Benzophenone-3 (BP-3) is an organic UV filter that is widely used in personal care products and has been indicated to have negative impacts on the environment and human health. The mammary glands of humans and rodents have been confirmed to be target organs affected by BP-3 exposure. However, limited information is available on the underlying mechanism currently. In this study, we hypothesized that low-concentration BP3 exposure during puberty might lead to a susceptibility to tumors through the mediation of mammary stem cells. Our findings revealed that BP-3 exposure at 50 mg/kg/day for 5 weeks during puberty led to reproductive outcomes such as reduced body weight, decreased serum estradiol and progesterone levels, and increased terminal end bud (TEB) numbers and areas. These effects were accompanied by a decreased fraction of basal mammary stem cells and decreased self-renewal and differentiation abilities of basal mammary stem cells in vitro and in vivo such as decreased sphere formation ability, a smaller 3D structure, increased branching points and hyperplastic lesions in regenerated mammary glands. Notably, for the regenerated mammary glands formed by the basal mammary stem cells of BP-3-treated mice, a decrease in the fraction of basal mammary stem cells and decreased expression levels of the milk protein β-casein and STAT5 were observed. Taken together, our data suggest that pubertal BP-3 exposure decreases the function of basal mammary stem cells such that they induce the abnormal development of mammary glands.