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Abstract
Objectives
The aim of this research was to determine the pH-dependent changes in F-ATPase activity and proton fluxes in Streptococcus mutans (S. mutans) as induced by varying the concentration of fluoride ±10 mM (0.058% (v/v)) ethanol.
Methods
S. mutans UA159 was grown in Brain Heart Infusion medium at pH 4.8, 6.8, or 8.8. The F-ATPase assay was initiated by the addition of ATP, and stopped by adding 10% trichloroacetic acid. For the proton flux assay, bacterial suspensions were titrated to pH 4.6 with 0.5 M HCl, and then 0.5 M HCl was added to decrease the pH values in units of approximately 0.4 pH. The subsequent increase in pH was monitored using a glass electrode. To disrupt the cell membrane, 10% (v/v) butanol was added to the suspensions after 80 minutes.
Results
At all pH levels, fluoride ±10 mM ethanol not only decreased F-ATPase activity but also increased the proton permeability of S. mutans. The largest effects were observed at pH 4.8. Ethanol enhanced these effects only at pH 4.8.
Conclusions
A very low concentration of ethanol enhanced the action of fluoride on F-ATPase activity and the proton permeability in S. mutans at acidic pH levels. We expect that low concentrations of ethanol may be used together with fluoride and/or other anticaries agents to develop more effective anticaries preparations.
References
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Fig. 1.
Growth curves of S. mutans UA159. The growth rates (c) were calculated as c=Δlog2(OD420)/Δt where OD420 is the optical density at 420 nm, and t is time. The c values of control groups were 0.008 min-1 (T=123 min), 0.019 min-1 (T=53 min) and 0.018 min-1 (T=56 min) at pH 4.8, 6.8 and 8.8, respectively. Data are reported as the mean±SEM of triplicate analysis of three preparations.
Fig. 2.
pH-dependent effects of combined use of fluoride and ethanol on the specific activities of F-ATPase in permeabilized cells of S. mutans UA159. Data are reported as the mean±SEM of triplicate analysis of three preparations. *P<0.05, **P<0.01, ***P<0.001 vs. control (0 ppm F-).
Fig. 3.
pH-dependent effects of combined use of fluoride and ethanol on the proton permeability in S. mutans UA159. Data are reported as the mean±SEM of triplicate analysis of three preparations.
Table 1.
pH-dependent effects of 10 mM ethanol on F-ATPase activity and proton permeability in fluoride-treated S. mutans UA159
|
Probabilitya |
|||
|---|---|---|---|
| pH 4.8 | pH 6.8 | pH 8.8 | |
| F-ATPase activity | 0.042* | 0.454 | 0.228 |
| Proton permeability | 0.032* | 0.642 | 0.914 |
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