Toothbrushes sterilization has strongly taken into consideration in order of prohibiting of transmission and reinfection. The differences of materials, procedures and duration, made the problem more complicated. Our objective of the present research was assessment of antibacterial effects of different procedures which are used for disinfection of those toothbrushes which are infected to Streptococcus mutans. 144 fresh toothbrushes were immersed in a 0.5 McFarland culture of S. mutans for 15 minutes. The toothbrushes were then classified into 8 groups ( 6 in each group) according to their disinfecting procedure including White vinegar 50%, Sodium hypochlorite %1, Ethyl alcohol and Povidone Iodine 10%. The disinfecting procedures which used were Microwave and Dishwasher. Bacterial cultivation was done and microbial colonies had been counted before toothbrushes were exposed to the above disinfectants. After exposure of toothbrushes to above disinfectants for 1, 5 and 10 minutes, the colonies were recounted. SPSS ver.19 (Kruskal–Wallis Test) was used for statistical analysis. Bacterial cultures of samples at Chlorhexidine group (negative control) were negative and White vinegar did not make a considerable sense. On the other hand, Sodium hypochlorite %1, Ethyl alcohol , Povidone Iodine 10% and dishwasher decreased the numbers of S. mutans considerably (p< %5). Microwave by increasing the time of exposition, decreased the numbers of microbes but it was not statistically significant. Povidone Iodine 10%, Sodium hypochlorite %1 and Ethyl alcohol, could be effective in disinfection of toothbrushes against S. mutans if be used for 10 minutes.