In order to clarify the biotoxic effects of oxytetracycline wastewater, the toxicity effects of oxytetracycline wastewater on zebrafish are studied by exposure test. The results show that oxytetracycline wastewater is a low toxic substance to zebrafish. The SOD activity and POD activity of zebrafish are differently inhibited at 3~9 d. At 12 d, SOD activity and POD activity are significantly induced (0.01<P<0.05). The SOD activity of 20% volume concentration group reaches the maximum (194.05 U/mg prot), and the POD of 60% volume concentration group reaches the maximum value (2.72 U/mg prot). The SOD activity and POD activity almost returns to control values at 15 d. In addition, there is significant correlation between SOD activity and POD activity in research. The MDA content shows a "Λ" trend in each volume concentration group, and the 80% volume concentration group reaches the maximum value (5.41 nmol/mg prot) at 15 d. The content of γ-H2AX shows a rising trend in zebrafish muscle tissue. At 15 d, the 40% volume concentration group shows extremely significant (400.02 ng/L, P<0.01), which is 1.57 times that of the control group. The DNA of zebrafish muscle cells has been damaged in a certain degree. This study shows that oxytetracycline wastewater is a low toxic substance to zebrafish, and the oxidative stress of zebrafish to oxytetracycline wastewater may be one of the mechanisms causing DNA damage. The biological toxicity of oxytetracycline can provide a scientific basis for further study of the ecotoxicity effect of oxytetracycline in water environment.