TEST FOR CARBOHYDRATES

THE MOLISCH TEST

Shows positive test for: All carbohydrates. Monosaccharides give a rapid positive test. Disaccharides and polysaccharides react slower. Reactions: The test reagent dehydrates pentoses to form furfural (top reaction) and dehydrates hexoses to form 5-hydroxymethyl furfural (bottom reaction). The furfurals further react with -naphthol present in the test reagent to produce a purple product (reaction not shown).

How to perform the test: Two ml of a sample solution is placed in a test tube. Two drops of the Molisch reagent (a solution of -napthol in 95% ethanol) is added. The solution is then poured slowly into a tube containing two ml of concentrated sulfuric acid so that two layers form. A positive test is indicated by: The formation of a purple product at the interface of the two layers.

Objectives: Test for Reducing Sugars,

CHEMICALS
:
glucose fructose sucrose potassium sodium tartrate sodium hydroxide

CuSO4 5 H20 Fehling I consists of 7 g of hydrated copper(II) sulfate dissolved in 100 mL of dist. water. Fehling II is made by dissolving 35 g of potassium sodium tartrate and 10 g of sodium hydroxide in 100 mL of dist. water. Fehling's reagent: Equal volumes of Fehling I and Fehling II are mixed to form a deep blue solution.

Apparatus and glass wares:

hot plate thermometer 3 conical measures, graduated, 500 mL 3 glass stirring rods 3 beakers 40 mL 3 beakers 150 mL beaker 1800 mL measuring cylinder 100 mL

Hazards and safety precautions:

Sodium hydroxide is very corrosive. Causes severe burns. May cause serious permanent eye damage. Very harmful by ingestion. Harmful by skin contact or by inhalation of dust. Copper(II) sulfate is harmful when swallowed.

Safety glasses, protective gloves, good ventilation.

Experimental procedure:
5 g of glucose, fructose and sucrose, respectively, are dissolved in 100 mL of dist. water. Three 500 ml conical measures are each filled with the sugar solutions. Each solution is made up to the 500 mL mark with dist. water warmed up to 60 C. Afterwards 8 mL of Fehling reagent are poured into each of the sugar solutions while stirring.

Results:
In two conical measures a yellow-red precipitate is formed. At first in the blue solution the precipitate may appear greenish. Finally the muddy green suspension yields brick red precipitate. Fructose reacts faster than glucose. In the third glass no change can be observed.

conical glucose solution measure 1 conical fructose solution measure 2 conical sucrose solution measure 3 no change brick red precipitate brick red precipitate

Discussion: Fehling's tests for aldehydes are used extensively in carbohydrate chemistry. A positive result is indicated by the formation of a brick red precipitate. Like other

aldehydes, aldoses are easily oxidized to yield carboxylic acids. Cupric ion complexed with tartrate ion is reduced to cuprous oxide.

Fig. 1: Redox reaction

BENEDICT'S TEST
Shows positive test for: Reducing sugars Reactions: Reducing sugars are oxidized by the copper ion in solution to form a carboxylic acid and a reddish precipitate of copper (I) oxide.

How to perform the test:

One ml of a sample solution is placed in a test tube. Two ml of Benedict's reagent (a solution of sodium citrate and sodium carbonate mixed with a solution of copper sulfate) is added. The solution is then heated in a boiling water bath for three minutes.

A positive test is indicated by: The formation of a reddish precipitate within three minutes.

a negative test (left) and a positive test (right)

RESULTS Give positive carbohydrates test in Chips

TESTS OF PROTEIN
BIURET TEST
(i) On adding a dilute of copper sulphite to alkaline solution of protein, a violet colour is developed.

(ii)

This test is due to the presence of peptide

linkage.

MILLONS TEST
(a) Millons reagent consists of mercury dissolved in nitric acid (forming a mixture of mercuric & mercurous

nitrates). (b) (c) When millons reagent is added to a protein, a white ppt is formed, which turn brick red on heating. This test is given by protein which yield tyrosine on hydrolysis (due to the presence of phenolic group).

NIHYDRIN TEST
(i) (ii) This test is given by all proteins.

When protein is boiled with a dilute solution of ninhydrin, a violet colour is produced.

RESULTS Show negative protein test in chips.

CHEMICAL TEST FOR STARCH OR IODINE:

STARCH: Plants store glucose as the polysaccharide starch. The cereal grains (wheat, rice, corn, oats, barley) as well as tubers such as potatoes are rich in starch. Starch can be separated into two fractions--amylose and amylopectin. Natural starches are mixtures of amylose (10-20%) and amylopectin (80-90%). Chemical Test for Starch or Iodine:

Amylose in starch is responsible for the formation of a deep blue color in the presence of iodine. The iodine molecule slips inside of the amylose coil.

Iodine - KI Reagent: Iodine is not very soluble in water, therefore the iodine reagent is made by dissolving iodine in water in the presence of potassium iodide. This makes a linear triiodide ion complex with is soluble. The triiodide ion ion slips into the coil of the starch causing an intense blue-black color. Starch Test: Add Iodine-KI reagent to a solution or directly on a potato or other materials such as bread, crackers, or flour. A blue-black color results if starch is present. If starch amylose is not present, then the color will stay orange or yellow. Starch amylopectin does not give the color, nor does cellulose, nor do disaccharides such as sucrose in sugar. Iodine Test: When following the changes in some inorganic oxidation reduction reactions, iodine may be used as an indicator to follow the changes of iodide ion and iodine element. Soluble starch solution is added. Only iodine element in the presence of iodide ion will give the characteristic blue black color. Neither iodine element alone nor iodide ions alone will give the color result. Results
Give positive Starch test in Chips.