Background: Metabolite profiling of novel psychoactive substances (NPS) is critical for
documenting drug consumption. N-(1-amino-3,3-dimethyl-1-oxobutan-2-yl)-1-(4-fluorobenzyl)-1Hindazole-
3-carboxamide (ADB-FUBINACA) is an emerging synthetic cannabinoid whose
toxicological and metabolic data are currently unavailable.
Methods: We aimed to determine optimal markers for identifying ADB-FUBINACA intake. Metabolic
stability was evaluated with human liver microsome incubations. Metabolites were identified after 1
and 3 h incubation with pooled human hepatocytes, liquid chromatography- high resolution mass
spectrometry in positive-ion mode (5600+ TripleTOF®, Sciex) and several data mining approaches
Results: Metabolite separation was achieved on an Ultra Biphenyl column (Restek®); full-scan TOFMS
and information-dependent acquisition MS/MS data were acquired. ADB-FUBINACA
microsomal half-life was 39.7 min, with a predicted hepatic clearance of 9.0 mL/min/kg and a 0.5
extraction ratio (intermediate-clearance drug). Twenty-three metabolites were identified. Major
metabolic pathways were alkyl and indazole hydroxylation, terminal amide hydrolysis, subsequent
glucuronide conjugations, and dehydrogenation.
Conclusion: We recommend ADB-FUBINACA hydroxyalkyl, hydroxydehydroalkyl and
hydroxylindazole metabolites as ADB-FUBINACA intake markers. N-dealkylated metabolites are
not specific ADB-FUBINACA metabolites and should not be used as definitive markers of
consumption. This is the first ADB-FUBINACA in vitro metabolism study; in vivo experiments
enabling pharmacokinetic and pharmacodynamics studies or urine from authentic clinical/forensic
cases are needed to confirm our results.