Standardization of analytical method to determination of phenylalanine exposure in casein hydrolysates by second derivative spectrophotometry

Abstract

Dietetic formulations for phenylketonurics generally contain free or low phenylalanine protein hydrolysates. Several hydrolytic conditions have been proposed in the literature to increase phenylalanine (Phe) exposure, in order to favor its subsequent removal. The second derivative spectrophotometry (SDS), a fast, simple and low cost analytical method, was used, in this work, for the determination of Phe exposure in different casein hydrolysates, prepared by using papain, and also for the characteriztion of aromatic amino acids. A temperature of 37 ºC, an enzyme:substrate ratio of 4% and a pH 7.5 showed to be the most suitable condition for preparing phenylketonuric diets, since they led to the highest Phe exposure. Concerning the characterization of aromatic amino acids, a good separation of tyrosine (Tyr) and tryptophan (Trp) peaks was obtained in pH 13.0, using the SDS. Moreover, the second derivative spectra of the aromatic amino acids were similar to those of the protein hydrolysates.