Key role of keratin matrix in toxicological analysis to determine the cause of death of a poly-drug addict

Introduction and aim: A man was discovered at home, lying on his couch with foam around his mouth; beside him was a cup containing a decoction, likely made by boiling San Pedro leaves. An autopsy later revealed that he had an aortic malformation, which was considered a possible contributing factor in his death; consequently, a toxicological analysis of the subject's biological matrices was ordered by the prosecutor. Methods: The initial toxicological screening using LC-MS/MS detected amphetamine, methamphetamine, and mescaline in biological samples, further analysis using SPE and GC- MS/MS in MRM mode identified a compound with characteristics similar to MDMA. Subsequently, a 3 cm hair sample was collected and prepared as follows: washing with dichloromethane, segmentation into 1-cm fragments (p=0-1cm; m=1-2cm; d=2-3cm), pulverization, incubation with MeOH at 37°C for 18 hours, extraction with solid-phase columns and finally analyzed in GC-MS scan mode. Using the NIST mass spectral library, two compounds from the benzofuran class were identified: N-methyl-5-(2-aminopropyl)- benzofuran (5-MAPB) and 6-(aminopropyl)benzofuran (6-APB). Finally, an analytical method using GC-MS/MS was developed and successfully validated for confirmation and quantification purposes. Results: The validation allowed the calculation of LOD and LOQ for 5-MAPB and 6-APB, the values are respectively: LOD= 0.03, 0,02 ng/mg and LOQ= 0,08, 0,05 ng/mg. Analysis of the hair by GC-MS/MS provided a more complete understanding of the individual's intake of substances, as hair mainly retains unaltered compounds. The results confirmed the presence of 5-MAPB and elevated levels of 6-APB: segmental analysis revealed concentrations for 5- MAPB as p=9,77 ng/mg; m=11,36 ng/mg; d=12,16 ng/mg and for 6-APB as p=7,15 ng/mg; m=10,58 ng/mg; d=7,15 ng/mg. Conclusion: The keratin matrix enabled the identification of the subject as a habitual user of 5-MAPB and 6-APB. Furthermore, the analysis of other biological matrices confirmed that the deceased had used both compounds prior to death.