Background and objectives <p>Fluorides are included in oral hygiene products to combat dental caries. But the potential threats in using a fluoridated toothpaste and its effect on oral buccal mucosal cells still remains a debate. The objective of this study was to evaluate the cytotoxic effects of fluoridated and non- fluoridated toothpaste on the buccal mucosa over a 12 week follow up period.</p> Materials and methods <p>This prospective double blinded parallel randomized clinical trial was conducted with 40 subjects. These participants were randomly assigned into two groups. 20 participants were given the fluoridated toothpaste and another 20 participants were given the non-fluoridated toothpaste. The participants were told to use the toothpaste for a period of 12 weeks. The buccal mucosal cell sampling was done after 12 weeks. All participants used the toothpaste for 12 weeks, which was already available in the market for this study (non-fluoride and 1400 ppm F for 12 weeks).</p> Results <p>The frequency of micronuclei showed statistically significant differences between the studied groups. The cellular changes in fluoridated subjects was not statistically significant (<i>p</i> = 0.087) and in non- fluoridated subjects also it was statistically not significant (<i>p</i> = 0.370). The nuclear changes in fluoridated subjects was not statistically significant (<i>p</i> = 1.000) but in non- fluoridated subjects it was statistically significant (<i>p</i> = 0.033). The intergroup comparison was not statistically significant (<i>p</i> = 1.000). Comparing two groups, statistically significant increase in the number of cells with cellular changes (<i>p</i> = 0.048) at 12 weeks of usage was observed, suggesting association between fluoride exposure and the presence of cellular changes. The micronuclei frequency between both the groups proved to be statistically significant (<i>p</i> = 0.0001*). There was a strong negative correlation between the type of toothpaste used and the number of micronuclei (<i>r</i> = − 0.853, <i>p</i> = 0.000), which was statistically significant. There was no significant correlation between gender and micronuclei (<i>r</i> = 0.098, <i>p</i> = 0.549) or cellular changes (<i>r</i> = 0.052, <i>p</i> = 0.752) Age shows no significant correlation with any of the parameters such as micronuclei (<i>r</i> = − 0.171, <i>p</i> = 0.291), nuclear changes (<i>r</i> = 0.038, <i>p</i> = 0.818), or cellular changes (<i>r</i> = − 0.051, <i>p</i> = 0.756).</p> Conclusion <p>Based on the results of this study, it can be concluded that simultaneous application of fluoridated toothpaste and fluoride based oral hygiene products does lead to cytogenetic damage in buccal mucosal cells.</p> Clinical relevance <p>The potential cytotoxic effects of fluoridated formulations on the oral mucosa, particularly under conditions of prolonged and repeated use, remain a matter of concern. This study evaluates whether fluoridated and non-fluoridated toothpastes exert differential effects on the buccal mucosa over an extended period, assisting clinicians in selecting dentifrices that ensure both caries prevention and the development of formulations suitable for individuals with mucosal sensitivity or existing oral pathologies.</p>