Extraction of casin from various milk sample & role in bioplastic formulation.

A Research Project submitted to the

C. G. Bhakta Institute of Biotechnology

(UKA TARSADIA UNIVERSITY, BARDOLI)

As partial requirement for the fulfillment of

The Degree of

MASTER OF SCIENCE IN MICROBIOLOGY

By

BODA MANSI H.

(201904101010001)

Under the guidance of

Dr.Naga Rathna Supriya

Mrs Shreya desai

C. G. Bhakta Institute of Biotechnology

UKA TARSADIA UNIVERSITY
Gopal Vidyanagar, Bardoli – Mahuva Road, Dist. SUrat – 394350

september 2019
 PART I: GENERAL INFORMATION

1. Name of the investigator : boda mansi h.

2. Institute : C. G. Bhakta Institute of Biotechnology

3. Semester : I

4. Project Title :Extrection of casin from various milk sample

& Role in bioplastic formulation

5. Specific Area :Microbial

6. Duration :four semester

7. Project Summary (Not to exceed one page. Please use separate sheet)
 PART II: TECHNICAL DETAILS OF PROJECT
(Under the following heads on separate sheets)

8. Introduction:
Histroically,petroleum based plastics have been formulation to increase the
durability and reduce photo,chemical and biological degradation. Popular
chemical formulation include polyethylene,polystyrene,and polyurethane.
However , recent concern about the persistence of plastic materials in the
environment, either as litter or through landfill disposal of municipal solid wastes
, has prompted a rethinking of the use of plastic based products. In several states
and communities legislation has been enacted requiring that biodegradable
plastics be used in common short term application such as packaging materials.
The use of petroleum based synthetic plastics have led to solid waste management
problems and secondary problems such as global warming, which is caused by
increased carbon dioxide levels in the atmosphere during the manufacturing and
disposal processes.conventional plastics are resistant to biodegradation , as the
surfaces in contact with the soil in which they are disposed are characteristically
smooth. Microorganisum within the soil are unable to consume a portion of the
plastic , which would, in turn , cause a more rapid breakdown of the supporting
matrix. This group of materials usually has an impenetrable petroleum based
matrix, which is reinforced with carbon or glass fibers. Biopolymers are
renewable, largely biodegradable and can have very similar properties to
conventional polyolefin polymers. These are all desirable properties given that
the use of conventional plastics is wide spread. Polyolefin are produced from
fossil fuels and resources depletion may become a determining factor in future
production. Understanding their environmental impact is important when
comparing then to biopolymers.
2. biodegradable plastic from milk protein:casein
Casein is a naturally occurring macromolecule that account for approximately
80%of the protein content of cow’s milk ; it is a phosphoprotein that can be
separated into various electrophoretic fraction, such αs- casein,κ-casein,β -casein,
and γ-casein in which each constituent differs in primary, secondary, and tertiary
structure, amino acid composition, and molecular weight. The casein family of
protein consists of several types of casein and each has its own amino acid
composition genetic variation, and functional properties. The casein are
suspended in milk in a complex called a micelle. The casein have a relatively
random, open structure due to the amino acid composition. the high phosphate
content of the casein family allows it to associate with calcium and from calcium
phosphate salts. But casein is not very strong, and water can wash it away. To
beef up casein, and boost its resistance to water , the scientists blended in a small
amount of clay and reactive molecule formaldehyde, which link casein’s protein
molecules together.
Milk contains 3.3% total protein. Milk protein contain all 9 essential amino acids
required by humans. Milk proteins are synthesized in the mammary gland, but
60% of amino acid used to build the protein are obtained from the cow’s diet.
Total milk protein content and amino acid composition varies with cow breed
and individual animal genetics there are 2 major categories of milk protein that
are broadly defined by their chemical composition and physical properties. The
casein family contains phosphorus and will coagulate or precipitate at PH 4.6.
the serum (whey) proteins do not contain phosphorus, and these protein remain
in solution in milk at PH 4.6. the principle of coagulation, or curd formation. In
cow’s milk approximately 82% of milk protein is casein and the remaining 18%
is serum, or whey protein.
2.2 casein structure
Protein are probably the most important class of biochemical molecules, along
with lipids and carbohydrates which are also essential of life. Casein is protein
that is found in milk which is used independently in many food as binding agent.
Amino acid have variety of chemically relative group like phenolic hydroxy
group, presence of peptide bond. Casein also includes amino group, ketones and
hydrazine group. Its structure is show as:

Fig:casein structure

3. extraction of casein from milk

When a sample of non-fat milk is precipitated using acetic acid after heating
milk, the casein separates from the sample leaving a clear solution by stirring the
solution while adding acid, the casein separates from the milky solution as an
amorphous mass which could be removed using suction filtration. When acetic
acid is added into the milk sample it from precipitates. The precipitates are then
washed 2 or 3 times using water after decanting off the filtrate which is then
rudded with sufficient amount of 0.1% sodium hydroxide solution and then the
resulting solution is filtered using a cloth. Precipitates terms as casein are then
treated with absolute alcohol in a breaker. The separated casein is then mixed
with the ether solution for a few minutes, the ether is allowed to settle, and the
process is repeated with a second potion of ether. After the second washing with
ether again filter the product. The ether washing remove any small quantities of
fat that may have precipitated with the casein.

3.1 confirmation of presence of casein

Since casein structure is composed of variety of functional group like carbonyl

group , phenolic hydroxyl group, amino acid, peptide bonds it can be confirmed
using confirmatory tests for various functional group present in it.

9. Objectives:
10. Work plan (methodology/experimental design to accomplish the stated aim)

Materials and Method: CASEIN PREPARATION

The milk solution is prepared by adding 80ml of distilled water and 20g of non-

fat milk powder by proper stirring. The formed solution of non-fat milk is

precipitated using 1.2milliliter of glacial acetic acid after heating it till a

temperature of 55 c . the casein separates as precipitates form the sample

leaving a clear solution. By stirring the solution while adding acid, the casein

separates from the milky solution as an amorphous mass which could be

removed using suction filtration. The precipitates are then washed 2 or 3 times

using water after decanting off the filtrate which is then rubbed with sufficient

amount of 0.1% sodium hydroxide solution and then the resulting solution is

filtered using a cloth. Precipitated termed as casein, then treated with absolute

alcohol in a beaker. The separated casein is then mixed with the ether solution

and then the resulting solution for a few minutes, the ether is allowed to settle,

and the process is repeated with a second portion of ether. After the second

washing with ether, again filter the product. The ether washings removed any

small quantities of fat that may have precipitated with casein.

PROCESS YIELD

Weight of milk powder used initially = 20gram

Weight of glacial acetic acid = 1.015gram

Weight of product obtained = 16.74gram

Process yield
dry weight of product obtained
thus a process yield of 79.657% is obtained for

11. Timelines: (Please provide quantifiable outputs)

Period of study Achievable targets

From starting date
Semester 7
Semester 8
Semester 9
Semester 10

12. List of references in the text