Aim: The study's aim was to compare the efficacy of 3% neem, 50% white vinegar, and a 40 mg/mL concentration of green tea, 0.2% chlorhexidine (CHX) gluconate, as disinfectants in reducing the number of Streptococcus mutans (S. mutans) in toothbrushes. Materials and Methods: The study was a parallel-arm, double-blinded, randomized, in vivo comparative experiment with 50 young adults selected at random by simple random sampling. The participants were divided into five experimental groups—group I with 3% neem extract, group II with 50% white vinegar, group III with 40 mg/mL of green tea extract, group IV with 0.2% CHX, and group V with plain water as the control. The participants were trained to immerse the toothbrush in the test solutions for 20 minutes after brushing daily for 7 consecutive days. All of the toothbrushes were tested for baseline values prior to intervention. The toothbrush bristles were collected after 7 days and underwent microbial analysis. The mean bacterial count for S. mutans was determined. Results: The Statistical Package for Social Sciences (SPSS) software version 2.1 was used to conduct the statistical analysis. Paired t-test and analysis of variance (ANOVA) were the two statistical tests utilized. All of the groups, with the exception of the control group, saw a statistically significant (p < 0.001) decrease in the number of S. mutans. The reduction in S. mutans is significantly different between groups I, II, III, and IV (p < 0.001). In group V, there were no significant changes. Conclusion: The bacterial counts of S. mutans in toothbrush bristles were successfully reduced using both herbal products and synthetic disinfectants. However, when compared to the other groups, 50% white vinegar performed the best. Clinical significance: It is essential for every individual to disinfect their toothbrush, either using herbal or synthetic solutions, in order to uphold good oral hygiene and systemic health, especially in immunocompromised patients.