Addition of hydrogen peroxide to methylene blue conjugated to β-cyclodextrin in photodynamic antimicrobial chemotherapy in S. mutans biofilm.

Abstract

This study evaluated the effect of hydrogen peroxide addition on β-cyclodextrin-conjugated methylene blue in antimicrobial photodynamic therapy(a-PDT) in S. mutans biofilm model using laser or light emitting diode (LED) (λ = 660 nm). A preliminary assay was performed to evaluate the cytotoxicity of hydrogen peroxide in oral fibroblasts by the colorimetric method (MTT). Afterwards, groups were divided into (n = 3, in triplicate): C (negative control), CX - chlorhexidine 0.2% (positive control), P (methylene blue/β-cyclodextrin), H (Hydrogen Peroxide at 40 μM), PH, L (Laser), LP, LH (Laser+Hydrogen Peroxide), LPH, LED, LEDP, LEDH, and LEDPH. The biofilm was formed in 24 h with BHI + 1% sucrose (w/v). Light irradiations were conducted with laser, 9 J, 323 J/cm2, 113 s or with LED, 8.1 J, 8.1 J/cm2 for 90 s. Microbial reduction was evaluated by counting the viable microorganisms of the biofilm after the respective treatments, in a selective culture medium, and laser confocal microscopy evaluation. LP, LH, LPH, LEDP, LEDH, and LEDPH groups statistically reduced the counts of S.mutans compared with the C group and the log reductions were of 1.87, 1.94, 2.19, 0.91, 0.92, and 1.33, respectively; the addition of hydrogen peroxide did not potentiate the microbial reductions (LPH and LEDPH) compared with the LP and LEDP groups. The association of hydrogen peroxide with the conjugated β-cyclodextrin nanoparticle as photosensitizer did not result in an enhanced effect of a-PDT; hydrogen peroxide behaved as a photosensitizer, since it reduced the number of S. mutans when associated with laser light.