Influence of photo-activation source on enamel demineralization around restorative materials

This study evaluated the effects of the photoactivation source and restorative material on the development of caries-like lesions on human enamel after an in vitro pH challenge. Enamel cavities were prepared in 36 blocks, which were assigned to two groups according to the restorative material: resin-modified glass ionomer (RMGI) and composite resin (CR). Samples were exposed to quartz-tungsten-halogen lamp, argon-ion laser, or light-emitting diode (n = 6). The Knoop microhardness (KHN) values of the top surface of all materials were evaluated. Restored enamel blocks were thermocycled and subjected to 10 demineralization-remineralization cycles at 37°C. KHN analysis of the superficial enamel was performed by four indentations located 100 μm from the restoration margin. The material KHN was not affected by the photoactivation source. No significant difference in KHN was noted between CR and RMGI. The enamel surface around RMGI exhibited a higher KHN (272.8 KHN) than the enamel around CR (93.3 KHN), regardless of the photoactivation source. Enamel demineralization around the dental restoration was not influenced by the photoactivation source. Less enamel demineralization was observed around the RMGI than around the CR restoration. Descriptors: Glass Ionomer Cement; Composite Resins; Tooth Demineralization. Introduction The longevity of dental restorations is related to the durability of the bond and the sealing of the cavity margins.1 Microspaces or gaps at the restoration/enamel interface may allow the penetration of fluid containing cariogenic microorganisms,2 resulting in secondary caries progression.2,3 Secondary caries are frequent in patients with high caries risk and may necessitate the replacement of dental restorations.4 To prevent restoration failure, researchers developed fluoride-releasing glass-ionomer (GI) restorative materials that inhibit secondary caries formation.5,6 The subsequently developed resin-modified glass ionomer (RMGI) materials showed some improved mechanical characteristics, such as better flexural strength and surface-wear resistance, compared to conventional GI cements,7 as well as superior handling properties due to their light-activation. Vitremer (3M ESPE) is a hybrid material of GI restorative material and composite resin (CR) with acid-base and light-cured reactions. VitDeclaration of Interests: The authors certify that they have no commercial or associative interest that represents a conflict of interest in connection with the manuscript. Submitted: Sep 05, 2012 Accepted for publication: Dec 10, 2012 Last revision: Jan 30, 2013 Influence of photo-activation source on enamel demineralization around restorative materials Braz Oral Res. remer sets via “triple-cure” mechanisms.8 The first two setting reactions are based on an acid-base neutralization and free-radical methacrylate. The third setting reaction ensures the continuous polymerization of the remaining monomers that are not excited during light exposure. In the third reaction, microencapsulated potassium persulfate and ascorbic acid form a patented redox catalyst system that provides the methacrylate cure of the glass ionomer in the absence of light. 9 Exposure of enamel to argon laser (AL) prior to cariogenic challenge has been shown to reduce the depth of caries lesions compared to enamel surfaces without prior AL exposure.2,3,10-14 This preventive effect has been attributed to the surface coating created by AL, which promotes changes in the mineral structure and organic component of the surface coating. The resulting coating consists of a reactive surface that is less susceptible to caries formation. In addition to the synergistic effect between topical fluoride and argon irradiation,12-14 increased fluoride retention (400%) has been reported on enamel irradiated with a low energy density of AL (10.72 J/ cm2).15 However, the exact mechanism of caries resistance by AL irradiation remains unknown.16 Based on the dental literature, it is reasonable to speculate that the activation of restorative materials with AL would prevent secondary caries formation, especially when a fluoride-releasing restorative material is used. The induction of caries-like lesions around restorations in vitro is an experimental approach that provides information about the clinical behavior of the restorative materials under the experimental conditions.6,17 The objective of this study was to evaluate the cariostatic potential of the light-activation source and restorative materials in vitro after a pH challenge. The response variable evaluated was the Knoop microhardness (KHN) profile of human enamel around and in the superficial area of the restorative materials. The following null hypotheses were tested: 1. the superficial enamel KHN values around two restorative materials cured by different light-curing units are not different; 2. the KHN values of the two restorative materials are not different. Methodology Ethical aspects This study protocol was approved by the Guarulhos University Research Ethics Committee (CEPUnG, process no. 104/2009). Experimental design The factors under study were the material (2 types) and photoactivation source (3 types) in a factorial 2 × 3 design for evaluation of the restorative material and the superficial enamel. The response variable was the KHN. The experimental units consisted of 36 dental blocks (n = 6 per group) obtained from 18 unerupted human third molars stored in 0.1% thymol solution at 4°C. Blocks of 4 × 4 × 2 mm were sectioned from the third molars with double-faced diamond disks (#7020; KG Sorensen, Barueri, Brazil) at low speed (Kavo, Joinville, Brazil) under water irrigation. Cavities of approximately 1.6 mm in diameter and 1.6 mm in depth were prepared with #2292 diamond burs (KG Sorensen, Barueri, Brazil) under water spray. Teeth were distributed into two groups according to the restorative material, and each group was divided into three subgroups (Table 1). Table 1 Experimental groups. Groups Subgroups Energy density (J/cm2) n samples Activation source / Activation time RMGI 1.1 Quartz-tungsten halogen lamp / 40 s 28 6 1.2 Argon laser (200 mW) / 20 s 12.8 6 1.3 Light-emitting diode / 20 s 24 6 CR 2.1 Quartz-tungsten halogen lamp / 40 s 28 6 2.2 Argon laser (200 mW) / 20 s 12.8 6 2.3 Light-emitting diode / 20 s 24 6 RMGI: resin-modified glass ionomer material; CR: composite resin. Ferla JO, Rodrigues JA, Arrais CAG, Aranha ACC, Cassoni A Braz Oral Res. the Sof-lex (3M ESPE) polishing system for 15 s with each disk. An 11-mm-diameter light tip was used with a quartz-tungsten-halogen (QTH)-based curing unit (Optilux 501; Demetron/Kerr, Danbury, USA), whereas the LED source (Radii Cal) had 1,200 mW/ cm2 and an 8-mm-diameter light tip. The power density of both curing units was constantly measured with a radiometer (Demetron/Kerr, Danbury, USA). The power selected for the AL (Accucure 3000; LaserMed, Salt Lake City, USA) was 200 mW for 20 s. A 6.3-mm-diameter spot size was measured with the knife-edge method. The energy density (J/cm2) of the light emitted by all curing units was calculated as the power density multiplied by the exposure time. Both products were quantitatively evaluated by five central indentations at 100-μm distance on the top surface (Figure 1). A microhardness tester (PanTec; Panambra Ind. e Técnica SA, São Paulo, Brazil) was used with a 25-g load for 20 s, with a dwell time of 15 s. The specimens were thermocycled 1,000 times between distilled water baths held at 5°C and 55°C. The dwell time was 60 s and 5 s of transfer time (MSCT-3e; Elquip, Equipamentos para All cavities were prepared and restored by the same calibrated operator. Table 2 shows composition, lot number, and application instructions of the selected materials. A commercial RMGI (Vitremer; 3M ESPE, St. Paul, USA; A3 shade) was tested in this study. Prior to the RMGI application, Vitremer Primer (3M ESPE, St. Paul, USA) was applied for 30 s, dried for 5 s, and light-activated for 20 s. The RMGI was mixed according to the manufacturer’s instructions in a 1:1 proportion and inserted in a single increment by syringe (Centrix Inc., Shelton, USA). Groups 1.1, 1.2, and 1.3 were light-activated according to each activation source. After RMGI insertion, Vitremer Finishing Gloss was applied and light-activated for 20 s with a light-emitting diode (LED) (Radii Cal; SDI Limited, Bayswater, Australia). For all groups, the adhesive was light-activated with a LED curing unit (Radii Cal; irradiance: 1,200  mW/cm2). After the bonding procedure, a nanofilled resin composite (Filtek Z350-OA3, 3M ESPE) was inserted in a single increment and activated by the respective photoactivation source. Restored enamel blocks were stored in 100% relative humidity at 37°C for 24 h and then polished with Table 2 Composition, lot number, and application mode of the selected materials. Material Manufacturer Composition Directions for use Adper Single Bond 2 (3M-ESPE, Irvine, USA; Lot: 9XL) HEMA, Bis-GMA, DMAs, functional methacrylate, copolymer of polyacrylic and polyitaconic acids, water, ethanol, nanofilller, photoinitiator Consecutively apply 2 coats, gently air-dry, and light-cure for 10 s Conditioner (3M-ESPE, Irvine, USA; Lot: 060808) Conditioner: etchant (37% phosphoric acid) Etch cavity for 15 s, wash and dry (do not desiccate) Z350-OA3 (3M-ESPE, Irvine, USA; Lot: 7CN) Bis-GMA, Bis-EMA, UDMA and camphorquinone. Fillers: Zirconia-sílica Light-activate each increment for 40 s for QTH source Vitremer (3M-ESPE; St. Paul, USA; Lot: 8HP; 8HH) Powder: fluoroaluminosilicate glass, microencapsulated potassium, Persulfate, ascorbic acid and pigments Liquid: aqueous solution of a polycarboxylic acid modified with pendant methacrylate groups, copolymer, water, HEMA and photoinitiators Light-activate each increment for 40 s for QTH source with 2-mm maximum thickness Vitremer Primer (3M-ESPE; St. Paul, USA; Lot: 8CB) Vitrebond copolymer, HEMA, ethanol and 


Introduction
The longevity of dental restorations is related to the durability of the bond and the sealing of the cavity margins. 1 Microspaces or gaps at the restoration/enamel interface may allow the penetration of fluid containing cariogenic microorganisms, 2 resulting in secondary caries progression. 2,3econdary caries are frequent in patients with high caries risk and may necessitate the replacement of dental restorations. 4To prevent restoration failure, researchers developed fluoride-releasing glass-ionomer (GI) restorative materials that inhibit secondary caries formation. 5,6The subsequently developed resin-modified glass ionomer (RMGI) materials showed some improved mechanical characteristics, such as better flexural strength and surface-wear resistance, compared to conventional GI cements, 7 as well as superior handling properties due to their light-activation.
Vitremer (3M ESPE) is a hybrid material of GI restorative material and composite resin (CR) with acid-base and light-cured reactions.Vit-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.remer sets via "triple-cure" mechanisms. 8The first two setting reactions are based on an acid-base neutralization and free-radical methacrylate.The third setting reaction ensures the continuous polymerization of the remaining monomers that are not excited during light exposure.In the third reaction, microencapsulated potassium persulfate and ascorbic acid form a patented redox catalyst system that provides the methacrylate cure of the glass ionomer in the absence of light. 91][12][13][14] This preventive effect has been attributed to the surface coating created by AL, which promotes changes in the mineral structure and organic component of the surface coating.The resulting coating consists of a reactive surface that is less susceptible to caries formation.In addition to the synergistic effect between topical fluoride and argon irradiation, [12][13][14] increased fluoride retention (400%) has been reported on enamel irradiated with a low energy density of AL (10.72 J/ cm²). 15However, the exact mechanism of caries resistance by AL irradiation remains unknown. 16ased on the dental literature, it is reasonable to speculate that the activation of restorative materials with AL would prevent secondary caries formation, especially when a fluoride-releasing restorative material is used.
The induction of caries-like lesions around restorations in vitro is an experimental approach that provides information about the clinical behavior of the restorative materials under the experimental conditions. 6,17The objective of this study was to evaluate the cariostatic potential of the light-activation source and restorative materials in vitro after a pH challenge.The response variable evaluated was the Knoop microhardness (KHN) profile of human enamel around and in the superficial area of the restorative materials.The following null hypotheses were tested:

Methodology Ethical aspects
This study protocol was approved by the Guarulhos University Research Ethics Committee (CEP-UnG, process no.104/2009).

Experimental design
The factors under study were the material (2 types) and photoactivation source (3 types) in a factorial 2 × 3 design for evaluation of the restorative material and the superficial enamel.The response variable was the KHN.
The experimental units consisted of 36 dental blocks (n = 6 per group) obtained from 18 unerupted human third molars stored in 0.1% thymol solution at 4°C.Blocks of 4 × 4 × 2 mm were sectioned from the third molars with double-faced diamond disks (#7020; KG Sorensen, Barueri, Brazil) at low speed (Kavo, Joinville, Brazil) under water irrigation.Cavities of approximately 1.6 mm in diameter and 1.6 mm in depth were prepared with #2292 diamond burs (KG Sorensen, Barueri, Brazil) under water spray.Teeth were distributed into two groups according to the restorative material, and each group was divided into three subgroups (Table 1). the Sof-lex (3M ESPE) polishing system for 15 s with each disk.An 11-mm-diameter light tip was used with a quartz-tungsten-halogen (QTH)-based curing unit (Optilux 501; Demetron/Kerr, Danbury, USA), whereas the LED source (Radii Cal) had 1,200 mW/ cm² and an 8-mm-diameter light tip.The power density of both curing units was constantly measured with a radiometer (Demetron/Kerr, Danbury, USA).The power selected for the AL (Accucure 3000; LaserMed, Salt Lake City, USA) was 200 mW for 20 s.A 6.3-mm-diameter spot size was measured with the knife-edge method.The energy density (J/cm²) of the light emitted by all curing units was calculated as the power density multiplied by the exposure time.
Both products were quantitatively evaluated by five central indentations at 100-µm distance on the top surface (Figure 1).A microhardness tester (Pan-Tec; Panambra Ind. e Técnica SA, São Paulo, Brazil) was used with a 25-g load for 20 s, with a dwell time of 15 s.The specimens were thermocycled 1,000 times between distilled water baths held at 5°C and 55°C.The dwell time was 60 s and 5 s of transfer time (MSCT-3e; Elquip, Equipamentos para All cavities were prepared and restored by the same calibrated operator.Table 2 shows composition, lot number, and application instructions of the selected materials.A commercial RMGI (Vitremer; 3M ESPE, St. Paul, USA; A3 shade) was tested in this study.Prior to the RMGI application, Vitremer Primer (3M ESPE, St. Paul, USA) was applied for 30 s, dried for 5 s, and light-activated for 20 s.The RMGI was mixed according to the manufacturer's instructions in a 1:1 proportion and inserted in a single increment by syringe (Centrix Inc., Shelton, USA).Groups 1.1, 1.2, and 1.3 were light-activated according to each activation source.After RMGI insertion, Vitremer Finishing Gloss was applied and light-activated for 20 s with a light-emitting diode (LED) (Radii Cal; SDI Limited, Bayswater, Australia).
For all groups, the adhesive was light-activated with a LED curing unit (Radii Cal; irradiance: 1,200 mW/cm²).After the bonding procedure, a nanofilled resin composite (Filtek Z350-OA3, 3M ESPE) was inserted in a single increment and activated by the respective photoactivation source.Restored enamel blocks were stored in 100% relative humidity at 37°C for 24 h and then polished with Pesquisa Odontológica, São Carlos, Brazil).The restored enamel blocks were covered with wax, except for the restoration area and 1 mm around this area, which remained exposed.The restored enamel blocks were submitted to pH challenge to induce caries-like lesions.The in vitro demineralization/remineralization dynamic model produced caries-like enamel lesions through a modified Featherstone model 18 to simulate the condition of high caries risk.Each restored block was placed in 15 mL of demineralization solution at 37°C (2.0 mmol/L of calcium, 2.0 mmol/L of phosphate in a buffer solution of 74 mM of acetate, pH 4.3).The pH-cycling regimen was performed over 14 days, with 10 daily cycles of 6 h in demineralizing solution at 37°C.The remineralizing solution contained calcium and phosphate at a known degree of saturation (50 mmol/L KCl, 1.5 mmol/L Ca, 0.9 mmol/L PO 4 , 20 mmol/L tri-hydroxymethil-aminomethan, pH 7.0) that was changed daily.The enamel blocks were washed with distilled water before an 18-h immersion in remineralizing solution.On the 6 th , 7 th , 13 th , and 14 th days of the cycle, the restored enamel blocks were kept only in the remineralizing solution. 6he quantitative KHN evaluation of the enamel caries-like lesions was performed by the PanTec microhardness tester, with a 25-g load for 5 s.Four indentations located 100 µm from the bonded surface were created on the enamel surface in each enamel block in the upper, lower, left, and right sides around the restorations (Figure 1).

Statistical analysis
Two-way ANOVA was performed to evaluate the influence of the two variables tested: photoactivation source and material for superficial enamel and material evaluations.The means of the KHN values were compared by a Tukey post-hoc test (α = 0.05).The software used for statistical analysis in both evaluations was SANEST (EPAMIG, Belo Horizonte, Brazil).Power was calculated with the G power 3.1.2software package (Heine, Universität Dusseldorf, Germany) (p = 1) for both material and enamel evaluation.

Results
ANOVA did not show differences for the "material" factor (p = 0.05), "photoactivation source" factor (p = 0.86), or interactions between factors (p = 0.25).Table 3 shows the KHN values and Tukey results of both materials exposed to the evaluated photoactivation sources for material evaluation.In the evaluation of the superficial enamel

Specimen
Delimited area with enamel demineralization Indentation located 100 µm of restored margins demineralization, ANOVA showed differences for the "material" factor (p < 0.05) but no differences for the "photoactivation source" factor or interactions between factors (both p = 0.7).Table 4 shows the KHN and Tukey results for the "material" and "photoactivation source" factors for the superficial enamel evaluation.

Discussion
According to the results of the present study, the first null hypothesis (the superficial enamel KHN values around two restorative materials cured by different light-curing devices are not different) was rejected.Null hypothesis no. 2 (the KHN values of the two restorative materials are not different) was accepted.Although several studies have shown that AL irradiation provides reduced depth of caries-like lesions, [10][11][12][13][14] and increased enamel microhardness 12 compared with control groups, in the current study, the AL irradiation promoted no significant changes in enamel KHN values.
A previous study showed that the use of halogen or AL activation with 200 mW for 20 s resulted in similar degrees of conversion for the same resin composite, whereas decreased superficial KHN values were obtained for AL activation with an exposure time of 10 s. 19 Therefore, this protocol was used in the present study.The exposure time of 20 s allowed an energy density of 12.8 J/cm².This lower energy density of AL was sufficient to promote higher fluoride retention for enamel, as demonstrated by Nammour et al. 15 Turbino et al. 20 recommended against the use of thickness increments of more than 1 mm for the AL with a power setting of 250 mW and an exposure time of 30 s.
The first AL was cleared for marketing and clinical use in 1991 by the U.S. Food and Drug Administration (FDA). 21Since that time, AL has been used clinically to light-activate dental materials 17 and has been shown to be capable of polymerizing compomers 2 or CR. 20,22The use of AL in dentistry reduces the chair time in dental offices.1][12][13][14] The enamel resistance to caries was also demonstrated in vitro 24,25 and in vivo 23 after AL irradiation.However, although AL was effective at activating the restorative materials in the present study, laser technology is more expensive than QTH or LED.
The dynamic pH-cycling model has been shown to be adequate for studies of enamel caries-like lesions. 6,18Various protocols (e.g., with 5 pH cycles 17 or 14 days in demineralization solution 16 ) have been used to simulate demineralization/remineralization phenomena in the oral environment with different restorative materials 2 and activation sources. 16Nevertheless, other important clinical variables that are not addressed in the in vitro setting should also be considered, such as the cariogenicity and frequency of the patient's diet or presence of saliva. 6oberts et al. 26 demonstrated that increments above 3 mm are not recommended for the same RMGI material evaluated in the present study.A 1.6-mm-deep standardized cavity preparation was used to ensure maximal setting from light activation.Importantly, in this study, the P-value for the "material" factor was 0.05 when the microhardness of the CR or RMGI was evaluated (Table 3).This borderline significant result may be due to the small sample size of our study (n = 6), which is a limitation of this paper.
The preventive effect of the evaluated RMGI recorded around this restorative material compared to enamel KHN values around the resin composite (Table 4).This result was related to the fluoride-release benefit of RMGI material. 27Fluoride release reduces demineralization on enamel around the restoration and along the cavity wall.The evaluated resin composite contains organic pigments.Because of its dark shade, the manufacturer recommends an exposure time of 40 s, which is similar to the recommended exposure time for the RMGI when a halogen-based curing unit is used.The dark shade (OA3) of the CR promotes light attenuation.

Conclusion
The photoactivation source did not influence enamel demineralization around dental restorations superficially.There was less development of enamel demineralization around RMGI restorations than around CR restorations.

Figure 1 -
Figure 1 -Schematic representation of the superficial enamel microhardness and indentation location on the restorative materials.

Table 2 -
Composition, lot number, and application mode of the selected materials.

Table 3 -
Material evaluation of the Knoop microhardness values for the material and photoactivation source factors, mean [standard deviation] and n samples.

Table 4 -
[5][6]was evident once higher enamel KHN values were Superficial enamel evaluation of the enamel Knoop microhardness values for the material and photoactivation source factors, mean [standard deviation] n samples and the results of Tukey's test.Means followed by different uppercase letters indicate significant differences (p < 0.05).RMGI: resinmodified glass ionomer; CR: composite resin; QTH, quartz-tungsten-halogen; AL, argon laser; LED, light-emitting diode.