We investigated the effect of photodynamic therapy (PDT) with hematoporphyrin monomethyl ether (HMME) on the viability of Streptococcus mutans (S. mutans) cells on biofilms in vitro. Streptococcus mutans is the primary etiological agent of human dental caries. Since dental caries are localized infections, such plaque-related diseases would be well suited to PDT. The diode laser used in this study had the wavelength of 635 nm, whose output power was 10 mW and the energy density was 12.74 J/cm2. HMME was used as photosensitizer. Samples were prepared and divided into five groups: (1) HMME; (2) Laser; (3) HMME+Laser; (4) Control group (+) with chlorhexidine; and (5) Control group (-) with sterile physiological saline. Inoculum of S. mutans incubated with HMME also examined with fluorescence microscopy. PDT exhibited a significantly (P<0.05) increased antimicrobial potential compared with 20 \mum/mL HMME only, laser only, 0.05% chlorhexidine, and 0.9% sterile physiological saline, which reduced the S. mutans of the biofilm most effectively. Laser and 0.05% chlorhexidine were caused reduction in the viable counts of S. mutans significantly different (P<0.05) also, but these two test treatments did not statistically differ from each other. HMME group did not statistically differ with negative control group. Fluorescence microscopy indicated that HMME localized primarily in the S. mutans of the biofilm. It was demonstrated that HMME-mediated PDT was efficient at killing S. mutans of biofilms and a useful approach in the treatment of dental plaque-related diseases.