Experiment 7

Title: Tests for carbohydrates

Aims:
1. To observe physical and chemical properties of some common carbohydrates
2. To use physical and chemical tests to distinguish between monosaccharide,
disaccharides, and polysaccharides.
3. To relate the process of digestion to the hydrolysis of carbohydrates.
4. To follow the reaction of glucose in base by use of Seliwanoff and Benedict
reagent.

Procedures:

A. Benedict’s Test For reducing Sugars

1. 10 drops of deionised water and 2% solution of glucose, fructose, sucrose,

lactose and starch are placed in separate test tubes. Benedict’s reagent to
each tube to a depth of 1 cm.

2. The test tubes are placed in boiling water bath for 3-4 minutes.The
observations are recorded.

B. Seliwanoff’s Tests for Ketoses

1. 10 drops of deionised water are placed and 2% if glucose, fructose, sucrose,

lactose, and starch are placed in separate test tubes. Seliwanoff’s reagent is
added to each tube to a depth of 1 cm.

2. The test tubes are placed in a boiling water bath and the time is noted.
After 1 minute, the colours in the test tubes are observed. The results as
fast colour change, slow colour change, or no change are recorded.

C. Iodine Tests For Polysaccharides

1. 10 drops of deionised water are placed and 2% solutions of glucose,

fructose, sucrose, lactose, and starch in separate test tubes are placed. 1
drop of iodine solution is added to each sample. The results are recorded.
 D. Hydrolysis of Disaccharides And Polysaccharides

1. 2% starch is placed in two test tubes to a depth of 2 cm, and 2% sucrose

solution is placed in two more test tubes to a depth of 2cm. 20 drops of 10%
HCl are added to one sample each sucrose and starch. 20 drops of H2O are
added to the other samples of sucrose and starch. The four test tubes are
heated in a boiling water bath for 10 minutes.
2. The test tubes from the water bath are removed and be cooled. 10% NaOH
(about 20 drops) are added until one drop of mixture turns litmus paper
blue. The samples for hydrolysis are tested for hydrolysis.

a) Iodine Test: 5 drops if each solution on a spot plate or watch

glass are placed. 1 drop of iodine reagent is added to each. The
observations are recorded. It is determined if hydrolysis has
occurred in each.

b) Benedict’s Test: 2mL of Benedict’s reagent is added to each of

the samples and heated in a boiling water bath for 3-4 minutes.
The observations are recorded. It is determined if the
hydrolysis has occurred in each.

Results:

Results Of carbohydrate Tests

Carbohydrate Benedict’s Test Seliwanoff’s Test Iodine Test

Glucose
Fructose
Sucrose
Lactose
Starch
Water (Control)
 Hydrolysis Of Disaccharides and Polysaccharides

Results Sucrose+H2O Sucrose+HCl Starch+H2O Starch+HCl

Iodine Test
Benedict’s Test
Hydrolysis
Products Present

Questions:
1. a) i) reducing sugars :
ii) non-reducing sugar :

b) Ketose Sugar :
c) Carbohydrates give a blue-black colour in the iodine test:

2. a) Carbohydrate(s) produces a reddish-orange solid with Benedict’s and red

colour with Seliwanoff’s reagent in 1 minute:

b) Carbohydrate(s) give a colour change with Benedicts’s test, a light orange

colour with Selliwanoff’s reagent after 5 minutes, and produces no bubbles
during fermentation:

c)Carbohydrate(s) gives no colour change with Benedict’s or Seliwanoff’s

test, but turns a blue-black colour with iodine reagent :

3. Positive(+) or negative(-) results:

Carbohydrate Benedict’s Test Seliwanoff’s Test Iodine Test

Glucose
Fructose
Galactose
Sucrose
 Lactose
Maltose
Amylose
Amylopectin

Conclusion:

1. The reducing sugar can turns the solution’s colour to brick red when reacts
with Benedict’s reagent. Glucose, fructose, lactose are reducing sugar, while
sucrose and starch are non-reducing sugar. The intensity of the red colour
higher after Benedict’s reaction, the concentration of reducing sugar is
higher.

2. The present of ketose can forms a deep red colour with the reaction of
Seliwanoff’s Test. Fructose and sucrose contain ketone group. The more
deep red colour of solution, the more concentration of ketose.

3. Amylose and amylopectin or polysccharides(starch, glucose, cellusoe) can be

identified by Iodine test by observing the colour change to blue-black which
is for amylase, while red or brown is for other polysaccharides. Glucose,
fructose, sucrose, and lactose indicate the presence of other
polysaccharides,while starch indicates present amylase.

4. Some carbohydrates can be hydrolysed with water and acid. Sucrose gives
glucose and fructose as the monosaccharides while starch gives glucose as
the monosaccharides after hydrolysis.

Discussion:

A. Benedict’s Test For Reducing Sugars

This test is a test to distinguish reducing from non-reducing sugars. Benedict’s
reagent contains Cu2+ ion that can be reduced to Cu+ as oxidation of sugar in the
solution occur, which the colour of the solution change from blue to red. In the
open-chain form, the carbon atom that contains the C=O bond is called the carbonyl
carbon. In the closed-ring (cyclic) structure, the carbonyl carbon is the one which is
attached to the O of the ring and an OH group. Only the carbons revert to the open
chain, then only oxidation can take place. When the cyclic structure opens, the
aldehyde group is available for oxifation. Sugars that can be oxidised by oxidising
agents are called reducing sugars because the oxidising agent is reduced in the
reaction. From the experiment, the red colour intensity of solution after reaction
of each varies from green to gold to red depanding on the concentration of
reducing sugar. Only sucrose and starch show obvious negative result of this test.
Sucrose it not reducing sugar since it cant revert to the open-chain form that
would provide the aldehyde group needed for reducing the Cu2+ ion. Precaution
should be taken in this experiment is rinse the hands or cloth as the bare hands or
lab cloth have directly contacted the Benedict reagent, since it may be caustic. And
also, handle the hot apparatus and test tube carefully

B. Seliwanoff’s Test For Ketoses

Seliwanoff’s test is used to test the presence of the ketose or differentiate

between hexoses with a ketone group and hexoses that are aldehydes. With
ketoses, a deep red colour is formed rapidly, while aldoses should give a light pink
colour, but takes a longer time to process. Fructose should be the obvious positive
result. From the experiment,.fructose showed the highest red colour intensity of
the solution after reaction take place, other than that, sucrose also show slightly
positive result instead of glucose, lactose, starch and water. The fructose contains
ketone group and sucrose also contains fructose that the reagent reacts most
sensitively of effectly with ketose or ketone group. Precaution can be taken is
handle the hot apparatus and test tube carefully.
 C. Iodine Test For Polysaccharides

Iodine test is used to identify glycogen and starch (amylase and amylopectin) which
are the polysaccharides. A dark blue colour is a positive test for amylose in starch.
A red or brown colour indicates the presence of other polysaccharides.
Polysaccharides will react with iodine to form a blue, red, violet, or purple product
which is a positive indicator of the presence of a polysaccharide. From the
experiment, the starch gave dark-blue colour change, while the rest of the samples
gave red colour change. When iodine is added to amylose, the helical shape of the
unbranched polysaccharide traps iodine molecules, producing a deep blue-black
complex. Amylopectin, cellulose, and glycogen react with iodine to give red to brown
colours. Glycogen produces a reddish-purple colour. Monosaccharided and
disaccharides are too small to trap iodine molecules and do not form dark colours
with iodine.

D. Hydrolysis Of Disaccharides and Polysaccharides

Hydrolysis of disaccharides is to get monoccharides. Disaccharides and

polysaccharides undergo acid hydrolysis with water to form monosaccharides
that compose them. Let say, like hydrolysis of maltose, it converts maltose
to glucose molecules. From the experiment, starch and sucrose do not give
positive test with Benedict’s. After hydrolysis, sucrose produces fructose
and glucose that give a positive Benedict’s test. Starch after hydrolysis
forms many glucose molecules that also give a positive test with the
Benedict’s reagent. In living systems, molecules known as enzymes facilitate
this breakdown. Hydrolysis of starch is necessary for the organism to utilize
the glucose monomers. Like in the body, enzymes in our saliva and from the
pancreas carry out the hydrolysis. Complete hydrolysis produces glucose,
which provides about 50% of our nutritional calories and also aids to fuel up
our daily activities. Check carefully with the samples added and do not mix
the solution with other substances.

The simplest forms of carbohydrate molecules are monosaccharides or single

sugars. One of the most important monosaccharides is glucose (C 6H12O6), the end
product of photosynthesis in plants and the molecule used by cells to produce ATP.
ATP is the molecule that supplies the energy to do cellular work. Other important
monosaccharides are fructose, galactose and ribose.

A disaccharide, or double sugar is two monosaccharides that have been joined

through dehydration synthesis. Some common disaccharides are lactose (from
milk), sucrose (table sugar) and maltose.

Polysaccharides are many monosaccharide molecules joined together through

dehydration synthesis. Amylose, also known as starch, is a polysaccharide made of
only glucose molecules. It is abundant in plants and is the plants’ way of storing the
glucose it makes during photosynthesis. Animals store glucose in glycogen, another
type of polysaccharide. Although starch and glycogen are both composed of
glucose subunits, the glucose monomers are bonded together in different ways so
these polysaccharides are not identical. Glucose subunits are bonded together a
third way in the polysaccharide cellulose. Cellulose, the most abundant
carbohydrate in the world, is not an energy storage molecule but is a structural
molecule. Cellulose makes up the cell walls of plants and is the primary component of
dietary fiber. For most animals it is completely indigestible.