ORIGINAL ARTICLE

Shear bond strength and ultrastructural interface analysisof different adhesive systems to Er:YAG laser-prepared dentin

Yeliz Guven & Oya Aktoren

Received: 23 April 2012 /Accepted: 15 August 2013# Springer-Verlag London 2013

Abstract The aim of this study was to evaluate the shearbond strength (SBS) of a microhybrid composite resin bondedwith three different adhesive systems to Er:YAG laser- (EL) orbur-prepared dentin surfaces and to analyze the quality andultrastructure of the adhesive–dentin interfaces by scanningelectron microscopy (SEM). The specimens prepared for SBStest and SEM analysis were randomly assigned to eight groups(G1–G8): G1, EL (Fidelis PlusIII, Fotona)+Clearfil S3 Bond(C3S); G2, EL+AdperSE Plus (SE); G3, EL+laser etch+Adper Single Bond2 (SB2); G4, EL+acid etch+SB2; G5,EL+SB2 (no etching); G6, bur+acid etch+SB2; G7, bur+S3; G8, bur+SE. Laser was used in very short pulse mode at asetting of 200 mJ/20 Hz for dentin preparation and at 80 mJ/10 Hz for dentin etching. Bond strength test: 3.5×2.0 mmcylindrical molds were placed onto adhesives and filled withthe composites. After 24 h in distilled water, SBSwas tested ata crosshead speed of 0.5 mm/min. SEM analysis: The dentin–adhesive interfaces were evaluated for the ultrastructure ofhybrid layer. Data of SBS (MPa) were statistically analyzedby ANOVA and Tukey HSD. ER:YAG laser-prepared dentinhas demonstrated significantly more SBS (p <0.01) for SEwhen compared to bur-prepared dentin. No significancies (p >0.05) in SBS have been determined between the total-etchadhesive applied groups with regard to etching types. SEManalysis revealed that hybrid layers obtained in Er:YAG laser-irradiated dentin exhibited more irregular and non-homogeneous pattern than the conventionally prepared den-tin. In conclusion, SE Bond demonstrated superior results inEr:YAG laser-ablated dentin compared to bur-prepareddentin.

Keywords Er:YAG laser . Adhesive . Bond strength . Hybridlayer

Introduction

Increased understanding of the caries process with the scien-tific researches and significant progress in caries diagnosis,dental materials, and cavity preparation techniques with thetechnological advances has led to the remarkable scientificadvance in dentistry over the past several decades. The adventof the use of acid etching technique on tooth surfaces pro-voked the studies of the adhesive restorative systems andrevolutionized the dental practice eliminating the need forthe removal of healthy tooth structure and thus allowing amore conservative and esthetic treatment [1, 2].

Despite continuous efforts in adhesive dentistry to developsimpler, more user-friendly, and less technique-sensitive ad-hesives, bonding to dentin and the complete sealing of theexposed dentinal surfaces remain problematic because of thecomplex and hydrated nature of this substrate [1, 3].

The surface created at the end of cavity preparation plays asignificant role in the bonding of the adhesive restorative ma-terials to tooth structure [4]. Drilling with a bur is the mostcommon way of cavity preparation, but bur preparation ofcavities often leads to more extensive cavities. This drawbackof rotary instruments for cavity preparation, along with thecurrent trend toward “minimal invasiveness,” has led to theintroduction of alternative methods for cavity preparation suchas laser irradiation [5, 6].

The Er:YAG laser is one of the most recommended types oflasers to be used on dental hard tissues, because its wavelength(2.94 μm) coincides with the main absorption peak of water(3.0 μm), and it is also well absorbed by OH− groups inhydroxyapatite. This provides a good interaction with all bio-logical tissues, including enamel, dentin, and cementum [7, 8].

Y. Guven (*) :O. AktorenDepartment of Pedodontics, Faculty of Dentistry, Istanbul University,Çapa, Istanbul 34093, Turkeye-mail: yelizgn@yahoo.com

Lasers Med SciDOI 10.1007/s10103-013-1424-0

The Er:YAG laser acts on dental substrate by thermo-mechanical ablation. The energy delivered by the laser irradia-tion heats and evaporates the water content in the target tissue,causing a rise of internal pressure within the tissue until theexplosive destruction of inorganic substance occurs [9, 10].

Erbium lasers produce minimal thermal damage to the pulpand surrounding tissues, especially when irradiated with con-tinuous water irrigation [8]. Other advantages of using lasers forhard tissue preparation include bactericidal effects and lessnoise, vibration, and discomfort for the patient than a rotaryhandpiece [11, 12]. Absence of smear layer on laser-treateddentin, open dentinal tubules, and microirregularities due to thepreferential removal of the intertubular dentin suggests that theresultant dentin surface is receptive to adhesive procedure [13].

Since the morphological appearance of lased dentin signif-icantly differs from that of conventionally treated dentin, therehas been a major interest to investigate the interaction patternbetween the currently available adhesive systems and thelaser-irradiated dentin [1, 14].

The bond strength of adhesive systems is one of the majorfactors to be considered when making an esthetic restoration. Aneffective adhesion to tooth structure is of great importance towithstand the stresses resulting from polymerization shrinkage,thereby enhancing retention and marginal integrity of the resto-rations [1]. Despite its efficiency, reported bond strengths ofcomposite resin to tooth substrate prepared by erbium laser areoften confusing and contradictory [11]. The morphological anal-ysis of resin–dentin interfaces gives a more reliable prognosisabout the bonding ability of adhesive systems to tooth surfaces,and therefore, it is required for the clarification of the interactionpatterns formed on lased dentin–adhesive bonding [15].

Since all adhesive systems were originally developed to acton tooth substrate prepared by conventional techniques, it isstill needed to investigate the adhesion performance of variousadhesive systems on laser-prepared dental surfaces.

Therefore, the aim of this study was to evaluate the shearbond strength (SBS) of a microhybrid composite resin bondedwith three different adhesive systems to Er:YAG laser- andbur-prepared dentin surfaces and to analyze the quality andultrastructure of the adhesive–dentin interfaces by scanningelectron microscopy (SEM). The tested hypothesis was thatthere is no significant difference in the interactionmorphologyand bond strength to dentin between composite and dentinsurfaces prepared by bur or Er:YAG laser ablation and bondedwith the tested adhesive systems.

Materials and methods

Tooth selection and preparation

One hundred and thirty-two freshly extracted, caries- andrestoration-free human third molars were selected for the

study. The teeth, thoroughly hand-scaled and cleaned withwater/pumice slurry using a low-speed handpiece, were storedin distilled water at 4 °C and used within 6 months afterextraction in accordance with the recommendations of theInternational Organization for Standardization (ISO) [16].

SBS test

One hundred and twenty teeth were used in the SBS test; theroots of the teeth were removed 2 mm below thecementoenamel junction, and the crowns were embedded inautopolymerizing acrylic resin with buccal surfaces posi-tioned for surface treatment and composite bonding. Afterpolymerization of the embedding resin, buccal surfaces wereabraded and then sequentially polished in a polishing machineusing 400- and 600-grit silicon carbide paper until a uniformlayer of peripheral dentin was observed.

After the dentin surfaces had been controlled for the ab-sence of enamel and/or pulp tissue with a stereomicroscope, asmall piece of insulating tape with a central hole was preparedin order to delimit the dentin bonding site. The tape perfora-tion was made by means of a rubber-dam punch modified toprovide 3.5-mm-diameter holes.

The specimens were randomly assigned to eight groups(n =15) according to the surface treatment and adhesive system.The first five groups were prepared with an Er:YAG laser, andlast three groups were prepared conventionally and served ascontrol groups.

Group 1 Er:YAG laser preparation, Clearfil Tri-S (C3S)Bond

Group 2 Er:YAG laser preparation, Adper SE Plus (SE)Bond

Group 3 Er:YAG laser preparation, laser etching, AdperSingle Bond 2 (SB2)

Group 4 Er:YAG laser preparation, acid etching, Adper Sin-gle Bond 2 (SB2)

Group 5 Er:YAG laser preparation, no etch, Adper SingleBond 2 (SB2)

Group 6 Diamond bur, acid etching, Adper Single Bond 2(SB2)

Group 7 Diamond bur, Clearfil Tri-S (C3S) BondGroup 8 Diamond bur, Adper SE Plus (SE) Bond

The Er:YAG laser used in this study was the Fidelis Plus III(Fotona Medical Lasers, Ljubljana, Slovenia) with a wave-length of 2.94 μm. Laser energy was delivered using a R02-Chandpiece with a spot size of 0.9 mm in diameter in a non-contact mode under continuous water spray (40–60 mL/min) ata focal distance of 7 mm from the target point. Avisible aimingbeam equipped with the handpiece guided the user to work onthis specific focal distance of 7 mm which coincides with thesmallest possible diameter of the aiming laser beam. The irra-diations were made by moving the handpiece continuously

Lasers Med Sci

above the tooth surface in order to obtain a pattern of rows andcolumns that overlapped. The laser parameters were chosenaccording to the manufacturer’s instructions: dentin, 200 mJat 20 Hz (energy density 62.9 J/cm2); and dentin etching, 80 mJat 10 Hz (energy density 15.73 J/cm2). The pulse duration was100 μs (very short pulse).

Conventional preparations were performed with a #10diamond fissure bur (Strauss & Co., Industrial DiamondsLtd., Ra’anana, Israel) in a high-speed handpiece (KaVo Den-tal, Bismarckring, Germany) using an air and water spraycoolant. New burs were used after every five preparations.

Following the dentin surface preparation, group 3 was laseretched in a focused, non-contact mode at a setting of 80mJ/10 Hz with continuous water irrigation; groups 4 and 6 werechemically etched with 35 % phosphoric acid (Scotchbond,3 M ESPE, St. Paul, MN, USA) for 15 s and rinsed with waterfor 15 s. Care was taken not to desiccate the treated surfacesby removing any excess water with a cotton pellet instead ofdrying with compressed air.

Details about the components and chemical formulationsof the adhesives and their application procedures according tothe manufacturer’s instructions are listed in Table 1. ClearfilTri-S Bond, a one-step self-etching adhesive, was applied ingroups 1 and 7; Adper SE Plus Bond, a two-step self-etchingadhesive, was applied in groups 2 and 8. In groups 3, 4, 5, and6, Adper Single Bond 2, an etch-and-rinse adhesive, was usedas the bonding agent. All adhesives were light cured for 10 s(Elipar FreeLight 2 LED, 3M ESPE, St Paul, MN, USA).

An adhesive tape with a 3.5-mm-diameter central hole wasplaced over the dentin surface to limit the bonding area.Following the respective adhesive applications, a split-bisected Teflon mold, with a circular hole (3.5 mm in diameterand 2 mm in height) was positioned over the hole in theadhesive tape. A composite resin (Filtek Z250, 3M ESPE,St. Paul, MN, USA) was placed and light cured in the mold,

forming cylindrical posts perpendicular to the dentin surface.After curing had been completed, Teflon matrix surroundingthe composite was opened and separated, leaving a resincylinder (3.5 mm in diameter×2 mm height) that adhered tothe delimited surface. A complementary 20-s polymerizationwas accomplished to ensure that the specimens were ade-quately polymerized. After storage in distilled water at37 °C for 24 h, the specimens were submitted to SBS test ina universal testing machine at a crosshead speed of 0.5 mm/min until fracturing occurred.

All materials and methods used complied with the recom-mendations of the International Organization for Standardiza-tion (ISO) (ISO/TS 11405:2003, Dental materials. Testing ofadhesion to tooth structure).

Statistical analysis was carried out using the NCSS 2007&PASS 2008 (UT, USA) statistical software program. One-wayanalysis of variance (ANOVA) and post hoc Tukey’s HSDmultiple comparisons were used to determine statistical dif-ferences in SBS between the groups at a significance level of0.05.

Fractured specimens were examined using a stereomicro-scope (Zeiss Stemi SV6; Carl Zeiss, Gottingen, Germany) at×30 magnification to assess the failure modes. Failure modeswere classified as adhesive failure (failure occurred at thespecimen/adhesive interface), cohesive failure (failure oc-curred in the material or the substrate with no damage to theinterface), and mixed failure (failure occurred in the interfaceand the material at the same time).

Scanning electron microscopy examination

The remaining 12 teeth were used for SEM investigation. Rootsof the teeth were removed 2 mm below the cementoenameljunction, and remaining fragments were embedded in clearepoxy resin (Epofix; Struers A/S, Copenhagen, Denmark)

Table 1 Adhesive systems used in the study

Product name Manufacturer Composition Application

Clearfil Tri-S Bond (S3) Kuraray Medical, Tokyo,Japan

10-MDP, Bis-GMA, HEMA, initiator,ethanol, water, stabilizer, nanofiller,hydrophobic dimethacrylate

Apply bond and leave for 20 s. Dry withhigh-pressure air blow for more than5 s. Light cure for 10 s

Adper SE Plus Bond (SE) 3M ESPE, St. Paul. MN,USA

Liquid A: water, HEMA, Surfactant,Pink colorant. Liquid B : UDMA,TEGDMA, TMPTMA (hydrophobictrimethacrylate), HEMA phosphates,MHP (methacrylated phosphates)Bonded zirconia nanofiller, Initiatorsystem based on camphorquinone

Apply Liquid A. Apply Liquid B for 20 sand see the color change. Air-dry for 10 s.Reapply the Liquid B and light-cure

Adper Single Bond 2(SB2)

3M ESPE, St. Paul. MN,USA

Adhesive: water, ethanol, Bis-GMA,HEMA, UDMA, bisphenol Aglycerolate, polyalkenoic silicananoparticles treated with acidcopolymer, dimethacrylate

Apply adhesive for 15 s. Gently air-dryfor 5 s. Light cure for 10 s

Lasers Med Sci

using 30-mm-diameter polyethylene molds. The occlusal thirdof the embedded tooth was removed with a horizontal cut 3mmbelow the occlusal surface, using a low-speed saw (Minitom,Struers, Copenhagen, Denmark) equipped with a diamondimpregnated disc under water cooling. No enamel was visibleexcept at the periphery.

Each dentin surface was divided into two equal areas, andstandardized Class I cavities with a mesiodistal andbuccolingual width of 2 mm and a depth of 1 mm wereprepared in each. Both cavities were at the same locationlabiolingually. A total of 24 cavities were assigned to the eightgroups (n =3) as previously described. In order to compare thecavity preparation methods in the same adhesive appliedgroups (in groups 1 and 7, groups 2 and 8, groups 4 and 6),one cavity of the specimen was prepared with a round dia-mond bur in a high-speed drill under air–water spray, and theother one was prepared with an Er:YAG laser R02-Chandpiece. The remaining three specimens including six cav-ities were treated with Er:YAG laser and prepared accordingto the protocols of groups 3 and 5.

After the application of adhesive systems and acid asdescribed in Table 1, the cavities were restored with amicrohybrid composite resin (Filtek Z250) by bulk techniqueand were light activated separately for 20 s with a LED curinglight. The samples were then left in distilled water at 37 °C for1 week.

Specimens were cross-sectioned longitudinally in a mesio-distal direction by a cut through the center of the restorationusing a low-speed diamond disc saw in order to expose theresin–dentin interface (Fig. 1). Exposed surfaces weresmoothened and polished with #1,000–#4,000-grit SiC paperson a polishing machine (Metkon GRIPO 1V, Bursa, Turkey).The teeth were cleaned in an ultrasonic device containingdistilled water for 10 min to remove any residues resultingfrom the sectioning and polishing procedures on the surface.The surfaces were then decalcified with 37 % phosphoric acidgel for 10 s and rinsed with distilled water. They were im-mersed in 3 % NaOCl solution for 60 s and rinsed with waterto dissolve all collagen dentinal tissue. One week later, thespecimens were mounted on aluminum stubs, desiccated,sputter coated with gold/palladium, and examined in a scan-ning electron microscope (JEOL JSM 6335F Field Emission;

JEOL Ltd., Tokyo, Japan). During the analysis, the mostrepresentative areas of dentin/adhesive system interfaces werephotographed at different magnifications (×500, ×1,000,×3,000). The photographs were assessed as related to thepresence or absence of hybrid layer, its integrity, thickness,homogeneity, and continuity along the interface, as well as theexistence of tags, their shape, and number.

Results

SBS test

The mean and standard deviations of SBSs in MPa are shownon Fig. 2. One-way ANOVA revealed significant differencesin mean SBS values between the groups (p <0.001). Thehighest SBS was observed in Er:YAG laser-prepared and SEBond applied group, while the lowest SBS was observed inbur-prepared and SE Bond applied group.

When the effects of laser and bur preparation methods onSBS of each adhesive system were compared, the only statis-tically significant difference was found between the SE ap-plied groups. In SE Bond applied groups, the specimenstreated with the Er:YAG laser demonstrated significantlyhigher mean bond strength values than those treated with thebur (p <0.01). No statistically significant differences wereobserved between mean SBS results of the SB2 and C3SBond applied groups, regardless of whether dentin wasEr:YAG laser ablated or bur treated (p >0.05).

In Er:YAG laser-treated groups, SE Bond showed thehighest scores, whereas the lowest scores were obtained withSB2 with or without acid etching (p <0.01). No statisticallysignificant differences were found between C3S and SE Bondas well as between C3S and SB2 with or without acid etching.In bur-treated groups, no statistically significant differenceswere observed among the tested adhesive systems (p <0.05).In Er:YAG laser-treated and SB2-applied groups, no signifi-cant difference was found onmean SBS values among etched,non-etched, and laser-etched groups (p >0.05).

The fracture failure modes of all groups were listed inTable 2. In all samples that were conventionally prepared(G6, G7, and G8), irrespective of the restorative system, the

Fig. 1 Preparation of specimensbefore SEM examination

Lasers Med Sci

predominant mode of failure was mixed failure. For theEr:YAG laser-prepared and SB2-applied samples, Er:YAGlaser-etched and no-etched groups (G3 and G5) mostlyshowed cohesive failure modes, while acid-etched group(G4) showed primarily mixed failure type. The predominantmode of failure in the Er:YAG laser-prepared and self-etchedapplied groups (G1 and G2) was mixed failure.

Scanning electron microscopy evaluation

Representative SEM micrographs of the resin–dentin inter-faces of the three adhesive systems bonded to Er:YAG laser-or bur-prepared dentin surfaces are shown in Figs. 3, 4, and 5.

Er:YAG laser-prepared cavities showed more irregular in-terfaces than the bur-prepared cavities. In Er:YAG laser-prepared cavities, the hybridization zones were hard to find,due to scarcity and discontinuity of such interdiffusion areaalong the resin–dentin interface. In addition, analysis of theresin–dentin interfaces demonstrated the presence of gaps inEr:YAG laser-prepared cavities.

Hybrid layer was observed in all groups except the Er:YAGlaser-prepared and SB2 applied (no etching) group. Also, thehybrid layer in Er:YAG laser-prepared cavities presented re-markable differences in thickness and continuity along theresin–dentin interface. With all the adhesives tested, a thinnerand non-homogeneous pattern of hybrid layer formation was

0 10 20 30 40

Er:YAG laser + Clearfil Tri-S Bond

Er:YAG laser + Adper SE Plus Bond

Er:YAG laser + laser etch + AdperSingle Bond 2

Er:YAG laser + acid etch + Adper SingleBond 2

Er:YAG laser + Adper Single Bond 2 (noetch)

Bur + acid etch + Adper Single Bond 2

Bur + Clearfil Tri-S Bond

Bur + Adper SE Plus Bond

21.38

26.45

20.45

17.46

15.2

17.81

18.82

13.18

4.08

5.41

6.93

5.11

4.68

4.24

3.55

2.59

Mean Shear Bond Strength Standard Deviation

Fig. 2 Mean shear bond strengths (MPa) and standard deviations of the groups

Table 2 Type of fracture failuresin groups Groups Cohesive failure Adhesive failure Mixed failure

Er:YAG laser+Clearfil Tri-S Bond 6 (40 %) 0 9 (60 %)

Er:YAG laser+Adper SE Plus Bond 4 (26.7 %) 1 (6.6 %) 10 (66.7 %)

Er:YAG laser+laser etch+Adper Single Bond 2 8 (53.4 %) 2 (13.3 %) 5 (33.3 %)

Er:YAG laser+acid etch+Adper Single Bond 2 4 (26.7 %) 2 (13.3 %) 9 (60 %)

Er:YAG laser+Adper Single Bond 2 (no etch) 7 (46.7 %) 1 (6.6 %) 7 (46.7 %)

Bur+acid etch+Adper Single Bond 2 1 (6.6 %) 6 (40 %) 8 (53.4 %)

Bur+Clearfil Tri-S Bond 2 (13.3 %) 4 (26.7 %) 9 (60 %)

Bur+Adper SE Plus Bond 2 (13.3 %) 3 (20 %) 10 (66.7 %)

Lasers Med Sci

observed after laser preparation and hybridization, whereasthe bur-treated surfaces show a thicker and homogeneoushybrid layer formation after hybridization.

Resin tags were observed in all the resin–dentin interfacesregardless of whether the cavities were Er:YAG laser ablatedor bur treated; however, resin tags in bur-prepared cavitieswere fewer than the Er:YAG laser-prepared cavities. Resintags appeared in a cylindrical shape except the acid etchingperformed groups that showed characteristic funnel-shapedresin tags.

In SE Bond applied groups, hybrid layer in Er:YAG laser-prepared cavities was thicker than the bur-prepared cavities.

Conversely, in C3S Bond applied groups, a thinner hybridlayer was detected in Er:YAG laser-prepared cavities com-pared to the bur-prepared cavities.

Discussion

Our study compared the in vitro SBS of a microhybrid com-posite resin bonded with three different adhesive systems toEr:YAG laser- or bur-prepared dentin and investigated thequality and ultrastructure of the resin–dentin interfaces bySEM. The null hypothesis claiming no difference between

Fig. 3 Representative SEMmicrograph of the resin–dentininterfaces of the specimenstreated with Er:YAG laser (a[×1,000], b [×3000]) and bur (c[×1,000], d [×3,000]) bondedwith Clearfil Tri-S Bond. a ,bNote the presence of gap alongthe interface, hybrid layerformation, and numerous resintags with lateral branches. c ,dNote the hybrid layer formation,fewer resin tags, and tightadhesion with no visible gap. Aadhesive system, C compositeresin, D dentin, HL hybrid layer,RT resin tags

Fig. 4 Representative SEMmicrograph of the resin–dentininterfaces of the specimenstreated with Er:YAG laser (a[×1,000], b [×3,000]) and bur (c[×1,000], d [×3,000]) bondedwith Adper SE Plus Bond. a ,bNote the non-uniform adhesivelayer, presence of gap along theinterface, non-homogeneoushybrid layer formation, andthicker resin tags with lateralbranches. c ,d Note the formationof a thin hybrid layer, fewer resintags, and tight adhesion with novisible gap. A adhesive system, Ccomposite resin, D dentin, HLhybrid layer, RT resin tags

Lasers Med Sci

Er:YAG laser- and bur-prepared cavity surfaces in dentinrestorative systems was rejected.

Although some investigators have used micro-tensile ortensile bond strength methods when evaluating the bondingability of adhesive systems to dentin, SBS testing was chosenin the present study. Shear stresses are believed to be majorstresses involved in in vivo bond failures of restorative mate-rials. Various investigators have suggested SBS testing as aviable method for predicting clinical performance [17, 18].

Ultramorphological and chemical characteristics of thetooth substrate have been considered to be important factorsfor adhesion. The use of the Er:YAG laser for cavity prepara-tion results in a surface with a significantly different morphol-ogy to that achieved by conventional techniques. Erbiumlasers created an irregular and microretentive morphologicalpattern of the surface with open dentinal tubules and free of asmear layer [19, 20]. These highly irregular cavity surfaceswithout a smear layer are believed to provide suitable surfacesfor strong bonding with composite resin materials, and laserirradiation is considered to be an alternative to conventionalacid etching [21]. In this regard, some of the previous inves-tigations have reported that the bond strength of composite tolaser-irradiated dentin was higher than or equal to that of acid-etched dentin [22, 23]. It was thought that openings of dentinaltubules after the Er:YAG laser treatment might facilitate theformation of a hybrid zone, since primer and adhesive canpenetrate the surface better when the smear layer is removed[21, 22]. On the contrary, some of the investigations havereported a decrease in bond strength to laser-irradiated dentin.Ceballos et al. explained the decreased bond strength in laser-ablated dentin by the formation of a laser-modified layer.Basal part of this layer appeared to lack of interfibrillar spaces

due to the fusion of the remnant denatured collagen fibrils.The presence of this fused layer without interfibrillar spaceswas thought to restrict the resin diffusion into the subsurfaceintertubular dentin and thereby resulting in lower SBS. Acidetching and water rinsing steps are believed to remove thismodified lased tissue [10]. In our study, data obtained from theetch-and-rinse adhesive applied groups demonstrated thatSBS of Er:YAG laser-etched group (G3) was higher thanthose of acid-etched groups (G4,G6), although the differencesdid not reach statistical significance. This result agrees withthe findings of Bertrand et al. [23] who did not find anysignificant difference between bur and Er:YAG laser with orwithout subsequent acid treatment. This result is also in agree-ment with the findings obtained by Celik et al. [24] who usedthe same etch-and-rinse adhesive system tested in this studyfound similar SBS values in Er:YAG laser-treated+acid-etched and bur-treated+acid-etched groups. The high variabil-ity in results can be attributed not only to the methodologiesand varying laser parameters but also the different character-istics of various adhesive systems tested.

Ramos et al. [14], Trajtenberg et al. [25], and Brulat et al.[26] observed that bond strength values obtained with self-etch adhesives on Er:YAG laser-treated dentin surfaces weregenerally lower than those measured on bur-treated dentinsurfaces. Ramos et al. [27] explained the limited effectivenessof self-etch adhesives on laser-treated dentin by the limitedcapacity of the acidic monomer to demineralize the laser-modified superficial layer and alter the morphological patternresulting from it. According to Brulat et al. [26], lower valuesof SBS measured with self-etch adhesive systems on Er:YAGlaser-prepared dentin surfaces could be ascribed to the ab-sence of a smear layer based on the findings of Tay et al. [28].

Fig. 5 Representative SEMmicrograph of the resin–dentininterfaces of the specimensbonded with Adper Single Bond2. a Laser-treated and acid-etchedgroup. Note the non-uniformadhesive layer, presence of gapalong the interface, hybrid layer,and funnel-shaped resin tagformation. b Bur-treated andacid-etched group. Note theuniform adhesive layer, thickerand conical-shaped resin tags. cLaser-treated and laser-etchedgroup. Note the non-homogeneous hybrid layer that insome parts is absent. d Laser-treated and non-etched group.Note the gap formation andabsence of hybrid layer. Aadhesive system, C compositeresin, D dentin, HL hybrid layer,RT resin tags

Lasers Med Sci

According to them, the smear layer residues form globularsubunits within the polymerized resin and would, thus, act liketrue mineral fillers. In addition, it has been reported that laserirradiation chemically modifies the tooth surfaces. Irradiationwith erbium lasers reduces the carbon-to-phosphorus ratio andleads to the formation of more stable and less acid-solublesurfaces [29]. According to Esteves-Oliviera et al., the weakacids present in the self-etching systems cannot sufficientlymodify this more acid-resistant surface to promote adhesivepenetration [30]. On the contrary, in the conducted research,self-etch adhesive applied groups yielded higher bondstrengths in the Er:YAG laser-treated groups than those ofbur-treated groups. However, the only statistically significantdifference was found between SE Bond applied groups. SEMmicrographs of the SE Bond applied groups showed thathybrid layer in Er:YAG laser-prepared cavities was thickerthan the bur-prepared cavities, and this observation was alsoconsistent with the SBS results of SE Bond applied groups.Improved adhesion of SE bond on laser-treated surfaces maybe explained by the resultant modified dental substrate. Laser-ablated surface demonstrated an increase in calcium andphosphate compounds and a reduction in carbonate and waterafter thermal effects and crystallography changes [31]. Car-boxyl groups in SE Bond can chemically bond to hydroxyap-atite and calcium and thus form stable calcium salts thatenhance resin adhesion by the formation of strong ionic inter-actions between the substrate and adhesive layer.

Fracture analysis of the bonded dentin surface in Er:YAGlaser-prepared groups revealed a high incidence of cohesivefailures compared to bur-prepared groups. These data are inagreement with the fracture pattern findings of Dunn et al. [12]and Martinez-Insua et al. [32]. Dunn et al. postulated thatcohesive dentin failures in laser-prepared specimens may beexplained by the lack of penetration of resin into Er:YAGirradiated dentin, creating a weak subsurface zone just belowthe interface [12]. SEM images in the present study demon-strating the gaps below the interface strongly confirm thisprediction. In a study conducted by Çelik et al. [24], self-etch (Clearfil Protect Bond and Clearfil tri-S Bond) appliedgroups mostly showed mixed failure modes which is consis-tent with the results of our self-etched applied groups.

The conducted study revealed that the Er:YAG laserinfluenced the morphology of the resin–dentin interface. Thisis probably due to the fact that the different kinds of surfacetreatment have different effects on the dentin surface morphol-ogy. In the present study, gap formation along the resin–dentininterface was observed in laser-prepared cavities. This obser-vation is in agreement with those of Aranha et al. [33] whoreported that such gaps could be a result of collagen alteration.Although the effect of laser on collagen network is still notcompletely clear, it is known that laser irradiation is able todevelopmicrostructural alteration as well asmicrorupture of thecollagen fibers [33]. The literature has strongly emphasized that

such alterations of the collagen structure could lead to incom-plete penetration of primers and monomers [34]. It was also notpossible in this study to evaluate the collagen network since thephosphoric acid and sodium hypochlorite were applied duringthe preparation of specimens for the SEM examination.Benezzato and Stefani [35] reported that Er:YAG laserablation denatures dentinal collagen fibers in deep regionsof dentin. However, in the first portion of the dentinaltubules, collagen appears with a normal aspect indicatingthat hybrid layer formation is possible. The presenting gapsjust below the interface can also be attributed to this weaksubsurface zone.

Resin–dentin interface analysis of Er:YAG laser-treatedgroups revealed an irregular and a discontinuous pattern ofhybrid layer along the interface. In several specimens, a typ-ical resin–dentin interdiffusion zone was hardly identified. Onthe other hand, bur-treated groups revealed well-defined, ho-mogeneous, and continuous hybrid layer formation. Thesefindings agree with those of previous studies [20, 36–38].According to Barceleiro et al., the irregular characteristics ofhybrid layer seem to be related to the effect of laser beamincidence on acid resistant of the tooth structures. When thelaser beam is perpendicular to the tooth substrate, an acid-resistant structure will form, and thus, the hybrid layer wouldsuffer damage in this part. On the other hand, in the regionaround the laser beam, a lower temperature increase willoccur, and less acid-resistant structure will form [37, 39].Aranha et al. [33] and Sassi et al. [36] observed thinner hybridlayer in laser prepared groups. In the present study, hybridlayer in laser-prepared specimens was thinner than that of bur-prepared specimes except the SE Bond applied groups whichshowed a similar thickness to the bur-prepared ones. Aranhaet al. [33] reported that the number and regularity of the resintags in laser-treated groups were higher than those observed inthe bur-treated groups for all the adhesives tested. This is inagreement with our SEM results. Conversely, Sassi et al. [36]and Schein et al. [39] reported that resin tags in laser groupswere less pronounced than the bur groups.

Many factors can influence the bonding performance ofadhesive systems to dentin including the dentin substrate, thesurface treatment, and the testing procedures. It is thereforesuggested that the bond strength data substantially varyamong the investigations [24]. It is of utmost importance tostandardize as many variables as possible or at least providethe details of all the variables used in the study [40]. ISOguidance on adhesive testing standardizes adhesion testing,detailing how tests should be performed [16]. In the presentstudy, the ISO guidance on adhesive testing was used for theselection of the tooth substrate, storage of the test specimens,designing the test equipment, and setting up the test parame-ters. In order to exclude possible influences of different re-storative resins on the bond strength, all the bonding agents inthis study were used with the Z250 composite resin. One

Lasers Med Sci

limitation of this study was that no additional aging procedurewas applied on the specimens.

The infiltration of resin monomers into demineralizeddentin results in the creation of a new structure called theresin–dentin interdiffusion zone or hybrid layer [41]. Hy-brid layer is the critical area of the adhesive bonding sincethe stability of the bonded interface relies on the creation ofa compact and homogenous hybrid layer. Morphologicalanalysis of this layer enables an assessment of the qualityof the bond [42, 43]. Therefore, in addition to the SBS test,ultramorphological analysis of adhesive system–dentin in-terface was performed in our study. Quantitative analysis ofthe hybrid layer was not possible due to the irregular anddiscontinuous pattern of hybrid layer in the laser-treatedgroups, and this can be regarded as another limitation ofthe study.

Different results obtained with the different adhesive sys-tems on Er:YAG laser-treated dentin surfaces in the presentstudy suggest that particular characteristics of the adhesivesystems, such as the physicochemical properties and interac-tion pattern with the lased substrate, have a strong influenceon the success of the resin–dentin bond. Therefore, furtherinvestigations should be undertaken to determine which ad-hesive protocol/systems should be preferred to yield an opti-mal bonding to laser-treated dental substrate. Moreover, in thefuture, new adhesive systems can be developed specific to thelaser-treated surfaces.

Conclusion

Based on the findings of the conducted research and within thelimitations of an in vitro study, it may be concluded that:

& When SE was used, mean SBS value measured onEr:YAG laser-prepared surfaces was significantly higherthan that observed on bur-prepared surfaces. When SBand C3S were used, no statistically significant differenceswere observed on SBS values when bur- and Er:YAGlaser-prepared dentin surfaces were compared.

& No significancies in SBS have been determined betweenthe total-etch adhesive applied groups with regard to etch-ing types

& SEM analysis revealed that hybrid layers obtained in laseddentin exhibited more irregular and non-homogeneouspattern than the conventional prepared dentin.

& SEMmicrographs of the SE Bond applied groups showedthat hybrid layer in Er:YAG laser-prepared cavities wasrelatively thicker than the bur-prepared cavities.

Acknowledgments We would like to thank Dt. Zafer Kazak for pro-viding the Er:YAG laser equipment.

References

1. Torres CP, Gomes-Silva JM, Borsatto MC, Barroso JM, Pécora JD,Palma-Dibb RG (2009) Shear bond strength of self-etching and total-etch adhesive systems to Er:YAG laser-irradiated primary dentin. JDent Child (Chic) 76:67–73

2. Murdoch-Kinch CA, McLean ME (2003) Minimally invasive den-tistry. J Am Dent Assoc 134:87–95

3. VanMeerbeek B, Yoshihara K, Yoshida Y,Mine A, DeMunck J, VanLanduyt KL (2011) State of the art of self-etch adhesives. Dent Mater27:17–28

4. Van Meerbeek B, De Munck J, Mattar D, Van Landuyt K,Lambrechts P (2003) Microtensile bond strengths of an etch&rinseand self-etch adhesive to enamel and dentin as a function of surfacetreatment. Oper Dent 28:647–660

5. Van Landuyt K, DeMunck J, Coutinho E, PeumansM, Lambrecht P,Van Meerbeek B (2005) Bonding to dentin: smear layer and theprocess of hybridization. In: Eliades G, Watts DC, Eliades T (eds)Dental hard tissues and bonding: Interfacial phenomena and relatedproperties, 1st edn. Springer, Berlin, pp 89–115

6. Lussi A, GygaxM (1998) Iatrogenic damage to adjacent teeth duringclassical approximal box preparation. J Dent 26:435–441

7. Gurgan S, Kiremitci A, Cakir FY, Yazici E, Gorucu J, GutknechtN (2009) Shear bond strength of composite bonded to erbium:yttrium-aluminum-garnet laser-prepared dentin. Lasers Med Sci24:117–122

8. Hibst R, Keller U (1989) Experimental studies of the application ofthe Er:YAG laser on dental hard substances: I. Measurement of theablation rate. Lasers Surg Med 9:338–344

9. De Moor RJ, Delme KI (2009) Laser-assisted cavity preparation andadhesion to erbium-lased tooth structure: part 1. Laser-assisted cavitypreparation. J Adhes Dent 11:427–438

10. Ceballos L, Toledano M, Osorio R, Tay FR, Marshall GW (2002)Bonding to Er-YAG-laser-treated dentin. J Dent Res 81:119–122

11. Obeidi A, Liu PR, Ramp LC, Beck P, Gutknecht N (2010) Acid-etchinterval and shear bond strength of Er, Cr:YSGG laser-preparedenamel and dentin. Lasers Med Sci 25:363–369

12. Dunn WJ, Davis JT, Bush AC (2005) Shear bond strength and SEMevaluation of composite bonded to Er:YAG laser-prepared dentin andenamel. Dent Mater 21:616–624

13. Manhães L, Oliveira DC, Marques MM, Matos AB (2005) Influenceof Er:YAG laser surface treatment and primer application methods onmicrotensile bond strength self-etching systems. Photomed LaserSurg 23:304–312

14. Ramos RP, Chimello DT, Chinelatti MA, Nonaka T, Pécora JD,Palma Dibb RG (2002) Effect of Er:YAG laser on bond strength todentin of a self-etching primer and two single-bottle adhesive sys-tems. Lasers Surg Med 31:164–170

15. Heintze SD (2007) Systematic reviews: I. The correlation betweenlaboratory tests on marginal quality and bond strength. II. The corre-lation between marginal quality and clinical outcome. J Adhes Dent9(Suppl 1):77–106

16. International Organization for Standardization ISO/TS 11405 (2003)Technical Specification Dental materials—testing of adhesion totooth structure, 2nd edition. Switzerland

17. Swift EJ Jr, Perdigão J, Heymann HO (1995) Bonding to enamel anddentin: a brief history and state of the art, 1995. Quintessence Int 26:95–110, 17

18. Gurgan S, Kiremitci A, Cakir FY, Gorucu J, Alpaslan T, YaziciE, Gutknecht N (2008) Shear bond strength of composite bondedto Er, Cr:YSGG laser-prepared dentin. Photomed Laser Surg 26:495–500

19. Freitas PM, Navarro RS, Barros JA, de Paula EC (2007) The use ofEr:YAG laser for cavity preparation: an SEM evaluation. MicroscRes Tech 70:803–808

Lasers Med Sci

20. Bertrand MF, Hessleyer D, Muller-Bolla M, Nammour S, Rocca JP(2004) Scanning electron microscopic evaluation of resin-dentininterface after Er:YAG laser preparation. Lasers Surg Med 35:51–57

21. Kohara EK, Hossain M, Kimura Y, Matsumoto K, Inoue M, Sasa R(2002) Morphological and microleakage studies of the cavities pre-pared by Er:YAG laser irradiation in primary teeth. J Clin Laser MedSurg 20:141–147

22. Visuri SR, Gilbert JL, Wright DD, Wigdor HA, Walsh JT Jr (1996)Shear strength of composite bonded to Er:YAG laser-prepared dentin.J Dent Res 75:599–605

23. Bertrand MF, Semez G, Leforestier E, Muller-Bolla M, Nammour S,Rocca JP (2006) Er:YAG laser cavity preparation and compositeresin bonding with a single-component adhesive system: relationshipbetween shear bond strength and microleakage. Lasers Surg Med 38:615–623

24. Celik EU, Ergücü Z, Türkün LS, Türkün M (2006) Shear bondstrength of different adhesives to Er:YAG laser-prepared dentin. JAdhes Dent 8:319–325

25. Trajtenberg CP, Pereira PN, Powers JM (2004) Resin bond strengthand micromorphology of human teeth prepared with an Erbium:YAGlaser. Am J Dent 17:331–336

26. Brulat N, Rocca JP, Leforestier E, Fiorucci G, Nammour S, BertrandMF (2009) Shear bond strength of self-etching adhesive systems toEr:YAG-laser-prepared dentin. Lasers Med Sci 24:53–57

27. Ramos RP, Chinelatti MA, Chimello DT, Borsatto MC, Pécora JD,Palma-Dibb RG (2004) Bonding of self-etching and total-etch sys-tems to Er:YAG laser-irradiated dentin. Tensile bond strength andscanning electron microscopy. Braz Dent J 15:I9–I20

28. Tay FR, Sano H, Carvalho R, Pashley EL, Pashley DH (2000) Anultrastructural study of the influence of acidity of self-etching primersand smear layer thickness on bonding to intact dentin. J Adhes Dent2:83–98

29. Apel C, Meister J, Schmitt N, Gräber HG, Gutknecht N (2002)Calcium solubility of dental enamel following sub-ablative Er:YAGand Er:YSGG laser irradiation in vitro. Lasers Surg Med 30:337–341

30. Esteves-Oliveira M, Zezell DM, Apel C, Turbino ML, Aranha AC,Eduardo Cde P, Gutknecht N (2007) Bond strength of self-etchingprimer to bur cut, Er, Cr:YSGG, and Er:YAG lased dental surfaces.Photomed Laser Surg 25:373–380

31. Moldes VL, Capp CI, Navarro RS, Matos AB, Youssef MN, CassoniA (2009) In vitro microleakage of composite restorations prepared by

Er:YAG/Er, Cr:YSGG lasers and conventional drills associated withtwo adhesive systems. J Adhes Dent 11:221–229

32. Martínez-Insua A, Da Silva DL, Rivera FG, Santana-Penín UA(2000) Differences in bonding to acid-etched or Er:YAG-laser-treatedenamel and dentin surfaces. J Prosthet Dent 84:280–288

33. Aranha AC, De Paula EC, Gutknecht N, Marques MM, RamalhoKM, Apel C (2007) Analysis of the interfacial micromorphology ofadhesive systems in cavities prepared with Er, Cr:YSGG, Er:YAGlaser and bur. Microsc Res Tech 70:745–751

34. Nakabayashi N, Saimi Y (1996) Bonding to intact dentin. J Dent Res75:1706–1715

35. Benazzato P, Stefani A (2003) The effect of ER:YAG laser treatmenton dentin collagen: An SEM investigation. J Oral Laser Appl 3:79–81

36. Sassi JF, Chimello DT, Borsatto MC, Corona SA, Pecora JD,Palma-Dibb RG (2004) Comparative study of the dentin/adhesivesystems interface after treatment with Er:YAG laser and acid etchingusing scanning electron microscope. Lasers Surg Med 34:385–390

37. de Barceleiro MO, de Mello JB, de Mello GS, Dias KR, de MirandaMS, Sampaio Filho HR (2005) Hybrid layer thickness and morphol-ogy: the influence of cavity preparation with Er:YAG laser. OperDent 30:304–310

38. de Barceleiro MO, Dias KR, Sales HX, Silva BC, Barceleiro CG(2008) SEM evaluation of the hybrid layer after cavity preparationwith Er:YAG laser. Oper Dent 33:294–304

39. Schein MT, Bocangel JS, Nogueira GE, Schein PA (2003) SEMevaluation of the interaction pattern between dentin and resin aftercavity preparation using ER:YAG laser. J Dent 31:127–135

40. al-Salehi SK, Burke FJ (1997) Methods used in dentin bonding tests:an analysis of 50 investigations on bond strength. Quintessence Int28:717–723

41. Eick JD, Gwinnett AJ, Pashley DH, Robinson SJ (1997) Currentconcepts on adhesion to dentin. Crit Rev Oral Biol Med 8:306–335

42. Heintze SD, Zimmerli B (2011) Relevance of in vitro tests of adhe-sive and composite dental materials. A review in 3 parts. Part 3:in vitro tests of adhesive systems. Schweiz Monatsschr Zahnmed121:1024–1040

43. Breschi L, Mazzoni A, Ruggeri A, Cadenaro M, Di Lenarda R, DeStefano DE (2008) Dental adhesion review: aging and stability of thebonded interface. Dent Mater 24:90–101

Lasers Med Sci