The invention discloses an HPLC-MS/MS method for detecting chlorzoxazone in animal plasma. The method comprises the following steps: weighing a sample to be detected, adding a glipizide internal standard solution and acetonitrile, performing whirling, ultrasonic and centrifugal treatment, extracting supernatant, and performing filtering and sampling for HPLC-MS/MS analysis; in addition, preparing stock solutions by dissolving chlorzoxazone and internal standard glipizide with methanol and performing volume fixation with acetonitrile respectively, preparing a series of standard working solutions by adopting treated blank plasma, and performing HPLC-MS/MS analysis to obtain a working curve and a correlation coefficient of chlorzoxazone; substituting measured detection data of the sample to calculate the chlorzoxazone concentration of the sample through the working curve and the correlation coefficient. The method is high in sensitivity, easy to operate, relatively high in repeatability and accuracy, and suitable for analyzing the content of chlorzoxazone in animal plasma, and the defects of the prior art are overcome.