Objectives Systemic diseases or oral situation changes can result in oral infections like candidiasis. Mouthwash is the most prevalent method to prevent or cure these infections. To formulate a more effective mouthwash, we combined Clotrimazole with a low dose of Chlorhexidine to investigate the in vitro effect against Candida spp.Methods and Materials Combinations of Chlorhexidine (0.03-16ug/ml) and Clotrimazole (0.03-16ug/ml) were tested against Candida spp. by microdilution chequerboard technique and disk diffusion method.Results From the chequerboard combination assay, the MICs of Chlorhexidine and Clotrimazole against Candida spp. decreased from >16µg/ml to 2–1µg/ml and from 2-0.25µg/ml to 0.5–0.125µg/ml, respectively, demonstrating favorable synergistic effects against 21 (84%) strains of Candida spp. The disk diffusion method showed an increase in halo size for the combination group when compared to the Clotrimazole alone group.Conclusions Studies have shown that combinations of antiseptic and antifungal agents are effective in nature. In our study, we found that low concentrations of Chlorhexidine can enhance the antifungal effect of Clotrimazole against Candida spp.. We predict that the mechanism of this synergism could be due to the increased penetration of Clotrimazole brought about by the binding of Chlorhexidine to the cell membrane. Further studies to determine the mechanism and in vivo effects could increase its probable usage in clinical studies.