Our previous study revealed that the photo-irradiation of rose bengal, erythrosine, and phloxine, xanthene photosensitizers, used as dental plaque disclosing agents, could exert bactericidal action on planktonic Streptococcus mutans via the singlet oxygen. In the present study, the photo-irradiation induced bactericidal activity of the three xanthene compounds against the experimental biofilm of S. mutans was investigated in combination with acid electrolyzed water (AcEW) and alkaline electrolyzed water (AlEW). As a result, only the photo-irradiated rose bengal in AlEW showed prominent bactericidal activity with a >3-log reduction of the viable bacterial count. Since our previous study showed that the affinity of rose bengal to bacterial cells was superior to that of erythrosine and phloxine, it was speculated that AlEW damaged the extracellular matrix of the experimental biofilm, which would let the rose bengal easily be bound to the bacterial cells. From these results, it is strongly suggested that rose bengal is a suitable photosensitizer for use as a plaque disclosing agent in photodynamic antimicrobial chemotherapy to treat dental plaque.