Biochemistry Laboratory Manual Experiment 2

Experiment 2
POLYSACCHARIDES
Polysaccharides are the complex carbohydrates. They are made up of chains of monosaccharides (the sugars)
which are linked together by glycosidic bonds, which are formed by the condensation reaction. The linkage of
monosaccharides into chains creates chains of greatly varying length, ranging from chains of just two
monosaccharides, which makes a disaccharide to the polysaccharides, which consists of many thousands of the
sugars.
As a group, the polysaccharides play diverse and important roles within the biology of life processes; however, they
can be divided into two broad groups.
Structural polysaccharides
Cellulose
Cellulose is a major component of plant cell walls. It is an unbranched polymer with about ten thousand glucose
units per chain. Hydroxyl groups (-OH) project out from each chain, forming hydrogen bonds with neighboring
chains which creates a rigid cross-linking between the chains, making cellulose the strong support material that it is.
Despite the combined strength of cellulose, it is fully permeable to water and solutes which makes it ideal for
allowing water and solutes into and out of the cell.
It is the most abundant organic substance in the living world and it has been estimated that more than half the total
organic carbon on the planet is in cellulose.
Chitin
Chitin is closely related in structure to cellulose, also being an unbranched polysaccharide. However, instead of the
hydroxyl groups (-OH), the chains have the following structure NH.CO.CH3 replacing it. Large amounts of chitin
are found in the cuticles of arthropods, with smaller amounts being found in sponges, mollusks and annelids. Chitin
can also be found in the cell walls of most fungi and in some green algae.
Polysaccharides used as energy stores
Polysaccharides make ideal storage molecules for energy for a number of reasons; a) they are large, this makes them
insoluble in water and therefore they exert no osmotic or chemical effect on the cell; b) they fold into compact
shapes; c) they are easily converted into the required sugars when needed.
Glycogen
Glycogen is a branched polysaccharide found in nearly all animal cells and in certain protozoa and algae. In humans
and other vertebrates it is principally stored in the liver and muscles and is the main form of stored carbohydrate in
the body, acting as a reservoir of glucose (which the glycogen can be broken down into) for when the body is being
starved of food. In an animal that has been fed well, as much as 10% of its liver weight may be glycogen. The livers
glycogen store can be almost completely depleted during a 24 hour fast or during muscular exertion but glycogen is
quickly re-synthesised from newly ingested carbohydrate.
DMSF-College of Dentistry
RMT

Prepared by: John Paul M. Banzon, RN and Zenaida M. Banzon,

Page 14

Biochemistry Laboratory Manual Experiment 2

Starch
Starch is similar to glycogen, however it is found in plant cells, protists and certain bacteria. The starch granules can
be seen in the cell through a light microscope. The starch granules are made up of two polysaccharides, amylose and
amylopectin. Amylose is an unbranched molecule made up of several thousand glucose units, coiled helically into a
more compact shape. Amylopectin is also compact but has a branched structure and is made up of twice as many
glucose units as amylose.
Combining polysaccharides with proteins and lipids
Polysaccharides can be combined with proteins or lipids to form other biological molecules. For example,
peptidoglycans, which are a combination of protein and polysaccharide and are found in the cell wall of certain
bacteria. Glycolipids, a combination of polysaccharides and lipids are found in the cell membrane.
Sources
Biological Science 1&2 by Green, Stout and Taylor, 2nd ed. Cambridge
Cell and Molecular Biology by Sheeler and Bianchi, 3rd ed. Wiley
A. Starch
Starch occurs in the form of granule. Under the microscope the particles appear as concentric layers
of different shapes and sizes, according to the species of the plant from which it is derived.

1.

Iodine Test on Starch Powder.

Place a small pinch of starch powder on a watch glass placed over a piece of white paper. Drop lugols
solution on it (Lugols solution is iodine potassium iodide solution). State your result.
Q1. What is the cause of the production of the color?
2.

Iodine Test on Starch Paste

Place 1 ml of starch paste (1%) in a test tube. Add a drop of iodine solution. Heat and note the color. Cool and
observe any change produced. Explain the principle involved in the different changes noted. State your results
B. Hydrolysis of Starch:
Procedure:
Place 50 ml of starch solution in a small beaker. Add 5 drops of concentrated hydrochloric acid. Boil gently.
At intervals of 4 minutes, test a drop with Iodine and a small portion with Benedicts test, until iodine test becomes
negative and the Benedicts test positive. Tabulate your results:
Q2. What changes did you notice in regards to the reaction with Iodine and Benedicts, as
hydrolysis progresses?
Q3. After how many minutes is the starch solution completely hydrolyzed?
Q4. What is the end product in the hydrolysis of starch by an acid?
Q5. Write the different stages in the hydrolysis of the starch witch acids
Q6. How would you differentiate this from theoretical knowledge of starch hydrolysis by
enzymes?

DMSF-College of Dentistry
RMT

Prepared by: John Paul M. Banzon, RN and Zenaida M. Banzon,

Page 15

Biochemistry Laboratory Manual Experiment 2

C. Phenylhydrazine Reaction:
Procedure:
Perform the phenlhydrazine test on the hydrolyzed starch. Examine the crystals under the
microscope and draw them. Compare the crystals with those obtained from glucose.
With this test how could you determine whether your starch has been completely hydrolyzed or
not?
D. GLYCOGEN:
Glycogen is the polysaccharide of the animal body. It is often called animal starch. It is non-reducing and
gives a red color with iodine.
Preparation of glycogen:
Procedure:
Cut two oysters/liver into small pieces. Mix with sand and grind in a mortar. Place in a beaker and add
25ml. of water. Heat for 25 minutes, adding water from time to time to maintain the volume. Acidify with
5% acetic acid.
Boil for 1 minute, cool and filter. Add 4 volumes 95% alcohol. Allow the precipitate to settle,
pour off the supernatant and the filter. Dissolve the precipitate in hot water to make a glycogen solution.
Test for Glycogen:
Test a small portion of the glycogen solution with Iodine and Benedicts reagent.
Results:
1. Iodine Test

2. Benedicts test:

Hydrolysis of Glycogen:
Procedure:
Acidify the remaining solution with HC1 and boil for ten minutes. Cool and neutralizing with 10% NaOH.
Test a small portion with Phenylhydrazine reagent. Compare your results of the Benedicts test with that
obtained previous to hydrolysis. Examine the osazone crystals under the microscope and draw.
Results:
1. Benedicts Test

2. Phenylhydrazine Test:

Summary and Conclusions

1.

What conclusions can you draw in the following experiments? What practical applications can you
think of?

DMSF-College of Dentistry
RMT

Prepared by: John Paul M. Banzon, RN and Zenaida M. Banzon,

Page 16

Biochemistry Laboratory Manual Experiment 2

DMSF-College of Dentistry
RMT

Prepared by: John Paul M. Banzon, RN and Zenaida M. Banzon,

Page 17