Determination of tramadol and its metabolite O-desmethyltramadol in vitreous humor. Is it helpful in forensic casework?

In recent years, there has been increasing interest on the use of alternative biological materials in forensic toxicology. Vitreous humor is one of them, which, due to the closed cavity it is contained, has a low degree of contamination and high purity that makes it ideal for use in postmortem specimens. The aim of this study was to investigate the distribution of tramadol and its active metabolite O-desmethyltramadol in vitreous humor and the usefulness of using this alternative biological matrix in tramadol-related forensic cases. For this purpose, a gas chromatography-mass spectrometric method for the determination of the two analytes in blood and vitreous humor samples, which included solid-phase extraction and derivatization using N,O-Bis(trimethylsilyl)trifluoroacetamide with 1% trimethylsilyl chloride, was developed. The method was fully validated according to international guidelines and was applied to blood and vitreous humor samples from 12 forensic cases. Both substances were found to be readily distributed in vitreous humor, since even in cases of very low concentrations of the analytes in blood, their detection was also possible in vitreous humor. In addition, the vitreous humor to blood concentration ratios were calculated for both substances and the mean values were found to be 0.91 for tramadol and 0.94 for O-desmethyltramadol. The results of our study indicate that the information that can be extracted from the analysis of vitreous humor samples is particularly useful during the investigation of tramadol-related cases. Nevertheless, the need for further study of this alternative material to establish therapeutic and toxic limits becomes apparent.