Dr.a.k.srivastava chemical and drug residues in dairy products (ayurvet seminar) 8.10.2014

Chemical and Drug Residues
in Dairy Products
Anil Kumar Srivastava
Director
ICAR - National Dairy Research Institute
Karnal-132001 (India)
Ayurvet Knowledge Symposium at Delhi; October 8 , 2014

Background
Residues in livestock
products
 Pesticide
 Antibiotic
 Hormones
 Anabolic drugs
 Mycotoxins
 Heavy metals
Chemicals in livestock
products
Adulterants in milk including
melamine
Have Potential risk of various life threatening diseases – cancer,
leukemia, reproductive disorder, disruption of body’s immune,
endocrine and nervous system…….
Ayurvet Knowledge Symposium at Delhi; October 8, 2014

Risk of Antibiotics Residue
 Consumer safety: Antibiotic resistance as “one of the
greatest threats to human health” (WHO, 2011)
 Regulatory Issues: Compliance to standards
 Processing difficulties in dairy processing: Economic loss
• Affects Rennet clotting time & improper ripening of cheeses
• Inhibits lactic acid fermentation & flavour development

Biggest Threat of Antibiotic Residue
Centers for Disease Control and Prevention,
USA noted that 2 million people develop
antibiotic resistance each year in the U.S.,
resulting in at least 23,000 deaths.
CDC report 2013

Standards for Antibiotic Residues
Name of antibiotic
MRL (ppb) in milk MRL as per FSSAI (ppb)
Codex EU in sea foods only
Benzyl penicillin 4 4 -
Ceftiofur 100 100 -
Oxytetracycline - 100 100
Tetracyclines 100 100 100
Sulfonamides 100 100 -
Gentamycin 200 100 -
Streptomycin 200 200 -
Enrofloxacin - 100 -
Bacitracin 500 100 -
Novobiocin 100 50 -
Trimethoprim 50 50 50
Oxolinic acid - - 300
FSSAI is silent about antibiotic residues in milk and milk products

Incidence of Antibiotic Residues in Milk
Type of drug
% of incidence
more than MRL
Codex (MRL) μg/Kg
β-Lactum 3.07% 4-200
Sulpha drug 0.87% 100
Tetracycline 1.31% 100
Amino – neomycin 0 1500
Amino-Streptomycin 1.31 200
Macrolide 0.43% 40-200
Multiple drugs 2.63 -
Total incidence 9.62% -
 A total of 228 samples were collected from individual farm and commercial dairies
around Karnal which consists of 122 raw milk, 77 Pasteurized and 29 Dried milk.
 Problem of antibiotic residues is more in organized dairies.
Kumar et al., 2012 (World J. Mirobiol Biotechnol, 28: 2559-2566)

MDR Test Kit: Detection of Antibiotic
Step -1 Step - 2
Add 75 μl of milk into ampoule
containing spores
Incubate at 64oC 3 h
Observe the colour change from
purple to yellow
No colour means
antibiotic positive
Colour change means
antibiotic negative
Step - 3
Residues in Milk

Novel Features of MDR Test Kit
• Cost effective (Rs 33 per test )
• Semi-quantitative detection at Codex MRL
• Validated with AOAC approved Charm 6602 Assay
• No interference of inhibitors other than antibiotic residues
• Stability of kit 12 months - refrigeration storage
• Test kit can perform at dairy farm, milk collection center,
dairy reception dock and R&D institutions
• Technology has been licensed to M/s Neugen Diagnostic Pvt.
Ltd. Secunderabad

Aflatoxin M1
 Naturally occurring mycotoxins
 Toxic metabolites produced by certain fungi, Aspergillus
flavus, A. parasiticus and A. nomius
 Four naturally occurring compounds: aflatoxin B1, B2, G1,
and G2
 Aflatoxin M1 (AFM1): hydroxylated metabolite of aflatoxin
B1
 AfAlatoxin B1 is biotransformed by hepatic microsomal
cytochrome P450 to aflatoxin M1
 AFM1 has been shown to be excreted in milk following
exposure to aflatoxin B1 contaminated feed

• AFM1 is relatively stable during the pasteurization,
storage and preparation of various dairy products
• AFM1 was shown to have genotoxic effects in mammalian
systems in vivo
• AFM1 was found to possess carcinogenic potential but it
was 10 times lower with respect to the parent molecule
(aflatoxin B1)
• AFM1, initially classified as a Group 2B agent, has now
been reclassified
Aflatoxin M1

Aflatoxin M1
AflatoxinM1 recommended limits in milk and infant formulae
 European Communities/Codex Alimentarius recommended
limits of aflatoxin M1 in milk 0.05 PPB and 0.025 PPB for
infant formulae.
 The US Food and Drug Administration have set levels
higher than the European Commission at 0.5 PPB in milk.
FSSAI - 30 PPB – all articles of food
- 0.5 PPB – Milk

Test for Detection of Aflatoxin M1 in Milk
A rapid and cost effective test working on principle of
inhibition of spore germination in presence of aflatoxin M1 in
milk.
1. Development of sky blue
colour within 45 min (tube-1)
indicates absence of
Aflatoxin M1 in milk.
2. No colour change within 45
min (tube-2) indicates
presence of Aflatoxin M1 at
0.5 ppb codex limit.
3. No colour change (tube-3)
indicates proper heat
treatment of milk (Negative
control )
1 2 3
Patent Application Number: # 3064/DEL/2010

Novel Features Test for Detection
 Results available within 45 min
 Cost effective (Rs.20 per test)
 Semi quantitative detection at 0.5 ppb limit set by Codex
 Validated with AOAC approved Charm 6602 and ELISA
assay
 Test can detect Aflatoxin M1 in raw, pasteurized & dried
milks
of Aflatoxin M1 in Milk

Pesticide Residues in Dairy Products
Organochlorine Organophosphate Carbamate
• Nonpolar
• More persistent
• Toxic
• Moderately polar
• Less persistent
• Most toxic
• Polar
• Less persistent
• More toxic
Examples
 DDT
 Dieldrin
 Chlordane
 Lindane
 BHC
 Endosulfan
 Heptachlor
Examples
 Parathion
 Malathion
 Methyl parathion
 Cloropyriphos
 Fenitothion
 Ethion
 Phorate
Examples
 Carbofuran
 Carbaryl
 Aldicarb

Pesticide Residues
• India is fourth largest supplier of
agrochemicals in global market
• Consumption of agrochemicals in
India is low
• India – 0.58 kg/Hectare (28% used
in paddy followed by cotton)
• Enough data is there to
indicate severity of problem
A report indicate that 51% of food commodities are contaminated with
pesticide residues and 20% of them are above MRL
http://www.eco-web.com/edi/070526.html

Oxytocin in Milk
• 9 amino acid neuropeptide hormone, synthesized in the
paraventricular and supraoptic nucleus of the hypothalamus
• OT has been used for induction or enhancement of uterine
contractions at parturition, treatment of postpartum retained
placenta and metritis
• In recent years – crude OT preparation used for quick and
complete evacuation of milk in buffaloes
Myth about OT
• Effect on adolescence
• Lack of nutritional element in such milk
• Adverse effect on immunity

Reality about Oxytocin
• Whether secreted endogenously in response to natural
stimuli or administered exogenously, OT produces the
desired effects within minutes and gets metabolized
rapidly, leading to inactive products
• If at all ingested would be digested like any other protein
• Till date not a single report demonstrates the appreciable
presence of OT in milk

• Milk samples collected from cows (n = 38)
administered either 25 or 50 IU OT prior to
milking were investigated for the presence of
OT in milk.
• There was no significant difference among
both groups with the mean concentrations of
OT being 15.8 and 14.9 pg/ml for cows
subjected to 25 and 50 IU OT administration
(basal conc. 10 pg/ml), respectively.
• Under normal circumstances administration of
only 0.1 IU of OT are required for milk let-down.
1 IU = 2 μg of pure peptide
n = 30
n = 19 n = 19
OT concentrations in milking just after OT
administration in cows.

Metal
Maximum level (PPM)
FSSAI Codex
Arsenic
• 0.1 – milk
• 0.05 – infant milk substitute & infant food
• 0.5 –ice-cream
-
Cadmium
• 1.5 – for other milk products
-
Lead
• 1.0 –ice-cream
• 2.5 – for other milk products
0.02 – infant milk
substitute & infant
food
Mercury • 1.0 – all milk products -
Copper
• 15 but not less than 2.8 – infant milk
substitute & infant food
• 30 – foods not specified
-
Tin • 250 – foods not specified -
Zinc
• 50 but not less than 25 – infant milk
substitute & infant food
-
Heavy Metals

Heavy Metal Residues
• Mainly from automobiles
• Problem is more in cities
• Use of sewage water to grow forage
• Almost all data that is available indicate levels of Pb, Cd,
Hg are within safe limit
• Arsenic in West Bengal indicate higher level in fodder
and milk but again within safe limit

Types of Adulterants Reported in Milk
 Carbohydrates : Sugar, Glucose, Starch, Maltodextrin, etc.
 Salts and fertilizers : Urea, Ammonium sulphate, NaCl etc.
 Neutralizers : NaOH, Na2CO3, NaHCO3 etc.
 Preservatives : H2O2, Formalin, Boric acid etc.
 Detergents : Liquid detergents, Washing powders etc.
 Pond Water : Heavier than tap water
 Miscellaneous : Vegetable fats and oils, Soymilk, Sorbitol,
Paint, Thiocyanate etc.

Why Milk Adulteration?
 Adulteration of milk by unscrupulous persons
is usually done for…..
• Increasing the quantity of milk
• Extending shelf-life
• Maintaining SNF
• Avoiding detection of sour milk.

Milk Adulteration in Other Countries
Name of
Country/Region
Type of Adulterant Reported
 Pakistan
 Bangladesh
 Sri Lanka
Water, Urea, detergent, formalin, H2O2,
Neutralizers, Starch, Flour, vegetable oil, formalin,
quaternary ammonium (QAC) compounds, Cane
sugar, sorbitol, salt, boric acid, hypochlorite etc.
 China Melamine
 Argentina
 Brazil
Water, Neutralizers, Sodium citrate, non-acid
cheese whey, synthetic milk
 Europe Cheese Whey, Milk fat adulteration, Species
adulteration of Milk

Adulteration of Milk Products
Milk product Adulterant (s) reported to be added
Ghee
• Vegetable oil,
• Hydrogenated vegetable oil,
• Animal body fat,
• Designer oil, mineral oil, colour, flavour etc.
Paneer • Edible and non-edible oil, starch etc.
Khoa and khoa based
sweets
• Edible and non-edible oil, starch etc.
Milk Powder
• Maltodextrin
• Neutralizers
• Whey powder
Ice-cream • Vegetable oil, starch

National Survey on Milk
Adulteration, 2011
Report, 2012
State/ UT No. of Samples Collected %Non-conformances
Bihar 75 100
Chhattisgarh 19 100
Daman & Diu 25 100
Jharkhand 25 100
Mizoram 25 100
Orissa 50 100
West Bengal 100 100
Uttar Pradesh 136 88
Punjab 109 81
Rajasthan 103 76
Delhi 71 70
Haryana 109 70
Madhya Pradesh 61 68
Maharashtra 126 65
 Goa 24 0
 Puducherry 25 0

• Total no of samples analysed: 1791
• Total non-conforming samples: 1226 (68.4%) – 28% - packed
milk
• 574 samples (47%) samples failed on account of Fat and SNF
• 2nd highest parameter of non-conformity was Skim Milk
Powder (SMP) in 548 samples (45%)
• Glucose in 477 (39%) samples
• Detergent was found in 103 samples (8%) – eastern part
(Orrissa, Jharkhand, W. Bengal)
 Haryana – 76 samples (70%) – Glucose and SMP
 Punjab – 88 samples (81%) – Glucose and SMP
Report, 2012
National Survey on Milk
Adulteration, 2011

Adulterant
% of sample
+ve
Sucrose 22%
Neutralizer 26%
Formalin 32%
H2O2 32%
Detergent 44%
Urea 60%
NaCl 82%
n = 50

NDRI is Working Proactively in
Developing Tools for Detection of Milk Adulteration
• Kit for detection of common adulterants in
milk
• A rapid method for detection of detergent in
milk – technology has been transferred
• Strip based tests for detection of other
common adulterants – detection limit is better
than liquid based tests
• Also developing tools for detection of
adulterants in milk products

Issues with Milk and Milk Products
 Fragmented Dairy Supply Chain
 Animals are diseased ? - FMD
 Outbreaks – not reported
 Feasibility of Traceability
• Possible in organized sector
 Validation of information on milk of buffaloes and
indigenous breeds
 Antibiotic residues – No MRLs
 Chemical residues – inadequate facilities for regular
monitoring

Other Issues
• Lack of Scientific information on the status of chemical
contaminants in food products.
• e.g 7th Session of the Codex Committee on Contaminants
- Lead contamination in milk - We did not have data to
resist the change in MRL of lead in juices
• Limited scientific surveys
• Limited scientific information on ethnic varieties of foods
• Lack of coherent action among stake-holders
• Lack of proper and dedicated mechanism for
epidemiological studies

Remedies/ Summary
• Harmonization of domestic standard and international
standards
• Redefine the role of Institutions in Codex-India for
domain specific issues
• Generation of data through Scientific Surveys -
Intervention of Govt. agencies e.g FSSAI
• Meta-analysis of existing data – particularly on data
regarding Indian foods - A working group under FSSAI
can take up this responsibility
• Regular interaction among industry, academia and Govt.

Thank You

12th Agricultural Science Congress
Theme: Sustainable Livelihood Security
for Smallholder Farmers
February 3-6, 2015
Organized by National Academy of Agricultural Sciences
&
National Dairy Research Institute, Karnal (Haryana), India

Dr.a.k.srivastava chemical and drug residues in dairy products (ayurvet seminar) 8.10.2014

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Dr.a.k.srivastava chemical and drug residues in dairy products (ayurvet seminar) 8.10.2014