This in vitro study evaluates the influence of application time of a one-step finishing system has on biofilm accumulation over nanoparticle filler-reinforced dental composites using a microcosm biofilm model in a constant depth film fermenter (CDFF). For that, sixty disk-shaped specimens (ø=5 mm × 2 mm thick) were made with nanoparticle filler reinforced dental composites (EsthetX™ and IPS Empress Direct). The specimens were manually polished with a finishing system (Enhance®) following the manufacturer's instructions for 5, 15 and 30 seconds, while unpolished specimens were used as controls. Oral biofilm was formed on the discs, using human saliva as inoculum and daily subject to 8 pulses of 10% sucrose solution. The biofilm was collected to determine the counts of total microorganisms. Data were analyzed by ANOVA-Tukey tests (α=5%). Increased time application of the finishing system significantly affected biofilm formation for tested materials (p ≤ 0.05). Biofilm formation over the nanoparticle filler-reinforced dental composites was greatly reduced after 15 s of application. No additional significant reduction in biofilm was observed when the specimens were finished for 30 seconds.However, differences related to the tested nanoparticle filler-reinforced dental composites were not statistically significant. By incorporating a minimum 15 s-time protocol into their everyday practices, dentists can positively reduce biofilm formation on nanoparticle fillereinforced dental composites and contribute to the longterm esthetic and integrity these restorations.