The contribution of veterinary growth promoters (VGP) to the environmental burden of endocrine-disrupting chemicals (EDCs) is largely unknown. At cattle feedlots, the excrement of cattle may contain VGPs, which can contaminate aquatic systems and pose reproductive health risks. The study identifies VGPs used at cattle feedlots in South Africa and confirms associated estrogenic activity in feedlot runoff water. Using a rat model, we investigate the potential reproductive health effects and thyroid function of an environmentally relevant mixture of VGPs. Collected water samples had low levels of selected VGPs, and estrogenic activity was detected in the T47D-KBluc bioassay. Rats exposed to VGP had significant adverse effects on male reproductive health, including shortened anogenital distance, lowered sperm counts, disorganized seminiferous tubules, and thyroid parameters. In conclusion, VGP can contribute to complex environmental EDC mixtures and may adversely affect the reproductive and thyroid health of both humans and wildlife. The varied topography of individual cattle feedlots will govern the rate and extent of effluent runoff, thus continuous monitoring of VGPs in aquatic systems surrounding cattle feedlots is necessary.