Qualitative Tests for Amino Acids
•
There are a number of qualitative tests to detect the presence of amino acids
•
These are largely dependent on the nature of R-group.
Ninhydrin Test
•
A color reaction given by amino acids and peptides on heating with the
chemical ninhydrin.
•
The technique is widely used for the detection and quantitation (measurement)
of amino acids and peptides.
•
Ninhydrin is a powerful oxidizing agent which reacts with all amino acids
between pH 4-8 to produce a purple colored-compound.
•
The amino acids proline and hydroxyproline also reacts but produce a yellow
color.
Principle
α-amino acid + 2 ninhydrin ---> CO2 + aldehyde + final complex (purple) +
3H2O
In summary, ninhydrin, which is originally yellow, reacts with amino acid and
turns deep purple. It is this purple color that is detected in this method.
Procedure:
1. 2 ml amino acid solution.
2. 1 ml Ninehydrine solution.
3. Heat in boiling WB for 5 min.
Test
Ninhydrine test
Observation
Result
Color complex
Amino acid present
Millon Test
•
This reaction is used to detect the presence of phenol (hydroxybenzene) which
reacts with Millon's reagent to form red complexes.
•
The only phenolic A.A is tyrosine.
Procedure:
1.
1 ml amino acid
2. 5 drops of Millon reagent
3. Heat the mixture in BWB for 10min
Test
Observation
Result
Millon's test
Brick red color.
Amino acid present
(tyrosine)