MODULE 14:

Constants of Fats and Other Fatty

Substances
Analysis of Fatty Substances
• ​Such methods usually consist of the determination
of physical and chemical properties or values known
as constants.
• ​When taken in conjunction with organoleptic tests
and special ID tests, such methods form the basis
upon which the purity and quality of fat samples are
judged.
• ​Physical constants such as specific gravity, solubility,
melting point, congealing point, refractive index and
optical activity determinations are discussed in the
 official compendia.
• ​Some samples employ tests for moisture and non
fatty tissue residues and are labeled under the
heading “Water and Sediment Content”. This is of
importance in fatty oils especially of animal origin.
Acid Value
• ​Number in mg of potassium hydroxide
necessary to neutralize the free fatty acids in
1g of oil, fat, wax, resin, balsam, or similar
organic substance of complex composition.
• ​May also be expressed as the number in mL of
0.1N NaOH required to neutralize the free fatty
acids in 10g of fat sample.
• ​Procedure carried out by dissolving the sample
 in alcohol or alcohol-ether solution and
titration with standard alkali solution.
• ​Balsams and resins are dissolved in alcohol
ether solution since the coloring matter is
dissolved in the ethereal layer.
• ​Solid fats and waxes are melted on a water
bath and titrated while hot.
Acid Value
• ​A high acid value, compared to the maximum official
requirement or maximum limit, indicate extensive hydrolytic
decomposition of the fat sample.
• ​It does not necessarily indicate rancidity since this is due to
action of air or bacteria.
• ​A low acid value, compared to the minimum official
 requirement or minimum limit indicates adulteration and an
inferior quality since fat samples inherently contain free fatty
acids.
• ​In titration of fatty substances:
– ​The sample should be clear.
– ​The sample should be introduced by a dropper or a Pasteur
pipet.
– ​It is necessary to shake the mixture thoroughly after addition of
each portion of alkali to secure complete extraction of the fatty
acids to be neutralized from the oily layer.
Sample Calculation: Acid Value
If a 2g sample of cod-liver oil required 4.5mL of
0.02N KOH in the titration of its free fatty acids,
what is its acid value?
mEq of KOH = ​39.098g + 15.9994g +
 1.0079g​ ​1 x 1,000
0.0561053g/mEq ​ ​mg/mEq
56.1053mg/mEq ​ ​56.11mg/mEq
Acid Value = ​V x N x 56.11mg/mEq
Sx wt. (g)
= ​4.5mL x 0.02N x 56.11mg/mEq
2g
= 2.52mg KOH/1g of cod liver oil
= 2.52mg/g
Saponification Value
• ​Number in mg of potassium hydroxide required to
neutralize the free fatty acids and saponify the esters
contained in 1 g of fat, fatty oil or volatile oil, wax, resin,
balsam or other substance of similar composition.
 • ​Quantifies the glycerides of acids and detects the presence
of unsaponifiable matter present as adulterant. ​• ​The sample
is treated with an excess of alcoholic 0.5N KOH ​and heated in
a flask with an air condenser, with constant ​shaking to
accelerate saponification.
• ​Alcoholic KOH is used in order to solubilize the sample and
the saponification products.
• ​The excess is back titrated with 0.5N HCl and a blank
determination is performed.
• ​H​2​SO​4 must
​ not be used as a substitute since K​2​SO​4​is
insoluble in alcohol.
Sample Calculation: Saponification Value
What is the saponification value of cottonseed
oil if a 1.532g sample treated with 25mL of
0.5N alcoholic KOH required 15.70mL of 0.51N
 HCl for the residual titration. The blank
determination required 26mL of the standard
acid.
Saponification Value = (​V​b –​ V​a​) x N x
56.11mg/mEq
Sx wt. (g)
= ​(26mL – 15.70mL) x 0.51N x 56.11mg/mEq
1.532g
= 192.39mg KOH/1g of cottonseed oil
= 192.39mg/g
Ester Value
• ​Number in mg of potassium hydroxide required to saponify the ​esters
in 1g of fat, fatty oil or volatile oil, wax, balsam, resin or similar
substance.
• ​In samples that contain free fatty acids, ester value is equal to the
difference between the saponification value and acid value. ​• ​In
samples that don’t contain free fatty acids, ester value is equal to the
saponification value.
• ​Important in the analysis of waxes to indicate the presence of
adulterants especially paraffin.
• ​Addition of acid value to the ester value will give the saponification
value.
Sample Calculation:
If a sample of beeswax is found to have an acid number of 20.4 and a
saponification value of 89.8, what is the ester value?
Ester Value = Saponification Value – Acid Value
= 89.8mg/g – 20.4mg/g
= 69.4mg KOH/1g of beeswax ​ ​69.4mg/g
Unsaponifiable Matter
• ​Substances present in oils or fats that are not
saponified by alkali hydroxides but are soluble
 in ordinary fat solvents.
• ​Usually added as an adulterant to fat samples.
• ​Normally are dissolved in alcoholic soap
solution after saponification with alcoholic
KOH, but floats on top of the reaction mixture
if large amounts are present.
• ​Indicates quality and purity.
• ​Method conducted by treatment of the sample
with alcoholic KOH with subsequent
evaporation of the alcohol. The residue that
remains is dissolved in water and the
unsaponifiable matter extracted by ether.
Iodine Value
• ​Number in g of iodine absorbed under specified
 conditions by 100g of oil, fat, wax or other
substance
• ​Quantifies the unsaturated fatty acids, both in the
free and combined form.
• ​It serves to characterize oils if they are pure or
admixtures.
Range of Iodine Values: (g/100g)
>120 – Drying Oils e.g. Linseed and Cod-Liver
Oil ​100-120 – Semi-drying Oils e.g. Cottonseed
and Sesame oil
< 100 – Non-drying Oils e.g. Olive and Almond Oil
Animal fat usually have iodine values below 90 ​• ​In
conjunction with saponification value, adulteration
can be detected as well as the nature of ​the
adulterant.
 Iodine Value
Examples:
Olive Oil Adulterated with Cottonseed Oil No
effect on saponification value but increased
iodine value of olive oil.
Castor Oil Adulterated with Olive Oil
No effect on iodine value but increased
saponification value of castor oil.
Methodologies: Hanus, Wijs, and Hϋbl
• ​The sample is dissolved in carbon tetrachloride and
treated with Iodine Monochloride/Iodochloride T.S.
(​I​Cl) in excess inside a glass-stoppered flask to
prevent loss and protected from light to prevent
other side reactions from occuring.
• ​If the solution is no longer yellow after such procedure, the test is
 repeated with a smaller amount of sample.
Iodine Value

• ​The excess ​I​Cl is treated with KI T.S. to liberate

Iodine.
I​Cl + KI ​ ​KCl + I​2
• ​An excess of KI is added to ensure complete removal
of free chlorine and prevent precipitation of iodine
in aqueous solution.
• ​The liberated I​2​is titrated with standard Na​2​S​2​O​3​. ​• ​A
blank determination is carried out to correct for
errors and to make it unnecessary to determine the
N of Iodochloride.
Sample Calculation: Iodine Value
Determine the iodine value of a sample of olive oil
weighing 0.2100g if 24.15mL and 12mL of 0.1100N
sodium thiosulfate solution are required for the
blank and residual titrations respectively. mEq of I
= ​126.9g
1 x 1,000
= 0.1269g/mEq
I​2 Value
​ = ​(V​b –​ V​a​) x N x 0.1269g/mEq x
100​ ​Sx wt. (g)
= (​24.15mL – 12mL) x 0.1100N x 0.1269g/mEq x
100​ ​0.2100g
= 80.76gI/100g of olive oil
= 80.76g/100g