EXERCISE 4

ORGANIC COMPONENTS: CARBOHYDRATES

Hannah Alexis M. Asilo

Abstract
Introduction
Most compounds in living things contain the element carbon, usually combined with
other elements. Most compounds that contain carbon are called organic compounds. These
organic compounds may form part of the organism or they may be cell products that do not
form part of the organism (Rabago et al., 2003). Most important groups of organic compounds
found in living things are proteins, lipids, nucleic acids and carbohydrates.
Carbohydrates are polyhydric alcohols possessing potentially active aldehyde or
ketone groups or derivative thereof. Carbohydrates are classified as simple or complex
depending on its chemical structure. Simple carbohydrates which contain just one sugar, such
as fructose, galactose and glucose are called monosaccharides while carbohydrates with two
sugars such as sucrose, lactose and maltose are called disaccharides. Carbohydrates with
three or more sugars are called polysaccharides.
Carbohydrates provide fuel for the central nervous system and energy for working
muscles. They also prevent protein from being used as an energy source and enable fatt
metabolism.
This exercise aims to become familiar with some of the methods commonly used in detecting
the presence of simple sugars or hydrolysed derivatives of polysaccharides based on their
color reaction.

Methods and Materials

Before the experiment, the fruit of Citrus maxima and the leaves of Ipomoea batatas were
homegenized separately using a blender. The extract were then used in the experiment
proper.
A. Characteristic color reactions of carbohydrates in some plant materials.
a. Molisch test
A total of eight clean test tubes were prepared and labelled accordingly. Each test
tubes was added with one ml of the following solutions using pipette: 1% glucose, 1%
fructose, 1% lactose, 1% sucrose, 1% starch, leaf extract, fruit extract and distilled
water as the control. In each test tube, two drops of -naphthol was added. After
shaking, each test tube was inclined as one ml of concentrated 𝐻2 𝑆𝑂4 was added by
allowing the acid to flow gradually on the side without mixing. Each test tube was
observed as a violet ring formed at the interphase.
b. Seliwanoff’s test
The same number of test tubes were used with three ml of the same solutions
(in Molisch test) were added in each test tube and labelled correspondingly. All test
tubes were then added with three ml of Seliwanoff solution and were heated
simultaneously in a boiling water noting the exact time when the test tubes were
placed in the water bath. Color development in each test tubes was observed. As

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 soon as the color developed, the test tubes were removed from the water bath. Time
required for the color to develop in the test tubes were recorded and was used to take
the difference between the time when the test tube was initially placed in the water
bath and the time the color developed in the test tubes in minutes.
c. Benedict’s test
The same number of test tubes were used with three ml of the same solutions
(in Molisch test) were added in each test tube and labelled correspondingly. Each
test tubes was added with three ml of Benedict’s reagent and mixed. The test tubes
were then heated in a boiling water bath. Each test tubes was observed and removed
as soon as a brick red precipitated formed.

Results and Discussion

I. Characteristic color reaction of carbohydrates
TESTS
Solution
Molisch’s Seliwanoff’s Benedict’s
(+) clear solution with (+) red precipitate; 21 (+) cloudy red solution
1% glucose
violet ring mins 34 s with precipitate
(+) clear solution with (+) red precipitate; 2 (+) cloudy red solution
1% fructose
violet ring mins and 34 s with precipitate
(+) cloudy brown
(+) clear solution with (+) red precipitate; 20
1% lactose solution with red
violet ring mins and 16 sec
precipitate
(+) 3-layered solution;
clear solution at the
(+) red precipitate; 3
1% sucrose top, violet ring and (-) clear blue solution
min 52 sec
cloudy white layer at
the bottom
(+) 3-layered solution;
clear solution at the
(+) red precipitate; 20 (+) clear blue solution
1% starch top, violet ring and
mins 41 sec with red precipitate
cloudy white layer at
the bottom
(+) cloudy green
(+) clear green (+) red precipitate; 7
Leaf extract solution with red
solution with violet ring mins 42 sec
precipitate
(+) 3-layered solution;
clear solution at the
(+) red precipitate; 4 (+) yellow solution with
Fruit extract top, violet ring and
min 51 sec red precipitate
cloudy white layer at
the bottom
Distilled water (-) clear solution (-) yellow solution (-) clear blue solution

Molisch’s test is a sensitive chemical test for the presence of carbohydrates, either
mono, di-, or polysaccharides. Thus, all the solution, except for distilled water gave a positive
result which was the formation of a violet ring. Leaf extract gave a positive result because of
the present of carbohydrates as part of the cell membrane or as a sugar, specifically glucose,
which is a product of photosynthesis and cellulose in its cell wall. Fruit extract also gave a
positive result because fruits basically have sugars which make them sweet and also for the
presence of cellulose.
Carbohydrates when treated with concentrated𝐻2 𝑆𝑂4 , undergoes dehydration to give
furfural or furfural derivative (5-hydroxymethylfurfural).

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These compounds condense with -naphthol to give reddish violet ring at the junction of two
solutions, which indicates the presence of carbohydrates.

In case of oligo and polysaccharides, in this case, starch, they are first hydrolyzed to
monosaccharides by acid, which are then dehydrated to form furfural or its derivatives.
Pentoses yield furfural and hexoses yield furfural derivative.
Seliwanoff’s test is used to differentiate ketose sugars from aldose sugars. If the sugar
contains a ketone group, it is a ketose. If a sugar contains an aldehyde group, it is an aldose.
This test relies on the principle that, when heated, ketoses are more rapidly dehydrated than
aldoses. When ketone sugar like fructose is heated with dilute HCl, it undergoes dehydration
to form 5-hydroxymethylfurfural, which on condensation with resorcinol gives cherry red color
that indicates the presence of ketone sugar.

Fructose and sucrose are two common sugars which give a positive test and was also
observed in the experiment. Sucrose gives a positive test as it is a disaccharide consisting of
fructose and glucose. Theoretically, glucose, lactose and starch should not give a positive
result because glucose is an aldose sugar, lactose is made of two molecules of glucose and
starch is made of amylopectin and amylose which are made of chains of glucose molecules.
The positive reaction of these solutions is due to the prolonged boiling of the solution. This
made aldose sugar (glucose or lactose) give the test positive due to their conversion to ketone
sugar by the HCl. And as observed in the recorded time the color changed, it took more than
20 minutes for the solutions to become red. In comparison to the time it took for fructose,
sucrose, leaf and fruit extract to become red, which is less than 10 minutes, the time the aldose
sugars took is 2x longer.
Benedict’s test is a test for the identification of reducing sugars. This test uses
Benedict’s reagent which is a complex mixture of sodium carbonate, sodium citrate, and coper
(II) sulphate pentahydrate. A positive test with Benedict’s reagent is shown by a color change
from clear blue to a brick-red precipitate.

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 The principle of Benedit’s test is that when reducing sugars are heated in the presence
of an alkali, they are converted to powerful reducing species known as enediols. Enediols
reduce the cupric compounds (𝐶𝑢2+ )present in the Benedict’s reagent to cuprous compounds
𝐶𝑢+ )which are precipitated as insoluble red copper (I) oxide (𝐶𝑢2 𝑂).

All monosaccharides are reducing sugars because of their free aldehyde o ketone
group. As observed from the experiment, all monosaccharides gave a positive result. Some
disaccharides are reducing sugars like maltose and lactose, which gave a positive result in
the experiment. But sucrose, which is a disaccharide did not give a positive result, thus is not
a reducing sugar. This is because its glycosidic bond is between its anomeric carbons and
thus cannot convert to an open-chain form with an aldehyde group and are stuck in the cyclic
form. Theoretically, starch is not a reducing sugar whether it’s in the form of amylose or
amylopectin. The first ring cannot open up because there’s no hydrogen on the circled oxygen
to allow for ring opening as well as the next ring, et cetera, cannot open up. so polysaccharide
such as starch cannot open up.

References:
Szalay, J. (2017) What are carbohydrates. Live Scince. Retrived from