Antibacterial Effect Of Chitosan And Its Derivative On Enterococcus Faecalis

Objective: The aims of our investigation were detections of antibacterial effect of chitosan and N-(2-hydroxyl)propyl-3-trimethyl ammonium chitosan chloride(HTCC)on the Enterococcus faecalis(E.faecalis)strains P25 RC,P52Sa and ATCC 29212,which were isolated from root canal and saliva of patients with refractory apical periodontitis,and standard strain,respectively.Meanwhile,the different antibacterial effect of chitosan and HTCC in different solvents was explored and the possible antimicrobial mechanisms were referred.Cytotoxicity assay was carried to provide reference for the selection of their concentrations in therapy.Methods: Chitosan and HTCC were serially diluted with double distilled water(DDW)or phosphate buffer solution(PBS)at concentrations of 20-2,500 ?g/ml.E.faecalis(P25RC,P52 Sa and ATCC29212 strains)in plankton was treated with chitosan and HTCC solutions,respectively.The positive control group was 2% NaClO,and the negative was DDW or PBS.Colony-forming unit assay was used to calculate the residual bacterium concentration after 24 h.E.faecalis(P25RC strain)biofilms were cultured on coverslips and dentin blocks,and then treated with experimental solutions for 24 h.The biofilms on coverslips and dentine blocks were observed under an inverted fluorescence microscope and scanning electron microscope(SEM),respectively.MC3T3-E1 pre-osteoblasts were seeded into 96-well plates and treated with chitosan or HTCC at various concentrations.The cytotoxicity of chitosan and HTCC on MC3T3-E1 pre-osteoblasts was detected using Cell counting kit-8 after 24,48 and 72 h of treatment.Results: The results revealed that the final minimum bactericidal concentrations(MBC)of chitosan and HTCC dissolved in DDW were 70 and 140 ?g/ml,respectively.Chitosan and HTCC in DDW exerted a significant antibacterial effect,as compared to those in PBS(P<0.05).The antibacterial rate of 78 ?g/ml chitosan in DDW on E.faecalis biofilms was 94%,which was slightly lower than 2% NaClO,and that of 156 ?g/ml HTCC was 79.56%.Cell disruption was happened after treatment of chitosan and HTCC solutions,which was observed under SEM.Chitosan had no cytotoxicity to pre-osteoblasts MC3T3-E1 at a concentration of <625 ?g/ml,while HTCC inhibited the proliferation of cells in this concentration range.Conclusion: Chitosan and HTCC exhibited prominent antibacterial properties on E.faecalis in planktonic state and biofilm,and solvents could affect the antibacterial effect via interfering with the charge interaction.Chitosan and HTCC have potential for application in root canal disinfection and fillings according to the cytotoxicity assay.