Question

Propose (3) three qualitative chemical tests that can differentiate the given peptides below:

Trp-Ala-Arg-Met-Ser (1)

Pro-Phe-Cys-Arg-Glu (2)

            Support your choice of tests with brief explanation

Answer 1

To differentiate the peptides, we have to test for different amino acids present in a particular peptide. For example, in peptide 1, tryptophan, alanine, arginine, methionine and serine are present. Here, we can look for tryptophan, alanine, methionine or serine but not for arginine since it is a common amino acid in both the given peptides.

Three qualitative chemical tests that can be used for differentiating the given peptides are:

1. Hopkins cole test

This test is specifically used for detecting tryptophan. The indole moiety present in tryptophan reacts with glyoxylic acid in the presence of concentrated sulphuric acid to give a purple colored product.

In this test, only peptide 1 (containing tryptophan) will give purple coloured adduct showing a positive reaction.

2. Lead sulphite test

This test is used for detection of sulphur contatining amino acid such as Cysteine. When cysteine is heated with NaOH (strong alkali), some of the sulphur is converted to sodium sulphide (Na2S) which can be detected by precipitating it to lead sulphide (PbS) using lead acetate solution. These precipitates are black in colour.

S (Cysteine) + 2NaOH          Na2S

Na2S + (CH3COO)2Pb            PbS + CH3COONa

Methionine gives negative result in lead sulfide test because sulfur present in methionine does not split under strong alkaline conditions.

Using this test, peptide 1 can be differentiated from peptide 2. Here, peptide 2 containing cysteine will give black reaction whereas peptide 1 will not.

3. Xanthoproteic acid test

This test is specific for aromatic amino acids, such as tyrosine and tryptophan. In the presence of concentrated nitric acid, the aromatic phenyl ring gives give yellow nitro-derivatives. At alkaline pH, the color of the reaction changes to orange due to the ionization of the phenolic group i.e. the salt of these amino acids give orange colour. Phenylalanine, however being aromatic amino acid, does not respond to this test, because of the absence of an activated benzene ring in phenylalanine.

In this test, only peptide 1 (containing tryptophan) will give yellow coloured product showing a positive reaction.

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