Reactions of Carbohydrates

Paulo T. Carpio
HUB32

De La Salle University-Dasmarias
Dasmarias, Cavite, Philippines

ABSTRACT

The reactions of given samples of carbohydrates were observed from different test that
gives different color of product to identify their specific type and distinguish from their
reactions. There are nine carbohydrate samples that were tested with two unknown who
were observed to determine their identity from comparing their results to the other given
samples. Six different tests were used and a 0.20ml of each carbohydrate sample was
placed in different test tube and added with 0.20ml of the test reagent. The identity of the
unknown sample 1 was determined to be Fructose while unknown sample 2 was
identified as Lactose.

INTRODUCTION

Carbohydrates are the main energy source of the human body and they are the single most
abundant class of organic molecule that can be found in nature. It is the structural building blocks
of the cells and components of numerous metabolic pathways. A broad range of cellular
phenomena such as cell recognition and binding depend on carbohydrates. (2)
Natural carbohydrates are subdivided into monosaccharides, or simple sugars containing
three to nine carbon atoms, polysaccharides, or polymers of monosaccharides, and an
intermediate category of oligosaccharides, with two to ten monosaccharide units joined. The most
important oligosaccharides to humans economically and biologically are the disaccharides.(3)

MATERIALS AND METHODS

A 0.20ml of known carbohydrate samples and two unknown samples were place in separate
test tubes. They were subjected to different test to observe their reactions. The different test
performed were shown on Table 1.1

Table 1.1 Qualitative tests for Carbohydrates

Test Amount of reagent Procedure

Molisch test 0.20ml Molisch reagent,

0.20ml conc. H2SO4
Iodine test 0.20ml iodine reagent
Benedicts test 0.20ml benedict reagent
Barfoeds test 0.20ml barfoed reagent
Seliwanoffs test 0.20ml seliwanoff reagent
Bials test 0.20ml bial reagent
Water bath at 100C
Water bath at 100C
Water bath at 100C
Water bath at 100C
 Visible color changes were observed on each reaction. The unknown samples were then
identified by comparing from the known carbohydrate samples.

RESULTS AND DISCUSSIONS

Carbohydrates can be identified or differentiated through different tests. One of these was
the use of condensation reagents which react with carbohydrates to produce highly colored
products. The tests that were used in the experiment with condensation reagents were Molischs,
Bials and Seliwanoffs test. In Iodine test, the carbohydrates must possess certain structural
features that allow it to form a condensation product. The condensation reaction tests were useful
to classify the type of carbohydrates and to identify the specific type of carbohydrates. The other
two tests used were for reducing sugars or saccharides were the Benedicts and Barfoeds test.
The determination of positive reactions from the different tests was shown in Table 2 with their
corresponding positive color change. Reactions of the carbohydrate samples to each test were
shown in Table 3 and the identity of two unknown carbohydrate sample were determined as
Fructose and Lactose.

Table 2. Positive Reactions for Carbohydrates Test

Test Positive Color Change

Molisch Deep purple color
Iodine Blue-black complex
Benedict Rust-colored precipitate
Barfoed Red precipitate
Seliwanoff Aldoses (blue green); Ketoses (red)
Bial Pentoses (blue/green); Hexoses (muddy
brown to gray

Table 3. Reaction of Carbohydrate Samples

Molisch Iodine Benedict Barfoed Seliwanoff Bial

Glucose ++ - ++ + - -
Galactose ++ - ++ + - -
Fructose ++ - ++ + ++ -
Lactose + - ++ - - -
Sucrose + - - - + -
Starch + ++ - - - -
Ribose ++ - ++ + - ++
Unknown 1 + - ++ + + -
Unknown 2 + - + - - -
Legend: ++ fast reaction; + slow reaction; - no reaction

Identity of Unknown 1: Fructose

Identity of Unknown 2: Lactose

A positive for Molischs test has a presence of carbonyl group which results to a deep purple
color product. It uses concentrated sulfuric acid as dehydrating acid to dehydrate carbohydrate.
All of the carbohydrate samples got a positive test because they obviously have carbonyl groups.
Monosaccharide gives a rapid positive test while disaccharide and starch were slower. The Iodine
test is used to distinguish starch so the starch sample got a positive test while the remaining
samples got a negative results or no reaction. An illustration of pertinent reaction for Molischs
test was shown in Figure 1.

Figure 1. Reaction of Carbohydrates in Molischs test

Benedicts test which is a mild basic solution reduces carbohydrates and it uses a mixture of
copper(II) sulfate, sodium nitrate and sodium carbonate. The presence of red copper(I) oxide
precipitate indicates that the saccharide has reduce the copper(II) ions. Only sucrose and starch
got a negative test. The Barfoeds test is a copper(II) ions in acidic medium that is use for
reducing monosaccharide and disaccharide. Samples with a negative test were lactose and
sucrose because they were disaccharides, starch because it was a polysaccharide and the
unknown sample 2. Illustrations for the reaction of carbohydrate in Benedicts and Barfoeds test
were shown in Figure 2 and 3.

Figure 2. Reaction of Carbohydrates in Benedicts test

Figure 3. Reaction of Carbohydrates in Barfoeds test

The Seliwanoffs test used hydrochloric acid as dehydrating acid and this test is for
distinguishing aldoses from ketoses. The formation of red product was the basis for a positive test
for the ketoses. Fructose got a positive result because it was basically a ketose, while the
sucrose was formed from glucose and fructose so it gives also a positive reaction and the rest of
the samples got negative result because they were all aldoses while the starch was a
polysaccharide. Bials test was the other test that identifies a specific type of carbohydrates
because it can distinguish pentoses from hexoses. In the experiment, positive result was for
pentoses which will give a blue or green product. Only the ribose got a positive result from all the
other sample because it was the only pentose while the other monosaccharide like glucose,
galactose were hexose and the two unknown sample got a negative results that signifies they
were not pentoses. Illustrations for the reaction of carbohydrates in Seliwanoff and Bials test
were shown in Figure 4 and 5. From the results of the experiment, the two unknown carbohydrate
sample were identified to be Fructose for unknown sample 1 and Lactose was the unknown
sample 2 because they got same results from the given known sample of fructose and lactose.

Figure 4. Reaction of Carbohydrates (aldoses & ketoses) in Seliwanoffs test

Figure 5. Reaction of Carbohydrates (pentoses & ketoses) in Bials test

 A faster way to identify the unknown carbohydrate is using a general test to a more specific
type of test. A good order of test that can be used to identify an unknown sample was shown in
Figure 6.

Molisch's Test

no reaction (Purple product)

not carbohydrate Iodine test

(no reaction) (Blue-black)

Barfoed's test Starch

reducing reducing
monosaccharide disaccharide
(Seliwanoff's test) (Benedict's test)

blue green-Aldose
red-Ketose
Bial's test

(blue/green) (brown-gray)
Pentose Hexose

Figure 6. Step of Tests for Identifying an Unknown Sample

REFERENCES

(1) Legaspi, G.A. 2009. Essentials of Biochemistry Laboratory

(2) McKee, T. McKee, J.R. 2003. Biochemistry-The Molecular Basis of Life. 3rd Edition.
McGraw-Hill Companies, Inc
(3) Langley R. Moore J.T. 2008. Biochemistry for Dummies. Wiley Publishing, Inc.
Indianapolis, Indiana
(4) Seeley, R.R. Stephens, T.D. Tate, P. 2005. Essentials of Anatomy & Physiology. 5 th
Edition. The McGraw-Hill Companies, Inc. New York