Oral hygiene is essential to prevent cavities and periodontitis associated with bacteria such as Streptococcus mutans and Actinomyces viscosus, controlling their balance through the use of mouthwashes. This study investigated the in vitro antibacterial efficacy of three different mouthwashes against Streptococcus mutans and Actinomyces viscosus. The antimicrobial effect was evaluated using the inhibition zone method after 24 and 48 h of incubation. The sample groups were distributed as follows: Experimental groups (30 = Listerine, 30 = Colgate Plax Soft Mint, 30 = povidone iodine 0.23%). Control group (+) 30 = 0.12% chlorhexidine and control group (−) 30 = distilled water. The statistical analysis considered ANOVA and Tukey’s test. The results indicated that Colgate Plax Soft Mint exhibited the highest inhibition zone of 21.38 mm against Actynomices viscosus and 20.37 mm against Streptococcus mutans at 24 h, while Listerine showed the lowest effect with an inhibition zone of 15.68 mm at 24 h and 8.84 mm at 48 h. The positive control group had an inhibition zone of 22.69 mm at 48 h in Actynomices viscosus and 18.98 mm at 24 h in Streptococcus mutans. It is concluded that the mouthwashes evaluated showed antimicrobial activity at 24 and 48 h, with no significant effect of time on the size of the inhibition zones. Colgate Plax Soft Mint generated the greatest halo and Listerine the least at 24 h, while Chlorhexidine 0.12% was more effective against Actinomyces viscosus.