Calcium phosphate deposition in human dental plaque microcosm biofilms induced by a ureolytic pH-rise procedure.
ABSTRACT The objectives were to develop and characterize a procedure based on a ureolytic pH rise to deposit calcium phosphate into microcosm dental plaque biofilms and to test the importance of the plaque pH range. Plaque biofilms were cultured in a multiplaque culture system ('artificial mouth') with a continuous supply of a simulated oral fluid (basal medium mucin; BMM) with 146 mmol/l (5% w/v) sucrose periodically applied over 6 min every 8h. After initial plaque growth, the biofilms were periodically exposed for up to 16 days to 6-min applications of calcium phosphate monofluorophosphate urea (CPMU) solution containing 20 mmol/l CaCl(2), 12 mmol/l NaH(2)PO(4), 5 mmol/l monofluorophosphate and 500 mmol/l urea (pH 5.0). Three application regimes were examined, one included a sucrose-induced acidic pH fluctuation. Plaque hydrolysis of the urea in CPMU caused the pH to rise to between 8.2 and 8.8, depositing fluoridated and carbonated calcium phosphates, and possibly some calcium carbonate, into the plaque. Calcium, phosphate and fluoride deposition was rapid for about 4 days and then slowed. After 10 days' treatment under standard conditions (BMM containing 1 mmol/l urea and 1 mmol/l arginine), plaque calcium and phosphate concentrations had increased up to 50-fold and 10-fold to approximately 2-4 and 1-2 mmol/g plaque protein, respectively. The calcium, phosphate and fluoride content increased steadily. Calcium phosphate deposition was proportional to the plaque resting pH, increasing over four-fold when the BMM urea concentration was increased from 0 to 20 mmol/l, which raised the resting pH from 6.4 to 7.2 and yielded a mean plaque calcium concentration of 14.3 mmol/g protein, one subsample reaching 20.8 mmol/g protein. Supplementation of BMM with 20% human serum inhibited deposition. These results support the hypothesis that an alkaline pH in plaque is critical in promoting plaque mineralization and that mineral deposition is modulated by serum. These factors are likely to be important in regulating calculus formation.
Article: Mechanisms of inhibition by fluoride of urease activities of cell suspensions and biofilms of Staphylococcus epidermidis, Streptococcus salivarius, Actinomyces naeslundii and of dental plaque.[show abstract] [hide abstract]
ABSTRACT: Fluoride is known to be a potent inhibitor of bacterial ureases and can also act in the form of hydrofluoric acid as a transmembrane proton conductor to acidify the cytoplasm of intact cells with possible indirect, acid inhibition of urease. Our research objectives were to assess the inhibitory potencies of fluoride for three urease-positive bacteria commonly found in the mouth and to determine the relative importance of direct and indirect inhibition of ureases for overall inhibition of intact cells or biofilms. The experimental design involved intact bacteria in suspensions, mono-organism biofilms, cell extracts, and dental plaque. Standard enzymatic assays for ammonia production from urea were used. We found that ureolysis by cells in suspensions or mono-organism biofilms of Staphylococcus epidermidis, Streptococcus salivarius or Actinomyces naeslundii was inhibited by fluoride at plaque levels of 0.1-0.5 mm in a pH-dependent manner. The results of experiments with the organic weak acids indomethacin and capric acid, which do not directly inhibit urease enzyme, indicated that weak-acid effects leading to cytoplasmic acidification are also involved in fluoride inhibition. However, direct fluoride inhibition of urease appeared to be the major mechanism for reduction in ureolytic activity in acid environments. Results of experiments with freshly harvested supragingival dental plaque indicated responses to fluoride similar to those of S. salivarius with pH-dependent fluoride inhibition and both direct and indirect inhibition of urease. Fluoride can act to diminish alkali production from urea by oral bacteria through direct and indirect mechanisms.Oral Microbiology and Immunology 01/2006; 20(6):323-32. · 2.81 Impact Factor