Biofilm accumulation on the polymethyl methacrylate (PMMA) restorations negatively affect the prognosis of the provisional restorations or the following treatment. This study developed a novel antibacterial PMMA resin containing low concentration of dimethylaminohexadecyl methacrylate (DMAHDM). Four resins were tested: (1) PMMA resin (Control), (2) 1.25% DMAHDM, (3) 2.5% DMAHDM, (4) 5% DMAHDM. Adding 1.25% DMAHDM into the PMMA resin did not influence the mechanical properties, degree of conversion, monomer releasing, and color stability of the specimens (p > 0.05). The incorporation of DMAHDM into PMMA resin could greatly prevent saliva-derived biofilms adhesion compared with the control group (p < 0.05). The metabolism level of saliva-derived biofilms on the 1.25%, 2.5%, and 5% DMAHDM resins were reduced by 20%, 54%, and 62%, respectively. And the mechanism of DMAHDM disturbing the integrity of bacterial cell walls was confirmed by flow cytometric analysis. Adding 1.25% and 2.5% DMAHDM did not compromise cytocompatibility of the modified resin (p > 0.05). Therefore, novel PMMA resin containing low concentration DMAHDM is promising as a future antimicrobial provisional restoration material for preventing microbial-induced complications in clinical settings.