Cymbopogon citratus essential oil: effect on polymicrobial caries-related biofilm with low cytotoxicity

Abstract The objective of this study was to evaluate the effects of Cymbopogon citratus essential oil and its main compound (citral) against primary dental colonizers and caries-related species. Chemical characterization of the essential oil was performed by gas chromatography/mass spectroscopy (GC/MS), and the main compound was determined. Antimicrobial activity was tested against Actinomyces naeslundii, Lactobacillus acidophilus, S. gordonii, S. mitis, S. mutans, S. sanguinis and S. sobrinus. Minimum inhibitory and bactericide concentrations were determined by broth microdilution assay for streptococci and lactobacilli reference, and for clinical strains. The effect of the essential oil on bacterial adhesion and biofilm formation/disruption was investigated. Negative (without treatment) and positive controls (chlorhexidine) were used. The effect of citral on preformed biofilm was also tested using the same methodology. Monospecies and microcosm biofilms were tested. ANOVA or Kruskal-Wallis tests were used (α=0.05). Cytotoxicity of the essential oil to human keratinocytes was performed by MTT assay. GC/MS demonstrated one major component (citral). The essential oil showed an inhibitory effect on all tested bacterial species, including S. mutans and L. acidophilus. Essential oil of C. citratus (10X MIC) reduced the number of viable cells of lactobacilli and streptococci biofilms (p < 0.05). The essential oil inhibited adhesion of caries-related polymicrobial biofilm to dental enamel (p < 0.01). Citral significantly reduced the number of viable cells of streptococci biofilm (p < 0.001). The essential oil showed low cytotoxicity to human keratinocytes. Based on these findings, this study can contribute to the development of new formulations for products like mouthwash, against dental biofilms.

Investigação das propriedades anticariogênicas de óleos essenciais: atividade antimicrobiana e caracterização química / Investigation of the anti-cariogenic properties of essential oils: antimicrobial activity, and chemical characterization

O objetivo desse trabalho foi avaliar os óleos essenciais de Thymus vulgaris, Hyptis spicigera, Cymbopogon citratrus, Cymbopogon martinii, Cymbopogon flexuosus bem como os terpenos (citral e cineoli), com relação ao seu potencial para aplicação em terapia anticariogênica. Inicialmente, e a seguir foram realizados testes microbiológicos para a determinação da concentração inibitória mínima (CIM), concentração bactericida mínima (CBM), utilizando os micro-organismos Actinomyces naeslundii ATCC 19039, Lactobacillus acidophilus ATCC 4356, Streptococcus gordonii ATCC 10558, Streptococcus mitis ATCC 9811, Streptococcus mutans ATCC 35688, Streptococcus sanguinis ATCC 10556 e Streptococcus sobrinus ATCC 33478, além de isolados clínicos de estreptococos do grupo mutans e Lactobacillus spp., ambos isolados de dentina cariada. Os três óleos essenciais com maior atividade antibacteriana foram selecionados e caracterizados quimicamente por cromatografia gasosa e avaliados frente ao biofilme de microcosmo, analisando a influência dos óleos essências na adesão de micro-organismo cariogênicos ao esmalte dentário. Além disso, foi analisada a capacidade de eliminação do biofilme de microcosmo sobre a lamínula de vidro. Os resultados revelaram a ação bactericida e bacteriostática de todos os óleos essenciais testados frente às cepas padrão e clínica, com CIMs variando de 0,78 a 3,125μl /mL. Para os terpenos os valores de CIM variaram de 3,125 to 50 μl/mL. No teste de adesão, os resultados apresentam 100% de inibição da adesão do microcosmo em esmalte dentário. Este resultado também foi observado no estudo frente ao biofilme. Diante disso, conclui-se que todos os óleos essenciais e os terpenóides testados possuem atividade promissora frente micro-organismos cariogênicos, além disso, os óleos essenciais de T. vulgaris, H. spicigera e C. citratus, foram capazes de impedir a adesão de micro-organismos bucais ao esmalte dentário

The aim of this study was to evaluate the essential oils of Thymus vulgaris, Hyptis spicigera, Cymbopogon citratrus, Cymbopogon martinii, Cymbopogon flexuosus, as well as the terpenes (citral and cineole),regarding their potential for application in anti-cariogenic therapy. The antimicrobial activity was determined by minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) for the following microorganisms: Actinomyces naeslundii ATCC 19039, Lactobacillus acidophilus ATCC 4356, Streptococcus gordonii ATCC 10558, S. mitis ATCC 9811, S. mutans ATCC 35688, S. sanguinis ATCC 10556 and S. sobrinus ATCC 33478, addition of clinical isolates of mutans streptococci and Lactobacillus sp., both isolated from carious dentin. The three essential oils with higher antibacterial activity were selected and chemically characterized by gas chromatography and evaluated against the biofilm microcosm, analyzing the influence of essential oils on the adhesion of cariogenic micro-organism to dental enamel. Furthermore, we analyzed the capacity of eliminating biofilm microcosm on glass coverslips. The results showed bactericidal and bacteriostatic action of all essential oils tested against standard and clinical strains, with MICs ranging from 0.78 to 3.125 μl / ml. Terpenes for the MIC values ranging from 3.125 to 50 μl / ml. In the adhesion test, the results showed 100% inhibition of the adhesion of tooth enamel in microcosm. This result was also observed when biofilm was evaluated. Therefore, it is concluded that all essential oils and terpenoids tested show promising activity against cariogenic microorganisms, mainly the essential oils of T. vulgaris, H. spicigera and C. citratus, were able to prevent the adhesion of oral microorganisms in dental enamel