A number of antiplaque agents have been available in dentifrices to decrease the bacterial number of dental plaque. S. mutans and L. casei were most closely associated with caries in humans. We have previously studied the in vitro mono-cultured microbial model. This study was performed to advance this model to the di-cultured microbial model into be suitable for testing the effect of dentifrices containing antiplaque agents, and to evaluate the efficacy of the di-cultured microbial model.
S. mutans and L. casei were cultured in tryptic soy broth medium for 14 hours at 37C, 5% CO;t. These bacteria were at the exponential phase for 12 hours. After culturing for 12 hours, the density was 7.57 Log,oCFU/ml for S. mutans, 7.51 Log,oCFU/ml for L. casei, and the optical density at 540 nm was 0.359, 0.258, respectively.
These ´Acteria were cultured in 7´S-MRS broth medium with 5% sucrose according to the mixed ratio of 1:1, 1:1/100, and 1/100:1 :for each bacterium. The growth of S. mutans and L. casei was not disturbed each other in the mixed culture, being the same growth rate as mono-cultures. Therefore, we selected the same density of bacteria in the mixed culture.
Incipient caries lesion was created by the solution containing O.1M lactic acid, 0.2% carbopol, and 50% saturated HAP for 44 hours at 37C. Specimen was selected in the range of 25VHN - 45VHN. Salivary pellicle was formed by immerging in human whole saliva for 24 hours. Plaque was formed on the enamel surface by the mixed culture in TS-MRS with 5% sucrose for 3 days. This medium contained the salivary salt or not, and the interval of medium change was 8 hours or 12 hours. Specimen hardness did not significantly changed and the density of bacteria adhered to enamel surface was 6.300.19 LogioCFU/mm2 for S. mutans, 5.040.55 Log,oCFU/mm2 for L. casei when medium with the salivary salt was changed at 8 hours interval for 3 days.
Salivary pellicle was formed on the enamel specimens at the above condition. Treatment regimens were a placebo, 1,100ppm F NaF/silica dentifrice, and 1,100ppm F NaF/silica plus 0.3% triclosan dentifrice. Artificial saliva contained gastric mucin(0.220/o), NaCl(0.038%), CaC12. 2H2O(0.02130/o), KH2PO4(0.0738%), and KCl(0.1114%). The specimens were treated with the artificial saliva, mixed whole saliva and dentifrice, and TS-MRS broth with 5% sucrose sequentially according to the microbial cyclic sequence for 15 days.
The delta VHNs of specimens treated with 1,100ppm F NaF/silica dentifrice plus 0.3% triclosan were statistically higher than those treated with 1,100ppm F NaF/silica dentifrice or placebo dentifrice. And S. mutans was fewer adhered to those treated with 1,100ppm F NaF/silica dentifrice plus 0.3% triclosan than. those treated with placebo dentifrice. L. casei was fewer adhered to those treated with 1,100ppm F NaF/silica dentifrice plus 0.3% triclosan than those treated with 1,100ppm F NaF/silica dentifrice or placebo dentifrice.
In. conclusion, it is suggested that the in vitro di-cultured microbial pH cycling model used in this study may be suitable for testing the effect of the dentifrices containing antiplaque agents.
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