Ninhydrin Test for amino acid

Procedure:

Add about 2 mg of the sample to 1 mL of a solution of 0.2 g of ninhydrin (1,2,3indanetrione monohydrate) in 50 mL of water. The test mixture is heated to boiling for 15-20 sec; This reaction is important not only because it is a qualitative test, but also because it is the source of the absorbing material that can be measured quantitatively by an automatic amino acid analyzer. This color reaction is also used to detect the presence and position of amino acids after paper chromatographic separation.

Positive Test:

A blue to blue-violet color is given by a-amino acids and constitutes a positive test.

Other colors (yellow, orange, red) are negative.

Complications:

Proline, hydroxyproline, and 2-, 3-, and 4-aminobenzoic acids fail to give a blue color but produce a yellow color instead.

Ammonium salts give a positive test.

Some amines, such as aniline, yield orange to red colors, which is a negative test.