The aim of this study was to validate a model of S. mutans biofilm formation, which simulated 'feast-famine' episodes of exposure to sucrose that occur in the oral cavity, showed dose-response susceptibility to antimicrobials and allowed the evaluation of substances with anticaries potential. S. mutans UA159 biofilms were grown for 5 days on bovine enamel slabs at 37°C, 10% CO2. To validate the model, the biofilms were treated 2x/day with chlorhexidine digluconate (CHX) at 0.012, 0.024 and 0.12% (concentration with recognized anti-plaque effect) and 0.05% NaF (concentration with recognized anti-caries effect). CHX showed dose-response effect decreasing biomass, bacterial viability and enamel demineralization (p < 0.05). Whereas, 0.05% NaF did not show antimicrobial effect but had similar effect to that of 0.12% CHX decreasing enamel demineralization (p < 0.05). The model developed has potential to evaluate the effect of substances on biofilm growth and on enamel demineralization.