Virtual LAB Output on Qualitative Analysis of Carbohydrate.

Introduction
Sugars, which are made up of carbon, hydrogen, and oxygen, are the most common
natural mixtures found in living organic species. Sugars are an important source of
energy for living beings to generate electricity. These are referred to as sugars because
they are carbohydrates. A large number of them have the overall Cx(H2O)y equation.
A. Reactions with Different Reagents
1. Molisch's test (named after Austrian botanist Hans Molisch) is a sensitive
chemical test for the presence of carbohydrates. It is based on the dehydration
of the carbohydrate by sulfuric acid or hydrochloric acid to produce an
aldehyde, which condenses with two molecules of phenol (usually -naphthol,
though other phenols also produce colored products) to produce a red- or
purple-colored compound.
2. Bial's test is a chemical method for detecting pentoses. It is named after
German physician Manfred Bial. Orcinol, hydrochloric acid, and ferric chloride
are among the ingredients. If there is a pentose present, it will be dehydrated to
produce furfural, which will then react with the orcinol to produce a colorful
material. A precipitate may form if the solution turns bluish. Two absorption
bands are visible in the solution, one in red between Fraunhofer lines B and C
and the other around the D line.
3. Seliwanoff’s test is a chemical method for determining the difference between
aldose and ketose sugars. The ketone/aldehyde functionality distinguishes
ketoses from aldoses. A ketose is a sugar that has a ketone group, while an
aldose is a sugar that contains an aldehyde group. This test is based on the fact
that ketones dehydrate more quickly than aldoses when heated. It is named for
Theodor Seliwanoff, the chemist who invented the test in the first place.
The reagents consist of resorcinol and concentrated hydrochloric acid:
● Polysaccharides and oligosaccharides are acid hydrolyzed to produce simpler
sugars, which are then followed by furfural.
 The resorcinol combines with the dehydrated ketose to give a deep cherry red
color. Aldoses may react in a minor way, resulting in a faint pink tint. Samples
should not be heated for too long during this test.
Two common sugars that result in a positive test are fructose and sucrose. Sucrose, a
disaccharide made up of fructose and glucose, results in a positive test.
B. Test based on the reducing property of sugars
A reducing sugar is one that possesses a free aldehyde group or a free ketone group
and hence can operate as a reducing agent. All monosaccharides, as well as some
disaccharides, oligosaccharides, and polysaccharides, are reducing sugars. The
aldoses, which have an aldehyde group, and the ketoses, which have a ketone group,
are the two types of monosaccharides. Galactose, glucose, and fructose are three
typical dietary monosaccharides that are all reducing sugars. Disaccharides are made
up of two monosaccharides and are divided into two categories: reducing and
nonreducing. Nonreducing disaccharides, such as sucrose and trehalose, have
glycosidic linkages between their anomeric carbons, preventing them from converting
to an open-chain form with an aldehyde group. Since only one of the two anomeric
carbons in reducing disaccharides like lactose and maltose is engaged in the glycosidic
link, they can change to an open-chain form with an aldehyde group. In the Tollens' or
Benedict's tests, for example, the aldehyde functional group permits the sugar to act
as a reducing agent. Certain ketoses can undergo tautomerization to create aldoses,
and their cyclic hemiacetal forms can open to reveal an aldehyde. Acetals, such as
those found in polysaccharide bonds, do not readily convert to free aldehydes.
Furthermore, reducing agents like carbohydrates are vulnerable to oxidizing
agents like Benedict's, Fehling's, and Barfoed's reagents during the oxidation process.
However, the reducing end is not present in all disaccharides or polysaccharides.
Sucrose, amylase, and amylopectin are three examples. The former has no hemiacetal
group on either end, whereas the two d have one per 2–10,000 monosaccharide units.
As a result, the reducing group concentration has increased.
1. Benedict's reagent (also known as Benedict's Solution or Benedict's Qualitative
Solution) is a chemical reagent named for Stanley Rossiter Benedict, an American
scientist.
Benedict's reagent is a chemical reagent that is usually used to detect the
presence of reducing sugar(s), although it can also detect other reducing compounds.
All monosaccharides and many disaccharides, such as lactose and maltose, are
included.
Benedict's test will typically detect the presence of aldehydes and alpha-
hydroxy-ketones, including those seen in some ketoses. Although ketose fructose is
not precisely a reducing sugar, it is an alpha-hydroxy-ketone that yields a positive test
because the base in the reagent converts it to the aldoses glucose and mannose.
A color change from clear blue to a brick-red precipitate indicates a positive
Benedict's reagent test.
2. The Barfoed's test is a chemical procedure for detecting monosaccharides. It
works by converting copper(II) acetate to copper(I) oxide (Cu2O), which results in a
brick-red precipitate.
RCHO + 2Cu2+ + 2H2O → RCOOH + Cu2O↓ + 4H+
(Disaccharides can also react, but it takes a lot longer.) The monosaccharide's
aldehyde group, which ordinarily forms a cyclic hemiacetal, is oxidized to carboxylate.
3. Tollens' reagent is a chemical reagent that is used to detect aldehyde or alpha-
hydroxy ketone functional groups. A solution of silver nitrate and ammonia makes up
the reagent. It was named after the German scientist Bernhard Tollens, who
discovered it. The precipitation of elemental silver, which typically produces a
characteristic "silver mirror" on the inner surface of the reaction jar, indicates a
positive test with Tollens' reagent.
4. Fehling's solution is a chemical reagent that can be used to distinguish between
water-soluble carbohydrate and ketone functional groups, as well as a test for
reducing and non-reducing sugars, in addition to Tollen's reagent. The presence of
aldehydes, but not ketones, is identified in this test by reducing a deep blue copper(II)
solution to a red precipitate of insoluble copper oxide. The test is often used to reduce
sugars, however it is not known to be aldehyde specific. Fructose, like a cation,
produces a positive test using Fehling's solution.

II. OBJECTIVE
To be able to identify types of monosaccharides using different specific chemical
tests.

Materials used
Samples:
∙ Glucose ∙ Sucrose ∙ Starch

Reagents:
∙ Distilled water ∙ Molisch’s reagent ∙ Fehling’s solution A
∙ Fehling’s solution B ∙ Benedict’s reagent ∙ Tollen’s reagent
∙ Iodine solution

Apparatus:
∙ Test tube ∙ Dropper ∙ Beaker

Procedure and Observation:

Solubility test
Take a small amount of the given sample in a test tube and add 3 ml of distilled water
into it and shake the test tube well.
Observation:

SI Sample Observation
No.
1 Glucose Soluble
2 Sucrose Soluble
3 Starch Insoluble

Inference:
Glucose and sucrose are water soluble sugars, whereas starch is not.
Benedict’s test
Take 1-2 ml of aqueous solution of the sample in a test tube and add 1-2 ml of
Benedict’s reagent. Keep the test tube in a boiling water bath.
Observation:

SI Sample Observation
No.

1 Glucose It is soluble because glucose is a reducing sugar that lowers the

CU(II) ions in the test reagent to red-precipitate copper (I) oxide.

2 Sucrose Since the test results are negative and the liquid remains
unchanged, the sucrose is insoluble.

3 Starch The test is negative, and the liquid remains the same, indicating
that it is insoluble.

Inference:
As a conclusion, in Benedict's test, the sucrose and starch are insoluble, whilst the
glucose is soluble.

Tollen’s test
Take 2-3 ml of aqueous solution of the sample in a test tube and add 2-3 ml of Tollen’s
reagent. Keep the test tube in a boiling water bath for 10 minutes.
Observation:

SI Sample Observation
No.

1 Glucose It is insoluble because there was no reaction to the test in terms

of lowering carbohydrates.

2 Sucrose Sucrose did not respond in the same way that glucose did. As a
result, Sucrose is insoluble.

3 Starch When the starch was added to the boiling water after 10
minutes, it did not show any changes. As a result, the starch is
insoluble.
Inference:
The results of the three tests revealed that glucose, sucrose, and starch did not
change after 10 minutes in boiling water, indicating that they are insoluble.

Iodine test
Take a small amount of aqueous solution of the sample in a test tube and add 1-2
drops of iodine solution into it.
Observation:

SI Sample Observation
No.

1 Glucose There will be no noticeable changes in starch, indicating that it

is insoluble.

2 Sucrose The presence of sucrose in the test suggests that no changes

have been done in terms of dissolving the starch, indicating that
it is insoluble.

3 Starch When the test was completed, the liquid changed color,
indicating that the starch was soluble.

Inference:
To summarize, glucose and sucrose are both insoluble sugars, whereas starch is.

Tollen’s test
Take 2-3 ml of aqueous solution of the sample in a test tube and add 2-3 ml of Tollen’s
reagent. Keep the test tube in a boiling water bath for 10 minutes.

Observation:

SI Sample Observation
No.

1 Glucose Tollen's reagent is reduced to elemental silver by the'reducing'

sugar glucose, leaving a silvery mirror surface on the inside of
 the test tube.

2 Sucrose There are no indicators of change that show the absence of

reducing carbohydrate.

3 Starch After 10 minutes, there are no characteristics of change that

indicate the lack of reducing carbohydrate.

Inference:
As a result, only the Glucose leaves a silver surface inside the test tube.

Iodine test
Take a small amount of aqueous solution of the sample in a test tube and add 1-2
drops of iodine solution into it.

Observation
SI Sample Observation
No.

1 Glucose After adding a solution to the Glucose, there are no

characteristics of change that indicate the absence of starch.

2 Sucrose There is no characteristics of change indicating the absence of

starch.

3 Starch Since starch is present, the color of the solution changes to

blue.

Inference:
As a result, only the Starch turns blue, indicating its presence.

Conclusion:
In conclusion, Benedict's test, Tollen's test, and Iodine test are used to assess
the solubility and reactivity of carbohydrates. However Benedict's test is not used to
identify the reducing carbohydrates in glucose, sucrose, and starch.
 Glucose is soluble in Benedict's test. Glucose, sucrose, and starch, on the other
hand, are not soluble in Tollen's test. Furthermore, the starch is soluble in an Iodine
test.
Lastly, only the glucose is a reducing sugar based on Tollen’s test. Since in the
Iodine test, it changed the solution to blue, the presence of starch in the starch
sample was determined.