Everyday denture cleansing is needed to prevent the mouth from denture stomatitis and esthetic concern. Enzyme cleansers are developed in attempt to break down the organic components of denture plaque. The aim of this research is to determine the efficient cleansing duration of papain solution to remove 24-hour denture plaque. The research was done on 16 patients with complete dentures. Plaque removal was determined by Lowry method and SDS PAGE. The result shows that papain dose required to hydrolyze 24-hour denture plaque is 15.66 TU/mg enzyme activity for 10 minutes soaking. Soaking the denture in papain solution for 10 minutes makes all of the detected plaque protein bands remove. In conclusion, efficient cleansing duration of papain solution to remove 24-hour denture plaque is papain activity of 15.66 YU/mg by soaking duration of 10 minutes. Further research is suggested to examine papain residue on the denture that may influence the denture wearer biocompatibility.