JOURNAL OF ENDODONTICS Copyright 2003 by The American Association of Endodontists

Printed in U.S.A. VOL. 29, NO. 9, SEPTEMBER 2003

Adhesion of Endodontic Sealers: Scanning Electron Microscopy and Energy Dispersive Spectroscopy
Iman M. Saleh, BDS, MSc, I. Eystein Ruyter, Dr. Rer. Nat., Dr. Philos., Markus P. Haapasalo, DDS, PhD, and Dag rstavik, DDS, PhD

The microscopic details of the debonded interfaces between endodontic sealers and dentin or gutta-percha were assessed in this study. Dentin, conditioned with 37% H3PO4 for 30 s, 25% citric acid for 30 s, 17% EDTA for 5 min, or a rinse with 10 ml of distilled H2O (control), and gutta-percha surfaces were coated with freshly mixed sealer: Grossmans sealer, Apexit, Ketac-Endo, AH Plus, RoekoSeal Automix, or RoekoSeal Automix with an experimental primer. The surfaces were pressed together and the sealers were allowed to set. After tensile bond strength testing, the morphological aspects of the fractured surfaces were assessed by scanning electron microscopy and energy dispersive spectroscopy. The energy dispersive spectroscopy successfully traced sealer components to the debonded surfaces. Some of the sealers penetrated into the dentinal tubules when the dentin surface had been pretreated with acids. However, these sealer tags remained occluding the tubules after bond failure in some instances only (Grossmans sealer, RoekoSeal Automix with an experimental primer, AH Plus/EDTA). Penetration of the endodontic sealers into the dentinal tubules when the smear layer was removed was not associated with higher bond strength.

smear layer was found to prevent the penetration of root canal sealers inside the dentinal tubules (3, 4). Therefore, its removal has been recommended to improve the obturation seal (5, 6) and reduce coronal leakage (7). On the other hand, presence or absence of the smear layer was found to have no significant effect on the apical seal (8). In a previous study (9) we also did not find improved bond strength of the various sealers after removal of the smear layer. Documenting the mode of failure of sealers to different dentin surfaces and gutta-percha might improve our understanding of the role of the smear layer. The purpose of this investigation was 2-fold: (a) to assess the microscopic details of the morphological aspects of the debonded surfaces by scanning electron microscopy (SEM); and (b) to identify elements of the sealers on the dentin and gutta-percha surfaces by energy dispersive spectroscopy (EDS) after debonding.

MATERIALS AND METHODS Human single-rooted teeth, stored in 0.01% NaOCl at 4C after extraction, were used in this study. Root dentin cylinders, 4 mm in diameter, were cut in a buccolingual direction at a right angle to the tooths long axis. The cylinders were mounted in brass holders and ground flat against 500-grit silicon carbide abrasive paper (Struers, Copenhagen, Denmark). Gutta-percha cylinders of 4-mm diameter were prepared from heat-softened gutta-percha (Roeko, Langenau, Germany). The cylinders were mounted in brass holders and their end surfaces ground flat in a similar manner to the dentin cylinders. The dentin specimens were randomly divided into four equal groups and their surfaces conditioned with 37% H3PO4 for 30 s, 25% citric acid for 30 s, 17% EDTA for 5 min, or 10 ml of distilled H2O (control). The conditioned dentin surfaces were then rinsed with 10 ml of distilled H2O and dried with an air stream for 5 s. Each group of conditioned specimens were further divided into six equal subgroups (n 4), according to the type of sealer used. For the Grossmans sealer (GS)/phosphoric acid subgroup, the pH of the acid-treated dentin surface was measured using pHindicator strips (Merck KGaA, Darmstadt, Germany) after rinsing the surface with 10 ml of distilled H2O. Five sealers of different chemical composition were tested (Table 1). An experimental primer supplied with RoekoSeal Automix (RS) also was investigated. The procedures for testing were previously described (9). Briefly, the dentin and gutta-percha surfaces were coated with a thin layer of the freshly mixed sealer and the
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The standard method of obturation of the root canal system is by using a core material in combination with a root canal sealer. A desirable property of the root canal sealer is to have adhesive strength both to the dentin and to the core material, which is usually gutta-percha. The sealer also must have cohesive strength to hold the obturation together (1). The ability of root canal sealers to adhere to dentin and gutta-percha may be expected to result in superior sealing ability, which should reduce leakage in clinical situations. Adhesion also should improve the stability of the root filling, e.g. during preparation for post space. The role of the smear layer in endodontics has been the subject of considerable debate since it was first described in 1975 (2). The

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Saleh et al. TABLE 1. Root canal sealers tested Sealer Code GS Chemical Composition Zinc oxide-eugenolbased Manufacturer Prepared according to Grossman, NIOM laboratory, Haslum, Norway Vivadent, Schaan, Liechtenstein Espe, Seefeld, Germany Dentsply DeTrey GmbH, Konstanz, Germany Roeko, Langenau, Germany Roeko, Langenau, Germany

Journal of Endodontics

Batch Number 2

Grossmans sealer

Apexit Ketac-Endo AH Plus RoekoSeal Automix RoekoSeal Automix

AP KE AH RS RP

Calcium hydroxide-based Glass-ionomerbased Resin-based Silicone-based Silicone-based

912697 FW0055094 9810000713 B239b/B240b M496

Primer

cylinders immediately pressed together. The test specimens were kept in an incubator at 37C and a relative humidity of 90 5%. The sealers were allowed to set for 1.5 times the manufacturers stated setting time. Tensile bond strength was measured in a universal testing machine (Instron, Instron Limited, Bucks, UK). The debonded dentin and gutta-percha specimens were separated from their brass holders and air-dried. Two specimens from each subgroup were selected for SEM and EDS examination. Each specimen was mounted on an aluminium stub and sputter-coated with carbon in a Balzers SCD 050 coating unit (Balzers AG, Balzers, Liechtenstein). The specimens were first observed under a scanning electron microscope (Philips XL 30, Eindhoven, The Netherlands) at an operating voltage of 10 kV and photographed at a magnification of 200, 800, and 2000. The debonded surfaces were examined to indicate the mode of failure: adhesive (at the dentin/sealer or gutta-percha/sealer interface), cohesive within the sealer, or a mixture of the two. The specimens were then analyzed by an energy-dispersive spectroscope (EDAX, DX-4-i, Mahwah, NJ) to identify elements of the sealers on the debonded surfaces. X-ray intensities in counts per second were set at 100 and the accelerating voltage at 20 kV. Control root dentin discs were prepared in the same manner as the experimental ones. Each was treated with one of the four pretreatment solutions used in the study and processed in a similar manner for SEM examination. Control discs were prepared for each sealer, as well as for gutta-percha and dentin, and processed for EDS analysis. Basic elements in each material were detected and used as a reference for their presence on the fractured surfaces. RESULTS Control SEM Specimens The control dentin specimen pretreated with distilled H2O showed the retained smear layer (with tracks from the carbide paper polishing). The smear layer was completely removed with exposure of the underlying dentinal tubules in the control specimens pretreated with phosphoric or citric acids. No packed material was found in the tubule openings, whereas the specimen pretreated with EDTA showed evidence of smear plugs in the tubules. Control EDS Specimens The elements detected in each of the control specimens dentin, gutta-percha, and the different sealersare shown in Table 2.

TABLE 2. Elements detected by EDS for each of the materials used Material Dentin Gutta-percha AH Plus Apexit Ketac-Endo RoekoSeal Automix Grossmans sealer Elements Ca, P, Mg Zn, Ba, S Ca, W, Zr Ca, Bi, Si Ca, Si, Al, F Si, Zr Zn, Ba, S

Experimental Specimens For AH Plus (AH) sealer, debonded dentin surfaces showed areas of cohesive failure within the sealer as well as areas of adhesive failure to dentin (Fig. 1a). In the adhesive failure area, EDS confirmed that particles remaining on the phosphoric acidand citric acid-treated dentin were not from the sealer (Fig. 1b). All gutta-percha surfaces showed remnants of the sealer, although macroscopically no sealer residues could be detected (Fig. 1c). Zr and Si were detected by EDS analysis on gutta-percha surfaces that appeared clean. Tags protruding from the sealer remaining on gutta-percha were observed where adhesive failure to dentin, which had been pretreated with phosphoric and citric acids, was obvious (Fig. 1d). This was not the case when dentin was pretreated with EDTA, where sealer tags occluded the dentinal tubules as evident by EDS (Fig. 1, e and f). For RS, SEM examination of the debonded surfaces revealed an adhesive failure to dentin treated with distilled H2O only (Fig. 2, a and b). The gutta-percha surface was completely covered by the sealer. The same mode of failure was observed when dentin was pretreated with phosphoric acid or EDTA. In this case the dentinal tubules were exposed with no sealer inside (Fig. 2c). EDS confirmed that particles detected on the dentin surface were not from the sealer (Fig. 2d). On the citric acid-treated dentin the sealer was partly detached from the surface (Fig. 2e). The undersurface of the sealer layer revealed Ca and P peaks (Fig. 2f). No sealer could be identified in the dentinal tubules. Examination of the distilled H2O-treated dentin surface, to which a primer was applied before RS, showed complete coverage by the sealer indicating an adhesive failure to gutta-percha (Fig. 3, a and b). The failure was adhesive to both dentin and gutta-percha when the dentin was pretreated with phosphoric acid, citric acid, and EDTA. The dentin surface was partly covered by sealer remnants and partly showing exposed dentinal tubules (Fig. 3c). Zr and Si were detected in the material occluding some of the tubules (Fig.

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FIG 1. Debonded AH specimen showing both adhesive failure to dentin (AD) and cohesive failure within the sealer (C). (a) Scanning electron micrograph ( 800) of dentin surface partly covered with sealer; citric acid pretreatment. (b) EDS analysis of area (white square in a). The area appears to be clear dentin with complete absence of sealer elements. (c) Scanning electron micrograph ( 200) of gutta-percha surface covered with a continuous layer of sealer; citric acid pretreatment. (d) Tags protruding from the sealer remaining on gutta-percha; phosphoric acid pretreatment ( 800). (e) Sealer tags occluding the dentinal tubules; EDTA pretreatment ( 800). (f) EDS of area (white cross in e) showing the presence of W and Zr in addition to sealer elements. The Zr peak is overlapped by P.

FIG 2. Debonded RS specimen. (a) Scanning electron micrograph ( 800) of dentin surface covered with smear layer; distilled H2O pretreatment. (b) EDS analysis shows clear dentin with complete absence of sealer elements. (c) Scanning electron micrograph ( 800) of dentin surface with exposed dentinal tubules and particles on the surface; phosphoric acid pretreatment. (d) EDS analysis of the particles reveals no sealer elements. (e) Scanning electron micrograph ( 800) of dentin surface with partly detached sealer; citric acid pretreatment. (f) EDS analysis of area (white cross) reveals Ca and P.

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FIG 3. Debonded RP specimen. (a) Scanning electron micrograph ( 800) of dentin surface completely covered with sealer; distilled H2O pretreatment. (b) Scanning electron micrograph ( 800) of gutta-percha surface; distilled H2O pretreatment. (c) Scanning electron micrograph ( 800) of dentin surface partly covered with sealer. Open dentinal tubules with sealer tags; citric acid pretreatment. (d) EDS analysis of the content of the dentinal tubules revealed Si and Zr. (e) EDS analysis of area (white square in c) revealed Ca and P. (f) Scanning electron micrograph ( 2000) of gutta-percha surface covered with sealer. Tags were protruding from the sealer surface; citric acid pretreatment.

FIG 4. Debonded KE specimen. (a) Scanning electron micrograph ( 800) of dentin surface completely covered with sealer; citric acid pretreatment. (b) Scanning electron micrograph ( 200) of gutta-percha surface; citric acid pretreatment. (c) EDS analysis of gutta-percha surface revealed traces of sealer. (d) Scanning electron micrograph ( 800) of dentin surface partly covered with sealer. Open dentinal tubules with no sealer tags; EDTA pretreatment. (e) EDS analysis of area indicated (white square in d). The area appears to be clear dentin. (f) Scanning electron micrograph ( 800) of gutta-percha surface partly covered with sealer; EDTA pretreatment.

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FIG 5. Debonded GS specimen. (a) Scanning electron micrograph ( 800) of gutta-percha surface completely covered with sealer. Tags were seen on the sealer surface; phosphoric acid pretreatment. (b) EDS analysis of sealer revealed Ca and P in addition to sealer elements. (c) Scanning electron micrograph ( 800) of dentin surface partly covered with sealer. Sealer tags are seen inside the dentinal tubules; phosphoric acid pretreatment. (d) EDS analysis revealed Zn inside the dentinal tubules. (e) Scanning electron micrograph ( 800) of dentin surface showing thread-like precipitations. (f) EDS analysis of the precipitated material revealed Ca and P.

3d). Some crystals, not sealer particles as confirmed by EDS, were precipitated on the phosphoric acid- and citric acid-treated dentin surfaces (Fig. 3e). Sealer tags protruding from the detached sealer surface were occasionally seen. The gutta-percha surface was only partly covered by sealer remnants showing protruding tags (Fig. 3f). Except for EDTA-treatment, SEM examination of the debonded dentin surfaces revealed a continuous layer of Ketac-Endo (KE) sealer (Fig. 4a). The gutta-percha surface occasionally revealed small particles identified as KE by EDS (Fig. 4, b and c). The surface of dentin that had been pretreated with EDTA was only partly covered by KE (Fig. 4d). Open dentinal tubules were otherwise seen with no evidence of sealer in them. No peaks other than dentin elements were identified (Fig. 4e). The debonded gutta-percha surface was similarly partly covered by KE (Fig. 4f). SEM examination of the debonded surfaces of GS revealed an adhesive failure to dentin that had been treated with distilled H2O. The gutta-percha surface was completely covered with the sealer, whereas a smear layer was covering the dentin surface. In case of acid treatment, the failure was cohesive within the sealer and adhesive to dentin. The gutta-percha surface was completely covered with the sealer showing protruding tags. Ca and P were identified in addition to the elements of GS (Fig. 5, a and b). Remnants of the sealer were partly covering the dentin surface and occasionally occluding the exposed dentinal tubules (Fig. 5c). Zn peaks were always detected inside the tubules (Fig. 5d). In phosphoric acid-treated dentin, Ca and P were detected in the threadlike particles remaining on the dentin surface (Fig. 5, e and f). The pH of the phosphoric acid-treated dentin was measured to approximately 5.5 after rinsing with distilled H2O.

For Apexit (AP), a mixed type of failure, cohesive within the sealer and adhesive to dentin, was observed for all pretreatment conditions. A continuous layer of the sealer covered the guttapercha surfaces (Fig. 6a). The debonded dentin surfaces were only partly covered by the sealer (Fig. 6b). No sealer was found to occlude the exposed dentinal tubules. This was evident by the tags that projected from the surface of the sealer covering the gutta-percha.

DISCUSSION This study is the sequel to an earlier investigation of the effects of dentin pretreatment on the tensile bond strength of various endodontic sealers (9). We found that the highest bond strengths for all sealers with the exception of GS were recorded when the smear layer was not removed. Pretreatment of dentin with phosphoric and citric acids resulted in the highest bond strength for GS. The SEM observations of the debonded surfaces were confirmed by EDS analysis, which provides qualitative identification of the elements present in a given SEM field. The endodontic smear layer is known to form on the surface of dentinal walls when the root canals are instrumented (2). Oksan et al. (3) and Kouvas et al. (4) observed that the smear layer obstructed the penetration of sealer tags into the dentinal tubules. The present study confirms these findings. SEM pictures revealed penetration of sealer tags only when the smear layer had been removed. Such mechanical interlocking, however, did not result in higher bond strengths as demonstrated in the first part of this study (9). The highest bond strengths were in fact recorded when dentin

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FIG 6. Debonded AP specimen/EDTA pretreatment showing both adhesive failure to dentin (AD) and cohesive failure within the sealer (C). (a) Scanning electron micrograph ( 800) of gutta-percha surface covered with a continuous layer of sealer. (b) Scanning electron micrograph ( 800) of dentin surface partly covered with sealer.

had been treated with distilled H2O in most instances. This was interpreted as an enhancement of adhesion of sealers to dentin in the presence of the smear layer. The findings of Lalh et al. (10) also suggest an important role played by the smear layer in adhesion; they found that for glass-ionomer based sealers higher bond strengths were recorded when the smear layer was not removed by EDTA. The smear layer, at low magnification, has a typical amorphous structure and, at higher magnification, a globular pattern (11). It has been suggested that the bond that develops essentially occurs between the material and the smear layer and not the underlying, mineralized dentin matrix. The strength of the bond may be determined and limited by the strength of the forces holding the globules to each other and to the underlying dentin (11). The bond strengths recorded in the case of distilled H2O-treated dentin were not the same for all sealers, which suggests a different mechanism of bonding to the smear layer for the different sealers. AH and AP had the same pattern of failure (adhesive to dentin and cohesive within the sealer), yet their bond strengths differed significantly. This difference can be attributed to the different cohesive strengths of the two sealers. The low bond strength and the adhesive failure to distilled H2O-treated dentin for GS reflect its weak bond to smear layer. However, the bond strength increased significantly after treatment of dentin with phosphoric and citric acids, and the mode of failure was predominantly cohesive within the sealer. This may be related to the effects from the acids causing precipitation of zinc phosphate aggregates at the interface, which is confirmed by the detection of Zn inside the dentinal tubules. It has been shown earlier that precipitation of aggregates

or crystals at the interface may increase bond strength (12). Adhesion between KE and the smear layer was strong enough so that failure occurred at the weakest adhesive bonds, which were formed at the sealer gutta-percha interface. For RS, the mode of failure was adhesive to dentin with all pretreatment conditions, yet the bond strength was higher in case of distilled H2O, which means a stronger bond to smear layer than to smear-free dentin. This bond was even stronger with the application of the primer, as demonstrated by the higher bond strength recorded and by the failure mode becoming adhesive to gutta-percha. This suggests a possible interaction of the primer with the smear layer resulting in improved adhesion of RS. In the present study, phosphoric and citric acids and EDTA were used as dentin pretreatments. Pretreatment of dentin with phosphoric acid or citric acid was found to be more effective than EDTA in removing the smear layer (13). The particles that were occasionally observed on the dentin surface in the case of citric acid pretreatment and confirmed by EDS not to originate from the sealers are presumably a result of the crystallization of the acid that had remained after final irrigation. Copious irrigation is necessary to avoid residual crystals, presumably of calcium citrate. AH, AP, and GS were able to penetrate into the dentinal tubules when the smear layer was removed. However, these sealer tags remained occluding the tubules after bond failure in some instances only (GS, AH/EDTA). Otherwise the sealer together with the tags was detached from the surface. There was no indication of penetration of KE into the dentinal tubules after EDTA pretreatment. In cases of pretreatment with phosphoric or citric acids, however, it was not possible to assess the penetration inside the dentinal tubules, because the failure was adhesive to gutta-percha. Similarly, RS did not penetrate inside the tubules. For RoekoSeal Automix with an experimental primer (RP), the use of the primer greatly enhanced the penetration of the sealer into the tubules but with no significant increase in bond strength. The opening of the tubules and the loss of the smear layer are apparently not favorable processes for improving adhesion. There may be at least two possible explanations for this. The opened dentinal tubules may act as stress raisers, which would promote failure in the adhesive joint, and/or the smear layer and plugs are rich in calcium and phosphate and are potential sites for strong adhesive bonding (14). The present study showed that not all sealers penetrated inside the exposed dentinal tubules and that the bond strength was not higher for all sealers that were able to penetrate inside the tubules. This confirms the suggestion that micromechanical retention by penetration of sealer tags inside the tubules is not the only, and maybe not an important, factor affecting adhesion of root canal sealers (3). Moreover, tubular penetration is clearly dependent on the chemical and physical properties of the sealer (3). Adhesion of sealers to dentin and gutta-percha is a complex process with separate mechanisms operating for the various sealers and different dentin surfaces. An overall variability was observed among sealers for adhesive properties to dentin and gutta-percha. The mode of failure for AH and AP did not vary with the different dentin pretreatments, whereas for GS, KE, RS, and RP more than one type of failure was observed. Some of the sealers penetrated into the dentinal tubules when the dentin surface had been pretreated with acids. However, these sealer tags remained occluding the tubules after bond failure in some instances only. The penetration of the sealer tags into the dentinal tubules was not associated with higher bond strength values (9).

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Drs. Saleh, Ruyter, and rstavik are affiliated with the NIOMScandinavian Institute of Dental Materials, Haslum, Norway. Drs. Saleh and Haapasalo are affiliated with the Department of Endodontics, Faculty of Dentistry, University of Oslo, Oslo, Norway. Address requests for reprints to Iman M. Saleh, NIOMScandinavian Institute of Dental Materials, P.O. Box 70, N-1305 Haslum, Norway.

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