Eugenol and thymol as potential inhibitors for polymicrobial oral biofilms: An in vitro study

Aim: Dysbiosis of polymicrobial biofilms causes dental caries. In a search for a new effective anticaries agent from eugenol and thymol, this study aimed to investigate the efficacy of eugenol and thymol on polymicrobial (Streptococcus sanguinis, Lactobacillus acidophilus, Actinomyces viscosus, and S. mutans) biofilms. Materials and Methods: Antibacterial and antibiofilm activities were tested using the microdilution method. Antibiofilm activities consisted of inhibiting biofilm formation and degradation of polymicrobial biofilms. Tests were conducted using the microdilution method on 96-well microtiter plates. Tests were done at concentration of 1% v/v, 0.5% v/v, 0.25% v/v, and 0.125% v/v. The compound for biofilm staining was 1% v/v crystal violet, and this study used a microplate reader at a wavelength of 595nm. The minimum biofilm inhibition concentration (MBIC50) and the value of minimum biofilm eradication concentration (MBEC50) were calculated to determine the effectiveness of antibiofilm test compounds against polymicrobial biofilms. The comparative compound used chloramphenicol and Listerine. Scanning electron microscope (SEM) was used to observe the morphological changes in the biofilm after the treatment. Results: Eugenol and thymol showed inhibitory activity against the formation of polymicrobial biofilms. In cells treated with eugenol on polymicrobial biofilms, the matrix of extracellular polymeric substance (EPS) became degraded. Thymol inhibited the biofilms' growth and damaged the EPS which protect bacterial biofilms. Conclusion: Based on these results, it can be concluded that eugenol and thymol have an inhibiting effect on the formation of polymicrobial biofilms at 24h and has a great potential in anticaries.