3-Nitro-l,2,4-triazol-5-onc (NTO) is an explosive developed at the Los Alamos National Laboratory in 1984. The thermal decomposition characteristics of NTO, including calculated detonation velocity and pressure, are similar to those of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX); however, NTO is less sensitive and more stable. Little or no open literature is available about the environmental toxicity of NTO and understanding of potential ecological impacts of its accidental release into aquatic ecosystems is necessary prior to its large-scale production. We conducted aquatic toxicity testing of NTO using the freshwater organism Ceriodaphnia dubia in a 7 day survival and reproduction assay and the unicellular green algae Selenastrum capricornutum in a 96 hr growth inhibition assay. The studies were conducted under pH-adjusted conditions due to a concentration-dependent increase in acidification of aqueous media by the addition of NTO. The resulting inhibitory concentration (IC) IC2 and IC50 values for reproduction of C. dubia were 51 and 57 mg/L, respectively. The estimated IC2o and IC50 values for S. capricornutum growth inhibition were 2195 and 3465 mg/L. Applying these results to the Chemical Scoring System for Hazard and Exposure Identification, NTO was ranked as practically nontoxic.