, shake again. Let stand for 5 mins and filter (with
milk sample, having high concentration of gelatin filtrate will be opalescent only, and no
clear filtrate can be obtained).
iii. Take a portion of the filtrate in a test tube and add equal volume of saturated aqueous
picric solution. Observe the change.
Interpretation
Formation of yellow precipitate indicates presence of gelatin in large concentration,
while yellow turbidity shows the presence of gelatin.
19.5.4.5 Urea
Reagents
a. Sodium acetate- acetic acid buffer- 1 N, pH 4.75,
b. Trichloroacetic acid (TCA)-24 %
c. Sodium hydroxide solution – 2 %
d. Sodium hypochloride solution – 2 %

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 Prepare fresh by passing chlorine gas into 2 %, NaOH solution and standardize to 2 %
available chlorine by usual idometric titration.
e. Phenol solution – 5 % - Prepare from almost colorless, reagent quality redistilled
phenol.
Procedure
i. Take 5 ml of milk sample in a 50 ml Erlenmeyer flask, add 5 ml sodium acetate acid
buffer or TCA solution and heat for 3 mins in boiling water bath using a stop watch (no
heating is required in case of TCA being used). Filter the ppt (whatman no 42 filter or
equivalent) and collect 1 ml of the filtrate in a test tube.
ii. To the filter add 1 ml NaOH solution followed by 0.5 ml sodium hypochloride solution,
mix thoroughly and finally add 0.5 ml phenol solution.
Interpretation
Formation of a characteristic blue or bluish green color indicates the presence of
extraneous urea in the milk sample. Filtrate from unadulterated milk remains colorles. The
blue color remains stable for at least 12 hrs. By this test as low as 0.1 % urea added to milk
can be detected.
19.5.4.6 Ammonium sulphate
Reagents
Same as under urea.
Procedure
Take 1 ml milk sample in a clean test tube, add 0.5 ml NaOH solution followed by 0.5 ml
sodium hypochloride solution and mix thoroughly. Add to this mixture 0.5 ml phenol solution
and heat for 20 seconds in boiling water bath (using a stop watch).
Interpretation
A bluish color immediately forms , which turns to deep blue afterwards, if the milk contains
extraneous ammonium sulphate added. The color remains stable for 12 hrs. In case of pure
milk only a salmon pink color is formed, which gradually changes to bluish in course of
about 2 hrs. Detection limit is 0.1 % added ammonium sulphate.
19.6 Determination of preservatives
In fluid milk preservation no any kind of preservatives can be used. The food law does not
permit to add preservatives for extending its self life. The officially permitted preservatives

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only in case of fluid samples which have to be stored for chemical examination. The
chemical preservatives commonly used to milk sample to prolong self life are benzoic acid,
salicyclic acid, boric acid and borates, formaldehyde, hydrogen peroxide, mercuric chloride
and potassium dichromate. The chlorine compounds and antibiotics are not used for
preservation of sample because of remaining residues these may create problems in
microbiological analysis of the sample. Some of the qualitative for the detection of such
preservatives are as follows.
19.6.1 Formalin
Formalin is a 40 % solution of formaldehyde will also detect addition of formalin as
preservative to milk.
19.6.1.1 Hehner test
Procedure
i. Take about 5 ml of the milk sample in a test tube.
ii. Add gently down the side of the test tube 2 ml of conc. H2SO4 containing a trace of
FeCl3 (the added acid should form the bottom layer of the mixture without mixing with
the milk).
iii. Observe the color of the ring formed at the junction of the two liquids – formation
of a violet to purple color indicates the presence of formaldehyde.
19.6.1.2 Hehner-fulton test
Procedure
i. To 10 ml cold conc. H2SO4 add in small portions equal volume of saturated bromine
water, cooling the mixture all the time.
ii. To 6 ml conc. H2SO4 add slowly with cooling 5 ml of distillate from milk prepared
thus: dilute 100 ml of milk with 100 ml water, acidify with a small quantity of H3PO4
and steam distil the mixture (or distil from a Kjeldhal flask with condenser through
trap)and collect 50 ml.
iii. Place 5 ml of the mixture under (ii) in a test tube, add slowly keeping the tube in a
cooling mixture (ice water bath) 1 ml aldehyde free milk followed by 0.5 ml oxidizing
solution of Br2 + H2SO4 (No.1) . Mix gently and observe the color.
iv. A purplish pink color indicates the presence of formaldehyde.

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19.6.2 Hydrogen peroxide (H2O2)
19.6.2.1 Paraphenylenediamine test
Procedure
a. Take 5 ml of milk sample in a test tube; add equal volume of fresh normal milk.
b. Add immediately to the mixture 5 drops of 2 % paraphenylenediamine aqueous
solution (freshly prepared) and mix gently.
c. Formation of deep blue color indicates the presence of H2O2.
19.6.2.2 Vanadium pentoxide (V2O5) test
Procedure
I. Prepare the reagent by dissolving 1 g vanadium pentoxide in 100 ml H2SO4 (6 vol.
H2SO4 + 94 volume water).
II. Take 10 ml sample in a porcelain basin.
III. Add 10-20 drops of V2O5 reagent to the milk, and mix gently with a glass rod.
IV. Formation of pink to red color indicates the presence of H2O2.
19.7 Detection of neutralizers
Alkali in various forms like sodium carbonates, sodium bicarbonate, sodium hydroxide and
lime are used to neutralize developed acidity in milk, although the practice is not permitted
by Public health laws. Detection of such neutralizers can be made by the following two
methods.
19.7.1 Rosalic acid test
Reagents
a. Ethanol – 90 % or rectified spirit.
b. Rosalic acid solution – 1 % in ethanol.
Procedure
I. Take in a test tube about 5 ml milk and mix with 5 ml ethanol followed by 2-3 drops of
rosalic acid solution- formation of rose red coloration indicates the presence of weak
alkali as neutralizer. Pure milk produces a brownish coloration only.
19.7.2 Ash alkalinity test
Reagents
a. Hydrochloric acid – standard, 0.1 N,
b. Phenolphthalein indicator.
Apparatus

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 a. Muffle furnace.
Procedure
I. Pipette 20 ml of milk into a porcelain basin and evaporate to dryness on boiling water
bath.
II. Remove the basin, cool to room temperature and ignite the residue by heating over
Bunsen flame (or in a muffle furnace) until grey-white ash is obtained.
III. Cool the basin to room temperature. Add to the residue 10 ml of water and disperse
the ash in water by stirring with a glass rod.
IV. Titrate the ash dispersate by standard HCl using phenolphthalein indicator – if the
volume of 0.1 N HCl required to neutralize the ash dispersate exceeds 1.20 ml the
milk is suspected to contain neutralizers.

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Chapter 20
Organization in Dairy

20.1 Organizational chart of dairy development corporation Central office,

Lainchour

Chairman of Managing committee

General Manager

KMSS Deputy General manager Deputy General manager

Administration Technical Appraisal and monitoring
HMSS section

Technical Management Quality control, Research

Administration Department
Department and Development
BMSS
Finance Department Training and Employment Internal Auditing
LMSS Department

MPP&SDS

Note:
KMSS = Kathmandu Milk Supply Scheme, Balaju
HMSS = Hetauda Milk Supply Scheme, Hetauda Industrial Estate, Hetauda
BMSS = Biratnagar Milk Supply Scheme
LMSS = Lumbini Milk Supply Scheme
MPP & SDS = Milk Product Production and Sales Distribution Scheme
20.2 Organizational chart of Biratnagar Milk Supply Scheme, Biratnagar
Project Manager

Assistant Quality Control Marketing management Computer Section

Project Manager
Chemical and Microbiological Sales Management
Employee Division laboratory
Administration section Sales & supply booth
General Section
Budget implementation
Accounts section Reconcillation & statement General store
Store Maintainance
Milk & milk product production Milk products store
Processing section
Milk powder production
Collection section Chilling center
Machine maintenance
Engineering section
Vehicles & other maintenance

356
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