Amino acids and their metabolites as potential biochemical markers in postmortem vitreous humour

Background Putrefaction is the microbial metabolism/degradation of autolysis end products like amino acids. The aim of this study was to identify and investigate amino acids and their bacterial degradation products/metabolites by LC-HRMS as possible markers of putrefaction. Methods Pig eyes ( n = 60) were stored in conical centrifuge tubes and petri dishes within a climate chamber at 25°C, 65% humidity over four days. Each day, eyes were macroscopically examined and vitreous humour was drawn from up to six eyes. Twenty-one amino acids/metabolites (lysine, histidine, arginine, N 2 -acetyllysine, N 2 -acetylarginine, N 6 -acetyllysine, N -acetylcadaverine, tyrosine, N -acetylagmatine, tyramine, phenylalanine, N 2,6 -diacetyllysine, 2-phenylethylamine, tryptophan, kynurenic acid, N- acetyltyrosine, N -acetyltyramine, N -acetylphenylalanine, N -acetyltryptophan, N -acetyl-2-phenylethylamine, N -acetyltryptamine ) were monitored by LC-HRMS. Results Opacification was observed and eyes stored on petri dishes dried out faster and turned dark compared to eyes stored in centrifuge tubes. Amino acids were universally present and showed a significant increase by up to 46-fold from day 0 to 4, except for arginine (no trend observed). Amino acid metabolites were first detected on day 2 with exception of N 2 -acetyllysine, N 2 -acetylarginine, and N 6 -acetyllysine (detected on day 0). All analytes showed a marked increase by days 3 and 4, especially kynurenic acid and N -acetylated compounds. Among the six eyes of each storage condition/day high variability was observed, between centrifuge tube and petri dish storage few variability was found. Conclusion LC-HRMS analysis of amino acids and their metabolites in vitreous humour could be a promising tool to evaluate the putrefaction status. However, further studies are required to better understand inter-individual metabolic differences and influencing environmental factors.