Characterization of extensive 2-fluorodeschloroketamine metabolism in pooled human liver microsomes, urine and hair from an addicted patient using high-resolution accurate mass spectrometry.

2-Fluorodeschloroketamine (2F-DCK) is a ketamine derivative involved in acute intoxications and deaths. The aim of this study is to investigate its metabolism using pooled human liver microsomes (pHLMs) and to apply it to authentic samples (urine, hair and seized materials) from a drug user. 2F-DCK (100 µM) incubates with pHLMs were analyzed by liquid chromatography-high-resolution accurate mass (LC-HRAM; Q-Exactive, Thermo Fisher Scientific®) according to a previously published protocol. Spectra annotation was performed using Compound Discoverer® software and the metabolic scheme was drawn using ChemDraw software. Urine (200 µL) and hair (previously decontaminated using dichloromethane and segmented into three segments: A, 0-3 cm; B, 3-6 cm; C, 6-9 cm) were extracted with a mixture of hexane:ethyl acetate (1:1) and chloroform:isopropanol (4:1). About 10 µL of both reconstituted residues were analyzed by LC-HRAM. Hair was also analyzed by LC-MS-MS (TSQ Vantage, Thermo Fisher Scientific®) for 2F-DCK and deschloroketamine (DCK) quantification. The two presumed 2F-DCK crystals consumed by the patient were dissolved in methanol (1 mg/mL) and 10 µL were analyzed by LC-MS-MS (Quantum Access Max, Thermo Fisher Scientific®). Twenty-six putative 2F-DCK metabolites were identified, 15 being reported for the first time. Thirteen metabolites were detected in pHLMs, 10 confirmed in both the patient's urine and hair and all were found in at least one of the two samples. Twenty-three metabolites were detected in urine and 20 in hair. Our research confirms the reliability of nor-2F-DCK as a target analyte and suggests OH-dihydro-nor-2F-DCK and dehydro-nor-2F-DCK as new target analytes in urine and hair, respectively. This is the first study to report DCK as a 2F-DCK metabolite using pHLMs and to determine its concentrations in hair (A/B/C, 885/1,500/1,850 pg/mg) following chronic use. Finally, the two seized crystals contained 2F-DCK at 67% and 96% with traces of DCK (0.4% and 0.6%) related to cross-contamination by container exchange.

Metabolic profiling of deschloro-N-ethyl-ketamine and identification of new target metabolites in urine and hair using human liver microsomes and high-resolution accurate mass spectrometry.

The aim of this study was to identify new markers of deschloro-N-ethyl-ketamine (O-PCE), a ketamine analogue that has been involved in acute intoxications with severe outcomes including death and whose metabolism has never been studied before. In vitro study after 2-h incubation with pooled human liver microsomes (HLMs) cross-checked by the analysis of urine and hair from a 43-year-old O-PCE user (male) were performed by liquid chromatography-high resolution mass spectrometry (LC-HRMS). Acquired data were processed by the Compound Discoverer® software, and a full metabolic profile of O-PCE was proposed. In total, 15 metabolites were identified, 10 were detected in vitro (HLMs) and confirmed in vivo (urine and/or hair), two were present only in HLMs, and the remaining three metabolites were identified only in biological specimens. While O-PCE was no longer detected in urine, nine metabolites were identified allowing to increase its detection window. In descending order of metabolites abundance, we suggest using 2-en-PCA-N-Glu (34%, first), M3 (16%, second), O-PCA-N-Glu (15.4%, third), OH-O-PCE (15%, fourth) and OH-PCE (11.9%, fifth) as target metabolites to increase the detection window of O-PCE in urine. In hair, nine metabolites were identified. OH-PCA was the major compound (78%) with a relevant metabolite to parent drug ratio (=6) showing its good integration into hair and making it the best marker for long-term monitoring of O-PCE exposure.