Antioxidant gene expression and metabolic responses of earthworms (Eisenia fetida) after exposure to various concentrations of hexabromocyclododecane

Highlights: • The earthworm were exposed to HBCD at sub-lethal concentrations. • Up-regulation of SOD gene expression level for earthworm exposed to HBCD was detected. • Metabolites of ATP, valine, lysine, glycine, betaine and lactate was increased. • Metabolomics(NMR) is more sensitive than gene expression for assessing HBCD toxicity. Hexabromocyclododecane (HBCD), a ubiquitous suspected contaminant, is one of the world's most prominent brominated flame retardants (BFRs). In the present study, earthworms (Eisenia fetida) were exposed to HBCD. The expression of selected antioxidant enzyme genes was measured, and the metabolic responses were assessed using nuclear magnetic resonance (NMR) to identify the molecular mechanism of the antioxidant stress reaction and the metabolic reactions of earthworms to HBCD. A significant up-regulation (p < 0.05) of superoxide dismutase (SOD) gene expression was detected, with the highest gene expression level of SOD appearing at a dose of 400 mg kg⁻¹ dw (2.06-fold, p < 0.01). However, the glutathione transferase (GST) gene expression levels did not differ significantly (p > 0.05). Principal component analysis (PCA) of the metabolic responses showed that all groups could be clearly differentiated, and the highest concentration dose group was the most distant from the control group. Except for fumarate, the measured metabolites, which included adenosine triphosphate (ATP), valine, lysine, glycine, betaine and lactate, revealed significant (p < 0.05) increases after 14 days of exposure to HBCD. HBCD likely induces high levels of anaerobic respiration, which would result in high levels of ATP and lead to the disintegration of proteins into amino acids, including valine and lysine, to produce energy. The observed changes in osmotic pressure were indicative of damage to the membrane structure. Furthermore, this study showed that NMR-based metabolomics was a more sensitive tool than measuring the gene expression levels for elucidating the mode of toxicity of HBCD in earthworm exposure studies.