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RESEARCH REPORT |
1 Paediatric Dentistry and Orthodontics, Faculty of Dentistry, The University of Hong Kong, Prince Philip Dental Hospital, 34 Hospital Road, Hong Kong SAR, China;
2 Department of Dental Materials, Nihon University School of Dentistry at Matsudo, Chiba, Japan;
3 Department of Research and Development, Shofu Inc., Kyoto, Japan;
4 Oral Biosciences, Faculty of Dentistry, The University of Hong Kong, Hong Kong SAR, China;
5 Department of Operative Dentistry, Tokyo Medical and Dental University, Tokyo, Japan; and
6 Department of Oral Biology and Maxillofacial Pathology, School of Dentistry, Medical College of Georgia, Augusta, GA, USA;
* corresponding author, kfctay{at}netvigator.com
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS & METHODSRESULTSDISCUSSIONREFERENCES |
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KEY WORDS: acidic monomer • water content • dentin • microtensile bond strength • nanoleakage
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS & METHODSRESULTSDISCUSSIONREFERENCES |
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Bonding to tooth structure via the self-etching approach is achieved by increasing the concentrations of acidic methacrylate monomers (Hayakawa et al., 1998). These monomers contain one or more acidic functional groups and have comparatively low, first-acid dissociation constants (pKa1). Water is required to dissociate these weak acids into ionized forms for permeation of the smear layer and demineralization of the underlying intact tooth substrates (Tay and Pashley, 2001; Pradelle-Plasse et al., 2004). A hydrophilic monomer, 2-hydroxyethyl methacrylate (HEMA), is often included as a transitional polymerizable solvent, since many of the acidic monomers are only mildly water-soluble (Chigira et al., 1989). Although the aggressiveness of self-etch adhesives is largely determined by the concentrations and the pKa values of these acidic monomers (Nishiyama et al., 2004b), differences in water concentration may also contribute to partial or total dissociation of the acid functionalities, creating a variable extent of fractional ionization of these monomers (Anseth et al., 1996).
The effect of water concentration on the aggressiveness and bonding efficacy of self-etching primers to dentin has not been thoroughly investigated, since commercially available systems differ in their resin monomer compositions, adhesive solvents, and water contents. Thus, it is difficult to study the contribution of a single factor without resorting to a modeling approach. In this study, we created experimental self-etching primers using one acidic resin monomer, one hydrophilic resin monomer, a fixed amount of ethanol, and variable amounts of water for simultaneous etching and priming of dentin. The null hypothesis tested was that altering the water concentration in two-step self-etching primers has no effect on their aggressiveness and bonding efficacy to dentin.
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MATERIALS & METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS & METHODSRESULTSDISCUSSIONREFERENCES |
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Preparation of Experimental Self-etching Primers
An anhydrous stock primer solution was first prepared by dissolving 4-acryloxyethyltrimellitic acid (4-AET) powder in HEMA. 4-AET was synthesized according to the method described by Ikemura and Endo (1998). The stock solution consisted of 40 wt% 4-AET, 59 wt% HEMA, and 1 wt% 2,4,6-trimethylbenzoyl-diphenyl phosphine oxide (Aldrich Chemical Company, Milwaukee, WI, USA) as the photoinitiator. The latter was used in lieu of camphorquinone because of its greater activity for photopolymerization of the acidic HEMA solution (Ikemura, unpublished results).
Five experimental two-bottle-type, self-etching primers were subsequently prepared, with the stock solution as component A, and absolute ethanol/water as component B. The resin-ethanol-water volume ratios after the 2 components were mixed were: 9-0-1 (water-free), 8-1-1 (10 vol% water); 7-2-1 (20 vol%), 5-4-1 (40 vol%), and 3-6-1 (60 vol%). Their pH values at ambient temperature (20–25°C), as determined with a digital pH meter (Model 501, Orion Research, Inc., Beverly, MA, USA), were 1.97 ± 0.06 (Group 8-1-1), 1.93 ± 0.06 (Group 7-2-1), 1.90 ± 0.04 (Group 5-4-1), and 1.86 ± 0.12 (Group 3-6-1). Group 9-0-1 could not be measured with the pH meter, since we could not completely dehydrate the electrolyte, and the pH value of 7.00 ± 0.00 was measured with litmus paper. The experimental self-etching primers were used immediately after the 2 components were mixed, to avoid hydrolysis of the acidic and hydrophilic monomers in the presence of water (Nishiyama et al., 2004a).
Tensile Bond Strength and Fractographic Evaluation
Three teeth were used for each of the 5 experimental groups. Immediately after the 2 components were mixed, each self-etching primer was applied with agitation on the dentin surface for 20 sec. Without being rinsed, the solution was gently air-dried for evaporation of the solvent. The primed dentin was bonded with Fluoro-Bond bonding agent (Shofu Inc., Kyoto, Japan) and light-cured for 20 sec via a halogen light-curing unit operated at 600 mW/cm2. Composite build-ups were constructed with a light-cured resin composite (Beautifil, Shofu Inc.) in 5 one-mm-thick increments.
After being stored in distilled water at 37°C for 24 hrs, the bonded teeth were sectioned occluso-gingivally into 0.9 x 0.9-mm composite-dentin beams, according to the ‘non-trimming’ technique of the microtensile test. Between 35 and 46 beams were available for bond testing in each group. Specimens were stressed to failure under tension by means of a universal testing machine (Model 4440; Instron, Inc., Canton, MA, USA) at a crosshead speed of 1 mm/min. Bond strength data were statistically analyzed by one-way ANOVA and Tukey’s multiple-comparison tests, with statistical significance set at 
= 0.05.
The failure modes of all specimens were initially examined with a stereomicroscope (Nikon SMZ10, Tokyo, Japan) at 30X magnification and were classified as adhesive, cohesive in dentin/composite, or mixed failures. For each experimental group, each type of failure mode was expressed as a percentage of the total number of specimens in that group. Four representative fractured beams from each group with tensile bond strengths that were close to the mean bond strength of that group were examined with scanning electron microscopy (SEM). In addition, 4 specimens from each group with tensile bond strengths that were either higher or lower than the corresponding mean bond strength of that group were also examined. The dentin sides of the fractured beams were air-dried, sputter-coated with gold/palladium, and examined by SEM (Cambridge Stereoscan 440, Cambridge, United Kingdom) at 10 kV.
Transmission Electron Microscopy (TEM)
Two teeth from each group were similarly treated with the self-etching primer solution and further bonded with Fluoro-Bond bonding agent. The bonded teeth were coupled to a microfilled resin composite (EPIC-TMPT, Parkell Inc., Farmingdale, NY, USA) to facilitate ultramicrotomy. The teeth were sectioned into 0.9-mm serial slabs. The 2 widest slabs from each tooth were used for TEM examination. For evaluation of the aggressiveness of various self-etching primers, 1 slab from each tooth was completely demineralized in ethylene diamine tetraacetic acid. After laboratory processing according to the protocol reported by Tay et al.(1999), 90-nm-thick epoxy-resin-embedded sections were prepared and stained with uranyl acetate and lead citrate.
For examining the extent of resin infiltration with resin-dentin interfaces formed by the self-etching primers, we immersed the second slab from each tooth in a 50 wt% ammoniacal silver nitrate solution for 24 hrs, according to the silver-impregnation protocol reported by Tay et al.(2002). After reduction of the diamine silver ions, the silver-impregnated slabs were processed for TEM examination without further demineralization and examined unstained. Both types of sections were examined by TEM (CM100, Philips, Eindhoven, The Netherlands) at 80 kV.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS & METHODSRESULTSDISCUSSIONREFERENCES |
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). Group 7-2-1 specimens exhibited the highest strength (47.6 ± 6.6 MPa) (P<0.05). This was followed by Groups 8-1-1 (43.1 ± 6.1 MPa) and 5-4-1 (42.3 ± 7.6 MPa), which were not significantly different from each other, but were significantly higher than Group 9-0-1 (35.9 ± 7.5 MPa). Group 3-6-1 exhibited the lowest bond strength (22.6 ± 8.1 MPa). Between 8 and 15% mixed and cohesive failures were observed in all groups, with the exception of Group 3-6-1, in which bond failures were exclusively adhesive.
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). Conversely, the fractured hybrid layer surface was porous in Group 3-6-1 (lower bond strengths), from which denuded collagen fibrils were detached from the resinous components (Fig. 1C). Specimens with bond strengths higher or lower than those of the corresponding mean bond strength of a particular group depicted either mixed failures or cohesive failures within the weaker dentin (not shown).
Increasing the water concentration from 0–60 vol% resulted in improved acidic monomer ionization, manifested as an increase in the depth of the hybrid layers created by the self-etching primers. Hybrid layer thickness varied from 0.2 µm in Groups 9-0-1 (Fig. 2A) and 8-1-1 (Fig. 2C), to 0.5–1 µm in Group 7-2-1 (Fig. 3A), to 1.5–2 µm in Groups 5-4-1 (Fig. 4A) and 3-6-1 (Fig. 4C).
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). In addition, an unusual reticular pattern of nanoleakage that was sporadically located in the mineralized dentin beneath the hybrid layers could frequently be identified in Group 9-0-1 (Fig. 2B), and occasionally in Group 8-1-1 (Fig. 2D).
Group 7-2-1 exhibited minimal nanoleakage within the partially demineralized zone (Fig. 3B) that was predominantly of the spotted type of nanoleakage (Tay et al., 2002) (Fig. 3C). A further increase in the water content of the self-etching primers was not accompanied by an improvement in the quality of dentin hybridization, with moderate (Group 5-4-1; Fig. 4B) to severe nanoleakage (Group 3-6-1; Fig. 4D) observed within the 1.5- to 2-µm-thick partially demineralized zones.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS & METHODSRESULTSDISCUSSIONREFERENCES |
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Despite the thinness of the hybrid layers (ca. 200 nm) in Groups 9-0-1 and 8-1-1, extensive nanoleakage could be identified within the zones of partially demineralized dentin. Conversely, minimal nanoleakage was observed in Group 7-2-1, with a thicker hybrid layer (0.5–1 µm). Water molecules form hydration clusters around functional groups on collagen peptides and proteoglycans, thereby preventing the interpeptide or peptide-proteoglycan hydrogen bonding that can collapse interfibrillar spaces in the demineralized matrix (Suzuki and Nakai, 1994; Nakaoki et al., 2000; Pashley et al., 2000, 2001). Thus, apart from its role in ionization of the acidic resin monomers, it appears that a minimal water concentration is necessary to keep the demineralized matrix in an expanded stage, even for self-etching primers in which resin infiltration occurs simultaneously with etching of the dentin. This could have accounted for the better resin infiltration and the minimal nanoleakage observed in Group 7-2-1.
The appearance of a reticular pattern of silver-infiltrated interfibrillar spaces beneath the hybrid layers in Group 9-0-1 and occasionally in Group 8-1-1 was unusual, and suggested that continuous etching of the dentin beneath the hybrid layer could have occurred, since the initially non-ionized acidic monomers were converted into their ionic states by water that was derived from the underlying dentin, or during the period of specimen storage in water prior to their laboratory processing. This phenomenon did not occur in the other 3 groups, probably because the acidic monomers were more optimally ionized with the water that was present in these mixed primer solutions, enabling the ionized form of the acidic monomers to exert an immediate etching effect on the underlying dentin. Conversely, delayed ionization of the acidic monomers in Groups 9-0-1 and 8-1-1 could have resulted in non-infiltrated spaces beneath the hybrid layers that were subsequently penetrated by the tracer solution. Water is also useful to solubilize the dissolved calcium and phosphate ions, thereby lowering their ion products below the solubility product constants for various forms of calcium phosphates (McDowell et al., 1977; Shellis et al., 1997). When insufficient water is present, re-precipitation of calcium phosphates within the very fine, partially demineralized interfibrillar spaces of the underlying dentin may have prevented resin infiltration into these regions, resulting in the unusual form of nanoleakage beneath the hybrid layers. The hypotheses of delayed ionization of acidic monomers under the condition of an inadequate supply of water, and its possible effect on etching and resin infiltration of dentin, are speculative and require further testing for their validity to be confirmed.
Increasing the water concentration beyond 20 vol% resulted in further increases in the degree of ionization of the 4-AET monomers, lowering of the pH of the self-etching primer solutions, and further increasing their depth of demineralization. However, increasing the water concentration also inadvertently diluted the concentrations of the adhesive monomers, thereby lowering their inward rates of diffusion. The water molecules also competed with the adhesive monomers for binding sites on collagen (Vaidyanathan et al., 2003). The excess water may also prevent optimal polymerization of the adhesive monomers. Using 62% BisGMA/37% HEMA-based bonding agent, Jacobsen and Söderholm (1995) reported the effect of increasing water concentration on the percent conversion of double bonds, bond strength, and phase changes. As water was added to the bonding agent, the percent conversion fell rapidly from 53.5% at zero water, to 22.7% when 20% water was added. Phase changes may also occur as water is added. Spencer and Wang (2002) examined the amount of water necessary to produce phase changes in 70% BisGMA/30% HEMA formulation dissolved in 45% ethanol. This formulation exhibited phase separations at 28 vol% water. Without ethanol solvation, the same 70% BisGMA/30% HEMA underwent phase separations at around 5 vol% water. These factors, individually or in combination, could have contributed to the continuous decline in bond strength, and the increasing severity of the nanoleakage in Groups 5-4-1 and 3-6-1, despite the formation of thicker hybrid layers (ca. 1.5–2 µm) by these more aggressive self-etching primers.
As the water concentration decreases, the adhesive monomer concentration increases. This tends to increase the viscosity of the formulation, permitting the creation of thicker adhesive concentrations. In a study that examined the microtensile bond strength of an acetone-based adhesive to phosphoric-acid-etched dentin, the highest bond strengths (64 MPa) were obtained with 37% acetone and 63% comonomers, while the lowest bond strength (38 MPa) was produced by 67% acetone and 33% comonomers (Cho and Dickens, 2004). Clearly, higher monomer concentrations are preferred.
Within the limits of this study, the null hypothesis—that altering the water concentration in two-step self-etching primers has no effect on their aggressiveness and bonding efficacy to dentin—has to be rejected. When self-etching primers are formulated, compromises must be made to provide sufficient water for adequate ionization of the acidic monomers, but without lowering the resin concentration too much to optimize their bonding efficacy to dentin.
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ACKNOWLEDGMENTS |
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Received June 2, 2004; Last revision February 27, 2005; Accepted April 8, 2005
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REFERENCES |
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