Methoxylated Modification of Glutathione-Mediated Metabolism of Halobenzoquinones In Vivo and In Vitro

Xenobiotics were generally detoxified in organisms through interaction with endogenous molecules, which may also generate metabolites of increased toxicity. Halobenzoquinones (HBQs), a group of highly toxic emerging disinfection byproducts (DBPs), can be metabolized by reacting with glutathione (GSH) to form various glutathionylated conjugates (SG-HBQs). In this study, the cytotoxicity of HBQs in CHO-K1 cells showed a wavy curve as a function of increased GSH dosage, which was inconsistent with the commonly recognized progressive detoxification curve. We hypothesized that the formation and cytotoxicity of GSH-mediated HBQ metabolites contribute to the unusual wave-shaped cytotoxicity curve. Results showed that glutathionyl-methoxyl HBQs (SG-MeO-HBQs) were identified to be the primary metabolites significantly correlated with the unusual cytotoxicity variation of HBQs. The formation pathway was initiated by stepwise metabolism via hydroxylation and glutathionylation to produce detoxified hydroxyl HBQs (OH-HBQs) and SG-HBQs, followed by methylation to generate SG-MeO-HBQs of potentiated toxicity. To further verify the occurrence of the aforementioned metabolism in vivo, SG-HBQs and SG-MeO-HBQs were detected in the liver, kidney, spleen, testis, bladder, and feces of HBQ-exposed mice, with the highest concentration quantified in the liver. The present study supported that the co-occurrence of metabolism can be antagonistic, which enhanced our understanding of the toxicity and metabolic mechanism of HBQs.