Six Spain Thymus essential oils composition analysis and their in vitro and in silico study against Streptococcus mutans.

BACKGROUND: Streptococcus mutans is a well-known oral pathogen that plays a critical role in the development of dental caries. Many studies have been directed to discover the chemical compounds present in natural products to inhibit the growth and biofilm formation activity of S. mutans. Thymus essential oils exhibit good inhibition on the growth and pathogenesis of S. mutans. However, details about the active compounds in Thymus essential oil and the inhibition mechanism still remain unclear. The aim of this study was to investigate the antimicrobial activity of 6 Thymus species (Three samples of Thymus vulgaris, two samples of Thymus zygis, and one sample of Thymus satureioides essential oils) on S. mutans, to identify the potential active components, and to reveal the underlying mechanism. METHODS: The composition of Thymus essential oils was analyzed by gas chromatography-mass spectrometry. And its antibacterial effect was evaluated based on the bacterial growth, acid production, biofilm formation and genetic expression of virulence factors by S. mutans. Potential active components of the Thymus essential oil were identified using molecular docking and correlation analysis. RESULTS: GC-MS analysis showed that the major components in the 6 Spain Thymus essential oils were linalool, α-terpineol, p-cymene, thymol and carvacrol. MIC and MBC analysis showed that 3 Thymus essential oils showed very sensitive antimicrobial activity, and were chosen for further analysis. The 3 Thymus essential oil exhibited a significant inhibitory effect on acid production, adherence and biofilm formation of S. mutans and the expression of virulence genes, such as brpA, gbpB, gtfB, gtfC, gtfD, vicR, spaP and relA. Correlation analysis showed that phenolic components, such as carvacrol and thymol, were positively related to DIZ value, which suggests that they are the potential antimicrobial components. Molecular docking between the Thymus essential oil components and virulence proteins also found that carvacrol and thymol exhibited strong binding affinity with functional domains of virulence genes. CONCLUSIONS: Thymus essential oil showed significant inhibition against the growth and pathogenesis of S. mutans depending on their composition and concentration. And phenolic compounds, such as carvacrol and thymol, are the major active components. Thymus essential oil could be used in oral healthcare products as a potential anti-caries ingredient.

Antibacterial activity of oregano essential oils against Streptococcus mutans in vitro and analysis of active components.

BACKGROUND: Streptococcus mutans (S. mutans) is considered the most relevant bacteria during the transition of the non-pathogenic commensal oral microbial community to plaque biofilms that promote the development of dental caries. Oregano (Origanum vulgare L.), is a universally natural flavoring and its essential oil has been demonstrated to have good antibacterial effects. However, the specific antibacterial mechanism of oregano essential oil (OEO) against S. mutans is still not completely understood. METHODS: In this work, the composition of two different OEOs was determined by GC‒MS. Disk-diffusion method, minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined to assess their antimicrobial effect on S. mutans. The inhibition of acid production, hydrophobicity, biofilm formation and real-time PCR for gtfB/C/D, spaP, gbpB, vicR, relA and brpA mRNA expression by S. mutans were assessed to preliminarily investigate the mechanisms of action. Molecular docking was performed to simulate the interactions with the virulence proteins and active constituents. MTT test using immortalized human keratinocytes cells was also performed to investigate cytotoxicity. RESULTS: Compared with the positive drug Penicillin /streptomycin 100X (DIZ: 34.13 ± 0.85 mm, MIC: 0.78125 µL/mL, MBC: 6.25 µL/mL), the essential oils of Origanum vulgare L. (DIZ: 80 mm, MIC: 0.625µL/mL, MBC:2.5µL/mL) and Origanum heracleoticum L. (DIZ: 39.67 ± 0.81 mm, MIC: 0.625µL/mL, MBC: 1.25µL/mL) could also exhibit similar effects to inhibit the acid production and reduce the hydrophobicity and biofilm formation of S. mutans at 1/2-1MIC concentration. And gene expression of gtfB/C/D, spaP, gbpB, vicR and relA were found to be downregulated. Due to the composition of essential oils from different sources being highly variable, through effective network pharmacology analysis, we found that OEOs contained many effective compounds, like carvacrol and its biosynthetic precursors γ-terpinene and p-cymene, which may directly target several virulence proteins of S. mutans. Besides, no toxic effect was instigated by OEOs at 0.1 µL/mL in the immortalized human keratinocytes cells. CONCLUSION: The integrated analysis in the present study suggested that OEO might be a potential antibacterial agent for the prevention of dental caries.