Dengue transmitted by mosquitoes of the genus Aedes, species aegypti, is a major public health concern in Brazil. The chemical control of the mosquito larvae has been performed with the larvicide temephos since 1967. However, vector resistance was reported to temephos in several Brazilian states, and the Ministry of Health ordered the replacement of this larvicide by diflubenzuron (DFB), an inhibitor of chitin synthesis. Both insecticides are diluted in water with larvae and are able to reach aquatic environments in which they subsequently adversely damage nontarget organisms. The aims of this study were to (1) determine the acute toxicity (EC50) and environmental risk (RQ) of DFB and temephos to the microcrustacean Daphnia magna, and (2) evaluate the chronic toxicity (no-observed-effect concentration [NOEC] and lowest-observed-effect concentration [LOEC]) of these larvicides to D. magna. The experiments were performed according to a completely randomized design. The estimated 48-h EC50 of temephos was 0.15 μg/L (lower limit = 0.1 and upper limit = 0.2 μg/L) and the 48-h EC50 of DFB was 0.06 μg/L (lower limit = 0.03 and upper limit = 0.1 μg/L). RQ values were 4.166.7 to DFB and 6.666.6 to temephos. NOEC and LOEC values were respectively 2.5 and 5 ng/L for DFB, and respectively 6.2 and 12.5 ng/L for temephos. Thus, temephos and DFB are classified as highly toxic to Daphnia magna and pose a high environmental risk to this species. Mortality of D. magna was observed at concentrations lower than those used in the field to control A. aegypti larvae.