As an environmental endocrine disruptor, Di-(2-ethylhexyl) phthalate (DEHP) affects blood-testis barrier (BTB)-associated proteins expression, which compromises BTB integrity and causes infertility. Notably, DEHP-induced testicular toxicity is related to oxidative stress, but the specific mechanism remains unclear. Therefore, we sought to investigate this mechanism and determine whether vitamin C and vitamin E administration would attenuate the BTB impairment induced by DEHP in vivo and by Mono-(2-Ethylhexyl) Phthalate (MEHP) in vitro, respectively. HE staining and EM found that DEHP exposure led to spermatogenesis dysfunction and BTB disruption, respectively. The Western blot and immunofluorescence results showed that DEHP exposure caused BTB impairment through oxidative stress-mediated p38 mitogen-activated protein kinase (MAPK) signaling pathway. Furthermore, Vitamin E and vitamin C could alleviate the oxidative stress, block DEHP-induced spermatogenesis dysfunction and BTB disruption by inhibiting p38 MAPK signaling pathway. In summary, vitamin E and vitamin C are good candidates for the treatment of DEHP-induced male infertility.