Comparative Toxicity of 6PPD and 6PPD-quinone on the Gut-Liver Axis in Zebrafish: Insights into Microbiota Dysbiosis and Oxidative Stress

The ubiquitous presence of the tire rubber antioxidant 6PPD and its toxic transformation product, 6PPD-quinone (6PPDQ), poses emerging risks to aquatic ecosystems. This study evaluated the comparative hepatotoxicity and enterotoxicity of environmentally relevant concentrations of 6PPD and 6PPDQ in adult zebrafish ( Danio rerio ), with a specific focus on the gut-liver axis. Histopathological analysis revealed that both compounds induced dose-dependent liver damage, characterized by hepatocyte vacuolation and nuclear degeneration; notably, 6PPDQ elicited distinct, and in some aspects comparable, tissue injury relative to the parent compound. Biochemical assays confirmed that oxidative stress is a key mechanism of toxicity, evidenced by elevated malondialdehyde (MDA) levels (increased by up to 62.2%) and glutathione peroxidase (GSH-Px) activity (increased by up to 257.1%), alongside inhibited superoxide dismutase (SOD) and catalase (CAT) activities (reduced by up to 15.2% and 25.3%, respectively). In the intestine, exposure compromised barrier integrity and triggered inflammation, indicated by villus structural damage and the upregulation of NF-κB p65 expression. 16S rRNA sequencing unveiled distinct dysbiosis patterns: 6PPD exposure reduced the abundance of Firmicutes and enriched Proteobacteria, whereas 6PPDQ significantly altered the abundances of Bacteroidetes and Proteobacteria. Functional prediction analysis indicated hypothetical functional shifts, suggesting that 6PPDQ may specifically impair immune-related pathways while enhancing energy metabolism. Collectively, these findings demonstrate that 6PPD and 6PPDQ induce concurrent multi-organ toxicity via the gut-liver axis, with the transformation product 6PPDQ presenting a distinct ecological risk profile. This study highlights the critical need to incorporate transformation products into the risk assessment of tire-derived contaminants.