Taking apparent enzyme activity of immobilized papain as the index, the effects of ultrasound frequency and power on the kinetics of immobilized papain chemical reaction were investigated. It was found that the decomposing reaction of casein by the immobilized papain is consistent with the first-order reaction rate formula. Ultrasonic treatment can significantly reduce the apparent activation energy. The ultrasonic power was 0.05 W/cm2, and the ultrasonic frequency was 28, 40, 50 and 135 kHz, the apparent activation energy decreased gradually, and the apparent activation energy of 38.23±0.53 kJ/mol at 135 kHz 0.25 W/cm2 was the lowest. When the ultrasonic frequency was 135 kHz and the ultrasonic power was increased from 0.05 W/cm2 to 0.45 W/cm2, the apparent activation energy of 32.81±0.09 kJ/mol was the lowest under the ultrasonic condition of 135 kHz 0.25 W/cm2, which was decreased by 52.48% compared with that before ultrasonic treatment.