INTRODUCTION

India’s sea food industry has become one of the leading suppliers of quality seafood to all the
major markets of the world. India has world class seafood processing plants that follow quality
control regimes compliant to stringent international regulatory requirements.

India contributes 6.3% of the total global aquaculture production and ranks 2nd after China.

“Increased production of L. vannamei, diversification of aquaculture species, sustained measures

to ensure quality, and increase in infrastructure facilities for production of value added products
were largely responsible for India’s positive growth in exports of seafood,” said Mrs. Nirmala
Sitharaman, Minister for Commerce and Industry.

ABOUT THE COMPANY

SRIKANTHINTERNATIONAL PVT LTD (SKIPL) is the private partnership company located

at 98A Someswaram village, alapadu PO, Kaikaluru Mandal, Krishna Dt., A.P.,India. Its
administration is being carried by the Board of Directors. It is aspiring to be the leading Indian
Shrimp Processor and Exporter. It has started by the technocrats educated from the first Fisheries
University in India, Hatchery owners and Farm owners. Its ultra modern processing facility with
state of art machineries were established in the year 1996 with the vertically integrated facilities
from Shrimp Hatchery, Shrimp Farming to Shrimp processing and Exports. Its vertically integrated
facility tuned to harness the requirements of food safety and quality standards.

Srikanthinternational pvt ltd is committed to produce and deliver safe, legal and quality marine
products to its ultimate customers according to their specifications and comply with regulatory
requirements ensuring utmost customer satisfaction through continual improvement. The company
is committed to adopt all trade practices, national and international standards such as ISO 22000,
IFS and BRC, which will ensure that the customer requirements are met at all times. Its primary
objective is “Food Safety” right from farm to plate. The company’s shrimp farms stocked with PL
from the hatchery are raised with eco-friendly inputs under technical supervision taking immense
care and protection, thus ensuring traceability and food safety.

1
Srikanthinternational Pvt ltd’s success lies in its committed, well-qualified, highly experienced
and motivated team.

VISION: To provide international customers with the highest quality, safest, and sustainably
produced marine-based products.

MISSION: To be a customer driven, quality conscious business enterprise providing value added
Seafood and marine-based products in socially and environmentally responsible manner.

QUALITY POLICY: To deliver to its customers, seafood as close to the condition as it is found
in, in its natural habitat as a Fresh, Safe and Legal product by the implementation of food safety
and quality standards.

COMPANY’S LOGO:

2
ORGANIZATIONAL CHART

3
PRODUCTS:

A) Head-On, Shell-On (HOSO) (Fig.I) F) Butter Fly cutting(Fig.V)

B) Head-Less, Shell-On (HLSO) (Fig.II) G) Peeled & Deveined (PD) (Fig.VI)

C) Easy Peel (EZPL) (Fig.III)

D) Peeled & Deveined, Tail-On (PDTO) (Fig.IV)

E) Pull Vein Peeled & Deveined (PVPD)

IMPORTING COUNTRIES:

4
 DESCRIPTION OF THE RAW MATERIAL:

• The raw material that is used in the production line of the plant is Litopenaeus vannamei.
• Its common names are “Pacific white shrimp, Whiteleg shrimp and King prawn”.
• Scientific name of the raw material: Litopenaeus vannamei, Penaeus vannamei
(formerly).
• It is a variety of prawn of Eastern Pacific Ocean commonly caught or farmed for food.
• The feed is purchased from the SHENGLONG bio-tech India pvt.ltd.
• The shrimps are being cultured in the GAYATRI hatchery and are also purchased from
other local aquaculture farms having the direct surveillance against environmental and
chemical contaminants and antibiotics or from the same farms through Coastal Aquaculture
Authority (CAA)/MPEDA registered suppliers.
• Shrimps are the swimming crustaceans with a cylindrical carapace, long narrow muscular
abdomens and long antennae.
• The structure of a shrimp consists of the following parts:
➢ Cephalo-thorax region with antennae,rostrum,compound eyes and pereiopods
➢ Abdomen (6 segments) with attached pleopods
➢ Tail portion (telson & uropod)

Fig. 02 A typical structure of the shrimp with labeled parts

5
PRODUCTION LINE:

In this company, the shrimps that are purchased are being processed, frozen, packed & labeled and
exported to the buyers by complying all the requirements that are given by the customers.

Shrimps (Litopenaeus vannamei) being perishable foods, should be processed by maintaining its
temperature below 3.8oC so as to preserve its quality.

The process done in the production line can be described in the following steps:

• Raw material receiving area

• Chill room
• Be-heading area & waste disposal area
• Filth washing area
• Grading
• Value addition area
• Chemical treatment/Soaking area
• Freezing area(Block/IQF)
• Primary packing and labeling
• Metal detection
• Secondary packing (Master cartons)
• Cold storage
• Shipment

RAW MATERIAL RECEIVING AREA

When the purchased raw material enters the plant, the cleanliness of the vehicle and the seal are
monitored. The raw material (HON Variety) is being unloaded from the vehicle, the samples are
taken as per the requirements for the Quality Assurance Lab (250g sample for each batch) and for
the raw material monitoring report (one sample for every 500kg of each batch). The temperature
of the raw material is checked and recorded for each batch. The presence of the sulphite is
examined by using the strip test method.

6
RAW MATERIAL SPECIFICATIONS (Table No. 01)

Appearance Satisfactory
Odor Satisfactory
Texture Soft & firm
Temperature <3.8oC
Dehydration Nil
Discoloration Nil
Deterioration Nil
Black spot/tail <3%
Broken/damaged <5%
Soft shell <10%
Foreign matter 2-3 vegetative matter
Antibiotics, pesticides Not detected
GMO Nil
Other allergens SMBS(<10ppm)
Ice condition Good

CHILL ROOM

The chill room is loaded with the raw material received in crates until the raw material is sent for
the processing unit. The temperature of the chill room is maintained at <4oC. Its capacity is around
4-5 tons. Ice should be on the raw material so as to maintain the temperature of the raw material
below 3.8oC.

BE-HEADING AREA & WASTE DISPOSAL AREA

It is the place where the heads of the shrimps are removed. The head waste is being sent to the
waste disposal area. After beheading, the shrimps are now termed as “headless variety (HLSO)”.

The yield% is being calculated by using the following formula:

Yield% = HLSO Wt. / HON Wt. X 100

7
The yield% should not be <70%. Otherwise, it would be a loss for the company. The loss of the
material is around 30% due to the removal of the head part.

FILTH WASHING AREA

After the completion of beheading process, the HLSO are dumped into the filth washing machine
containing chilled chlorinated water (2ppm). It has a washing capacity of 2000kg/h. The material
is being thoroughly washed and is sent to the grading section.

GRADING

Now the material is passed through the grading machine via belt conveyor and the grading is done
according to the size set in the machine.

There are different grades classified as per the no. of pieces present in one lb (454g). They are:

U5 U10
8/12 13/15
16/20 21/25

26/30 31/40

41/50 51/60

61/70 71/90

91/100 100/200
In order to check whether the material is properly graded or not, we calculate the uniformity by
using the formula i.e., uniformity = wt.of big pieces/wt. of small pieces. The no. of pieces taken
as per count is as follows (Table No. 02):

Count No. of pieces taken(big or small)

>50 10
40-50 7
30-40 6
20-30 5
<20 4

8
The difference in wt. between big and small pieces taken should be <20g.

To check whether the HLSO count for the specific HON count is attained properly& to get the
count per 1lb, a conversion factor is used. (1Kg/454g = 2.202 X 0.70 = 1.54)

HLSO Count = HON Count/1.54

VALUE ADDITION

After the grading is done, the HLSO is then processed into different varieties as per the buyers’
requirements. The following are the varieties processed in the plant:

A. EASY PEEL (EZPL)

It is the variety which is processed by cutting the HLSO shrimp variety with the shell
attached from 1st to 5th segment with the help of the scissors and the vein is pulled out.

Fig.03 Easy Peel variety (EZPL)

B. PEELED & DEVEINED, TAIL-ON (PDTO)

It is the variety which is processed by peeling off the shell attached to the abdomen and
leaving that of the tail portion, cutting the meat from 2nd to 5th abdominal segment with the
help of the pencil knife and then the vein is pulled out.

Fig.04 Peeled & Deveined, Tail-On variety (PDTO)

C. PULL VEIN PEELED & DEVEINED (PVPD)

9
 It is the variety which is processed by peeling off the shell completely and pulling out the
vein with the help of the needle.

D. PEELED & DEVEINED (PD)

It is the variety which is being processed by peeling off the shell completely and cutting
the meat portion from 2nd to 5th abdominal segment with the help of the pencil knife and
the vein is then pulled out.

Fig. 05 Peeled & Deveined variety (PD)

The waste (shell, vein, etc.,) is sent to the waste disposal area with the help of the crates.
After the value addition process is done, the varieties are being monitored so as to find the
defects if any are present or not. The following are the some of the defects found:
1. Semi-soft shell
2. Soft shell
3. Black head
4. Red hanging meat
5. Black spots
6. Discoloration (DC)
7. Brushed piece
8. Loose shell
9. Broken piece
10. Attached legs
11. Attached vein
12. Tail cut (broken tail)

10
 Fig. 06 some of the defects found in the processing area

The yield of the value added varieties from the HLSO variety is as follows as (Table No. 03):

YIELD% (FROM HLSO

VARIETY VARIETY) LOSS%
EZPL 99% 1%
PDTO 88% 12%
PVPD 86% 14%
PD 84% 16%
In order to check whether the meat count is attained or not, the following formula is used:

Meat count = HLSO Count / Yield% (taken in chemical point)

CHEMICAL TREATMENT (SOAKING AREA)

After the value addition of the meat, the chemical treatment is either done or not as per the
requirements of the buyer. It is done generally in order to gain the yield lost due to the removal of

11
the shell and other attached parts. It is the process that takes around 90 minutes in which the
chemical composition is added as follows (for 100kg of the meat):

a) Carfosel(Sodium tri polyphosphate) 2%

b) Salt 2%
c) Ice 36%
d) Water 60%

The soaking process is carried out in a soaking machine and in order to get uniformly treated, the
stirring is done accordingly either manually or with a mechanical stirrer. The product now is
termed as “treated” if it undergoes soaking process. Otherwise, it is called as “non-treated”.

The yield% of the following after the completion of the soaking process is as follows (Table
No. 04):

VARIETY(FROM
HON) TREATED NON-TREATED
HLSO 71% 69% OR 70%
EZPL 73% 69%
PDTO 67% 60%
PD 65% 57%

FREEZING AREA

It is the area of the processing line where the meat is either block frozen or IQF frozen. The freezing
of the product is done so as to preserve the quality of the product as it is a perishable food product.

12
In the block freezing process, the meat is sorted, weighed, pan set, filled with glaze water, froze at
-40oC for about 1hr in the contact freezers and depanned and the blocks are sent for the primary
packing.

In the IQF (Individual Quick Freezing) process, the meat is sorted, weighed, passed on the belt
conveyor where froze at -40oC (IQF unit), glazed, glaze hardened at -40oC (Hardener unit) and
weighed and sent for the primary packing. The belt speed is set accord to the variety passed through
the IQF equipment.

PACKING AND METAL DETECTION AREA

After the product is either block frozen or IQF frozen, it is weighed, graded (in case of IQF frozen
product) and then packed in the master cartons as per the packing style required by the buyer. The
primary packing material used is polythene bag covers and the secondary packing material is the
corrugated fiberboard box (master carton). Before packing in the master cartons, the product is
being passed through the metal detector so as to ensure the absence of any metal in the product. If
any product is detected, the product is removed from the packing it in the master carton.

COLD STORAGE

After the product is packed in the master cartons, it is stored in the cold store having capacity 3241
metric tons until it is being loaded in the container for the shipment purpose. The temperature of
the cold store should be below -18oC so as to maintain the temperature of the product at -18oC.

SHIPMENT

After the confirmation of the test reports and other norms required by the buyers, the reefer
container is checked for proper hygienic conditions. The reefer container to be loaded for the
shipment is cleaned thoroughly to be a foreign material free, insect/pest free one. It is then loaded
with specific no. of master cartons of the specified product as per the “break-up” (i.e., the no. of
the master cartons to be loaded as per the buyer’s order). Data loggers are attached at the center of
the reefer container so as to monitor the temperature of the container at -18oC. The company’s &
customs seals are attached to the reefer container before it is sent out.

ANALYSIS OF COST OF PRODUCTION

13
The following analysis is done using estimated values of the particulars

I. Fixed costs:

Land 19.5 crores

Building 10.0 crores
Machinery 19.2 crores
Furniture 1 crore
Vehicles 7 crores
Prepaid expenses 1 crore
Total 57.8 crores

II. Variable costs (per annum):

Raw materials 328 crores

Cash for paying wages 5.5 crores
Salaries 1.2 crores
Repairs 1 crore
Interest 4 crores
Taxes 38.4 crores
Transportation 6 crores
Total 384.1 crores

TOTAL COSTS = fixed costs + variable costs

= 57.8 crores + 384.1 crores

= 441.9 crores

BREAKEVEN POINT (BEP)

14
 BEP in terms of unit sales = Total fixed cost/(unit sale price – unit variable cost)

= 57.8 crores/(700 – 461.5)

= 15,38,784.1 units

REFRIGERATION SYSTEM

Refrigeration system in a food industry especially, in case of perishable food products plays a vital
role in the maintenance of quality of the food products. It plays an important role in the processing
of frozen foods.

The refrigerant used in the company is “anhydrous ammonia” (R-717). It is a clear, colorless
liquid or gas, free from visible impurities but has a sharp pungent odor. It is commercially available
at low price and commonly used as refrigerant in various refrigeration units. However, exposure
to it at small concentrations may cause irritation to eyes, throat and burning sensation but high
exposure to this refrigerant may lead to death.

Refrigeration process is a cyclic process in which the refrigerant is stored temporarily in the vessels
before the cycle starts. From the vessels, it is sent to the evaporator (i.e., plate freezer or IQF belt)
through pipes where the cold refrigerant liquid turns to hot vapor due to the heat transfer between
the product and refrigerant. Now the hot vapor is passed through insulated pipes to reach the
compressors and then to the condensers to cool the vapor and when the refrigerant is in liquid state
(i.e., cooled down to normal state), it is then sent to the receiver and then to the vessels. And again
the process is started when required.

VESSELS EVAPORATORS COMPRESSORS

(Plates/belt)

RECEIVER CONDENSERS

Fig.07 Schematic representation of refrigeration process in the plant

15
POWER SUPPLY

The power supply for the running of the company is provided around 33kVA power line distributed
by Southern Power Distribution Company of Andhra Pradesh Limited and the company is
facilitated with a power back up of 1500 kVA capacity generators supplied by Cummins Powerica.

Fig.08 POWER LINE Fig.09 GENERATOR

WATER TREATMENT PLANT

In the water treatment plant, bore well (raw water) is collected in the overhead tank subjected to
filtration through slow sand filter, carbon filter and again passed through resin softener. Water is
chlorinated to the level of 2ppm by automatic chlorine dozer. The filtered water is pumped to the
overhead tanks and then passing through the UV filter before it is being distributed to the plant. It
has a reverse osmosis (RO) plant with capacity 10,000litres/h within. Its storage capacity is around
8, 00,000litres.

Fig.10 Water Treatment Plant & Reverse Osmosis plant within it

PROCESSING EQUIPMENT

The plant is well-equipped with different equipments at different processing sections.

16
 I. Weighing Balances
These are used to weigh the specific quantity of the material in the plant. They are
usually present at the sections like raw material receiving area, grading section,
packing area, etc.,

Fig.11 Weighing Balance

II. Work Tables

Work tables are provided at the areas like beheading, value addition, arranging the
meat in the slabs (Block freezing), IQF, packing & labeling etc., in the plant.
These tables are equipped with an outlet so as to drain the water if present on them

Fig.12 Work Tables in the plant

III. Filth Washing Machine

It is the equipment used to cleanse or wash the shrimps with chilled chlorinated
water (2ppm). It has a capacity of 2000kg/h.

Fig.13 Filth washing machine

17
IV. Grading Machine
Grading machine is used to grade the shrimps according to the size. It is supplied
by Yantai Jiuhua, China. It has a belt speed setting and the wheel adjusters to set
the size for the shrimp to pass through and around six belt conveyors to collect the
shrimps after grading.

Fig.14 Grading Machine

V. Soaking machine
It is the equipment used in the soaking area. It consists of a mechanical stirrer and
square box at the base.

Fig.15 soaking machine used in the plant

VI. Plate Freezers

Plate freezers are the contact type freezers used to freeze the value added shrimp
product into frozen blocks. There are 3 plate freezers in this plant, in which two
plate freezers have a capacity of 324 slabs each while the other one has a capacity
of 440 slabs. They are supplied by Arico, Vietnam.

18
 Fig.16 Plate freezer

VII. IQF line

IQF line is the equipment line consisting of the belt conveyors, IQF unit and
hardener. It is operated at the temperatures below or at -40oC. There are 2 IQF lines
in the plant, each having a capacity of 925kg/h per each IQF unit. The equipments
are supplied by JBT (Advantech), Sweden.

Fig.17 IQF LINES

VIII. Metal Detector
Metal detectors are the equipment used for the detection of any metal in the food
product. There are 2 metal detectors in the packing sections of block & IQF area.
Before the food product is passed, the test kits are passed so as to ensure the proper

19
 working of the metal detector. Later, the food product is passed through and packed
in the plant.

Fig.18 Metal detector

IX. Sealing machine
Sealing machines are used to seal the packing material so that there is no wastage
of food product or for ensuring the safety of the food material in the packing
material. There are around 3-4 sealing machines in the plant.

Fig.19 sealing machines used in the plant

X. Pallet truck
Pallet trucks are the equipment used to convey the load from one place to another.
There are two kinds of pallet trucks present in this plant. They are hand pallet trucks
and mobile pallet trucks.

Fig.20 Hand pallet truck

XI. Refrigeration system
The refrigeration system in this plant is PLC & SCADA application controlled
automatic type. It consists of a receiver, 2 vessels, 3 high-stage compressors, 4
boosters, condensers and pipeline supply i.e., either insulated pipes or not. The

20
 equipments in the refrigeration system are supplied by FRICK and JOHNSON
CONTROL, USA.

Fig.21 Equipment present in the refrigeration system

XII. Flake Ice machine
It is the equipment used to manufacture the flake ice so as to spread on the material
in the processing section in order to maintain the temperature of the product in low
ranges. It has a production capacity of 60 tons/day. It is supplied by GEA Ecoflex,
Germany.

Fig.22 Flake Ice collected from the flake ice machine

XIII. Reefer & Insulated containers
These are used to load the tons of food product and transport from farms to plant,
from plant to shipment boarding port for the proper maintenance of cold chain
throughout the transport line. Their capacities vary accord to the range in between
10 to 40 tons of load.

Table No. 05 Estimated prices of the following equipment

21
S.No. Name of the Specification Supplier Cost/price Purpose of the
equipment or per unit equipment/machinery
machinery
1. Filth/washing 2 Tonnes per JBT Food 1 crore Washing headless
machine hour Tech shrimp
2. Grading 2 Tonnes per JBT Food 2 crores Grading of shrimp
machine hour Tech
3. Soaking 250 Kg per JBT Food 1.2 crores Soaking of shrimp
machine hour Tech with stirring
4. Plate freezers 1.6 Tonnes JBT Food 3 crores Block freezing of
per hour Tech shrimp
5. Individual 2 Tonnes per JBT Food 3.5 crores Individual Quick
quick freezing hour Tech Freezing (IQF) of
machine shrimp
6. Sealing 2 Tonnes per JBT Food 0.5 crores Sealing of packages
machine hour Tech
7. Metal detector 2 Tonnes per JBT Food 1 crore Detection of ferrous,
hour Tech non ferrous, stainless
steel metals in packed
shrimps
8. Refrigeration 250 Tonnes Frick & 5 crores To maintain
system Jhonson refrigeration cycle
control
9. Flake ice 3 Tonnes per GEA 2 crores Production of flake ice
machine hour Ecoflux

STORAGE FACILITIES

22
There should be an adequate storage facility as the company is processing and exporting the
shrimps, which are perishable food products as its temperature and quality should always be
monitored. In this plant, there are around 3 chill rooms having a capacity of 10 tons each, two cold
store having capacity of 3241 metric tons together and packing store so as to store the cartons,
polythene pouches & duplex etc.,

Fig.24 Racks in the cold store

Fig.25 Packing store

PLANT CAPACITY

The following is the plant capacity per day:

BLOCK FREEZING – 26 Tons/day (8+8+10)

IQF FREEZING – 33 Tons/day

TOT. PLANT CAPACITY PER DAY = 59 Tons

SHELF-LIFE OF THE FROZEN PRODUCT: 2 YEARS (24 Months)

23
NUTRITIONAL LABELLING ON THE PACKING MATERIAL

The following is the nutritional information that is being labeled on the packing material
of the frozen food product (Table No. 06):

 Servings size 4 oz (112g)

 Servings per container: 8

Calories per serving: 60 calories

%Daily Value
Total Fat 0g 0%
Saturated Fat 0g 0%
Trans Fat 0g
Cholesterol 125mg 42%
Sodium 350mg 15%
Total Carbohydrates 0g 0%
Dietary fiber 0g 0%
Sugars 0g
Proteins 15g
Vitamin-A 0% Vitamin-C 0%
Calcium 8% Iron 0%
SHRIMP WASTE UTILIZATION

The waste from the shrimp processing section includes the head waste, shell waste and other wastes
of the food product. The waste is collected from various areas of the processing line is being sent
to the waste disposal area. It should be sent out from the plant before 24 hours. Otherwise, the
microbial load present in the waste starts to grow rapidly and this may lead to spoilage of the waste
and it may contaminate the fresh food material. The waste sent out is being dried, crushed into
powder and is commonly used in the animal feed.

These are the some of the applications of the shrimp waste:

➢ As fish feed, animal feed supplement ,bait or fertilizer

➢ Nutrient media for certain microbes like Bacillus sps. (protease enzyme)
24
 ➢ Chitin extraction – stabilizer, natural biopolymer
➢ Chitosan extraction – bioactive compound used in different fields like agriculture, food,
biotechnology, cosmetics, medicine, etc.,
➢ Glutamic acid extraction – essential amino acid for brain development

TRACEABILITY

When a particular lot of raw material is received in the raw material receiving section, it
will be subjected to raw material evaluation. On approval of the material, a lot number is
allotted to it. This number will contain all required information such as name & address of the
farmer/ supplier, location of the pond, variety & type of the product and the time of arrival.

During subsequent processing of the material, the same lot number will be retained in all the
relevant records. The finished product will be given a code number denoting the date of
production. This code number is affixed on the primary containers as well as on the master
cartons. Apart from that, Lot numbers will also be marked on the packed cases. All the
packaging material such as Master cartons, duplex, trays, pouches & ingredients are procured
from approved resources & traceability is maintained.

CODING SYSTEM

The coding system followed by the company for each batch of production done is as
follows:

For example, let us take a code number i.e., 8M25

Here, 8 indicates the year of the production i.e., 2018

M indicates the code letter given for a month i.e., December

25 indicates the day of the production i.e., 25th day

The code letters for the months in a year are as follows:

NAME OF THE MONTH

25
 CODE LETTER
A JANUARY
B FEBRUARY
C MARCH
D APRIL
E MAY
F JUNE
G JULY
H AUGUST
J SEPTEMBER
K OCTOBER
L NOVEMBER
M DECEMBER
Whereas for the traceability code is different, as it is the registration number given to a farm by
CAA/MPEDA. Eg. AP006854-004 indicates the CAA/MPEDA Farm Registration number.

MANPOWER

The following table indicates the estimated no. of manpower present in the plant:

Table No.07 Estimated no. of manpower present in the company

DEPARTMENT MANPOWER
Pre-processing section 200
Block section 35
IQF section 35
Cold store section 20
Quality assurance 25
Administration office dept. 20
Maintenance& hygiene 20

PLANT LAYOUT

26
MARKETING SECTION

Marketing is defined as “the activity, set of institutions, and processes for creating,
communicating, delivering, and exchanging offerings that have value for customers, clients,
partners, and society at large” by the American Marketing Association.

In the marketing section, the purchasing of the raw material, packing material & other required
materials in the plant, selling the products to the buyers, money transactions, etc., are to be carried
out effectively.

The process of export marketing that takes place from the enquiry of the products to dispatch can
be described as follows:

1. Enquiry- done by the buyer to know the details of the food products in the company

27
 2. Quotation – a standard business process to supply the buyers’ required products at
specified price in a specified time period, sent by the company
3. Negotiation – a bargaining process between seller and buyer seeking to discover a
common ground & reaching to an agreement to settle to a matter of mutual concern
4. Confirmation of order
5. Pro forma invoice – an estimated invoice sent by the seller to the buyer, which contains
all the particulars of the products to be sold
6. Code list – prepared by the company in which the buyer’s requirements like kind of
variety, specific packing style, no. of the master cartons needed, etc.,. it is sent to all the
departments concerning in the processing of the product
7. Processing & checking – the area where the products are processed and inspected to
ensure the quality of the product. In order to ensure the safety of the food product, the
samples are sent to the external labs for the testing procedures
8. Packing & labeling –the area where the products are packed and labeled according to the
specifications set by the buyer
9. Storing – the area where the products are being stored temporarily until the dispatch
process is started
10. Pre-shipment inspection – the inspection that is being carried out to ensure the quality of
the product and to maintain the safety of the food products.
11. Shipment - the process of loading specified quantity of the products into the container,
sending the shipment documents (like antibiotics report, PHT report, test reports, pro
forma invoice, etc.,) as per the buyer’s requirements is done

In a food company, mainly in case of perishable ones, there is a higher level of risk taking in the
selling of the food products to customers whose satisfaction is to be met. Otherwise, it leads to a
loss to the company as it involves many factors that may cause the damage to the food products.

28
 QUALITY CONTROL & ASSURANCE IN THE PLANT

QUALITY
It can be defined as “the degree of excellence” of some material or thing.

Quality of sea food products is attached to physical, biochemical and Microbial

characteristics.

QUALITY CONTROL

It is described as “the system of maintaining of the standards of the processed

products by testing or checking a sample of the output against the specification required”.

QUALITY ASSURANCE
It is “the maintenance of the desired level of the quality in a product, by means of
attention to every stage of the process of the production”
It is the way of preventing the mistakes or defects in the processed products and
avoiding the risks when delivering them to the customers.
QUALITY POLICY OF THE COMPANY

To deliver to its consumers, seafood as close to the condition as it is found in, in its
natural habitat a Fresh, Safe and Legal product by implementation of food safety and
quality standards.

MAINTENANCE OF QUALITY IN THE PLANT

Royale marine has in-house laboratory to conduct special tests to detect the presence of
micro organisms and anti-biotic to ensure the end product is absolutely contamination free.

Quality is monitored at every level and quality is maintained throughout the process right
from shrimp’s farms till export of the finished product. Stringent quality measures are
implemented at every stage

The system of traceability adopted by Royale Marine ensures tracking its products from
hatchery to the consumer.

29
Quality department in the company is divided into two teams:

1. Quality Assurance

2. Online Quality Control

In order to maintain the quality standards and requirements, the company follows certain
manuals. They are:

Table No.08 The manuals followed by the company

S.NO. ABBREVIATED MANUAL NAME

FORM
1. HACCP Hazard Analysis Critical Control Point
2. SOP Standard Operating Procedure
3. SSOP Sanitary Standard Operating Procedure
4. CGMP Current Good Manufacturing Practices
5. FS&QMS Food Safety and Quality Management System

Royale Marine is in compliance with BRC - Grade A and FSSC 22000, CGMP 21 CFR
Part110 of the USFDA, HACCP system based on Codex Alimentarius Commission’s
guidelines. It has the following certifications:

➢ HACCP
➢ FSSC 22,000 (Food Safety Standard Certificate)
➢ BRC – British Retail Consortium

30
Fig.26 FSSC 22000 & HACCP Certifications

31
 Fig.27 BRC & BAP Certifications

PRINCIPLES OF SOP

The “eight” principles of SOP are as follows:

i. Safety of water
ii. Condition & cleanliness of food contact surface
iii. Prevent cross contamination
iv. Hand washing, hand sanitizing & toilet facilities
v. Protection from adulterants, microbes
vi. Labeling, storage & using of toxic compounds
vii. Control of employee health conditions
viii. Exclusion of pests

ONLINE QUALITY CONTROL IN THE PLANT

In the plant production line, the quality is being monitored by the team of QC at every stage of the
production line so as to ensure the quality of the product is made to meet the specifications or the

32
standards. There are reports to monitor the quality of the product, which are recorded or monitored
by the QC at each section of the processing line.

From the raw material receiving section to value addition section, the temperature of the material
is monitored so to maintain its temperature below 4 oC, checking the presence of the sulphite by
using sulphite strip test, taking the samples from every batch of the raw material & checking them
and sending the samples of each batch for the analysis purpose, inspecting the cleanliness of the
areas of the production, the hygienic conditions of the personnel, checking the levels of the
chlorinated water whether the level of chlorination is met or not and monitoring the defects,
uniformity of the meat pieces, checking the meat count and monitoring whether the material is
being covered by ice in the pre-processing sections of the production line or not.

The following are the formulae related to the above concerned processing sections:

➢ YIELD% = HLSO Wt./HON Wt. X 100

Where, HLSO Wt. = Wt. of headless variety obtained after the removal of head part

HON Wt. = Wt. of head-on variety taken (as in 1 or 2kg)

➢ HLSO Count = HON Count / 1.54

Here, this is known as the conversion factor in the grading section to get the count of
headless variety per lb.
➢ Meat Count = HLSO Count / Yield%(in decimal points)
It is checked whether the meat count is accurate according to the HLSO count in the value
addition section
➢ Veins% = No. of veins / Tot. no. of pieces x 100
It is checked in order to know the percentage of veins present in the respective sample
taken. It should be <3% to met the specifications
➢ Soft shell% = No. of soft shells / Tot. no. of pieces x 100
It is monitored to minimize the number of soft shells in a certain lot for better attainment
of the food product quality
➢ Grade = Wt. of 1lb (454g) / Piece Wt.

33
 This is to cross-check the count by taking the sample of a batch in the production line.

Table No.09 Grade & its approximate min. and max. wt. of each piece
Grade /count range Minimum wt. of each Maximum wt. of each
piece(g) piece(g)
U5 >90.8 -
U10 45.4 75.6
8/12 37.8 56.75
13/15 30.2 35
16/20 23 28.3
21/25 18.2 22
26/30 15.1 17.5
31/40 11.35 15
41/50 9.08 11.1
51/60 7.5 9
61/70 6.5 7.44
71/90 5.04 6.6
91/110 4.12 5
100/200 2.3 4.5

Reports like raw material monitoring report, temperature inspection log, and vehicle
cleaning report, beheading section log, grading section log, value addition report, etc are
being recorded for each and every batch of the production.

In the block section, the meat is being block frozen as per the order by the buyer. The samples are
taken for checking the meat count, defects identifying, etc by QC

The meat count is obtained by taking a sample, measuring its wt. and counting the no. of pieces
present and the count either for 1lb or for 4lb is calculated by using the following formulae:

➢ Count per one lb = No. of pieces x 454(1lb) / Net quantity taken (4lb or 1816g)
➢ Count per 4 lb = No. of pieces x 1816(4lb) / 454g or 1 lb

34
In the IQF section, the glaze% is calculated by following the glaze procedure:

1. Fix the water bath 8 times weight of the sample and held at 20oC (Frozen product)
2. Take the sample in a drainer held at -18oC for glaze checking
3. Immerse it in the water and agitate for 20-25s until the glaze is removed by feel
4. Allow the product to drain for about 2 minutes
5. Re-weigh the sample product (Deglazed product)
6. Calculate the glaze% by the following formula

• Glaze% = Frozen wt. of the product – Deglazed wt. of the product X 100
Frozen wt. of the product

The other formulae used in knowing the net count of the product, frozen count of the product
and filling wt. per carton are as follows:

➢ Net count = Tot. no. of pieces x 454g / Deglazed weight of the product
➢ Frozen count = Tot. no. of pieces x 454g / Frozen wt. of the product
➢ Filling wt. per carton = Frozen wt. / Deglazed wt. x Packing style

The monitoring of the temperatures at each section is very necessary as the temperature is a factor
in the maintenance of the quality of the product in the production line, the limits of temperature at
different sections in the production line is as follows:

Table No. 10 Limits of the temperature to be maintained at various sections in the plant

Name of the section / area/ product Temperature to be maintained (oC)

Raw material <3.8
Processing section <16
Chill room <4
Block Freezing -40
IQF -40
Cold store <-18

Application of HACCP system

35
It is the scientific, rational and systematic approach to identify, assess and control hazards during
production, processing, manufacturing and use of food. It controls hazardous elements in the food
system such as containments, pathogenic microorganisms, and physical objects, chemicals, etc for
the safety of customer or the storage condition.

The procedures for the implementation of HACCP system in the company are being followed.

There are 3 CCPs that are being checked by the company in the production line. They are:

i. CCP 1 – RAW MATERIAL RECEIVING ( SULPHITE)

ii. CCP 2 –PACKING AND LABELING
iii. CCP 3 – METAL DETECTION

At the metal detection area, the test kits used for checking the functioning of the detector are:

a) Ferrous test kit (Fe)– 1.5*1.5*1.5 mm

b) Non-ferrous test kit (N Fe) - 2*2*2 mm
c) Stainless steel test kit (SS) - 2.5*2.5*2.5 mm
QUALITY ASSURANCE LAB

In the quality assurance lab, various procedures for different analyses are followed so as to meet
the standards set or the requirements of the buyer for a better quality food product.

The equipment present in the lab are weighing balances, sealing machine, turbo evaporator,
heating mantle, distillation unit, distillation unit, vortex mixers, centrifuges, digital colony counter,
pH meter, freezers, hot air oven, microwave oven, autoclaves, hot plate, water bath, incubators,
BOD incubator, laminar air flow chamber, TDS meter, etc.

The tests or procedures that are done in the lab are follows:

➢ Water analysis
➢ pH analysis & TDS measurement
➢ Sulphite analysis
➢ Chlorine% estimation method
➢ Chloroscope test
➢ Filth analysis

36
 ➢ Fumigation
➢ Antibiotics test
➢ Swab analysis
➢ Microbial analysis

WATER ANALYSIS

Water analysis is done for all the water samples collected from various processing sections and
bore wells present in the plant

Principle

1. Ethylene di-amine tetra acetic acid (EDTA) or its sodium salts form a chelated soluble
complex when added to a solution of certain metal cautions.
2. If a small amount of a dye such as Eriochrome black T (EBT) or Calmagite is added to an
aqueous solution containing Ca & Mg ions at pH of 10.0±0.1, the solution becomes wine
red.
3. If EDTA is added as a titrate, the Calcium & Magnesium will be complexed.
4. The solution turns from wine red to blue, making the end point of the titration.
5. Mg ions must be present to yield a satisfactory end point. to ensure this, a small amount of
complexmatrically neutral magnesium salt of EDTA is added to the buffer, this
automatically introduces sufficient Mg.
6. The sharpness of endpoint increases with increasing Ph. The specified Ph of 10.0±0.1 is
quite satisfactory, a limit of 5 min. is set for the duration of titration to minimize CaCO 3
precipitation

37
Method

1. Take n/50 EDTA ( 3.72g/1000mL distilled water) in the burette

2. Transfer 20 mL of water in a beaker
3. Add 2 ml ammonia buffer
4. Dissolve 1 tot. Hardness indicator tablet / EBT (0.5g/100mL distilled water) /
Calmagite in the above water sample.
5. Add drop by drop EDTA from the burette
6. The end point will be when the color of the water becomes blue.

Calculation

Degree of Hardness (ppm) = [Vol. of EDTA ×50×0.02×1000] /Vol. of water taken

Fig. 28 Color indication of water before and after titration (water analysis)

38
pH ANALYSIS

A convenient scale for the reporting of hydrogen ion concentration is derived from

pH = log (10) 1 / (H)

Where (H) is the hydrogen ion activity

The value of the unknown solutions may be presented directly in pH units or mV.

Rules of pH

a. Strong acids & bases are dangerous and will cause serious chemical burns; safety
is a must
b. Solution must be aqueous to have pH
c. Because of the high resistance circuit (100 mega ohms). Moisture in connectors,
sensor or cable will shunt the pH meter
d. In using the two buffers, be careful not to cross contaminate. Two buffers are used
to check the span of the pH electrode.
e. Indication of broken glass electrode is that the pH meter will read about 6.5 pH
units.
f. In case of coated liquid junction or depleted electrolyte, pH meter will drift off
scale.
g. Coated glass electrode – slow response to pH change; pH meter will drift off scale
h. On installation, shake sensor down to get rid of bubbles in glass electrode. Wait
until the sensor reaches the same temperature as the solution before calibrating
i. Proper mixing of the process is essential in pH control.

Method

1. The calibration of pH electrode is done using buffer solution of pH 4

&7
2. Connect the combination pH electrode to the input socket, wash it with
water and switch on the instrument
3. Dip the electrode in the water sample to be tested

39
 4. Now observe the readings displayed on the panel
5. Record the values observed
6. Switch off the pH meter

Fig.29 pH meter

TDS MEASUREMENT

TDS means “Total Dissolved Solids”. The measurement of TDS is done the TDS meter. By
dipping the meter in the water sample, the TDS value is displayed and can be recorded.

Fig.30 TDS meter

ESTIMATION OF SULPHITE

The methods followed for the sulphite analysis in the lab are as follows:

40
a. Titration method
b. Monier Williams method

a. Titration method
1. Collect the sample, peel the shrimp and then cut it into pieces
2. Weigh 50g meat and transfer in 250mL conical flask and add 100 mL distilled water
3. Keep it in constant agitation during 10 min.
4. Withdraw with a pipette 10 mL of this maceration water and pour it in a conical
flask.
5. In the same flask, add 1mL HCl (37%) and add 1 mL starch solution (1%)
6. Titrate the sample with the iodine solution until the solution changes to blue color
7. Calculate the rate of SO2 residual according to the following formula

Formula

SO2 = Volume of iodine solution×0.5×1000×100 ppm

Weight of flesh sample ×10mL

Preparation of iodine solution as follows:

Potassium iodide 4.35g

Potassium iodate 0.45g Add 100mL distilled water & keep it in

Sodium bicarbonate 0.51g protected flask

Starch solution: dissolve 1g starch in 100mL distilled water (100oC heat) & cool to 50oC

b. Monier Williams Method

1. Collect the sample
2. Assemble the apparatus
3. Take 65 mL of 0.1N Sodium bicarbonate in gas purifying bottle
4. Add 400mL of distilled water, 90mL of HCl ( 30mL HCl & 60mL dist. water) in
round flask

41
 5. Take 30mL of 3%H2O2 and 2 drops of methyl red solution in gas collecting bottle
6. Give N2 gas connection through sodium bicarbonate solution
7. Apply power to the heating mantle. Provide the water flow
8. After 15 min., add 50g of crushed sample meat and then add 100mL of 5% ethanol
in round flask
9. Power setting that causes 80-90 drops/min.condensate to return the flask
10. Let the contents of the flask boil for 105 minutes
11. Remove the H2O2 flask
12. Immediately titrate with sodium hydroxide 0.01N, the end point is the appearance
of pale yellow color
13. Calculate the SO2 residual by the following formula:

SO2 = Volume of NaOH solution ×32.03×1000×0.01 ppm

50

Preparation of solutions:

0.1N Sodium bicarbonate – Dissolve 5.3g in 1000mL distilled water

0.01 N NaOH – Dissolve 0.4g in 1000mL distilled water

42
 Fig.31 Monier Williams Apparatus

ENUMERATION OF CHLORINE%

It is the procedure to test the chlorine% present in the chemical whenever a new lot/batch of
chlorine chemical is arrived in the plant

Procedure

1. Take 20mL distilled water in a 100mL conical flask and add 1mL of chlorine to it
2. Add 1g of Potassium iodide (KI)
3. Add 1ml acidified with acetic acid
4. Titrate against sodium thiosulphate
5. The color of solution turns to pale yellow color
6. Add 1mL of starch
7. Again titrate against sodium thiosulphate
8. Continue the titrations until the clear solution appears
9. Note down the end point
10. Calculate the chlorine% by using the following formula

43
 % Chlorine =Volume of Sodium thiosulphate x1000 x0.3546 / 100 x 10

CHLOROSCOPE TEST METHOD

 Take the test tube fill water sample up to 10mL (Mark).

 Add 1-2 drops of O-Toludine solution into the sample with help of dropper.

 Shake well and keep the tube for ten minutes to develop color.

 Compare developed color with chloroscope.

 Note down the reading of same from “ppm” scale marked in front of chloroscope.

 When testing is completed, pour out the sample, clean with fresh water and cleaning brush.

The water that is supplied to the plant is being treated with chlorine in order to reduce the
microbial load present in it. To add the required amount of chlorine in water, the following
formula is used for the calculation

Amount of chlorine to be added (mL) = Req.ppm X Qty. of water (in lt.)

Avail.Cl% X 100

Table No. 11: Chlorination levels

1. Processing 2ppm or below

2. Glazing 2ppm or below

3. Ice manufacturing 2ppm or below

44
 4. Hand Sanitization dip 20ppm

5. Foot Sanitization dip 50-100ppm

6. Washing of tables/utensils 50-100ppm

7. Washing of floor 100-200ppm

FILTH ANALYSIS

Filth Analysis is done to examine the microbial load and extraneous material, present in
the sample

Method

i. Take one half of shrimp block or IQF

ii. Put on a 12 diameter standard no.8 mesh sieve.

iii. Keep no.8 sieve on no.14 mesh sieve.

iv. Wash the shrimp block or IQF thoroughly with a forced shower of water
till the block is completely thawed.

v. Transfer all the material. Collected on no.14 mesh sieve to Wildman trap
flask

vi. Make the volume of water in flask to about 600mL

vii. Add 30mL Hexane. Tilt the flask about 400 angle by hand rotate it briskly
to 200-250 strokes per minute.

viii. Allow the mixture to stand for 30minutes. Frequently stirring the bottom
layer.

ix. Spin the stopper to remove sediment and trap off by raising the stopper to
remove sediments and trap off by raising the stopper to the neck of the flask.

45
 x. Hold the stopper in place and transfer the liquid into a beaker and add 30mL
of hexane to trap flask again and repeat the above procedure.

xi. Filter the contents of beaker by suction using a Buchner funnel and filter
paper. Moisten the filter paper with 1.1 glycerin and alcohol.

xii. Examine the filter paper at 30X magnification for extraneous material.

FUMIGATION PROCEDURE

• Put “OFF” the A/C unit

• Take Potassium permanganate (15g) in a petri dish and keep it in the area where fumigation
is to be carried.

• Place the petri plate on a polyethene bag

• Add about 25mL of 35% Formaldehyde solution to potassium permanganate (2:1).

• Immediately close the area

• Label the area as “Area Under Fumigation” so that nobody enters room.

• Keep the room under fumigation for a minimum of 8-12 hours

• Defumigate the room by putting on the A/C unit.

Hazards of Formaldehyde:

i. It is toxic to inhalation & ingestion and causes irritation to eyes and

respiratory tract
ii. Skin contact can cause dermatitis, also cause skin burns
iii. Exposure to a concentration of 10 to 20ppm can cause difficulty in breathing
iv. Slightly corrosive in nature, suspected carcinogen

ANTIBIOTICS TEST (BIO-SCIENTIFIC METHOD)

46
General information

In this method, ELISA procedure is used. It provides a competitive enzyme immunoassay for the
quantitative analysis of nitrofurans in feed, fish, shrimp, meat (beef, chicken and pork), eggs,
honey, milk, serum and urine.

Salient features of ELISA:

➢ Simple
➢ Rapid
➢ Sensitive
➢ Cost-effective

In the lab, the antibiotics tested are of 5 paramaters in which one is chloramphenicol (CAP) and
other 4 belong to a group termed as “nitrofurans” (AOZ,AMOZ,AHD,SEM)

Chloramphenicol (CAP) is abroad spectrum antibiotic which is frequently employed in animal

production for its excellent antibacterial & pharmacokinetic properties. However, in humans, it
leads to haematoxic side effects.

Nitrofurans (AOZ, AMOZ, AHD and SEM) have been widely &effectively used for the prevention
and treatment of gastrointestinal infections caused by E.coli & Salmonella spp.in cattle, pigs,
poultry and used as growth promoters in food producing animals.

However, both WHO & EU are unable to assign a maximum residue limit for nitrofurans
because of the potential carcinogenic effects of their residues on human health. Consequently, the
administration of nitrofurans to food producing animals has been prohibited.

Internal limits of antibiotics:

➢ CAP – 0.15ppb
➢ NITROFURANS:
• AOZ,AMOZ – 0.35ppb
• AHD,SEM – 0.5ppb

Sample extraction procedure

47
 i. Chloramphenicol (CAP)
1. Take 3g of homogenized sample
2. Add 6mL of ethyl acetate to the above homogenized sample
3. Shake for 5min.upside down by using cyclomixer
4. Centrifuge 10mins/4000rpm at room temperature (20-25oC)
5. Transfer 4mL of supernatant into a new tube and evaporate at 60oC to
complete dryness by N2 or air
6. Add 2mL of n-hexane to the dried residue
7. Add 1mL of sample extraction buffer (SEB) and vortex with cyclomixer for
1min.
8. Centrifuge at 4000rpm for 10 min.at room temperature (20-25oC)
9. Use 100µL of the lower aqueous phase per well in the assay
ii. Nitrofurans ( AOZ, AMOZ, AHD & SEM)
a. Mix 1g of homogenized sample with 0.5mL sample extraction buffer (SEB) and
3.5mL distilled water, 0.5mL 1M HCl and 80µL of 2-Nitrobenzaldehyde (in
DMSO) by shaking vigorously
b. Incubate at 60oC for 3h.
c. Add 5mL of 0.1M K2HPO4, 0.4mL 1M NaOH and vortex for about 10s
d. Add 5mL ethyl acetate and shake vigorously for 30s.
e. Centrifuge 10min/4000rpm at room temperature (20-25oC)
f. Transfer 3mL of ethyl acetate upper layer into a new vail and evaporate to dryness
at 60oC
g. Dissolve the residue in 2mL n-hexane and add 1mL sample extraction buffer (SEB)
& vortex for 30s
h. Centrifuge 10min/4000rpm at room temperature (20-25oC)
i. Use 100µL of the lower aqueous phase per well in the assay.

Test Procedure

1. Insert sufficient no. of wells into the microwell holder for all standards and samples
2. Add 100µL of each standard and 100µL of sample to the wells
3. Add 50µL of the conjugate to the bottom of each well

48
 4. Mix gently by shaking the plate manually
5. Incubate for 30 min.at room temperature (20-25oC)
6. Pour the liquid out of the wells and tap the microwell holder upside down
vigorously (3 times in a row) against absorbent paper to ensure complete removal
of liquid from wells
7. Fill all the wells with 250µL washing buffer and pour out the liquid again
8. Repeat two more times the above step
9. Add 100µL TMB substrate to each well.
10. Mix gently by shaking the plate manually
11. Incubate for 20 min.at room temperature (20-25oC) in the dark
12. Add 100µL of the stop solution to each well
13. Mix gently by shaking the plate manually
14. Measure the absorbance at 450nmwith the help of the equipment “Bio-rad ELISA
reader”. Read within 30 min.after the addition of stop solution

Preparation of chemicals

• 1M HCl : Take 8.3mL conc. HCl & make upto 100mL distilled water
• 1M NaOH: Dissolve 2g in 50mL distilled water
• 0.1M K2HPO4: Add 1.74g in 100mL distilled water
• SEB: Take 10mL SEB in 90mL distilled water (1:9)
• Wash Buffer: Add 10mL wash solution to 190mL distilled water

SWAB ANALYSIS

Swab collection:

• Collecting the samples of the areas which come in contact with the food
product
• To test the level of microbial contamination/load on various surfaces in the
plant.
A. WATER

49
 1. Water sample is collected from taps ( tap number to be mentioned
in the sample covering note) in sterile bottles/conical flasks of 1 liter
capacity with ground flask stoppers having an overhanging rim
2. They are sterilized at 160oC for 1h after being covered by kraft paper
3. The opening and closing of the sterile bottle must be done with
meticulous care to avoid any contamination
4. When water sample is drawn from a tap, flame the tip of the tap
using spirit and allow water to flow for 5 min.before collection
5. In case the test is to be undertaken after 3h, the bottle must be kept
in ice.
6. If sample is to be taken from chlorinated water supply, it is important
that any trace of chlorine should be neutralized immediately after
collection
7. A crystal of sodium thiosulphate or 0.1mL of 2% solution of sodium
thiosulphate introduced into the sampling bottle prior to sterilization
serves neutralization of chlorine
8. Immediately before testing, the water sample should be mixed by
inverting the bottle several times
9. Thereafter some of the contents are poured off, the stopper is
replaced and the bottle is shaken vigorously up & down
B. ICE
1. A minimum of 1kg of ice used for processing shall be collected
aseptically in a sterile stainless steel container & transported to the
laboratory
2. If there is a considerable delay from the time of withdrawal of
samples and actual analysis (>4h), the samples shall be kept in cool
condition
C. WORKER’S HAND & FOOD CONTACT SURFACES
a. 25cm2 area are swabbed using a square 5 X 5 cm steel template of
correct size, which can be readily sterilized by alcohol flaming can
be used to outline the area

50
 b. First wipe the swab slowly & firmly in an interior direction through
a distance of 12.5cm
c. Rotate the swab against the direction of overall wiping movement
d. Then stroke the area in the same direction 3 times, turning the swab
slightly between the strokes
e. Finally roll the swab once over the wiped area, but in the opposite
direction from that in which the original strokes were made
f. This will serve to pick up whatever maybe adhering to the surface
g. Place the swab immediately into bottle containing 100mL of the
diluents, in a wide mouthed bottle
h. Pull the stick free if the swab in the medium is to be transported,
hold it under the same condition as water samples are being
transported i.e., hold it below 5oC until analyzed
i. For the analysis of V.cholerae from worker’s hand, a separate swab
shall be collected
j. Swab from the worker’s hand shall be drawn from different parts of
hand and aseptically transported immediately into a flask
containing 100mL of sterile alkaline peptone water at pH 8.6
k. The sample collected shall be transported to the laboratory in the
usual manner under sealed condition and accompanied by covering
note containing the details of the tests to be carried out

Maximum permissible limits ( Table No.12)

S.No Sample TPC (37OC) TPC(22OC) Coliforms V.cholerae
1. Water 20/mL 100/mL Nil Absent
2. Ice 20/mL 100/mL Nil Absent
3. Table/utensil/trays 100/cm2 - Nil Absent
4. Worker’s hand 100/cm2 - Nil Absent

MEDIA PREPARATION AND AUTOCLAVE METHOD

51
 The method for the preparation of media is provided on the pack. It gives the quantity of
powder to be suspended per liter of water for injection. Weigh required quantity of powder and
add required quantity of freshly collected water for injection. Note down the weights mix well to
dissolve and check the pH of the media before sterilization.
The equipment shall be autoclaved at 121oC for 15 minutes. The following precautions
shall be observed here:
a) Cover the plugs of cotton wool with kraft paper. Similarly wrap the items whose
external surface is to be kept sterile
b) Loosen cap closures before autoclaving and tighten again on cooking
c) Distribute the items within the autoclave so that steam will be able to circulate
freely. Ensure that steam will have access to all containers and that there will be
no pockets of trapped air
d) Make sure that there is enough water in the autoclave that coil is fully immersed
in water
e) Ensure that all air is expelled from the autoclave before it is close. If the air is not
discharged fully, the difference in temperature for the same pressure is
considerable

The following table (Table No. 13) will clearly show this:

Temp. (inoC) when air is expelled

Pressure 100% 50%
15 lbs 121 112
20 lbs 126 116
Allow slight bleed of steam during sterilization
MICROBIAL ANALYSIS
Microbial analysis is done to check the microbial loads present in the samples brought to
the lab. In the lab, for the microbes like E.coli, Salmonella typhi, Staphylococcus aureus, Vibrio
cholera, V.parahaemolyticus, etc; the test procedures are carried out.

52
 The media that are used for the microbial analysis, are prepared by adding the required
quantity in the known volume of distilled water, dissolving the media either by manually shaking
or by keeping them on the hot plate and then autoclaved at 121 oC for about 15-20 minutes at 15-
20lbs pressure.

And later, they are further proceeded for inoculation process by using the serial dilution
method or by directly inoculating a certain amount, kept in the incubators & transfer to the
sterilized petri plates either by streaking process or by spread plate technique. The plates are
incubated for specific time period at specific temperature. The results are recorded as per the
observations of the tests.

The media are prepared as per the batch samples received for each day. They are prepared
either for a single batch (PB) or for as composite of two batches (APW, LB)

The following are the procedures followed in the lab for the microbial analysis:

PROCEDURE FOR TOTAL PLATE COUNT (TPC)

25g of homogenized sample + 225mL sterile phosphate buffer (10-1)

10mL of above mixture

90mL sterile phosphate buffer (10-2)

1mL

9mL sterile phosphate buffer (10-3)

1mL

53
 Transfer 1mL of 10-2 & 10-3 dilutions each into sterile petri dishes & pour 12-15mL of
plate count agar (at 45±1oC) & mix well

[Note: Duplicate plates should be prepared for each dilution]

Let the media solidify, invert & incubate at 35±2oC for 48±2h

Count no. of colonies on all plates with 25 to 250 CFU presents & calculate for CFU present
& calculate for TPC/g by substituting in below formula

For plates with 30-300 CFU:

Calculate the TPC as follows

N = Sum of all colonies on all plates counted

[( 1 X n1 ) + (0.1 X n2)] X (d)

Where, N = no. of colonies per g of sample

n1 = no. of plates in first dilution

n2 = no. of plates in second dilution

d = dilution from which the first count were obtained

Note:

If any plate is having <30 CFU’s, consider it as TLTC (too low to count) and if any plate is having
>300 CFU’s, consider it as TNTC (too numerous to count)

In case of presence of TNTC or TLTC plates in one dilution, consider only other dilution plates.
In that case, calculate TPC as follows.

N = Total no. of colonies in appropriate plates X D

54
 No. of duplications

Where, N = no. of colonies per g of sample

D = appropriate dilution factor

Note:

In case of all TNTC or all TLTC plates, do the re-sampling by increasing or decreasing the dilution
respectively. [ AOAC. 1990]

PROCEDURE FOR E.COLI AND TOTAL COLIFORM COUNT

Presumptive test:

25g of homogenized sample is mixed with 225mL of phosphate buffer (10-1)

10mL sample is added to 3 1mL sample is added 0.1mL sample is added to 3 tubes of
tubes of 10mL double strength to 3 tubes of 10mL 10mL of single strength sterile LSTB
single strength sterile with durham’s tube

55
 sterile LSTB with durham’s LSTB with durham’s
tube tube
Incubate at 37oC for 24h

Acid + gas production is a +ve result

Confirmative test:

A loopful of culture from the +ve tubes is transferred to 5mL of sterile EC broth with
durham’s tubes Incubate at 45.5±0.2oC for 24h

Acid + gas production is a +ve result

A loopful of culture from the positive tubes is streaked with EMB agar plates

Incubate at 37oC for 24h

The EMB agar plates appear dark purple with metallic shine colonies are counted, isolated
& purified to Tryptose Soya Agar (TSA) plates and further subjected to biochemical test
for identification

Transfer 1mL of each dilution to 3 LST tubes, for 3 consecutive dilutions

Biochemical test:

1. Eijikman test +ve

2. Indole production test +ve
3. Methyl red +ve
4. Citrate utilization +ve
5. Voges proskauer test +ve

Isolation and identification of E.coli by “direct method”:

56
25g of composite samples is mixed with 225mL of PB or physiological saline, which gives 10 -1
dilution, 0.1mL inoculums from the dilution is spread plated on Terigitol-7 agar (T-7)

The plates are incubated at 37oC for 48h. E.coli colonies on T-7 plates appear as circular, non-
mucoid, flat, and yellow with pinkish tinge.

E.coli (cfu/g) = No. of positive colonies X 100

Weight of the sample

Suspected typical colonies are sub-cultured onto TSA plates and subjected to the following tests
for confirmation.

Biochemical tests:

1. Eijikman test +ve

2. Indole production test +ve
3. Methyl red +ve
4. Citrate utilization +ve
5. Voges proskauer test +ve

PROCEDURE FOR VIBRIO CHOLERAE & VIBRIO PARAHAEMOLYTICUS

25g of homogenized sample + 225mL of sterile alkaline peptone water (APW)

Primary enrichments:

Incubate at 37±1oC for 16h

57
 Streak a loopful of incubated APW suspension on 24h old TCBS (with 3%
salt or without salt) plates to obtain isolated colonies

Secondary enrichments:

After incubation, 0.1mL culture is transferred to 10mL sterile alkaline peptone water
(APW)

Incubate the broth at 37oC for 6h

Streak on 24h old TCBS (with 3% salt or without salt) plates

Incubate the broth at 37oC for 24h

Observe the without salt TCBS plates for colonies which are large, smooth, yellow &
slightly flattened with opaque centre and translucent centre, as V.cholerae, and 3% salt
TCBS plates, moderate greenish colonies are referred to Vibrio parahaemolyticus

If typical colonies were observed, proceed for the following biochemical tests

Presumptive tests for VC & VP

Subculture, three or more typical colonies from each of the TCBS (Thiosulphite Citrate
Bile salt Sucrose) plates into nutrient agar slants

Incubate each suspected culture into separate tubes containing TSI & KI agars by streaking
the slant & stabbing the butt

58
 Incubate overnight at 37±1oC

Observe for typical reactions of VC & VP

Presumptive test & biochemical reaction of VC & VP (Table No. 14)

REACTIONS V.CHOLERAE V.PARAHAEMOLYTICUS

Gram stain Gram –ve rods Gram –ve rods
Motility Motile Motile
Growth in tryptone at 37±1oC Indole +ve Indole +ve
TSI (Triple Sugar Iron)agar Acid slant & acid butt H2S – Acid slant & acid butt H2S –ve
ve (yellow color) (yellow color)
KI (Kligler Iron) agar Alkaline slant (red) & acid Alkaline slant (red) & acid butt
butt (yellow) H2S -ve (yellow) H2S -ve
Lysine decarboxylase +ve +ve
Arginine dihydrolase -ve -ve
Ornithine decarboxylase +ve -ve/+ve
Inositol -ve -ve
Mannitol +ve (acid) +ve
Oxidase +ve +ve
Catalase +ve +ve
Kanagawa reaction -ve +ve
Voges proskauer -ve -ve
Growth in peptone water +ve -ve
containing no NaCl
Growth in peptone water -ve +ve
containing 8% NaCl
Growth in peptone water -ve -ve
containing 11% NaCl

59
PROCEDURE FOR SALMONELLA

25g of homogenized sample + 225mL sterile fluid lactose broth (LB) and blend for 2 min.
let stand for 60 min. at room temperature

Incubate at 35±2oC for 24±2 h

Transfer 0.1mL to 10mL RV (Rapport Vasilidus) broth and 1mL to TTB (Tetra Thionate
Broth) respectively

Incubate them at 42±0.2oC for 24±2h (water bath) & at 35±2oC for 24±2h (water bath)
respectively

Streak a loopful of culture on BSA, XLDA & HEA agar and incubate at 35oC for 24±2h

Examine for typical Salmonella colony morphology

I. HEA (Hektone Enteric Agar) plates: Blue green to blue colonies with or without black
centres. (may be appearing as glossy black center or almost completely black colonies)
II. XLDA (Xylose Lysine Desoxycholate Agar) plates: Pink colonies with or without black
centers. (may be appearing as large glossy black center or almost black colonies)

60
III. BSA (Bismuth Sulphite Agar) plates: Brown, grey or black colonies (with metallic shine
sometimes) Surrounding medium is usually brown at first. But may turn black for longer
incubation (halo effect)

Procedure for presumptive testing for typical Salmonella

1. Lightly touch the very center of the colony by sterile needle and inoculate TSI slant by
streaking slant & stabbing butt
2. Without flaming, inoculate LIA slant by stabbing deep butt (4cm) twice & then streak slant
3. Incubate TSI & LIA slant at 35oC for 24±2h. cap tubes loosely to maintain aerobic
condition
4. For TSI (Triple Sugar Iron Agar): Salmonella typical produce alkaline (red) slant & acid
(yellow) butt. With or without H2S production
5. For LIA (Lysine Iron Agar): Salmonella typical produces alkaline (purple) reaction in
butt of tube. Consider only distinct yellow color in butt of tube as positive (acid reaction).
Don’t silminate culture that produce discoloration in butt of tube solely on this basis. Most
Salmonella cultures produce H2S in LIA. For future confirmations, proceed for
biochemical tests & then serological test.

Biochemical & serological reaction of Salmonella(Table No. 15)

Test or substrate Result Salmonella sps.

+ve -ve reaction
Glucose (TSI) Yellow butt Red butt +
Lysine decarboxylase Purple butt Yellow butt +
(LIA)
H2S (TSI & LIA) Blackening No blackening +
Ureases Purple-red color No color change -
Lysine decarboxylase Purple color Yellow color +
broth
Phenol red dulcitol Yellow color &/ or No gas; no color +
broth gas change
KCN broth Growth No growth -

61
 Malonate broth Blue color No color change -
Indole test Violet color at surface Yellow color at -
surface
Polyvalent flagellar Agglutination No agglutination +
test
Polyvalent somatic Agglutination No agglutination +
test
Phenol red lactose Yellow color &/ or No gas; no color -
broth gas change
Phenol red sucrose Yellow color &/ or No gas; no color -
broth gas change
Voges proskauer test Pink to red color No color change -
Methyl red test Diffuse red color Diffuse yellow color +
Simmons citrate Growth;blue color No growth; no color V
NOTE: +, 90% or more +ve in 1 or 2 days; -, 90% or more –ve in 1 or 2 days; v,variable

Sample collection for USA & other than USA

Procedure for pre-enrichment

For countries other than USA

a) Aseptically weigh 25g of the sample & transfer into a sterile blending container
b) Add 225mL of lactose broth & blend for 2 min at high speed
c) Aseptically transfer the homogenized mixture to a sterile wide-mouthed screw cap
jar of 500mL capacity or other appropriate container & let the homogenate stand
for 60 min at room temperature
d) Ensure well mixing by swirling & determine the pH with test paper. If necessary,
adjust the pH to 6.8±0.2
e) Loosen the jar caps by ¼ turn & incubate the sample mixture for 24±2h at 37±1oC

For samples drawn from consignments meant for export to USA

a) Draw 375g of sample for Salmonella analysis

62
 b) Add 3750mL of a sterile lactose broth
c) Proceed further as described above under (i) above from (a-e) (AOAC.2000)

PROCEDURE FOR STAPHYLOCOCCUS AUREUS

25g of homogenized sample + 225mL sterile phosphate buffer (10-1)

For each dilution to be plated, aseptically transfer 1mL sample suspension to 3 plates of
BPA (Baird Parker Agar), distributing 1mL of inoculum equitably to 3 plates (e.g., 0.4mL,
0.3mL, & 0.2mL). Spread the inoculums over surface of agar plate, using sterile bent glass
streaking rod

Retain the plates in upright position until inoculum is absorbed by agar (about 10min on
properly dried plates). If inoculum is not readily absorbed, place the plates upright in
incubator for about 1h. Invert the plates & incubate 45-48h at 35-37oC

Select the plates containing 20-200 colonies, unless only plates at lower dilutions (> 200
colonies) have colonies with typical appearance of S.aureus.

Colony morphology:

Colonies of S.aureus are circular, smooth, convex, moist, 2-3mm in diameter on uncrowded plates,
gray to jet-black, frequently with light-colored (off-white) margin, surrounded by opaque zone &
frequently with an outer clear zone; colonies have buttery to gummy consistency when touched
with inoculating needle.

Typical characteristics of S.aureus, S.epidermidis, & Micrococci (Table No. 16)

63
 Characteristics S.aureus S.epidermidis Micrococci

Catalase activity + + +

Coagulase production + - -

Thermonuclease + - -
production

Lycostaphine + + -
sensitivity

Anaerobic utilization + + -
of
a) Glucose + - -
b) Mannitol

NOTE: +, most (90% or more) strains are positive; -, most (90% or more) strains are negative
(AOAC.1995)

The following table (Table No. 17) describes the media used, incubation time, colony
morphology and effects on humans of the respective microbes that are being tested:

S.No Tests Media Incubation Colony morphology Effects

on
humans
1. TPC PB 35±2oC for -- --
48±2 h
TGB

64
2. E.coli PB 37oC for 48h Circular, non-mucoid, flat, Cold
yellow with tinge
LSTB
TSA
3. Salmonella typhi LB 35±2oC for 24h Brown/grey/black colonies Typhoid
42±2oC for 24h
RV,TTB
35±2oC for 24h Pink colonies with/without
BSA,XLDA,HEA
black centres

Blue-green to blue colonies

with/without black centres
4. Staphylococcus aureus PB 48h at 37oC Circular,smooth,convex,mois Cough
t,2-3mm in dia., light color
BPA
margin surrounded by opaque
zone,buttery with inoculating
needle
5. Vibrio cholera, APW 37±1oC for 16h Withoutsalt-large,smooth, Cholera
37oC for 16h yellow & slight flattened with
V.parahaemolyticus TCBS,TCBS with
37oC for 16h opaque centre & translucent
3% salt
With salt –moderate greenish

Internal microbial standards(Table No. 18)

Parameter Raw product limit Cooked product limit

Total Plate Count <1, 00,000 cfu/g <10,000 cfu/g
Total coliforms <100/g <10/g
E.coli <20/g Negative/g
Salmonella sps. Negative/25g Negative/25g
Staphylococcus sps. <100/g < 10 MPN/g
Vibrio cholera Negative/25g Negative/25g

65
 Vibrio parahaemolyticus Negative/25g Negative/25g

DISCARDING PROCEDURE

1. Incubated media should not be discarded directly.

2. Certain type of media has to be kept in autoclave at 121°C showing 15 lbs pressure.

3. Fresh deodorizing pearls (Tablet) which is used in order to reduce bad smell.

4. Take out the glassware from autoclave after discarding procedure.

5. Dispose the discarded media.

6. Wash all the glassware under running tap water and then with soap oil with proper
scrubbing to shiny layer using scrubber or brush.

7. Soak overnight in acid water (0.25% HCl or H2SO4)

8. Again wash with soap oil and let all the water drain-off and clean with spirit.

9. Keep in hot air oven at 161°C for 1hour

10. Let the glassware cool and use them for practical purpose aseptically

EFFLUENT TREATMENT PLANT

The ETP is kept running round the clock to treat the water used in the processing plant before
being discharge into the canal. The scope of ETP is that the effluent is to meet the standards set by
Pollution Control Board (PCB) and be environment eco-friendly.

66
 Fig.32 Flow diagram of the process of the effluent treatment plant

The procedure followed in the Effluent Treatment Plant (ETP) can be described as below:

• The water from the processing plant goes through a grit chamber to remove items like
sticks, poly pouches, polythene sheet, paper and other debris.
• The processing water from grit chamber falls into the collection tank or settlement tank
• In the settlement tank, lime is added at the rate of 10kgs/day in 2 lots to control pH & help
in the settlement of oil if any present.
• Water from the settlement tank is now pumped to the aeration tank or chamber.
• From the aeration tank, water is passed through the sludge beds, sand filter, activated
carbon filter and the chlorine dosing is done.
• Later, this water is used for client scope i.e., a) Gardening purpose & b) Flushing and
bathroom purpose.

CONCLUSION

In this four month In-plant training programme with all the support from the staff in the plant, we
had the opportunity to explore different sections of the ROYALE MARINE IMPEX PRIVATE
LIMITED, Chirala. We were given the opportunity to observe & practically experience the
different sections in the production line and quality related aspects in the plant.

While working with the company of such magnitude where there is no margin of error, we had the
opportunity to understand the importance of quality & how even the tiny things matter so much in
a highly competitive market. The company not only works towards the efficiency of the processes
but also takes the responsibility in using the resources & also in the treatment of the water & the
effluents that are being produced in the plant.

The following are some of the features of the company:

67
 • The quality system that is based on HACCP, SOP, SSOP, GMP, etc., is well
documented and implemented & gives the possibility to ensure food safety.
• The company has an efficient recall procedure, which is important in case of a problem
with the product, as it can help to locate the problem.
• The traceability system in this company is based on a good labeling system & record
keeping detailing the history of the product.
• The use of microbiological testing is also an important tool to verify the quality system
related to hygiene and the safety of the product and the production line.

Understanding how the record keeping regarding traceability works in this company was also a
practical experience.

68