Mono(2-ethylhexyl)-phthalate (MEHP) is the primary metabolite of di(2-ethylhexyl)-phthalate (DEHP), which is widely used in industry as a plasticizer. Previous studies showed that both DEHP and MEHP have been found in not only human urine samples but also natural aquatic environments, and well documented as toxicants for reproduction and endocrine disruptors, However, the effects of MEHP exposure on aquatic organisms, including invertebrates such as Daphnia magna (D. magna), are still scarce. In the present study, the lipid alterations caused by MEHP in D. magna have been identi ed by analyzing lipid accumulation and nontarget metabolomics. Additionally, the expressions of reproduction were investigated. Ecotoxicologically relevant concentrations (1 to 2 mg/L) and exposure time ranges (24 h to 21 days) have been tested. MEHP resulted in no mortality to D. magna with all exposure conditions, but the number of lipid droplets increased after 96 h of exposure and enhanced reproduction of female adult daphnids was observed in the 21-day of exposure. Besides, we observed that MEHP enhanced lipid metabolism with the identi cation of 283 potential lipid metabolites, including glycerolipids, glycerophospholipids, and sphingolipids, following 48 h of exposure. MEHP treated group exhibited signi cantly higher expression of ecdysone receptor (EcR) and vitellogenin 2 (Vtg2) at 6 h and 24 h. Meanwhile at 48 h, EcR and Vtg2 were downregulated in 1 and 2 mg/L MEHP exposure. Our data show that the changes in EcR pathway with different exposure time could be associated with the lipid accumulation due to lipids increment subsequently to increased reproduction in MEHP-exposed D. magna.