Abstract: In order to improve the quality of beef,papain was adopted to optimize the beef tenderness. Based on the single factor experiments,the enzyme concentration,pH,processing temperature,processing time,which had a significant influence on the shear force and sensory evaluation were optimized by response surface optimization using Box-Behnken experimental design. The optimum tenderizing conditions were determined as follows:enzyme concentration of 0.07%,pH6.9,processing temperature of 60℃,processing time of 1.8 h. Under the optimum conditions,the water holding capacity,elasticity,cohesion,chewiness of beef were significantly raised,while the cooking water loss rate,shear force and hardness were remarkable reduced,compared to the control group(p<0.01). The sensory evaluation reached 94.20 points after optimization,which increased by 39.56%. The preliminary study have shown that papain could degrade beef muscle fiber to improve tenderness of beef.