Influence of pH, bleaching agents, and acid etching on surface wear of bovine enamel

ABSTRACT Development of new materials for tooth bleaching justifies the need for studies to evaluate the changes in the enamel surface caused by different bleaching protocols. Objective The aim of this study was to evaluate the bovine dental enamel wear in function of different bleaching gel protocols, acid etching and pH variation. Material and Methods Sixty fragments of bovine teeth were cut, obtaining a control and test areas. In the test area, one half received etching followed by a bleaching gel application, and the other half, only the bleaching gel. The fragments were randomly divided into six groups (n=10), each one received one bleaching session with five hydrogen peroxide gel applications of 8 min, activated with hybrid light, diode laser/blue LED (HL) or diode laser/violet LED (VHL) (experimental): Control (C); 35% Total Blanc Office (TBO35HL); 35% Lase Peroxide Sensy (LPS35HL); 25% Lase Peroxide Sensy II (LPS25HL); 15% Lase Peroxide Lite (LPL15HL); and 10% hydrogen peroxide (experimental) (EXP10VHL). pH values were determined by a pHmeter at the initial and final time periods. Specimens were stored, subjected to simulated brushing cycles, and the superficial wear was determined (μm). ANOVA and Tukey´s tests were applied (α=0.05). Results The pH showed a slight decrease, except for Group LPL15HL. Group LPS25HL showed the highest degree of wear, with and without etching. Conclusion There was a decrease from the initial to the final pH. Different bleaching gels were able to increase the surface wear values after simulated brushing. Acid etching before bleaching increased surface wear values in all groups.


INTRODUCTION
Among the techniques to reestablish the good aesthetics for discolored vital teeth, vital tooth bleaching is a more conservative and less invasive alternative than direct and indirect conventional restoration techniques and has been well accepted among patients because they consider it a safe and effective technique 22 . The effectiveness of dental as the bleaching system used, bleaching agent concentration, application time and the application of light 1,16,26 .
Light sources can be used to optimize the bleaching procedure in an attempt to increase the rates of peroxide dissociation and reduce the bleaching session 2,7,16 . Analyzing the bovine enamel wear caused by different bleaching techniques, the enamel wear because similar results were obtained with and without light sources 13,23 . In addition to the above-mentioned factors, the pH of the bleaching agents 19 and acid etching before treatment 6,12,16,25 whitening process. At the present, manufacturers have shown concern about the acidity of bleaching gels because a pH below the critical point (pH 5.5 to 6.5) 21 may result in the dissolution of enamel, microhardness alterations 14 , increased sensitivity, increased enamel wear and surface roughness 13,23 .
In an endeavor to control demineralization, which is able to increase the penetration of the bleaching gel and, consequently, tooth bleaching 16 , the acid etching technique can be used before the bleaching procedure 6 . This technique works by producing changes in the enamel surface in two distinct ways. First, the acid removes the surface layer and, consequently, the plaque and acquired film are removed and, second, the remaining enamel with a more porous surface layer is also removed 20 . A greater ability for peroxide penetration was obtained when the previous acid etching was performed with a light source 6 . An increasing number of tooth-bleaching products are commercially available with different compositions and concentrations 11 , and nowadays, there is a trend towards decreasing the gels' concentration values and diversifying their formulations 1,3,23,24 .
the need for studies to evaluate the changes in the enamel surface caused by different bleaching protocols.
Thus, the aim of this in vitro study was to evaluate the bovine dental enamel wear in function of different bleaching gel protocols, the acid etching and pH variation.

MATERIAL AND METHODS
This is an in vitro study, and the experimental design for pH analisys was divided into two study agents based on hydrogen peroxide (H 2 O 2 ), two at 35%, one at 25%, one at 15%, and one at 10%] and time periods (at two levels: before and after bleaching). Wear analysis consisted of one study factor: treatment [in 11 levels: Control (did not receive the acid etching and bleaching gel), and two at H 2 O 2 35%, one at 25%, one at 15%, and one at 10%, with or without acid etching]. Different factors because different bleaching treatments use different light sources following the protocol indicated by the manufacturer. The control group

Material
The materials and equipments used for this study are outlined in Figure 1.

Specimen preparation
A total of 60 bovine mandibular central incisors were chosen for the study. They were cut into rectangular shapes and embedded in slow-setting surface was produced by wet polishing with 320, 400, 600 and 1200-grit silicon carbide abrasive papers. To control the individual variation in the enamel surface characteristics among the tested teeth, each specimen was divided into two halves, providing a control area and a test area 13,18,23 . The test area was further divided, with one-half receiving 37% phosphoric acid (Alpha Etch; Nova DFL Indústria e Comércio S.A., Rio de Janeiro, Brazil) for 30 seconds followed by the bleaching gel protocols, and the other half receiving only the bleaching gel protocols ( Figure 3).

Bleaching procedure
The specimens were randomly divided into six groups (n=10) described in Figure 2.

Measurement of pH levels of bleaching gels
To analyze the pH levels of the bleaching gels, a portable pHmeter with a digital display (Model 1001 Sentron, Sentron, WA, USA) was used. This device has an electrode of small dimensions, compatible with the size of the specimen. It could make a readout with an accuracy of 0.01 in a few seconds and with minimal waste of bleaching gel. Before beginning each group of readouts, the pHmeter was calibrated with double standard substances the bleaching gels were obtained from the average was taken 30 seconds after the contact between the bleaching gel and the bovine enamel, and, the second readout, 30 seconds after the end of the time used, with activation with a hybrid light source 23 .
Brushing abrasion All test areas, of the 60 studied specimens ---37% phosphoric acid for 30 seconds ---37% phosphoric acid for 30 seconds ---37% phosphoric acid for 30 seconds ---37% phosphoric acid for 30 seconds applications: One minute after the gel application, three consecutive minutes of diode laser/blue LED hybrid light activation were performed followed by a one minute interval and another three minutes of hybrid light activation. random, three readouts were performed on each portion (with or without acid etching) and the mean wear value was calculated (Figure 3). These surface (protected area) on one side to the opposite one, crossing the surface exposed to the experimental procedures. Thus, the wear readout the distance in micrometers between the graph midline, corresponding to the specimen plane line (intact area) and the deepest valley corresponding to the ground area 8,13,18,23 .

Statistical analysis
For correlation analyses between pH and wear, the Pearson correlation test was used. For pH analysis, two-way repeated measures ANOVA followed by the Tukey`s test were used. For wear, one-way ANOVA followed by the Tukey's test were adopted.

RESULTS
There was no correlation between pH and wear. When analyzing the pH values (Table 1)  peroxide Groups activated with hybrid light at the end of the bleaching agent application. The group that showed more alkaline was the experimental 10% hydrogen peroxide Group with nitrogen-doped titanium oxide nanoparticles, activated with violet hybrid light, which had a lower concentration of measurements.
The one-way ANOVA test for evaluating the surface wear showed statistically significant differences between the tested groups (p<0.05). Next, the Tukey's test was applied for the values obtained ( Table 2). All groups, except the 35% Total increase when acid etching was previously performed. The lowest wear value was found in control group (Group C), which did not undergo the acid etching and dental bleaching, and the 25% Lase Peroxide Sensy II Group (LPS25HL) with prior acid etching showed the highest wear value. The Group LPS25HL without acid etching showed similar results (p<0.05) to the other groups where the etching was done (Figure 4).

DISCUSSION
Most studies on dental bleaching investigates the morphological changes in the enamel 10,13,14,23 and the pulp damage 5,17 caused by different techniques, however, the changes caused by differences of the bleaching gels' pH is still controversial 23 . In this study when observing the results obtained by pH analysis of bleaching agents, one could conclude that there was a trend towards a decrease in the periods, except for the 15% Lase Peroxide Lite Group activated with hybrid light [diode laser/blue differences (Table 1). Another important fact that has already been described in the literature 23,27 and corroborates with this study, is that the higher the concentration of hydrogen peroxide, the more acidic is the pH in the bleaching gels.
In this study, the EXP10VHL experimental group (gel with nitrogen-doped titanium oxide nanoparticles) activated with hybrid light [diode laser/violet LED (VHL) (experimental)], presented the highest pH values in comparison to the other groups evaluated. The HL and VHL hybrid light sources employed to activate the bleaching gels presented a minimum increase (less than 2°C) in pulp chamber temperature 1 , assuring   effective dental bleaching with low pulp risk 6 . The values obtained for the pH of hydrogen peroxide, regardless of the group studied, remained above the level considered critical for enamel dissolution (pH 5.5 to 6.5) 21 . However, it was noted that as the concentration of the gel diminished, its pH increased (Table 1). Therefore, the pH of these bleaching gels is unlikely to cause severe morphological changes on the surface of the enamel. Acid conditioning prior to bleaching is a procedure still used in dental practice, however, 6 showed that there is no difference in the color analysis when acid conditioning is previously done, added to this, this study showed an increased wear due to the acid conditioning when the same bleaching gel concentration was analyzed. Only the Group TBO35HL showed no increased wear in the area with acid etching, in comparison with the area that received only the bleaching gel (Table 2). In the comparison between groups, Group LPS25HL showed the highest wear for the area with acid etching ( Table 2).
Comparing the control group, which underwent brushing cycles only, with the other groups, it was less wear (Table 2) than the experimental groups in the area treated with both the acid etching and bleaching gel and the area treated only with bleaching gel. Certainly, these differences in wear are due to the changes caused by the bleaching agents on the dental enamel, results already found in the literature, in which tooth-bleaching techniques led to increased surface roughness and wear if subjected to simulated tooth brushing, when compared with the control group that was only brushed 13,16,23 .
Note that in this study the dental bleaching may have left the surface of the bovine enamel more susceptible to wear caused by the 100,000 simulated brushing cycles, which is in agreement with the makes the enamel surface more susceptible to abrasion and morphological changes 1,9,13,23,28 . The adopted simulated tooth brushing (100,000 simulated brushing cycles) caused wear between because other factors contribute to the loss of this structure. These cycles are equivalent, on average, to two years of brushing in vivo 4,15 , a period that was adopted for being considered as a safe interval between bleaching sessions.
The analysis of this research's results, associated evaluation that, in general, the 25% Lase Peroxide Sensy II Group activated with hybrid light (HL) (diode laser/blue LED) presented the highest wear values, causing greater aggression to the tooth structure. The acid etching was more severe in all Group. Although a direct correlation was not found with the results of wear, it was observed that the lower the bleaching gel concentration, the more alkaline it tended to be. Acid etching prior to bleaching can be indicated obtaining satisfactory results after attempts by conventional whitening techniques, especially in patients with a high degree of pigmentation, seeking a greater penetration of the gel to the enamel. However, few studies evaluate this procedure and its consequences, thus justifying the objective of this study, which, despite being an in vitro study, seeks to answer questions that may assist in the clinical practice of the procedure. The results of this study show that etching with 37% phosphoric acid can lead to increased enamel wear, and should not be used as a routine procedure, only when necessary in an attempt to avoid more invasive procedures such as cavity preparation and production of indirect restorations. However, further studies related to the effects of acid etching, pH, and different concentrations extrapolate the results to the clinical context.

CONCLUSIONS
Within the limitations of this in vitro study, the following conclusions can be drawn: There was a trend towards a decrease in the initial pH at the end, except in the Lase Peroxide Lite 15% Group. There was no direct correlation between the values of pH and surface wear.
Different bleaching gels are able to increase the surface wear values after simulated brushing, mainly in the Lase Peroxide Sensy 25% Group.
Acid etching before bleaching increased the surface wear values in all groups.