Test for Carbohydrates

Molisch test

 Molisch’s Test: Objectives, Principle, Reagents, Procedure and Result

 Objective:
 to identify carbohydrate from other biomolecules
 Principle of Molisch’s test:
 Molisch’s test is a general test for all carbohydrates. In this test,
carbohydrates when reacted with conc. H2SO4 get dehydrated to form
furfural and its derivatives. When monosaccharide are treated with conc
H2SO4 or conc HCl, -OH group of sugar are removed in the form of water
and furfural is formed from pentose sugar and hydroxymethyl furfural is
formed from hexose sugar. These products reacts with sulphonated α-
naphthol to give a purple (violet red) colored complex.
 Result Interpretation of Molisch’s test:
 Positive Molisch’s test: purple color complex
 Negative Molisch’s test: no purple color
Iodine Test for Sugar

 Principle of Iodine test for carbohydrate:

 Starch when reacted with I2 forms absorbed compound that gives blue
color. On heating or on addition of alkali like NaOH or KOH, color
disappears. This reaction is only physically association where I2 traps in the
coiled structure of polysaccharide. On heating or on addition of alkali; the
coiled structure becomes linear and the I2 molecules are released and the
color disappears. The test will be answered by fructose, sucrose and other
keto containing carbohydrates.
 Result interpretation of Iodine test:
 Positive iodine test: dark blue color (starch)
 Negative iodine test: glucose, fructose and sucrose
Fehling’s Test

 Objective:
 to detect reducing sugar in a given solution
 Principle of Fehling’s test:
 Fehling’s test is one of the sensitive test for detection of reducing sugars.
Fehling’s reagents comprises of two solution Fehling’s solution A and solution B.
Fehling’s solution A is aqueous copper sulphate and Fehling’s solution B is
alkaline sodium potassium tartarate ( Rochelle salt). Rochelle salts (sodium
potassium tartarate) present in the reagent acts as the chelating agent in this
reaction.These two solution are mixed in equal amount before test.
 On heating an aldehyde or reducing sugar with Fehling’s solution give reddish
brown prepitate. Formation of red precipitate of cuprous oxide denotes the
presence of reducing sugar
 Result interpretation:
 Positive Fehling’s test: reddish brown ppt ( glucose, fructose, lactose)
 Negative Fehling’s test: No red ppt (sucrose, starch)
 Benedict’s test
 Objective:
 to detect reducing sugar ( carbohydrate having free aldehyde or ketone
functional group)
 Principle:
 The Reducing sugar under alkaline condition form enediols. Benedict’s
solution contains milder alkali Na2CO3. Enediols are powerful reducing
agents. They can reduce cupric ions to cuprous ions which is the basis for
Benedict’s reaction. The cuprous hydroxide during the process of heating is
converted to red cuprous oxide.
 positive benedict’s test: color change from blue to brick red ppt ( glucose)
 Negative Benedict’s test: no change in color( sucrose)

Color observed Sugar % Result interpretation

Blue Nil Absent of suagr
Green color 0.5% +
Green ppt 0.5-1% ++
Yellow ppt 1-1.5% +++
Orange ppt 1.5-2% ++++
Brick red ppt >2% +++++
Barfoed’s Test

 Objective:
 to detect reducing sugar
 to distinguish monosaccharides from reducing disaccharides
 Principle of Barfoed’s test:
 Barfoed’s test is used for distinguishing monosaccharides from reducing
disaccharides. Monosaccharides usually react in about 1-2 minute while
the reducing disaccharides take much longer time between 7-12 minutes
to react with the reagent. Brick red color is obtained in this test which is due
to formation of cuprous oxide.
 Result interpretation of Barfoed’s test:
Seliwanof’s Test

 Principle of Seliwanoff’s test:

 Seliwanoff’s test is used to distinguish aldoses from ketoses. On treatment
with conc. Acid, ketoses are dehydrated more rapidly to give furfural
derivatives and on condensation with resorcinol give cherry red complex.
The test will be answered by fructose, sucrose and other keto containing
carbohydrates.
 Result Interpretation of Seliwanoff’s test:
 Positive seliwanoff’s test: Fructose and sucrose
 Negative seliwanoff’s test: glucose, distilled water
 *If the reaction is allowed for longer time, aldoses also produce positive
results
 Bial’s Test
 Principle of Bial’s test:
 Bial’s test is useful in distinguishing pentoses sugar from hexoses sugars.
Pentosses ( such as ribose sugar) form furfural in acidic medium which
condense with orcinol in presence of ferric ion to give blue green colored
complex which is soluble in butyl alcohol.
MUCIC ACID TEST

 The mucic acid test is used to identify the presence of the sugar galactose
in food or in synthetics manufacture. When concentrated nitric acid is
heated together with galactose, a dicarboxylic acid called mucic acid
forms as a white precipitate, which counts as a positive result.
 The mucic acid test gives a positive result with several galactose-containing
substances such as lactose and dulcite. With these other sugars, there is no
formation of a white precipitate when heated with nitric acid, but an acid
that is soluble in water. To test the presence of other sugars, the Benedict's
test, Barfoed's test and other tests in sugar analysis are usually conducted.