Application of l-methionine γ-lyase in chiral amino acid analysis

Abstract

Here, a conventional chiral amino acid analysis method using high-performance liquid chromatography was coupled with a sample pretreatment using l-methionine γ-lyase from Pseudomonas putida ICR 3460 for the selective analysis of l-methionine and l-tryptophan. The sample was analyzed after the degradation of l-methionine with l-methionine γ-lyase, as l-methionine coelutes with l-tryptophan under the standard chiral amino acid analytical conditions used for precolumn derivatization with o-phthalaldehyde and N-acetyl-l-cysteine. The l-tryptophan in the sample was then eluted as a clearly separated peak and analyzed further. Since the l-methionine γ-lyase did not act on l-tryptophan, we were able to calculate the l-methionine or l-tryptophan concentration based on the data obtained from 2 individual runs: the sample with and without l-methionine γ-lyase pretreatment. The concentration of l-tryptophan was calculated from the data obtained from the sample with l-methionine γ-lyase pretreatment, while the concentration of l-methionine was calculated using the following equation: l-methionine concentration = {the data from the sample without l-methionine γ-lyase pretreatment}-{the data from the sample with l-methionine γ-lyase pretreatment}. Model samples containing authentic amino acids and a fermented food sample were analyzed by our method, and the calculated concentrations of l-methionine and l-tryptophan were consistently in agreement with the theoretical values.