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Year : 2022  |  Volume : 25  |  Issue : 4  |  Page : 509-515

Can the hydrogel form of sodium ascorbate be used to reverse compromised resin infiltrant penetration after bleaching?

1 Department of Restorative Dentistry, European University of Lefke Faculty of Dentistry, Lefke, Mersin-10, Turkey
2 Pharmaceutic Technology Department, Ege University Faculty of Pharmacy, İzmir, Turkey
3 Department of Restorative Dentistry, Ege University Faculty of Dentistry, İzmir, Turkey
4 Department of Oral and Maxillofacial Surgery, Cyprus Health and Social Sciences University, Faculty of Dentistry, Morphou, Mersin-10, Turkey

Date of Submission10-Sep-2021
Date of Acceptance14-Dec-2021
Date of Web Publication19-Apr-2022

Correspondence Address:
Prof. H Kemaloglu
Associate Professor, Department of Restorative Dentistry, Ege University Faculty of Dentistry İzmir
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/njcp.njcp_1805_21

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Aims: The aim of this study was to investigate the effects of an antioxidant on the bleaching-induced reduction in the penetration depth of infiltrant resins. Materials and Methods: White spot lesions (WSLs) were created on 105 bovine tooth samples, each measuring 6 × 4 × 4 mm. Five samples were randomly selected for the examination of lesion characteristics. The remaining 100 samples were then divided into four groups (n = 25). In Group I, the WSLs were treated with resin infiltration (RI) only. RI was performed on Group II immediately after bleaching. In Group III, an antioxidant was applied for 2 h after bleaching, and this was immediately followed by RI. The Group IV samples were treated with RI at the end of a 1-week waiting period after bleaching. The penetration depths were evaluated through confocal laser scanning microscopy. Results: The lowest penetration rate, which was approximately 57%, was observed in Group II. This was followed by Group III (87%), Group IV (90%), and Group I (92%). Group II, in which the samples were infiltrated immediately after bleaching, had the lowest mean penetration percentage. All the bleached groups exhibited significantly lower penetration percentages than the nonbleached group (Group I) (P < 0.05). Antioxidant application increased the penetration significantly (P < 0.05). Conclusion: Application of sodium ascorbate was found to reverse the reduced resin penetration depth and penetration percentages resulting from bleaching. The postponement of adhesive procedures after bleaching yielded similar results.

Keywords: Antioxidants, bleaching, confocal laser scanning microscope, resin infiltration, scanning electron microscope

How to cite this article:
Sadikoglu I S, Arici M, Kemaloglu H, Turkun M, Caymaz M G. Can the hydrogel form of sodium ascorbate be used to reverse compromised resin infiltrant penetration after bleaching?. Niger J Clin Pract 2022;25:509-15

How to cite this URL:
Sadikoglu I S, Arici M, Kemaloglu H, Turkun M, Caymaz M G. Can the hydrogel form of sodium ascorbate be used to reverse compromised resin infiltrant penetration after bleaching?. Niger J Clin Pract [serial online] 2022 [cited 2022 May 18];25:509-15. Available from:

   Introduction Top

White spot lesions (WSLs) are earliest stage of tooth decay.[1] Several treatment options are available for these enamel lesions. Increasing remineralization by using fluoride or casein phosphopeptide–amorphous calcium phosphate has been shown to be effective in stopping caries.[2] However, according to the International Caries Detection and Assessment System (ICDAS), clinical studies of these remineralization processes have not yet produced cosmetic improvements or significant reductions in caries lesions.[3],[4]

In recent years, another popular treatment for caries lesion has been resin infiltration (RI), which is defined as a microinvasive approach. This technique has been found to reduce microporosity. Moreover, the demineralized tissue is mechanically supported, and microorganisms with caries-forming potential are trapped in deep regions of the lesion and deprived of nutritional support.[5],[6] In situ[7] and in vivo studies[8],[9] have demonstrated the effectiveness of RI techniques.

When WSLs are masked by RI, the opaque chalky appearance of the enamel disappears, and the restored area regains its original color. However, after the application of RI, patients might not be satisfied because of their being accustomed to the white opaque appearance.[10] In such cases, bleaching could be performed before the application of RI.[11]

Several studies have found that adhesion quality is adversely affected by attempts to bind the composite to the enamel immediately after bleaching. The result is low bond strength and microleakage.[12],[13],[14],[15] The general approach has been to have a delay between the administration of the bleaching and adhesive procedures because the reduction in bond strength has been found to be transient.[16] However, in some clinical situations, postponement of the restoration session to complete the treatment is impossible because of patient schedules.[17] Therefore, the effectiveness of several methods has been investigated to ensure that restorative procedures can be performed immediately after bleaching.[18] The deterioration of the adhesion quality of the enamel surface to which bleaching is applied can be improved with the administration of a 10% sodium ascorbate solution before the adhesive process. Lai et al.[13] applied a sodium ascorbate solution to enamel for 3 h after bleaching. They reported that reduced bond strength was increased. Türkün and Kaya[19] reduced the duration of the sodium ascorbate application. They found that 10-min application was sufficient to increase bond strength.

One of the success parameters in RI is the achievement of efficient penetration depths. A recent study found that the whitening process, which is known to adversely affect the adhesive bonding of composite resins, had a negative effect on the penetration of resin infiltrants.[12] The aim of the current in vitro study was to investigate the effects of the postbleaching application of 10% sodium ascorbate hydrogel on the success of the RI treatment of artificially created WSLs in bovine teeth. The initial hypotheses were as follows:

H1: The bleaching process would adversely affect the penetration depth of the resin infiltrant applied to WSLs.

H2: The use of sodium ascorbate after bleaching would increase the penetration depth of the resin infiltrant.

   Method and Materials Top

Specimen preparation

All specimens (n = 105, 6 × 4 × 4 mm) were prepared from the bovine incisors of the second dentition Bovine teeth were obtained from a state slaughterhouse in North Cyprus-Famagusta region. Since only teeth of animals that died for other reasons were used for the study, our university did not find it necessary to obtain an ethics committee report. After being embedded in acrylic resin blocks (MelioDent Self Cure, Heraeus Kulzer, Hanau, Germany), enamel surfaces were successively polished with 1,200-, 2,400-, and 4,000-grit abrasive paper. An adhesive tape measuring 3 × 3 mm was placed on the center of each sample, and an acid-resistant varnish was applied to the entire enamel surface. After the varnish was dry, the adhesive tape was removed. Thus, a 3 × 3 mm unprotected area was obtained to simulate a WSL.

Development of white spot lesions

Queiroz solution,[20] which is classified as a short-term demineralizing solution, was used to create artificial enamel subsurface lesions in unprotected areas. The proportion of demineralizing solution per area of exposed enamel surface was calculated at 2 mL/mm2.

To simulate oral conditions, all specimens were subjected to the 8-day pH cycle described by Queiroz et al.[20] It involved 4 h of acidic challenge with demineralizing solution and 20 h in a remineralizing solution[21] each day in an incubator at 37°C.

To evaluate the development of WSLs, an opaque whitish appearance was determined by visual examination after the removal of a section of the varnish adjacent to the exposed surface after air drying. WSLs with scores of ICDAS 2 were obtained after these cycles. Five specimens were randomly selected for confocal laser scanning microscopy (CLSM) of the lesion characteristics. The remaining 100 samples were then divided equally into four groups (n = 25).

Experimental groups

Group I (positive control): RI

The enamel surfaces were first etched by the application of 15% hydrochloric acid gel (Icon-Etch, DMG, Hamburg, Germany) for 2 min followed by a water rinse for 30 s. The lesions were then dried with ethanol (Icon-Dry, DMG, Hamburg, Germany) for 30 s. For 3 min, a low-viscosity resin infiltrant (Icon-Infiltrant, DMG, Hamburg, Germany) was subsequently applied to the lesions, which were then light cured for 40 s (Elipar S10, 3M ESPE, St. Paul, MN, USA). Last, polishing was performed with the polishing wheels under water.

Group II: RI immediately after bleaching

The artificial WSLs were bleached with a chemically activated in-office bleaching system (Opalescence boost 40% hydrogen peroxide, Ultradent, South Jordan, UT, USA) in accordance with the manufacturer's instructions. Upon completion of the bleaching process, the RI was immediately applied to the bleached WSLs, as described for Group I.

Group III: Antioxidant gel application prior to RI of bleached specimens

Immediately after bleaching was completed, a hydrogel containing 10% sodium ascorbate was applied to the enamel surfaces for 2 h. Each sample was then washed with pressurized water for 30 s and air-dried. After the completion of antioxidant gel therapy, RI was performed as described for Group I.

Group IV: RI 1 week after bleaching

The specimens in this group were bleached with the same bleaching agent as described for Group II and then incubated in deionized water at 37°C for 1 week. The lesions were then infiltrated with resin, as described for Group I.

Image Analysis

In preparation for imaging, all specimens were divided into two halves perpendicular to the surface of WSLs. A low-speed water-cooled cutting device (Isomet, Buehler, Lake Bluff, USA) was used. To obtain better images, thinner (approximately 1–2 mm) samples were subsequently obtained, by incision, from the incisal and cervical areas adjacent to the artificially created 3 × 3 mm WSL area.

Confocal laser scanning microscopy

For the visualization of the pore structures of the remaining lesions, the specimens were immersed in 50% ethanol solution containing 100 μM sodium fluorescein (SF; Sigma Aldrich) for 10 min. The samples were then thoroughly washed in deionized water for 3 min and observed through a confocal laser scanning microscope (TCS SPE, Leica Microsystems, Heidelberg, Germany) in combined fluorescence and transmission mode. The excitation light had a maximum wavelength of 488 nm, and fluorescence was detected through a 525/50 nm band-pass filter. The transmitted light was detected without filters. The images were recorded with a lateral resolution of 1024 × 1024 pixels. The recorded confocal microscope images were examined with ImageJ image analysis software (ImageJ, NIH, Bethesda, MD). The lesion and penetration depths were measured at 10 different points at equal intervals (approximately 300 μm), and the mean penetration was calculated for each lesion.

Scanning electron microscopy

The structure of WSLs and the changes in the specimens after infiltration were examined with a scanning electron microscope (Apreo S, Thermo Fisher Scientific Inc., Waltham, MA, USA). The specimens were then placed on microscope holders with a two-sided adhesive tape, and a 7-nm-thick gold coat was applied with a Leica EM ACE600 (Leica Microsystems, Heidelberg, Germany) coater. The coated samples underwent SEM, and images were recorded.

Statistical analysis

After the completion of the experimental applications, the data were analyzed in IBM SPSS Statistics for Windows, Version 25.0 (IBM Corp., Armonk, NY, USA). Descriptive statistics were performed, and the results were presented as the means, standard deviations, and minimum and maximum values. Normal distribution was assessed through the Kolmogorov–Smirnov test. The differences were analyzed with the t-test. A value of P < 0.05 was considered significant.

   Results Top

On confocal microscopic images, porous structures, such as enamel lesions, appeared green because of the absorption of the SF solution. In contrast, the intact enamel tissue and infiltrated portions of the lesions did not exhibit any fluorescence; thus, they appeared black [Figure 1]a,[Figure 1]b,[Figure 1]c,[Figure 1]d. In the two-dimensional images of lesions and infiltrated sections, green shadows were frequently observed in the dark infiltrated areas. This was attributed to the nonhomogeneous distribution of the cavities and infiltrated areas in their real three-dimensional structures. Eight samples were damaged during preparation for microscope examination. Therefore, 97 samples were examined.
Figure 1: CLSM Images: Resin infiltrated areas appear black, while noninfiltrated areas remain green in group 1 (a), group 2 (b), group 3 (c), and group 4 (d)

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The mean penetration, standard deviation, and minimum and maximum penetration in the experimental groups are presented in [Table 1]. The mean lesion depth of the samples was approximately 107 μm. Group I, to which only RI (i.e., no whitening) was applied to WSLs, had the highest mean penetration percentage value. The lowest penetration rate, which was approximately 57%, was observed in Group II. This was followed by Group III (87%), Group IV (90%), and Group I (92%) [Figure 2]. The differences among the groups were statistically significant (P < 0.05). Group II, in which the samples were infiltrated immediately after bleaching, had the lowest mean penetration percentage. All the bleached groups exhibited significantly lower penetration percentages than the nonbleached group (Group I) (P < 0.05). The results for Group IV were significantly higher than those for Group III, the antioxidant group (P < 0.05).
Figure 2: Penetration graphic: average penetration percentages of experimental groups

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Table 1: Penetration depth scores of the groups

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In SEM images, contrasts were observed in the infiltrated resin, demineralization areas, and healthy enamel tissue. The enamel prisms on demineralized enamel surfaces were wider and darker. They appeared almost black. In contrast, the healthy enamel texture had a lighter grayish color [Figure 3]. The infiltrated resin appeared as extensions that filled these enlarged prisms. The resin extensions were seen intensely only in Group I images. However, because almost all the demineralized lesion areas in these samples were filled with resin, distinguishing between the healthy and demineralized enamel was difficult [Figure 3]a. The resin penetration in the Group II SEM images was much shallower than that observed in Group I. The results were similar to those for the CLSM images. Under the penetrated resin tags, the lesion region that could not be reached by the resin was distinguishable from the healthy enamel because of the clear monitoring of the enlarged enamel prisms [Figure 3]b. The SEM indicated that this area was the largest in the Group II samples. Similar to CLSM findings, SEM images for the Group III and IV samples [Figure 3]c and [Figure 3]d indicated that the penetration was deeper than that in Group II. In addition, some demineralized areas where the resin could not progress were also observed.
Figure 3: SEM images of experimental groups: (a) successful infiltration in group 1, (b) resin infiltrated and noninfiltrated lesion areas in group 2, (c) represents group 3, and (d) represents group 4

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   Discussion Top

A review of the literature indicated that comparisons of caries development in human and bovine teeth have yielded similar results, thereby confirming the effectiveness of using bovine teeth as an alternative in human dental caries. Paris et al.,[2] Askar et al.,[22] and Horuztepe and Baseren[11] applied RI to artificially created WSLs on bovine teeth. Because of their availability, standardization, and larger flat surface area, the teeth of same-aged bovines were used in the present study.

Hydrogen and carbamide peroxide are commonly used bleaching agents. Briso et al.[23] stated that 35% hydrogen peroxide had more negative effects than 10% carbamide peroxide in terms of resin tag formation. Studies have shown that high concentrations of hydrogen peroxide sodium ascorbate were more efficacious than home bleaching agents. In accordance with the abovementioned studies, 35% hydrogen peroxide solution containing an office bleaching agent was used to determine the effects of high levels of peroxide.

Several studies have shown that composite bond strength decreased in adhesive restorations performed immediately after the application of in-office and home bleaching agents.[12],[13],[14],[15],[24],[25] However, a review of the current literature indicated that very few studies have discussed the effects of bleaching on the success of RI.[11],[26] In the present study, the lowest penetration percentage was detected in Group III, to which RI was applied immediately after bleaching. This supported the findings of Horuztepe and Baseren,[11] who applied RI to bovine teeth samples with artificial initial caries lesions immediately after bleaching. They found that the application of bleaching prior to RI negatively affected the penetration depth of the resin infiltrant.

Several explanations have been proffered for the decreased adhesion quality after bleaching. Josey et al.[27] reported that bleaching caused surface and subsurface changes. They indicated that the postbleach application of phosphoric acid in adhesive restorations can destroy enamel prisms and, thus, negatively affect adhesion quality. In the present study, one of the reasons that the groups that underwent bleaching exhibited lower penetration than the group with only RI might have been the loss of enamel prisms, as explained by Josey et al.[27] Another theory that has gained acceptance is that the degraded adhesion might be related to oxygen radicals that occur after bleaching. These radicals enter the enamel–resin interface, block resin penetration, and disrupt resin polymerization. In the present study, the infiltrant resin that was used in bleached groups was similarly affected by the peroxides; therefore, full penetration was not possible.

Lai et al.[28] demonstrated that the decrease in bond strength that followed the application of hydrogen peroxide and sodium hypochlorite was reversed with sodium ascorbate. Türkün and Kaya[19] reduced the antioxidant application time observed by Lai et al.[28] from 3 h to 10 min. They found that even a 10-min application of sodium ascorbate was sufficient to increase bond strength. Sodium ascorbate produces this healing effect by reacting with the oxygen radicals in the environment and restoring the oxidized adhesive monomer. Thus, polymerization barriers are eliminated, and adhesion is less compromised. The application of an antioxidant application eliminates oxygen radicals and supports the development of hybrid layers and resin tags, which are the basic components of adhesion.

The present study used 10% sodium ascorbate hydrogel under the hypothesis that the penetration depth of the resin infiltrant would increase. The sodium ascorbate significantly eliminated the negative effects of bleaching on adhesion quality. These results were similar to those of previous studies.[29],[30] The present study also confirmed previous findings regarding increased bond strength in the samples that were soaked in distilled water or artificial saliva following bleaching. The percentage of penetration in the samples in which the RI was administered 1 week after bleaching was found to be significantly greater than that in the samples that were infiltrated immediately after bleaching. These results indicate that postbleach delays in the administration of RI or adhesive restorations can prevent a decrease in resin penetration. The statistical evaluation indicated that the difference between Group IV, for which RI was postponed for 1 week, and Group III, to which 10% sodium ascorbate was applied after bleaching, was significant. The postponement group had a higher penetration percentage than the antioxidant-applied group.

Soviero et al.[31] and Prajapati et al.[32] reported that SEM is a useful method for measuring lesion depth and resin penetration. In contrast, Paris et al.[33] reported difficulties in detecting and measuring the resin. This was attributed to insufficient radio opacity and the possibility that the comparisons of SEM measurements could be misleading because of the use of procedures, such as drying and coating. Consequently, SEM was used to support CLSM results and to provide more detailed examination of the lesion–resin relationship.

The results from the SEM supported those from the CLSM. The analysis of SEM images indicated that resin penetration depths in the samples in which RI was performed immediately after bleaching were shallower than those in the other groups. The samples that had been treated with only RI (i.e., no bleaching) were determined to have the deepest penetration. The findings were similar to those of Horuztepe and Başeren.[11] SEM revealed that the enlargement resulting from mineral loss was in demineralized and interprismatic areas. However, enamel prisms could not be followed in RI areas. The reason might be demineralization, which would have resulted from the application of hydrochloric acid to eliminate the smear layer before SEM. With SEM images in the present study, distinguishing demineralized areas characterized by enlarged enamel prisms in the positive control group was more difficult than doing the same in the other groups to which bleaching had been applied.

These findings, which were supported by CLSM results, might be attributable to the fact that the lesion was almost completely filled with resin. It is noteworthy that with CLSM, green areas were observed. This indicated that lesions were very narrow and the RI covered very large areas. The lower penetration depths observed in bleached samples supported Horuztepe and Başeren's SEM findings.[11] An examination of CLSM measurements indicated that the negligible numerical difference between Groups III and IV was statistically significant. The samples for which the RI was postponed had greater infiltration percentages, which were statistically significantly different from those observed in antioxidant-treated samples (P < 0.05).

Both initial hypotheses were confirmed by the results. The method of measuring infiltrant penetration is the limitation of this study. More precise results can be obtained with an imaging and measurement method in which both the resin and pores can be stained. Whether use of an antioxidant gel for a longer period can increase penetration. Therefore further studies needed to investigate this topic.

   Conclusion Top

The results of this in vitro study have led to the following conclusions:

  1. The application of RI immediately after bleaching negatively affected the penetration depth of the resin infiltrant.
  2. The application of 10% sodium ascorbate gel for 2 h increased the penetration depth.
  3. Within the limits of this study, the postponement of RI for 1 week allowed the infiltrant to reach significantly higher penetration values than those for the antioxidant application (P < 0.05).

Although there are many studies on the effect of antioxidants on compromised bond strength after bleaching, there is no research other than this one investigating on the effect of the RI method. Future studies are needed before we can draw definite conclusions about this untouched question. The authors of this study believe that the dual staining method will yield results with richer visuals in future studies. Another question that needs to be answered is how different antioxidants will give results in different times.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

   References Top

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  [Figure 1], [Figure 2], [Figure 3]

  [Table 1]


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