ON TYROSINE AND TRYPTOPHANE DETERMINATIONS IN PROTEINS

TLDR

The article reviews existing methods for quantifying tyrosine and tryptophan in proteins. The authors improved protein hydrolysis by 18‑20 h NaOH digestion followed by rapid neutralization and acidification, and developed a more accurate Millon‑based colorimetric assay for tyrosine using mercuric sulfate and sodium nitrite, including a protocol for preparing the reagent. The improved methods yield accurate amino‑acid contents, reporting 1.4 % tryptophan and 6.4 % tyrosine in casein, with comparable values for egg albumin, edestin, gliadin, and zein.

Abstract

The article summarizes research into the existing methods for the quantitative determination of tyrosine and tryptophane in proteins. Limitations to the accuracy of the Folin-Looney method (reaction of a phosphotungstic phosphomolybdic acid in a phenol solution, evaluated using colorimetry) have been solved by an improved method detailed in the text. The hydrolysis of proteinaceous material to allow chemical analysis of tryptophane has also been improved; the method is based on digestion with sodium hydroxide for 18-20 hours, followed by rapid neutralization and acidification with sulfuric acid. A more accurate test for tyrosine based on Millon's reaction has been developed; acidified mercuric sulfate solution is used to dissolve precipitated tyrosine and sodium nitrite is added to produce the colored product which is assessed colorimetrically. Two types of casein analyzed by these methods contained 1.4% tryptophane and 6.37-6.55% tyrosine. Tryptophane and tyrosine content of various materials were: casein 1.4%, 6.4-6.6%; egg albumin 1.3%, 4.0%; edestin 1.5%, 4.5%; gliadin 0.84%, 3.1%; zein 0.17%, 5.9%. A method for preparation of the pure mercuric sulfate reagent is described.