OSAZONE TEST – For differentiation of reducing sugar

Osazone test is performed to differentiate the reducing sugars on the basis of crystal formation.

REAGENTS:

1. Phenylhydrazine mixture consisting of:

i. Phenylhydrazine, and

ii. Anhydrous sodium acetate.

2. Original solution (0.8.) – containing a carbohydrate

PRINCIPLE:

Aldehyde or keto group containing sugars reacts with the amine group of phenylhydrazine to give phenylosazone.

PROCEDURE:

• Take phenylhydrazine mixture and till about half an inch of the test tube with it.
• Add 3 to 5ml of sugar solution (original solution) to the test tube.
• Mix thoroughly and place it in a boiling water bath.
• Note the appearance of a yellow crystalline precipitate in the test tube.
• Remove and allow the test tube to cool slowly.
• Do not cool the test tube under tap water.
• Examine the crystals under the microscope and make a drawing.
• Draw only what you see.

NOTE:

Osazones of glucose are formed in 10-15 minutes whereas those of disaccharides take up to 45 minutes. Sucrose does not react with phenylhydrazine to form crystalline compounds called osazones.

Observation and Result:

SUGAR	OSAZONE	SHAPE
Glucose	Glucosazone	Needle-shaped
Fructose	Fructosazone	Needle-shaped
Galactose	Galactosazone	Fluffy ball shaped
Lactose	Lactosazone	Fluffy ball shaped
Maltose	Maltosazone	Sunflower shaped

 

Precautions:

• Wash the apparatus before and after the experiment.
• Carefully Observe the Crystals under the microscope.
• Carefully handle all the chemicals.

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