The increasing resistance of microorganisms to conventional antimicrobial compounds requires the development of innovative solutions, such as antimicrobial nanoparticles, to combat antibiotic-resistant infections. This study aimed to optimize the synthesis of a cellulose/gum Arabic/silver (cellulose/GA/Ag) bionanocomposite and evaluate its antibacterial properties against S. mutans, a key contributor to dental caries. Using the Taguchi method, we designed nine experiments with varying levels of cellulose (2, 4, and 6 mg/mL), gum Arabic (1, 2, and 3 mg/mL), and silver nanoparticles (2, 4, and 6 mg/mL). The nanocomposite synthesized under optimal conditions (2 mg/mL cellulose, 3 mg/mL gum Arabic, and 6 mg/mL silver nanoparticles) demonstrated the most potent antibacterial activity, reducing the bacterial survival rate of S. mutans to 0 Log10 CFU/mL, indicating complete inhibition. Variance analysis revealed that silver nanoparticles had the most significant impact on bacterial survival (53.22%), followed by gum Arabic (35.55%) and cellulose (8.86%). Characterization techniques confirmed the successful formation of the nanocomposite: FTIR analysis indicated hydrogen bonding between cellulose and silver nanoparticles, while XRD confirmed the crystalline structure of the nanocomposite. SEM and TEM images revealed a uniform distribution of silver nanoparticles within the cellulose–gum Arabic matrix. TGA-DSC analysis showed enhanced thermal stability, with a significant weight loss at 375 ℃, corresponding to the degradation of cellulose and gum Arabic. The results demonstrate that the cellulose/GA/Ag nanocomposite, synthesized under optimal conditions, exhibits exceptional antibacterial properties and stability, making it a promising candidate for antimicrobial applications in medical and dental fields.