Organic matrix cultivation is one of the main forms of soilless cultivation in facility agriculture. In the process of repeated utilization of organic matrix, continuous cultivation obstacles need to be disinfected, etc. Based on the analysis of the current disinfection technology and disinfection equipment, the ozone disinfection technology was proposed to solve the problem. Combining with rotating barrel research status, the rotary drum of ozone disinfection equipment parameters, the main structure and key operation parameters for the theoretical analysis and preliminary specified were designed, due to the dynamics and kinematics of organic matrix in the rotating barrel was complex, the EDEM software was adopted for organic matrix particle modeling and SolidWorks for rotating barrel with lifting board and screw conveyers cover, selecting Hertz-Mindlin contact model. The EDEM software was used with organic substrate bending angle as the evaluation index, lifting board bending angle, rotating barrel rotational speed and fill rates as test factors. Numerical simulation of three factors and five levels of orthogonal rotation was carried out. By using the regression analysis method of the DesignExpert software, a regression mathematical model of bending angle was established to optimize and verify the parameters. The results showed that the lifting board angle and rotating barrel rotational speed had significant influence on the bending angle, but the filling rate was not significant in the range of 10%~30%. The optimal structure parameters and working conditions of the organic matrix ozone disinfection device were: lifting board bending angle was 124.23° and drum rotational speed was 6.29r/min, the organic substrate bending angle was 89.3°. According to theoretical analysis and simulation results, the development of ozone disinfection equipment rotating barrel diameter was 1000mm, the length was 1000mm, and eight lifting boards were distributed in the rotating board radial uniformly, and its height was 170mm, and bending angle was 124°. The screw conveyers outside diameter was 160mm, the screw conveyers inside diameter was 60mm, the pitch was 160mm, and screw conveyers opening angle was 60°. Validation results showed that, in the disinfection stage, an effective “material curtain” and second collision with screw conveyers cover of the organic matrix was formed. In the unloading stage, the organic matrix can fall back to the inside of the screw conveyers cover. After the ozone disinfection equipment was sterilized for 60 minutes, the sterilization performance test showed that the number of bacteria was 31CFU/mL, the number of fungi was 3CFU/mL, the sterilization rate of bacteria was 88.9%, and the sterilization rate of fungi was 97.9%, which met the actual production needs.