Biochem Lab 2
Biochem Lab 2
2.1.
OBJECTIVE
2.2.
INTRODUCTION
Lipids are relatively insoluble in water. They are soluble in non-polar solvents,
like ether, chloroform, methanol. Lipids have high energy content and are metabolized to
release calories. Lipids also act as electrical insulators, they insulate nerve axons. Fats
contain saturated fatty acids, they are solid at room temperatures. Example, animal fats.
Plant fats are unsaturated and are liquid at room temperatures. Pure fats are colorless,
they have extremely bland taste. The fats are sparingly soluble in water and hence are
described are hydrophobic substances. They are freely soluble in organic solvents like
ether, acetone and benzene. The melting point of fats depends on the length of the chain
of the constituent fatty acid and the degree of unsaturation. Geometric isomerism, the
presence of double bond in the unsaturated fatty acid of the lipid molecule produces
geometric or cis-trans isomerism. Fats have insulating capacity, they are bad conductors
of heat. Emulsification is the process by which a lipid mass is converted to a number of
small lipid droplets. The process of emulsification happens before the fats can be
absorbed by the intestinal walls. The fats are hydrolyzed by the enzyme lipases to yield
fatty acids and glycerol. The hydrolysis of fats by alkali is called saponification. This
reaction results in the formation of glycerol and salts of fatty acids called
soaps. Hydrolytic rancidity is caused by the growth of microorganisms which secrete
enzymes like lipases. These split fats into glycerol and free fatty acids.
Lipids are soluble in some organic solvents. This property of
specific solubility in non polar solvents is utilized for extracting lipid
from tissue. In biological materials, the lipids are generally bound to
proteins and they are, therefore, extracted either with mixture of
ethanol and diethyl ether or a mixture of chloroform and methanol.
2.3.
Conical flasks
Separatory funnel
Seeds (mung bean)
Anhydrous sodium sulphate
Diethyl ether : ethanol mixture (2:1,v/v)
1% sodium chloride
Water bath
Rotavapor
Agitator
Pestle and mortar
2.4.
PROCEDURE
sodium chloride.
Take the pooled fractions in a separatory funnel, shake it
thoroughly and allow it to stand for 5 minutes. The lipids will be
recovered in diethyl ether layer while soap, glycerol and other
2.5.
RESULTS
2.6.
2.7.
2.8.
Group
2.28.
of
beaker
5.415
0.940
4.171
3.703
4.418
5.482
0.975
4.212
3.754
4.436
of
B 2.9.
.0670
.0359
.0406
.0516
.0180
empty beaker)
2.29.
2.36.
2.37.
2.38.
2.39.
Grou
p
2.46.
Perc
2.40.
2.41.
2.42.
2.43.
2.44.
2.45.
B1
B2
B3
B4
B5
B6
2.47.
2.48.
2.49.
2.50.
2.51.
2.52.
3.36
3.35
1.79
2.03
2.58
0.90
entage of
crude
lipid
5
weight of crude lipid
100
2 g of seed
2.53.
2.54.
2.55. DISCUSSION
2.56. From the experiment, lipid can be extracted efficiently
using rotavapor which than the fat content can be determined by
filtrates to remove the solvent. Our group had chosen mung bean as
the sample for the experiment. The sample is prepared by mixing
with anhydrous sodium sulfate using a ratio 4 to 1. Sodium sulfate is
important to absorb any water from the sample. This mixture is
inserted into cellulose made thimble and placed inside the
rotavapor. The solvent used in this experiment is hexane which has
boiling point at 70 C. The rotavapor glassware with a roundbottomed flask is filled with boiling chips so that no splashing during
the evaporation. The evaporation and extraction took about an hour
and then the round-bottomed flask is removed and placed in the
heated water bath of the concentrator to remove all the solvent. The
fat content is calculated by this formula;
2.57.
The
combination
of
these
characteristics
and
the
2.62.
CONCLUSION
2.63.
2.64.
extract the total lipid content from the sample of mung bean.
Furthermore we are also succeeded to determine the fat content which
is 2.580 % of that sample. From the experiment we know how to use
the rotavapor, its function and principle so that we can determine fat
content from any sample by ourselves.
2.65.
REFERENCES
2.66.
2.67.
McKee, T and Mckee, J.M. (2003). Carbohydrates in Biochemistry The
molecular basis of life, Third Edition. McGraw-Hill. New York. pp. 201-231.
2.68.
2.69.
Sawhney, S.K. and Randhir, S. (2005). Introdcutory practical Biochemistry.
Alpha Science International Ltd.,Harrow. U.K. pp.15-32.
2.70.
Harwood,
Laurence
M.;
Moody,
Christopher
J.
(1989).