MINDANAO UNIVERSITY OF SCIENCE AND TECHNOLOGY

C.M. Recto Ave., Lapasan, Cagayan de Oro City

Laboratory Report
in
Isolation and General Tests of Glycogen
from Chicken Liver
(Experiment 4)

Submittted by:
Swetsel M. Capito
Jamaica G. Mariano
Reyna Y. Panuncio
Jelly Jean L. Paye
Shaira Jhann L. Rosales
John Mark C. Saburao
BS Chem- 3H1

July 27, 2015

1 INTRODUCTION
1.1

Theories and Principles

From Biochemistry (5th ed.) by Berg, Tymoczko & Stryer (2002), glycogen is a readily

mobilized storage form of glucose. It is a very large, branched polymer of glucose residues that
can be broken down to glucose molecules when energy is needed. Glycogen is not as reduced as
fatty acids are and consequently not as energy rich. However, it is an important fuel reserve for
several reasons. The controlled breakdown of glycogen and release of glucose increases the
amount of glucose that is available between meals. Hence, glycogen serves as a buffer to maintain
the blood-glucose levels, which is especially important since glucose is virtually the only fuel used
by the brain, except during prolonged starvation. Moreover, the glucose from glycogen is readily
mobilized and is therefore a good source of energy for sudden, strenuous activity. Unlike fatty
acids, the released glucose can provide energy in the absence of oxygen and can thus supply
energy for anaerobic activity.
The two major sites of glycogen storage are the liver and skeletal muscle. The
concentration of glycogen is higher in the liver than in muscle (10% versus 2% by weight), but
more glycogen is stored in skeletal muscle, because of its much greater mass. In the liver,
glycogen synthesis and degradation are regulated to maintain blood-glucose levels as required to
meet the needs of the organism as a whole. In contrast, in muscle, these processes are regulated
to meet the energy needs of the muscle itself.
Molisch's test is a general experiment for the presence of carbohydrates, in which
compounds are dehydrated to furfural in the presence of the concentrated sulfuric acid. The naphthol from the Molisch reagent, a solution of -naphthol in 95% ethanol, reacts with the cyclic
aldehydes to form purple-colored condensation products (Kofffuor, 2012).
Iodine test is useful when distinguishing starch and glycogen from other polysaccharides.
When iodine solution is added, iodine atoms fit into the helices to form complexes (Harisha,
2005). While monosaccharides and disaccharides are too small to trap iodine molecules, starch
form a more intensified color of blue-black from as glycogen produces a reddish color (Timberlake,
2007).

1.2

Objectives

To isolate and purify glycogen from a chicken liver sample by means of heating and
precipitation

To analyze and test the presence of glycogen by carrying out general tests such as
Molischs test and iodine test

2 METHODOLOGY
2.1

Materials, Chemicals and Apparatus

The material mainly used in this experiment was chicken liver. The apparatus used were

water bath, Petri dish, 50 mL beaker, mortar and pestle, distilled water, test tubes. The chemicals
used in this experiment were 95% ethanol, Molischs reagent, 12M H2SO4, 0.01M Iodine and 0.1%
CH3COOH.
2.2

Procedure
The chicken liver sample was cleaned and minced in a petri dish, in which 6 grams was

transferred to a beaker and constantly stirred in addition of 24-mL boiling distilled water. The
mixture was then grounded with mortar and pestle, and placed back to the beaker containing 6mL distilled water before putting in a boiling water bath for 30 minutes. Afterwards, 2-mL of 0.1%
acetic acid was added into the mixture before filtration. The collected filtrate was then divided
into 3 test tubes, and to be subjected in the following polysaccharide tests.
In Molischs test, 1-mL of glycogen solution was placed in a test tube, and added with 510 drops of 95% ethanol solution, 1-mL Molischs reagent and 2-mL 12 M sulfuric acid.
In iodine test, 1-mL of glycogen solution was placed in another test tube, and added with
a few drops of 0.01 M iodine. The mixture was then warmed in a boiling water bath and observed
for a change in color.
Observations and data gathered throughout the experiment were then recorded in the lab
notebook.

RESULTS AND DISCUSSION

3.1

Results

3.1.1

Extraction and Precipitation of Glycogen

After addition of hot water, the liver became light brown.

While boiling, a yellow liquid was produced.

The first filtration was done without addition of acetic acid. Hence, the mixture was
filtered repeatedly, producing a clear yellow solution.

3.1.2

General Tests for Polysaccharides

3.1.2.1

Molischs Test

Color of reagent: Light, cloudy violet

After addition of 95% ethanol: a slight warming effect alongside a pale yellow solution

After addition of Molischs reagent: a cloudy yellow surface

After addition of 12 M sulfuric acid: a cloudy white upper layer, a middle purple ring
and a cloudy yellow bottom layer

3.1.2.2

Iodine Test

Color of Reagent: Brown

After addition of iodine: light yellow solution

After warming in a water bath: a reddish solution (1 min); if stayed longer, reverses
into light yellow

3.2

Discussion
Glycogen, a polysaccharide stored mainly in liver and muscles, was extracted from the

chicken liver sample at a high temperature, since it contains glycosidic bonds which are resistant
to hydrolytic activity of OH even at elevated temperatures. The light yellow liquid produced is
soluble to water due of the branched structure of glycogen and free water molecules it contains
(Wynick, 2012).
On the other hand, peptide bonds in proteins, ester bonds in lipids and phosphodiester
bonds in ribonucleic acids in the liver undergo hydrolysis at high temperature in alkaline pH.
Under these conditions, the glycogen solution, now only slightly contaminated with other
polysaccharides, fragments of denatured DNA and low molecular weight compounds, can be
obtained through addition of ethanol which results to glycogen precipitation, acquiring a relatively
purified glycogen (Medical University of Lodz, 2012).

In the experiment, however, ethanol was not used in the precipitation of glycogen.
Instead, dilute acetic acid was used as required by the procedure. Either way, both acetic acid
and ethanol are weak acids, still resulting to the same glycogen precipitation.
Molischs test is one of the general tests used for polysaccharides. In this test,
carbohydrates form a purple colored ring at the junction. In the experiment, formation of purple
colored ring at the junction of the glycogen solution was observed after the addition of
concentrated H2SO4, which shows a positive result. This is because carbohydrates, when treated
with concentrated H2SO4, undergo dehydration to form furfural derivatives which condenses with
-naphthol in Molischs reagent that results to the formation of colored products.
Iodine is useful in distinguishing starch and glycogen from other polysaccharides.
Glycogen reacts with iodine to give a reddish brown color as starch gives blue-black. It is thought
that starch and glycogen form helical coils. Iodine atoms can then fit into the helices to form a
starch-iodine or glycogen-iodine complex. Starch in the form of amylose and amylopectin has less
branches than glycogen. This means that the helices of starch are longer than glycogen, therefore
binding more iodine atoms. The result is that the color produced by a starch-iodine complex is
more intense than that obtained with a glycogen-iodine complex (Chemistry Laboratory, 2014).
4

CONCLUSION
Glycogen is a carbohydrate found in chicken liver. It can be extracted by means of

precipitation with dilute CHCOOH and purified using 95% ethanol. The extracted glycogen from
the experiment showed positive results on the general test for polysaccharides. In Molischs test,
the glycogen formed a purple colored ring and in the iodine test, a reddish-brown coloration was
obtained, verifying that glycogen is a polysaccharide carbohydrate.
5

REFERENCES

All Medical Stuff. (2014, November 26). Molischs test - A Qualitative Test for the Presence of
Carbohydrates. Retrieved 2015, July 26 from http://allmedicalstuff.com/molischs-testqualitative-test-presence-carbohydrates/
Berg, J.M., Tymoczko, J.L. & Stryer, L. (2002). Chapter 21 - Glycogen Metabolism. Biochemistry
(5th ed.). W. H. Freeman, New York

Chemistry Laboratory. (2012). Iodine Test for Starch and Glycogen. Retrieved 2015 July 26 from
http://generalchemistrylab.blogspot.com/2011/12/iodine-test-for-starch-andglycogen.html
Harisha, S. (2005). Iodine Test for Starch and Glycogen. An Introduction to Practical
Biotechnology. Firewall Media
Kofffuor, G. (2012). Molisch's Test - Qualitative Test in Carbohydrates. Retrieved 2015, July 24
fromhttp://web.knust.edu.gh/oer/pages/index.php?siteid=knustoer&page=find_material
s&cou=41
Medical University of Lodz. (2012, October 12). Glycogen. Retrieved 2015, July 26 from
csk.umed.lodz.pl/~luska/6yearprogramme/lab3a.pdf
Timberlake, K.C. (2007). Test for Carbohydrates. Laboratory Manual for General, Organic, and
Biological Chemistry. Pearson Education, Inc.
Wynick,

D.

(2012).

Glycogen.

Retrieved

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http://www.askabiologist.org.uk/answers/viewtopic.php?id=8458

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from