Assay of L-phenylalanine as phenylethylamine after enzymatic decarboxylation; application to isotopic studies.

Procedures have been reported for the use of specific bacterial decarboxylases to assay individual amino acids by measurement of the CO2 evolved (1). Since 1 mole of amine is formed for each mole of amino acid decarboxylated, it should also be possible to estimate the amount of amino acid from the quantity of amine formed. When L-phenylalanine is decarboxylated by L-phenylalanine decarboxylase, phenylethylamine is formed. Measurement of the phenylethylamine offers certain advantages over measurement of the COz. The amine can still be identified structurally with the parent amino acid and has solubility characteristics which permit it to be separated from all unchanged amino acids and from many other amines. The employment of enzymatic CO2 evolution as a specific assay method necessitates the use of highly purified enzyme preparations to remove other decarboxylases and other sources of COz. Formation of phenylethylamine can only be due to the presence of phenylalanine. Since the amine can be separated from unchanged amino acids and from other amines, it is possible to use unpurified extracts as sources of decarboxylase activity. A further advantage of the amine procedure is the increased sensitivity (0.01 PM) compared with that of the CO2 method (1 .O PM). With larger samples and more decarboxylase it is possible to form mg. quantities of phenylethylamine from L-phenylalanine. In radioactive studies it is therefore possible to obtain from an amino acid mixture the phenylethylamine, corresponding to the L-phenylalanine, in sufficient quantity and purity to perform isotopic measurements. In this communication there are reported two analytical applications of L-phenylalanine decarboxylase : (1) the determination of L-phenylalanine in tissues and protein hydrolysates and (2) the isolation of pure phenylethylamine hydrochloride for isotopic measurement from amino acid mixtures containing isotopic L-phenylalanine.