The purpose of this study was to provide an effective toothbrush management method by comparing the number of bacteria according to the toothbrush rinsing water temperature and the toothbrush storing method, and to recognize the importance of the toothbrush management method. To identify the changes in the number of bacteria according to the toothbrush rinsing water temperature, the toothbrushes of 50 students were collected after they brushed their teeth, then 20 ml cold water (10°C) and hot water (50.0°C), respectively, were sterilized right before the experiment, and were used for rinsing the toothbrushes. To identify the changes in the number of bacteria by toothbrush storing method, the toothbrushes were stored at one of the following locations and under the corresponding condition: a sunny windowsill, indoors without a cap, or indoors with a cap. To observe the types of bacteria present in the toothbrushes, a phase microscope was used, and to identify the variations of Streptococcus mutans (S. mutans) present, Brain Heart Infusion (BHI) broth agar was used for the culture. Statistical analysis was performed using IBM SPSS Ver. 21.0 (IBM Co., Armonk, NY, USA) at a 0.05 significance level through a t-test and one-way ANOVA and Tukey’s post hoc test. When the types of bacteria present in each toothbrush were determined using a phase microscope, cocci with motility, which contribute to the activation of dental caries, were mostly found. In addition, the number was found to have significantly deceased when the toothbrush was rinsed in hot water rather than in cold water and decreased in the sunny windowsill group, followed by the “indoors with a cap” group and the “indoors without a cap” group, in that order. Therefore, rinsing a toothbrush with hot water and storing it at a sunny place will decrease the bacterial proliferation in the toothbrush.