The present study aims to evaluate the effect of different Tyrosol concentrations on the hydrolytic enzymes production by Candida albicans and Candida glabrata, on the lactic acid production by Streptococcus mutans and on the ability to form single and mixed biofilms of the three species mentioned. First, according to the microdilution method, it will determined the minimum inhibitory concentration of Tyrosol capable of inhibiting the growth of each strain in the planktonic state. After, single and mixed biofilms of the three species will be formed (for 48 hours) in the presence of different Tyrosol concentrations on specimens of acrylic resin and hydroxyapatite within 24-wells microtiter plates. Subsequently, the enzymatic activity (proteinase, phospholipase and hemolytic) of Candida species will be evaluated using specific culture medium for each type of hydrolytic enzyme, while the lactic acid production by S. mutans will be measured by the Clinpro Cario L-Pop Kit. The effect of Tyrosol on the biofilm formation ability will be quantified by counting the number of culturable cells, total biomass (crystal violet staining assay) and metabolic activity of the cells. Moreover, the effect of Tyrosol on preformed biofilms will be assessed through the previously mentioned assays. Biofilm structure after treatment with Tyrosol will be visualized by scanning electron microscopy. Controls devoid Tyrosol will be included in all analyzes, and all tests will be performed in triplicate on three different occasions. Data will be analyzed to employ the most appropriate statistical test. P < .05 will be considered statistically significant.
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