# Study on Phase I Metabolic Processes and Metabolite Biomarker Identification of Synthetic Cannabinoids 5F-ADB-PINACA and 5F-ADBICA in Human Liver Microsomes and Zebrafish Model

## TL;DR

This study explores how two synthetic cannabinoids are metabolized in human and zebrafish models, identifying key metabolite biomarkers and differences in their metabolic processes.

## Contribution

The first use of correlation analysis in temporal metabolic profiling of synthetic cannabinoids and the first in vitro and in vivo detection of 5F-ADBICA metabolites.

## Key findings

- M13 is a potential biomarker for 5F-ADB-PINACA metabolism via hydrolytic defluorination.
- M19 is a potential biomarker for 5F-ADBICA metabolism via hydrolytic defluorination.
- Structural differences between the two cannabinoids lead to distinct metabolic pathways despite similar reactions.

## Abstract

Synthetic cannabinoids (SCs) are a rapidly developing kind of novel psychoactive substance, frequently associated with acute intoxication and public health concerns. This study aimed to elucidate and compare the phase I metabolic pathways of two structurally related SCs, 5F-ADB-PINACA and 5F-ADBICA, using in vitro and in vivo models. Temporal metabolic profiling was performed to identify potential signature metabolites. Temporal abundance patterns and correlation cluster analysis of metabolites were analyzed to determine metabolite biomarkers. The two SCs were incubated with pooled human liver microsomes for 24 h and were also evaluated in vivo in zebrafish. Metabolite profiles were characterized using UHPLC-QE Orbitrap-MS. HLM analysis identified 21 5F-ADB-PINACA metabolites and 28 5F-ADBICA metabolites. Metabolites of 5F-ADBICA were detected for the first time in vitro and in a zebrafish model. Zebrafish studies confirmed the presence of all key metabolites observed in HLM. Comparative analysis of their metabolic pathways revealed differences in metabolism driven by structural differences between the indazole and indole cores. This is the first time that correlation analysis has been used in the temporal metabolic profiling of SCs. This study comprehensively characterized the metabolism of 5F-ADB-PINACA and 5F-ADBICA, identifying M13 (hydrolytic defluorination) as a potential metabolite biomarker for 5F-ADB-PINACA and M19 (hydrolytic defluorination) as a potential metabolite biomarker for 5F-ADBICA. The metabolic reactions of the main metabolites of the two synthetic cannabinoids are consistent. However, their metabolic processes (i.e., the overall metabolic pathways and temporal progression of these reactions) are different, which illustrates the metabolic similarity of structurally similar synthetic cannabinoids and the impact of different structures on the metabolic processes.