COMPANY PROFILE

Dairy was stared in 1977 with a Handling capacity of 1.0 LLPD

Under BSDC in operation FLOOD-1

Management of the Dairy handed over to NDDB on 01.10.1981

Management handed over to VPDUSS Ltd, Patna by NDDB on

01.07.1988.

Plant capacity expanded from 1.0 LLPD to 1.5 LLPD in operation

Flood-II in 1994.
 BACKGROUND

Patna was one of the milk shed identified under operation Floo1 for
Implementation of programme. A100, 000 It/day capacity Feeder Balancing
Dairy (FBD) and 100MT / day capacity Feed plant (CFP) were set up under
this programme.

In order to implement the programme properly and also organizing the

milk procurement activity and for the management of above two plants. The
Bihar State Dairy Corporation (BSDC) was found in the year 1972.The
Dairy Corporation was to develop the dairy corporation both at the village
level and milk shed level on “Anand Pattern” and it was expected that the
milk shed level co-operation would take over the entire infrastructure
created in due course.

The co- operation after recruitment and training of necessary staff

positioned the procurement and input wing from 1975. A spear head team
(SHT) was deputed from National Dairy Development Board (NDDB) from
the same year for helping the co-operation in organizing and developing the
co-operation. Through the progress could not achieve the goals for which it
was established.

Subsequently, the stable government felt it necessary to request the

NDDB for taking over the infrastructure created on management basis. The
NDDB took over the management of the infrastructure with effect from first
October 1981 under the banner of Patna Dairy Project (PDP).

Progress of Patna Dairy Project

 The National Dairy Development Board (NDDB) immediately
after Taking over the project positioned an integrated Spear Head Team
(SHT) to restructure the milk procurement activities and also for steam
lining the Working of the Feeder Balancing dairy (FBD) and Capacity Feed
plan(CFP) under the management of national Dairy Development Board
(NDDB) the project has not only made excellent progress but had been able
to establish the fact that the co-operative could function equally well in
Bihar to and what is essential in proper atmosphere and guidance.

Along with the organization of milk procurement activities and

Management of both t he plants on commercial lines, the National
Dairy Development Board (NDDB) took special care to develop the
Vaishal Patliputra Dugha Utpadak Sahkari Sangh Limited (VPDUSS), the
milk shedLevel co-operative for taking over the project once the dairy board
withdrawsIt management. National Dairy Develoment Board (NDDB)
handed overThe Patna Dairy project (PDP) to vaishal patliputra
Dugha UtpadakSahkari Sangh Limited (VPDUSS) with effect from first
July 1988.

Feeder Balancing Dairy

The Feeder Balancing Dairy (FBD) with a capacity to

handle 1.5Lakhs litre per day has facilities for manufacture of milk
powder, butter, ghee, ice cream, peda, paneer and plain / misti dahi and
lassi.
 The production and marketing of table butter the brand name
“SUDHA” was introduced from 1st October 1993 and the response has been
encouraging.

The marketing of “SUDHA” brand of ice cream in Patna after test

marketing in August- September 1994 was formally launched from April
1995. The initial response has been more than satisfactory.

The marketing of ‘SUDHA’ brand plain / misti dahi in Patna was

started in October – November 2001 and was formally launched from
2001.The initial response for this product too has been overwhelming.

Cattle Feed Plant

The role of balanced feed is not only increasing milk production

but also sustaining the same by ensuring regular conception need not be
cover Emphasized. Realizing the same union has been making consistent
efforts for popularizing the consumption of balance feed by the milk
producers.

In addition to catering the needs of the dairy co- operation societies

Cattle feed is sold through dealers in rest of the state for beter capacity
Utilization of the plant. Further realizing the importance of introduction of
Latest technologies in the field, the production and sale of by pass protein
Feed was started from the year 1989-90. The response for this feed is
encouraging.

MILK MARKETING
The marketing of liquid milk in sachet was introduced from the year1981
it’s off. However initially the thrust was for organizing the milk

Procurement activities to stabilize the same at reasonable level. Nevertheless

there was some natural growth in the milk marketing over the years.
However for various reasons there was some stagnation for few years in the
quantity of milk marketed with certain modifications in the policy decisions
and because of the concerted efforts, the quantum of milk being marketed
steadily growing.

QUALITY AND PRODUCTIVITY ACTIVITIES

The Dairy plant Management programme (DPMP) was introduced in the

year 1992 followed by Quality Assurance Programme (QAP) in the year
1993 with the help of National Dairy Development Board (NDDB). This
Resulted in bringing about a positive change leading to viability of the
project coupled with lowering of operational costs on one hand and
improved quality of production on the other. Consequent to the liberalization
and globalization of Indian economy in early 90’s it was felt that the
Organization should strive to make its total outlook approach and system of
highest standards. Accordingly it was decided in the year 2001 that the
organization should go in for ISO certification both in quality management
System and food safety. This process was successfully completed leading to
ISO 9001-2000 and HACCP (15-15000_ certification by Bureau of Indian
Standard, in March 2002.

THRUST AREAS

1. To make SUDHA Board, a market leader by making Sudha milk

and Milk product consumer delight and ensuring that the esteemed
customers get value for money.

2. Consolidation of DCSs already organized leading to increase milk

Prodocurement.
3. Further improvement in the involvement / participation of members in
their co- operatitives.

4. Bridging the flush / clean gap further.

5. Popularization all the input programme.

6. Increasing the through puts and sale of milk and milk products as Well as
cattle feed, by pass protein feed and UMB.

7. Reducing further the handling losses and increasing the utilization of

plant capacities.

8. Optimizing the utilization of all consumables.

9. Human resource Development through training, orientation etc to the

Employees at all level for ensuring better motivation for involvement
leading to all round progress of the organization.

OBJECTIVE OF THE PLANT

The VPDUDD Ltd. is committed to continually and proactively

Seeking to live up to the expectations and need of its esteemed milk
produces and customers and delight them with quality products and
services. It shall strive to delight its customers by making available high
quality and safe food with innovative features.

Its endeavor shall be to bring qualitative charge in the socioeconomic

Status for its member producers by ensuring remunerative price for their
Produce and offering quality products at competitive price to consumers
through professional excellence.
 The milk union based on ANAND pattern shall achieve this
novel goal through-

a) Selecting appropriate, cost–effective and eco-friendly process and

technology.

b) Improving the sanitation in the city area by forcing the cattle out
of The city area to rural area.

c) Developing the dairying in Bihar to meet the nutritional requirement of

the people either by supply fluid milk or by the product.

d) Empowering its employees to excel in every sphere through continual

skills, process and system.

e) Providing vibrant work environment that result in excellence.

f) Applying HACCP principles & Q.A activities in quality management

system that results in production of sale food.

Pathogens in Milk & Milk products

1. Introduction
Milk is an excellent food and protective medium for pathogens, whose
Growth depends mainly on temperature and competing microorganism and
their metabolites. Several of them produce toxins, and many are spore
formers. Their disease producing capacity depends upon the initial load of
infection in the milk and on the subsequent dilution, processing, time lapse
before the milk is consumed and other factors.

Pathogenic microorganisms in milk are derived from the dairy animal

itself, the human handler or the environment. One of the most important
extraneous, sources of infection is a contaminated water supply. Insects, dirt,
manure and poor sanitation are other pathogen in milk and its products.

With due care in milk production and handling the modern processing
facilities and good hygienic practices, pathogens can be controlled.

There have, however, been reported cases of outbreak of Disease

attributed to pathogens in milk products. It is therefore necessary to take
maximum care to insure that the product is safe for consumption. This needs
through understanding of products, process, equipment, environment and
pathogens, among others. This issue of the techniques and next one,
catalogue the important pathogens in milk with brief description of their
Characteristics.

THOGENS IN MILK & MILK PRODUCTS

AND DISEASES CAUSED BY THEM.

S.NO NAME OF PATHOGEN DISEASE

1. Bacillus anthrasis Skin infection

2. Bacillus cereus Diarrhoeal illness

3. Brucella abortus Brucellosis

4 Camphylobacter jejuni Bacterial diarrhea

5. Clostridium botulinum Respiratory failure

6. Clostridium Perfringes Gastroenteritis

7. Coxiella burnetti Q. fever

8. Escherichia coli Gastroenteritis, Colitis

9. Listeria monocytogens Meningitis & Abortion

10 Salmonella spp Enteric fever & Typhoid

11 Shigella spp Dysentery

12. Staphylococcus aureus Gastroenteritis

13 Mycobacterium tuberculosis Tuberculosis

.

1.Bacillus anthracis
Characteristies Description
(i)General Useally large rod shaped, non-
motile,spore

Formation capsulated. Gram positive

bacteria
(ii) Source Diseased animal, soil air
(iii) Pathogenicity Anthrax (a fatal disease)

Human
Cutaneous (Skin infection)
Inhalation (affecting lungs) and
gastrointestinal forms of infection
(Animals) Anthrax
(iv) Growth parameter
# temperature 7°c to 49°c
(v) Shedding in milk No any animal with Anthrax. Either
ceases tolactate or gives Milk that is
body.

Growth in Milk
Associated Dairy foods
(vi) Inactivation parameter
Yellowish.visibly abnormal
No
May be raw material
Vigorous boiling for 2 to 3 minutes
Elimination of infected additives from
the food chin. Products from diseased
and dying animals should be rejected
animals should be rejected for human
consumption .through cooking of animal
products offers protection from
vegetative cells of

B. anthracis.
2. Bacillus cereus
Characteristics Description

(i) General Rod shaped aerobic spore froming gram

+ve bacteria
(ii) Source Air, water, fodder. Food
 .soil.udder,milking equipment etc
(iii) Pathogenicity Food born illness (Infection) intoxication
# humans Diarroheal illness (resembles

C. perfringens food poisoning

# Infection Dose Food poisoning due to enterotzin

Production at high population level (more

than 106/g ) particularly in starchy food
#Toxin Type Heat labile enterotoxin (produce in small
intestine) Heat stable (120°c for 90°c
miniutes)

Enterotoxin
(iv) Growth parameter
# Temperate 7°c to 49°c( mesophillic organism capble of
growing at 7°c to 12°c )
#Water Activity 0.93 minimum
#PH 4.3 to 9.3
(v) Shedding in Milk No
#Growth in Milk Yes
# Associated Dairy Foods These organism are common contaminants

Of raw milk and invariably present in pasteurized

milk and product. Rapid growth

Of vegetative cells during periods of temp. abuse is

presumably responsible for high incidence if this
organism in milk during summer months.

Presence of B.cereus in powered milk probably

Possess the greatest puplic health concern because

both pasteurization and spray-drying induce
germination and out growth od spores in the
reconstituted products.

B.Cereus can give rise to “bitty” cream and sweet

curdling in pasteurized milk. Due to the spores
 uriving the pasteurization treatment and the
vegetative cells. Spores of

B. cereus can spoil UHT milk

(vi)Inactivation parameters Heat sterilization done in an autoclave or by ultra
high treatment, is enough to reduce.

B. cereus spore population to a level to ensure no

public health risk and microbiogical stability

Of the product Increase in pasteurization temp.

above 78°c reduces the shelf life of milk due to

Activation of spores of B cereus

(vi) Control measures Widespread occurrence of B.cereus in the nature

environment ensure its continuous recovery from
milk and other dairy products during all stages of
production .However dairy related outbreak of
B.cereus poisoning are readily prevented by
sanitary handling. Prevented by sanitary handling.
Minimizing contamination of raw milk at the

BACTERIOLOGICAL STANDARDS OF MILK & MILK

PRODUCTS. (as per PFA, effective from Dec.2005)

PASTEURIZED MILK
PARTICULARS STANDARDS
SPC Not more than 30000/gm

Coliforms Absent in 0.1 ml.

E.Coil Absent in 1 gm
Salmonella Absent in 25gm

Shigella Absent in 25gm

Staphylococci aureus Absent in 1 gm

Yeast & Molds Absent in 1gm

Anaerobic spore formers Absent in 1gm

Listeria monocytogenes Absent in 1 gm

PASTEURIZED FLAVOURED MILK

PARTICULARS STANDARDS

SPC Not more than 30000/gm

Coliforms Absent in 0.1 ml.

E.Coil Absent in 1 gm

Salmonella Absent in 25gm

Shigella Absent in 25gm

Staphylococci aureus Absent in 1gm

YEAST & Molds Absent in 1 gm

Anaerobic spore formers Absent in 1gm

Listeria monocytogenes Absent in 1gm

Milk power
PARTICULARS STANDARDS
SPC Not more than 50000/gm

Coliforms Absent in 0.1 ml.

E.Coil Absent in 1 gm

Salmonella Absent in 25gm

Shigella Absent in 25gm

Staphylococci aureus Absent in 0.1 gm

Yeast & Molds Absent in 1gm

Anaerobic spore formers Absent in 1gm

Listeria monocytogenes Absent in 1 gm

DAHI
PARTICULARS STANDARDS

SPC Not available

Coliforms Absent in 10 gm

E.Coil Absent in 1 gm

Salmonella Absent in 25gm

Shigella Absent in 25gm

Staphylococci aureus Not more than 100/gm

Yeast & Molds Not more than 100/gm

Anaerobic spore formers Absent in 1gm

Listeria monocytogenes Absent in 1 gm

GHEE

PARTICULARS STANDARDS

SPC Not more than 5000/gm

Coliforms Absent in 0.1 ml.

E.Coil Absent in 1 gm

Salmonella Absent in 25gm

Shigella Absent in 25gm

Staphylococci aureus Absent in 1 gm

Yeast & Molds Absent in 1gm

Anaerobic spore formers Absent in 1gm

Listeria monocytogenes Absent in 1 gm

ICE-CREAM

PARTICULARS STANDARDS

SPC Not more than 250000/gm

Coliforms Not more than10/gm

E.Coil Absent in 1 gm

Salmonella Absent in 25gm

Shigella Absent in 25gm

Staphylococci aureus Absent in 1 gm

Yeast & Molds Absent in 1gm

Anaerobic spore formers Absent in 1gm

Listeria monocytogenes Absent in 1 gm

PANEER

PARTICULARS STANDARDS

SPC Not more than 5000/gm

Coliforms Not more than 90/gm

E.Coil Absent in 1 gm

Salmonella Absent in 25gm

Shigella Absent in 25gm

Staphylococci aureus Not more than 100/gm

Yeast & Molds Not more than 250/gm

Anaerobic spore formers Absent in 1gm

Listeria monocytogenes Absent in 1 gm

 ICE-CREAM MIX

PARTICULARS STANDARDS

SPC Not more than 25000/gm

Coliforms Not more than 10/gm

E.Coil Absent in 1 gm

Salmonella Absent in 25gm

Shigella Absent in 25gm

Staphylococci aureus Absent in 1 gm

Yeast & Molds Absent in 1gm

Anaerobic spore formers Absent in 1gm

Listeria monocytogenes Absent in 1 gm

ICE-CANDY

PARTICULARS STANDARDS

SPC Not more than 25000/gm

Coliforms Not more than 10/gm

E.Coil Absent in 1 gm

Salmonella Absent in 25gm

Shigella Absent in 25gm

Staphylococci aureus Absent in 1 gm

Yeast & Molds Absent in 1gm

Anaerobic spore formers Absent in 1gm

Listeria monocytogenes Absent in 1 gm

DRINKING WATER STANDARD AS PER IS:10500

(Mg/Lit)

PARAMETERS BIS LIMIT (Mg/ lit)

Aluminum 0.03

Arsenic 0.05

Cadmium 1.00

Chloride 75.0

Chlorine 250

Chlorine(total) 0.20

Cobalt 0.05

Coliforms (total/ml) 0.00

Colour 10.0m

Copper 5.00

Cyanide 0.05
Hardness 1.00

Hydrogen peroxide 300

Lead (Pb) 0.30

Magnesium (Mg) 30.0

Manganese(Mn) 0.10

Mercury(Hg) 0.001

pH 6.5- 8.5

phenols 0.001

Selenium 0.05

TDS 500

Sulphate 200

Surfactants 0.2

Turbidity(non-microbial) 5 NTU

LACTOMETER

PRINCIPLE:

A lactometer is used to find out the amount of water in the milk. It works on
the principal of specific gravity of milk. It consists of a test tube and a meter
bulb.

Toned milk = 8.5%

Double toned milk = 9.0%

Standardized milk = 8.5%

Sudha Gold = 9.0%

At 27c lactometer reading = 30

29c lactometer reading = 28

CLR = Lactometer reading

= CLR / 4 x.2 x fat +.66

 = 28/4x.2x54.6x0.66

SNF = 7x1.78 = 8.78%

Total Solid = Fat +SNF = 5.6 + 8.78 = 14.38

Moisture content = 100-14.38 = 85.62%

Gram Staining of Plain Dahi &Misti

Dahi
Procedure:-

Prepare Smear on a glass slide

↓ Heat fix using a spirit lamp

Bacterial cell fixed on slide

↓ put a drop of crystal violet and wait

For 1 miniute

Remove excess stain with tap water

↓

Bacterial cell are stained

↓ Stain with gram’s iodine and wait

For 1 minute
 Remove excess stain with tap water
↓

Decolourize the stain with 95% ethanol

↓ put 1 drop of saffraning and wait

For 1miniute

Put 1 drop of saffranine and wait for 1 mimiute

Wash with tap water.

Result:- Plain Dahi- Gram +ve coccous

Misti Dahi- Gram + ve coccous and some are diplococcus.

Aim: Methylene blue reduction test for microbial contamination of milk.

Requirement: test tubes, Test tube stand, pipette, Milk sample (Raw milk,
pasteurized sudha milk), Methylene blue solution.

Principal: Methylene blue reduction test is done to judge the shelf life of
milk, to now the probable quality of milk, to know the sanitary condition of
lant.herincipal behind is that a milk sample that contains a large population of
actively metabolizing microorganisms will contain a markedly decreased
concentration of dissolved oxygen because of growth of organisms. In other
words, the oxidation-reduction potential of the sample is greatly lowered. The
dye MB a redox indictor, loses its color in an anaerobic environment & is
said to be reduced. The Methylene blue reductase said to be reduced. The
ethylene blue test is designed to screen the quality of milk, which may
contain large population of enteric organisms & streptococcus lactis, which
are potent reducer of dye. The speed at which Reduction occurs following
addition of ethylene blue to a sample of milk indicates quality of milk. This
determination id made as follows;

1. Reduction within 30 minutes-very poor quality.

2. Reduction occurring within 30 mins to 2 hrs-poor quality of milk
3. Reduction occurring between 2& 6hrs- fair quality of milk
4. Reduction occurring between 6to 8 hrs- good quality of milk.

Procedure:

Methylene blue solution was prepared.

The temperature of water bath was fixed at 37°c
10ml of each 2 sample of milk (Raw milk, pasteurized milk) were poured in
test tube.
Then 1 ml of Methylene blue solution was added into the test tubes.
The test tubes were incubated at 37°c in water bath & observation was taken

Observation:

Raw milk: decolorized (blue to white) within 30 mins.

Pasteurized milk: decolorized after 6 hrs.

Result

Methylene Milk sample Classification of Approximate

blue Milk numberof
bacterial/ml
Reduction
time
0 to 30 mins Raw milk Verypoor quality >20,000

361 to 480 Pasteurized Good quality <500,000

milk
mins

Conclusion: from above observation it can be conclude raw milk was poor
quality of milk because microbial load in that milk was more,
as it was not boiled as it decolorized after 6hrs Microbial load
was least in this milk.

Precaution:

• Methylene blue was prepared carefully.

• Proper aseptic condition were made (sterilized tubes, sterilized rubber,
pipette should be used.)
 • Temperature of water bath should be maintained properly as required
& test tube should be appropriately labeled.

Aim: To detect the antibiotic residue in milk.

Requirement: Bromocresol purple agar embedded with Bacillus

Stereothermophilic var. calid ollactis, raw milk, 1 nutrient table, and pipette.

Pricipal: The Delvo test is one of the tests used to detect the presence
drugresidue in milk. The principle of the method involves germination
growth of spores of specific bacteria (Bacillus Stereothermophilic var.
calidollactis,) embedded in agar upon the addition of nutrients & following
incubation at a specified temperature.

If milk is free of inhibitory substances the growth of these

spores produces acid. Which changes the colour of the pH indicator
(bromocresol purple) in agar from purple to yellow? However, if milk
contains certain inhibitory subsequently the acid production by the bacteria
the medium remains purple in colour.

Procedure:

• Ml of milk is added to bromocresol purple agar embedded with B

Stereothermophilic.
• 1 nutrient tablet is added with forceps.
• Leaved foe 4hrs at 64.5°c in water bath as it is a growth temperature
essential for these bacteria.
• Then after seeing the colours of tubs further observation is made.
 Observation: As raw milk samples from 5 different places Islampur,
Bidouli, Sukhmal, Sarojnagar, Bhadura were used. It was observed that
bromocresol purple agar turned to yellow colour.

Result : All samples from 5 different places give antibiotic –ve test.
Conclusion: Thus, by seeing above observation we can say that, all samples
are antibiotic free & it shows that as antibiotic free & it shows that as
antibiotic is absent in milk samples it mean bacteria is surviving in it & able
to convert lactose of media to lactic acid & colour turn ti yellow.

Precaution:
• Proper aseptic conditions were made
• Water bath temperature is set accurately so that proper growth
environment is given to bacteria.

Aim : To check the cleaning efficiency of the tank / tanker / pipelines.

Principal: proper sanitary condition of the plant is the most important step in
a food processing industry. There must be proper cleaning of the tank,
tankers & pipelines. Micro- orgaisms are very important in proper
fermentation processes. Apart from a few microorganisms which are
hazardous for human health. Therefore proper conditions should be
maintained to inhibit or check the growth of microorganism.

Raw material from different sources is collected in large tanks

& tankers. Pasteurized are passed from one to another through various
pipelines. SO, proper care should be taken to prevent post-pasteurization
contamination of milk.

Requirement: Cotton swab, dilution, nutrient agar, violet red agar media,
sterilized pipettes, sterilized petriplates, colony counter etc.

PROEEDURE:-

Preparation of diluents

Stock solution- 34 gm potassium dehydrogen phosphate is dissolved in 1000

ml distilled water. 1.25 ml of the above solution was then diluted to make
1000 ml. The solution was sterilized at 121°c & 151b / inch2 pressure for
15-20 minutes.

Preparation of decimal dilution

5 ml of dilution was poured on swab.

It was sterilized at 121°c 151b/inch² pressure for 15 to 20 minutes.
With the help of swab, microbial sample were obtained from different tanks,
tankers pipelines.

Inoculation & incubation

1ml of diluents was transferred to sterilized petriplates.

10ml of the melted cooled nutrient agar & violet red media was added to each
petriplates.

Each pert plate was rotated gently, immediately after addition of the media,
for inform distribution of the organisms & allowed to solidify the agar.

Second layer of violet red agar medium was poured after solidification for
anaerobic conditions.

All the plants were incubated in inverted position for 24-48 hrs at 37°c.

Silo Uncountable No growth

Cream tank No growth No growth

Vat Uncountable Uncountable

Tank Uncountable 2× 45=90

36× 45=1620 No growth

Silo pipe
Blank No growth No growth

Calculation & result:

It has concluded that containers containing raw milk have uncountable

bacterial colonies whereas containers in which pasteurized milk is kept free
from any type of contamination. Therefore, we can say that
sanitaryCondition of the plant is good.

Precaution:
 • Aseptic condition should be maintained.
• Media should be properly prepared.
• All plates should be sterilized.

EXPERIMENT

Aim: To count the coliform of pasteurized milk

Principle: The coliform group of bacteria comprises all aerobic, facultative

anaerobic gram negative non sore forming rods able to ferment lactose with
the production of acid & gas at 30°c, 35°c or 37°c within 48hrs. Common
source of these organism‘s intestinal tract of warm blooded animals. Certain
bacteria of non-feacal origin are also members of this group.

Typically, three bacteria are classified in the genera Escherichia,

Enterobacteria & Klebsiella. The presence of these coliforms in dairy
products is suggestive of unsanitary conditions or practices during
production, processing or storage of milk & milk products.

The coliform estimates are performed on raw milk to determine the degree
of contamination during milk production. While the tests on pasteurized milk
are referred to detect post pasteurization contamination.

Requirement: milk sample, sterile pipettes, sterile test tubes, sterile petri
plates, autoclave, colony counter.

Procedure:

Preparation of diluents

Stock solution -34 gm potassium dihyrogen phosphate is dissolved in 1000ml

distilled water. 1.25ml of the above solution was then diluted to make 1000
ml .the solution was sterilized at 121°c & 151b/ inch² pressure for 15-20
minutes.
Preparation of decimal dilution

1ml of the test sample was pipette & added to 9ml diluents.

This primary dilution was shaked using mechanical shaker for 5 to 10

seconds.

1ml of this primary dilution was transferred into another tube containing 9
ml sterile diluents, contact between pipette & the diluents & mixed to obtain
10-1 diluents.

Inoculation & incubation

1ml of diluents was transferred to sterilized petriplates.

10ml of the melted cooled nutrient agar & violet red agar media was added to
each petriplates

.Each petriplates were rotated gently, immediately after addition of the

media, for inform distribution of the organism & allowed to solidify the agar.

Second layer of violet red medium was poured after solidification for
anaerobic conditions.

All the plates were incubated in inverted position for 24-48 hrs at 37°c

Observation:

Bacterial colonies were observed on the plates, which were counted using
colony counter of both the milk samples.

67,66 & 63 colonies on respective three plates, which give an average of 65.

Therefore, CFU colony forming unit = no of colonies × 1 0 =65

×10=650/ml

Result:

As the number of bacterial cells per ml is less than 30,000 therefore, milk is
property pasteurized.

Precaution:

• Septic condition should be maintained.

• Media should be properly prepared.
• All plates should be sterilized.
 EXPERIMENT

Aim: Alkaline phosphates test to check the efficiency of passsteurized milk.

Requirement: Constant temperature, water bath maintained at 37.5 ± 0.5oc

Pipettes, volumetric flask, measuring cylinder, test tubes with s toppers, milk
samples (raw milk, pasteurized milk)

Chemical reagents

Buffer solution: 1.5 gm of sodium bicarbonate & 3.5 gm of sodium carbonate

in distilled water & volume is made up to 1000ml.
Buffer substrate solution: 0.15 gm of substrate (disodium p nitro phenyl
phosphate) into a 100ml volumetric flask & volume is made up with buffer
solution.

Principal: Phosphates enzyme, at pH 9.5 & temperature 37°c spits the

substrate, p nitro phenyl phosphate to give p nitro phenol, which is yellow
colored in alkaline solution.
Phosphatase present in milk is destroyed during pasteurization, Therefore,
Phosphatase test is performed to determine the efficiency of milk in dairy
plant.

Procedure:

• 1ml of each 2 sample of milk (raw milk & pasteurized milk poured in
test tubes.
• Then 5 ml of buffer substrate solution is added into the test tubes.
 • The test tube was incubated at 37°c in water bath & observation was
made.

Observation:

Seeing the colour of milk made observation. If, colour changes to yellow then
considered as positive test. If milk is pasteurized properly then colour of milk
not change it remains white. First reading is made at 10mins & next at 30
mins.

Raw milk: Raw milk became in a faction of second.

Pasteurized milk: Remains white.

Result: As raw milk’s colour changes to yellow colour it means phosphatase

+ve test. Pasteurized milk gives –ve test.

Conclusion: Thus, by observing the colour of milk sample, we can say that
(pasteurized milk sample provided to us is properly processed, as Alkaline
phosphates enzyme present in milk denatured at pasteurization temperature.

Precaution:
Aseptic condition solution should be maintained.
Buffer substrate should not be exposed to direct sunlight.
Standard plate count (SPC) of pasteurized milk (Smart,
Shakti)

Principal:

The standard plant count (SPC0 is one of the oldest and most widely
Used method for enumerating bacteria. Classically SPC procedures are used
to determine the total plate count (TPC) or Aerobic plate count (APC) or
total viable count (TVC).SPC is the standard, method to which other
screening tests are compared.

The methods involves preparation of the decimal dilution of the

milk sample, transferring a know volume (generally 1ml) of the appropriate
dilution of the sample into petri dishes adding a prescribed nutrient agar
into each dish and incubating the dishes at a specified incubation period.
After the incubation period, the colonies developed in each plate are counted
and the total viable count is calculated using the dilution factor employed in
the tests.

Requirements:
Milk sample (Smart, Shakti), sterile pipettes, sterile test tubes, sterile
petridishes, autoclaves, colony counter.

Procedure:

Preparation of diluents

Stock solution:

1. 1000 ml distill water in flask

2. Add 34gm potassium dihydrogen phosphate
3. 1.25 ml of the above solution was then diluted to make 1000ml
4. The solution was sterilized at 121°c and 151b/ inch2 pressure for 15-20
minutes.

Preparation of decimal dilution

1ml of the test sample was pipette and added to 9ml diluents.
This primary dilution was shaked using mechanical shaker for 5-10 seconds.

1ml of this primary dilution was transferred into another tube containing
diluents, avoiding contact between pipette and the diluents, and mixed to
obtain 10-2 dilution.

Similarly 10-³ dilution was made adding 1ml of 10-² dilution to 9ml sterile
diluents.

Inoculation and incubation

1ml of 10-3 dilution was transferred to sterile labeled petriplates.

10ml of the melted, cooled agar medium was added to each pertriplates.
Each petriplates was rotated gently, immediately after addition of the
medium, for uniform distribution of the organism and allowed to solidify the
agar.
All the plants were incubated in inverted position for 48hours at 37°c

Observation:
Bacterial colonies were observed on the plates which were counted colony
counter of both milk samples.
10, 15, and 18 colonies on plantes incubated with shakti
12, 16, and 20 colonies on plantes incubated with smart

Calculation and result

The number of bacteria per ml of the milk sample was calculated.

Viable cell per ml=mean plant count x10³
Direct microscopie somatic cell count (DMSCC)
Principle:
The direct microscopic estimation of somatic cell count in milk is
referred to as the DMSCC. The number of somatic cells in raw milk
provides a measure of the presence and the extent of mastitis or certain other
abnormal milk secretion. Direct microscope somatic cell count is the
officially recognized procedure for confirming the somatic cell count in milk
previously estimated by one of several screening tests. Test results are
reported in actual counts of somatic cells per milliliter of milk.

Requirement: milk sample, slides, Newman’s strain, distill water

Procedure:
1. A square of 1cm2 areas was drawn on the slide
↓
2. 0.1ml of milk sample was spread on that area, and was heat fixed
↓
3. The slide was dipped in Newman’s stain for 20 minutes.
↓
4. The slide was rinsed in water until all the surplus dye was washed off.

5. The slide was air-dried before examine the film under microscope.

Calculation and expression of result:

Compute somatic cell count as follows and express the result as direct
microscopic somatic cell count (DMSCC) per ml of milk.

DMSCC per ml= no. of somatic cells in a single strip x5333

Observation:

S.N Sample collected No of cells observed DMSCC per ml

1 Bhagwanpur Nil 0

2 Bairichak 5 27775

3 Gangachak Nil 0

4 Daulatpur 6 33330

5 Bhadaura 3 16665

Fluorescence image of somatic cells before image processing

Result:

Milk collected from sudha dairy does not come from diseased animals.

Precaution:

1. Milk sample should be fresh

2. Raw milk should be used
3. Excess of stain should be properly washed off.
 Air exposure of media on different section of the plant

Principal:

The atmosphere contains all the major group of microbes ranging from
the algae to the viruses. The microbial flora of air is transient and variable.
Air is not a medium in which microorganisms can grow but is a carries
particulate matter, dust and droplets, which may be laden with microbes. The
no, and kinds of microorganisms containing the air are determined by the
source of contamination in the environment.

Medium used for air exposure was made acidic for the proper growth of yeast
and moulds and also to restrict the growth of bacteria. Each plate was
exposed for a given period of time in different section of the plants such as
paneer section, dahi section, ice –cream section, peda section.

Requirements: PDA media, sterile Petri plates, tartaric acid etc.

Procedure:

1. 10ml of the melted cooked PDA media was added to each Petri plate.
 ↓

2. Plates were allowed to solidify

↓
3. Plates were exposed to air for 5 minutes
↓
4. All the plates were incubated in inverted position for 72 hours at room
temp.

Observation:

Fungal colony observed on the plates. Numbers of yeast and molds colonies
were counted.

S.N Sites No of colonies Time of No of mold/

yeasts / molds exposure min m3
1 Ice cream section 16 5mins 16x5=80
2 Paneer packing 13 5mins 13x5=65
room
3 Peda packing room 13 5mins 13x5=65
4 Dahi packing room 8 5mins 8x5=40

Result:

A large number of different molds and fungi were obtained at different

section of plant.

Precaution:

Aseptic condition should be maintained.

Media should be properly prepared.

 DETERMINATION OF PROTEN IN MILK BY
FORMAL TITRATION (PYNE’S METHOD)

PRINCIPLE-

When formaldehyde is added in milk which was previously titrated against

Standard alkali to the end point of an indicator like phenolphthalein, it bind
with the amino group of the milk protein and releases equivalent amount of
protein and releases equivalent amount of protein which could be titrated
against the alkali to the same end point.

The amount of alkali used in second titration is the measure of the amino
group originally present in the protein.

APPARATUS-

Pipette-10ml, 2ml,1ml

Granduated burette.

Erlenmeyer flask-100ml
REAGENT-

Neutral formalin, saturated potassium oxalate solution,

N/10 sodium hydrocide, phenolphthalein indicator

PROCEDURE-

1. Take 10gm of milk into 100ml flask.

2. Add 5 drops of phenolphthalein indicator.
3. Add 0.4ml of saturated potassium oxalate and keep this solution
undisturbed for 4-5 minutes.
4. Titrate the milk against the standard alkali solution to its end point.
5. Add 2 ml of neutral formalin and mix well.
6. Titrate against the standard alkali to the same end point as before.
7. Record the volume of alkali in second titration.

Result-
Percentage of protein=R*1.7

COMPOSITION OF MARKET MILK PER 100 gm

S.N PARTICULERS UNITS TONED STD DOUVLE SUDHA

GOLD
MILK MILK TONED MILK
MILK
1 FAT Grams 3.10 4.60 1.60 6.10

2 SNF Grams 8.60 8.6 9.10 9.10

3 PROTEIN Grams 3.20 3.20 3.55 3.55

4 LACTOSE Grams 4.70 4.70 4.80 4.80

5 MINERAL Grams 0.70 0.70 0.75 0.75

6 CALCIUM Mg 118 118 128 128

7 PHOSPHORUS Mg 90.0 90.0 96.0 96.0

8 WATER Grams 88.3 86.8 89.3 84.8

9 VITAMIN A I.U 150 200 75.0 200

10 CALORIES K cal 59.0 73.0 48.0 90.0

POISONOUS METAL LIMITATION

METALS LIMITS (Maxm) REMARKS

Lead 0.5ppm All milk /milk pdts.
Maxm 1.0
ppm.lollies)

Arsenic 0.1ppm milk.

Maxm 0.5ppm

Icecream / lollies)

Copper 30ppm All milk / milk pdts.

Zinc 50ppm All milk / milk pdts.

Cadmium 1.5 ppm All milk/milk pdts.

Mercury 1ppm Milk.

Methyl mercury 0.25ppm All milk / milk pdts.

MILK ENZYMES

Nomenclature, classification general properties and salient characteristics of

some important milk enzymes.

An enzyme is a biological catalyst elaborated by the living cells of

mammary tissue gain entrance into milk accidentally or unavoidably during
the secretory process. Enzymes are proteins they are denatured or
inactivated by high temperature process, a pH of optimum activity and
exhibit specificity for certain substance.

Milk contains a number of enzyme about 21 enzymes have been purified,

isolated or definitely identified in bovine milk, milk enzymes play a vital
role in assessing milk quality and shelf-life. Enzymes in milk have
extensively utilized as yard-stick. For evaluation a variety of biological
events operative in milk, these specific enzyme catalyzed a variety of
biological events operative in milk, these specific enzyme catalyzed
operations in milk have been summarized below.

1. Correlation with existence of pathogens.

The enzymes in milk are heated sensitive. The presence of certain

enzymes in milk is of great importance in relation to heat treatment e.g.
Alkaline phosphates activity in milk has time honoured cheek for the
degree of pasteurization.
The thermal death point of Mycobacterium tuberculosis and the enzyme
alkaline phosphates coincide with each other hence the activity of latter
serves as an index for the presence of former in milk.

2. Correlation with spoilage.

The enzyme lipase acts on milk fat, produces hydrolytic rancidity and
thereby causes the spoilage of butterfat and products containing fat.

3. Correlation with under diseases.

Increase in level of A. esterase, aldolase, and catalase are the established
consequences of Udder disease and some other physiological disorder.
Leucocytes count of milk has a correlation with the level of catalase in
milk.

4. Correlation with flavor defects.

Xanthine oxidase has been implicated in the oxidative degradation of
dairy products resulting in flavor defects.

5. Correlation with bacterial action

Lysozyme has antibacterial or immunological significance and is
associated with keeping quality of milk.
6. Correlation with stability of emulsion.

Ribonuclease may influence the stability of fat emulsion since it is

associated with the microtonal component of the fat globule membrane.

The principal enzymes present in milk are-

1. Aldolase EC 4.1.2.13
2. α Amylase EC 3.2.1.1
3. β Amylase EC 3.2.1.2
4. Carbonic anhydrase EC 4.2.1.1
5. Catalase EC 1.11.1.6
6. Cytochrome –c-reductase EC 1.6.99.3
7. Diapheraes EC 1.6.4.3
8. A-esterase EC 1.6.4.3
9. B-estrase EC 3.1.1.8
10. C-estrase EC 3.1.1.8
11. Lactose synthetase EC 2.4.1.22
12. Lipase EC 3.1.1.3

13. Ly sozyme E C 3.2.1.17

14. Phosphoprotein phosphalase E C 3.1.3.16
15. Phosphtase alkaline EC 3.1.23.1
16. Peroxidase EC 3.4.4
17. Protease EC 3.4.4
18. Rhodonase EC 2.8.1.1
19. Ribonucease EC 2.7.7.16
20. Solase
21. Xanthine oxidize EC 1.2.3.2

Other enzymes present in milk include.

Sulphydril restuetase, sulphydryl oxidase, Oleinase, Transaminase, Sorbitol
anhydrase, phosphohexose isomerase, β Glueuronidase, Flavo kinase etc.

These enzymes depending upon their mode of action on milk

Constituent’s physiological components can be categorized as follows.

1. Hydrolytic group of enzymes e.g. lipase, esterase and protease

2. Enzymes having physiological signification e.g. eatalase, lysozyme
etc.
3. Enzymes associated with the microsomal particles of milk e.g.
alkaline phosphates, xanthine oxidease, cytochrome –c- reductase etc.
4. Enzyme whose roles are yet to be defined confirmed e.g. ribonuelease,
rhodonase, carbonic.

Milk enzyme can be classified broadly according to the substrate

they act upon carbohydrates, proteases act upon protein, lipase act upon
lipids and so forth. Their classification proposed by International union of
biochemistry (IUB) However appears to be the one most widely used
according to which enzyme are divided among six major classes.

1. Oxidoreduclases which catakyze oxidation or reduction reaction

2. Transterases which catalyze the transfer of specific chemical moiety.
3. Hydrolase Which hydrolyse substrate with concomitant uptake of water
4. Lyases which catalyze the addition of a group to double bonds or
conversely removes groups from substrate leaving double
bond.
5. Isomerases which cause isomerization.

1. Ligases [synthetase] Which catalyses the condensation [bonding

together] of two Molecules coupled with the cleavage of pyrophosphate
bond of ATP or similar triphosphate

Each class is then divided into sub-class, each sub-class sub-subclass and
finally each sub-subclass contain several enzymes. Or example the serial
classification number the first digit 3 from the left represent the class of
hydrolases. The second digit 1, represent sub-class of carboxylic ester
hydrolyses and the final digit 3, represent the enzymes glycerol ester
hydrolyses.

It is believed that these enzymes are normal constituent of cell or tissue

and during milking process concomitant with the cell rupture these
enzymes are spilled into milk. It has been suggested that these enzymes
are secreted in milk for the benefit of the young having rather
underdeveloped or in compete digestive system.
 Milk is not a homogenous solution of enzymes. Enzymes occur in milk
in four distinct phases.

1. Water soluble
2. Associated with cream or lipid
3. Bound to easein
4. Enzymes present in microsomal particles.

A brief account of milk enzymes present in SUDHA DAIRY:

1. ALDOLASE- A glycolytic enzyme associated in the sequence of

reaction involved in the metabolism of carbohydrates i.e. hydrolyses
fruclose-1, 6-diphosphate into dihydroxy- acetone and
phodphoglyceric aldehyde milk exhibit significant aldolase activity
serum. The enzymes are associated with the fat globules, on separation
isconeentrated in cream in cream layer. Aldolase is rather unstable in
milk, the activity decrease rapidly at 37° and 45°c.
.

2. AMYLASE- This enzymes catalyses the hydrolysis of α-1,4-D

glycosidic linkages in starch and glycogens α-amylase content of milk
is very low but high in mastitis milk.

Α-amylase is inactivated by the pasteurization temperature but β-amylase

is fairly heat resistant, α-amylase is considered normal constituents of
cows milk white β-amylase is only in some milk.

3. CARBONIC ANHYDRASE- This enzyme reversibly catalyzes the

hydration of CO2 & dehydration of carbonic acid.

Carbonic anhydrase

CO2+H2O → Η 2CO2

This enzyme is a Zine containing protein

4 CATALASE- In catalyzes decomposition of H2O2 into water and

oxygen.

catalase
2H2O2 → 2H2O+O2
 Its concentration in milk is directly proportional to lencoyle eoant catalases
content of colostrums and mastitis milk is significantly higher then that in
normal milk and its appearance in milk is dependent upon the physiological
condition of the animal. For this reason the Catalase content of milk has been
proposed as a means of detecting mastitis, catalases activity and particularly
with feed. Catalase activity is present in both cream & shine and its co-
precipitation with cascin have also been reported like from other sources.
Catalase of milk also appears to be a haemato protein .Heating at 60-75 for
30 minuts destroys the catalase and its activity is greatly impaired if it is
heated for the same length of time at some what lower temperature.

1. CYTOCHROME-C-REDUCTASE- The enzyme appears to be

concentrated in milk microsomes its activity is measured
spectrophotometically. The rate of reduction of Cytochrome C

2. DIAPHORASE- This enzyme catalyses the hydrogenation of

lipoamide in the presence of a hydrogen acceptation. Its activity is
measured by following the reduction of 2,6- dichlorophenol
indophenols. In the enzyme appears to be associated with
microsomes.

3. ESTERASES- the term esterase embraces a variety of enzymes

which catslyse the hydrolysis of esters. Although lipase is also an
esterase its activity.

Generally is regarded to be confined to glycerol esters. Three esterase

namely A, B and C have reported.

A esterase (aryl esterase) is a typical esterase which hydrolyzes

phenyl acetate at a higher rate than phenyl butyrate. Aliphatic esters
normally are attacked.

B esterase (carboxy or glycerol ester hydrolyser) hydrolyzes aliphatic

and aromatic esters but not the choline esters. It is sensitive to
organophosphates but not to serine.

C esterase (Choline) splits esters more rapidly than aromatic and

aliphatic esters. It is sensitive to organospates and serine.

A & C esterases activity is high in colestrum and she formers. A

esterase activity of milk appears to be related to mastitis. B esterase
contains glycerol hydrolase also lactase synthetase.It eatalyzes the
synthesis of lactose from UDP galaelose and α-D glucose. This
enzyme originates from the microbes of the mammary gland cell.
Brodbeek and Ebner (1966) while purifying this enzyme from bovine
milk demonstrated it resolve into two fraction designated as A and B
neither fraction was active as enzyme, but gained the activity when
 the two fraction combined together. Fraction A and B were identied
as galactosy transferase and α facalbumin.

The galtosy transferase is found in much body tissue but α -

laealbumin only in lactating mammary gland. This protein modifies
he galactosy transferees in such a manner that it transfers galactose to
glucose only and hence the synthesis of lactose in mammary gland.

4. LIPASEE- It catalyzes hydrolysis of glycerol esters (fat & oils) in

emulsion several workers have isolated many lipases each differing
from each other in many respects, in milk there are two forms of
lipases Viz.

a) Plasma lipase: associated with casein and requires activating

treatment like homogenization, agitation or foaming before it
produces raneidity.
b) Membrane lipase: adsorbed on the fat globule membrane which is
catalyzed by cooling of milk.
Purified lipase possesses a single pH optimum and a temperature
optimum of 37°c.
It is unstable and highly sensitive towards light, heat and serval
reagents.
Various milk constituents have a profound effect upon its activity.
Saltsand casein being inhibitory, the observed lipase activity in a
complete milk system may be the net result of the inhibitory
action of various milk constituents.

Milk lipase has drawn particularly and preferential attention over other
enzymes in milk because of its established role in the development of
flavor in milk. Milk processing and storange invole lipase action with
a resultant production of undesirable rancid, flavor. This enzyme is
responsible for inducing altreration; however it is a notorious enzyme
in regard to its hydrolytic rancidity production in milk and milk
products.

1. LYSOZYME – Milk of large number of species contain lysozyme

and human milk is the richest source. Bowing milk contain 13µg of
lysozyme per 100 ml while human milk contain 36 µg/ ml. the
enzyme from bowing and human milk differs considerably in their
physiochemical properties. The isoeletric point of bovine and human
milk lysozymes are pH 9.5 and 11.0 with optimum ph for activity at
7.9 and 6.85 respectively.

This enzyme has specific behavior or lying certain bacteria such

function is achieved by the hydrolysis of the β-(1-4) linkagae
 between acetyl glucosamine and n-acetyl muramic presenting
polymer from in its probable role described as associated with

a) Natural antibacterial factor in fresh milk.

b) In relation to infant nutrition.
c) The mechanism of natural immunity
d) Keeping quality of milk

It is remarkably stable to heat (human milk lysozyme 1 more heat sensitive

particularly at acid pH . It is relatively a small protein with a milexular
weight approximately 18,000 which is slightly higher than that of egg white
lysozyme.

1. PEROXIDASE- It catalyzes decomposition of H.O in the presence of

an oxidizable compound or hydrogen donor milk peroxidase often
referred to as lactoperoxidase perhaps the first enzyme reported in
milk.
Milk is one of the best sources of peroxidase. It represents as much as
1% of the total serum protein of the milk It appears to be associated
with albumin or when protein compound of milk. This enzyme is
more heat stable when compared with other enzyme in milk. It
exhibit regeneration properly when inactivated by metod. This
enzyme can be used to detect 11.0 when added as a preservative in
milk. It is a home protein with a content of about 0.07%.

11 PHOSPHATASE- This class of enzyme hydrolyzes phosphoric acid esters

A large number of phosphor of phosphates exists in nature such as
phosphorous esterase phosphoric – esterase’s, phosphorylases,
pyrophosphatases, phosphoprotein phosphatases etc. Milk contains several
phosphatases but most of the work done has been on two phosphatases.

Alkaline Phosphatase: It is a phosphomonoesterase with a pH optimum of

9.6. Its activity in normal cow’s milk vaties considerably.The relative
distribution of the enzyme in milk fraction is established. About 30-40
present of the enzyme is convent rated in cream where it is adsorbed on the
fat globule as microtonal particles, the balance is (60-70) present) is
distributed throughout. The skim milk probably in the liproteins particles.

It is a native enzyme of milk and plays an important role in energy transfer

mechanism of living brings. The complete loss of activity of this enzyme
synchronizes with pasteurization temperature of milk. Therefore its activity is
eidely used as yard- stick of pasteurization efficiency of a milk processing
plant. The well known phosphatase test is based on this characteristic. The
main substrates used for assessing its activity include disodium phenyl
phosphate,p-nitrophenyl phosphate phenolphthalein phosphate etc. The
enzyme exhibits the characteristic of reappearing or becoming activated in
heated milk especially in HTST processes.

Acid phosphates: Its concentration is low in milk its optimum pH is 4.0. It is

unstable when exposed to sunlight or UV radiation but it is very heat
resistance requiring 96°cFor 5 minute for its complete destruction.

PROTEASE- It catalyzes hydrolysis of the peptide linkages of protein to

small fragments. Milk protease acts in slightly alkaline medium; its action is
retarded in acidic- medium. Its optimal activity is at pH 8.5. It is inactivated
by heat at 75-80°C and in acid medium it is destroyed at 72° For 10 minutes.
Its activity is not likely to have any serious effect in milk processing however
while preparing evaporated milk by HTST method reactivation of protease is
possible it may however not be a milk protease.

RIBONUCLEASE- Bovine milk contain 25mg / 1 of this enzyme, which

catalyzes the hydrolysis of ribonucleic acid like ribonuclese A from bovine
pancrease, milk ribonuclease is also a basic low molecular weight protein. It
exists primarily in whey. It is believed to be related with the immunological
characteristic.

SALOLASE- It catalyzes the hydrolysis of phenyl salicylate. It has also been

detected in bovine mammary gland..

XANTHINE OXIDASE- It is rather non- specific, enzyme since it

Catalyzes the oxidation of purines, phyrimidines aldehyde. Xanthine
oxidase is a prominent enzyme in milk .It is present in the milk of mare and
human. It is part of microsomal particles located on fat globule
Membrane.It is a metallo –flavor protein containing molybdenurm and
iron as the metals in the ratio of
1:2: (1.3+.5):8 as Protein: FAD: MO”Fe.

The enzyme has been implicated in the oxidative deterioration of milk fat

is another characteristic feature of this enzyme. It appears to be activated

by variation in temperature treatment such as cooling, heating, agiation
and various chemical agents such as detergents. Heat sensitivity of this
enzyme increases on storage homogenization and treatment with
proteolytic and lipolytic enzyme.

CONTROLPOSTPROCESS
CONTAMINATION OF MILK

INTRODUCTION

Raw milk received in dairy plant contain bacteria mostly from the
contamination during its handling and bacterial growth.
Milk is therefore pasteurized as soon as possible after milking in which
processes not of other bacteria are killed .This makes milk safe for
human consumption and
Increases its shelf life.
However proper processing of milk alone is not adequate to ensure the
safety and quality of final products, all the bulk in the subsequent “milk
chain” need to be property manage. In the absence of this, a dairy plant
runs the risk of unsafe and a poor quality product with attendant
disastrous results. Consider for example the following incidences:-

• There was an outbreak of staphylococcal entertain poisoning due

to
Contaminated processed milk of one dairy company in Japan in
2000
 Over 11000 people were affected with 165 requiring
hospitalization. Infection was traced to one plant where a single
value in the production line had been inadequately cleared. The
company suffered a loss of over 125500 crores.

• In the U.S.A, 29 outbreak were recorded between 1995 and 1997

Related to dairy product of which 34% were caused by
contaminated milk. A Salmonella spp. was isolated in outbreak
and one each due to
Campylobacter and Listeria.

• A market survey of processed milk of 10 brands in India in 1999

Reported high level of E. coli in milk of several brand.

• In an incident in U.S.A in 1985, 16000 persons suffered

salmonella due to drinking of pasteurized low fat milk of 2
brands. The milk was found to be contaminated with Salmonella
typhinurium. The contamination of milk occurred due to cross
contamination with raw milk in the pasteurizer.

There are only a few example many more have been reported and many
have remained unreported.
Inadequately processed milk or recontamination caused poor quality and
unsafe milk, which cost the dairies heavy economic losses and dairies of
reputation.
Reason od low quality and unsafe processed milk include inadequate
pasteurization process or post pasteurizations process contamination .In
most of cases of unsafe pasteurized milk the reason found has been post
Process contamination (PPC). Therefore while it is important to ensure
proper pasteurization of milk, it is equally important to take all
precaution to avoid PPC. Its major reason of PPC and their control
measure.

COMMON RECONTAMINATING MICRO-ORGANISM

Pasteurized milk has been responded to be contaminated with several

types of spoilage causing and / or pathogenic micro-organism.

Some microbial hazards identified in pasteurized milk

Campylobacter jejuni, Listeria monocytogenes, Salmonella spp
Staphylococcal entertoxin,yresinia entercolitica, shiga toxin producting
E. coli (STEC).
Gram negative (psychrotroph)
 Pseudomonades
Bacillus cereus

Recontamination of pasteurized milk gram- negative psychotropic

bacteria (GNP), responsible for spoilage of milk has been reported to
occur in filluing step. Pseudomonades have been found to be the most
frequently occurring bacteria in refrigerated pasteurized milk.
Gram- positive spore (GPS), such as Bacillus cereus spores, also
recontamination processed milk and are sometimes alone responsible for
its spoilage. The contamination site could be dead ends, pockets and
traps where bacteria in case the CTP system are ineffective.
Enter pathogenic E.coli and salmonella have been reported to be present
in pasteurized milk due to recontamination. Likewise campylobacter spp,
staphylococcus aureus and Listeria monocytogenes have been found in
recontaminated milk.

POST PROCESS CONTAMINATION ROUTES

Milk may be contaminated via a myriad of contact surface of processing and

packaging equipment and plant environments. Milk residues on inadequately
cleaned surface; tanks, pipes and valves can support the

Survival and growth of microbial contaminants. Spores of Bacillus cereus

are very hydrophobic and will attach to the equipment surfaces where they
may germination and form biofilm at sites that are difficult to clean.

Contamination of milk by Bacillus cereus has been demonstrated in silos,

Pasteurizer, milk pipelines with bad welding and packaging machines. Very
often recontamination has been found to occur in process by the rinsing water
inside and allows milk to come to contact with the surrounding air and with
its aerosois. Condensed water on the equipment may also find its way into
the milk and packaging material might be contaminated.

Recontamination of milk could take place at various sites in equipment and

pipe work such as dead ends, pockets and traps and mixing processed milk
receiving back from stores.

Outer Environment
↓
Plant Environment
 Pasteurizer Packaging machine

Pasteurised

Personnel Packaging film

Raw Milk

Modern pasteurizers are often complex and although efficient these do

present possibilities of cross contamination of milk, plant treated cooling
milk is passed through regeneration and cooling water section where it is
separated by relatively thin plates from raw milk and chilled water,
Respectively. Should any of these is a signification risk of recontamination of
milk with pathogen or spoilage micro- organism.

Further, the cleaning in place (CIP) lines and other associated lines in the
pasteurizer might, if incorrectly designed or installed, allow raw milk to by
pass the pasteurizer completely.

PRECAUTIONS AGAINT RECONTAMINATION

Prevention of recontamination of pasteurized milk is of major importance in

Production of pasteurized milk that is both safe and of satisfactory shelf- life.
Some control measures against contamination of milk with specific
pathogens are listed below. However pasteurizers and filling machines are
usually designed and constructed to minimize the possibility of the
pasteurized product being contaminated, however adequate precaution are
required to ensure that post process contamination does not take place. Some
important precautions are elaborated below:-

1)SOURCE: plant environment

PATHWAY:- Usually indirect via contamination of equipment. Also

possible via personnel and packaging.
 PRECAUTION:- Eliminate contamination of pasteurized milk side
of regeneration by leakage etc. Correct environmental sanitation.

2)SOURCE:- Equipment

ROUTE: - Direct as following:-

i) Contamination of equipment by raw milk, coolant leaked in
refrigeration or cooling section, respectively, due to gasket
failure, split or pin hones in pasteurizer plates improper
pipelines / valves arrangement.
ii) Contamination by stagnant milk deposits at dead ends, values
gaskets etc.
iii) Inadequate CIP and manual cleaning where necessary.

PRECAUTIONS:-

Vent interspaces between seals to atmosphere to provide an ingredient visual

in designs of gasket failure.

• Maintain a positive pressure balance at least 0.5 bar (0.5kg /sq.cm),

between pasteurized milk and raw milk in the regeneration section.
• Ensure correct positioning of flow diverter and associated pipe work
to avoid contamination of flow resumes after diversion.
• Restrict operating period to 8 hrs.
• Testing for corrosion cracks and pinholes by a lithium injection test
twice a year.
• Visual inspection every day for backpressure control.
• Milk contract surface of pasteurizing plant should be fabricated from
high grade stainless steel and polished by electro-polishing to avoid
crevices and
Consequent entrapment of soil, hands etc. should be insured to the
highest possible standard.
• Follow suitable cleaning and sanitizing programmers for pasteurizer,
storage tanks, silos etc.
• Extra cleaning in case of more than 3days between processing runs.
• Dismantling and inspection every 5 years.
SOURCR:- Raw milk

PATHWAY: Direct or indirect via contamination of plant environment,

passive transfer or hands of personnel etc.

PRECAUTIONS

I) Correct design of equipment and related pipe work

II) Correct operation and maintenance of pasteurizer
III) Correct plant layout
IV) Control of personnel movement and avoidance of ‘hand on’ during
operation involving milk or milk contact surfaces.

4) Source:- personnel

ROUTE:

i) Direct due to personnel suffering clinical illness or being chronic

carrier of pathogens.
ii)Indirect due to introduction to plant of contamination from outside
environment.

PRECAUTION:-
i) Follow appropriate medical and exclusion policies.
ii) Ensure good personnel hygiene and correct use of protective clothing
and footwear.
iii) Prohibit raw products such as eggs being brought into the plant by
farmer or worker for sale to fellow farmers.

5) SOURCE :-Packing

ROUTE

I. Failure to adequately sterilize the packaging film in the packing

machine.
II. Contamination of packaging from plant environment etc.

PRECAUTIONS:-

I. Ensure correct functioning of required ultra-violet light system so that

the packaging film is properly sterilized
II. Protect packaging from contamination.
III. Cleaning and disinfections of packaging machine and buffer storage
tank according to conformed procedures.
IV.Disinfection of specific machine part by alcohol spraying.
 V. Adherence to keep hygienic condition during filling.
VI.Follow conformed procedure regarding start, stop and interruptions.
VII.Remove first 20 packs.

It is of particular importance to ensure that there are no cross contamination.

It should be ensured that pipelines and valves cannot be arranged and / or
fail in such a way that Pasteurized product or pasteurized products line could
be contaminated. It is critical to ensure that the flow diversion value is
correctly installed and operated so that under- processed

Milk is not carried forward. The operation of the flow diversion should be
checked each time plant is started up and the correct operation of recorded
per assured.

Recording thermometer should check daily against calibrated mercury in –

glass thermometer and holding time validated annually.

Unnecessary product handling steps between processing and filling should be

avoided. In addition, the lines leading to the filler should be designed for
efficient cleaning in place.

Packaging machines the selves should be designed with emphasis on clean

ability and potential for contaminations during filling.

The location of the filler is also important .It is undesirable to place milk
filler in the same room as an open cheese vat when the environment could
contain significant bacterial load. It is a good practice if the filler is placed in
a designated filler room that operates under specific white room guidelines
i.e. filtered air –employed dress codes etc. The packaging materials should be
stored in a clean and sanitary manner to avoid contamination.

CONCLUSION

It was really fun as well as a new experience for me to make this project.

I felt much more informed and learned after completing this project. With my
raid of variety of dairy products, Patna Dairy project (PDP) has Emerged as a
unique name in the field of dairy industry. The quality of products, the plants
are easily determined from the satisfaction of its loyal customers.

For Patna Dairy plant, there is still a lot to be done and achieved. The way
PDP has penetrated the lanes and by lanes of Patna and its suburbs reflects
nothing but its success and commitment.

My informants were very useful and it was really fun gathering information
and doing experiments and various tests in bacteriolocal department.