Toxicity and behavioural effects of ocfentanil and 2-furanylfentanyl in zebrafish larvae and mice

The introduction of the so-called New Psychoactive Substances represents a problem of global concern due to several factors, including multiplicity of structures, poorly known activity, short half-life in the market, lack of pure standards etc. Among these problems, of the highest relevance is also the lack of information about metabolism and adverse effects, which must be faced using simple and low-cost animal models. On these grounds, the present work has been carried out on 5 days post fertilization zebrafish (Danio rerio) larvae in comparison with adult mice (Mus musculus). Ocfentanil and 2-furanylfentanyl were administered at different concentrations to zebrafish larvae (1, 10 µM) and mice (0.1, 1, 6, 15 mg/kg). The behavioural assay showed a decrease in basal locomotor activity in zebrafish, whereas in mice this effect was evident only after the mechanical stimulus. Larva extracts and mice urine were analysed by using liquid chromatography coupled to high resolution mass spectrometry to identify the metabolic pathways of the fentanyl analogs. For 2-furanylfentanyl, the most common biotransformations observed were hydroxylation, hydration and oxidation in zebrafish larvae, whereas mice produced mainly the dihydrodiol metabolite. Hydroxylation was the major route of metabolism for ocfentanil in zebrafish larvae, while in mice the O-demethylated derivative was the main metabolite. In addition, a study was conducted to evaluate morphological effects of the two drugs on zebrafish larvae. Malformations were noticeable only at the highest concentration of 2-furanylfentanyl, whereas no significant damage was observed with ocfentanil. In conclusion, the two animal models show similarities in behavioral response and in metabolism, considering the different biological investigated.