The present studies were undertaken to determine the effects of reactive oxygen metabolites on erythropoietin (Ep) biosynthesis in Ep-producing renal carcinoma (RC) cells using a sensitive radioimmunoassay for Ep. Xanthine (10-5M) and increasing concentrations of xanthine oxidase (8 x 10(-7) to 5 x 10(-4) units/ml) produced a significant dose-related increase in Ep production at a concentration of greater than or equal to 4 x 10(-6) units/ml, whereas xanthine alone had no effect. Catalase, a scavenger of hydrogen peroxide (H2O2), in concentrations of 50 to 500 micrograms/ml produced a significant inhibition of the increase in Ep production induced by xanthine-xanthine oxidase; while no effect was seen on basal levels of Ep production and the growth of RC cells. Glucose oxidase (greater than or equal to 0.032 mU/ml), a direct H2O2 generator, and exogenous H2O2 (greater than or equal to 4 x 10(-6)M) added to the incubation mixture, caused a significant enhancement of Ep production in a dose-dependent manner. Xanthine-xanthine oxidase, glucose oxidase, and H2O2 in the above concentrations did not produce significant cytotoxicity (51Cr release or trypan blue dye exclusion). The present data suggests that H2O2, a reactive oxygen metabolite may play a significant role in Ep production.