Cymbopogon Citratus (Lemongrass Oil) Oral Sprays as Inhibitors of Mutans Streptococci Biofilm Formation

Introduction: Dental caries that occurs in young children or Early Childhood Caries (ECC) remains highly prevalent worldwide. Streptococcus mutans is not only one of the pioneer groups in plaque formation but is also crucial for its continuous development. It is important to prevent the presence of Mutans Streptococci (MS) for more effective prevention of dental caries. Aim: To evaluate the susceptibility of Streptococcus mutans (ATCC 25175), Streptococcus sobrinus (ATCC 6715) and three Streptococcus mutans clinical isolates from Thai children to three oral spray formulations (2%L38, 4%L40, 6%L42) of Cymbopogon citratus extracted oil, and evaluate the inhibitory effect of each formulation on growth and biofilm formation of each strain. Materials and Methods: S. mutans clinical isolations were obtained from children (12 males, 5 females aged 3.5-10-years-old) who came to the Paediatric Dental Clinic, Faculty of Dentistry, Chulalongkorn University, Bangkok. The inhibitory effect of each formulation on growth and biofilm formation was performed using a 96-well polystyrene plate biofilm assay. The inhibition effect was further analysed using fluorescent dyes (SYTO9 and propidium iodide) and Confocal Laser Scanning Microscopy (CLSM). All experiments were performed in triplicate. Data was expressed as mean +/- SD. For the determination of live/dead cells at the excitation wavelength of 485 nm, the green emission at 530 nm (live cells) and the red emission at 630 nm (dead cells) were measured and their ratio was calculated. The Kruskal-Wallis Test (Bonferroni and Dunn Test) was used to compare the experimental and control groups (significance at 95%). Results: All oral spray formulations showed excellent antibacterial activity. Results showed that 3 and 6 hours. preformed biofilms of all strains were sensitive to all formulations (100%) when compared to the controls. Biofilm formation of all strains was inhibited (100%) when compared to the controls by all formulations at different concentrations. Six percent (L42) oil formulations showed the maximum inhibition effect, followed by 4% (L40) and 2% (L38), respectively. Results from CLSM analysis of S. mutans (ATCC 25175) showed that 6% (L42) oil formulations gave the maximum inhibition effect, followed by 4% (L40) and 2% (L38), respectively. For S. sobrinus (ATCC 6715), 2% (L38) oil formulations showed the maximum inhibition effect, followed by 4% (L40) and 6% (L42). For all S. mutans clinical isolates, all formulations inhibited biofilm effectively and were not significantly different. Conclusion: In this study, all oral spray formulations from Cymbopogon citratus essential oil showed excellent antibacterial activity and they might be an alternative inexpensive natural medication to prevent dental caries in the near future.