Ketamine-derived designer drug methoxetamine: metabolism including isoenzyme kinetics and toxicological detectability using GC-MS and LC-(HR-)MSn.

Methoxetamine (MXE; 2-(3-methoxyphenyl)-2-(N-ethylamino)-cyclohexanone), a ketamine analog, is a new designer drug and synthesized for its longer lasting and favorable pharmacological effects over ketamine. The aims of the presented study were to identify the phases I and II metabolites of MXE in rat and human urine by GC-MS and LC-high-resolution (HR)-MS(n) and to evaluate their detectability by GC-MS and LC-MS(n) using authors' standard urine screening approaches (SUSAs). Furthermore, human cytochrome P450 (CYP) enzymes were identified to be involved in the initial metabolic steps of MXE in vitro, and respective enzyme kinetic studies using the metabolite formation and substrate depletion approach were conducted. Finally, human urine samples from forensic cases, where the ingestion of MXE was suspected, were analyzed. Eight metabolites were identified in rat and different human urines allowing postulation of the following metabolic pathways: N-deethylation, O-demethylation, hydroxylation, and combinations as well as glucuronidation or sulfation. The enzyme kinetic studies showed that the initial metabolic step in humans, the N-deethylation, was catalyzed by CYP2B6 and CYP3A4. Both SUSAs using GC-MS or LC-MS(n) allowed monitoring an MXE intake in urine.

Liquid chromatography-tandem mass spectrometry screening method using information-dependent acquisition of enhanced product ion mass spectra for synthetic cannabinoids including metabolites in urine.

The total number of synthetic cannabinoids (SCs) - a group of new psychoactive substances (NPS) - is increasing every year. The rapidly changing market demands the latest analytical methods to detect the consumption of SCs in clinical or forensic toxicology. In addition, SC metabolites must also be included in a screening procedure, if detection in urine is asked for. For that purpose, an easy and fast qualitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) urine screening method for the detection of 75 SCs and their metabolites was developed and validated in terms of matrix effects, recovery, and limits of identification for a selection of analytes. SC metabolites were generated using in vitro human liver microsome assays, identified by liquid chromatography-high resolution tandem mass spectrometry (LC-HRMS/MS) and finally included to the MS/MS spectra in-house library. Sample preparation was performed using a cheap-and-easy salting-out liquid-liquid extraction (SALLE) after enzymatic hydrolysis. Method validation showed good selectivity, limits of identification down to 0.05 ng/mL, recoveries above 80%, and matrix effects within ±25% for the selected analytes. Applicability of the method was demonstrated by detection of SC metabolites in authentic urine samples.

In vitro metabolism of the synthetic cannabinoids CUMYL-PINACA, 5F-CUMYL-PINACA, CUMYL-4CN-BINACA, 5F-CUMYL-P7AICA and CUMYL-4CN-B7AICA.

Synthetic cannabinoid consumption trends underlie fast changes and provide several challenges to clinical and forensic toxicologists. Due to their extensive metabolism, parent compounds are hardly detectable in urine. Therefore, knowledge of the metabolism of synthetic cannabinoids is essential to allow their detection in biological matrices. The aim of the present study was the elucidation of the metabolism of CUMYL-PINACA, 5F-CUMYL-PINACA, CUMYL-4CN-BINACA, 5F-CUMYL-P7AICA, and CUMYL-4CN-B7AICA with a focus on the analytical and interpretational differentiation of the compounds. Microsomal assay mixtures containing co-substrates, 10 µg/mL substrate and 1 mg/mL pooled human liver microsomes were incubated for 1 hour at 37°C. Investigation of the metabolites was performed on a Thermo Fischer Ultimate 3000 UHPLC system coupled to a Sciex 6600 QTOF System. Hydroxylation was observed to be a major biotransformation step for all 5 cumyl-derivatives, followed by dihydroxylation. For CUMYL-PINACA, a major metabolic pathway was hydroxylation at the pentyl moiety, followed by a second hydroxylation at that pentyl moiety or oxidation to ketone. A major metabolic pathway for the compounds containing a nitrile function was nitrile hydrolysis followed by carboxylation and further hydroxylation. For the fluorinated compounds, oxidative defluorination and carboxylation were abundant metabolic steps. Some of the metabolic transformations lead to structurally identical metabolites, which should not be used as marker for the intake of a particular parent compound. In addition, several constitutional isomers containing either an indazole or azaindole core structure were detected, which should be differentiated by retention time rather than by their mass spectra alone.

Structural characterization of the new synthetic cannabinoids CUMYL-PINACA, 5F-CUMYL-PINACA, CUMYL-4CN-BINACA, 5F-CUMYL-P7AICA and CUMYL-4CN-B7AICA.

Synthetic cannabinoids are a group of new psychoactive compounds (NPS) that act as agonists at the cannabinoid receptor. First reported in 2008, they currently represent one of the largest groups of NPS that are monitored by the European Monitoring Centre for Drugs and Drug Addiction (EMCDDA). Five samples (4 from the European RESPONSE project and one from daily casework) containing different synthetic cannabinoids were analyzed by a complex of analytical methods including gas chromatography-electron ionization mass spectrometry (GC-EI-MS), liquid chromatography-high resolution mass spectrometry (LC-HRMS), infrared spectroscopy (IR) and nuclear magnetic resonance spectroscopy (NMR). Five new synthetic cannabinoids containing a cumyl moiety as a linked group were identified: CUMYL-PINACA, 5F-CUMYL-PINACA, CUMYL-4CN-BINACA, 5F-CUMYL-P7AICA, CUMYL-4CN-B7AICA. 5F-CUMYL-PINACA and 5F-CUMYL-P7AICA as well as CUMYL-4CN-BINACA and CUMYL-4CN-B7AICA are constitutional isomers and only differ in the position of a nitrogen atom. The article contains all analytical data for a proper identification and differentiation of the five cumyl compounds.

Validation of an ion mobility spectrometry (IMS) method for the detection of heroin and cocaine on incriminated material.

The validation of a qualitative ion mobility spectrometry (IMS) procedure for the detection of trace amounts of heroin and cocaine on incriminated material using a vacuum cleaner for sampling is presented. The limit of detection, the limit of decision, selectivity and robustness were determined. As an approach, robustness was determined using ionizational interferences and matrix effects. By using this simple sampling procedure, a positive result for incriminated clothes needs a contamination of 250ng cocaine and 1000ng heroin, respectively.