Synergistic antibacterial effect of ginsenoside Rh2 and calcium hydroxide on Enterococcus faecalis

Treatment of root canal infections becomes more challenging due to the extremely high tolerance of Enterococcus faecalis (E. faecalis) to calcium hydroxide (Ca(OH)₂). Ginsenoside is a Chinese herbal extract that has been proven to have antimicrobial properties and synergistic activities. And this study evaluated the antibacterial activity of ginsenoside Rh2 in combination with Ca(OH)₂ against E. faecalis and its preliminary mechanism of action. Broth microdilution method, checkerboard dilution method, time-inhibition curve, drug resistance assays, scanning electron microscopy, and biofilm inhibition and removal assays indicated that Rh2 in combination with Ca(OH)₂ exhibited potent antibacterial activity against E. faecalis. Rh2 exerted significant in vitro antibacterial activity against E. faecalis, with a minimum inhibitory concentration (MIC) of 3.125 μg/mL and minimum bactericidal concentration (MBC) of 6.25 μg/mL, and significantly enhanced the susceptibility of E. faecalis to Ca(OH)₂ (FICI = 0.5). Furthermore, cell membrane permeability assays, surface hydrophobicity assays, ATPase activity assays, and intra-biofilm extracellular polysaccharides (EPS) assays revealed that Rh2 and Ca(OH)₂ synergistically inhibit bacteria mainly by increasing membrane permeability. Ultimately, cytotoxicity assays showed that Rh2 exhibited only low toxicity, the half maximal inhibitory concentration (IC50) of Rh2 was 19.75 μg/mL. This study confirmed the synergistic antibacterial activities of Rh2 and Ca(OH)₂ against E. faecalis. Our findings indicate that the Rh2 and Ca(OH)₂ combination may be a promising alternative approach to treating root canal infections.