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Refrigerated chicken meat freshness. Correlation between easily hydrolisable

nitrogen, ph value and biogenic amine contents

Article · July 2002

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 The Annals of the University Dunarea de Jos of Galati
Fascicle VI – Food Technology, New Series Year II (XXXI)

REFRIGERATED CHICKEN MEAT FRESHNESS. CORRELATION BETWEEN EASILY

HYDROLISABLE NITROGEN, pH VALUE AND BIOGENIC AMINE CONTENTS

Octavian BASTON*, Ioan TOFAN*, Alexandru Lucian Stroia**,

Daniela MOISE**, Octavian BARNA*

* Dunarea de Jos University of Galati, Faculty of Food Science and Engineering, 111,
Domeasca St., Tel./Fax: +40 236 460165
**Timişoara Mihai Eminescu University, 19, Aries St., Tel: +40 256 466360
**Institute for Research-Development of the Horticultural Products Marketing
and Industrialization – Horting, Bucharest, 1A, Intrarea Binelui St., Tel: +40 21 4610706

Received 24 June - Accepted 10 August

Abstract
In this study we evaluated the variation of highly hydrolysable nitrogen, pH and the biogenic amines from
the refrigerated chicken meat and we calculated the freshness indices based on biogenic amine content.
We correlated the obtained results (pH and ammonia content) with Romanian legislation in order to
determine the storage time limit for the refrigerated chicken that can be acceptable for consumption.
Finally we determined the freshness indices proposed by different authors for the refrigerated chicken
meat based on the biogenic amine content. The meat pH increased constantly from the first (5.92) up to
the 20th day of storage in refrigerated state (7.33). Also, the easily hydrolysable nitrogen increased from
20.5 to 39.68mg NH3/100g. From the biogenic amine content point of view, the first day spermine is
preponderant (50% of the total amines). The 20th day, cadaverine is the preponderant amine followed by
putrescin (17%) and β-phenylethylamine (13%). Based on the biogenic amine content, three freshness
indices (FI) were calculated as per the relationships proposed by: Mietz and Karmas, Veciana-Nogues
et.al., and by Silva and Gloria.
Correlating the obtained results for easily hydrolysable nitrogen, pH and Romanian legislation norms,
we determined that the maximum freshness limit for chicken meat is at the 3rd day of storage, hereby FI
are as follows: FI (Mietz and Karmas) = 0.27, FI (Veciana-Nogues) = 12.2, FI (Silva and Gloria) = 0.24.
Keywords: biogenic amines, spoilage, quality index, freshness index, chicken meat, refrigerated meat,
pH, easily hydrolysable nitrogen.

1. Introduction purpose. In order to obtain products with high

conservation durability and to increase the
The refrigerated chicken meat spoilage when stored refrigeration effect, it is necessary to have as less
for a long period is due to the microorganism action initial microbial load as possible (Tofan, 2005).
and the biochemical transformations inside the
Initially, chicken meat quality was evaluated by
product. If the refrigerating chain from producer to
determination of microbiological and sensorial
consumer is not ensured, or if the seller overpasses
attributes. For the identification of the early signs of
the shelf life, the consumer can have an unpleasant
meat alteration, some chemical indices were
surprise of buying an altered product. After chicken
proposed: volatile nitrogen basis, composites
slaughter, the muscular tissue suffers irreversible
resulted after breaking the nucleotides, volatile
physical, chemical and biochemical transformations
acidity and the biogenic amine content (Halsz et.al,
which determine the muscle to convert in meat. The
1994).
microbial spoilage processes occurr later.
Microorganism activity is reduced by using The biogenic amine occurrence is a consequence of
refrigeration temperatures for meat conservation the enzymatic decarboxylation of the precursor

Corresponding author: octavian.baston@ugal.ro

37
 Baston et.al. / The Annals of the University Dunarea de Jos of Galati
Fascicle VI – Food Technology, New Series, II (XXXI), 2008, 37-43

aminoacids because of the microorganism activities. Romanian Recommendation Norm 24/01/2005

Polyamines: spermine and spermidine are natural (***, 2005).
amines produced by the body. The biogenic amines:
The samples were analysed the first day when the
putrescin, cadaverine, histamine, tyramine,
meat was received, recorded as day 1, then the 3rd,
tryptamine, β-phenylethylamine can be formed 5th, 7th, 13th, and 20th day. The dry matter
when storing the chicken meat due to microorganism determination was done according to STAS 9065/3-
action. The biogenic amine determination is 73. The pH was measured using a standardized
important not only because of their toxicity but also method (STAS 9065/8-74) with a WTW Ino Lab pH
their potential use as freshness indicators 730 pH-meter.
(Balamatsia et.al, 2006)
The easily hydrolysable nitrogen was determined
Different authors’ studies regarding the refrigerated according with STAS 9065/7-74. The purpose was
chicken meat showed that some of the previously to determine the ammonia quantity that is formed in
mentioned biogenic amine concentrations are the product after protein degradation by the spoilage
increasing in time, while others are decreasing microorganism activity.
during storage (Vinci et Antonelli, 2002, Apostolos
et.al, Sarinen et.al, 2002, Balamatsia et.al, 2006, The measurement of biogenic amines content using
2007). The freshness index of meat calculated on the high performance liquid chromatography, was
basis of biogenic amines is a topic to be debated. performed according to the method proposed by
The occurrence of these amines is dependant on Food Research Institute from Helsinki, Finland
different factors that vary in time. The microbial (Eerola et.al, 2001). All the reagents used were
population influences the profile of biogenic amines. analytic pure, for HPLC use. Te water used was
Spoilage responsible microorganisms might not have deionised. The necessary reagents were purchased
the capacity of amine forming. It is difficult to from the Merck and Sigma-Aldrich companies.
establish quality limits universally accepted based Installations and equipment used for biogenic amine
on the biogenic amine content. The above- determination: Philips 7768 food processor,
mentioned reasons are partially justifying the homogenisation device 7011S, Kern 770-60
relative dispersal of biogenic amine values for meat, analytical balance, Silent CrusherM homogenisation
in various researches. From a practical point of device, centrifuge EBA 21, filter paper for quick
view, the relative simplicity and quickness filtering with 55 mm diameter, syringe filters with
identification and quantification of the biogenic porosity of 0,45 µm and 13 mm diameter, Heidolph
amines (compared to the micro-biological
REAX control agitator, ultrasonic water tank
measurement) besides the economical advantages
Aquawave TM, incubator BMT INCUCELL 55,
(for example the quick test for determining the
water deionising system EASY pure RoDi, filtering
diamines described by Hall et.al, 1999), are reasons
assembly with vacuum pump. The device for the
for using these substances as chemical indices for
HPLC determination was a liquid chromatograph
animal origin product freshness.
model SURVEYOR produced by Thermo Electron
The purpose of the study is evaluation of company, configured with detector model PDA
refrigerated chicken meat freshness using biogenic PLUS DETECTOR, auto-sampler model
amine index. AUTOSAMPLER PLUS, pump model LC PUMP
PLUS and detector UV-VIS. Chromatography
2. Materials and Methods column is type BDS Hipersyl C18.
The biogenic amines quantification: quantitative
The chicken carcases were purchased from the
measurement was performed depending on the
Agricola International Bacau company
internal standard using peaks for each biogenic
slaughterhouse. The meat was analysed after
amine. The 254nm wavelength absorbance was
cooling, packaging and transportation from the plant
measured and the resulted peaks were integrated
the first day after slaughter. The carcases were
with CromQuest software. The concentration of
stored aerobically for 20 days at a temperature of
each biogenic amine was expressed in mg/kg.
4±1°C in the refrigerator. The refrigerator used is
Electrolux ENB43691S. The carcases weight varied The statistical analysis of the obtained data was
between 1.2÷1.5 kg. Sampling was done as per done using SPSS 13 software for 10 samples in each

38
 The Annals of the University Dunarea de Jos of Galati
Fascicle VI – Food Technology, New Series Year II (XXXI)

of the storage days. The results obtained are value is not yet constant, it decreases within the first
presented as the mean ± standard deviation (SD). few hours and then increases in time (Debut et.al,
The standard deviation is a measure of the 2003, Duclos et.al, 2007). During the ageing
dispersion of outcomes around the mean. The process, the pH rises, due to proteolysis.
differences among means were determined using the The sampling has an important influence concerning
method of the smallest squares and the significance the results because the global pH was influenced by
level was p< 0.05. the skin pH, meat/skin ratio and the type of meat
from the anatomical parts where the sampling was
3. Results and Discussion performed.
Only the pH value as a freshness indicator is
3.1. Evaluation of the pH during chicken meat controversial because its value varies depending on
storage many factors. Yet, according to this factor, and in
concordance with Romanian regulations
The pH values are shown in table 2. We obtained an (pH=5.8…6.0), good quality fresh meat is up to the
initial pH value of 5.82, that is fitting well in the 3rd day of storage.
limits of 5.8…6, which indicates that the product is
fresh in accordance with Romanian Directive 86 3.2. Evaluation of easily hydrolysable nitrogen
(***, 2002). In time, it can be noticed that the content of refrigerated chicken meat
chicken meat pH increases. Starting with the 3rd day,
the meat pH is over 6.0, meaning that it starts In Table 3, there are shown the easily hydrolysable
spoiling. The spoilage continues up to the last day of nitrogen values in the refrigerated chicken at 4ºC for
measurements. 20 days.
The pH increase is due to the bio-chemical reactions In Table 3, it can be noticed that the first day of
in post-mortem chicken meat. There are many evaluation, the easily hydrolysable nitrogen value is
factors that influenced the pH initial chicken meat 20.5 mgNH3/100g, increasing up to the 20th day to
value. The chicken meat was evaluated the second 39.685 mgNH3/100g.
day after slaughter. The first day of slaughter the pH

Table 1. Operating conditions of HPLC instalation

Gradient Wave Column Column Sample room Injected
Time, Flow,
Ammonia Nitrile length, pressure, temperature, temperature, sample
min ml/min
acetate, % acetate, % nm bar ºC ºC volume, µl
0.01 40 60
15 40 60
20 30 70 1.00 254 min. 70 40 7 20
25 5 95
30 40 60
Table 2. pH variation of chicken meat stored at refrigerated conditions
Storage time (days)
1 3 5 7 13 20
pH value Mean SD Mean SD Mean SD Mean SD Mean SD Mean SD
5.82 0.1 6.05 0.08 6.41 0.09 6.52 0.08 6.81 0.06 7.33 0.19
SD - standard deviation of 10 determinations.

Table 3. Easily hydrolysable nitrogen content variation during storage of refrigerated chicken
Easily Storage time (days)
hydrolysable 1 3 5 7 13 20
nitrogen Mean SD Mean SD Mean SD Mean SD Mean SD Mean SD
content, 20.5 1.26 22.2 0.68 24.9 1.11 25.7 0.87 33.7 0.75 39.68 0.40
mg NH3/100 g
SD - standard deviation of 10 determinations.

39
 Baston et.al. / The Annals of the University Dunarea de Jos of Galati
Fascicle VI – Food Technology, New Series, II (XXXI), 2008, 37-43

Correlating the obtained values with the imposed but during the first 7 days has the highest values
limits by the Romanian Recommendation Norm compared with the other amines.
(***, 2002) of 25 mgNH3/100g, the 5th day of Calculating each amine ratio out of the total amount
storage refrigerated chicken meat is considered to be (table 5), it can be noticed that from the first day up
fresh. The accumulation in time of easily to the 7th day spermine was the dominant amine, its
hydrolysable nitrogen is due to protein hydrolysis ratio decreasing gradually (from 50% down to
catalysed by meat enzymes. Also, microorganisms 38%). The first day, cadaverine, putrescin and
with proteolytical activity can act on proteins histamine had the lowest content out of the total
transforming them into smaller compounds, such as amines. The 20th day, tryptamine, serotonin and
free aminoacids. The aminoacids can suffer histamine had the lowest level of amines. The first
oxidative deamination, decarboxylation and day, spermine had the highest percentage of the
desulphuration resulting gases as ammonia, carbon biogenic amines while during the 20th day the first
dioxide, hydrogen sulphide etc. Meat itself contains place belongs to cadaverine followed by putrescin
free aminoacids or they can occur from proteins by and phenylethylamine. These three amines have a
hydrolysis. Subsequently, they can be degraded negative influence on the food product odour.
partially or totally to simple compounds as CO2,
The biogenic amine occurrence is mainly a
H2O, NH3, H2S etc. Also, easily hydrolysable
consequence of the activity of decarboxylases,
nitrogen in food Romanian legislation is a very
enzymes produced by microorganisms. These
important marker for raw meat freshness.
enzymes act on the free aminoacids or on the
aminoacids resulted from protein hydrolysis.
3.3. Biogenic amine accumulation in refrigerated
Polyamine spermine and spermidine are amines
chicken meat
existing in the body and are naturally produced by
it. The biogenic amines: putrescin, cadaverine,
Table 4 shows the biogenic amine content,
histamine, tyramine, tryptamine,β-phenylethylamine
measured from the raw chicken meat, refrigerated
are formed during storage of chicken meat due to
and stored for 20 days.
microorganism activity. The decrease in time of
In the table it can be noticed that: the first day of
spermidine and spermine is due to their use as
measurement of the biogenic amines, cadaverine
nitrogen sources by microorganisms (Balamatsia
and putrescin were not detected in any of the
et.al, 2006, 2007).
analysed samples. Tryptamine content is increasing
in time having a small variation during the 3rd day of
3.4. Raw chicken meat freshness index evaluation
storage. Phenylethylamine is increasing in time,
mainly after the 7th storage day the quantity is highly
In a series of researches, the limits of freshness meat
increasing (approximately threefold after the first
food products were determined with index
day). Putrescin has a similar variation as the
calculated based on the biogenic amine content. The
phenylethylamine. Cadaverine is increasing in time
researchers proposed some biogenic amines as an
and after the 7th day has a very high increase from
index or freshness index based on biogenic amine
3.15 mg/kg (day 7) up to 30.07 mg/kg (day 13) and
content for meat evaluation (Apostolos et.al, 2006,
57.98 mg/kg in the 20th day. Histamine during the
Balamatsia et.al, 2006, 2007, Silva et. Gloria, 2002).
first day of storage has the lowest content of the all
studied biogenic amines in chicken meat. It The advantage is the small concentration of biogenic
increases slowly in time; the 20th day the quantity is amines that can be detected with HPLC long before
6.1 mg/kg. Serotonin is increasing in time, thus its they can be sensorially identified especially by
accumulation is low: 5,16mg/kg during the first day smell. That’s why biogenic amines can be chemical
and 7.93 mg/kg during the 20th day. Initially, indicators of meat spoilage, and therefore can be
tyramine is not so high (2.99 mg/kg) as compared used for evaluation of the freshness status of the
with the other biogenic amines. However the animal origin products. Initially, those freshness
concentrations increase in time up to 13.67 mg/kg. indicators were used on fish meat.
Spermidine values are limited approximately
between 5 and 6 mg/kg for all the 20 days of
chicken meat storage. Spermine decreases in time,

40
 The Annals of the University Dunarea de Jos of Galati
Fascicle VI – Food Technology, New Series Year II (XXXI)

Table 4. The biogenic amine content variation during storage of refrigerated chicken
Biogenic amine Storage time (days)
content, mg/kg 1 3 5 7 13 20
Mean SD Mean SD Mean SD Mean SD Mean SD Mean SD
Tryptamine 4.04 0.88 3.98 0.56 5.40 1.21 5.94 1.17 6.50 1.89 10.34 0.85
Phenylethylamine 4.19 0.36 4.85 0.90 5.21 2.64 5.40 3.39 17.62 1.36 23.14 3.48
Putrescin nd 0.00 3.05 4.86 3.19 0.19 3.47 2.63 16.67 7.72 28.84 10.59
Cadaverine nd 0.00 2.74 3.63 2.75 0.79 3.15 2.23 30.07 10.47 57.98 12.33
Histamine 2.36 0.12 2.64 0.78 2.39 0.81 3.00 0.84 2.91 0.13 6.10 0.44
Serotonin 5.16 0.84 6.57 1.98 6.50 1.54 6.72 0.46 7.65 1.23 7.93 0.28
Tyramine 2.99 1.52 3.79 1.82 3.24 1.80 3.93 1.57 8.08 2.84 13.67 2.12
Spermidine 4.84 0.33 5.67 1.15 4.89 2.91 6.01 1.10 5.92 1.72 5.68 2.38
Spermine 23.01 2.55 24.00 2.93 23.55 1.42 22.94 1.36 20.52 1.10 17.93 3.74
nd-not detected, SD - standard deviation (10 determinations).

Table 5. Storage variation percentage of each biogenic amine from the total content
Storage time (days)
Biogenic amine content, %
1 3 5 7 13 20
Tryptamine 9 7 9 10 6 6
Phenylethylamine 9 8 9 9 15 13
Putrescin 0 5 6 6 14 17
Cadaverine 0 5 5 5 25 34
Histamine 5 5 4 5 3 4
Serotonin 11 11 11 11 7 5
Tyramine 6 7 6 6 7 8
Spermidine 10 10 9 10 5 3
Spermine 50 42 41 38 18 10

The freshness index (FI) most used in the literature

4,00
4.0
Karmas index va

for refrigerated chicken meat are:

Karmas index value

3,00
3.0
histamine + cadaverine + putrescin
a) , proposed 2,00
2.0
1 + spermine + spermidine
1,00
1.0
by Mietz and Karmas (Balamatsia et.al, 2006, Silva
et Gloria, 2002) 0,00
0.0
b) Cadaverine + Putrescin + Tyramine + Histamine, 0 2 4 6 8 10 12 14 16 18 20
expressed in mg/kg, and proposed by Veciana- Storage time (days)
Nogues and other (Balamatsia et.al, 2006, Silva and Figure 1. Mietz and Karmas freshness index variation for
Gloria, 2002) refrigerated chicken meat
spermidine
c) proposed by Silva and Gloria (Silva
spermine The refrigerated meat freshness is reduced in time as
et Gloria, 2002) a result of biochemical, physico-chemical and
We made the necessary calculations for the microbiological transformations. The loss of
freshness index expressions mentioned above in freshness indicates that meat has started spoiling.
order to establish the variation curve for the
refrigerated chicken meat. The results are presented 3.5. Correlating the experimental values and
in figure 1, figure 2 and figure 3. establishing the freshness limit

We know that meat freshness is a quality indicator Based on the performed chemical measurement, and
that decreases in time because of meat spoilage. taking into account the limit imposed by the

41
 Baston et.al. / The Annals of the University Dunarea de Jos of Galati
Fascicle VI – Food Technology, New Series, II (XXXI), 2008, 37-43

Romanian standards regarding pH and the easily Correlating the pH variation and the easily
hydrolysable nitrogen, we can asses that the 3rd day hydrolysable nitrogen with the quality indices
is the maximum limit for the refrigerated chicken measured based on the biogenic amines, we
meat best quality (freshness). consider that all three freshness indices can be used
as quality indicators for the refrigerated chicken
120.00
120.0 meat.
100.00
100.0
References
Veciana index value
Veciana index va

80.00
80.0
Apostolos, P. Chouliara, I. Paleologos, EK. Savvaidis, I.
60.00
60.0 Kontominas, M., 2006, Relation of biogenic amines
40.00
to microbial and sensory changes of precooked
40.0 chicken meat stored aerobically and under modified
20.00 atmosphere packaging at 4 C, European Food Res.
20.0 and Tech., 223, 5, 683-689.
0.00
0.0 Balamatsia, C.C. Paleologos, EK. Kontominas, MG.
0 2 4 6 8 10 12 14 16 18 20
Savvaidis, IN., 2006, Correlation between microbial
Storage time (days) flora, sensory changes and biogenic amines
F formation in fresh chicken meat stored aerobically or
igure 2. Veciana-Nogues freshness index variation under modified atmosphere packaging at 4C:
for refrigerated chicken meat possible role of biogenic amines as spoilage
indicators, Antonie van Leeuwenhoek, Springerlink,
89, 9-17.
Sliva and Gloria index value

0.35
0.35
0.30
0.30 Balamatsia, C.C. Paleologos, EK. Kontominas, MG.
Index value S&

0.25
0.25 Savvaidis, IN., 2007, Possible role of volatile amines
0.20
0.20 as quality indicating metabolites in modified
0.15
0.15 atmosphere packaged chicken fillets: Correlation
0.10
0.10 with microbiological and sensorial attributes, Food
0.05 Chemistry, 104, 4, 1622-1628.
0.05
0.00
0.00 Debut, M. Berri, C. Baeza, E. Sellier, N. Arnould, C.
0 2 4 6 8 10 12 14 16 18 20 Guemene, D. Jehl, N. Boutten, B. Jego, Y.
Storage time (days) Beaumont, C. and Le Bihan-Duval, E., 2003 ,
Variation of chicken technological meat quality in
relation to genotype and preslaughter stress
Figure 3. Silva and Gloria freshness index variation for
conditions, Poultry Science, 82, 12, 1829-1838.
refrigerated chicken meat
Duclos, M. J. Berri, C. and Le Bihan-Duval, E., 2007,
Correlated with the biogenic amine content Muscle Growth and Meat Quality, J. Appl. Poultry
expressed by the values of freshness index, during Research, 16, 107-112.
the 3rd day of storage, the refrigerated chicken meat Hall, M. Sykes, PA. Fairclough, DL. Lucchese, LJ., 1999,
FI are as follows: A test strip for diamines in tuna, Journal AOAC
International, 82, 5, 1102-1108
- the FI proposed by Mietz and Karmas is 0.27.
Halasz, A. Barath, A. Simon-Sarkadi, L. Holzapfel, W.,
- the FI proposed by Veciana-Nogues is 12.2. 1994, BAs and their production by microrganisms in
- the FI proposed by Silva and Gloria is 0.24. food, Trends of Food Sci. Technol. 5, 42-49.
Mervi Rokka s.a., 2004, Monitoring of the quality of
4. Conclusions modified atmosphere packaged broiler chicken cuts
stored in different temperature conditions B.
Based on the evaluation quality indicators of (the Biogenic amines as quality-indicating metabolites,
easily hydrolysable nitrogen and pH) the freshness Food Control, 5, 8, 601-607.
limit of the raw refrigerated chicken meat is three Saarinen M.T., 2002, Determination of biogenic amines
days after slaughter. as dansyl derivatives in intestinal digesta and feces

42
 The Annals of the University Dunarea de Jos of Galati
Fascicle VI – Food Technology, New Series Year II (XXXI)
by reversed phase HPLC, Chromatographia, 55, 5-6, ***, 2002, Romanian Directive no. 86 from 30/08/2002
297-300. for approving the Norms regarding the chicken meat
selling published in the Official Gazette, Part I no.
Silva C.M.G. and Gloria M.B.A., 2002, Bioactive amines
699 from 24/09/2002, issued by the Agricultural,
in chicken breast and thigh after slaughter and during
Alimentation and Forests Minister, of the Health and
storage at 4±1 °C and in chicken-based meat
Family Minister and of the Chairman of the National
products, Food Chemistry, 78, 2, 241-248.
Consumers Protection Authority.
Eerola, S., Hinkkanen, R., Lindorfs, E. and Hurvi, T.,
***, 2005, Romanian Recommendation Norm
2001, Liquid chromatographic determination of
24/01/2005 published in the Official Gazette, Part I
biogenic amines in dry sausages, Journal AOAC
no. 230 bis from 18/03/2005 – Sanitary vet norm and
International, 76, 3, 575-577.
for food safety that establishes the rules for animal
Tofan, I., 2005, Refrigerating chain of the perishable origin sampling for the laboratory examination.
food products, Agir, Bucharest, 9, 20, 197-198.
Vinci, G. Antonelli, M.L., 2002 Biogenic amines: quality
index of freshness in red and white meat, Food
Control, 13, 8, 519-524.

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