title: Characterization and in vitro phase I microsomal metabolism of designer benzodiazepines - an update comprising adinazolam, cloniprazepam, fonazepam, 3-hydroxyphenazepam, metizolam and nitrazolam.
abstract: Designer benzodiazepines represent an emerging class of new psychoactive substances. While other classes of new psychoactive substances such as cannabinoid receptor agonists and designer stimulants are mainly consumed for hedonistic reasons, designer benzodiazepines may also be consumed as 'self-medication' by persons suffering from anxiety or other psychiatric disorders or as stand-by 'antidote' by users of stimulant and hallucinogenic drugs. In the present study, five benzodiazepines (adinazolam, cloniprazepam, fonazepam, 3-hydroxyphenazepam and nitrazolam) and one thienodiazepine (metizolam) offered as 'research chemicals' on the Internet were characterized and their main in vitro phase I metabolites tentatively identified after incubation with pooled human liver microsomes. For all compounds, the structural formula declared by the vendor was confirmed by nuclear magnetic resonance spectroscopy, gas chromatography-mass spectrometry (MS), liquid chromatography MS/MS and liquid chromatography quadrupole time-of-flight MS analysis. The detected in vitro phase I metabolites of adinazolam were N-desmethyladinazolam and N-didesmethyladinazolam. Metizolam showed a similar metabolism to other thienodiazepines comprising monohydroxylations and dihydroxylation. Cloniprazepam was metabolized to numerous metabolites with the main metabolic steps being N-dealkylation, hydroxylation and reduction of the nitro function. It has to be noted that clonazepam is a metabolite of cloniprazepam, which may lead to difficulties when interpreting analytical findings. Nitrazolam and fonazepam both underwent monohydroxylation and reduction of the nitro function. In the case of 3-OH-phenazepam, no in vitro phase I metabolites were detected. Formation of licensed benzodiazepines (clonazepam after uptake of cloniprazepam) and the sale of metabolites of prescribed benzodiazepines (fonazepam, identical to norflunitrazepam, and 3-hydroxyphenazepam) present the risk of incorrect interpretation of analytical findings. Copyright © 2016 John Wiley & Sons, Ltd.
authors.affiliation: Institute of Forensic Medicine, Forensic Toxicology, Medical Center - University of Freiburg, Faculty of Medicine, University of Freiburg, Albertstr. 9, 79104, Freiburg, Germany. bjoern.moosmann@uniklinik-freiburg.de.
authors.forename: Bjoern
authors.initials: B
authors.lastname: Moosmann
authors.affiliation: Institute of Pharmaceutical Sciences, University of Freiburg, Albertstr. 25, 79104, Freiburg, Germany.
authors.forename: Philippe
authors.initials: P
authors.lastname: Bisel
authors.affiliation: Institute of Forensic Medicine, Forensic Toxicology, Medical Center - University of Freiburg, Faculty of Medicine, University of Freiburg, Albertstr. 9, 79104, Freiburg, Germany.|Hermann Staudinger Graduate School, University of Freiburg, Hebelstr. 27, 79104, Freiburg, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Franz
authors.affiliation: Institute of Forensic Medicine, Forensic Toxicology, Medical Center - University of Freiburg, Faculty of Medicine, University of Freiburg, Albertstr. 9, 79104, Freiburg, Germany.
authors.forename: Laura M
authors.initials: LM
authors.lastname: Huppertz
authors.affiliation: Institute of Forensic Medicine, Forensic Toxicology, Medical Center - University of Freiburg, Faculty of Medicine, University of Freiburg, Albertstr. 9, 79104, Freiburg, Germany.
authors.forename: Volker
authors.initials: V
authors.lastname: Auwärter
mesh_terms: D001569:Benzodiazepines / Q000737:chemistry / Q000378:metabolism*; D000697:Central Nervous System Stimulants / Q000737:chemistry / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D015198:Designer Drugs / Q000737:chemistry / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D006213:Hallucinogens / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D050216:Metabolic Detoxication, Phase I; D008862:Microsomes, Liver / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
chemical_list: D000697:Central Nervous System Stimulants; D015198:Designer Drugs; D006213:Hallucinogens; D001569:Benzodiazepines
keywords: LC-MS/MS; flunitrazepam; human liver microsomes; legal highs; new psychoactive substances