Abstract
Objectives
MDP (10-methacryloyloxydecyl dihydrogenphosphate) has been proven to possess chemical bonding ability to tooth hard tissues, but its contribution to the enamel bond strength has not been recognized. The aim of this study was to investigate the contribution of chemical bonding to the short- and long-term bovine enamel micro-tensile bond strengths (μTBS).
Methods
The acid-etched enamel surfaces were treated without any primer (control) or with one of three MDP-containing primers (containing different ratio of MDP/HEMA/Bis-GMA, Kuraray Co.) for 5 s, water-sprayed and air-dried. Subsequently, the pretreated enamel surfaces were applied with an etch-and-rinse adhesive Durafill Bond (Heraeus Kulzer) and placed with composite resin Durafill VS (Heraeus Kulzer). The specimens were prepared for μTBS tests after 24-h or 1-yr water storage. The etched enamel surfaces treated with or without MDP-containing primers were analyzed by scanning electron microscopy (SEM) and X-ray photoelectron spectroscopy (XPS).
Results
The acid-etched enamel treated with the MDP-primers for a very short time could produce the greater enamel μTBS than the control did ( p < 0.05), and change enamel micromorphology. No significant different μTBS were found between 24-h and 1-yr water storage ( p > 0.05). The chemical bonding of MDP on the enamel surfaces was re-confirmed by XPS.
Significance
The additional chemical bonding of MDP around the enamel crystallites of the etched enamel substrate could significantly increase the short- and long-term enamel μTBS, and their μTBS surpass those of the etch-and-rinse adhesive alone.
1
Introduction
Chemical bonding of carboxylic acids and phosphoric acid esters (PAEs) to enamel/hydroxyapatite (HAp) surfaces has been studied by numerous researchers . Chemical bonding at the resin–tooth hard tissue interfaces has been presumed to provide a strong and durable bond without any additional tooth preparations for macro-mechanical retention . Whether carboxylic acids either decalcify or chemically adsorb onto HAp depends on the dissolution rate of the respective calcium salts in their own acidic solutions, irrespective of concentration and pH . Some PAEs used in contemporary self-etch dental adhesives could decalcify and chemically adhere to HAp simultaneously . Hannig et al. demonstrated the inter- and intra-HAp crystallite hybridization by self-etch adhesives at the enamel–resin interfaces without any nano-gaps . Taken together, these findings suggest a physisorption or chemisorption of self-etch primers onto enamel/HAp surfaces after the interaction of self-etch primers with the enamel crystallites . Chemical bonding at the enamel/HAp surfaces has been investigated using X-ray photon spectroscopy (XPS), X-ray diffraction (XRD), liquid- and solid-state nuclear magnetic resonance (NMR), etc. in previous studies . Ten-methacryloyloxydecyl dihydrogen phosphate (MDP) is one of the most promising PAEs used in self-etch adhesives such as Clearfil SE Bond and Clearfil S3 Bond . Recently, Yoshihara et al. detected a 4-nm layered structure around HAp powder after reaction with MDP . Likewise, our previous research revealed that the self-etch adhesives containing either carboxylic acids or PAEs could decalcify and chemically adhere to HAp simultaneously . However, the previously published data did not demonstrate that self-etch adhesives containing acidic functional monomers produce stronger bond strengths than etch-and-rinse adhesives do . Yoshihara et al. examined the chemical stability of new functional monomer-calcium salts in combination with their previously determined bond strength results , and inferred that formation of stable monomer-calcium salts enhanced the immediate bond strength of self-etch adhesives . Based on the hypothesis that similar surface micromorphology would result in similar bond strengths, Erickson et al. matched the enamel surface micromorphologies that resulted from treatments with self-etch adhesives or various concentrations of phosphoric acid, analyzed the corresponding enamel bond strengths, and deduced that chemical bonding of some self-etch adhesives cannot compete with the micromechanical interlocking produced by phosphoric acid etching .
All previously published data about chemical bonding at the enamel/HAp surfaces were not directly associated with the bond strengths obtained by micromechanical interlocking. We previously assumed that chemical bonding of self-etch adhesives on the residual HAp crystallites would make a minor contribution to the immediate mechanical bond strengths . Up to now, it has not been clarified how much chemical bonding at the enamel surfaces contributes to the short- and long-term mechanical bond strengths. The null hypothesis tested in this study was that chemical bonding on the etched enamel substrate would make a minor contribution to the short- and long-term enamel bond strengths obtained by micromechanical interlocking, and long-term water storage would deteriorate the resin–enamel interfaces.
The aim of this experimental study was to investigate (1) the synergistic contribution of chemical bonding on the etched enamel surface to short- and long-term micromechanical bond strengths; (2) the effect of MDP-containing experimental primers on the micromorphology of the etched enamel surfaces; and (3) the resin–enamel interfaces after short- and long-term storage in water.
2
Materials and methods
2.1
Specimen preparations
Thirty-two non-carious bovine mandibular incisors were extracted from 8 cows in a local slaughterhouse, stored in the respective bottle containing 0.1% thymol solution at 4 °C, and were used for this study within 1 month after extraction. Four incisors extracted from the same cow were randomly divided into 4 groups according to the different surface treatments. Twenty-four incisors from 6 cows were randomly assigned into 4 groups as above-mentioned. All labial enamel surfaces were ground under copious running water serially with 300-, 600-, 1200- and 2500-grit SiC paper for 30 s, each ending with 4000-grit SiC paper for 1 min. The enamel surfaces were etched with 37% phosphoric acid (ETCH-37, Bisco, Inc., Schaumburg, IL, USA) for 15 s, water-sprayed for 30 s, and thoroughly air-dried. Subsequently, they were treated with one of three experimental (MDP-containing) primers (Ex-1, Ex-2, Ex-3, Table 1 ) for 5 s or without any experimental primer (control). Afterwards, the enamel surfaces were thoroughly water-sprayed for 1 min and totally air-dried. The etch-and-rinse adhesive Durafill Bond (non-acidic functional monomers, thus, no chemical reaction with enamel/hydroxyapatite) (Heraeus Kulzer, Hanau, Germany) was applied to the pre-treated enamel surfaces, gently air-dried, and light-cured for 20 s. Finally, two 2-mm increments of the composite resin Durafill VS (Heraeus Kulzer, Hanau, Germany) were placed, and light-cured for 40 s, respectively. All the light-curing procedures were done using a light-curing unit (MACO, SLC-VIIIB, Hangzhou, China) with a light output of 800 mW/cm 2 . All the materials used in the study are summarized in Table 1 .
| Product and manufacturer (LOT No.) | Principal ingredients | pH | Steps of application |
|---|---|---|---|
| Etch-37 | 37% Phosphoric acid etchant, bonzalkonium chloride | – | Applied and left untouched for 15 s, rinsed, totally dried |
| Bisco, Inc., Schanmburg, IL, USA (0900005544) | |||
| Durafill bond Heraeus Kulzer, Hanau, Germany (232.08) |
Methacrylate solvent, silicon dioxide, benzoi methyl ether, photopolymerization catalyst | – | Applied and gently air-blown to a film, light-cured for 20 s |
| Ex-1 Kuraray, Co., Japan (040218) |
30% (w/w) MDP/HEMA/Bis-GMA = 1:1:1 (w/w), solvent: distil water/ethanol=1:1 (v/v) | ≈2 | Applied for 5 s, water-rinsed for 1 min, totally dried |
| Ex-2 Kuraray, Co., Japan (040218) |
30% (w/w) MDP/HEMA/Bis-GMA = 2:1:1 (w/w), solvent: distil water/ethanol = 1:1 (v/v) | ≈2 | Applied for 5 s, water-rinsed for 1 min, totally dried |
| Ex-3 Kuraray, Co., Japan (040218) |
30% (w/w) MDP/Bis-GMA = 1:1 (w/w), solvent: distil water/ethanol = 1:1 (v/v) | ≈2 | Applied for 5 s, water-rinsed for 1 min, totally dried |
| Durafill ® VS Heraeus Kulzer, Hanau, Germany (21206) |
Urethanedimethacrylate, silicon dioxide (0.02~0.07μm) | – | Two 2 mm increments light-cured for 40 s, respectively |
2.2
Micro-tensile bond strengths (μTBS)
After water storage at room temperature for 24 h, 12 specimens were perpendicularly sectioned through the resin–enamel interfaces using a low-speed saw (Isomet 1000, Buehler, Lake Bluff, IL, USA) under continuous water cooling. Subsequently, they were prepared into beams of about 0.9 mm × 0.9 mm × 8 mm. The μTBS tests were performed with a Micro Tensile Tester (Bisco Co., USA) at a tensile speed of 1 mm/min until fracture. The dimension of the fractured surface was measured. The μTBS was calculated in MPa.
After water storage at room temperature for 1 yr (water change every week), another 12 specimens were prepared and subjected to μTBS tests as mentioned above.
After either 24-h or 1-yr water storage, the μTBS tests were repeated three times using 4 bovine incisors per cow per time (24 incisors from 6 cows).
2.3
Scanning electron microscopy (SEM)
The labial enamel surfaces of another 4 bovine incisors were treated with phosphoric acid and experimental primers as above-mentioned. These specimens were treated neither with adhesive nor composite resin. Furthermore, two debonded specimens were randomly selected from each group after μTBS tests. All the specimens were dried with a series of alcohol solutions, gold-sputtered, and analyzed with an SEM (ZEISS ULTRA 55, Germany).
During specimen preparation for μTBS tests, an additional 1 mm thick specimen was prepared from each group. After the cross-sectioned surfaces of these specimens were treated with 0.1 M HCl for 20 s to delineate the resin–enamel interfaces, the specimens were prepared as above-mentioned for the SEM observations.
2.4
X-ray photoelectron spectroscopy (XPS)
The labial enamel surfaces of another 4 bovine incisors were treated in the same way as those prepared for SEM analysis. The specimens were dried with a series of alcohol solutions, and subsequently prepared in high vacuum for XPS (AXIS ULTRA DLD , Kratos, England). XPS was performed using a monochromatic Al Kα X-ray source under the emission current of 8 mA, and accelerating voltage of 15 kV.
2.5
Statistics
Statistical analysis was performed with the SPSS software (version 17.0, SPSS Inc., Chicago, IL, USA). Factorial design ANOVA was used to statistically analyze all μTBS data. Post hoc LSD multiple comparisons were applied to evaluate the statistical differences in μTBS among the four groups after 24-h or 1-yr water storage.
2
Materials and methods
2.1
Specimen preparations
Thirty-two non-carious bovine mandibular incisors were extracted from 8 cows in a local slaughterhouse, stored in the respective bottle containing 0.1% thymol solution at 4 °C, and were used for this study within 1 month after extraction. Four incisors extracted from the same cow were randomly divided into 4 groups according to the different surface treatments. Twenty-four incisors from 6 cows were randomly assigned into 4 groups as above-mentioned. All labial enamel surfaces were ground under copious running water serially with 300-, 600-, 1200- and 2500-grit SiC paper for 30 s, each ending with 4000-grit SiC paper for 1 min. The enamel surfaces were etched with 37% phosphoric acid (ETCH-37, Bisco, Inc., Schaumburg, IL, USA) for 15 s, water-sprayed for 30 s, and thoroughly air-dried. Subsequently, they were treated with one of three experimental (MDP-containing) primers (Ex-1, Ex-2, Ex-3, Table 1 ) for 5 s or without any experimental primer (control). Afterwards, the enamel surfaces were thoroughly water-sprayed for 1 min and totally air-dried. The etch-and-rinse adhesive Durafill Bond (non-acidic functional monomers, thus, no chemical reaction with enamel/hydroxyapatite) (Heraeus Kulzer, Hanau, Germany) was applied to the pre-treated enamel surfaces, gently air-dried, and light-cured for 20 s. Finally, two 2-mm increments of the composite resin Durafill VS (Heraeus Kulzer, Hanau, Germany) were placed, and light-cured for 40 s, respectively. All the light-curing procedures were done using a light-curing unit (MACO, SLC-VIIIB, Hangzhou, China) with a light output of 800 mW/cm 2 . All the materials used in the study are summarized in Table 1 .
| Product and manufacturer (LOT No.) | Principal ingredients | pH | Steps of application |
|---|---|---|---|
| Etch-37 | 37% Phosphoric acid etchant, bonzalkonium chloride | – | Applied and left untouched for 15 s, rinsed, totally dried |
| Bisco, Inc., Schanmburg, IL, USA (0900005544) | |||
| Durafill bond Heraeus Kulzer, Hanau, Germany (232.08) |
Methacrylate solvent, silicon dioxide, benzoi methyl ether, photopolymerization catalyst | – | Applied and gently air-blown to a film, light-cured for 20 s |
| Ex-1 Kuraray, Co., Japan (040218) |
30% (w/w) MDP/HEMA/Bis-GMA = 1:1:1 (w/w), solvent: distil water/ethanol=1:1 (v/v) | ≈2 | Applied for 5 s, water-rinsed for 1 min, totally dried |
| Ex-2 Kuraray, Co., Japan (040218) |
30% (w/w) MDP/HEMA/Bis-GMA = 2:1:1 (w/w), solvent: distil water/ethanol = 1:1 (v/v) | ≈2 | Applied for 5 s, water-rinsed for 1 min, totally dried |
| Ex-3 Kuraray, Co., Japan (040218) |
30% (w/w) MDP/Bis-GMA = 1:1 (w/w), solvent: distil water/ethanol = 1:1 (v/v) | ≈2 | Applied for 5 s, water-rinsed for 1 min, totally dried |
| Durafill ® VS Heraeus Kulzer, Hanau, Germany (21206) |
Urethanedimethacrylate, silicon dioxide (0.02~0.07μm) | – | Two 2 mm increments light-cured for 40 s, respectively |
2.2
Micro-tensile bond strengths (μTBS)
After water storage at room temperature for 24 h, 12 specimens were perpendicularly sectioned through the resin–enamel interfaces using a low-speed saw (Isomet 1000, Buehler, Lake Bluff, IL, USA) under continuous water cooling. Subsequently, they were prepared into beams of about 0.9 mm × 0.9 mm × 8 mm. The μTBS tests were performed with a Micro Tensile Tester (Bisco Co., USA) at a tensile speed of 1 mm/min until fracture. The dimension of the fractured surface was measured. The μTBS was calculated in MPa.
After water storage at room temperature for 1 yr (water change every week), another 12 specimens were prepared and subjected to μTBS tests as mentioned above.
After either 24-h or 1-yr water storage, the μTBS tests were repeated three times using 4 bovine incisors per cow per time (24 incisors from 6 cows).
2.3
Scanning electron microscopy (SEM)
The labial enamel surfaces of another 4 bovine incisors were treated with phosphoric acid and experimental primers as above-mentioned. These specimens were treated neither with adhesive nor composite resin. Furthermore, two debonded specimens were randomly selected from each group after μTBS tests. All the specimens were dried with a series of alcohol solutions, gold-sputtered, and analyzed with an SEM (ZEISS ULTRA 55, Germany).
During specimen preparation for μTBS tests, an additional 1 mm thick specimen was prepared from each group. After the cross-sectioned surfaces of these specimens were treated with 0.1 M HCl for 20 s to delineate the resin–enamel interfaces, the specimens were prepared as above-mentioned for the SEM observations.
2.4
X-ray photoelectron spectroscopy (XPS)
The labial enamel surfaces of another 4 bovine incisors were treated in the same way as those prepared for SEM analysis. The specimens were dried with a series of alcohol solutions, and subsequently prepared in high vacuum for XPS (AXIS ULTRA DLD , Kratos, England). XPS was performed using a monochromatic Al Kα X-ray source under the emission current of 8 mA, and accelerating voltage of 15 kV.
2.5
Statistics
Statistical analysis was performed with the SPSS software (version 17.0, SPSS Inc., Chicago, IL, USA). Factorial design ANOVA was used to statistically analyze all μTBS data. Post hoc LSD multiple comparisons were applied to evaluate the statistical differences in μTBS among the four groups after 24-h or 1-yr water storage.
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