Dimercaptosuccinic acid (DMSA) coating improves the uptake efficiency presumably by engendering the Fe2O3-NPs.In the present study, we investigated the possible environmental safety concentrations of Fe2O3-NPs using different assay systems in nematode Caenorhabditis elegans with lethality, development, reproduction, locomotion behavior, pharyngeal pumping, defecation, intestinal autofluorescence and reactive oxygen species (ROS) production as the endpoints.After exposure from L4-larvae for 24-hr, DMSA coated Fe2O3-NPs at concentrations more than 50 mg/L exhibited adverse effects on nematodes.After exposure from L1-larvae to adult, DMSA coated Fe2O3-NPs at concentrations more than 500 mg/L had adverse effects on nematodes.After exposure from L1-larvae to day-8 adult, DMSA coated Fe2O3-NPs at concentrations more than 100 mg/L resulted in the adverse effects on nematodes.Accompanied with the alterations of locomotion behaviors, ROS production was pronouncedly induced by exposure to DMSA coated Fe2O3-NPs in the examined three assay systems, and the close associations of ROS production with lethality, growth, reproduction, locomotion behavior, pharyngeal pumping, defecation, or intestinal autofluorescence in nematodes exposed to DMSA coated Fe2O3-NPs were confirmed by the linear regression analysis.Moreover, mutations of sod-2 and sod-3 genes, encoding Mn-SODs, showed more susceptible properties than wildtype when they were used for assessing the DMSA coated Fe2O3-NPs-induced toxicity, and the safety concentrations for DMSA coated Fe2O3-NPs should be defined as concentrations lower than 10 mg/L in sod-2 and sod-3 mutant nematodes.