AMITY UNIVERSITY

SUMMER TRAINING PROJECT TESTING FOR QUALITY CONTROL OF MILK

Submitted By Priyanka Goyal IMT/08/9057 Amity Institute of Biotechnology

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MOTHER DAIRY
DELHI

SUMMER TRAINING PROJECT ON

TESTING FOR QUALITY CONTROL OF MILK

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ACKNOWLEDGEMENT
It gives me immense satisfaction in completion of summer internship project entitled Testing for Quality Control of Milk. On the submission of my project report I would like to express my sincere gratitude to my guide Mr. S.K. Mittal for mentoring me & taking active interest throughout the project and for sharing his insights on the topic & for being a constant source of inspiration and courage during the entire project work. He was always available, correcting mistakes, intelligently directing me to proper sources of information advising to aim for simplicity, brevity, clarity and accuracy. I am deeply indebted to him for his support and guidance and for providing me with an environment to complete my project successfully. I would also like to thank all the laboratory staff members for sharing their immense experience and extending their support in carrying out this project work. I am greatly acknowledged for their kind help.

Priyanka Goyal

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AMITY INSTITUTE OF BIOTECHNOLOGY BONAFIDE CERTIFICATE

Certified that the training report entitled Testing for Quality Control of Milk which is being submitted by Priyanka Goyal (IMT/08/9057), in partial fulfilment of award of the degree of Int. M. Tech in Biotechnology from Amity Institute Of Biotechnology, Noida in a record of bonafide work carried out by her under my supervision and guidance, the matters embodied in this training report have been worked out independently. The matters presented in this report have not been submitted for the award of any degree, diploma or certificate.

Internal Guide

Head of Department

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BONAFIDE CERTIFICATE
Certified that this project report TESTING FOR QUALITY CONTROL OF MILK is the bonafide work of PRIYANKA GOYAL (IMT/08/9057), who carried out the project work from 23rd May 2011 to 11th July 2011 under our supervision.

SIGNATURE Mr. S.K. Mittal Deputy Manager Production Dept. Mother Dairy, Delhi

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DECLARATION
I hereby declare that the project work entitled Testing for Quality Control of Milk submitted to Mother Dairy, Delhi is a original work done by me under the guidance of Mr. S.K. Mittal and this project work have not been submitted in a part or full for any other degree or diploma of this or any other organization/institute/university.

Priyanka goyal (IMT/08/9057)

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CONTENTS
TOPIC
INTRODUCTION

PAGE NO.
10-14

REVIEW OF LITERATURE

15-25

OBJECTIVE

26

MATERIAL METHODS

27-43

RESULT

44-45

CONCLUSION

46-47

BIBLIOGRAPHY

48-53

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LIST OF TABLES
TABLE NO.
1

TITLE
PRODUCTION AND PER CAPITA AVAILABILITY OF MILK

PAGE NO.

12

WORLDS TOP 20 MILK PRODUCING COUNTRIES

12

AVERAGE COMPOSITION OF MILKS OF VARIOUS MAMMALS

21

STANDARDIZED COMPOSITION OF VARIOUS MILK TYPES IN MOTHER DAIRY

25

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LIST OF ABBREVIATIONS
R.M. Value B.R. Test SNF PPM M.B.R.T. C.O.B. Reichert Meissl Value Butyro-Refractometer Test Solids Not Fat Parts Per Million Methylene Blue dye Reduction Test Clot On Boiling

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INTRODUCTION

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INDIAN DAIRYING
India has emerged today as the largest milk producer in the world with an annual production of more than 100 million tones. Over the years, milk has been transformed from insufficient production to selfsufficient production, from rationing to plentiful availability, from loose unhygienic milk to milk that is pure from subjugation to a symbol of farmers economic independence. We cannot forget that there is something very special about milk. It requires that any brand of milk and milk products to act not simply as a seller, but as a trustee. Milk is not a white good or a brown good. Milk is not a status symbol; rather it is the symbol of nutrition. It is not only food , but an essential ingredient of life itself and by its very indispensable nature , it has one of the biggest markets, both nationally and globally. The main reasons for the world focus on India are ; one, the low cost economy and two, the continuing economic liberalization process initiated in 1991. Other important factors include: y y y y y Inexpensive labour Presence of worlds third largest pool of technical manpower Worlds largest democracy Well established independent judiciary free from government interference Ease in communication due to widespread use of English by the educated and professional class.

In 2010 , requirement for food grains touched 266 million tones which will rise to 343 million tones by 2020. For milk estimated consumption will be 271 million tones by 2020.

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TABLE NO.1

PRODUCTION AND PER CAPITA AVAILABILITY OF MILK

TABLE NO.2 WORLDS TOP 20 MILK PRODUCING COUNTRIES

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MOTHER DAIRY
Mother Dairy, Delhi was set up in 1974 under the Operation Flood Programme. It is now a wholly owned company of the National Dairy Development Board (NDDB). Mother Dairy markets & sells dairy products under the Mother Dairy brand (like Liquid Milk, Dahi, Ice creams, Cheese and Butter), Dhara range of edible oils and the Safal range of fresh fruits & vegetables, frozen vegetables and fruit juices at a national level through its sales and distribution networks for marketing food items. Mother Dairy sources significant part of its requirement of liquid milk from dairy cooperatives. Similarly, Mother Dairy sources fruits and vegetables from farmers / growers associations. Mother Dairy also contributes to the cause of oilseeds grower cooperatives that manufacture/ pack the Dhara range of edible oils by undertaking to nationally market all Dhara products. It is Mother Dairys constant endeavor to (a) Ensure that milk producers and farmers regularly and continually receive market prices by offering quality milk, milk products and other food products to consumers at competitive prices and; (b) Uphold institutional structures that empower milk producers and farmers through processes that are equitable. At Mother Dairy, processing of milk is controlled by process automation whereby state-of-the-art microprocessor technology is adopted to integrate and completely automate all functions of the milk processing areas to ensure high product quality/ reliability and safety. Mother Dairy is an ISO 9001:2008 (QMS), ISO 22000:2005 (FSMS) and ISO 14001:2004 (EMS) certified organization. Mother Dairy has Certificate of Approval from Export Inspection Council of India also. Moreover, its Quality Assurance Laboratory is certified by National Accreditation Board for Testing and Calibration Laboratory (NABL)-Department of Science and Technology, Government of India. Mother Dairy markets approximately 2.8 million liters of milk daily in the markets of Delhi, Mumbai, Saurashtra and Hyderabad. Mother Dairy Milk has a market share of 66% in the branded sector in Delhi where it sells 2.3 million liters of milk
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daily and undertakes its marketing operations through around 14,000 retail outlets and 845 exclusive outlets of Mother Dairy. The companys derives significant competitive advantage from its unique distribution network of bulk vending booths, retail outlets and mobile units. Mother Dairy ice creams launched in the year 1995 have shown continuous growth over the years and today boasts of approximately 62% market share in Delhi and NCR. Mother Dairy also manufactures and markets a wide range of dairy products that include Butter, Dahi, Ghee, Cheese, UHT Milk, Lassi & Flavored Milk and most of these products are available across the country. The company markets an array of fresh and frozen fruit and vegetable products under the brand name SAFAL through a chain of 400+ own Fruit and Vegetable shops and more than 20,000 retail outlets in various parts of the country. Fresh produce from the producers is handled at the Companys modern distribution facility in Delhi with an annual capacity of 200,000 MT. An IQF facility with capacity of around 75 MT per day is also operational in Delhi. A state-of-the-art fruit processing plant of fruit handling capacity of 120 MT per day, a 100 percent EOU, setup in 1996 at Mumbai supplies quality products in the international market. With increasing demand another state-of-the-art fruit processing plant has been set up at Bangalore with fruit handling capacity of around 250 MT per day. Mother Dairy has also been marketing the Dhara range of edible oils for the last few years. Today it is a leading brand of edible oils and is available across the country in over 2,00,000 outlets. The brand is currently available in the following variants: Refined Vegetable Oil, Refined Soybean Oil, Refined Sunflower Oil, Refined Rice Bran Oil, Kachi Ghani Mustard Oil and Filtered Groundnut Oil. Mother Dairy has also launched extra virgin Olive Oil under the Daroliva brand. Mother Dairy has over the last 3 decades, harnessed the power of farmer cooperatives to deliver a range of delicious products and bring a smile on your face. In times to come, Mother Dairy shall strive to remain one of Indias finest food companies.

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REVIEW OF THE LITERATURE

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HISTORY

1959 MILK SUPPLY IN OBERLECH, VORARLBERG, AUSTRIA

Humans first learned to regularly consume the milk of other mammals following the domestication of animals during the Neolithic Revolution[1] or the invention of agriculture. This development occurred independently in several places around the world from as early as 90007000 BC in Southwest Asia[2] to 35003000 BC in the Americas.[3] The most important dairy animalscattle, sheep and goatswere first domesticated in Southwest Asia, although domestic cattle has been independently derived from wild auroch populations several times since.[4][5] Initially animals were kept for meat, and archaeologist Andrew Sherratt has suggested that dairying, along with the exploitation of domestic animals for hair and labor, began much later in a separate secondary products revolution in the 4th millennium BC.[6] Sherratt's model is not supported by recent findings, based on the analysis of lipid residue in prehistoric pottery, that show that dairying was practiced in the early phases of agriculture in Southwest Asia, by at least the 7th millennium BC.[7][8] From Southwest Asia domestic dairy animals spread to Europe (beginning around 7000 BC but not reaching Britain and Scandinavia until after 4000 BC),[9] and South Asia (70005500 BC).[10] The first farmers in central Europe[11] and Britain[12] milked their animals. Pastoral and pastoral nomadic economies, which rely predominantly or exclusively on domestic animals and their products rather than crop farming, were developed as European farmers moved into the PonticCaspian steppe in the 4th millennium BC, and subsequently spread across much of the Eurasian steppe.[13] Sheep and goats were introduced to Africa from Southwest Asia, but African cattle may have been independently domesticated around 7000 6000 BC.[14] Camels, domesticated in central Arabia in the 4th millennium BC, have also been used as a dairy animal in North Africa and the Arabian peninsula.[15] In the rest of the world (i.e., East and Southeast Asia, the Americas
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and Australia) milk and dairy products were historically not a large part of the diet, either because they remained populated by hunter-gatherers who did not keep animals or the local agricultural economies did not include domesticated dairy species. Milk consumption became common in these regions comparatively recently, as a consequence of European colonialism and political domination over much of the world in the last 500 years. In 1863, Louis Pasteur of France developed a method of heating wine to kill the microorganisms that cause wine to turn into vinegar[16]. Later, this method of killing harmful bacteria was adapted to a number of food products and became known as pasteurization. The first milk processing plant in the United States to install pasteurizing equipment was the Sheffield Farms Dairy in Bloomfield, New Jersey, which imported a German-made pasteurizer in 1891. Many dairy operators opposed pasteurization as an unnecessary expense, and it wasn't until 1908 that Chicago became the first major city to require pasteurized milk. New York and Philadelphia followed in 1914, and by 1917 most major cities had enacted laws requiring that all milk be pasteurized. In 1884, Doctor Hervey Thatcher, an American inventor from New York, invented the first glass milk bottle, called 'Thatcher's Common Sense Milk Jar', which was sealed with a waxed paper disk.[16] Later, in 1932, plastic-coated paper milk cartons were introduced commercially as a consequence of their invention by Victor W. Farris.[16] The town of Harvard, Illinois celebrates milk with a summer festival known as "Milk Days". Theirs is a different tradition meant to celebrate dairy farmers in the "Milk Capital of the World."[17]

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DEFINITION OF MILK
Milk is defined in the Milk Ordinance and Code recommended by the U.S. Public Health Services[18] as the lacteal secretion, practically free from colostrum obtained by the complete milking of one or more healthy cows, which contains not less than 81/4% of milk solids-not-fat and not less than 31/4% of milk fat. Since this definition was promulgated as a basis for the enforcement of regulatory laws, it limits milk to that of the cow and prescribes only minimal percent of two chief components. Minimal standards in the states vary from 8.0 to 8.5% for milkSNF and from 3.0 to 3.8% for milk-fat[19]. Cow's milk has a pH ranging from 6.4 to 6.8, making it slightly acidic.[20][21] The udder, immediately after parturition, secretes a fluid known as colostrum, which differs considerably in composition from the later secretion; hence the exclusion of colostrum is above definition. Milk is the liquid food secreted by the mammary glands for the nourishment of newly born, containing water, fat, proteins, lactose and minerals (referred to as ash). An average gross composition of cows milk would be as follows[22] : water, 87%; fat, 3.5-3.7%; lactose, 4.9%; proteins, 3.5% and ash (minerals), 0.7%. In 60 specified marketing areas under U.S. Federal and State regulations in 1970, average fat content of market milk(about 90,000 samples) was 3.47% varying between 3.22 and 3.84%[23]. The term milk is also used for white colored, non-animal beverages resembling milk in color and texture such as soy milk, rice milk, almond milk, and coconut milk. In addition, a substance secreted by pigeons to feed their young is called crop milk and bears some resemblance to mammalian milk.[24]

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TYPES OF CONSUMPTION
There are two distinct types of milk consumption: a natural source of nutrition for all infant mammals and a food product for humans of all ages that is derived from other animals[46]. In almost all mammals, milk is fed to infants through breastfeeding, either directly or by expressing the milk to be stored and consumed later[47][48]. Some cultures, historically or currently, continue to use breast milk to feed their children until they are seven years old. Human infants sometimes are fed fresh goat milk. There are known risks in this practice, including those of developing electrolyte imbalances, metabolic acidosis, megaloblastic anemia, and a host of allergic reactions.[25] In many cultures of the world, especially the Western world, humans continue to consume milk beyond infancy, using the milk of other animals (especially cattle, goats and sheep) as a food product. For millennia, cow's milk has been processed into dairy products such as cream, butter, yogurt, kefir, ice cream, and especially the more durable and easily transportable product, cheese. Modern industrial processes produce casein, whey protein, lactose, condensed milk, powdered milk, and many other food-additive and industrial products. Humans are an exception in the natural world for consuming milk past infancy, despite the fact that many humans show some degree (some as little as 5%) of lactose intolerance, a characteristic that is more prevalent among individuals of African or Asian descent.[26] The sugar lactose is found only in milk, forsythia flowers, and a few tropical shrubs. The enzyme needed to digest lactose, lactase, reaches its highest levels in the small intestines after birth and then begins a slow decline unless milk is consumed regularly.[27] On the other hand, those groups who do continue to tolerate milk often have exercised great creativity in using the milk of domesticated ungulates, not only of cattle, but also sheep, goats, yaks, water buffalo, horses, reindeers and camels. The largest producer and consumer of cattle and buffalo milk in the world is India.[28]

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COMPOSITION OF MILK
The major constituents of milk are: water, fat, protein, lactose, ash or mineral matter. The minor constituents are: phospholipids, sterols, vitamins, enzymes, pigments, etc.

The true constituents are milk fat, casein and lactose[49][51]. WATER It constitutes the medium in which all the milk constitutes are either dissolved or suspended[50]. Most of it is free and only a very small portion is in bound form, being firmly by protein and phospholipids. MILK FAT The bulk of milk fat occurs in the form of globules, which average approximately 2 to 5 microns in size (range 0.1 to 22 microns)[53][55]. It is an oilin-water type of emulsion[52]. The surface of the globules is coated with a layer of material commonly known as fat globule membrane. It is composed of phospholipids and stabilizes the fat emulsion by preventing the milk fat globules from coalescing[68][69].

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Species

Water

Fat

Proteins

Lactose

Ash

SNF

Total Solids

Woman[29] Cow[30] Cow[31] Goat[32] Ewe[33] Egyptian Buffalo

[34]

87.43 87.20 86.61 87.00 80.71 82.09

3.75 3.70 4.14 4.25 7.90 7.96

1.63 3.50 3.58 3.52 5.23 4.16

6.98 4.90 4.96 4.27 4.81 4.86

0.21 0.70 0.71 0.86 0.90 0.78

8.82 9.10 9.25 8.75 11.39 9.95

12.57 12.80 13.39 13.00 19.29 17.91

Chinese Buffalo[35] Philippine Carabao[36] Indian Buffalo[37] Camel[38] Ass[39] Mare[40] Reindeer[41] Llama[42]

76.80

12.60

6.04

3.70

0.86

10.60

23.20

78.46

10.35

5.88

4.32

0.84

11.19

21.54

82.76

7.38

3.60

5.48

0.78

9.86

17.24

87.61 89.03 89.04 63.30 86.55

5.38 2.53 1.59 22.46 3.15

2.98 2.01 2.69 10.30 3.90

3.26 6.07 6.14 2.50 5.60

0.70 0.41 0.51 1.44 0.80

7.01 8.44 9.37 14.24 10.30

12.39 10.97 10.96 36.70 13.45

TABLE NO.3 AVERAGE COMPOSITION PERCENTAGE OF MILKS OF VARIOUS MAMMALS

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MILK PROTEINS Proteins are amongst the most complex organic substances. The proteins of milk consist of Casein, -Lacto globulin and -Lacto albumin[54]. It is in a colloidal state. Casein is only found in milk and exists as calcium caseinate phosphate complex. It is easily coagulated by acid, rennet, alcohol, heat and concentration. Kiermeier[43] has published an extensive review of the non-protein components of milk, including the influence of many factors of milk production upon them. More than 600 reports were evaluated of which 352 were cited[56][57]. In view of the mass of data being reported Kiermeier suggests that some systems of cataloging the data be set up. LACTOSE (MILK SUGAR) It is found only in milk and exists in true solution phase in the milk serum. It is one-sixth as sweet as sucrose[58]. On crystallization, it forms hard gritty crystals. Chemically, lactose is composed of one molecule each of D-glucose and Dgalactose[59][60]. It is fermented by bacteria to yield lactic acid and other organic acids. MINERAL MATTER (ASH) Although it is present in small quantity, it is very essential for human body[78][79]. The major salts present are of K, Na, Mg, Ca PO43-, citrate, sulphate, bicarbonate etc. It considerably influences physio-chemical properties of milk and also affects the nutritive value[67][71]. The ash content of Holstein Friesen milk in Japan[44] over a yearly period averaged 0.712% higher[45] and lower[70] than this have been reported. 0.015%. Ash contents both

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PHOSPHOLIPIDS There are three types of phospholipids (Lecithin, Cephalin[72] and Sphingomylin[74]). Lecithin is important constituent of the fat globule membrane and contributes to the richness of milk flavor of milk and other milk products[61]. PIGMENT There are two type of pigment present in milk, fat soluble and water soluble. Carotene[73] is fat soluble and is responsible for the yellow color of milk and milk products. The other two are xanthophyll and riboflavin[62][63]. Carotene also acts as an anti oxidant and as a source of Vitamin A. Riboflavin contributes to the white colour of milk[69]. MILK ENZYMES The important milk enzymes and their specific action are as follows:

Anise (diasterase) Lipase Phosphatase

Starch splitting Fat splitting, leads to rancid flavor It is capable of splitting certain phosphoric esters

Protease Peroxidases

It is capable of splitting proteins Decompose hydrogen peroxides and Catalase

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VITAMINS Vitamins are present in very minute quantities, but play a very important role in vital functioning of human body[64][65][66]. There are 25 vitamins present in milk[75][76][77]. These are fat soluble e.g. Vitamin A, D, E, K. as well as water soluble vitamins such as B-complex (B1, riboflavin or B2, pantothenic acid, niacin, pyridoxine or B6.)

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Particulars

Toned Milk (Bulk Vending Milk)

Full Cream

Standardized Toned

Double Toned

Skim Milk

Fat Protein Lactose Minerals SNF Water Calories (K.Cal) Calcium (mg) Phosphorus (mg) Vitamin A

3.05 3.10 4.76 0.70 8.56 88.37 58.00 118.00 90.00 350.00

6.10 3.55 4.80 0.75 9.10 84.80 90.00 128.00 96.00 240.00

4.60 3.20 4.70 0.70 8.60 86.80 73.00 118.00 90.00 200.00

3.10 3.20 4.70 0.70 8.60 88.30 58.00 118.00 90.00 150.00

1.60 3.55 4.80 0.75 9.10 89.30 48.00 128.00 96.00 75.00

0.05 3.40 4.70 0.70 8.80 91.15 33.00 121.00 88.00 70.00

TABLE NO.4 Standardized composition of various milk types in mother dairy

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OBJECTIVE
To learn the laboratory testing for quality control of milk.

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MATERIAL METHODS

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TESTING FOR QUALITY CONTROL OF MILK

RAW MILK RECEPTION : Milk procured from various co-operative societies is received in this section. It also includes the weighing of milk and preparation of truck sheet. The reception of milk has been illustrated by following schematic flow chart. Flow Chart for Reception of Milk : Unloading of Cans Placing the cans on the conveyor Dumping the cans of a particular society in the weighing bowl Drawing of sample from the milk of a particular society Passing the cans to can washer Trafficking of milk to the processing section with suction motor Washing the cans with water mixed with soda or detergent* Cleaning the cans by hot water at 90oC Drying the cans by circulating air at 90oC * Trisodium phosphate
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The place where the raw milk is received from the vehicles is called dock. Various chemical and organoleptic tests are conducted at this place for checking the quality of milk and termed as dock testes or platform tests. A sample of 200ml is kept in laboratory for 24 hours as a satisfying measure so that is the vendor is not satisfied with tests, then the same may be carried out again. The various testes carried out for first hand quality checking of milk are listed below : y y y y y y y y y y y y y y y y y y y ORGANOLEPTIC TETS COB ALCOHOL TEST ROSALIC ACID ACIDITY AMMONIUM COMPOUND MBRT UREA STARCH SALT SUGAR GLUCOSE MALTODEXTRIN SODIUM IONS R.M. VALUE B.R. TEST FAT PERCENTAGE SNF PERCENTAGE PROTEIN PERCENTAGE

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ORGANOLEPTIC TEST The organoleptic test permits rapid segregation of poor quality milk at the milk receiving platform. No equipment is required, but the milk grader must have good sense of sight, smell and taste. The result of the test is obtained instantly, and the cost of the test are low. Milk which cannot be adequately judged organoleptically must be subjected to other more sensitive and objective tests. Abnormal smell and taste may be caused by:
y y y y y

Atmospheric taint (e.g. barny/cowy odour). Physiological taints (hormonal imbalance, cows in late lactationspontaneous rancidity). Bacterial taints. Chemical taints or discolouring. Advanced acidification (pH < 6.4).

CLOT ON BOILING Boil a small amount of milk in a spoon, test tube or other suitable container. If there is clotting, coagulation or precipitation, the milk has failed the test. The test is quick and simple. It is one of the old tests for too acid milk(pH<5.8) or abnormal milk (e.g. colostral or mastitis milk ). If a milk sample fails in the test, the milk must contain many acid or rennet producing microorganisms or the milk has an abnormal high percentage of proteins like colostral milk. Such milk cannot stand the heat treatment in milk processing and must therefore be rejected. ALCOHOL TEST The alcohol test determines the susceptibility milk of coagulate due to developed acidity or unbalance salt. This test is of prime importance in milk to detect milk which has a tendency to curdle during processing sterilization or pasteurization. Procedure : 5 ml of raw milk is mixed with 5ml of 8% absolute alcohol, if precipitation occurs then the alcohol test is positive i.e. milk is least heat stable. Absence of any precipitation indicates appropriate heat stability of milk.

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ROSALIC ACID TEST (for neutralizers) Add 2 ml of rosalic acid solution in a test tube containing 2 ml of the milk sample subjected to testing. Rose red colour development indicates presence of neutralizers (carbonates/bicarbonates)in the powder and formation of flakes indicates disturbed salt balance i.e., alcohol test positive. Light rose red colour or light brown colour formation indicates absence of neutralizers. ACIDITY Ten ml of milk is mixed well with as much amount of distilled water in a flask and the 1-2 drops of phenolphthalein indicator are added. Then N/10 NaOH is run down from a burette to the content is the flask until a light pink color appears. The amount of N/10 NaOH utilized is noted down. 1 ml of N/10 NaOH solution is equivalent to 0.01 g of lactic acid. Good quality milk has 0.12-0.15% lactic acid, sour milk has 0.15-0.20% lactic acid and curded milk has 0.7 to 2.0% lactic acid. AMMONIUM COMPOUND To 1 ml of milk sample, add 2 ml of Neslers reagent. Formation of orange colour with brown tinge indicates the presence of ammonia in the milk. Neslers reagent make 3 solutions of the following : 8 grams of mercuric chloride in 150ml of distilled water ; 60 grams of sodium hydroxide in 150ml of distilled water ; 16 grams of potassium iodide in 150ml of distilled water. Add mercuric chloride and sodium hydroxide solutions and mix well. Then to this mixture add potassium iodide solution and make the volume upto 500ml. MBRT (METHYLENE BLUE REDUCTION) MBRT is one of the most important tests for quality assessment of milk. It is an indicator of shelf life or keeping quality of milk in addition of checking whether milk is properly pasteurized or not. To determine the test, 10ml milk is taken in a test tube and to it 1 ml of methylene blue dye is mixed. The content is heated and kept in a water bath at a temp. of 37oC. Methylene blue dye is prepared in oxidized form. Bacteria Present in milk reduce this dye in a short time. So if blue colour in the test tube (dye+milk) is changed to white in short time, this shows that larger count of bacteria is still present in the milk which is attributed to improper pasteurization.
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UREA Urea is a natural constituent of milk and is present to the extent of 70 mg per 100 ml (700 ppm). The test based on the use of para dimethyl amino benzaldehyde can be followed for determination of added urea. 5 ml of milk is mixed with 5 ml of 1.6 % DMAB prepared by dissolving 1.6 gm of p Dimethyl amino benzaldehyde in 100 ml of 10 % HCl. Distinct yellow colour is observed in milk containing added urea. The control (normal milk) shows a slight yellow colour due to presence of natural urea. STARCH Take about 5 ml of milk in a test tube. Bring to boiling condition and allow the test tube to cool to room temperature. Add 1-2 drops of iodine solution to the test tube. Development of blue colour indicates presence of starch which disappears when sample is boiled and reappears on cooling. SALT Take 5 ml of silver nitrate (0.134%) solution in a test tube and add 2 drops of potassium chromate (10%) solution in it. Brick red colour formation is observed. To this add 1 ml of milk and mix well. Observe the colour, if yellow colour is obtained then it indicates the presence of added salt. SUGAR Take 3 ml of milk sample in a test tube and add 5 ml dilute HCl (1:2) containing resorcinol (0.1 grams resorcinol dissolved in 100 ml of dilute HCl). Gently mix the test tube and keep it in boiling water bath for 5 minutes. Formation of brick red colour indicates presence of sugar. Sugar is added to increase the density of milk to prevent the detection of added water. GLUCOSE To 1 ml of milk sample in a test tube add equal volume of acetate buffer and filter. To 0.2 ml of filterate add 2.8 ml water and 2 ml of barfords reagent. (1) Heat the tube in boiling water for 4 minutes. After cooling for 2 minutes add 3 ml of barfords reagent (2) and mix the contents. Development of deep blue colour indicates the presence of glucose. Filter the contents of the tube through Whatman No. 42 filter paper. Collect the filtrate in a colorimetric tube, after
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discarding first 1 ml. Measure the absorbance in a photoelectric colorimeter, using red filter or determine absorption maxima in a spectrophotometer between 620- 780 um against blank prepared identically from a pure milk sample. The concentration of glucose in the sample can be determined with the help of a standard curve prepared from milk samples containing known amounts of added glucose i.e., 0.5, 1.0, 2.0, 5.0 percent glucose in milk. MALTODEXTRIN Maltodextrin is a type of dextrin, a non-sweet nutritive saccharide polymer that consist of D-glucose. Take 20 ml of homogenous milk sample in a 100ml beaker. First the sample will be checked for the presence of glucose using Diastix strip. Hold the plastic end of the strip without touching the green coloured reagent area. Dip this reagent area into the milk sample and gently remove the excess milk sample from the strip by single jerk. Compare the colour change from the colour chart exactly after 30 seconds. If sample shows presence of glucose then maltodextrin test is not conducted. If glucose is absent then the pH of the sample is adjusted to 4-4.5 by adding dilute lactic acid. Usually 0.8 to 1.2 ml of lactic acid is used to maintain pH in this range. Then 1 ml of enzyme solution is added and finally the sample is incubated at 62 2oC temperature for 5minutes. Sample is then cooled to room temperature and again checked for glucose with diastix strip. Change in colour of the strip indicates presence of glucose but after enzyme treatment indicates presence of maltodextrin and sample is rejected. SODIUM IONS It is done by direct measurement with a sodium ion selective electrode. Take 25 ml milk in a 50 ml beaker and add 2.5 ml of ISA to it. Add the magnetic stirrer and place the electrode in the sample. Sample is rejected if sodium ions are present more than 600ppm. R.M. VALUE (REICHERT-MEISSL VALUE) It may be defined as the number of milliliters of 0.1 N KOH solution required to neutralize the volatile, soluble fatty acids distilled from 5 g of fat under specified conditions. It is primarily a measure of butyric acid. The value for milk fat ranges between 17 and 35 and this value is well above that for all other fats and oils. A weighed amount of fat is completely saponified with alkali (KOH solution). The resulting solution is then acidified with dilute sulphuric acid and then steam
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distilled. The distillate is titrated against 0.1N KOH solution and RM value is calculated. R.M. Value = no. of ml of 0.1 N KOH 5/ weight of oil or fat B.R. TEST (BUTYRO REFRACTOMETER TEST) Extract ghee from the milk sample and put few drops of melted ghee on cleaned prism of the B.R. which is maintained at 40oC. allow it to stand for 1 minute and then note down the reading. Milk is accepted only when the reading comes in the range of 40 43. FAT PERCENTAGE GERBER METHOD Principle : The milk is mixed with sulphuric acid and iso-amyl alcohol in a special Gerber tube, permitting dissolution of the protein and release of fat. The tubes are centrifuged and the fat rising into the calibrated part of the tube is measured as a percentage of the fat content of the milk sample. The method is suitable as a routine or screening test. It is an empirical method and reproducible results can be obtained if procedure is followed correctly. Procedure : Measure 10 ml of sulphuric acid into a butyrometer tube, preferably by use of an automatic dispenser, without wetting the neck of the tube. Mix the milk sample gently but thoroughly and fill the milk pipette above the graduation line. Wipe the outside of the pipette and allow the milk level to fall so that the top of meniscus is level with the mark. Run the milk into the butyrometer tube along the side wall without wetting the neck, leave to drain for three seconds and touch the pipette's tip once against the base of the neck of the butyrometer tube. Add 1 ml of Amyl alcohol, close with a lock stopper, shake until homogeneous, inverting it for complete admixture of the acid. Keep in a water bath for 5 min. at 652oC taking care to have casein particles if any to dissolve fully, and centrifuge for 4 min. at 1100 rpm. The tubes should be put in centrifuge, so as to conform to radial symmetry, and as evenly spaced as possible, in order to protect bearings of the centrifuge. Allow the centrifuge to come to rest. Remove the butyrometer tubes and place in water bath for 5 minute at 652oC . Read the percentage of fat after adjusting the height in the tube as necessary by movements of the lock stopper with the key. Note the scale reading corresponding to the lowest point of the fat meniscus and the surface of separation of the fat and acid. When readings are being taken hold the butyrometer with the graduated portion vertical, keep the point being read in level with the eye, and then read the butyrometer to the nearest half of the smallest scale division.
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BUTYROMETER USED IN THE GERBER TEST FOR FAT PERCENTAGE DETERMINATION

SNF PERCENTAGE The estimates the amount of solid-not-fat (SNF) present in the milk sample. For this purpose Lactometer is used. Prior to filling the metallic cylinder upto brims with milk sample the temperature of milk (29 oC) is noted. The lactometer is then inserted inside the cylinder containing milk, and the temperature of the milk is noted again. The scale of lactometer projecting above the milk gives the lactometer reading. If the temperature of the milk is 29oC then the lactometer reading itself is taken as CLR and if the temp. is below or above 29 oC, then for each degree rise or fall of temp. o.5 is added or deducted from the observed lactometer reading. Solid-not-fat is determined by using following expression. SNF = CLR/4 + 0.21 fat + 0.66
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Suppose CLR is = 28.0 and Fat% = 6% then. SNF = 28/4 + 0.21 x 6 + 0.66 = 7 + 1.26+0.66 SNF = 8.92% PROTEIN PERCENTAGE Principle : Total nitrogen and non protein nitrogen content of test sample is determined separately. Subtract non protein nitrogen content from total nitrogen content, in the sample and multiply by 6.38 to get milk protein nitrogen content. Procedure : Weigh accurately about 5 gm of sample and transfer to the Kjeldahl flask. Digest with sulphuric acid by using Copper sulphate as catalyst and potassium sulphate as boiling point elevator to release nitrogen from protein and retain nitrogen as ammonium salt. Concentrated NaOH is added to release ammonia which is absorbed in HCl and back titrated. Multiply percent nitrogen by 6.38 to calculate total protein nitrogen. For determining non protein nitrogen, protein is precipitated by addition of trichloro acetic acid (TCA) solution. Final concentration of TCA in the mixture is about 12 %. Precipitated milk protein is removed by filteration. Filterate contains non protein nitrogen components of milk. Nitrogen content of filterate is determined by Kjeldahl method Reagents : (a) Trichloro acetic acid solution - 15% (w/v). CCl3COOH TCA is soft deliquescent crystal which should be stored in container protected from light and moisture (b) Hydrochloric acid standard solution 0. 01 M HCl Procedure : Reconstitute sample ( about 5-10 gm) with warm water at 40 oC and make upto 100 ml. Pipette 10 ml into a preweighed 125 ml erlenmeyer flask and weigh. Add 400.5 ml TCA in the flask. Weigh flask and contents. Swirl to mix. Let precipitate settle for 5 minutes. Filter through Whatman No1 paper or equivalent, 15 cm, Nitrogen free, and collect entire filterate. Filterate should be clear and free from particulate matter. If it is not, repeat entire sample preparation. Swirl filterate to mix. Pipette 200.2 ml into a 50 ml beaker, and weigh. Pour filterate from beaker into Kjeldahl digestion flask that contains boiling chips, potassium sulphate and copper sulphate. Immediately reweigh empty beaker. Add H2SO4, and digest. Prepare a blank (16 ml of 15 % TCA and no test sample) and keep record of blank value.

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Calculate percent nitrogen which is non protein nitrogen expressed as protein equivalent. Protein equivalent % = Nitrogen % 6.38 Calculation Substract total non protein nitrogen expressed as protein equivalent from total protein to determine milk protein in Milk. Protein percent = total protein % total non protein nitrogen (protein equivalent %)

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BACTERIOLOGY OF MILK
Milk is synthesized by cells within the mammary gland and is virtually sterile when secreted into the alveoli of the udder. Beyond this stage of milk production, bacterial contamination can generally occur from three main sources; within the udder, outside the udder, and from the surface of equipment used for milk handling and storage. Cow health, environment, milking procedures and equipment sanitation can influence the level of microbial contamination of raw milk. Equally important is the milk holding temperature and length of time milk is stored before testing and processing that allow bacterial contaminants to multiply. All these factors will influence the total bacteria count (SPC) and the types of bacteria present in raw bulk tank milk.

MICROBIAL CONTAMINATION FROM WITHIN THE UDDER

Raw milk as it leaves the udder of healthy cows normally contains very low numbers of microorganisms and generally will contain less than 1000 colonyforming units of total bacteria per milliliter (cfu/ml). In healthy cows, bacterial colonization within the teat cistern, teat canal, and on healthy teat skin does not significantly contribute total numbers of bacterial neither in bulk milk, nor to the potential increase in bacterial numbers during refrigerated storage. This natural flora of the cow generally will not influence the SPC, PI, LPC, or Coliform counts. While the healthy udder should contribute very little to the total bacteria count of bulk milk, a cow with mastitis has the potential to shed large numbers of microorganisms into her milk. The influence of mastitis on the total bacteria count of bulk milk depends on type of bacteria, the stage of infection and the percent of the herd infected. Quarters from infected cows have the potential to shed in excess of 10,000,000 bacterial cfu/ml of milk produced. Mastitis organisms found to most often influence the total bulk milk bacteria counts are Streptococci (primarily Strep agalactiae and Strep uberis) although other mastitis pathogens have the potential to influence the bulk tank count as well. Staphylococcus aureus is not thought to be a frequent contributor to total bulk tank counts although counts as high as 60,000/ml have been documented. While Staph aureus and Strep ag are rarely found outside of the mammary gland, environmental mastitis pathogens (Strep uberis and coliforms) can occur in milk as a result of other contributing factors such as dirty cows, poor equipment cleaning and/or poor cooling. Increases in SCC can sometimes serve as supportive evidence

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that mastitis bacteria may have caused increases in the bulk tank counts. This correlation seems to apply more for Streptococci than for Staph aureus. Correlations between increases in somatic cell counts and other environmental mastitis organisms, including coliform bacteria, and coagulase-negative Staphylococci, were found to be poor as well. Staph aureus and Strep ag do not grow significantly on soiled milking equipment or under conditions of marginal or poor milk cooling. In general, mastitis organisms will not influence PI or LPC though in some cases of coliform mastitis, Coli counts may be elevated.

MICROBIAL CONTAMINATION FROM OUTSIDE THE UDDER

The exterior of the cows udder and teats can contribute microorganisms that are naturally associated with the skin of the animal as well as microorganisms that are derived from the environment in which the cow is housed and milked. In general, the direct influence of natural inhabitants as contaminants in the total bulk milk count is considered to be small and most of these organisms do not grow competitively in milk. Of more importance is the contribution of microorganisms from teats soiled with manure, mud, feeds or bedding. Teats and udders of cows inevitably become contaminated while they are lying in stalls or when allowed in dirty lots. Used organic bedding has been shown to harbor large numbers of microorganisms often exceed 100,000,000 to 10,000,000,000 per gram of bedding. Organisms associated with bedding materials that contaminate the surface of teats and udders include streptococci, staphylococci, spore-formers, coliforms and other Gram-negative bacteria. Both thermoduric and psychrotrophic strains of bacteria are commonly found on teat surfaces indicating that contamination on the outside of the udder can influence PI, LPC, and Coli counts. The influence of dirty cows on total bacteria counts depends on the extent of soiling of the teat surface and the udder prep procedures employed. Milking heavily soiled cows could potentially result in bulk milk counts exceeding 10,000 cfu/ml. Several studies have investigated premilking udder hygiene techniques in relation to the bacteria count of milk. Generally, thorough cleaning of the teat with a sanitizing solution (predip) followed by thorough drying with a clean towel is effective in reducing the numbers of bacteria in milk contributed from soiled teats.

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MICROBIAL CONTAMINATION FROM EQUIPMENT CLEANING AND SANITIZING PROCEDURES

The degree of cleanliness of the milking system probably influences the total bulk milk bacteria count as much, if not more, than any other factor. Milk residue left on equipment contact surfaces supports the growth of a variety of microorganisms. Organisms considered to be natural inhabitants of the teat canal and teat skin are not thought to grow significantly on soiled milk contact surfaces or during refrigerated storage of milk. This generally holds true for organisms associated with contagious mastitis (Staph aureus and Strep ag) though it is possible that certain bacteria associated with environmental mastitis (coliforms) may be able to grow significantly. In general, bacteria from environmental contamination (bedding or manure) are more likely to grow on soiled equipment surfaces. Water used on the farm might also be a source of bacteria, especially psychrotrophs, which could seed soiled equipment. Cleaning and sanitizing procedures can influence the degree and type of bacterial growth on milk contact surfaces by leaving behind milk residues that support growth, as well as by setting up conditions that might select for specific microbial groups. Even though equipment surfaces may be considered efficiently cleaned with hot water, more resistant bacteria (thermodurics) may endure in low numbers. If milk residue is left behind (milk stone) growth of these types of organisms, although slow, may persist. Old cracked rubber parts are also associated with higher levels of thermoduric bacteria. Significant build-up of these organisms to a point where they influence the total bulk tank count may take several days to weeks though increases would be detected in the LPC. Less efficient cleaning, using lower temperatures and/or the absence of sanitizers tends to select for the faster growing, less resistant organisms (psychrotrophs), principally Gram-negative rods (coliforms and Pseudomonas) and some Streptococci. This will result in a high PI and in some case an elevated LPC. Effective use of chlorine or iodine sanitizers has been associated with reduced levels of psychrotrophic bacteria that cause high PI counts. Psychrotrophic bacteria tend to be present in higher bacteria count milk and are often associated with neglect of proper cleaning or sanitizing procedures and/or poorly cleaned refrigerated bulk tanks.

MILK STORAGE TEMPERATURE AND TIME

Refrigeration of raw milk, while preventing the growth of non-psychrotrophic bacteria, will select for psychrotrophic microorganisms that enter the milk from soiled cows, dirty equipment and the environment. Minimizing the level of contamination from these sources will help prevent psychrotrophs from growing to
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significant levels in the bulk tank during the on-farm storage period or at the processing plant. In general these organisms are not thermoduric and will not survive pasteurization. The longer raw milk is held before processing (legally up to 5 days), the greater the chance that psychrotrophs will increase in numbers. Holding milk near the PMO legal limit of 45C allows much quicker growth than milk held below 40C. Although milk produced under ideal conditions may have an initial psychrotrophic population of less than 10% of the total bulk tank count, psychrotrophic bacteria can become the dominant bacteria after 2 to 3 days at 40C, resulting in a significant influence on PI counts. Colder temperatures 3436C will delay this shift, though not indefinitely. Under conditions of poor cooling with temperatures greater than 45C, bacteria other than psychrotrophs are able to grow rapidly and can become predominant in raw milk. Streptococci have historically been associated with poor cooling of milk. These bacteria will increase the acidity of milk. Certain bacteria are also responsible for a "malty defect" that is easily detected by its distinct odor. Storage temperatures greater than 60C tend to select for these types of contaminants. The types of bacteria that grow and become significant will depend on the initial contamination of the milk. Once milk leaves the farm, raw milk handling as well as the sub-sample collected by the milk hauler is beholden to the same sets of rules. If the raw milk or the sample used to run the regulatory tests are maintained at the proper temperature, bacterial counts can be significantly altered. Since most bulk tank milk is commingled in an over-the-road milk truck, the only way to determine each producers contribution to the commingled milk is the sub-sample collected at pickup. Sample integrity must be maintained from the original bulk tank of milk, through hauling to the milk processor, and eventually through the end of the diagnostic procedures. As more milk processors are using bacteria counts within their premium programs, competent subsample handling is essential. In addition, much like SCC, one or two samples within a 30-day span are inadequate to provide a proper measure of a farms management practices.

TESTS FOR BACTERIA IN RAW BULK TANK MILK

STANDARD PLATE COUNT The Standard Plate Count (SPC) of raw milk gives an indication of the total number of aerobic bacteria present in the milk at the time of pickup. Milk samples are plated in a semi-solid nutrient media and then incubated for 48 hrs at 32C (90F) to encourage bacterial growth. Single bacteria (or clusters) grow to become visible colonies that are then counted. All plate counts are expressed as the number
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of colony forming units per milliliter (cfu/ml) of milk. Newer filmsbased tests have allowed for automation of this procedure. The nutrient agar system remains the gold standard. Aseptically collected milk from clean, healthy cows generally has an SPC less than 1,000 cfu/ml. Higher counts suggest bacteria are entering the milk from a variety of possible sources. Though it is impossible to eliminate all sources of contamination, counts of less than 5,000 cfu/ml are possible, while counts of 10,000 cfu/ml should be achievable by most farms. The most frequent cause of a high SPC is poor cleaning of the milking system. Milk residues on equipment surfaces provide nutrients for growth and multiplication of bacteria that contaminate the milk at subsequent milk times. Other procedures that can elevate bulk-tank SPC are milking dirty udders, maintaining an unclean milking and housing environment, and failing to rapidly cool milk to less than 40F. On rare occasions, cows with mastitis can shed infectious bacteria into the milk can also cause a high SPC. In these circumstances, a concurrent elevation in SCC should be evident. PRELIMINARY INCUBATION COUNT The Preliminary Incubation Count (PI) reflects milk production practices. This procedure involves holding the milk at 55F for 18 hours prior to plating. This step encourages the growth of groups of bacteria that grow well at cool temperatures (psychrotrophs). Bacteria in the incubated sample are counted with the SPC procedure and compared to the SPC from the same sample to determine if a significant increase has occurred. PI counts are generally higher than the SPC. Counts with a 3-4-fold increase are considered significant. Some consider counts greater than 50,000 cfu/ml to be of concern regardless of the SPC, though in some cases the counts may be equal and in rare cases the PI may be lower. High PI counts are most often associated with a failure to thoroughly clean and sanitize either the milking system or the cows. Bacteria considered to be natural flora of the cow, including those that cause mastitis, are not thought to grow significantly at the PI temperature. However, PI equal to or slightly higher than a high SPC (greater than 50,000 cfu/ml) may suggest that the high SPC is possibly due to mastitis pathogens. Marginal cooling (milk held at temperatures over 40F) or prolonged storage times may also result in unacceptable PI levels by allowing organisms that grow at refrigeration temperatures to multiply. LAB PASTEURIZED COUNT Though most bacteria are destroyed by pasteurization, there are certain types and certain bacterial stages that are not. The Lab Pasteurized Count (LPC) estimates the number of bacteria that can survive the pasteurization process. Milk samples are heated to simulate batch pasteurization at 145F for 30 minutes. Bacteria that
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survive pasteurization (thermodurics) are then counted using the SPC procedure. LPC are generally much lower than SPC, with counts greater than 300 considered high. Bacteria found in the natural flora of the cow, as well as those associated with mastitis, are generally not thermoduric. High LPC values are most often associated with a chronic or persistent cleaning failure in some area of the system or significant levels of contamination from soiled cows. Other common causes of high LPC are leaky pumps, old pipeline gaskets, inflations and other rubber parts, and milk stone deposits. COLIFORM COUNT The Coliform Count (Coli Count) procedure selects for bacteria that are most commonly associated with manure or environmental contamination. Milk samples are plated on a selective nutrient media that encourages the growth of coliform bacteria, while preventing the growth of others. Although coliforms are often used as indicators of fecal contamination, there are strains that commonly exist in the environment. Coliforms may enter the milk supply as a consequence of milking dirty cows or the claw becoming soiled with manure during milking. Generally, counts greater than 100 cfu/ml would indicate poor milking hygiene or other sources of contamination. Higher coliform counts more often result from dirty equipment and in rare cases result from milking cows with environmental coliform mastitis.

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RESULT

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Mother Dairy maintains stringent measures to ensure the quality and purity of the milk provided to its consumers. Each batch of incoming and outgoing milk is subjected to 21 quality tests including presence of foreign matter and bacteriological tests. The quality of milk accepted and dispatched meets certain predetermined standards. The milk goes through various processes such as Clarification, Homogenization, Standardization and Pasteurization, to ensure that it is safe for human consumption. Their motive for following such strict quality measures is to ensure that there is no contamination while processing or packaging. Mother Dairy promises its consumers that it will continue to produce products of the highest quality standard. Its Dairy products are processed & packed in ISO certified plants & strict controls are exercised by quality assurance department on all the plants. Our plants are certified for FOOD SAFETY MANAGEMENT SYSTEM ISO 22000. Mother Dairy Quality Assurance Laboratory is certified by National Accreditation Board for Testing and Calibration Laboratory (NABL)-Department of Science and Technology, Government of India. All the operations in this organization are manned by qualified & highly experienced personnel. Mother Dairy not only focus on having a motivated & hard working sales team but also focus on having a team of well qualified & trained personnel operating on field .This team operating on field ensures that the products manufactured at the plant are handled properly once the products leave the factory, to avoid any type of spoilage of the product.

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CONCLUSION

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Mother Dairy is committed towards delivering products which meet all regulatory, industrial, consumer quality and food safety standards to our valued consumers. We at Mother Dairy are focused on building a sustainable Quality and Food Safety Program using state of the art processes and innovative technologies towards delivering wide range of Dairy & Food products with best in class quality. Mother Dairy systems are designed to have process monitoring and controls at each stage of food chain. It is our endeavor to create a culture of Total Quality where continuous improvement of our people, processes and products becomes a way of life. At its Innovation Centre we have a dedicated team of Research Scientist who are constantly collaborating international standards and best practices for Quality & Food Safety in its products.

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BIBLIOGRAPHY

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