In vitro Characterization of NPS Metabolites Produced by Human Liver Microsomes and the HepaRG Cell Line Using Liquid Chromatographyhigh Resolution Mass Spectrometry (LC-HRMS) Analysis: Application to Furanyl Fentanyl

Abstract

HLMs represent an acceptable first choice tool for a single NPS metabolism study in forensic laboratories. Dihydrodiol-Fu-F and despropionylfentanyl could be proposed as reliable metabolites to be recorded in HRMS libraries in order to improve detection of Fu-F users. Nevertheless, additional verifications of in vivo data remain necessary to confirm relevant blood and urinary metabolites of Fu-F.

In vitro metabolism of the novel synthetic opioid agonist cyclopropylfentanyl and subsequent confirmation in authentic human samples using liquid chromatography‐high resolution mass spectrometry

“…Metabolites were identified in a two‐step investigative process; first with FS accurate mass analysis of the in vitro incubate and data mining for postulated metabolites, followed by targeted MS 2 analysis for any metabolites identified. Examination of the metabolism of fentanyl and other fentanyl‐analogues allowed the postulation of potential metabolites of CycP‐F . The possible pathways included N ‐dealkylation, amide hydrolysis, hydroxylation and potential sequential transformations.…”

Detection of the designer benzodiazepine flunitrazolam in urine and preliminary data on its metabolism

“…In order to confirm the purity and the amount of the FNTZ tablet (0.25 mg), a liquid chromatography high‐resolution mass spectrometry detection (LC–HRMS) analysis (see section 2.6) was complemented by nuclear magnetic resonance (NMR) analysis, according to previous applications . Briefly, the NMR spectra were recorded on AVANCE 300 (Bruker Biospin, France) operating at 300 MHz equipped with a 5‐mm quadruple nucleus probe (QNP) probe at 295 K. The 1H spectra were recorded with 64 scans, a 32‐K time‐domain data points with 4800 Hz spectral width, an acquisition time of 3.42 seconds, a relaxation delay of 2 seconds and a flip angle of 30°.…”

“…The formation of FNTZ metabolites were investigated by in vitro incubation with human liver (HLMs) and retrospective analysis of urine samples by LC–HRMS using a method previously reported . Once thawed, HLMs (2 mg protein/mL for a final volume of 100 μL) were pre‐activated by alamethicin (at 0.1 mmol/L) on ice in an intermediate volume of 50 μL in 0.1 M Tris–HCl‐MgCl 2 (10mM MgCl 2 and 100mM Tris–HCl solution) at pH 7.4.…”

“…In order to confirm the purity and the amount of the FNTZ tablet (0.25 mg), a liquid chromatography high-resolution mass spectrometry detection (LC-HRMS) analysis (see section 2.6) was complemented by nuclear magnetic resonance (NMR) analysis, according to previous applications. 11,12 Briefly, the NMR spectra were recorded on AVANCE Linearity was observed for FNTZ concentrations ranging from 0.1 to 100 ng/mL, with a correlation coefficient of r 2 = 0.992. QC samples (1 and 10 ng/mL), analyzed in duplicate in ten independent experimental assays, were used for determination a coefficient of variation for precisions and accuracy.…”

“…The formation of FNTZ metabolites were investigated by in vitro incubation with human liver (HLMs) and retrospective analysis of urine samples by LC-HRMS using a method previously reported. 11,12 Once After oral administration of one FNTZ tablet, the overall behavior of the subject was not modified. A slight sensation of weakness was noticed (feeling that it was time to go to bed for sleeping), but no physical or behavioral impairment was observed.…”