Effect of Iron Oxide Nanoparticle Incorporation on the Cytocompatibility and Antimicrobial Behavior of a Calcium Silicate-Based Endodontic Sealer

Background: Persistent intraradicular infection and biofilm survival remain major challenges in endodontic treatment, particularly because residual microorganisms may remain within dentinal tubules despite chemomechanical preparation. The antimicrobial efficacy of sealers may be insufficient against resistant bacteria. This study evaluated the effect of incorporating red and black iron oxide nanoparticles into BioRoot RCS on its antimicrobial activity and cytocompatibility. Methods: BioRoot RCS was modified with red or black iron oxide nanoparticles at 0.5 wt% and 2.0 wt%, generating 5 groups: unmodified sealer, 0.5% red, 2.0% red, 0.5% black, and 2.0% black. Surface morphology was analyzed using scanning electron microscopy, while elemental composition was determined by energy-dispersive X-ray spectroscopy. Antibacterial activity against Enterococcus faecalis and Fusobacterium nucleatum was assessed using a direct contact test, antibiofilm activity by colony-forming unit reduction on infected dentin discs, and cytocompatibility using human gingival fibroblasts and the AlamarBlue assay. Results: Iron was detected in the modified formulations, and elemental mapping showed homogenous distribution of calcium and iron. The 2.0% formulations showed significantly higher antibacterial and antibiofilm effects than the corresponding 0.5% groups (p < 0.05), with 2.0% black showing the lowest bacterial counts. Cytocompatibility differed at 1 and 3 days but not at 7 days, and all groups remained close to the control level with no significant difference (p > 0.05). Conclusions: Within the limitations of this in vitro study, experimental modification of BioRoot RCS with iron oxide nanoparticles, particularly at 2.0 wt%, improved the antimicrobial and antibiofilm efficacy of BioRoot RCS while maintaining acceptable cytocompatibility. However, physicochemical and handling properties must be evaluated before the clinical relevance of this modification can be determined.