Metabolism of synthetic cathinones through the zebrafish water tank model: a promising tool for forensic toxicology laboratories

Abstract

The aim of this study was to identify in vivo phase I metabolites of five psychoactive substances: N-ethylpentylone, ethylone, methylone, α-PVP and 4-CDC, using the in house developed experimental setup zebrafish (Danio rerio) water tank (ZWT). High-resolution mass spectrometry allowed for metabolite identification. A pilot study of reference standard collection of N-ethylpentylone from the water tank was conducted. Methods ZWT consisted in 8 fish placed in a 200 mL recipient-containing water for a single cathinone. Experiments were performed in triplicate. Water tank samples were collected after 8 h and pretreated through solid-phase extraction. Separation and accurate-mass spectra of metabolites were obtained using liquid chromatography-high resolution tandem mass spectrometry. Results Phase I metabolites of α-PVP were identified, which were formed involving ketone reduction, hydroxylation, and 2″-oxo-pyrrolidine formation. The lactam derivative was the major metabolite observed for α-PVP in ZWT. N-Ethylpentylone and ethylone were transformed into phase I metabolites involving reduction, hydroxylation, and dealkylation. 4-CDC was transformed into phase I metabolites, reported for the first time, involving N-dealkylation, N,N-bis-dealkylation and reduction of the ketone group, the last one being the most intense after 8 h of the experiment. Conclusions ZWT model indicated to be very useful to study the metabolism of the synthetic cathinones, such as N-ethylpentylone, ethylone, α-PVP and 4-CDC. Methylone seems to be a potent CYP450 inhibitor in zebrafish. More experiments are needed to better evaluate this issue. Finally, this approach was quite simple, straightforward, extremely low cost, and fast for "human-like" metabolic studies of synthetic cathinones. Keywords Synthetic cathinones • Zebrafish • In vivo metabolism • High-resolution mass spectrometry • Human-like experimental model • Reference standard collection in zebrafish water tank Electronic supplementary material The online version of this article (

Etazene induces developmental toxicity in vivo Danio rerio and in silico studies of new synthetic opioid derivative

“…The D. rerio model has been frequently used in studies of NPS, including behavioral assessment of opioid addiction, anxiety, memory, and toxicological and pharmacological targets 7 . Recently, the D. rerio model has been successfully used to identify metabolites for various NPS groups, including cannabinoids 8 , opioids 9 , and cathinones 10 . Comparing metabolic processes of D. rerio to humans, consistent results have been obtained, which may be the basis for replacing the existing in vitro methods, characterized by substantial limitations 11 .…”

Forensic Impact of the Omics Science Involved in the Wound: A Systematic Review

“…In this context, forensic molecular pathology includes applying omics sciences to investigate the genetic basis and the cause of death at the molecular biological level. Today many genomic investigations carry out an analysis of the genetic background (9)(10)(11), study the dynamics of gene expression (transcriptomics) is also playing an important role (12)(13)(14), as well as vital phenomena involving activated biological mediators and their degenerative products (proteomics) (15), and finally, the analysis of the different metabolites involved (metabolomics) (16)(17)(18).…”

The metabolism of valerylfentanyl using human liver microsomes and zebrafish larvae

“…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.…”

“…14,15 Additionally, they have opioid receptors similar to humans and have been used as a human disease model. [16][17][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.…”