Bench-scale experiments were carried out to optimize ozone oxidation conditions for copepod inactivation by response surface methodology. Independent variables were ozone dosage (1, 3 and 5 mg/L), contact time (5, 15 and 25 min) and organic matter concentration (2, 5 and 8 mg/L). The results showed that all independent variables significantly influenced copepod inactivation rate. The observed inactivation rate was increased with the increasing ozone dosage and contact time. However, the trend of response gradually stabilized when the contact time was beyond 20 min. Copepod inactivation was more sensitive to high ozone dosage and short contact time under the same CT (dosage × contact time) value conditions. There was a negative effect of organic matter concentration on copepod inactivation. The polynomial response model for inactivation rate of copepod was established with R2 = 0.9933 and adjusted-R2 = 0.9865. The F-value of 165.93 implied the model significance and the p-value lower than 0.0001 indicated its good fit to use in the matrix. The ‘lack of fit (LOF) F-value’ of 0.0099 implied that the LOF was significant relative to pure error. The evaluation of the model showed it is able to make suitable predictions for the intended application.