Evaluation of Antimicrobial Mouthrinses Using a Microplate-Based Flow-Through Biofilm Model

Objective: The purpose of this study was to evaluate the antimicrobial potential of commercially available essential-oil (EO), cetylpyridinium chloride (CPC) and chlorhexidine (CHX) mouthrinses using an established multi-treatment biofilm model (MTBM). Methods: Stimulated saliva was collected from donors who refrained from oral hygiene. The saliva was pooled, homogenized and dispensed into a 12 channel bio-cassette. The bio-cassette was attached to a 24 channel continuous flow peristaltic pump system in a 30.5ºC chamber. A 96 peg polystyrene lid was placed in the bio-cassette, and a pellicle formed on the pegs before media was added to the system, allowing biofilms to form for 24 hours. The MTBM consisted of treatments over the course of 60 hours. The MTBM followed mouthrinse label instructions (30 or 60 seconds, once or twice daily). Biofilms subjected to twice daily “rinsing” had 5 hours between treatments. The peg biofilms were harvested immediately after the final treatment by sonication. An aliquot from each well was analyzed for ATP bioluminescence. Results were reported as mean log10 RLU (relative light units) as an expression of ATP concentration. Results: Calculated log10 RLU differences from the water control showed that the EO mouthrinse (EO) produced > 2.5 log₁₀ reduction, the 0.12% CHX mouthrinse (CHX) showed >2.0 log₁₀ reduction and all CPC mouthrinses tested showed <1.7 log₁₀ reduction. Conclusion: These results indicate that in the MTBM, the EO mouthrinse exhibited superior biofilm kill to all CPC rinses tested and similar kill to CHX, well documented for its ability to control plaque and gingivitis. Previous publications cited in the FDA tentative final monograph on oral health care products for plaque and gingivitis have linked CPC efficacy to bioavailability in specific formulations, which may account for results reported here. These data suggest that the EO mouthrinse tested may be more efficacious than the CPC mouthrinses.