Inibição in vitro de biofilme multiespécie da cavidade oral com o óleo essencial de Syzygium aromaticum

Abstract

The oral cavity is an important site regarding the complexity and diversity of its amphibiotic microbiota and the co-adhesion of these microorganisms facilitates the formation of biofilms, especially multispecies biofilms, which are associated with prevalent infections that affect the oral cavity. Therefore, the search for compounds that have good antimicrobial action, with broad spectrum of action is encouraged. The aim of this study was to evaluate, in vitro, the inhibitory activity of Syzygium aromaticum essential oil against the formation of a multispecies biofilm from the oral cavity. The essential oil was characterized according to its phytochemical profile by Gas Chromatography coupled to a Mass Spectrometer and its antioxidant activity was evaluated by the DPPH (2,2 diphenyl-1-picrylhydrazyl) reduction assay. The antimicrobial activity of the essential oil was initially evaluated against strains of Candida and species of bacteria to stablish the Minimum Inhibitory Concentration (MIC), minimum bactericide (MBC) and minimum fungicide (MFC) using the broth microdilution assay, then it was determined the influence of the essential oil on the growth kinetics of Candida species. A biofilm formed with human saliva was used to stablish antibiofilm activity, biofilm morphology by fluorescence microscopy, biofilm viability by CFU / mL count and metabolic activity by XTT reduction assay (2,3-Bis- (2-Methoxy-4-Nitro-5-Sulfophenyl)-2H-Tetrazolium-5-Carboxanilide). The major compound of S. aromaticum essential oil was eugenol (74.54%) and it showed strong antioxidant activity in the first time evaluated (45 minutes). It also showed antimicrobial activity against all Candida species (MIC 500-1000 µg/mL, MFC 1000-2000 µg/mL) and bacteria (MIC 2000 µg/mL, MFC> 2000 µg / mL). The growth kinetics evaluation showed that concentrations 2000 µg / mL (from 2 hours) and 1000 µg/mL (from 4 hours) of the essential oil presented similar results to the positive control (Nystatin). In the morphological analysis of the biofilm it was observed, for both tested concentrations of S. aromaticum a smaller number of structures, which appear fragmented and amorphous, additionally the antibiofilm activity was confirmed by the significant reduction of the number of UFC/ mL when compared to the control and the reduction in biofilm metabolic activity 91.40% (2000 µg/mL) and 92.19% (4000 µg/mL), with no statistical difference between the concentrations evaluated and the positive control of the test (Chlorhexidine). Syzygium aromaticum essential oil inhibits biofilm formation, with no difference from positive control. Its major compound is eugenol, it has expressive antioxidant and antifungal activity, with action on cell growth kinetics similar to the gold standard.