The effect of O₃ oxidation on the flotation recovery of chalcopyrite and molybdenite was investigated. The results of single-mineral flotation tests showed that under the conditions that O₃ concentration was about 90 g/m³ and the gas flow rate was 4 L/min, after chalcopyrite and molybdenite were oxidized for 2 min, the flotation recovery of the chalcopyrite dropped to 16%, while the flotation recovery of molybdenite did not change much. The results of flotation tests for the artificially mixed ores showed that O₃ oxidation pretreatment could replace the Na₂S reagent as a chalcopyrite depressant in the selective flotation separation of chalcopyrite and molybdenite. The contact angle measurements revealed that the O₃ oxidation treatment selectively rendered the surfaces of the chalcopyrite to be hydrophilic. The Zeta potential and X-ray photoelectron spectroscopy analyses revealed that the chalcopyrite surface most likely became hydrophilic due to the formation of water-insoluble hydrophilic oxides and hydroxides on the mineral surface after the oxidation treatment. In addition, although the surface of the molybdenite was slightly oxidized, its surface remained good hydrophobicity.