43 Dairy Year Book-2008
Recent Trends In Detection Of Adulteration Of Milk
And Dairy Products
Monika Gupta And M.P. Gupta
Research Scholar, Deptt. Of Chemistry U.P. College Varanasi,
Professor and Head, Deptt. Of Dairy Science
U.P. Pt. D.D. Upadhyay Pashu Chikitsa Vigyan Vishwavidyalaya
& Go–Anusandhan Sansthan, Mathura-281001 (U.P.)
Adulteration of foodstuffs is commonly practiced in our
country by the trade. From the view point of protecting
the health of the consumer, the Government of India
promulgated the ‘Prevention of Food Adulteration Act’
(PFA Act) in 1954. The Act came into force from 1st
June, 1955. It prohibits the manufacture, sale and
distribution of not only adulterated foods but also
foods contaminated with toxicants and misbranded
According to PFA Act, an article of food shall be
deemed to be adulterated:
(a) If the article sold by the vendor is not of the
nature, substance or quality demanded by the
purchaser or is not of the nature, substance or
quality which it proports or is represented to be,
(b) If the article contains any other substance which
affects injuriously the nature, substance or quality
(c) If any inferior or cheaper substance has been
substituted wholly or partly which affects the
quality of food.
If Any Constituent Of The Food Has Been
Abstracted Or Partly So
As to affect injuriously the quality of food.
(d) If the food article has been prepared, packed or
kept under insanitary conditions whereby it has
become contaminated or injurious to health ,
(e) If the article consists wholly or in part of any
filthy, putrid, rotten, decomposed or diseased
animal or vegetable substance or is insect infested
or otherwise unfit for human consumption.
(f) If the article is obtained from a diseased animal,
(g) If the article contains any poisonous or other
ingredient which renders it injurious to health,
(h) If the container of article is composed , whether
wholly or in part, of any poisonous or deleterious
or deleterious substance which renders its
contents injurious to health,
(i) If any colouring matter other than that prescribed
or if the amounts of prescribed colouring matter
are not within the prescribed limits of variability.
(j) If the article contains any prohibited preservative,
or permitted preservatives in excess of the
prescribed limits,
(k) If the quality or purity of the article falls below the
prescribed standard or its constituents are present
in quantities not within the prescribed limits of
variability which renders it injurious to health.
(l) If the quality or purity of the article falls below the
prescribed standard or its constituents are present
in quantities not within the prescribed limits of
variability but which does not render it injurious to
Provided that, where the quality or purity of the article,
being primary food, has fallen below the prescribed
standards or its constituents are present in quantities
not within the prescribed limits of variability, in either
case, solely due to natural causes and beyond the
control of human agency, then, such article shall not
be deemed to be adultered
The rules laid down under PFA Acct apply equally and
essentially to all articles of food, including milk and
milk products. It is evident from the PFA Act that food
adulteration includes:
1. Intentional addition, substitution or abstraction of
substances which adversely affect the quality of
2. Incidental contamination of foods with deleterious
constituents such as toxins, pesticides, pathogenic
micro organisms etc., due to ignorance, negligence
or lack of proper storage facilities,
3. Contamination of food with harmful
microorganisms during production, packaging,
storage and distribution.
Fraudulent Character
As to the character of adulteration, generally
encountered in market foods, 9 out of 10 adulterated
foods are so classed by reason of the addition Of
cheaper though harmless ingredients added for
commercial profit rather than by the addition of
actually poisonous or injurious substances, though
occasional instances of the latter are found. Authentic
instances of actual danger to health from the presence
of injurious ingredients are rare so that the question of
food adulteration should logically be met largely on the
ground of its fraudulent character.
Nature Of Adulteration
Before going into the details of hazards of adulteration,
we will have a look on the nature of adulterants
generally encountered in milk and milk products. RECENT TRENDS IN DETECTION OF ADULTERATION
44 Dairy Year Book-2008
Milk and milk products Adulterants
Milk water, removal of fat, addition of skim
Milk, reconstituted milk, skim milk
Powder thickening agents such as
Starch, arrowroot, flour, cane sugar
Glucose, urea, fertilizers pesticides,
NaCl, chlorine and pesticide residues,
Antibiotics and other drugs
Preservatives neutralizers, kerosene
Oil petroleum products heavy metals etc.
Cream Other fats, thickening agents.
Butter Hydrogenated fats, thickening agents animal Fat,
margarine, pea nut butter
Ghee Vegetable oils, fats, animal fats, thickening Agents
Condensed milk Preservatives, colour, skim milk, Homogenized foreign fats
Ice cream Prohibited colour, artificial sweeteners, Prohibited flavours,
foreign fats.
Milk powder starch dextrins
Khoa, Rabari and other Starch, arrowroot, blotting paper arti
Concentrated indigeneous Ficial sweeteners, colouring matters. Milk products
Milk based sweets Starch, colouring matter, artificial Sweeteners.
Cheese Starch, prohibited colouring matters, hydrogenated fats
It is obvious from the above table that not only
harmless adulterants but toxic and injurious
substances also find their way in milk and milk
products. Many cases of food poisoning from these
dairy products have been noticed in the past. Despite
food legislation, these have remained uncontrolled.
Unscrupulous producers and traders in our country
have never shirked to deal with adulterated or sub
standard stuff for human consumption to make
monetary gains. Where millions of producers and
traders are involved where consumers are not fully
initiated in checking malpractices and value of
wholesome food, legal steps as laid down in PFA Act
are extremely difficult to maintain Furthermore without
adequate trained man power and laboratory facilities.
It is almost impossible to deal with millions of
producers and traders. More often than not the
application of the PFA Act therefore, remains confined
to large traders and organized dairies, the number of
which is relatively small.
Health Hazards
Sub standard qualities of dairy products which are
outcome of malpractice of adulteration besides
affecting adversely the nutrient value of these products
also pose potential health hazards to consumers owing
to unhygienic practices followed during their
preparation, storage and distribution. Food poisoning
out breaks through consumption of Indian milk
products and milk based sweets have been reported
(Ghodekar et al. 1974 Batis et al 1981 The types of
toxins elaborated by some micro organisms in milk and
milk products have been listed below:
S. No. Causative organisms Toxins
1. Staphylococus Enterotoxins A,B,C,D, E and F
2. Clostridium botulinum C, perfringens Neurotixin, A to F A to E Enterotoxin
3. E, Coli LT (Heal labile ) and ST (heat stable) enterotoxins
4. B. cereus Hemolysin, lecithinase and enterotoxin
5. Aspergillus flavus Aflatoxins M1, M2, M4 etc
Cheese, milk powders, butter cream frozen milk
products like kulfi and ice cream are all potential
carrier or staphylococcal enterotoxins as revealed by
several food poisoning outbreaks (Batish et al 1981) In
these circumstances, it would be appropriate to
suggest that the Public Health Authorities should see
that regulations are strictly enforced at all stages
beginning from procurement of good quality milk free
from pathogens to marketing of final products passing
through manufacture, handling, storage and
Adulteration Rampant:
The malpractice of adulteration in milk products has
reached an alarming stage, Besides milk the menace of
adulteration is rampant in almost all dairy products but
it is too widely rampant in such products as ice cream,
butter and ghee, which are by and large, in great
demand by consumers. Substandard qualities of these
products containing various types of harmful
adulterants including pesticide residues are available in
the market. Indigenous milk products are also widely
adulterated Khoa and paneer besides milk based
sweets are widely adulterated as these are in great
demand, Ghee he costliest dairy product is the most
adulterated dairy product and most of the research
work has been carried out to find out simple, rapid and
reliable tests to detect various adulterants in ghee
(Kumar et al 2002) Reports on survey of market
guality of dairy products have revealed that about 50 RECENT TRENDS IN DETECTION OF ADULTERATION
45 Dairy Year Book-2008
percent of market samples were adulterated (Kumar et
al 1981 subrahmanyam, 1981 Sharma and Gupta 1982
Dubey and Gupta 1986 ; Sharma 1991; Rao et al 2002
and Arora et al 2004) Rao et al (2002) found 95% of
samples from venders to be adulterated .
Recently, Arora et al (2004) conducted a survey to
detect adulteration in milk in some states of North
India, using rapid test kit developed at NDRI, Karnal.
The prominent adulterants detected were urea, starch,
sugar, neutralizer, common salt and added water. The
most common adulterant encountered was water
followed by neutralizers and sugar. Adulteration was
detected in all the states of North India which were
surveyed and in some milk samples more than one
adulterant was detected the following table depicts the
status of milk adulteration in some states of North
Table – 1: Status Of Milk Adulteration In Some States Of North India,
Name of
the State
Total No
d (%)
No. of milk samples found containing
Urea Starch Glucose Sugar NeutraLiz
(NaCI) Added Formain
More than
Panjab 337 91 (25%) 1 1 3 0 22 0 51 1 12
U.P. 254 58(22%) 1 5 0 1 11 1 20 3 13
Delhi 64 25(39%) 0 0 0 1 16 0 0 0 8
Rajasthan 20 10(50%) 0 0 0 1 8 1 0 0 0
Haryana 321 120(37%) 4 2 4 20 35 4 46 0 5
Total 996 304 9 8 7 23 92 6 117 4 38
30.5% 0.9% 0.8% 0.7% 2,3% 9.2% (0.6%) (11.7%) 0.1% (3.8%)
Arora et al (2004)
It is obvious that Rajasthan, Delhi and Haryana are the
prominent states, where adulteration is rampant. It is
interesting to note that out of 321 samples collected
from Haryana, 120 samples were found to be
adulterated the samples from Haryana U.P. and Panjab
also contained urea/ synthetic milk.
Not only private sector but public sectors including
organized dairies have also been found to be involved
in this malpractice. Even today, quite a good
percentage of samples bearing ISI or Agmark
Standards would be found to be of substandard
quality. Even reputed brands of milk products
manufacturers are trying to maintain just the minimum
prescribed standards and not the optimum standards
of quality. Unless the laws are strictly enforced, it
would be impossible to get rid of the heinous crime of
Synthetic Milk Versus Adulterated Milk
Adulteration of milk has assumed new heights with the
emergence of synthetic milk in few states of northern
India. However the synthetic milk phenomenon in
India is quite recent with report appearing from
Kurukshetra, Haryana. Over the years, the reports of
synthetic milk menace have also emerged from the
states of Himachal Pradesh, Punjab, Rajasthan and
Uttar pradesh (Kumar et al 1998).
The word synthetic milk is a misnomer. By synthetic
milk, one would normally understand a product
analogous to natural milk in appearance physical
chemical and nutritional properties but there is
absolutely no similarity between the two. It is a high
degree of adulteration to increase the volume or
quantity of milk for increasing the profit Normally it is
mixture of water pulverized soap / detergent
vegetable/ refined oil caustic soda salt urea etc An
expert can detect synthetic milk in its pure form on the
basis of its appearance, flavour and consistency but
when it is mixed with natural milk it is difficult to
detect through organoleptic testes the smell, flavour,
colour, taste and consistency appear natural in skil
fully adulterated milk Table 2 depicts the differences
between synthetic and natural milk.
Table -2: Comparison Of Synthetic Milk With Natural Milk
S. No Characteristics Synthetic milk Natural milk
1. Appearance White White
2. Taste Bitter Palatable
3. Odour Soapy becomes distinct on boiling Characteristic milky odour
4. Texture When rubbed between fingers, gives a
soapy feeling
No soapy feeling when rubbed between
5. Ph Alkaline, 8.0-10.0 6.6-6.8
6. Urea test Distinctly positive (Deep yellow colour) Faint yellow due to natural urea in milk
7. Sugar test (Sucrose, glucose) Positive Negative
8. Neutraliser Test Positive Negative
9. Vegetable oil/fat test Positive Negative
10. On heating Turns yellowish Remains white RECENT TRENDS IN DETECTION OF ADULTERATION
46 Dairy Year Book-2008
Adulterated milk differs from synthetic milk in the
sense that the bulk of mixture is natural milk to which
some additional components, viz. water skim milk
sugar salt preservatives and / or neutralizers have
been added to enhance the bulk or extend the shelf life
of raw milk. On the other hand, the synthetic milk
does not contain any natural milk constituent synthetic
milk poses great risk to human health and life because
of excessive use of harmful detergents/ chemicals like
caustic soda urea ammonium sulphate formalin
hydrogen peroxide etc sodium ions in caustic soda and
detergents increase the risk of coronary heart
diseases. And urea in synthetic milk causes deleterious
effects on kidneys. In brief, synthetic milk is a toxic
Detection Of Adulterants In Milk
Simple and rapid methods have been developed to
detect various adulterants in milk. The ingredients of
synthetic milk are also detected by specific tests for
urea, ammonium sulphate, detergents, vegetable oils
Some of these tests have been delineated below:
1. Detection Of Removal Of Fat By Skimming
The following indicates this:
(i) Lower percentage of fat
(ii) Higher density reading
(iii) Higher ratio of SNF: fat
2. Detection Of Added Waster
The following indicates this
(i) Lower percentage of fat.
(ii) Lower percentage of SNF
(iii) Lower density reading
(iv) Depression of freezing point.
Water is the most common adulterant and its presence
can be detected by testing the freezing point of milk
the AOAc Specifies a freezing point for normal milk of
–0.550C and the percentage of dded water is
calculated as follows:
Percentage of added water = 0.550- T x 100
T is the freezing point depression (FPD) of suspected
milk sample. FPD of pure milk is 0.550.
A tolerance of 3 percent is allowed, which is equivalent
to specifying a minimum FPD for authentic milk of
0.53350C. The addition of preservatives and other
soluble matters like sugar and salts decrease the
freezing point of watered milk and thus escape the
detection of adulteration.
3. Detection Of Starch
Starch, cereal flours or arrowroot are added to make
up the density of milk to prevent detection of added
water. It is detected by starch- iodide test.
Three ml well mixed sample is taken in a test tube. It
is heated to boil over flame, cooled to room temp. A
drop of 1 percent iodine solution is added and mixed.
Appearance of blue colour indicates the presence of
starch which disappears on boiling and reappears on
4. Detection Of Cane Sugar
It is added to raise the density to prevent detection of
extraneous water.
To about 10 ml milk in a test tube, add 1 ml conc. HCL
and 0.1 g resorcinol and mix Place the test tube in
boiling water bath for 5 min, In the presence of cane
sugar (sucrose), red colour is produced.
5. Detection Of Glucose
Whereas the test for detection of cane sugar is simple,
that of glucose is not so. For this reason, glucose may
be added to milk instead of sucrose.
Take 1 ml milk or protein- free filtrate and add 1 ml.
modified Barfoed reagent. Heat in boiling water bath
for 3 min and cool under tap water for 2 min then add
1ml phosphomolybdic acid reagent and mix.
Development of deep blue colour indicates the
presence of glucose in milk. Pure milk shows faint
bluish due to diluted Barfoed reagent.
6. Detection Of Sodium Chloride
Sodium chloride (common salt) is added to make up
the density (lactometer reading) of watered milk.
Take 2 ml of milk and add 0.1 ml of 5 percent
potassium chromate and 2 ml of 0.1 N silver nitrate.
Appearance of Yellow precipitate indicates the
presence of sodium chloride.
7. Detection Of Ammonium Sulphate
Like urea, ammonium sulphate is a chemical fertilizer,
which is added to milk to raise the density of watered
Take 2 ml. milk in a test tube and add 0.5 ml NaOH
(2%) 0.5 ml sodium hypochlorite (2%) and 0.5 ml
phenol (5%) Heat in boiling water bath for 20 sec. A
bluish colour forms immediately, which turns deep blue
Pure milk shows salmon pink colour which gradually
changes to bluish after 2 hours.
8. Detection Of Urea
Like ammonium sulphate, urea is a chemical fertilizer,
which is added to watered milk to make up its density
(lactometer reading) Being an important ingredient of
synthetic milk, it is also used in milk to raise its SNF
content Several methods have been developed to
detect adulteration of milk with added urea. It is
noteworthy that urea is also a natural constituent of
milk. The average content of urea in cow milk is about
50 mg/100 ml whereas in buffalo milk it is present to
the extent of 35 mg/100 ml (average). It is also
important to note that feeding of urea as a protein
supplement in the ration of dairy animals does not help
to increase the urea content of milk substantially.
However, concerted investigations need to be taken up
in this direction as the menace of urea adulteration in
milk is rising day by day.
Test (i)
Take 5 ml. milk and add equal volume of 24 percent
trichloroacetic acid (TCA) to precipitate fat and
proteins of milk. Filter and collect filtrate take 1 ml. RECENT TRENDS IN DETECTION OF ADULTERATION
47 Dairy Year Book-2008
filtrate and add 0.5 ml. sodium hypochlorite (2%),
0.5ml. sodium hydroxide (2%) and 0.5 ml phenol
solution (5%) and mix.
A characteristic blue or bluish green colour develops in
presence of added urea whereas pure milk remains
Test (ii)
Take 5 ml milk in a test tube, add 0.2 ml urease (20
mg/ml) Shake well at room temperature and then add
0.1 ml Bromothymol Blue (BTB) solution (0.5%)
Appearance of blue colour after 10-15 min. indicates
the presence of urea in milk. Normal milk shows faint
blue colour due to natural urea present in milk.
Test (iii)
Take 5 ml milk in a test tube and add 5 ml of p –
Dimethyl Amino Benzaldehyde (DMAB) reagent (1.6%
in ethyl alcohol containing 10% HCI) Development of
distinct yellow colour denotes the presence of added
urea. The pure milk sample shows a slight yellow
colour due to the presence of natural urea in milk.
Processing treatments such as chilling, pasteurization
and boiling of milk as well as adulterants and
neutralizers do not affect the determination of added
urea in milk (Bector et al 1998)
The test is more sensitive when it is conducted on
protein free filtrate obtained as in case of test (i).
9. Detection Of Detergent In Milk
Take 5 ml in a test tube and add 0.1 ml Bromocresol
Purple (BCP) solution (0.5%) Appearance of violet
colour indicates the presence of detergent in milk pure
normal milk shows only faint violet colour.
10. Detection Of Pulverized Soap
It is also an ingredient of synthetic milk like detergents
.Soaps are defined as sodium or potassium salts of
fatty acids. Hence, to detect the presence of pulverized
soaps, iodine value, refractive index, fatty acid
composition, salt ratio etc. are excellent methods. The
presence can also be detected by qualitative method
as follows.
To 10 ml. of milk in a test tube, 10 ml. hot water is
added followed by 2-3 drops of phenolphthalein
indicator. Development of red/pink colour denotes the
presence of soap in milk.
11. Detection Of Synthetic Milk
Take 5 ml milk in a test tube and add 0.2ml urease (20
mg. per ml) Shake well and then add 0.1 ml of BTB
solution (0.5%) Appearance of dark blue colour
indicates the presence of synthetic milk. The methods
for detection of urea and synthetic milk are same; the
only difference is appearance of dark blue colour in
case of synthetic milk.
12. Detection Of Neutralizers In Milk
Neutralisers such as caustic soda, caustic potash
sodium carbonate, sodium bicarbonate and lime water
etc. are commonly added to milk to neutralize the
developed acidity in milk. Some of these chemicals
(neutralizers) are also ingredients of detergents which
are major components of synthetic milk. The
neutralizers added to milk are detected as follows:
Test (i):
To above 5 ml milk in a test tube, add 5 ml of alcohol
and a few drops of rosolic acid (1 percent alcoholic
solution) and mix well. Appearance of rose red colour
indicates the presence of sodium carbonate or
bicarbonate neutralizer in milk. Pure milk shows only a
brownish colouration.
Test (ii) Determination Of pH:
The pH of milk to which neutralizers have been added
is generally alkaline. The pH of such milk is always
more than 8.0, which can be determined by using
indicator dyes, pH paper or electrometrically using pH
Test (iii) Alkalinity Of Ash:
Neutralisation of milk with lime, soda ash or caustic
soda increases the ash content, and total alkalinity of
the ash from a fixed quantity of milk. This is detected
by ashing accurately measured 20 ml of milk and
titrating the ash after dispersing in 10 ml water. If the
amount of standard 0.1 N hydrochloric acid required to
neutalise the alkalinity exceeds 1.20 ml, it indicates
the presence of neutralisers in milk.
13. Detection Of Colouring Matter
It is a common practice to adulterate buffalo milk with
water and sell it as cow milk after adding some yellow
colour to it. The following colours are generally used:
(a) Artificial colours
(b) Coaltar dyes
(c) Annatto
(d) Turmeric
Some of these dyes are permitted only in some dairy
products but none in milk. Thee are often detected as
Test (i)
To 10 ml milk in a test tube, add 10 ml diethyl ether
and shake vigorously. Allow to stand. Presence of any
colour is indicated by yellow colour of the ethereal
Test (ii)
Add sodium bicarbonate to milk to make it alkaline.
Immerse a strip of filter paper for 2 hours. Red yellow
colour observed on filter paper indicates the presence
of annatto. Treatment of paper with stannous chloride
turns pink.
Test (iii)
Add a few drops of hydrochloric acid to milk.
Development of pink colour indicates azo (coaltar)
14. Detection Of Buffalo Milk Added To Cow Milk
Where there is a great demand for cow milk the
buffalo milk is generally diluted with water and mixed
with cow milk to meet the shortages in demand. It is RECENT TRENDS IN DETECTION OF ADULTERATION
48 Dairy Year Book-2008
easily easily detected by Hansa test for this test Hansa
test serum is required.
First dilute the milk 1/10. Put a drop of diluted milk on
the centre of a glass slide. Now place a drops of Hansa
test serum (duly preserved) on the drop of milk and
mix together with a glass rod or clean tooth pick.
Curdy particles develop within half a minute in milk
containing buffalo milk.
15. Detection Of Formaldehyde
Formalin (40 percent aqueous solution of
formaldehyde) is the most common preservative
added to milk. The addition of any kind of preservative
to milk is legally prohibited. Yet, market samples of
milk are occasionally found adulterated with
formaldehyde or hydrogen peroxide. Formalin
(formaldehyde) added to milk is detected by Hehner
test as follow:
To about 10 ml milk is a test tube. About 5 ml
concentrated sulphuric acid containing traces of ferric
chloride is added slowly along the side of the test tube
so that it forms a layer at the bottom, without mixing
with the milk. The development of a violet or blue
colour ring at the junction of the two liquids indicates
the presence of formaldehyde the test may be
combined with the determination of fat nothing
whether a violet colour forms on addition of sulphuric
acid in the butyro meter.
16. Detection Of Hydrogen Peroxide
This is another preservative which is frequently used in
milk to prolong its keeping quality.
Add to about 5 ml of milk (suspected sample) in a test
tube, an equal volume of raw milk and 5 drops of a 2
percent solution of paraphenylene diamine. A blue
colour is developed in presencec of hydrogen peroxide.
Note: Hydrogen peroxide in destroyed when milk is
heated or stored for a long period.
17. Detection Of Nitrates (Pond Water) In Milk
Sodium and potassium nitrates are oxidizing agents
and hence act as preservative Pond water also
contains appreciable quantities of nitrates and such
water is usually admixed with milk by rural milk
producers or vendors.
(i) Take 10 ml milk in a beaker and add 10 ml
mercuric chloride solution (2.5%in 1% HCI) Mix
well and filter through what man No 42 filter
(ii) Take 1 ml filtrate in a test tube and add 4 ml of
diphenyl amine sulphate or diphenyl benzidine
reagent Development of blue colour indicates the
presence of nitrates.
18. Detection Of Vegetable Fat
In synthetic milk milk fat is replaced by vegetable fat
or oil (refined oil) Thus, vegetable fat/oil is the chief
source of fat in synthetic milk. When synthetic milk is
admixed with cow or buffalo milk, the presence of
vegetable oil/fat becomes evident, which can be easily
detected by one or more of the following methods:
(1) Detection By Measuring Analytical
The adulteration of vegetable fat in milk can be
detected by extracting the fat either by Rose- Gottlieb
method or fat extracted in butyrometer (special
butyrometer having both end open) and measuring its
physico- chemical characteristics such as Butyro
refractometer (BR) reading, Reichert – Meissi and
Polenske values.
(2) Baudouin Test:
Hydrogenated vegetable oil (vanaspati) is a common
adulterant in milk fat. Its presence in milk fat can be
detected by the fact that sesame oil (minimum 5%) is
added in vanaspati by the law. Thus the presence of
this oil in milk fat indicates the presence of vanaspati
or sesame oil.
To 5 ml melted milk fat in a test tube, add 5 ml conc.
HCI and 0.4 ml furfural solution (2% distilled not
earlier than 24 hr. in alcohol.) Shake vigorously for 2
minutes and allow the mixture to separate. The
development of red or pink colour in acid layer
indicates the presence of sesame oil, which is
confirmed by adding 5 ml water and shaking again. If
colour in acid layer persists, sesame oil/ vanaspati is
19. Detection Of Adulterants By Using Kits
Several test kits for detecting various adulterants viz
urea, neutralizers, sucrose, glucose, pesticides
antibiotics, aflatoxins have been developed in our
country at National Dairy Research Institute, Karnal,
Central food technological Research Institute, Mysore,
PCDF, Lucknow and elsewhere . For detection of
mastitis, simple strip test has been developed further
M/s Gist Brocades, Netherlands, have developed
Delvotest kits testing presence of antibiotics and
sulpher drug residues in milk.
Causes Of Adulteration
1. Demand And Supply Gap: More acute during
summer due to low milk production and increased
2. Physical Nature Of Milk: Aqueous and opaque
nature of milk can accommodate many adulterants
in milk.
3. Degraded Moral Society: Wrecked moral status
coupled with passion for profiteering.
4. Spoiled Socio-Economic Structure: Poor
persons engaged in the business do so to increase
their income and raise socio- economic status.
5. Perishable Nature Of Milk: The unscrupulous
producers / traders use preservatives neutralizers
etc. to prolong the shelf life of sub standard milk.
6. Low Purchasing Power Of Customer:
Encourages the supplier to adulterate milk and sell
at cheaper rate.
7. Unorganised Condition Of Dairy Industry:
Most of the milk is procured and traded by
unorganized dairies, which freely adulterate the
8. Low Legal Standards And Their Improper
49 Dairy Year Book-2008
9. Lack Of Suitable, Rapid And Sure Tests:
Consumers have no access to public analytical
laboratories to get their samples analysed.
Adulteration Of Ghee And Its Detection
Ghee is an expensive product costing 3 times as much
as edible vegetable oils Thus its adulteration is a very
profitable proposition and one widely practiced variable
nature of the product is a further incentive to
adulteration The analytical characteristics of ghee
cover a very wide range permitting fairly high degrees
of adulteration while still keeping within these natural
limits .
Aqueous emulsions of coconut, groundnut and other
oils, or various starches are added to milk. On souring
and churning. These pass into butter and thence into
ghee, the latter still exhibiting its characteristic flavour.
Various vegetable oils are in common use. Coconut oil
approaches close to ghee in its analytical
characteristics and is the bugbear of the analyst
Tallows and other animal body fats are procured from
slaughter houses and being hard and firm and difficult
to be detected visually in ghee even when present in
considerable amount In fact blends of tallow and
vegetable oil can be prepared which look remarkably
like ghee.
Several methods have been developed for the
detection of adulteration in milk fat (ghee) which are
based on the differences in the nature and extents of
major / minor components of milk fat and adulterant
fat / oil No single test can detect all types of adulterant
fat/ oil. Hence, often more than one test has to be
employed o confirm the purity of milk fat.
Adulterants In Ghee Are Generally Detected By
Following Methods
1. Using different fat constants (analytical
2. Using specific tests for different adulterants viz (a)
vegetable fats (b) Animal fats (c) Non fatty
1. Using Different Fat Constants:
Fats and oils exhibit certain physico chemical
characteristics which have long been employed for
their characterization and differentiation These
analytical characteristics indicate the nature of
constituents which make up the fat Ghee is unique
among natural fats in that it contains a large
proportion of fatty acids of chain length lower than 12
C atoms and therefore its characteristics are quite
Five analytical characteristics are considered to be of
prime importance. Of these, 3 characteristics, the
Reichert Meissl Value (RV) Polenske Valve (PV) and
iodine value (IV) measure certain specific constitnents
of milk fat. Two other characteristics the specific value
(SV) and Butyro refract meter (BR) reading (or
refractive index) give an indication of the overall
average nature of the constitnent fatty acids present.
Based on some of these analytical characteristics, the
legal (PFA) and Agmark standards have been provided
to keep a check on the adulteration of ghee.
Table – 3 describes the standards of ghee under
prevention of food Adulteration (PFA) act.
Table 3: Standards Of Ghee Under PFA Rules
S.No. Name of the State & U.T B.R Reading
At 400 C
RM (Reichert)
Meissl value (Min)
Percentage of
FFA (as Oleic
acid) (Max)
Percentage of
Moistur (Max)
1. Bihar, Chandigarh, Delhi, Punjab,
Haryana (Areas other than cotton tract)
40-43 28 3 0.5
2. Manipur Meghalaya Mizoram Arunachal
Pradesh Orissa Nagland Tripura Assam
Goa Kearla Himachal Pradesh U.P. J&K
Maharashtra Rajasthan (Areas other than
jodhpur Division) Haryana (Cotton Tract
Areas) Lakshadweep
40-43 26 3 0.5
3. Kamataka Belgaum Distt Madhya
Pradesh Areas sotehr than cotton tract
areas Pondicherry
40-44 26 3 0.5
4. Andhra Pradesh Daman & Diu Dadar &
Nagar Haveli Karataka (Areas other than
Belgaum Distt)
40-43 24 3 0.5
5. Andaman & Nicobar Island Tamil Nadu 41-44 24 3 0.5
6. Gujarat (Areas other than cotton tract ) 40-43.5 24 3 0.5
7. Gujarat (Cotton tract areas) Madhya
Pradesh (Cotton tract areas) Maharashtra
(cotton tract areas) Rajasthan (Jodhpur
sub division)
41.5-45 21 3 0.5
Baudouin test shall be negative
By cotton tract is meant the areas in the state where cotton seed is extensively fed to the cattle and so notified by the
state Govt. concerned
Usually such cotton tract areas ghee has low RM value and high BR reading compared to other areas RECENT TRENDS IN DETECTION OF ADULTERATION
50 Dairy Year Book-2008
Ghee May Contain BHA Not More Than 0.02% As Antioxidant.
Table -4: Agmark Standards Of Ghee
S. No Tests All India Winter Regional Summer
1. Baudouin Negative Negative Negative
2. Phytosterol Acetate Negative Negative Negative
3. B.R.reading(400C) 40.0-43.0 41.5-44.0 42.5-45.0
4. R.M. Value (Minimum) 28 23.0 21.0
5. Polenske Value 1.0-2.0 0.5-1.2 0.5-1.0
6. Moisture (%) Maximum 0.3
7. Free fatty acids ( as %Oleic acid)
(a)Special Grade (Red Label ) Not more than 1.4 ———
(b) General Grade (Green Label) Not more than 2.5
(C) Standard Grade (Chocolate Label)
Where cotton seed is exclusively fed to milch animal.
Not more than 3.0 ———-
Analytical Characteristics Of Adulterant Oils And Fats
It is necessary to understand the physico chemical characteristics of adulterant oils and fats to assess the nature
and extent of changes in these characteristics of ghee as a consequence of adulteration. Table 5 describes some
of these analytical characteristics.
Table 5: Some Characteristics Of Ghee And Adulterant Oils And Fats
Oils / Fats B.R.Reading
(at 400C)
cation Value
Opacity Time
(at 230C)
Time**(at 170C)
Cow ghee 40-43 28-33 2-3 225-235 32-35 63-64 18-19 min —
Buffalo ghee 40-43 30-36 1.5-2.5 230-237 31-37 63-64 14-15 min 18-20min
(cottonseed fed) 44-45 15-22 0.5-0.7 210-220 41-44 66-68 11-12min 10.5-12.5 min
Groundnut oil 53-58 – – 188-195 84-102 – >40min >35 min
Hydrogenated oil 52-55 – – 197-199 70-79 66-67 1.5-2.0 min >35min
Coconut oil 38-39 6- 8 15-20 245-262 6-10 – >40 min >35min
Palm Oil 39-40 4-8 7-12 243-255 10-18 – >40 min >35min
Sesame Oil 60-64 – – 185-195 100-120 – >40 min >35min
Buffalo body fat 45-46 0.5 0.5 192-199 30-36 68-69 5-7, 10-12 sec Instant
Cow body fat 47-49 – – 193-199 38-54 – – Instant
Goat body fat 44-46 0.5 0.5 197-198 33-37 68-69 30-40 sec Instant
Pig body fat 48-51 0.5 0.5 197-200 56-64 75-76 — Instant
Sheep body fat 45-46 0.5 0.5 199-200 36-41 68-69 Instant
*Time required to acquire opacity of 0.15 O.D **Time required for the appearance of crystals.
Source: Kumar et al (2002)
Besides analytical characteristics, some of other tests
like opacity test, Bomer valve and crystallization test
have been found highly useful in detecting adulteration
of ghee with vegetable fat and / or animal body fat
opacity test and crystallization tests are particularly
useful and described below in some detail.
2. Using Specific Test
(1) Opacity Test:
Singhal (1980) developed an opacity test to detect the
Adulteration of ghee with animal body fat. Test is
performed by taking a clear melted fat sample (5g) in
a test tube and maintained at 500C for 30 min. Test
tube is then transferred to 230C Water bath and the
opacity time (time taken by the clear melted fat
sample to become opaque i.e., optical density > 0.5) is
recorded at 590 nm in a colorimeter . Normal ghee
takes more than 35 minutes while animal body fat
(buffalo, goat and sheep) take only 10 to 20 seconds
to become opaque.
In a modified test, Panda and Bindal (1998a) recorded
the opacity time (table 5) as the time required by a fat
sample at 230C to acquire the O.D in the range of 0.14
to 0.16 and consequent transmittance of 68 to 72
Opacity time of pure ghee (14-15 min) was found to
be much higher than that of ghee adulterated with
animal body fats (2-9 min at 10% level and 3-11 min
at 5% level of adulteration ) and much lower than that
of ghee adulterated with vegetable oils (21-25 min at
10% level and 19-21 min. at 5% level .
(2) Crystallisation Test:
The crystallization test has been developed to detect
the adulteration of ghee / milk with animal body fat
and vegetable oils separately or in combination (Panda
and Bindal 1998 b) this test obviates the use of any
spectrophotometer /colorimeter and can be easily used
as a platform test.
Melted and filtered fat (0.8ml) is taken in a glass
stoppered tube, to which 2.5 ml of the solvent mixture
(acetone: benzene, 3.5:1) is added and mixed RECENT TRENDS IN DETECTION OF ADULTERATION
51 Dairy Year Book-2008
thoroughly. The tube is placed in waterbath maintained
at 170C till the onset of csrystallisation. The time
required for the occurrence of crystallization in fat
sample is noted.
Table 6 & 7 show that ghee adulterated (as low as 5%
level) with animal body fats (cow buffalo and pg)
crystallizes within 8 to 15 min. In comparison to
genuine ghee which shows crystallization time of 19
min (Panda and Bindal 1998 b).
Table 6: Crystallization Time * At 170C Of Ghee Before And After Adulteration With Animal Body Fats
At 5% And 10% Level.
Ghee Adulterant Time (min) of crystallization at
0% Level 5% Level 10% Level
Buffalo ghee
— 19 – —
Pig body fat – 15 12
Cow body fat – 10 5
Buffalo body fat – 8 3
Cotton tract ghee 11 – —
*Based on 8 observation
Table – 7: Crystallization Behaviour of Normal Ghee, Cotton Tract Ghee and Adulterated Ghee
Time /Temp Normal Ghee Cotton Tract Ghee Adulterated Ghee with 5% Cow Body fat
At 170C
After 5.5 min No crystals No Crystals Crystals appeared
After 6.0 min –do– Crystals appeared Crystals increased
After 6.5 min –do– Crystals increased —-do—
After 8.0 min –do– 5.5 ml crystals 4.5 ml crystals
At 250C
After 11min —do– Crystals disappeared Crystals persisted
After 25 min —do— —do— –do–
Ghee samples prepared from milk adulterated with
animal body fats (crystallization time 4 to 12 min) and
these of milk adulterated with vegetable oils (23 to 27
min) were also detected by this test. The
crystallization time of all these samples was wither
considerably lower (animal body fats as well as
hydrogenated fats) or higher (vegetable oils) than that
of ghee sample prepared from normal milk (19 min) as
is clear from table-8)
Table -8: Crystallization Time At 170C Of Ghee prepared From Normal Milk And Milk Adulterated With
Animal Body Fats And Vegetable Oils (10% Level)
Milk Fat Adulterant Time (Min) of Crystallization
Ghee (Control)
Pig body fat
Cow body fat
Buffalo body fat
Safflower oil
Cottonseed oil
Soybean oil
Groundnut oil
Sunflower oil
Mustard oil
Coconut oil
Hydrogenated oil (Dalda)
* Based on 8 observation
3. Baudouin Test:
Addition of 5 percent sesame oil to vanaspati is
compulsory as a marker for detecting the adulteration
of latter in ghee by Baudouin test the method has been
described in detail earlier.
4. Phytosteryl Acetate (PA) Test:
The detection of plant sterols (phytosterols) in
association with cholesterol is important in detecting
the adulteration of milk fat with vegetable fat.
The melting point of cholesteryl acetate (114-150C) is
lower than that of phytosteryl acetates (125-1370C). If
the melting point of steryl acetate is 1150C the ghee
sample is genuine and free from vegetable fat If the
melting point is higher than 1170C, the fat sample is
considered to contain vegetable fat.
5. Halphen’s Test For Cotton- Seed Oil
This test can detect as low as one percent cotton seed
oil. The ghee samples from cows fed with cotton seed
52 Dairy Year Book-2008
To 5 ml melted ghee in a test tube, add 5 ml iso amyl
alcohol and 5 ml 1 percent solution of sulphur in
carbon bisulphide Heat in boiling water bath for 30-35
min. The development of rose red colour indicates the
presence of cotton seed oil.
Note: If cotton seed oil has been heated above 2500C,
the test fails completely.
6. Methylene Blue Reduction (MBR) Test For
Cotton Seed Oil :
This test is based on the cylopropenoic fatty acids
present in cotton seed oil which quickly reduce the
methylene blue dye. Reduction of methylene blue
denotes either the presence of cotton seed oil in milk
fat or ghee from cotton tract area. Normal ghee does
not reduce methylene blue.
7. Lodine Test For Presence Of Starch:
Starchy materials are used when liquid oil are added to
ghee to increase its consistency. These materials are
insoluble in fat and therefore easily detected by mixing
the ghee with hot water, removing aqueous layer and
adding a few drops of 1 percent iodine solution. The
formation of blue colour indicates the presence of
Remedial Measures:
(1) There is need for rationalization of the standards
prescribed under PFA Act. Buffalo milk, for which
minimum requirement for fat % is 6 in most of the
states, is hardly available as such in the market for
sale. Either it is watered and sold as cow milk or
admixed with cow milk and sold as mixed milk. In
lieu of buffalo milk, full cream milk has been
introduced containing minimum of 6.0% fat and
9.0% SNF.
(2) The PFA Act and Rules must be strictly enforced
and offenders punished adequately. The manpower
limitation, lack of adequate training to the food
inspectors and apathy of consumers encourages
the menace of adulteration.
(3) The adulterated substandard and injurious food
stuffs (including dairy products) should be
discouraged from trade. The enforcement of the
act should be rigidly carried out particularly against
unorganized dairies and small traders and vendors,
which are the root causes of this malpractice.
(4) Certain discrepancies exist in the standards
prescribed under PFA Act and ISI and Agmark
standards. These anomalies should be rectified and
a uniform standard should be prescribed.
(5) Rapid, reliable and inexpensive tests to detect
various harmless and harmful adulterants should
be worked out so that cases of adulterations are
detected readily.
(6) The milk producers should be given incentives for
clean milk production and should be encouraged to
supply the milk to the registered village societies,
milk unions and dariries. The various
intermediaries should be eliminated.
(7) Special provision should be made for packaging
and distribution / sale of dairy products. Most of
the market samples of dairy products are stored
under unhygienic conditions and sold loose without
any specification with regard to the nature and
content of the product.
(8) Special provisions should be framed for rigorous
control over the production, distribution and sale of
milk and milk products including registration of
premises where they are manufactured,
maintenance of premises in a sanitary condition
and maintenance of healthy states of human
beings associated with the production, distribution
and sale of such foods.
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(2004). Status of milk adulteration in some states of North
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(1981) .Indian Dairyman 33,435.
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PFA Act (1954) Prevention of food Adulteration Act and
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Agra City M.Sc Thesis submitted to Agra Univ. Agra.
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