Abstract:Investigate the effects of combined environmental estrogens (EES) treatment on reproductive behaviors of male zebrafish ( Danio rerio ). Four groups were set as follow: Control (without EES in water), EE2-low (with EE2: 5.55 ng L -1 in water), EE2-high (with EE2: 11.1 ng·L -1 in water) and EES (water containing 4-NP: 62.2 ng·L -1 , BPA: 250 ng·L -1 , E1: 4.56 ng·L -1 , E2: 5.53 ng·L -1 , E3: 39.6 ng·L -1 ). Adult male zebrafish were exposed to the drugs mentioned above for 60 d and then paired with unexposed adult females individually. Reproductive behaviors of both male and female were counted and analyzed thereafter. The results showed that compared with the control group, there was no significant difference in the time and chasing frequency of single and simultaneous entry into the spawning area between the EE2-low group and the male zebrafish. However, the EES-high group and EES group showed a significant decrease in the above four indicators for both female and male zebrafish ( p <0.05). The research results indicate that the combined treatment of multiple environmental doses of EES has stronger reproductive toxicity compared to 5.55 ng·L -1 with the same estrogenic effect, and the reproductive toxicity of 11.1 ng·L -1 EE2 is basically equivalent and can significantly inhibit the reproductive behavior of male zebrafish.