In vitro antimicrobial activity of chitosan mouthwash against Streptococcus mutans and Lactobacillus casei

Abstract

Objective: The aim of this study was to investigate the antimicrobial effect of chitosan mouthwash against Streptococcus mutans and Lactobacillus casei in the laboratory.

Materials and Methods: The chitosan mouthwashes were prepared from 3 different molecular weights (30 kDa, 890 kDa and 1,700 kDa). The solutions of the chitosan mouthwash were prepared for 10 mg/ml, 5 mg/ml, 2.5 mg/ml and 1.25 mg/ml by dissolving chitosan powder with 3 molecular weights in each concentration. Chitosan mouthwash was formulated from chitosan powder, acetic acid, saccharin, blueberry flavor, sodium hydroxide and glycerol. Streptococcus mutans and Lactobacillus casei were cultured on BHI agar and MRS agar respectively and were transferred to 5 ml of BHI broth and MRS broth respectively and incubated for 24 hours. Measure the optical density (OD) of both bacteria and adjusted the initial load of bacteria to 2x10⁶ CFU/ml. Determined the antimicrobial effect of chitosan mouthwash against both bacteria using time-dependent killing assay by viable count technique with various concentrations of 5mg/ml, 2.5 mg/ml and 1.25 mg/ml. Samples were collected at 0 hour, 30 mins, 2 hours, 6 hours, and 12 hours. The inhibition of bacterial growth was determined by standard plate count technique. The tube with 0.12% chlorhexidine mouthwash was used as positive control and mouthwash solution without chitosan and Phosphate buffer saline (PBS) were used as negative controls.

Result: The results showed 5mg/ml, 2.5mg/ml and 1.25mg/ml of the chitosan mouthwash have bactericidal effect against Streptococcus mutans and Lactobacillus casei. All concentrations of the chitosan mouthwash showed an ability to inhibit the bacterial growth more than 3 log CFU/ml against Streptococcus mutans within 12 hours. For Lactobacillus casei, the lowest concentration that could inhibit growth of bacteria more than 3 log CFU/ml within 12 hours is 2.5 mg/ml. The lower molecular weight (30 kDa and 890kDa) of chitosan mouthwash could inhibit the growth of Streptococcus mutans more than the higher molecular weights (1700 kDa). Interestingly, the higher molecular weight (890 kDa and 1,700 kDa) of chitosan mouthwash could inhibit the growth of Lactobacillus casei more than the lower molecular weight (30 kDa).