Fluoride rinse effect on retention of CaF 2 formed on enamel / dentine by fluoride application

Calcium fluoride-like materials (“CaF2”) formed on dental surfaces after professional fluoride application are unstable in the oral environment but can be retained longer with a daily NaF mouthrinse. We tested the effect of twice daily 0.05% NaF rinses on the retention of “CaF2” formed on enamel and dentine after applying acidulated phosphate fluoride (APF). “CaF2” formed on enamel/dentine by APF application significantly decreased after exposure to artificial saliva and the 0.05% NaF rinse was ineffective to avoid this reduction. These findings suggest that the combination of APF and 0.05% NaF is not clinically relevant, either for caries or dental hypersensitivity.


Introduction
The anticaries benefit of acidulated phosphate fluoride (APF) application is well established. 1It results in the formation of a high concentration of calcium fluoride-like materials ("CaF 2 ") on enamel and dentine surfaces, which has been considered responsible for the anticaries effect of professionally applied topical fluoride treatments. 2 Nevertheless, 80%-90% of "CaF 2 " formed is dissolved in the oral environment during the first week because saliva is undersaturated with respect to "CaF 2 ". 3 Therefore, retaining "CaF 2 " on enamel or root dentine for a prolonged period of time is desirable, mainly for patients with a high risk for caries. 4he dissolution of "CaF 2 " formed on enamel and dentine after applying APF could be decreased by the daily use of fluoride products, such as a 0.05% NaF mouthrinse.A 0.05% NaF mouthrinse may compensate for the dissolution of "CaF 2 " because it contains 225 ppm F, which forms a small amount of "CaF 2 " on enamel-dentine; however, this subject has not been explored.
Therefore, we evaluated if twice daily exposure to 0.05% NaF solution in vitro (simulating F mouthrinse) would prolong the retention of "CaF 2 " formed on enamel and dentine after applying APF.

Methodology Experimental design
An in vitro 2 × 3 factorial study was conducted, including the following factors: 1) APF application at two levels, i.e., applied (APF + ) or not (APF − ) Declaration of Interests: The authors certify that they have no commercial or associative interest that represents a conflict of interest in connection with the manuscript.and 2) 0.05% NaF treatment at three levels, i.e., not treated (NT), treated with NaF during saliva immersion (NaF + ), or treated with purified water (NaF − ) during saliva immersion.

Preparation of enamel and dentine slabs
The enamel and dentine slabs (3 × 3 × 2 mm) were obtained from sound bovine incisors, and their external surfaces were polished flat.Then, the slabs were isolated with wax; thus, only the external surface of the enamel or dentine was exposed to the treatment.

Treatments
The enamel and dentine slabs were immersed into either the control or APF solution for 4 min under agitation (120 rpm).The slabs were washed for 30 s with a stream of purified water and gently dried with absorbent paper.The slabs in the APF − /NT and APF + /NT groups were immediately analyzed for alkali-soluble F concentration.The remaining slabs were individually immersed in artificial saliva 5 (1.7 mL/mm 2 ) and maintained at 37°C for 7 days.The slabs from both groups were exposed to purified water or a 0.05% NaF solution for 1 min under agitation (120 rpm) twice daily at 7 am and 5 pm.Following each treatment, the slabs were flushed with purified water (10 s), dried, and immersed again in artificial saliva.The artificial saliva was changed every morning after treatment.

Determination of "CaF 2 "
Slabs were individually immersed in 1 M KOH (0.4 mL/block) for 24 h under agitation.An additional 6-h extraction with 1 M KOH (0.3 mL/block) was performed to deplete all alkali-soluble fluoride.The extraction solution was buffered with TISAB II containing 1 M HCl.Fluoride was measured with an ion-selective electrode (Orion 96-09; Thermo Scientific, Inc.) and an ion analyzer (Orion EA-940, Thermo Scientific) against standards prepared as the samples, and the alkali-soluble fluoride concentration (sum of the two alkali extractions) was expressed as µg F/cm 2 .

Statistical analysis
A factorial 2 × 3 analysis was conducted.Enamel and dentine data were independently analyzed.The alkali-soluble fluoride concentrations in the enamel and dentine slabs were transformed to square root and log10, respectively.Tukey test was used for the post-analysis of variance comparisons.The data were analyzed using SAS software version 8.01 (SAS Institute, Cary, USA), and a p-value of < 0.05 was considered significant.

Results
APF resulted in significantly higher "CaF 2 " concentrations in the enamel (Table 1) and dentine (Table 2) slabs (p < 0.05) than the control."CaF 2 " concentrations significantly decreased by 85 and 90% in the enamel and dentine after exposure to saliva and purified water, respectively (p < 0.05).The "CaF 2 " concentration retained in the enamel and dentine after treatment with 0.05% NaF did not differ from the control group treated with water (p > 0.05).

Discussion
Our results confirmed that loosely-bound f luoride ("CaF 2 ") was formed after APF was applied to enamel, 2 but also showed that the same occurs with dentine.The "CaF 2 " concentration formed on dentine was 7-fold higher than that on enamel, which may be explained by the smaller hydroxyapatite crystals in dentine resulting in a larger surface area to crystallite volume ratio and therefore a more reactive mineral phase. 6dditionally, dentine is more acid-soluble than enamel, 7 resulting in more calcium being released by the APF treatment, which reacts with fluoride and precipitates as "CaF 2 ".Nevertheless, our data confirmed that "CaF 2 " formed on the enamel and dentine was not stable in a saliva-like solution. 3CaF 2 " concentration decreased similarly (85%-90%) in enamel and dentine after 7 days of exposure to artificial saliva, and the 0.05% NaF twice daily treatment did not prevent this decrease.However, the amount of "CaF 2 " remaining was higher than the concentration found in slabs pre-treated with a non-fluoridated control solution (APF − ).The findings for enamel show that these reservoirs may last for a significant period of time, as previously shown in vitro 3 and in situ 8 and the same is expected for dentine.
The finding that the 0.05% NaF rinse did not prevent the dissolution of "CaF 2 " formed by APF may be explained by the very small amount of fluoride deposited on sound dental tissues from a mouthrinse. 9n fact, the twice daily NaF treatment significantly increased alkali-soluble fluoride concentrations on the dentine slabs that were not pre-treated with fluoride, but the concentration after 7 days was only approximately 1% of that resulting from the APF treatment (Table 2).
The present results are also relevant when APF is recommended as a dentine hypersensitivity treatment. 10Although "CaF 2 " may block dentinal tubules and decrease sensitivity for some time, the daily use of a fluoride mouthrinse may not be suitable to prolong this effect.

Conclusion
Our findings suggest that 0.05% NaF treatment twice daily does not prevent oral dissolution of "CaF 2 " formed on enamel or dentine by APF application.