Study of the in vitro and in vivo metabolism of a novel synthetic cannabinoid PX-2 in human liver microsomes and zebrafish

Synthetic cannabinoids are the second largest group of new psychoactive substances reported by the United Nations Office on Drugs and Crime in the last decade and case reports bring attention to its high potency effects and its severe toxicity, including fatalities. Moreover, synthetic cannabinoids are usually entirely metabolized and metabolic pathways for many new generation synthetic cannabinoids are still unknown. In this study, the metabolism of five third generation synthetic cannabinoids was evaluated using zebrafish (Danio rerio) larvae as 24-h in vivo model studied within 5 days after fertilization. The studied synthetic cannabinoids were MMB-CHMICA, ADB-CHMICA, ADB-CHMINACA, MDMB-CHMCZCA, and NNL-3, and the respective metabolites were identified by liquid chromatography-high resolution tandem mass spectrometry. Eleven, six, fourteen, eleven, and four metabolites were identified for MMB-CHMICA, ADB-CHMICA, ADB-CHMINACA, MDMB-CHMCZCA, and NNL-3, respectively, and metabolic pathways have been proposed.The use of zebrafish larvae, with a high degree of physiological and genetic homology to humans, is an emerging tool very useful for the identification of metabolic pathways of psychoactive substances. Results obtained in this study compared well with metabolites obtained previously for the same target molecules or structural analogous after in vitro incubation with human or rat hepatocytes. Thus, potential biomarkers for the evaluated compounds are the O-demethylated metabolite for MMB-CHMICA; the oxidative deamination to hydroxyl metabolite for ADB-CHMICA; hydroxyl metabolites at cyclohexylmethyl, tert-butyl, and indazole moieties for ADB-CHMINACA; hydroxyl metabolites at carbazole core, tert-butyl, or cyclohexylmethyl tail moieties for MDMB-CHMCZCA; and amide hydrolyzed, defluorinated, and dihydroxilated metabolite for NNL-3.

Section: Resultsmentioning

confidence: 59%

Section: Resultsmentioning

confidence: 99%

Section: Characterization Of Adb-chmica Metabolitesmentioning

confidence: 96%

Section: Characterization Of Adb-chmica Metabolitesmentioning

confidence: 99%

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Metabolism of third generation synthetic cannabinoids using zebrafish larvae

Morales-Noé

Esteve‐Turrillas

Armenta

2021

“…Additionally, they have opioid receptors similar to humans and have been used as a human disease model 16–18 . Studies have been performed to investigate metabolite markers for synthetic cannabinoids, 19 human performance‐enhancing drugs, 20,21 synthetic cathinones, 22 and opioids 23,24 using the zebrafish model. More recently, we utilized the zebrafish model as a single assay for the toxicity and metabolism of fentanyl and detected norfentanyl, β‐hydroxyfentanyl, and 4‐anilino‐ N ‐phenethylpiperidine (4‐ANPP) from 24 to 96 h postfertilization, 25 providing additional support for their ability to metabolize opioids.…”

Section: Introductionmentioning

confidence: 99%

The metabolism of valerylfentanyl using human liver microsomes and zebrafish larvae

Cooman

Hoover

Sauvé

et al. 2022

Valerylfentanyl, a novel synthetic opioid less potent than fentanyl, has been reported in biological samples, but there are limited studies on its pharmacokinetic properties.The goal of this study was to elucidate the metabolism of valerylfentanyl using an in vitro human liver microsome (HLM) model compared with an in vivo zebrafish model. Nineteen metabolites were detected with N-dealkylation-valeryl norfentanyl and hydroxylation as the major metabolic pathways. The major metabolites in HLMs were also detected in 30 day postfertilization zebrafish. An authentic liver specimen that tested positive for valerylfentanyl, among other opioids and stimulants, revealed the presence of a metabolite that shared transitions and retention time as the hydroxylated metabolite of valerylfentanyl but could not be confirmed without an authentic standard. 4-Anilino-N-phenethylpiperidine (4-ANPP), a common metabolite to other fentanyl analogs, was also detected. In this study, we elucidated the metabolic pathway of valerylfentanyl, confirmed two metabolites using standards, and demonstrated that the zebrafish model produced similar metabolites to the HLM model for opioids.

Quantitative determination and metabolic profiling of synthetic cathinone eutylone in vitro and in urine samples by liquid chromatography tandem quadrupole time‐of‐flight mass spectrometry

Yeh

Wang

2022

In Taiwan, synthetic cathinones are the most prevalent new psychoactive substances, and their use is growing continuously. Urine samples are currently analysed to determine drug abuse, but the metabolic profiles and metabolites of these compounds are not widely reported. Given that cases of eutylone abuse have been growing since 2020, this study established a method employing supported liquid extraction combined with liquid chromatography tandem quadrupole time‐of‐flight mass spectrometry to identify and quantify eutylone and its metabolites in urine samples. Method validation was performed, and eight authentic samples were analysed. Moreover, in vitro metabolism experiments were conducted, and metabolites were generated by incubating eutylone with human liver microsomes and cytosol. Metabolite characterisation was achieved by confirming the accurate mass and product ions in full MS/MS spectra. Five metabolites were identified in in vitro experiments; they resulted from eutylone N‐dealkylation, β‐ketone reduction, demethylenation, aliphatic hydroxylation and sequential demethylenation and O‐methylation. The metabolic profile was obtained evaluating the metabolites at different incubation times: Demethylenation occurred first, followed by N‐dealkylation, β‐ketone reduction and aliphatic hydroxylation. Three additional metabolites were identified in authentic samples. Based on in vitro and in vivo evidence, we propose that the demethylenation and O‐methylation metabolite, the β‐ketone reduction metabolite, and the β‐ketone reduction, demethylenation and O‐methylation metabolite are the most appropriate biomarkers of eutylone consumption. Using these markers can help expand the eutylone detection window and provide information for toxicology research.