Spectrophotometric analysis of amino acids and peptides with their copper salts.

The Pope and Stevens method (1) for quantitative determination of amino acids and peptides is based on iodometric analysis of the combined copper, a procedure that is somewhat inconvenient. Woiwood (a), using the color produced by diethyldithiocarbamate, has described a spectrophotometric modification of the Pope and Stevens method. Martin and Mittelmann (3) have used polarographic analysis of copper. Schroeder, Kay, and Mills (4) have improved the Pope and Stevens method to obtain precise and accurate results for amino acids, but they also use iodometric analysis of copper. The present paper describes a simple and rapid spectrophotometric method for determining amino acids and peptides which is based on the color obtained when the combined copper is converted to the copper salt of alanine, together with an application of the procedure in studying the rate of protein hydrolysis. Color intensities of equimolar concentrations of copper complexes of amino acids vary. Therefore, any one amino acid cannot be used as a reference standard for the determination of the other amino acids. However, an excess of an amino acid added to solutions of the copper complexes of other amino acids converts their copper largely to the complex of the added amino acid which can then serve as a standard. Alanine was chosen as standard because of its relative high solubility, the ease of its purification, its cheapness, and because it gives a color intensity near to or intermediate between those of the other amino acids.