Antibiofilm activity of α-pinene and berberine against Enterococcus faecalis: an in vitro study

Background This in vitro study aimed to evaluate the antibacterial and antibiofilm effects of the natural compounds α-pinene and berberine, compared with 2.5% sodium hypochlorite, against Enterococcus faecalis biofilms. Methods Forty-nine extracted mandibular premolars were prepared, sectioned, and inoculated with Enterococcus faecalis. For antibacterial testing, specimens were incubated with Enterococcus faecalis for 7 days and then treated with α-pinene (2 mM), berberine (2 mM) for 24 h, or sodium hypochlorite (2.5%) for 5 min. Untreated specimens served as controls. For biofilm inhibition testing, specimens were pretreated with the test agents prior to bacterial inoculation. Outcomes were evaluated by colony-forming unit (CFU) counts and confocal laser scanning microscopy (CLSM). Data were analyzed using ANOVA and Tukey tests. Results All tested agents eliminated mature biofilms, resulting in complete bacterial killing (0.00 log₁₀ CFU), whereas untreated specimens showed significant bacterial growth (8.08 log₁₀ CFU). In the inhibition model, berberine and sodium hypochlorite prevented biofilm formation (0.00 log₁₀ CFU), while α-pinene did not (7.47 log₁₀ CFU). Conclusions Both α-pinene and berberine exhibited strong antibacterial effects against mature Enterococcus faecalis biofilms. Notably, berberine also inhibited biofilm formation, highlighting its potential as a natural adjunctive antimicrobial agent for endodontic disinfection.