Analytical characterization of novel psychoactive substances and adulterants: ortho-methyl fentanyl, N,N-dimethyltryptamine, and medetomidine
TUScholarShare October 19, 2026 DOI: 10.34944/22br-ry12 (opens in new tab)
Study at a glance
AI-extracted from the abstract| Characteristics | Preclinical experimental study (in vivo animal dosing and in vitro incubations) Peer reviewed |
|---|---|
| Population | Rats (in vivo dosing) and human liver microsome preparations (in vitro) |
| Interventions | ortho-methyl fentanyl fentanyl N N-dimethyltryptamine (DMT) medetomidine dexmedetomidine levomedetomidine xylazine |
| Topics | 5-MeO-DMT DMT |
| Key findings | Reports the first in vivo characterization of ortho-methyl fentanyl, showing approximately three-fold lower potency than fentanyl in rats. Poly-drug human liver microsome incubations showed altered rates of fentanyl conversion to norfentanyl when ortho-methyl fentanyl or medetomidine were added separately, suggesting potential competitive metabolism, whereas xylazine did not significantly change fentanyl metabolism. DMT metabolites were identified and quantified, with intranasal DMT effects comparable to human studies. |
Abstract
Forensic toxicology relies heavily on foundational analytical chemistry techniques for the identification and characterization of drugs and toxins, and this work specifically focused on novel psychoactive substances (NPS). NPS are drugs that have newly emerged on the illicit or recreational drug market for use that circumvents legislation (i.e., “legal highs”) and cause significant risk to drug users with little knowledge of the activity, potency, or pharmacokinetics of the NPS. This research sought to analytically characterize two NPS (ortho-methyl fentanyl, N,N-dimethyltryptamine) and one adulterant (medetomidine) to allow for deeper understanding of these dangerous and newly emergent drugs. This research focused on in vivo, in vitro, and analytical characterization of these NPS using drug dosing with animal models, human liver microsome (HLM) incubations, and method development via liquid chromatography tandem quadrupole mass spectrometry (LC-QQQ-MS), respectively. In Chapters 4 and 5, N,N-dimethyltryptamine (DMT) was evaluated via in vivo rat dosing using intranasal (IN) and subcutaneous (SC) routes of administration as well as in vitro metabolite generation and elucidation via HLM incubation. It was determined through these analyses that DMT and its metabolites could be identified, elucidated, and quantitated using HLM and LC-QQQ-MS with pharmacodynamic comparison of IN DMT comparable to studies reported in humans. Additional metabolites (oxidation, di-oxidation) not previously confirmed for DMT were identified and characterized but were unable to be confirmed due to lack of standard reference material. In Chapters 6 and 7, medetomidine was successfully characterized via LC-QQQ-MS and HLM incubation via liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QTOF-MS). Medetomidine enantiomers, dexmedetomidine and levomedetomidine, were successfully chromatographically resolved to allow for confirmation of illicit medetomidine not being diverted from human use (i.e., illicit use from veterinary or clandestine origins). Few biotransformations were identified for medetomidine (e.g., oxidated metabolites) and comparison of metabolism of medetomidine and its enantiomers showed significant differences in rate of metabolism with dexmedetomidine showing the quickest clearance using in vitro methods. In Chapters 8 and 9, ortho-methyl fentanyl was evaluated using in vivo rat dosing with subcutaneous administration in comparison to fentanyl and the first reported metabolic comparison of single-drug versus poly-drug use. Through this research, the first in vivo characterization of ortho-methyl fentanyl is reported, showing an approximate three-fold decrease in potency in comparison to fentanyl. Through poly-drug HLM incubations, a significant difference in the rate of fentanyl conversion to norfentanyl was identified with the addition of ortho-methyl fentanyl and medetomidine separately, showing potential competitive metabolism. The addition of xylazine did not significantly alter fentanyl metabolism. The constant identification of new NPS on the illicit drug market causes significant challenges for forensic toxicologists; however, as shown through this research, new techniques utilizing analytical chemistry methodology can allow for a more complete and deeper understanding of NPS risks, activity, potency, pharmacodynamics, and pharmacokinetics. While this research is not often completed for NPS, it is necessary to characterize these new drugs to understand the potential risks to public health and safety a new uncontrolled drug can cause on drug using populations.
Comparable studies
Other preclinical and animal studies on 5-MeO-DMT, most cited first.
| Study | Year | Design | Participants |
|---|---|---|---|
| Psychedelic N,N-dimethyltryptamine and 5-methoxy-N,N-dimethyltryptamine modulate innate and adaptive inflammatory responses through the sigma-1 receptor of human monocyte-derived dendritic cells. Human primary monocyte-derived dendritic cells | 2014 | In vitro study | |
| Effects of N,N-Dimethyltryptamine on Rat Behaviors Relevant to Anxiety and Depression Adult male Sprague-Dawley rats | 2018 | Experimental study | |
| Hallucinogen-like actions of 5-methoxy-N,N-diisopropyltryptamine in mice and rats. Mice and rats | 2006 | Preclinical study | |
| Agonist Properties of N,N-Dimethyltryptamine at Serotonin 5-HT2A and 5-HT2C Receptors Rats and transfected fibroblasts | 1998 | Experimental study | |
| Subtypes of the 5-HT receptor mediating the behavioural responses to 5-methoxy-N,N-dimethyltryptamine in the rat Intact rats and rats depleted of monoamines with reserpine | 1985 | Experimental study |