High concentrations of HP that promote enamel surface alterations soften the superficial layer of the enamel surface, increase surface porosity, and release more Ca2+ than low concentrations of HP and carbamide peroxide (CP) [4, 15, 17]. Thus, 38% HP has been recruited for the present study to investigate the Ca2+ released from the bleached enamel surfaces after an acidic challenge. With this designated high concentration, it was aimed to observe the maximum Ca2+ release after a further demineralization process.
Alterations in the inorganic component of hydroxyapatite might be an indicator of the changes in Ca2+ levels of enamel. Rotstein et al. demonstrated that most bleaching agents may cause various changes in the levels of calcium, phosphorus, and potassium in dental hard tissues; whereas Tezel et al. demonstrated that 35% and 38% HP caused Ca2+ loss from the enamel surfaces. In the present study, Ca2+ released from the enamel of specimens treated with 38% HP was significantly higher than the untreated control group (p < 0.05). Based on the Ca2+ values, this result suggests that high concentrations of bleaching agents cause surface alterations after an acidic challenge.
The application of highly concentrated fluoride favors the formation of a CaF2-like layer . This deposit is later dissolved, allowing fluoride to diffuse into the underlying enamel, saliva, or a plaque layer covering the tooth. It is assumed that some of the fluoride supports the remineralization of enamel. The results of a previous study confirmed that phosphates and proteins from saliva coated the calcium-fluoride layer on the enamel as a pH-controlling reservoir. This layer acted to decrease demineralization and promote remineralization .
Al-Qunaian et al. investigated the effects of whitening agents on caries susceptibility of human enamel and reported that no significant differences in caries susceptibility were observed between the untreated control specimens and those specimens treated with 10% CP, 20% CP with fluoride, and 35% HP. There were no significant differences between the treated and controlled specimens for teeth treated with 10% CP or 35% HP. However, specimens treated with whitening gel containing 20% CP with fluoride had significantly reduced caries susceptibility when compared with their untreated controls. It was claimed that this effect could be related to fluoride incorporation in 20% CP gels containing fluoride, and the results were in agreement with laboratory studies that fluoride enhanced enamel remineralization.
In the present study, fluoride agents were applied to the bleached enamel and then subjected to further demineralization. When the test groups that were bleached with 38% HP were compared, the decrease in Ca2+ loss of the 1.5% TiF4-treated group was detected to be the lowest (Table 2). Regarding this result, it can be assumed that TiF4 may be effective in protecting the bleached enamel surface against acid attack. Interestingly, no Ca2+ release was detected from three specimens of TiF4 group during the first 4 days, and there was also no Ca2+ release from two specimens during the second 4-day interval (Table 2). We assume that this effect might be the result of the glaze formation after topical TiF4 application. It is known that formation of a glaze layer takes less than 10 s after the application of TiF4. The ability of TiF4 to strongly protect enamel against the action of acid is a synergistic effect of glaze formation and increased enamel fluoride content. The high fluoride content and great reduction in solubility found in TiF4-treated enamel suggests that a fluoride reaction with the enamel is involved . In a previous study, Tezel et al. reported that TiF4 was found to be more effective than Duraphat (NaF, 2.26% F) or Elmex (amine fluoride, 1.25% F) in preventing artificial enamel lesion formation. Attin et al. reported that fluoridation was effective in increasing the resistance of enamel against demineralization by erosive substances. Similarly, the findings of this present study demonstrated that the resistance of bleached enamel against acid attack increased after 1.5% TiF4 treatment.
The comparison of the Ca2+ losses from the test groups that were bleached with 38% HP revealed that the decrease in Ca2+ losses of the 2.1% NaF-treated group was also lower, indicating that NaF could also protect enamel surfaces against acid attack. When the NaF-treated group was compared with the control group, it was seen that the amount of Ca2+ lost from the NaF group was significantly different during the first 4 days (p < 0.05) (Table 3). However, when the NaF-treated group was compared with the 38% HP group, the difference was statistically significant during the whole test period (16 days) (p < 0.05).
However, when the effect of NaF treatment against acid attack was compared with TiF4 treatment, it was observed that its influence was not as strong as TiF4 (Table 3, Figure 1). Tveit et al. assumed that complexes were formed between TiF4 and hydroxyapatite, based on a strong binding of the titanium compound and the oxygen atom of the phosphate group. Mundorff et al. suggested that TiF4 acted with enamel both chemically, by decreasing enamel solubility, and physically, owing to the formation of a protective glaze on the enamel surface. van Rijkom et al. compared the erosion-inhibiting effect of topical fluoride treatment based on the deposition of CaF2-like material using 1% NaF and 4% TiF4. It was concluded that the reduction of Ca2+ loss was more stable for TiF4 than for the NaF group, and the reduction appeared to be smaller with longer acid exposure times.
Generally, fluoride uptake of demineralized enamel is higher when compared with sound enamel . It is assumed that the applied fluoride can easily penetrate through the porous structure of demineralized enamel and that can create a higher number of possible retention sites [27, 28]. According to the results of one study , the bleached and fluoridated enamel was more resistant against erosive attacks than the bleached/unfluoridated and unbleached/unfluoridated enamel. In the present study, at the end of the test period, the total amount of calcium released from the bleached/fluoridated specimens was lower than the control group (unbleached/unfluoridated) and the difference was significant in TiF4-treated specimens (p < 0.05) (Table 3). This result is noteworthy in that bleached and fluoridated teeth may be more resistant to acid attack than sound teeth. These results would need to be investigated with further studies.
In the last decade, there has been a growing interest in demineralization and remineralization studies because of the demand for minimally invasive treatment techniques. When scanning through the literature, we encountered a number of different techniques applied in these types of studies. In vitro demineralization using acid buffers and in vitro demineralization/remineralization using a pH-cyling model are the most frequently used techniques that possess both advantages and disadvantages. It is important to choose the simplest and most practically appropriate model. Similar to the previous studies, we preferred using AAS for in vitro demineralization to observe the impact of fluoride agents on the Ca2+ loss following further demineralization. This method is a very sensitive but reliable method for calcium analysis, which avoids the interaction of other solutes [30, 31]. It can be used with confidence to quantify erosion of both enamel and dentine, and their chemical analyses of mineral release [32–34].
Based on the results of the present study, the null hypothesis that states there would be no difference between the amount of Ca2+ released from the enamel surfaces that were treated with NaF and TiF4 after an acidic challenge was rejected. It was also shown that topical fluoride application decreased the amount of Ca2+ released from the 38% HP-treated enamel surfaces after further demineralization. TiF had a significantly more pronounced effect than NaF in protecting enamel surfaces against acidic attack after bleaching with 38% HP.