Abstract
Toxicometabolomics, essentially applying metabolomics to toxicology of endogenous compounds such as drugs of abuse or new psychoactive substances (NPS), can be investigated by using different in vitro models and dedicated metabolomics techniques to enhance the number of relevant findings. The present study aimed to study the toxicometabolomics of the two NPS α-pyrrolidinobutiophenone (1-phenyl-2-(pyrrolidin-1-yl)butan-1-one, α-PBP) and α-pyrrolidinoheptaphenone (1-phenyl-2-(pyrrolidin-1-yl)heptan-1-one, α-PEP, PV8) in HepaRG cell line incubates. Evaluation was performed using reversed-phase and normal-phase liquid chromatography coupled with high-resolution mass spectrometry in positive and negative ionization mode, respectively, to analyze cells and cell media. Statistical evaluation was performed using one-way ANOVA, principal component discriminant function analysis, as well as hierarchical clustering. In general, the analysis of cells did not mainly reveal any features, but the parent compounds of the drugs of abuse. For α-PBP an increase in N-methylnicotinamide was found, which may indicate hepatotoxic potential of the substance. After analysis of cell media, significant features led to the identification of several metabolites of both compounds. Amino acid adducts with glycine and alanine were found, and these have not been described in any study before and are likely to appear in vivo. Additionally, significant changes in the metabolism of cholesterol were revealed after incubation with α-PEP. In summary, the application of metabolomics techniques after HepaRG cells exposure to NPS did not lead to an increased number of identified drug metabolites compared to previously published studies, but gave a wider perspective on the physiological effect of the investigated compounds on human liver cells.
Highlights
Several in vitro models have been developed and used for investigating the metabolism of drugs of abuse (DOA) and new psychoactive substances (NPS), and for xenobiotics in general (Manier et al 2018; Richter et al 2017a, b; Sinz 2012; Sinz and Kim 2006)
Concerning extract 1 after α-PEP incubation, any analysis using positive ionization led to two significant features, while negative ionization did not lead to any significant feature
Toxicometabolomics of the two NPS α-PBP and α-PEP after HepaRG cell line incubation were evaluated by analyzing the cells and the cell medium separately
Summary
Several in vitro models have been developed and used for investigating the metabolism of drugs of abuse (DOA) and new psychoactive substances (NPS), and for xenobiotics in general (Manier et al 2018; Richter et al 2017a, b; Sinz 2012; Sinz and Kim 2006). PHLM are limited to reactions catalyzed by membrane-bound enzymes such as CYP enzymes, FMOs, and uridine 5′-diphospho-glucuronosyltransferases (UGTs) Since these reactions are the most relevant ones in drug metabolism, pHLM are widely applied in such studies (Sinz and Lyubimov 2011). In pHLS9, an unseparated mixture of cytosol and microsomes, both phase I and phase II metabolites, are present, but it generally suffers from lower enzyme activities compared to isolated fractions (Brandon et al 2003; Zhang et al 2012) Immortalized cell lines such as HepaRG and HepG2 are often applied to avoid the high costs of primary human hepatocytes and to apply a model that is better suited to represent the physiology of liver cells (Sinz and Kim 2006)
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