Metabolomics Study with Gas Chromatography-Mass Spectrometry of Hepatotoxicity on Adult Zebrafish (Brachydanio Rerio) after Exposure to Diquat

Although diquat is a widely used water-soluble herbicide in the world, its toxicity to freshwater fish has not been well characterized. In this study, gas chromatography-mass spectrometry (GC-MS) based metabolomics approach combined with histopathological examination and biochemical assays was applied to comprehensively assess the hepatotoxicity in zebrafish (Brachydanio rerio) after diquat exposure at two dosages of 0.34 and 1.69 mg·L−1 for 35 days. The results indicated that 1.69 mg·L−1 diquat exposure cause serious cellular swell and vacuolization with increased nuclear abnormality, and lead to obvious disturbance of antioxidative system and dysfunction in liver; while no obvious pathological injury could be detected, and changes in liver biochemistry were less pronounced at the dose level of 0.34 mg·L−1. Multivariate statistical analysis and pattern recognition showed different GC-MS profiles of zebrafish liver following exposure to diquat, the cluster of the treated groups were both clearly separated from the control samples. The differentially abundant metabolites mainly include carbohydrates, amino acids, lipids, nucleotides, and their derivatives. In the exposure group of 1.69 mg·L−1 diquat, severe disturbances of amino acid metabolism played important biological roles associated with inhibition of energy metabolism, reduced immunity, and disorders in neurotransmitters as pathway analysis revealed. Additionally, fluctuation of inositol, creatine, and pantothenic acid, substances associated with stress regulation and signal transduction, participating in metabolic abnormalities in zebrafish with diquat-triggered hepatic damage. Energy metabolism of zebrafish exposed on 0.34 mg·L−1 diquat more inclined to rely on anaerobic glycolysis than the normal ones. Amino acid metabolism responses were less affected, but obvious interference effects on lipid metabolism were observed with 0.34 mg·L−1 diquat exposure. These results imply increased sensitivity of metabolomics versus histopathology and clinical chemistry in recognizing liver toxicity of diquat. This study will contribute to explore possible mechanism of hepatic damages on nontarget freshwater fish induced by diquat and provide important basis for its environmental risk assessment.