PROTEINS 



143 



color is due to the presence of two peptide groups, — CO'NH — . When 

 three amino acids are joined, two such groups are formed, for example, 



HNH-CH- CO-NH 'CH' CO-NH -CH'COOK 



I I I 



Ri xvt Rt 



Tripeptides, with the exception of glycyl-glycyl-glycine, give the test, 

 as do also peptones, proteoses, and all native proteins. It is one of the 

 most general of the protein color tests. 



The Hopkins-Cole Test. A purplish color is developed when a protein 

 containing tryptophan radicals is treated with the Hopkins-Cole reagent 

 (magnesium glyoxylate). The cause of the color development is the 

 indole group, which exists in the amino acid, tryptophan. It has been 

 assumed that the color is due to the formation of indigo by the action 

 of the reagents on indole groups. 



The Ninhydrin Test. All amino acids (except proline and hydroxy- 

 proline) and, hence, all proteins give a blue to purple color with ninhydrin. 



CO 

 CeH. C(OH)j 

 CO 



Only a free amino and a free carboxyl group are required for the test. 

 All amino acids except the two mentioned possess such groups. The blue 

 color results from a condensation of two molecules of the reagent with 

 ammonia, which splits off from the amino group of the amino acid. 

 The color compound is the anion of a salt and has the following formula: 



CeH. C— N=C ^CeH* 

 C-0- CO 



Precipitation 



The precipitatin of proteins varies considerably with the reagent that 

 is used. Some reagents precipitate only the globulins, whereas others 

 precipitate a larger number of proteins but do not precipitate the pep- 

 tones. Some reagents precipitate not only the proteins and peptones 

 but also carry down certain amino acids. Among the most effective 

 precipitants are ammonium sulfate, magnesium sulfate, mercury salts, 

 trichloracetic acid, phosphotungstic acid, tungstic acid, tannic acid, col- 

 loidal iron, and strong solutions of alcohol. Phosphotungstic acid appears 

 to be the reagent which precipitates the largest percentage of nitrogen, 

 whether this is in the form of proteins, peptones, amino acids, or other 

 nitrogenous compounds. 



