Efficacy of Ozone in Inhibiting Biofilm Induced Secondary Caries

Objective: The aim of this study was to evaluate the efficacy of ozone gas on multi-species biofilm to inhibit secondary caries. Methods: Approximately 4x5x4 mm³ sized specimens were prepared from the mid-labial portion of bovine incisors and mid-buccal portion of human molars. Enamel surfaces were flattened with SiC paper and coated with nail varnish. Approximately 2x3x2 mm³ sized dentino-enamel class-I cavity was prepared at the middle of each specimen. Cavities of the specimens were then partially filled with CLEARFILL AP-X with SE-BOND (Kuraray Medical, Tokyo, Japan) keeping 0.8~1 mm deep unfilled space for biofilm accumulation at the top. Artificial biofilms were then formed on the resin composite filled surfaces using multi-species cariogenic bacteria (Streptococcus mutans MT8148, Streptococcus sobrinus 6715 and Streptococcus gordonii ATCC10558) in an oral biofilm reactor for 20 hrs followed by 7 or 30 day incubation period in sucrose added heart infusion broth (Becton Dickinson, Sparks, MD). Gaseous ozone was applied from HealOzone device (Kavo, Germany) for 240 sec, two times a day for 5 days during the first seven days of incubation. Morphology of the biofilm induced lesions at resin-enamel interface was investigated by using a fluorescence microscope (FM; CKX41, Olympus, Japan) and SEM followed by image analysis of the lesions. Data were statistically analyzed by using one way ANOVA and Tukey's HSD method. Results: The caries lesions were clearly visible at the interface of restorations by FM in all samples due the loss of fluorescent mineral particles of enamel and dentin. However, in FM images and data on image analysis, it was appeared that the lesion size was smaller with ozone compared to non-ozone samples and that were confirmed by SEM data. The size of the demineralized enamel at the interface of the resin composite was reduced after ozone treated compared to non-ozone treated specimens. Conclusion: It was appeared that application of gaseous ozone inhibits progression of secondary caries. This study was supported by Center of Excellence (COE) Program for Frontier Research on Molecular Destruction and Reconstruction of Tooth and Bone at Tokyo Medical and Dental University.