CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS

PRADESHIK COOPERATIVE DAIRY FEDERATION LTD. 29, PARK ROAD, LUCKNOW

Training Period: June 1st, 2012 July 15th, 2012

TRAINING INCHARGE: Mr. Satish Yadav

Asst. Chemist Central lab, PCDF

SUBMITTED BY: Kshitiz C. Srivastava

Dept. of Biotechnology Enrollment No: 09101038

JAYPEE INSTIUTE OF INFORMATION TECHNOLOGY NOIDA

CONTENT
S No Topic 1. PCDF : ABOUT THE COMPANY ORGANIZATION STRUCTURE 2. 3. 4. 5. DIFFERENT SECTIONS PROCESS EQUIPMENTS INTRODUCTION : MILK INDUSTRY PROCESSING OF MILK AT PARAG DAIRY ANALYSIS OF MILK PRELIMINARY TEST QUALITY TEST ADULTRATION TEST MILK PRODUCT ANALYSIS BUTTER CURD PANEER PEDA GHEE MICROBIOLOGICAL ANALYSIS MBR TEST COLIFORM TEST STANDARD PLATE COUNT WATER QUALITY TEST OIL & GREASE TOTAL HARDNESS BOD COD Page No. 5 6 7 8 9

6.

10 11 12

7.

13

8.

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CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

ACKNOWLEDGEMENT
It gives me immense pleasure and satisfaction to state that my project Analysis of milk and milk products has been successfully completed while undergoing training at Pradeshik Cooperative Dairy Federation Ltd. (PARAG), Lucknow. I express my sincere gratitude to my esteemed and reverend Mr.Satish Yadav (Asst. Chemist), PCDF, Lucknow, U.P. under whose guidance this project was completed successfully. My certain thanks to Mrs. Sunita Sahu, Mrs.Rekha Rani(Asst. chemist), Mr.Surendra Singh (Asst. Chemist), Mrs.Kalpana (Asst. Chemist), Mr. S.K. Prajapati (Lab Asst.), and all other staff of Parag, Lucknow for supporting and guidance in preparing this project.

- Kshitiz C. Srivastava

CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

DECLARATION
I Kshitiz Chandrika Srivastava, hereby declare that the research work presented in the report entitled Chemical and Microbiological Analysis of Milk & Milk Products is to fulfill the requirement of degree of Btech. Biotechnology. The report includes original work and studies carried out by myself and the content of the project do not form the basis for the degree to me or anybody else.

Signature of Student: ... Name of Student: Enrollment No: Kshitiz Chandrika Srivastava 09101038

CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

PRADESHIK COOPERATIVE DAIRY FEDERATION LTD. (PARAG)

ABOUT THE COMPANY PCDF was formed in 1962 with the aim to develop organized dairying in the State on Cooperative lines, which is a cohesive body that successfully established direct link between the producer and the ultimate consumer. This Apex Milk Cooperative draws its inherent strength from the farmers committed participation, and injects corporate skills and dynamic professionalism into what is fundamentally a traditional institution.

Uttar Pradesh is the largest milk producing state of India contributing 17 % of the total milk production of India. In the year 2010-2011, the total milk production in the state was 21033.3 thousand kg. per day. Successful efforts gave way to formation of Lucknow Milk Union in 1938-the only Milk Union in the country -giving Uttar Pradesh the credit of being a pioneer state in the country in this segment. PCDF was the chosen agency to implement the World Banks prestigious Operation Flood programme in the state. At present PCDF lends its support and services to 6,00,000 rural milk producers through 59 District Milk Unions and about 13,500 Village Dairy Cooperatives in the State PARAG is the brand name for a range of milk and milk products including- Milk, Skimmed Milk Powder, Whole Milk Powder, Butter , Ghee, and an array of indigenous milk products like Paneer,Curd,Peda,Milk Cake, Khoa, Laddu, Mattha and Chhachh etc. A Jersey Cattle breeding unit is located in Rae Bareilly, for rearing of Jersey Bulls. At FFHC Unit in Moradabad, Frozen Semen Doses of Good quality breed of milch animals are prepared for Artificial Insemination Services. Fodder Seed Processing Plant located in Aligarh, produces good quality Fodder Seeds for distribution to farmers for cultivation of good quality fodder feed for milch animals. ORGANISATION STRUCTURE

DAIRY MINISTER MANAGING DIRECTOR CHIEF GENERAL MANAGER GM ADMINISTRATION SECTIONAL HEADS

CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

DIFFERENT SECTIONS: PCDF

1. Regeneration Section: This section is used for transferring heat from warm milk to chilled milk. It consists of different sections i.e., regeneration 1, 2& 3. 2. Heating Section: In this section, milk is heated upto 71.70C for 15 sec., At this temp., bacteria like mycobacterium tuberculi or coxielleaburnett gets destroyed. 3. Holding Section: Complete pasteurization is done in this section. 4. Chilling Section: Here milk is cooled upto 50C.

PROCESS EQUIPMENT
Chiller: In this equipment milk is cooled upto 5C. Capacity > 20,000lt/hr. Silo Tanks: Milk from chiller is stored in silo tanks. They are completely strong insulated tanks. There are three silo tanks in Parag Dairy: Tanks 1. 2. 3. Capacity 5,000 lt. 10,000 lt. 30,000 lt.

Cream Separator: It is based on the principle of centrifugation. Law of centrifugation is based upon density of particles. By this method, cream is separated from raw milk Ghee Kettle: Ghee is prepared from cream. It is a double jacket steel vessel. There are three valves- Safety valve, Tube valve, and Water valve. It has agitator on the upper side and supply of steam from boiler. Churner: In churner, cream is rotated at high speed with milk to form butter. Boiler: They help in pasteurization. They produce steam which is utilized for pasteurization and by ghee kettle, etc. Boiler Type: Fire tube horizontal boiler. Refrigeration: This device uses liquid ammonia for chilling water. Chilled water is transferred throughout the dairy through pipelines. Milk Pasteurizer: In this raw milk is heated to the temp. above 700C for destructing pathogenic microbes in the milk. Homogenizer: These are used for processing recombined milk during lean; period by adding liquefied fat. Here fat globules are mixed by high pressure.

CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

INTRODUCTION: MILK
Milk is a whole, fresh, clean, lacteal secretion obtained by complete milking of one or more healthy milk animals, excluding that obtained within 15 days before and 5 days after calving or such periods as may be necessary to render milk practically colostrums free and containing minimum prescribed percentages of milk fat and milk solids not fat(SNF). The price of milk is determined by its fat and SNF contents. Milk is an ideal food for all. It is rich in carbohydrate like lactose, proteins mainly casein and fat. Further minerals like calcium, phosphorus, sodium, potassium and magnesium and present in appreciable quantities. Milk provides considerable quantity of thymine, riboflavin, niacin, pyridoxine, pantothenic acid, biotin, folic acid and vitamin B12. MILK COMPOSITION CONSTITUENTS Water Total Solids Fat SNF Proteins Lactose Ash COW MILK % 86.36 13.64 4.90 8.74 3.40 4.60 0.74 BUFFALO MILK % 83.22 16.78 7.30 9.49 3.80 4.90 0.78

PROPERTIES OF MILK Acidity pH Density Avg. Specific gravity Microbial load

7

0.12 0.15 6.4 6.8 15.6 16.0 1.028 1.032 30,000/ml

CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

PROCESSING OF MILK AT PARAG DAIRY

Raw milk receiving dock Grading samples and testing Weighing and filtration Chilling (5C or below) Raw milk storage

Pasteurization (78 C +/-2C) HTST

Cream Separator

Cream (White Butter) Pasteurized milk storage Cream Storage Tank Pouch machine balance tank

Crating and Cold Storage

STANDARDIZATION OF MILK Milk is standardized in three standard packaging of 500ml/1000ml pouch at Parag Dairy: 1) Full Cream Milk 2) Toned Milk 3) Skimmed Milk S No.
1. 2.

Content
FAT% SNF%

Full Cream Milk

6% 9%

Toned Milk
3% 8.5%

Skimmed Milk
NIL 8.7%

CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

ANALYSIS OF QUALITY OF MILK

PRELIMINARY TEST
S no 1. 2. 3. Test Organoleptic Clot on boil Phosphotase Protocol Check the colour, appearance, texture, odour and flavour of the product. Take small sample of milk and boil. Take 1ml of pasteurized milk in a test and add phosphotase solution. Keep it in a water bath at 37 C for 5 min. ( To test correctness of pasteurization). Observations Preliminary test to observe any physical change in the product. Formation of clots indicates increase of acidity. Yellow colour indicative of presence of phosphotase enzyme, i.e. presence of raw milk in past. sample.

QUALITY TEST S No.

1.

Content
Acidity

Test
Take 9 ml milk sample in a test tube. Add few drops of phenolphthalein indicator. Titrate it by N/10 NaOH till light pink color. Record the reading. GERBERs TEST Take 10ml of H2SO4 in a lock stopper burette. Add 10.75ml of milk and 1ml isoamyl alcohol. Centrifuge the reaction mix at 12000 rpm for 3min. Record the reading. Warm milk to 40-45C for 5 min in a water bath and cool it to 29C(for pasteurized milk) / 15.5C (for raw milk). Transfer the sample to lactometer. Put the lactometer jar by holding from the stem slowly.

Observations
Acidity of Milk= 9 NV/W Standard range: 0.10-0.17 N= 0.1, V= 1.5, W= 9ml Acidity = 0.15 Standard range = 4-6% Reading = 5.5 % Fat = 5.5%

2.

Fat%

3.

SNF%

Standard range = 9% SNF% = CLR+ 0.2*Fat% + 0.50 (raw milk) SNF% = CLR+ 0.2*Fat%+0.29(pas. milk) CLR =LR+ (temp. of milk)/3 or 5.

ADULTERATION TEST
S No. 1. 2. 3. Adulterant CAUSTIC SODA UREA SUGAR Test 5ml milk + 5ml Rosalic acid(1%) 5ml milk + 5ml DMAB 3ml milk + 6ml Resorcinol Observations Pink color, test +ve. Orange color, test ve. Bright yellow color, test +ve. Light yellow color, test ve. Brick red color, test +ve.

CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

4. 5. 6.

STARCH SALT GLUCOSE

3ml milk + 1% Iodine sol. (3 drops) 5ml milk + 0.5 Potassium chromate+ 0.1342% Silver nitrate. 1ml milk + 1ml bradfoeds reagent (boil for a min.) + 1ml phosphomolybdic acid 5ml milk + Neslers Reagent 1ml milk + 1ml conc. HNO3

Blue color, test +ve. No color change, test ve. Brown color, test +ve. Yellow color, test ve. Dark blue color, test +ve Slight blue color, test -ve Brown color, test +ve Black color, test -ve White solution, test +ve transparent solution test ve

7. 8.

DAP POSTER COLOR

MILK PRODUCT ANALYSIS

a) BUTTER Process Flow Chart:
Receiving cream Sterilization Standardization (35%-40% Fat) Pasteurization( 82C- 83C)

Packaging & Storage (5-10C)

Salting

Washing

Churning

Aging (5-10C)

Cooling (5-10C)

Analysis of quality of butter: Standards for table butter: 80% fat + 3% salt + 16% moisture +1%curd + anneto (natural color). S No.
1.

Content
Moisture%

Test
Heat 10g of butter over hot plate and remove the moisture by continuous heating. Place the beaker in desiccator to absorb the moisture. Measure the final and initial weight of the butter. Take the precipitated sample without moisture. Wash the sample three times with 20ml petroleum ether and decant it. Measure the weight of the sample. Dissolve the ppt. in distilled water. Make up vol. upto 250 ml and take 17.6 ml of this solution. Add 0.5ml of K2Cr2O4 and titrate with 0.1204 N AgNO3 till brick red colouration.

Observation
Initial wgt. = 64.64g Final wgt. = 62.86 Total moisture = 1.77 %Moisture = (1.77/10)*100 = 17.7% Initial wgt. = 64.64g Final wgt. = 54.63g %Fat= (8.03/10)*100 = 80.3% Titrate Vol.= 2.2ml %Salt= 2.2%

2.

Fat%

3.

Salt %

10

CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

b) CURD: Process Flow Chart:

Receiving milk Packaging &Storage 5C Preheating Cooling Filtration Incubation Standardization Pasteurization( 82C- 83C)

Addition of Starter (1%)

Cooling (5-10C)

Analysis of quality of Curd: Standards for quality of curd: Acidity should range from 0.6 to 0.82
S No. 1.

Property
Acidity

Test
Take 20ml d/w in a beaker and add 9 gm of curd to it. Dissolve the mixture and add drops of phenolphthalein indicator and titrate with N/10 NaOH till light pink color. Note the reading.

Observation
Acidity = 9 MV/W M = 0.1, W= 9g, V= 7.7ml Acidity = 0.77

c) PANEER: Process Flow Chart:

Raw milk chilled Heating upto 85C & Cooling at 75C. Add coagulant (1.5%-1.6% citric acid) Collect solid and transfer to hoop and drain the whey.

Packaging & Storage (5-10C)

Cutting Slabs 100/1000g pieces.

Paneer blocks immersed in pasteurized chilled water for 30 min.

Pressing for 5min under 20 kg weight.

Analysis of quality of Paneer: Standards for Paneer: A) Moisture not less than 50%. B) Fat% = 25, extracted on 50% dry matter. S No.
1.

Content
Moisture%

Test
Frying Method: Heat of ghee in a dish over hotplate. Now add 5g of finely chopped paneer till it becomes brown in color. Note the final and initial weight

Observation
Initial wgt. = 43.77g Final wgt. = 41.24g %Moisture= (2.63/5)*100 = 52.6%

2.

Fat%

11

Reading = 4 Take 10 ml of commercial H2SO4 in a lock stopper % Fat = reading*6.6 = 26.4% close burette. Add 1.69 g of sample and 1ml isoamyl alcohol. Centrifuge at 12000 rpm for 3min. Record reading.

CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

d) PEDA: Process Flow Chart:

Raw milk chilled Heating milk in pan & stirring continuously. Cooling at room temp. and transferring to another pan. Addition of Sugar (7% of milk) in pan.

Packaging &sStorage in dry place

Cooling at room temp.

Scrapping & Molding

Addition of flavors ( Ellaichi)

Cooling at room temp.

Analysis of quality of Peda: Standards for quality of peda: Fat should not be less than 20 %.

S No
1.

Content
Fat %

Test

Observations

Take 10 ml of commercial H2SO4 in a lock Fat% = Reading * 6.6 Reading = 3.17 stopper close burette. Add 1.69g of sample and 1ml isoamyl Fat% = 20.92% alcohol. Centrifuge at 12000 rpm for 3min. Record reading

e) GHEE: Process Flow Chart:

Milk Centrifugal separator Packaging & Storage Cream Cooling and granulation Ripened/ Unripened Churning

Clarification

Creamy butter

Analysis of quality of ghee: S no.

1.

Content
Moisture%

Test
Weigh about 10g of sample in moisture dish . Heat the sample on hot plate. Cool it in a desiccators. Note the final and initial reading. Take 2ml of ghee in a test tube. Add 2ml of chloroform and 2ml of H2SO4.

Observation
Moisture content = loss in wgt./initial wgt.*100 If a transparent sol. occurs, test is +ve.

2.

Cholesterol

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CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

MICROBIOLOGICAL ANALYSIS OF MILK

MICROBIOLOGICAL STANDARDS FOR MILK AND MILK PRODUCTS AS PER BIS S No
1. 2. 3. 4. 5. 6. Particulars Pasteurized Milk Pasteurized Cream Butter Curd Peda Paneer SPC/ ml or g. 30,000 60,000 30,000 5,00,000 Coliform/ ml or g. Absent in 1ml Absent in 1ml 10 10 90 YMC 50 100 10 250

Microbiological test: Pasteurization involves heating milk to a temperature that kills disease-causing microorganisms and substantially reduces the levels of spoilage organisms. Thus pasteurization treatment renders milk safe for human consumption, but to ensure the correctness of pasteurization and decrease of microbial load in the milk it is essential to perform various microbiological tests. S No
1.

Test
MBR

Protocol

Result

Take 10ml milk (Past. & Raw Milk) in Pasteurized milk: Color of the milk does change different test tubes. after 4.45 hrs; hence the Add 1ml of methylene blue solution to it. Now keep the test tube in water bath at microbial load is decreased significantly. 370C. Observe the color change in test tube every Raw milk: Color of milk changed in 1.30 hours, half an hour. Continue test for 5 hours. indicates the high microbial load in milk. Prepare serial dilution of the sample. In a Petri plate add 10-15ml of MacConkeys agar media. Inoculate 1ml from each dilution in the Petri plate. Incubate for 24hrs and observe the coliform colony. Prepare serial dilution of the sample. In a Petri plate add 10-15 ml of LB agar. Inoculate 1ml from each dilution into the petri plate. Incubate for 24 hrs and observe the bacterial colony. No colonies were observed

2.

Coliform

3.

SPC

Colonies Observed: 10 : NIL 10 : 1 10 : 3

Inference: a) Excessive bacterial count is indicative of improper pasteurization treatment. b) Presence of high thermophillic bacteria indicates faultiness in the pasteurizer.
13 CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

WATER TEST
The effluents produced in the water used by the dairy is treated before it is been drained into Gomti river, a local river body of Lucknow city. There are various test performed to check the quality of water and on this basis the polluted water is being treated. S No. 1. Test Oil & Grease Protocol
Take 250ml of sample in separatory funnel. Shake well & add small amounts of ethyl alcohol. Keep for some time to separate two distinct layer a) upper petroleum ether, b) lower sample. Discard the lower layer. Filter the petroleum ether in a beaker over filter paper. Dry and weigh. Pipette 25ml sample and add 2ml buffer solution. Add EB indicator. Titrate with 0.01 M EDTA sol.

Result
Calculations: Oil & Greece (mg/L): A-B*10^6/Vol. = 100 mg/L

2. Total Hardness 3. BOD

Total Hardness as CaCO3: EDTA used*1000/Vol.

= 1.3*1000/25 = 52 mg/L
BOD mg/l= D1-D2/ Vol. Before Incubation (BI) Initial DO = 0.0 Final DO = 7.2 D1-D2= 7.2 Diff (BI-AI) : BOD : After Incubation (AI) Initial DO = 4.4 Final DO = 7.5 D1-D2= 3.1 Diff (BI-AI) : 4.1 BOD : 2050 mg/L

Take 25ml of sample and add 1ml

of phosphate buffer, MgSO4, CaCl2, and FeCl3 solutions/L of water. Before use bring dilution water temperature to 20 3C. Store in cotton-plugged bottles long enough for water to become saturated with DO. Make two sample bottle Determine initial DO by titrimetric iodometric method in one bottle and close second bottle tightly; incubate for 5 days at 20C.

4. COD

After 5 days incubation determine final DO in sample dilutions.

Take 50ml of sample and very slowly add 5ml H2SO4 + Ag2SO4 and shake slowly. Add 25ml of 0.041N K2Cr2O7. Reflux for 2 hr on hot plate. Disconnect reflux condenser and dilute it with twice its vol. distilled water and cool. Titrate with 0.25N ferrous ammonium sulphate. Take reading on sharp color change from blue green to reddish brown.

COD mg/l= A-B*1000*8/Vol. Initial (A) : 0.0 Final (B) : 5.6 COD = 2880 mg/L

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CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.

REFERENCE
1) Marth, E. H., and Steele, J. L., 1998. Applied dairy microbiology: Marcel Dekker, Inc. PROTOCOL REF: 2) Protocol manual: Pradeshik Dairy Cooperative Dairy Federation Ltd. 3) Strategy and Action Plan for Ensuring Safety of Milk and Milk Products, Food Safety and Standards Authority of India. SITE REF: 4) National Dairy Development Board http://www.nddb.org/Compositional-Analysis-of-Milk-Product.aspx 5) Pradeshik Cooperative Dairy Federation Ltd. http://paragmilkup.in/Static.aspx

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CHEMICAL AND MICROBIOLOGICAL ANALYSIS OF MILK & MILK PRODUCTS.