REACTIONS OF CARBOHYDRATES
Introduction:
• Carbohydrates are hydrophilic substances with a potential free aldehyde or keto group and
a number of hydroxyl groups.
• Monosaccharides, the simple sugars in this series, have one sugar carbon chain.
• Glucose (an aldo hexose) & Fructose (a keto hexose) are the most common
monosaccharides.
• Glucose is present in fruits, sugarcane, milk and other biologically important carbohydrates.
• Fructose is present in fruit juices & honey (richest source). It is also a constituent of cane
sugar and inulin.
GENERAL REACTIONS OF CARBOHYDRATES
EXPERIMENT OBSERVATION INFERENCE
1. Molisch’s Test:
To 2ml of sugar solution, 2-3 drops of A purple colored ring was
Indicates the presence
alcoholic α– naphthol (Molischs reagent) formed at the junction of
of carbohydrates in the
was added & mixed. Then 2ml of conc. the interface of the two
given solution.
H2SO4 was added carefully along the layers of the test tube.
sides of the tube without shaking.
Principle: Carbohydrates are dehydrated in the presence of conc. sulphuric acid to form furfural
derivatives (hydroxyl methyl furfural for hexoses and furfurals for pentoses). The ring is due to
the condensation product of furfural derivatives with α- naphthol.
2. Iodine Test: Indicates the presence
Blue color solution was
To 2 ml of sugar solution 2-3 drops of of starch in the given
formed.
iodine solution was added. solution.
Principle: Iodine forms a colored adsorption complex on the surface of the polysaccharides like
starch, dextrin and glycogen.
3. Benedict’s Test: Green to Brick Red
5 ml of Benedict’s reagent was taken in a colored precipitate was
test tube and boiled for 1 minute. Then 8 formed. Indicates the presence
drops (0.5ml) of sugar solution was Green = 0.5-1.0 gm% of reducing sugars in
added & boiled again for 1 minute and Yellow = 1.0-1.5gm% the given solution.
cooled. Benedicts reagent consists of: Orange = 1.5- 2.0 gm%
(Na2 CO3; CuSo4 & Sodium citrate) Red or brick = >2.0gm%
Principle: Carbohydrates with free aldehyde or keto groups in alkaline medium form enediols.
Enediols are powerful reducing agents which reduce cupric ions to cuprous ions. The red color
is due to the formation of cuprous oxide (Cu2O).
4. Barfoed’s Test:
To 3 ml of Barfoed’s reagent 1 ml of Scanty red scum was Indicates the presence
sugar solution was added & boiled for 1 formed at the top and of monosaccharaides in
minute. Allow the test tube to cool in the sides of the tube. the given solution.
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EXPERIMENT OBSERVATION INFERENCE
test tube stand. Barfoeds reagent consists This test is specific for
of: monosaccharides
(Copper acetate in glacial acetic Acid)
Principle: Monosaccharides act also as reducing agents in weak acidic conditions. Barfoed’s
reagent is weakly acidic and is only reduced by monosaccharides (cupric ions to cuprous oxide).
5. Seliwanoff’s Test:
To 3ml of Seliwanoff’s reagent 0.5 ml of
Indicates the presence
sugar solution was added & boiled for 30 Cherry red color was
of ketose sugar in the
seconds and allowed to cool. formed.
given solution.
Seliwanoffs reagent consists of:
(Resorcinol in HCl)
Principle: Ketohexoses on treatment with HCl forms furfural derivatives. Cherry red color is
due to the condensation of furfural derivatives with resorcinol.
6. Rapid furfural test:
To 3ml of sugar solution 2-3 drops of 1% Indicates the presence
alcoholic α-naphthol (Molischs reagent) of ketose sugar in the
Deep purple color was
was added and mixed, then 3ml of given solution. It is a
observed.
concentrated HCl was added and boiled confirmatory test for
for ten seconds and allowed to cool. ketoses.
Principle: Keto hexoses on treatment with HCl forms furfural derivatives. Deep purple color is
due to the condensation of furfural derivatives with alcoholic α-naphthol.
7.Benedict’s test after acid
hydrolysis(Special test for sucrose):
To 5ml of sucrose solution, 5 drops of
Indicates the presence
conc. HCl was added and Boil for 2 mins
of reducing sugars after
& Cooled. Then Neutralized with 20% A brick red precipitate
acid hydrolysis.
Na2CO3 till the effervescence ceases. was formed.
Sucrose consists of
Perform Benedict’s test with this
glucose and fructose.
neutralized solution as follows: 0.5ml
Solution + 5ml Benedict’s reagent. Boil
for 2 min over a low flame and cooled.
Principle: Sucrose gives a negative Benedict’s test as it is a non-reducing sugar. But when
Benedict’s test is repeated with the hydrolysate, after acid hydrolysis it gives a positive
Benedict’s test, indicating the hydrolysis of sucrose to its constituent monosaccharides glucose
and fructose. Free glucose and fructose are reducing in nature and thus give a positive Benedict’s
test.
8.Osazone Test:
Take 1 spatula of phenyl Needle shaped – Confirms the presence of
hydrazine hydrochloride, 2 Glucosazones/Fructosazones. respective sugars in the
spatula of sodium acetate & 8 given solution.
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EXPERIMENT OBSERVATION INFERENCE
drops of glacial acetic acid. Add Sunflower shaped –
5 ml of sugar Solution. Mix, Maltosazones.
label &keep in boiling water
bath for 20 minutes Hedge – hog/ powder puff
(monosaccharides); 40 minutes shaped – Lactosazones.
(disaccharides).
Let it cool on the test tube rack.
Place a drop of yellow crystal on
a clean glass slide. Cover it with
cover slip and examine under a
microscope.
Principle: Osazones are yellow crystalline compounds formed when a solution of sugar with
a free carbonyl group is heated with excess of phenyl hydrazine hydrochloride. Each sugar
will give rise to an osazone of a definite crystalline form and has its own melting point.
First, the two carbon atoms take part in the reaction. Phenyl hydrazine reacts with carbonyl
group of the sugar to form phenyl hydrazone, which then reacts with two further molecules of
phenyl hydrazine in which rearrangement between nitrogen and hydrogen molecules takes
place and finally results in the formation of osazones.
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PERFORM THE FOLLOWING TESTS WITH GLUCOSE AND FRUCTOSE:
1. Molischs test
2. Benedicts test
3. Barfoeds test
4. Seliwanoffs test
5. Rapid furfural test
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REACTIONS OF DISACCHARIDES
Disaccharides are the carbohydrates, which on hydrolysis forms two monosaccharide units.
Maltose, lactose and sucrose are the common disaccharides. The former two are reducing
disaccharides, whereas Sucrose is a non-reducing disaccharide.
Maltose:
• It is a disaccharide made up of two glucose units joined together by α-1, 4 glycosidic bonds.
• Since, one of the aldehyde groups remains free, it is a reducing sugar.
• It is also called Malt sugar and is seen in Malt, Germinating seeds & obtained on hydrolysis
of starch.
Lactose:
• It is also a reducing disaccharide. It is made up of a molecule of α-D-glucose united to β-D-
galactose by β-1, 4 glycosidic bonds.
• It is present in milk (Milk sugar) and can be hydrolysed to glucose & galactose by the
enzyme lactase present in intestinal juice.
Sucrose:
• It is a Non-reducing disaccharide. It is made up of one molecule of α-D-glucose and one
molecule of β-D fructose united by α, β-1, 2 glycosidic bond (between the aldehyde and
keto groups of glucose and fructose respectively).
• Hence there is no free functional group in sucrose and so it is a Non-reducing sugar. It does
not form osazones.
• It is also called Invert sugar & Table sugar.
• Sucrose occurs widely in nature as cane sugar, beet sugar & in several ripe fruits.
Trehalose:
• It is a Non-reducing disaccharide formed of two glucose units joined by α-1,1glycosidic
bond. It is seen in hemolymph of some insects, fungi & yeasts.
PERFORM THE FOLLOWING TESTS WITH MALTOSE, LACTOSE AND SUCROSE.
1. Molischs test
2. Benedicts test
3. Barfoeds test
4. Hydrolysis of sucrose
5. Benedicts test in hydrolysate
6. Seliwanoffs test in hydrolysate
7. Rapid furfural test in hydrolysate
8. Osazone test
NAME OF TEST MALTOSE LACTOSE SUCROSE
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MOLISCHS TEST Positive Positive Positive
BENEDICTS TEST Positive Positive Negative
BARFOEDS TEST Negative Negative Negative
Hydrolysed to
HYDROLYSIS OF
SUCROSE
__ __ glucose and
fructose
BENEDICTS TEST
IN __ __ Positive
HYDROLYSATE
SELIWANOFFS
TEST IN __ __ Positive
HYDROLYSATE
RAPID FURFURAL
__
TEST IN __ Positive
HYDROLYSATE
OSAZONE TEST Petal shape Woolen ball shape __
TEST FOR POLYSACCHARIDES:
EXPERIMENT OBSERVATION INFERENCE
1. MOLISCHS TEST Positive General test
Intense blue colour Starch
2. IODINE TEST Brown red colour Glycogen
Purple colour Dextrin
3. BENEDICTS TEST Negative Non reducing sugar
4. Hydrolysis of starch: add 5ml of 2N HCL to 20 ml of 1% starch solution. Divide the
solution equally in 5 test tubes. Heat all the test tubes keeping in boiling water bath. remove
the tube one by one after 1, 5, 8, 12 and 20 minutes of boiling. Simultaneously cool it and
go for iodine test putting a drop of solution on iodine in a tile. With rest of the solution do
benedicts test (after neutralizing with a pinch of NaHCO3) and see the result.
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ACID HYDROLYSIS OF STARCH :
TIME COLOUR WITH REDUCTION STATE OF
IODINE HYDROLYSIS
1 minute Intense blue Nil Nil
5 minute Light blue - Slight
(soluble starch)
8 minute Purple + More
(amylodextrin)
12 minute Reddish Brown ++ Still more
(erythro dextrin)
20 minute No colour +++ Nearly complete
(acrodextrin)
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