A New Method for the Microdetermination of Free and Total Nonprotein-bound Tryptophan in Plasma

A new technic for the determination of tryptophan in plasma, based on the tryptophan-dimethylaminobenzaldehyde reaction, is described. It provides a method for the determination of as little as 20 i.g. of tryptophan and also permits the de-termination of bound tryptophan in nonprecipitable peptides, using 10 % perchloric acid as the deproteinizing agent. rFHE EVER-INCREASING APPLICATION of chromatographic methods allows most of the free amino acids present in plasma to be assayed with relative simplicity. However, tryptophan determination is difficult by these methods because of its rapid destruction in an acid medium. This fact creates a serious problem when it is necessary to know the com-plete plasma amino acid pattern in nutrition experiments or Some clinical studies. This circumstance led us to study a useful procedure for the microdetermination of free tryptophan in plasma. Rhode (1) was the first to point out the possibility of determining protein tryptophan by using the reaction between the amino acid and dimethylanlinobenzaldehyde lii the presence of an oxidizing agent. Using this reaction as a basis, several technics were published for the determination of tryptophan in proteins and foods (2-6). However, the determination of free tryptophan is not satisfactorily performed by this reaction because the amino acid is partially destroyed in the strongly acid reaction medium, as it was pointed out by one of us in a previous paper (4). Moreover, that technic requires samples containing at least 100-200 pg. of tryptophan. Since free tryptophan concentration in plasnia rarely exceeds 5 mg./100 ml., the procedure is unsuitable for such assays