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Keywords:

  • Bioactive glass;
  • bleaching;
  • demineralization;
  • enamel;
  • remineralization

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Conclusions
  8. References

Background:  The aim of this study was to determine the effects of bleaching on the structure of the enamel layer of teeth and the potential of the commercial bioactive glass NovaMin® in two different toothpastes to remineralize such regions of the enamel. Three aspects were considered: the extent and nature of the alterations in the enamel after application of the bleaching agents; the extent of remineralization after application of two commercial toothpastes containing bioactive glass; and whether or not there were differences between the toothpastes in terms of their effectiveness in promoting remineralization.

Methods:  Bleaching agent based on 16% carbamide peroxide was applied to the enamel surface of freshly extracted human molars for 8 minutes, once a day for 7 days. After the bleaching cycles, the enamel surface was analysed by SEM and EDX.

Results:  The results obtained in the study lead to the conclusion that application of 16% carbamide peroxide causes distinct morphological changes to the enamel surface which vary from mild to severe. Subsequent treatment with either of the toothpastes containing the bioactive glass NovaMin® resulted in the formation of a protective layer on the enamel surface, consisting of bioactive glass deposits, with only slight differences between the two brands. Application of these dentifrices also caused increases in the Ca and P content of the enamel layer, returning it to that of undamaged enamel.

Conclusions:  Remineralizing toothpastes should be used after bleaching, in order to repair any damage to the mineral tissue caused by these procedures.


Abbreviations and acronyms:
CP

carbamide peroxide

HP

hydrogen peroxide

Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Conclusions
  8. References

Bleaching has become a popular procedure with patients seeking improvements in the perceived appearance of their teeth. Generally, bleaching agents are based on either hydrogen peroxide (HP) or carbamide peroxide (CP). Their activity leads to improvement of the discoloured tooth structure through decomposition of peroxide into free radicals1 which break down large pigmented molecules. It is the chromophores within these large molecules which absorb light in the visible region, and are thus responsible for the colour stain in the enamel. Fragmentation of these molecules into smaller species by free radicals alters the light absorption and thus reduces or eliminates the stain.2

Carbamide peroxide is a chemical adduct of urea and HP, which upon dissolving in water or saliva dissociates back into HP and urea. Thus, CP can be considered as a precursor of the active bleaching species HP.1–4

Although the bleaching treatment offers aesthetic improvement, the effect of bleaching agents on dental hard tissues is still being debated. Some studies have claimed that there are no significant effects of bleaching agents on human enamel.5 However, other studies have obtained contrary results, with treated enamel showing morphological changes suggesting that bleaching is an erosive process.6–9 Bleaching directly affects the organic (protein) content of the tooth, but this leads to changes in the mineral phase, resulting in the observed morphological changes to the tooth surface.10

It may be possible to reverse this damage by the deployment of mineralizing agents to treat the affected tooth surface. One type of mineralizing agents are the bioactive glasses. These materials are capable of bonding chemically to hard dental tissues and their components are oxides of calcium, sodium, phosphorus and silica in ratios that impart bioactivity. In vivo, these glasses are able to form a layer of hydroxyapatite on their surface.11,12

A commercial bioactive glass that has been used in the treatment of dental hypersensitivity and enamel remineralization is NovaMin®, a material which was originally developed as a bone regeneration material. NovaMin® is a ceramic material consisting of amorphous sodium-calcium-phosphosilicate which is highly reactive in water, and consists of a fine particle size powder which can physically occlude dentinal tubules.13–15 In the aqueous environment of the tooth, sodium ions from the NovaMin® particles rapidly exchange with hydrogen cations (in the form of H3O+), and this brings about the release of calcium and phosphate (PO43−) ions from the glass.16,17 A localized, transient increase in pH occurs during the initial exposure of the material to water due to the release of sodium. This increase in pH helps to precipitate the extra calcium and phosphate ions provided by the NovaMin® to form a calcium phosphate layer. As these reactions continue, this layer crystallizes into carbonate-enriched hydroxyapatite (HCA).18 The combination of the residual NovaMin® particles and the newly formed HCA layer results in remineralization of the enamel surface and prevents further demineralization.19

Several NovaMin® containing toothpastes, such as Dr. Collin’s Restore™ (available in USA and Australia), X-PUR® (Canada) and Vantej (India) have recently been introduced to the market worldwide. In the present study, two commercial toothpastes which contain NovaMin® glass particles were used, namely Mirawhite® tc and Nanosensitive® hca (both manufactured by Hager & Werken, GmbH & Co KG, Germany) in order to provide potential remineralizing formulations. Their overall ability to repair the enamel surface following bleaching was examined.

The purpose of the present study was to determine the effects of bleaching on the structure of tooth enamel and explore the role of NovaMin® in toothpastes for its potential to remineralize any damaged regions of the enamel. In addition, any differences in effectiveness between the two commercial toothpastes were examined.

Materials and methods

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Conclusions
  8. References

Fifteen young immature permanent human molars, extracted for orthodontic reasons, were used in the study. The roots were cut with a high-speed dental handpiece at the level of the cemento-enamel junction, and the remnants of the pulp tissue discarded. The coronal segments were thoroughly ultra-sonicated and polished with pumice and polishing toothpaste. The excess toothpaste was cleaned by water-spray for 3 minutes. The teeth were randomly divided into 3 groups, consisting of 5 teeth each and stored in physiological saline during the experimental period. All of the groups were treated with Mirawhite® Pro (Hager & Werken, GmbH & Co KG, Germany), a commercial bleaching agent which contains 16% CP, for 8 minutes a day for 7 days. After each bleaching cycle, all of the teeth were cleaned by water-spray for 3 minutes. The first group served as a control, while the second and the third groups were treated for possible remineralization. The second group was treated with Mirawhite® tc (Hager & Werken, GmbH & Co KG, Germany), which contains 5.5% NovaMin® and the third with Nanosensitive® hca (Hager & Werken, GmbH & Co KG, Germany) which contains 7% NovaMin® for 5 minutes. Afterwards, the dentifrices were cleaned with a toothbrush for 5 minutes under copious water-spray to eliminate possible leftovers.

The teeth were cut in half along the longitudinal axis in a vestibulo-oral direction. One half of each sample was gold-sputtered and analysed under SEM (Cambridge Stereoscan 360 High-Resolution Scanning Electron Microscope, Cambridge Instruments Co., UK), while the other half was cast in Epo-Thin resin (Buehler®, USA, Batch No. 20-8140-032) with the cut surface facing the bottom of the mould. The cast specimens were cured in a vacuum desiccator for 24 hours, polished with different sizes of carborundum grits up to 1 μm diamond, carbon coated and analysed under SEM (Jeol, JSM 5310LV Scanning Electron Microscope, Japan). Also, scans of the enamel surface and linescans along the line that goes through the enamel into the resin were collected (ISIS 300 Systems, Oxford Instruments Co., UK). Finally, quantitative EDX point analysis (ISIS 300 Systems, Oxford Instruments Co., UK) was performed on the enamel surface to determine the elemental levels (%) of strontium (Sr), sodium (Na), magnesium (Mg), aluminium (Al), silicon (Si), phosphorus (P) and calcium (Ca). For each sample, five points were selected and the mean values calculated. The statistical analysis was performed by one-way ANOVA. When statistically significant differences appeared (at the level of significance p < 0.05), the post hoc– Tukey’s honest significant difference test was applied.

Results

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Conclusions
  8. References

The SEM micrographs of the first group, treated with bleaching agent only, show changes in the enamel surface that suggest that a demineralization process has occurred. This varies in severity from mild, i.e. increased surface porosity (Fig 1a) and slight etching with increased surface irregularities (Fig 1c, d), to severe, i.e. clear destruction of the enamel surface resulting in significant depressions and irregularities (Fig 1b, e).

image

Figure 1.  Representative SEM photomicrographs of the specimens from the control group: enamel surface. (a) increased depth of enamel irregularities; (b) depressions; (c)–(d) areas of increased surface porosity, etching like appearance; (e) erosions; (f) longitudinal section of the tooth – surface demineralization is visible; (g) scan of the enamel surface – presence of C, O, Ca and P; (h) line scan through the enamel surface – arrows towards the declined levels of Ca and P in the superficial zones of the enamel (E-enamel, R-resin).

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The two groups treated first with bleaching agent then with one of the potential remineralizing agents are characterized by completely different surface appearances (Figs 2 and 3). In both cases, the SEM micrographs indicate that deposits of bioactive glass particles are dispersed throughout the enamel surface and almost completely seal the irregularities created by the bleaching agent (Fig 2e). The sample treated with Nanosensitive® hca (presented in Fig 3a, b) has a ‘flower-field’-like appearance, with amorphous deposits that seem to be growing out of the enamel. The longitudinal sections demonstrate the formation of a superficial protective layer consisting of bioactive glass deposits in the experimental groups.

image

Figure 2.  Representative SEM photomicrographs of the specimens from the Mirawhite® tc group: (a)–(d) amorphous deposits on the enamel surface; (e) incorporation of bioactive glass particles onto the enamel surface; (f) longitudinal section of the tooth – surface deposition of bioactive glass; (g) scan of the enamel surface – presence of C, O, Ca, Ti, Si, Cl and P; (h) linescan through the enamel into the resin: arrows – surface deposits firmly attached to the enamel surface and composed of Ca and P (E-enamel, R-resin).

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image

Figure 3.  Representative SEM photomicrographs of the specimens from the Nanosensitive® hca group: (a)–(b) loss of interprismatic substance, while enamel rods are preserved; (c)–(e) bioactive glass particles deposited on the enamel surface; (f) longitudinal section of the tooth – surface deposition of bioactive glass; (g) scan of the enamel surface – presence of C, O, Ca, Na, Si, Cl and P; (h) linescan through the enamel into the resin: arrows towards the surface deposits, with high levels of Ca and P in their composition and attached to the enamel (E-enamel, R-resin).

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Additionally, the linescan analyses indicate reduction in Ca and P in the enamel surface of the group treated with a bleaching agent only, whereas in the remineralized groups this protective ‘shell’ is firmly attached to the enamel surface and has a similar composition to the enamel (Ca and P).

The results obtained from the EDX elemental analysis (Table 1) confirmed that remineralization occurs in the experimental groups, since there are statistically significant increased levels of Ca and P, compared to the control group. In the samples treated with Nanosensitive® hca, there is also an increase in levels of Mg and Na.

Table 1.   EDX analysis of enamel surfaces
Element (%)Group 1 16% CP Mean ± (SD)Group 2 16% CP+ 5.5% NovaMin® Mean ± (SD)Group 3 16% CP+ 7% NovaMin® Mean ± (SD)
  1. a. Statistically significant differences at p < 0.05 between group 1/group 2.

  2. b. Statistically significant differences at p < 0.05 between group 1/group 3.

  3. c. Statistically significant differences at p < 0.05 between group 2/group 3.

Silicon ab0.39 (0.15)0.37 (0.13)0.19 (0.13)
Magnesium bc0.15 (0.10)0.10 (0.05)0.27 (0.16)
Aluminium0.05 (0.02)0.06 (0.04)0.07 (0.04)
Sodium bc0.37 (0.20)0.48 (0.14)0.83 (0.30)
Strontium0.21 (0.11)0.32 (0.10)0.30 (0.28)
Phosphorus abc15.60 (5.27)19.07 (2.25)21.71 (2.68)
Calcium ab28.43 (8.30)35.74 (3.25)37.56 (2.02)

Discussion

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Conclusions
  8. References

Although bleaching is a widespread dental procedure, it is still very controversial. In the literature, contradictory results regarding the ability of bleaching agents to cause morphological changes of the enamel are reported, and the differences seem to be associated with variations in the duration of the bleaching process and the concentration of active bleaching agents. Bitter20 reported that 10% CP can affect the tooth surface and underlined the importance of considering the effect of these changes on enamel integrity since, in the long-term, they could be the cause of abrasions or cusp fractures, particularly in teeth that have been restored or weakened by other dental treatments. In another study, the authors reported that after 10% CP night-guard bleaching, porosity of the enamel and a reduction in the bond strength appeared.21 Covington et al. reported the development of focal areas of very shallow erosion rather than pitting.7

Against this, several studies have reported that 10% CP gels are safe for enamel and do not result in modifications to the enamel surface.22–24 However, in general, these have been of shorter duration than those studies that have found damage.

Lopes et al.25 established that erosions on the enamel surface following bleaching were not uniform and their intensity varied depending on the sample, which is similar to the results we have obtained. Although the gel used in our study was only 16% CP and was applied for a relatively short period of time, the SEM micrographs demonstrate that enamel alterations do appear. They fluctuate from the very delicate to more severe demineralization. The latter might be capable of progressing into cavity formation. These relatively pronounced defects could result from the storage medium used (physiological saline) which did not encourage remineralization, or the nature of the teeth that were used, i.e. immature permanent teeth, which are less mineralized than the mature ones.

The present study employed a recommended regime for bleaching, namely application of the Mirawhite® Pro bleaching agent for 8 minutes per day for 7 days. When this was done, clear changes were observed in the surface enamel layers of the treated teeth.

Results in the present study also show that toothpastes containing bioactive glass are able to bring about remineralization of the damaged enamel surfaces. The SEM images demonstrate the success of this approach. Bioactive glass deposits were found on the enamel surface in both of the experimental groups, which suggests that they may act as a reservoir of ions available for remineralization at sites of possible demineralization. This is confirmed by the presence of a protective layer on the longitudinal sections of the enamel surface. The deposits are firmly attached, and were not removed by thorough washing and brushing. The linescans confirm this close attachment to the surface, a feature which is more pronounced in the group treated with Nanosensitive® hca.

Enamel remineralization by the toothpastes containing bioactive glass occurred by incorporation of different elements into the enamel structure.19 The basis of their bonding is the chemical reactivity of the glass in the presence of body fluids. The surface chemical reaction results in the formation of an HCA layer.16

The remineralization process is a slow dissolution/precipitation process of mineral constituents into the hard dental tissues.26 This process can be clearly seen on the sample treated with Nanosensitive® hca (Fig 3a, b). These specimens show a ‘flower-field’-like pattern, which results from precipitation of a new mineral phase on enamel that has lost inter-prismatic substance, while retaining enamel rods essentially unaltered. This confirms the results of Hegedus et al.10 who stated that peroxide-containing bleaching agents affect the organic phase of the enamel.

The results obtained by the EDX analysis confirm that mineral depletion has occurred on the enamel surface of teeth treated with 16% CP. This shows that bleaching agents cause not only removal of the organic phase, but also some of the mineral phase.27 A previous study found that teeth exposed to 10% CP solution for 6 hours lost small amounts of surface calcium as determined by atomic absorbtion spectroscopy, but this was considered to not be clinically significant.1 In an in vitro study, Goo et al.28 reported a slight loss of mineral content in the enamel when 10% CP was used. Rotstein et al. found that whitening agents can affect the hard tissues by altering the calcium concentration.29 This mineral loss can be reversed, as we have shown, by the use of toothpastes containing bioactive glass particles, since they are able to replenish the Ca and P content of damaged enamel and return it to that of undamaged enamel.

Another study comparing the remineralization potential of NovaMin® and CPP-ACP containing toothpastes found that the group treated with CPP-ACP increased the levels of calcium and phosphorus in the surface enamel and the resulting calcium-phosphate deposits were found to be amorphous.19 CPP-ACP enhances the remineralization of artificially formed dentinal lesions. The suggested mechanism for this is the stabilization of calcium phosphates on the tooth surface by the casein phosphopeptides, which leads to high concentration gradients of calcium and phosphate ions, thus promoting the remineralization of hard tissues.30

Therefore, we conclude that bleached enamel surfaces have distinctly different appearances from untreated enamel, and that the bleaching process is damaging to the mineral content of the tooth, as well as to the protein component. However, such damage can be reversed with the use of toothpastes containing bioactive glass powders which provide the ions necessary to remineralize these damaged surfaces. Of the two dentifrices studied, Nanosensitive® hca gave slightly better results.

Conclusions

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Conclusions
  8. References

The use of the commercial bleaching system containing 16% CP bleaching agent and applied under the recommended treatment regime causes distinct but variable morphological changes to the enamel surface. Subsequent treatment of damaged teeth with toothpastes containing the bioactive glass NovaMin® resulted in repair processes taking place. A protective layer of bioactive glass deposits is formed on the enamel surface, and there is an increase in the Ca and P content of the enamel layer, returning it to that of undamaged enamel.

Within the limitations of this in vitro study, we may conclude that although clinically effective, bleaching regimes adversely affect the enamel surface. This damage can be repaired by subsequent use of toothpastes containing bioactive glass powders as remineralizing agents. Further investigations should be undertaken in order to evaluate the long-term effect of the application of bioactive glasses on enamel remineralization, and their implication on enamel bonding.

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Conclusions
  8. References
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