Organic wastewater contaminants, including pharmaceuticals, caffeine, and nicotine, have received increased scrutiny because of their detection in water bodies receiving wastewater discharge. Despite recent measurement in United States streams, caffeine's effect on freshwater organisms is not well documented. The present study measured caffeine's lethal and sublethal effects on the freshwater species, Ceriodaphnia dubia, Pimephales promelas, and Chironomus dilutus. These organisms, which are used in standard testing or effluent monitoring, were exposed to aqueous caffeine solutions under static exposure for 48 hours and daily renewed static exposure for 7 days. Averaged responses of 48-hour acute end points indicated that C. dubia was more sensitive to caffeine exposures (LC 50 = 60 mg/L) than either P. promelas (LC 50 = 100 mg/L) or C. dilutus (LC 50 = 1,230 mg/L). Exposure-response slopes confirmed these findings (3% mortality/mg/L for C. dubia; 0.5% mortality/mg/L for P. promelas; and 0.07% mortality/mg/L for C. dilutus). Comparative 7-day responses between C. dubia and P. promelas (LC 50 = 46 and 55 mg/L, respectively) were more similar than the broad range of acute values.Sublethal effects measured for caffeine exposure included impaired C. dubia reproduction (IC 50 = 44 mg/L) and inhibited P. promelas growth (IC 50 = 71 mg/L). According to the results of this study, combined with earlier studies reporting environmental concentrations and product halflives, caffeine should pose negligible risk for most aquatic vertebrate and invertebrate organisms.