Metabolite markers for three synthetic tryptamines N‐ethyl‐N‐propyltryptamine, 4‐hydroxy‐N‐ethyl‐N‐propyltryptamine, and 5‐methoxy‐N‐ethyl‐N‐propyltryptamine

• Published in: Drug testing and analysis 2024-12, Vol.16 (12), p.1544-1557
• Main Authors: Bergh, Marianne Skov‐Skov, Bogen, Inger Lise, Grafinger, Katharina Elisabeth, Huestis, Marilyn A., Øiestad, Åse Marit Leere
• Format: Article
• Language: English
• Subjects: 4‐hydroxy‐N‐ethyl‐N‐propyltryptamine (4‐OH‐EPT); 5‐methoxy‐N‐ethyl‐N‐propyltryptamine (5‐MeO‐EPT); Biomarkers - metabolism; Humans; Hydroxylation; Metabolism; metabolite; Metabolites; microsomes; Microsomes, Liver - metabolism; new psychoactive substances (NPS); N‐ethyl‐N‐propyltryptamine (EPT); Psychotropic Drugs - chemistry; Psychotropic Drugs - metabolism; Substance Abuse Detection - methods; synthetic tryptamines; Tandem Mass Spectrometry - methods; Tryptamines - chemistry; Tryptamines - metabolism; ultra‐high performance liquid chromatography–quadrupole time‐of‐flight mass spectrometry (UHPLC‐QTOF‐MS)
• ISSN: 1942-7603; 1942-7611; 1942-7611
• DOI: 10.1002/dta.3668

N‐Ethyl‐N‐propyltryptamine (EPT), 4‐hydroxy‐N‐ethyl‐N‐propyltryptamine (4‐OH‐EPT), and 5‐methoxy‐N‐ethyl‐N‐propyltryptamine (5‐MeO‐EPT) are new psychoactive substances classified as tryptamines, sold online. Many tryptamines metabolize rapidly, and identifying the appropriate metabolites to reveal intake is essential. While the metabolism of 4‐OH‐EPT and 5‐MeO‐EPT are not previously described, EPT is known to form metabolites by indole ring hydroxylation among others. Based on general knowledge of metabolic patterns, 5‐MeO‐EPT is also expected to form ring hydroxylated EPT (5‐OH‐EPT). In the present study, the aim was to characterize the major metabolites of EPT, 4‐OH‐EPT, and 5‐MeO‐EPT, to provide markers for substance identification in forensic casework. The tryptamines were incubated with pooled human liver microsomes at 37°C for up to 4 h. The generated metabolites were separated and detected by ultra‐high performance liquid chromatography–quadrupole time‐of‐flight mass spectrometry analysis. The major in vitro EPT metabolites were formed by hydroxylation, N‐dealkylation, and carbonylation. In comparison, 4‐OH‐EPT metabolism was dominated by double bond formation, N‐dealkylation, hydroxylation, and carbonylation in vitro and hydroxylation or carbonylation combined with double bond loss, carbonylation, N‐dealkylation, and hydroxylation in vivo. 5‐MeO‐EPT was metabolized by O‐demethylation, hydroxylation, and N‐dealkylation in vitro. The usefulness of the characterized metabolites in forensic casework was demonstrated by identification of unique metabolites for 4‐OH‐EPT in a human postmortem blood sample with suspected EPT or 4‐OH‐EPT intoxication. We characterized the major metabolites of the synthetic tryptamines EPT, 4‐OH‐EPT, and 5‐MeO‐EPT after incubation with pooled human liver microsomes. The usefulness of the discovered metabolites in forensic casework was demonstrated by identification of unique metabolites for 4‐OH‐EPT in a human postmortem blood sample with suspected EPT or 4‐OH‐EPT intoxication.