INTERNATIONAL ISO

STANDARD 660

Third edition
2009-06-15

Animal and vegetable fats and oils —

Determination of acid value and acidity
Corps gras d'origines animale et végétale — Détermination de l'indice
d'acide et de l'acidité

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Reference number
ISO 660:2009(E)

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ISO 660:2009(E)

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Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies
(ISO member bodies). The work of preparing International Standards is normally carried out through ISO
technical committees. Each member body interested in a subject for which a technical committee has been
established has the right to be represented on that committee. International organizations, governmental and
non-governmental, in liaison with ISO, also take part in the work. ISO collaborates closely with the
International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization.

International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2.

The main task of technical committees is to prepare International Standards. Draft International Standards
adopted by the technical committees are circulated to the member bodies for voting. Publication as an
International Standard requires approval by at least 75 % of the member bodies casting a vote.

Attention is drawn to the possibility that some of the elements of this document may be the subject of patent
rights. ISO shall not be held responsible for identifying any or all such patent rights.

ISO 660 was prepared by Technical Committee ISO/TC 34, Food products, Subcommittee SC 11, Animal and
vegetable fats and oils.
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This third edition cancels and replaces the second edition (ISO 660:1996), which has been technically revised.
It also incorporates the Amendment ISO 660:1996/Amd.1:2003.
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INTERNATIONAL STANDARD ISO 660:2009(E)

Animal and vegetable fats and oils — Determination of acid

value and acidity

1 Scope
This International Standard specifies three methods (two titrimetric and one potentiometric) for the
determination of the acidity in animal and vegetable fats and oils, hereinafter referred to as fats. The acidity is
expressed preferably as acid value, or alternatively as acidity calculated conventionally.

This International Standard is applicable to refined and crude vegetable or animal fats and oils, soap stock
fatty acids or technical fatty acids. The methods are not applicable to waxes.

Since the methods are completely non-specific, they cannot be used to differentiate between mineral acids,
free fatty acids, and other organic acids. The acid value, therefore, also includes any mineral acids that may
be present.

2 Normative references
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The following referenced documents are indispensable for the application of this document. For dated
references, only the edition cited applies. For undated references, the latest edition of the referenced
document applies.
ISO 660:2009
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ISO 661, Animal and vegetable fats and oils — Preparation of test sample
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ISO 3696, Water for analytical laboratory use — Specification and test methods

3 Terms and definitions

For the purposes of this document, the following terms and definitions apply.

3.1
acid value
number of milligrams of potassium hydroxide required to neutralize the free fatty acids present in 1 g of fat,
when determined in accordance with the procedure specified in this International Standard

NOTE The acid value is expressed in milligrams per gram.

3.2
acidity
content of free fatty acids determined according to the procedure specified in this International Standard

NOTE The acidity is expressed as a percentage by mass. If the result of the determination is reported as acidity
without further explanation, this is, by convention, the acidity based on the oleic acid content.

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ISO 660:2009(E)

4 Principle
The sample is dissolved in a suitable solvent mixture, and the acids present are titrated with an ethanolic or
methanolic solution of potassium or sodium hydroxide.

The methods specified in 9.1 and 9.2 are reference methods.

5 Reagents
WARNING — Attention is drawn to the regulations which specify the handling of hazardous
substances. Technical, organizational and personal safety measures shall be followed.

Use only reagents of recognized analytical grade, unless otherwise specified.

5.1 Solvent A for solvent mixture (5.3): ethanol, volume fraction, ϕ ≈ 96 %.

As a replacement, propan-2-ol, volume fraction, ϕ ≈ 99 %, can be used.

5.2 Solvent B for solvent mixture (5.3): diethyl ether, peroxide-free.

As a replacement, tert-butyl methyl ether, light petroleum (boiling range 40 °C to 60 °C) or toluene can be
used.

WARNING — Diethyl ether is very flammable and may form explosive peroxides. Use with great
caution.
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5.3 Solvent mixture,
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mix equal volumes of solvent A and B, (e.g. ϕ A = 50 ml/100 ml and
ϕ B = 50 ml/100 ml).
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For hard or animal fats, a solvent mixture of one volume of solvent A (e.g. 25 ml) and three volumes of tert-
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butyl methyl ether or toluene (e.g. 75 ml) is recommended.

Neutralize, just before use, by adding potassium hydroxide solution in the presence of 0,3 ml of the
phenolphthalein solution per 100 ml of solvent mixture.

For the titration with aqueous KOH, the solvent propan-2-ol can be used.

5.4 Ethanol or methanol, of minimum volume fraction, ϕ = 95 %.

5.5 Sodium hydroxide or potassium hydroxide, ethanolic or methanolic standard volumetric solutions,
amount of substance concentration c(NaOH) or c(KOH) = 0,1 mol/l and 0,5 mol/l. The concentration shall be
checked with a standard volumetric HCl solution.

NOTE The ethanolic/methanolic sodium/potassium hydroxide solution may be replaced by an aqueous

sodium/potassium hydroxide solution, but only if the volume of water introduced does not lead to phase separation.

5.6 Phenolphthalein, solution in ethanol, mass concentration, ρ = 1 g/100 ml.

5.7 Thymolphthalein, solution in ethanol, mass concentration, ρ = 2 g/100 ml.

5.8 Alkali blue 6B, solution in ethanol, mass concentration, ρ = 2 g/100 ml.

For dark-coloured fats, alkali blue or thymolphthalein shall be used.

5.9 Water in accordance with ISO 3696, grade 3.

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6 Apparatus
Usual laboratory equipment and, in particular, the following.

6.1 Burette, capacity 10 ml, graduated in 0,02 ml, ISO 385[1] class A.

6.2 Burette, capacity 25 ml, graduated in 0,05 ml, ISO 385[1] class A.

6.3 Analytical balance, capable of being read to the nearest 0,001 g.

6.4 Automatic titration apparatus (based on potentiometric electrode) or potentiometer.

6.5 Combined pH electrode for non-aqueous acid/base titrations.

6.6 Graduated volumetric flasks, volume 1 000 ml, ISO 1042[2] class A.

7 Sampling
A representative sample should have been sent to the laboratory. It should not have been damaged or
changed during transport or storage.

Sampling is not part of the method specified in this International Standard. A recommended sampling method
is given in ISO 5555 [3].
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Preparation of test sample
Prepare the test sample in accordance with ISO ISO661, except that if the sample contains volatile fatty acids, the
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test sample shall not be heated and filtered.
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9 Procedure

9.1 Cold solvent method using indicator (Reference method)

9.1.1 Depending on the expected magnitude of the acid value, select the test portion mass and alkali
concentration from Table 1.

9.1.2 According to Table 1 weigh the test portion into a 250 ml conical flask.

9.1.3 Add 50 ml to 100 ml of the neutralized solvent mixture (5.3) and dissolve the test portion if necessary
with gentle warming.

For high melting point samples, use an ethanol-toluene mixture.

9.1.4 After the addition of an indicator (5.6, 5.7 or 5.8), titrate with constant swirling using standard
potassium hydroxide solution (5.5). The endpoint of the titration is reached when the addition of a single drop
of alkali produces a slight but definite colour change persisting for at least 15 s.

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ISO 660:2009(E)

Table 1 — Test portion masses and alkali concentrations

Accuracy of
Product group Acid value Mass of test Concentration of
weighing of the
(examples) approx. portion KOH
test portion
g mol/l g

Refined vegetable oils

0 to 1 20 0,1 0,05
Animal fats
Crude vegetable oils 1 to 4 10 0,1 0,02
Technical grade animal fats 4 to 15 2,5 0,1 0,01
0,5 0,1
Soap stock fatty acids 15 to 75 0,001
3,0 0,5
0,2 0,1
Technical fatty acids > 75 0,001
1,0 0,5

9.2 Cold solvent method using potentiometric titration (Reference method)

9.2.1 According to Table 1, weigh the test portion into a 150 ml beaker.

9.2.2 Add 50 ml to 100 ml of the neutralized solvent mixture (5.3) and dissolve the sample, if necessary with
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For high melting point samples, use an ethanol-toluene mixture.

9.2.3 Introduce the combined electrode in the solvent mixture and connect it with the automatic titration
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9.2.4
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Start the stirrer for at least 30 s and then titrate with constant swirling using standard potassium
hydroxide solution (5.5).

9.2.5 As soon as the equivalence point is reached, record the amount of standard solution used.

9.3 Hot ethanol method using indicator

9.3.1 Under the conditions specified in this method, short-chain fatty acids, if present, are volatile.

9.3.2 Weigh into a flask a sufficient mass of the test sample as shown in Table 1, according to the colour
and expected acid value.

9.3.3 Heat to boiling 50 ml of the ethanol containing 0,5 ml of the phenolphthalein indicator in a second
flask. While the temperature of the ethanol is still above 70 °C, neutralize it carefully with a solution of 0,1 mol/l
sodium or potassium hydroxide.

The endpoint of the titration is reached when the addition of a single drop of alkali produces a slight but
definite colour change persisting for at least 15 s.

Larger volumes of ethanol and indicator may be necessary for dark-coloured fats. Moreover, for dark-coloured
fats, alkali blue or thymolphthalein shall be used.

9.3.4 Add the neutralized ethanol to the test portion in the first flask and mix thoroughly. Bring the contents
to the boil and titrate with the sodium or potassium hydroxide solution, agitating the flask contents vigorously
during the titration.

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10 Calculation
The acid value, wAV, or the free fatty acid content, wFFA, is reported as follows:

a) to two decimal places for values between 0 up to and including 1;

b) to one decimal place for values between 1 up to and including 100;

c) as a whole number for values > 100.

In addition to the following calculations, the approximate free fatty acid content (acidity) is calculated from:

wFFA = 0,5 × wAV

10.1 Acid value

The acid value, wAV, expressed as a mass fraction, is equal to

56,1 × c V
w AV =
m

where

c is the exact concentration, in moles per litre, of the standard volumetric sodium or potassium
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hydroxide solution used;

V (standards.iteh.ai)
is the volume, in millilitres, of standard volumetric sodium or potassium hydroxide solution used;

m is the mass, in grams, of the test portion.

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The acidity or free fatty acid content, wFFA, expressed as a percentage mass fraction, and according to fat
type (see Table 2), is equal to

V c M × 100
wFFA =
1 000 × m

where

V is the volume, in millilitres, of the standard volumetric sodium or potassium hydroxide solution used;

c is the concentration, in moles per litre, of the standard volumetric sodium or potassium hydroxide
solution used;

M is the molar mass, in grams per mole, of the acid chosen for expression of the result (see Table 2)
according to the fat type;

m is the mass, in grams, of the test portion.

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