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ABSTRACT
Carbohydrates are the most abundant class of organic compounds found in living organisms. It is defined as any of a
group of organic compounds that includes sugars, starches, celluloses, and gums and serves as a major energy source
in the diet. The objective of this experiment is to isolate the polysaccharide glycogen from chicken liver and explain
the principle involved in it and in the general tests done to determine the polysaccharide content of the sample.
Initially, the glycogen from chicken liver is isolated by heating and adding 0.1% acetic acid and then adding 5-10
drops of ethanol. The successful isolation of starch was proven from the Molisch's Test and I 2 Reaction. The sample is
also hydrolyzed via acidic hydrolysis. Quantitative analysis of carbohydrates was also performed in this experiment by
mixing 12.5 mL of Nelsons A with 0.5 mL Nelsons B and transferring the measured amounts of glucose solution to
the test tubes. The concentration of the unknown in mg/tube and mg/mL was determined after constructing a glucose
standard curve by plotting absorbance readings against concentrations of standard solutions. In this determination,
the amount of free reducing sugars in the sample is directly related to the molybdenum blue formed via a series of
oxidation/reductions reactions, and is measured colorimetrically.
INTRODUCTION
Carbohydrates, also known as saccharides,
are carbon compounds that contain large
quantities of hydroxyl groups, and are also the
most important sources of energy. They have the
basic general formula Cn(H2O)n and they are the
most commonly found organic compounds in
living organisms. They are classified into several
groups, namely monosaccharides, disaccharides,
and polysaccharides, depending on the number of
their monosaccharide.
EXPERIMENTAL
A. Compounds tested ( or Samples used )
A. Molisch's Test
Few drops of Molisch's reagent were added into 1
mL glycogen solution. 2 mL concentrated H 2SO4
was poured down the side of the tube to form a
layer. The color at the junction of the two liquids
was observed.
B. I2 Reaction
Few drops of 0.01 M I2 was added into 1 ml
sample solution. The red color indicates the
presence of glycogen. The mixture was warmed
in a water bath and observed if there was any
change in color. The result was then noted.
3. Acid Hydrolysis of Polysaccharides
5 drops of conc. HCl was added to 5 mL of the
glycogen solution contained in a test tube. The
tube was covered with a cotton boil and was
boiled in a water bath for 30 minutes. The
viscosity of the solution before and after heating
was noted. The solution was then neutralized
using dilute NaOH and the hydrolysate was then
kept in a refrigerator until it was subjected for
Benedicts Test
4. Quantitative Analysis
The Nelson's reagent was prepared by mixing
12.5 mL Nelson's A with 0.5 Nelson's B. 8 test
tubes were labeled and measured amounts of
standard glucose solution were transferred into
the test tubes according to the following protocol.
Tube No.
Glucose
Distille
Unknow
Standar
d
n Sample
d
Water
(mL)
(mL)
(mL)
1
0
1.0
0
2
0.1
0.9
0
3
0.2
0.8
0
4
0.4
0.6
0
5
0.6
0.4
0
6
0.8
0.2
0
7
1.0
0
0
8
0
0.6
0.4
Fig. 2 (Glucose standard, Distilled water,
Unkown sample amounts)
1.0 mL Nelson's reagent was added to each test
tube and shaken well. The tubes were heated
simultaneously in a boiling water bath for 20
minutes. Then, the tubes were removed and
cooled in a beaker of water. 1.0 mL
arsenomolybdate reagent was added to each
tube and shaken occasionally for 5 minutes or
until the Cu2O precipitate dissolves. After that,
the absorbance of the standards and unknown
were read against a reagent blank at 480 nm. A
glucose standard curve was then constructed by
plotting
absorbance
readings
against
concentrations of standard solutions. The
concentration of the unknown was determined in
mg/tube and mg/mL.
REFERENCES
From books
University of Santo Tomas Faculty of Pharmacy.
(2014). Laboratory Manual in Organic Chemistry
Revised Edition. Manila: Author
From the internet (on-line)
[1]Retrieved : May 10 2016
http://oregonstate.edu/instruct/bb450/450ma
terial/stryer7/21/figure_21_02.jpg
[2]Retrieved : May 10 2016
https://d2gne97vdumgn3.cloudfront.net/api/file/
0QFD9Ud8RfKt2fdMGkJd