It is well known that antioxidants and reactive oxygen species play an important role in carcinogenesis. In this sudy, we attempted to evaluate antioxidant enzyme activities and lipid peroxidation levels in cancerous bladder tissue and to determine their relationship with bacterial infection. Bacterial culture was made from all urine samples using Blood and Eosin Methylene Blue agars for checking the presence of bacterial infections. We measured thiobarbituric acid reactive substances (TBARs) and activities of xanthine oxidase (XO), superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), and catalase (CAT) in cancerous tissues of 25 bladder cancer patients, in noncancerous adjacent bladder tissues of 13 out of these 25 patients, and in control bladder tissues of 15 patients with a non-neoplastic genitourinary disease. TBARs levels increased and XO, SOD, GSH-PX, and CAT activities decreased significantly in cancerous bladder tissues. TBARS, XO, and SOD levels were not significantly different between noncancerous adjacent tissue and control bladder tissue. Statistically significantly lower GSH-PX and higher CAT activities were observed in noncancerous adjacent bladder tissue compared with cancerous tissue. GSH-PX level of tumor tissue was correlated significantly with tumor grade (r=−0.425, P=0.034). Results suggested that pathway activity of free radicals were accelerated in the cancerous human bladder tissues via increased TBARs levels and decreased enzyme activities of XO, SOD, GSH-PX, and CAT, which implicated a severe exposure of cancerous tissues to oxidative stress. J. Clin. Lab. Anal. 24:25–30, 2010. © 2010 Wiley-Liss, Inc.