Objective: To investigate the antibiofilm activity of alpha- mangostin (AMG) loaded nanoparticle (nanoAMG) against dental caries pathogen Streptococcus mutans. Methods: AMG was isolated from the peels of Garcinia mangostana L. using silica gel columns and chemically analysed by high performance liquid chromatography and nuclear magnetic resonance. NanoAMG was prepared using the solvent evaporation method combined with high-speed homogenization. The nanoparticles were characterized using dynamic light scattering, field emission scanning electron microscopy (FE-SEM) and Fourier transform infrared spectroscopy (FTIR). The toxicity of nanoAMG in fibroblast NIH/3T3 cell line was determined using MTT method. The antibiofilm effect of nanoAMG was determined through the evaluation of biofilm formation by Streptococcus mutans using a 96-well plate. Biofilm biomass was quantified using crystal violet. Cell viability was observed under confocal microscopy using LIVE/DEAD BacLight staining. Moreover, gene expression was determined by quantitative real-time PCR and membrane permeabilization activity by measuring the uptake of o-nitrophenol- β-D-galactoside. Results: NanoAMG size was in a range of 10-50 nm with a polydispersity index of < 0.3 and zeta potential value of -35.2 mV The size and the incorporation of AMG in the nanoparticles were confirmed by FE-SEM and FTIR analyses. The IC