Take 5 drops of extract in an incubation tube and add 5 drops of 1% starch solution, mix it and then cover the mixture with a layer of toluene.
In another incubation tube take 5 drops of boiled extract, add 5 drops of starch solution. This will serve as the control experiment.
Incubate both the tubes in a bath at 37°C for about one hour. After one hour take a small part of the incubated mixture and add iodine solution.
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No change in colour of iodine solution will indicate the presence of amylase enzyme. Take another part of the extract, add few drops of Benedict’s solution and heat it till it boils.
A brick-red precipitate will confirm the presence of amylase enzyme in the extract. Repeat the above two tests with the control mixture.
In the first test violet colour will confirm the presence of starch as such which has not been digested.
Starch always gives bluish-violet colour with iodine solution. In the experimental solution the enzyme extract is mixed with iodine solution, the amylase present in the enzyme extract hydrolyses the starch into maltose and glucose, therefore, there is no change in the colour of iodine.
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In the control experiment amylase is destroyed by boiling the enzyme extract, the starch is not hydrolyzed into maltose and glucose, therefore, it gives bluish-violet colour with the iodine solution.
In the second test with Benedict’s solution a red precipitate of cuprous oxide appears. In this glucose is formed. Since glucose is a reducing sugar, it reduces copper sulphate into cuprous oxide.