The occurrence of phthalate esters (PE) was examined in biota, ambient water, and sediments of two man-made lakes (Asejire and Eleyele) in southwestern Nigeria. Five fish species (Tilapia zillii, Hepsetus odoe, Parachanna obscura, Chrysichthys nigrodigitatus, and Mormyrus rume) were analyzed for PE levels and used for calculating bioconcentration factors (BCF) and biota–sediment accumulation factors (BSAF). In addition, measured PE levels were thereafter used to calculate the phthalate pollution index (PPI) in biota and the environment. At both lakes, all sampled species had k-factor > 1, showing apparently normal growth and health condition. Higher PE levels were found in sediments compared with water at both lakes, with a pattern showing that di(2-ethylhexyl) phthalate (DEHP) was predominant PE. While there were no unique patterns of PE concentrations in both lakes, differences were observed in organ concentration patterns that were evident at both lakes. For T. zillii, the BSAF was higher for dibutyl phthalate (DBP) compared to diethyl phthalate (DEP) and lowest for DEHP. The concentration pattern demonstrated that DBP concentrated more in gills (BCF: 6.7), while DEHP concentrated more in liver (BCF: 15.2) of T. zillii at Asejire. At Eleyele, T. zillii liver and gills concentrated less DEP and DEHP. The PPI value was significantly higher in sediment with respective values of 0.27 and 0.44 at Asejire and Eleyele lakes compared with water with respective values of 0.1 and 0.18 at Asejire and Eleyele lakes. Overall, our findings suggest a broader environmental and human health implication of high PE levels in these lakes, since they provide vast water volumes that are used for municipal domestic water supply. Further, these lakes support intense artisanal fisheries, representing significant sources of aquatic food resources for neighboring communities.