Purpose: The effects of arginine as a toothpaste additive were assessed on oral streptococci with and without a known arginine deiminase system (ADS) and cariogenic biofilms. Materials and Methods: Suspensions of Streptococcus mutans, S. sobrinus and the ADS-positive (ADS+) S. sanguinis and S. gordonii were cultured with or without 1.5% L-arginine for 24 h. Thereafter, biofilms consisting of the four species were formed on polystyrene surfaces with or without 1.5% L-arginine for up to 10 d. Finally, biofilms that formed on enamel surfaces were exposed to a daily mechanical cleaning with an arginine and sodium monofluorophosphate (SMF+Arg)-containing toothpaste, a sodium monofluorophosphate fluoride (SMF)-containing toothpaste or a negative control for up to 10 weeks. At different incubation times, the pH in the culture media, the citrulline production and the percent of ADS+ bacteria within the biofilms were determined. Microsurface hardness loss was quantified in the experiments using enamel specimens. Results: In the presence of 1.5% arginine, S. sanguinis and S. gordonii showed a high level of production of citrulline after 6 h of incubation, together with an increase in the pH when compared to S. mutans and S. sobrinus. With arginine supplementation, the percentage of ADS+ species was higher at 1, 2 and 4 days and citrulline production was higher at all days of biofilm formation on polystyrene surfaces. After 4 and 10 weeks of treating biofilms on enamel surfaces, the SMF+Arg group had a higher proportion of ADS+ strains than the SMF group; at 4 weeks, the pH was higher in the SMF+Arg group. Loss of enamel hardness was the lowest in the SMF+Arg group and was significantly less in the SMF+Arg group than in the control group after 2, 4 and 10 weeks of treatment. Conclusion: Toothbrushing using an arginine-containing toothpaste may protect against dental caries. Keywords: arginine, citrulline, multispecies biofilm, microsurface hardness, oral streptococci