Hexabromobenzene (HBB) is a novel brominated flame retardant (BFR) with ample evidence of its ubiquitous existence in the aquatic ecosystems. However, to date, the toxicological effects of this BFR on fish have been inadequately researched. The present study was conducted, based on an in vivo model, to investigate HBB-induced biochemical changes in liver and gill tissues of Carassius auratus after medium-term exposure to different concentrations (10, 150, and 300 mg/kg) for 7, 14, and 25 days. Oxidative stress was evoked evidently for the prolonged exposure, demonstrated by significant inhibition in antioxidant enzymes activities including superoxide dismutase, catalase, glutathione reductase, glutathione peroxidase, and glutathione S-transferase, and a decrease in reduced glutathione level, as well as simultaneous elevation in malondialdehyde content. Moreover, Na+, K+-ATPase activity, and protein level were remarkably reduced in fish tissues. Based on the integrated biomarker response, the toxic potency in each treatment was distinguished, and the more severe stress was mainly noted with the increasing concentrations and the extending durations. It was also observed that liver exhibited more pronounced alterations in biochemical parameters than gill, probably indicating the vulnerability of liver to HBB-triggered oxidative stress. Taken together, the results of this study clearly showed that HBB was capable of inducing oxidative stress and inhibiting Na+, K+-ATPase activity in different tissues of C. auratus after medium-term exposure. © 2013 Wiley Periodicals, Inc. Environ Toxicol 29: 1460–1470, 2014.