Abstract: In this study, the cells of Klebsiella sp. O852 were disrupted by ultrasonic extraction. The location and the conversion conditions of the key enzymes for the conversion of limonene to trans-dihydrocarvone were investigated. Then, the effects of ultrasonic time, material-liquid ratio, and amplitude on cell disruption were studied using the single factor experiments. Additionally, the Box-Behnken central combination test was designed to determine the optimal extraction conditions. The results showed that the enzyme was an intracellular enzyme. The optimal cultivation time was 8 h, which was the mid-logarithmic phase of cell growth. The optimum conversion time was 4 h, and the highest enzyme activity was obtained in phosphate buffer solution. The single factor experiments and response surface test showed that the optimal ultrasonic time was 21 min, material-liquid ratio was 1:13, and amplitude was 36%. And the content of the enzyme activity was (40.97±2.02) U/g under these optimal conditions. This study would provide a reference for further study on the purification and enzymatic properties of this enzyme.