RESUMO
AIM: Black extrinsic tooth stain (BETS) is a health challenge that commonly affects children. Aggregatibacter actinomycetemcomitans (Aa) presents in higher prevalence within the polymicrobial community of BETS. In this study, the anti-planktonic and anti-sessile activities of cinnamon essential oil (CEO) and its individual compounds against Aa were evaluated. The preventive effect of CEO and its active substances on BETS formation was also studied in vitro. METHODS: Aa was isolated from a preschool child with BETS and was identified based on the morphological characteristics, MALDI-TOF mass spectroscopy and 16S rRNA sequencing. The effect of CEO and its individual compounds on the growth kinetics of planktonic and sessile Aa cells as well as their antibacterial efficacy and their rate of bacterial killing were examined. The preventive effect of CEO and its active substances on the formation of BETS was evaluated using an ex vivo model. The data were analysed using one-way analysis of variance (ANOVA) and the significance level was set at p < 0.05. RESULTS: Out of eight individual compounds of CEO, only eugenol, cinnamaldehyde and α-methyl cinnamaldehyde showed anti-Aa activities. The values of the minimum inhibitory concentrations (MICs) were in the following order: CEO (421.5 mg/ml) > α-methyl cinnamaldehyde (26.37 mg/ml) > cinnamaldehyde (0.209 mg/ml) > eugenol (0.052 mg/ml). CEO, eugenol, cinnamaldehyde and α-methyl cinnamaldehyde, respectively, exhibited two-, four-, four- and eightfold increase of sessile MIC compared to their planktonic MIC. The growth kinetics of both planktonic and sessile Aa in the presence of CEO, eugenol, cinnamaldehyde and α-methyl cinnamaldehyde revealed a complete inhibition at the MICs and 5.3%-37.4% biofilm inhibition at sub-MICs. The time-killing study demonstrated that CEO, eugenol and cinnamaldehyde were capable of reducing the survival rate of both planktonic and sessile Aa cells after 15-20 and 25-30 min, respectively. However, α-methyl cinnamaldehyde showed a superior anti-planktonic to anti-biofilm activity. The daily incorporation of CEO, eugenol and cinnamaldehyde at their MICs for 14 days totally prevented the formation of BETS in the ex vivo model; however, in the case of α-methyl cinnamaldehyde, BETS was visually detectable after 10 days. CONCLUSION: CEO and its individual compounds have marked antibacterial activity against Aa. The effective results against planktonic and sessile Aa within reasonable time indicate that they can be used to prevent BETS.
