Investigation of the antibacterial effects of hinokitiol on Fusobacterium nucleatum

Hinokitiol (HNK), which is known to have stress-relieving properties in living organisms and to show antimicrobial activity against both bacteria and fungi, has been used as a form of aromatherapy in oral care products, such as bath additives, toothpaste, and mouthwash. This study was performed to assess the antibacterial effects of HNK on Fusobacterium nucleatum isolates from patients with periodontitis. F. nucleatum was cultured on Gifu Anaerobic Medium (GAM) for 24 h. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of HNK against F. nucleatum were determined using the liquid dilution method. Furthermore, the bactericidal effects of HNK on F. nucleatum were evaluated using adenosine triphosphate (ATP) bioluminescence assay and flow cytometry. The MIC and MBC of HNK against F. nucleatum were 50 µg/mL and 200 µg/mL, respectively. When HNK solution was added to F. nucleatum cultures adjusted to 106 CFU/mL at the MIC, the ATP levels within the F. nucleatum cells decreased significantly in the experimental group compared with the controls after 24 h. Moreover, staining with propidium iodide (PI) and fluorescence intensity measurements using flow cytometry revealed that the addition of HNK increased the number of dead cells. In summary, HNK demonstrated significant antibacterial effects against F. nucleatum, suggesting its potential as a plant-derived agent for prevention of periodontitis. This study highlighted the potential of HNK as a component of dental care for prevention of periodontitis.