Effect of Chemomechanical Caries Removal on Bonding of Self-etching Adhesives to Caries-affected Dentin Hamdi Hosni Hamdan Hamamaa / Cynthia Kar Yung Yiub / Michael Frances Burrowc
Purpose: To evaluate the effect of enzyme-based (Papacárie) and sodium-hypochlorite–based (Carisolv) chemomechanical caries removal methods on bonding of self-etching adhesives to caries-affected dentin, in comparison to the standard rotary-instrument caries removal method. Materials and Methods: Seventy-eight carious permanent molars exhibiting frank cavitation into dentin were used. Forty-eight teeth were randomly divided into three groups, according to the caries excavation methods: (i) Papacárie, (ii) Carisolv and (iii) a round steel bur. After caries removal, each group was subdivided into two groups for two-step (Clearfil SE Bond) or one-step (Clearfil S3 Bond) self-etching adhesive application and resin composite buildups. Bonded specimens were sectioned into beams for microtensile bond strength testing. Bond strength data were analyzed using three-way ANOVA and Tukey’s test. For interfacial nanoleakage evaluation using a field-emission scanning electron microscope, caries was similarly removed from the remaining thirty carious molars, bonding was performed as for bond strength testing, and the teeth were sectioned. Results: Results of three-way ANOVA revealed that bond strength was significantly affected by “adhesive” (p < 0.001) and “dentin” (p < 0.001), but not “caries excavation methods” (p > 0.05). The bond strength of the two-step self-etching adhesive was significantly higher than that of the one-step self-etching adhesive (p < 0.001). Conversely, the bond strength of self-etching adhesives to sound dentin was significantly higher than to residual caries-affected dentin (p < 0.001). Greater silver penetration was observed in the bonded interfaces of residual caries-affected dentin and in interfaces bonded with the one-step self-etching adhesive vs those bonded with the two-step self-etching adhesive. Conclusion: Chemomechanical caries removal did not affect the bonding of self-etching adhesives to cariesaffected dentin as compared to caries excavation with rotary instruments. Keywords: chemomechanical caries removal, Carisolv, MDP, Papacárie, self-etching adhesives, caries-affected dentin. J Adhes Dent 2014; 16: 507–516. doi: 10.3290/j.jad.a33250
M
inimally invasive caries excavation techniques, including laser ablation, air abrasion, sonic abrasion, and chemomechanical caries removal, are characterized
a
Clinical Assistant Professor, Department of Operative Dentistry, Esthetic and Restorative Dentistry, Faculty of Dentistry, Mansoura University, Egypt. Ideas, hypothesis and experimental design, performed experiment in partial fulfillment of PhD degree, wrote manuscript.
b
Clinical Professor, Faculty of Dentistry, The University of Hong Kong, Prince Philip Dental Hospital, Hong Kong SAR, China. Idea, hypothesis and experimental design, proofread manuscript.
c
Professor and Chair, Biomaterials Department, Melbourne Dental School, The University of Melbourne,Victoria, Australia. Proofread manuscript.
Correspondence: Professor Cynthia Yiu, Paediatric Dentistry and Orthodontics, Faculty of Dentistry, The University of Hong Kong, Prince Philip Dental Hospital, 34 Hospital Road, Sai Ying Pun, Hong Kong SAR, China. Tel: +8522859-0251, Fax: +852-2559-3803. e-mail: [email protected]
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Submitted for publication: 03.05.14; accepted for publication: 09.10.14
by a common feature of selective removal of cariesinfected tissue, leaving the caries-affected tissues intact.14 The chemomechanical caries removal method was introduced by Goldman18 in 1967, with the first chemomechanical caries removal agent being GK-101,18 which was developed to GK101E (Caridex National Patent Dental Products; New Brunswick, NJ, USA) in 1984. The clinical application of Caridex in caries removal was limited due to the complexity of its delivery equipment and the relatively long time needed for caries excavation compared to conventional methods.6 To overcome these problems, Carisolv (Medi Team Dentalutveckling / Rubicon Life Science; Göteborg, Sweden) was introduced in 1998 and further developed in 2004.16 Carisolv is a sodium-hypochlorite-based (NaOCl) agent, which chlorinates and disrupts hydrogen bonds of the partially degraded collagen in carious dentin, facilitating its removal.27 507
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In 2005, a new chemomechanical caries removal agent ‘Papacárie’ (Formula and Acao; São Paulo, Brazil), which is a papain enzyme-based agent, was introduced by Bussadori et al.8 Papain is a proteolytic enzyme with bactericidal and anti-inflammatory actions. The precise mechanism of action of papain is unknown. It has been reported that infected carious tissues have lost the antiprotease α-1-anti-trypsin, which inhibits protein digestion in sound collagen-based tissues.8 In 2009, Bertassoni et al4 reported that papain enzyme could partially degrade intact nonmineralized type I collagen fibrils from rat tail tendon. Furthermore, papain has been shown to reduce the mechanical properties of intact mineralized dentin as a result of degradation of matrix proteoglycans, suggesting that the action of papain might be nonspecific.4 Self-etching adhesives are classified into two groups according to the number of clinical steps needed for bonding, ie, one- or two-step self-etching adhesives. In onestep self-etching adhesive systems, the acidic primer and the bonding agent are supplied in one bottle, whereas in two-step self-etching adhesive systems, the acidic primer and the bonding agent are supplied and applied separately in two steps. The self-etching adhesives are now classified into four types according to their acidity: ultramild (pH > 2.5), mild (pH 2–2.5), intermediate (pH 1–2), and strong (pH < 1).47 Bonding to caries-affected dentin is one of the most controversial topics in adhesive dentistry. Caries-affected dentin is characterized by a marked reduction in mineral content, loss of crystallinity, and alteration of the organic matrix.50 This complex substrate poses a great challenge to achieving reliable bonding. Previous studies have reported higher bond strengths of etch-and-rinse adhesives than self-etching adhesives to caries-affected dentin.10,40 Conversely, other studies have claimed that acid etching might lead to further loss of inorganic content of the caries-affected dentin.28,48 This may result in poor hybridization of the demineralized caries-affected dentin by adhesive resin.8 In contrast, Omar et al34 considered self-etching adhesives the “preferred adhesive” for bonding to the wetter, caries-affected dentin, due to its hydrophilic nature. Selecting the appropriate selfetching adhesive system compatible with caries-affected dentin raised another area of controversy, particularly with the development of one- and two-step self-etching adhesives. Several studies have evaluated the nanoleakage patterns22 and microtensile bond strengths5,7,17,25 of self-etching adhesives bonded to residual dentin following chemomechanical caries removal. Previous studies have mainly compared the effect of sodium-hypochlorite– based (NaOCl) chemomechanical caries removal agents, mainly Carisolv, on bonding to residual caries-affected dentin with the conventional rotary caries removal method. These studies reported that application of Carisolv did not compromise bonding to caries-excavated dentin.3,7,25,40,43 However, very few studies have examined the effect of enzyme-based chemomechanical caries removal agents (Papacárie or the experimental Biosolv [SFC-V and SFC508
VIII, 3M-ESPE; Seefeld, Germany] gels) on bonding to caries-affected dentin.3,31,32 Botelho Amaral et al5 evaluated the effect of bonding etch-and-rinse and self-etching adhesives to in-situ–formed, caries-affected dentin after the use of enzyme-based chemomechanical caries removal. The results of the study showed that bonding of the tested adhesives to caries-affected dentin was not affected by the application of Papacárie gel. Gianini et al17 examined the bond strength of etch-and-rinse and self-etching adhesives to demineralized dentin after the use of a papain-based chemomechanical caries removal method and showed that the papain-based caries removal method did not interfere with the bonding of the adhesives to demineralized dentin. However, the in-situ–formed demineralized dentin differed from the natural carious lesions due to the lack of vital odontoblasts. Hence, the aim of our current study was to compare the effect of enzyme-based (Papacárie) and sodium-hypochlorite–based (Carisolv) chemomechanical caries removal methods on bonding of self-etching adhesives to caries-affected dentin from extracted carious molars with the rotary caries removal method. The null hypotheses tested were that (i) chemomechanical caries removal has no effect on bonding of self-etching adhesives to caries-affected dentin and (ii) there is no difference in bond strength between the two-step self-etching (Clearfil SE Bond, Kuraray; Tokyo, Japan) and one-step self-etching (Clearfil S3 Bond, Kuraray) adhesives.
MATERIALS AND METHODS Seventy-eight permanent molars exhibiting frank moderate cavitation (severe decay, stage 5, ICDAS [International Caries Detection and Assessment System]) on the occlusal surface extending to the dentin were used. The molars were stored in 0.5% chloramine-T solution at 4°C for a period of less than one month following extraction. Forty-eight carious molars were used for microtensile bond strength testing, while the remaining thirty were used for nanoleakage evaluation. The study was approved by the local Institutional Review Board (IRB ref no: UW 11-355). Tooth Preparation Using a slow-speed diamond impreganated disk (Isomet, Buehler; Lake Bluff, IL, USA), the occlusal enamel was removed perpendicular to the long axis of the tooth to expose a flat dentin surface. The teeth were then randomly divided into three groups (n = 16) according to the caries excavation method: y Group 1: The carious cavity was treated with Papacárie (batch No 8996, Formula and Acao). The gel was left for 30 s prior to excavating the dentin using a #4 Carisolv noncutting instrument (Medi Team Dentalutveckling / Rubicon Life Science). Once the gel became cloudy, it was rinsed off with distilled water for 20 s and the process was repeated until successive applications of the gel failed to become cloudy. The Journal of Adhesive Dentistry
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Table 1
Adhesive systems used in the study
Brand name
Type
Composition
pH
Solvent
Manufacturer
Batch #
Clearfil SE
Two-step self-etching
Primer: 10-MDP, HEMA, hydrophilic dimethacrylate di-camphorquinone, N-N diethanol P-toluidine, water Bond: 10-MDP, bis-GMA, HEMA, hydrophobic dimethacrylate, di-camphorquinone, N-N diethanol P-toluidine, silanated colloidal silica
approx. 2*
Water
Kuraray Noritake Dental; Tokyo, Japan
11563
Clearfil S3
One-step self-etching
10-MDP, bis-GMA, HEMA, di-camphorquinone, ethanol, water, silanated colloidal silica
approx. 2.7**
Water, ethanol
Kuraray Noritake Dental
11113
Abbreviations:10- MDP,10-methacryloxydecyl dihydrogen phosphate; HEMA, 2-hydroxyethyl methacrylate; bis-GMA, bis-phenol diglycidyl methacrylate. *Chiaraputt et al.11 ** Botelho Amaral et al5 and Shinoda et al.41
y Group 2: The carious lesion was treated with Carisolv multimix gel (batch No. 10489, Medi Team Dentalutveckling / Rubicon Life Science), supplied as a twin syringe two-gel system. The gel was mixed automatically (Static Mixer, Medi Team Dentalutveckling / Rubicon Life Science) in the syringe tip prior to application on the carious lesion. The gel was left for 30 s prior to excavating the dentin using the #4 Carisolv noncutting instrument. Similar to Papácarie, once the gel became cloudy, it was rinsed away and the process was repeated until successive applications of the gel failed to become cloudy. y Group 3: Caries removal was performed with a lowspeed handpiece (NSK; Tochigi, Japan) at 35,000 rpm using a round steel bur (#012, Midwest, Dentsply; Surrey, UK) after application of a caries detector dye (batch No. 758AA, Kuraray) to define the carious lesion and verify the removal of “caries-infected” dentin, leaving lightly stained pink “caries-affected” dentin as per the manufacturer’s instructions. Bonding Procedures After caries removal, each group was subdivided into two groups (n = 8) according to the self-etching adhesive used. A two-step self-etching adhesive, Clearfil SE Bond (Kuraray) was used for the first group, and a one-step self-etching adhesive, Clearfil S3 (Kuraray), was used for the second group (Table 1). The adhesives were applied following the manufacturer’s instructions, followed by light curing of the adhesive for 10 s using a quartz-halogen light-curing unit (Elipar 2500, 3M ESPE; St Paul, MN, USA) with a light intensity output of 670 mW/cm2. The bonded surface was covered with a resin composite (Filtek Z250, 3M ESPE; St Paul, MN, USA). The resin composite was applied in 2-mm-thick increments and light cured for 20 s at the same light intensity. The teeth were stored in distilled water at 37°C for 24 h. Microtensile Bond Strength Testing All the bonded teeth were cut perpendicularly and vertically in the “x” and “y” directions through the bonded interface to obtain beams (0.9 mm x 0.9 mm wide; 5 to 7 mm long), each consisting of composite, adhesive, Vol 16, No 6, 2014
and dentin. At least sixteen bonded beams from each tooth were selected; eight of the bonded beams were from the flattest central region of the caries-affected dentin and the remaining from the sound dentin area of the same tooth to act as the control beams. The cross-sectional area of each specimen was measured with a pair of digital calipers (Model CD-6BS, Mitutoyo; Tokyo, Japan). The beams were attached to a Bencor Multi-T device (Danville Engineering; San Ramon, CA, USA) using a cyanoacrylate resin (Zapit, Dental Ventures of America; Corona, CA, USA) and subjected to tensile stress in a universal testing machine (Model 4444, Instron; Norwood, MA, USA) at a crosshead speed 1 mm/ min. The experimental unit of the current study was the tooth; this means that the recorded value of each group represented the mean microtensile bond strength value of the eight teeth. The number of beams which failed prior to testing was recorded for each group, and they were replaced by intact bonded beams for bond strength testing. The beams with pre-test failures were excluded from the statistical analysis. Analysis of Failure Modes All fractured beams were observed under polarized light microscopy (Nikon polarizing Microscope Eclipse LV100POL, Nikon; Tokyo, Japan) at 100X magnification to determine the mode of failure: cohesive in dentin (CD), adhesive (AD), mixed (MI), or cohesive in composite (CC).20 Representative specimens from each group were examined by field-emission scanning electron microscope (FE-SEM) (Hitachi S-3400N, Hitachi High Technologies America; Schaumburg, IL, USA) to confirm the failure mode as observed by polarized light microscopy. Nanoleakage Evaluation The remaining 30 carious molars were randomly allocated, and caries removal and bonding were performed following as for microtensile bond strength testing, except that the subgroup sample size was 5. Each bonded tooth was sectioned into halves through the cavity along the mesio-distal plane using a low-speed water-cooled diamond saw (Isomet, Buehler). The bonded specimen was coated with two layers of nail varnish within 1 mm of the bonded interface. The specimens were rehydrated 509
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Table 2 Percentages of beams with pre-test failures and microtensile bond strength of one-step and two-step self-etching adhesives to sound and residual dentin following caries excavation with Papacárie, Carisolv, or rotary instrument Caries Removal Methods Papacárie Sound dentin
Carisolv
Residual cariesexcavated dentin
Sound dentin
Rotary
Residual cariesexcavated dentin
Sound dentin
Residual cariesexcavated dentin
Beams with pre-test failures/ (percentage) Clearfil SE Bond
4 / (6.3%)
6 / (9.1%)
4 / (6.3%)
7 / (10.4%)
3 / (4.8%)
7 / (10.4%)
Clearfil S3 Bond
5 / (7.7%)
11/ (15.5%)
6 / (8.8%)
12 / (16%)
6 / (8.8%)
10 / (14.3%)
Microtensile Bond Strength (MPa) * Clearfil SE Bond
38.01a ± 2.4
31.5b ± 2.8
43.0a ±
3.4
30.8b ± 2.7
40.4a ± 4.2
26.1b ± 3.7
Clearfil S3 Bond
26.5b,c ± 1.6
23.2c ± 1.9
23.9c ± 2.1
20.1c ± 2.2
29.2b ± 4.7
17.4c ± 1.0
*Values are means ± standard deviation, n = 8 teeth/group. Multiple comparison test, all groups compared to all other groups; groups identified by different superscript letters (both in rows and columns) are significantly different at p < 0.05.
in distilled water for 10 min, immersed in 50% (W/V) ammoniacal silver nitrate solution and kept in total darkness for 24 h, and then rinsed with distilled water for 5 min. Specimens were subsequently immersed in photodeveloping solution (Kodak GBX fixer and replenishers, Kodak; Rochester, NY, USA) and exposed to fluorescent light for 8 h in order to reduce the diamine silver ions into metallic silver grains within voids along the bonded interface. Specimens were further rinsed with distilled water for 5 min. The observed surface was wet polished with 600-, 800-, 1200-, 2400-, and 4000- grit silicon carbide papers (Microcut, Buehler), followed by polishing with a polishing cloth using 6-, 3-, 1-μm diamond pastes (Diamat, Pace Technologies; Tucson, AZ, USA). Specimens were then ultrasonically cleaned, air dried for 24 h, mounted on aluminum stubs, and carbon coated. The resin/dentin interface was analyzed using FE-SEM operated in backscattered electron mode at 10 mm working distance and 12 KV accelerating voltage (ACCV). Statistical Analysis The bond strength data were analyzed using a statistical software package (SPSS, V.19, IBM; Armonk, NY, USA). Since the bond strength data were normally distributed (Kolmogorov-Smirnov test) and homoscedastic (Levine test), they were analyzed using three-way ANOVA to examine the effect of three variables (caries excavation method, adhesive, and dentin) and the interaction of the three factors on the microtensile bond strength. Tukey’s post-hoc multiple comparison test was performed to compare the means between the groups, with statistical significance set at α = 0.05. A Kruskal-Wallis test with Dunnett’s post-hoc test was used to compare the incidence of different failure modes among the different groups. A significance level of 5% was employed for all analyses. 510
RESULTS Microtensile Bond Strength Test The mean microtensile bond strengths of two-step and one-step self-etching adhesives to residual dentin following caries excavation with Papacárie, Carisolv, or rotary instruments are shown in Table 2. Results of the three-way ANOVA revealed that the microtensile bond strengths were significantly affected by the factors “adhesive” (p < 0.001) and “dentin” (p < 0.001), but not the factor “caries excavation method” (p > 0.05). The interaction of all three factors, “caries excavation method”, “adhesive”, and “dentin” was significant (p < 0.05). In general, the bond strengths of the two-step selfetching adhesive were significantly higher than those of the one-step self-etching adhesive (p < 0.001). The bond strength of self-etching adhesives to sound dentin was higher than that to residual caries-excavated dentin (p < 0.001). The method of caries removal had no effect on the bond strengths of two-step and one-step self-etching adhesives to residual dentin following caries excavation (p > 0.05). The percentage of beams with pre-test failures was higher in the residual caries-affected dentin in comparison to sound dentin areas. Moreover, it was found that the percentage of the beams with pre-test failures was higher in the one-step self-etching adhesive group than in the corresponding two-step self-etching adhesive group (Table 2). As shown in Table 3, statistical analysis (KruskalWallis test with Dunnett’s post-hoc test) revealed an increase in the percentage of cohesive failures in dentin when bonded to residual caries-excavated dentin vs sound dentin in Papacarie/S3 group, Rotary/SE, and Rotary/S3 groups. The mixed failure mode was commonly observed in the sound dentin groups. The percentage of cohesive failure in composite failure was very low. RepreThe Journal of Adhesive Dentistry
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Table 3 Failure mode analysis (Kruskal-Wallis with Dunnett’s post-hoc test) Group
Fracture mode
(%)
Statistical difference
P SE-SD
CD AD MI CC
9.38 31.25 56.25 3.13
A, B
P SE-CAD
CD AD MI CC
7.81 26.56 64.06 1.56
A, B
P S3-SD
CD AD MI CC
9.38 43.75 46.94 0.00
B
P S3-CAD
CD AD MI CC
28.13 32.81 39.06 0.00
C
C SE-SD
CD AD MI CC
6.25 43.75 46.88 3.13
B
C SE-CAD
CD AD MI CC
26.56 32.81 31.25 9.38
B, C
C S3-SD
CD AD MI CC
12.50 60.94 26.56 0.00
C
C S3-CAD
CD AD MI CC
37.50 34.38 28.13 0.00
C
R SE-SD
CD AD MI CC
4.51 17.01 75.35 3.13
A
R SE-CAD
CD AD MI CC
16.84 18.58 64.58 0.00
B
R S3-SD
CD AD MI CC
1.39 19.17 73.37 6.08
A
R S3-CAD
CD AD MI CC
19.97 26.22 53.82 0.00
B
CAD
b
CAD
CD: cohesive failure in dentin; AD: adhesive failure; MI: mixed failure; CC: cohesive failure in composite. P: Papacárie: C: Carisolv; R: rotary: S3: Clearfil S3 bond: SE: Clearfil SE bond; SC: sound dentin; CAD: cariesaffected dentin. Groups identified by different letters (left column) are significantly different at p < 0.05.
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a
Fig 1 SEM micrographs showing cohesive failure in residual caries-affected dentin following (a) NaOCl-based (Carisolv) and (b) enzyme-based (Papacárie) chemomechanical caries removal. More destruction of dentin surface was observed in Carisolv (arrow) than in Papacárie groups. CAD: caries-affected dentin.
sentative SEM micrographs from the chemomechanical caries removal groups with cohesive failure in dentin are shown in Fig 1. Nanoleakage Analysis Representative FE-SEM micrographs showing nanoleakage patterns of the bonded interfaces from the Papacárie, Carisolv, and rotary groups are shown in Figs 2 to 4. In general, the hybrid layers in the residual cariesaffected dentin groups were thicker than those in the sound dentin groups. Silver nitrate uptake was higher in the bonded interfaces of the residual caries-affected dentin groups than in the sound dentin groups. Similarly, the silver uptake at the bonded interfaces formed by the one-step self-etching adhesive was higher than those formed by the two-step self-etching adhesive. In the Papacárie caries-excavated groups, silver penetration was observed within the intertubular dentin; while 511
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b
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in the rotary caries-excavated groups (Rotary SE and Rotary S3), silver penetration was observed on the top of the excavated dentinal surface. However, both nanoleakage patterns were observed in the Carisolv caries-excavated groups (Carisolv/SE and Carisolv/S3).
DISCUSSION Previous studies which evaluated the microtensile bond strength of residual caries-affected dentin following enzyme-based chemomechanical caries removal used demineralized dentin to avoid the effect of caries lesion size, shape, depth, and surface irregularities on the results of the bond strength test.5,17 However, results obtained from demineralized dentin should be interpreted with caution, because natural carious dentin contains a wide variety of pathogenic materials, organic acids, hydrolytic enzymes, and whitlockite occluding the dentinal tubules.30 Kinney et al22 reported that artificial loss of mineral content led to collapse of the collagen fibers, which might affect resin hybridization into dentin and bond strength results. The microtensile bond strength test used in the current study considered the tooth as the test unit and calculated the mean bond strength value using the beams from each tooth. This reduced bond strength variation and permitted testing a broad area of carious lesion. The flattest portion of the residual caries-affected dentin was selected to reduce the microshear forces on the adhesive interface during microtensile bond strength testing.40 Furthermore, using the sound dentin from the same tooth as control decreases the variation of results in comparison with using another sound tooth as the control group. 512
Fig 2 Back-scattered SEM images showing the nanoleakage patterns of the Papacárie group. a: Papacárie/Clearfil S3/caries-affected dentin. b: Papacárie/ Clearfil S3/sound dentin. c: Papacárie/ Clearfil SE/caries-affected dentin. d: Papacárie/ Clearfil SE/sound dentin. The residual caries-affected dentin (CAD) (a and c) showed higher silver uptake than did sound dentin (b and d). The one-step self-etching adhesive groups (a and b) showed higher silver uptake than did the two-step self-etching adhesive groups (c and d). Silver penetration was observed within the intertubular dentin in the residual CAD following enzyme-based (Papacárie) chemomechanical caries removal (hand pointers). Ad: adhesive; HL: hybrid layer; D: dentin.
The present results showed that there was no significant difference in bond strength among the three caries excavation methods. The null hypothesis that chemomechanical caries removal has no effect on bonding of selfetching adhesives to caries-affected dentin could not be rejected. Previous studies which evaluated the morphological changes of dentin following chemomechanical caries removal reported that the excavated surface following Papacárie excavation was rough and characterized by a total absence of smear layer, with mostly patent dentinal tubules.19 In contrast, the excavated surface following Carisolv was irregular and partially covered with a smear layer with most of the dentinal tubules being patent or partially occluded.19,25 These morphological characteristics may increase the dentinal surface area for micromechanical retention of adhesive resin following caries removal. Conversely, many studies reported that the excavated surface following caries removal using burs was relatively smooth, and covered with a smear layer with occlusion of tubules.15,19,44 Some studies reported that the smear layer might act as a natural barrier preventing resin infiltration into dentin.12,25 However, Lohbauer et al26 reported that hybridization of self-etching adhesive resin with dentin mainly depended on bonding of hydrophilic monomers to the exposed collagen scaffold, and that the resin tags did not influence bond strength. Furthermore, Oliveira et al33 found that the smear layer formed following cutting with carbide burs was thin, with moderate surface roughness, and that the bond strength to dentin surfaces prepared by carbide bur was higher than to surfaces prepared by diamond bur and silicon paper. Abdebayo et al1 mentioned that application of adhesive resin with a scrubbing motion enhanced resin penetration into dentin. Therefore, the thin smear layer following caries excavation The Journal of Adhesive Dentistry
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Fig 3 Back-scattered SEM images showing the nanoleakage patterns of Carisolv groups. a: Carisolv/Clearfil S3/caries-affected dentin. b: Carisolv/ Clearfil S 3/sound dentin. c: Carisolv/ Clearfil SE/caries-affected dentin. d: Carisolv/ Clearfil SE/sound dentin. The residual caries-affected dentin (a and c) showed higher silver uptake, compared to sound dentin (b and d). The one-step self-etching adhesive groups (a and b) showed higher silver uptake, compared to the two-step self-etching adhesive group (c and d). The hand pointers indicate silver penetration on the top of the excavated dentinal surface and within the intertubular dentin in the residual CAD following NaOCl-based (Carisolv) chemomechanical caries removal. Ad; adhesive, HL: hybrid layer, and D: Ad; adhesive, HL: hybrid layer, and D: dentin.
*
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and the scrubbing motion used in self-etching adhesive application may explain the nonsignificant difference in bond strength between the rotary and chemomechanical caries excavated groups observed in the current study. In addition to micromechanical bonding, both self-etching adhesives contained the functional monomer group MDP (10-methacryloyloxdecyl dihydrogen phosphate), which can chemically bind to hydroxyapatite.15 This chemical bonding mechanism may increase the bond strength of residual caries-affected dentin in all groups. The results of this study are in agreement with the majority of previous studies, which reported that the caries excavation method did not affect bond strength to residual caries-affected dentin.3,5,25,43 The results revealed that sound dentin showed higher bond strength than caries-affected dentin in all but the Carisolv and Papacárie one-step self-etching adhesive groups. This marked reduction in bond strength of cariesaffected dentin could be attributed to the nature of this type of dentin, which is partially demineralized, porous, and wetter, and also contains more exposed collagen fibrils than sound dentin.1,37,50 Moreover, the partial demineralization in caries-affected dentin may reduce the MDP-dentin bonding sites and affect chemical bonding of self-etching adhesives to caries-affected dentin vs sound dentin.7,46 The higher percentage of beams with pre-test failures in the one-step self-etching adhesive groups than the two-step self-etching adhesive groups confirmed the bond strength results. From a practical and clinical viewpoint, it is very difficult to limit the application of the chemomechanical gel to the carious lesion without spreading to the surrounding sound dentin. The gel may also flow to the surrounding sound dentin during the rinsing process. This may reduce Vol 16, No 6, 2014
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the bond strength of sound dentin in the one-step selfetching adhesive chemomechanical caries removal group, particularly Carisolv. This reduction can be explained in light of recent studies by Kunawarote et al24 and Prasansuttiporn et al,36 who reported that dentin bond strength of one-step self-etching adhesives was adversely affected by NaOCl products. The NaOCl products increased the “intrinsic” pH of dentin, even after rinsing-off with water, which buffered the acidity of the ultra-mild one-step selfetching adhesives (Clearfil S3, pH ca 2.7); however, this dentinal buffering action had a minimal effect on the mild two-step self-etching adhesives with a relatively lower pH (Clearfil SE, pH ca 2). The single adhesive layer of one-step self-etching adhesive might also be easily contaminated by the NaOCl byproducts (oxides), thereby affecting the free-radical polymerization reaction. In contrast, the two-step self-etching adhesive is covered with an additional resin layer, which may reduce the adverse effects of NaOCl by-products. The results of the current study are in agreement with De Oliveira et al,13 who reported that mild self-etching adhesives (pH > 2) could easily be buffered by the high alkalinity of Carisolv-pretreated dentin (pH ca 11). Conversely, the nanoleakage results also revealed that silver penetration was higher in Carisolv/Clearfil S3/sound dentin and Carisolv/ Clearfil S3/caries-affected dentin groups, compared to Carisolv/Clearfil SE/sound dentin and Carisolv/ Clearfil SE/ caries-affected dentin groups, respectively (Fig 3). This higher silver uptake may be attributed to the contamination of the hybrid layer by NaOCl by-products (oxide), which act as voids within the hybrid layer. The free-radical polymerization reaction of the resin adhesive may also be affected, leading to areas of uncured resin, allowing silver ions to diffuse through. 513
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c
d
The results of this study showed that the two-step selfetching adhesive (Clearfil SE) had higher bond strength than the one-step self-etching adhesive (Clearfil S3). Therefore, the second hypothesis that there is no difference in bond strength between the two-step self-etching (Clearfil SE Bond) and one-step self-etching adhesives (Clearfil S3 Bond) has to be rejected. The observed lower bond strength of one-step self-etching adhesive could be due to the difference of pH between Clearfil S3 bond, which is less acidic than that of Clearfil SE primer (2.8 vs 2, respectively).35 The pH of Clearfil SE is sufficient to produce a superficial demineralization and form a uniform/shallow hybrid layer. In contrast, the demineralization caused by Clearfil S3 was not sufficient to achieve proper resin hybridization.35 The relatively higher percentages of pre-test failures in the one-step self-etching adhesive groups, compared to the two-step self-etching adhesive groups confirmed the results of the bond test. In view of the lower bond strength from Clearfil S3, use of Clearfil SE Bond is preferred clinically. The bond strength results were also supported by nanoleakage observations, which revealed higher silver impregnation in the one-step self-etching adhesive groups. This may be attributed to the ethanol solvent of Clearfil S3, which left entrapped air molecules during the volatilization process.39 Tay et al45 reported that it was more difficult to remove the last trace of water from demineralized dentin bonded with ethanol-solvated adhesive, due to the great affinity of ethanol with hydrogen ions of the residual water molecules. Furthermore, Clearfil S3 forms a relatively thin semi-permeable hybrid layer, which may lead to distinct silver penetration, in comparison to Clearfil SE.49 Tay et al44 attributed this semi-permeability of the hybrid layer to the non-uniform chain mobility structure of the hydrophilic monomers. 514
Fig 4 Back-scattered SEM images showing the nanoleakage patterns of rotary groups. a: Rotary/Clearfil S3/cariesaffected dentin. b: Rotary/Clearfil S 3/ sound dentin. c: Rotary/Clearfil SE/ caries-affected dentin. d: Rotary/Clearfil SE/sound dentin. The residual cariesaffected dentin (a and c) showed higher silver uptake, compared to sound dentin (b and d). The one-step self-etching adhesive groups (a and b) showed higher silver uptake compared to the two-step self-etching adhesive group (c and d). Silver penetration was observed on the top of the excavated dentinal surface in the residual CAD following rotary caries removal (hand pointers). Ad: adhesive; HL: hybrid layer; D: dentin.
The analysis of the fractured beams showed a high percentage of cohesive failure within the residual dentin of the chemomechanical caries removal groups. This could be due to the high degree of porosity, loss of crystallinity and partial degradation of collagen fibers, leaving a weak dentin structure (Fig 1).50 Furthermore, Pugach et al37 reported that caries-affected dentin showed marked alteration of collagen cross linking and reduction of the nanomechanical properties. The high percentage of cohesive dentin failure may also be attributed to the increased wetness of the caries-affected dentin,25 affecting the bonding efficacy of hydrophobic resin (bis-GMA).38 This was also supported by the nanoleakage results, which revealed higher silver penetration in caries-affected dentin, following both chemomechanical caries removal, when compared to the corresponding sound dentin (Figs 2 and 3). Nayif et al28 reported that despite the miscibility of hydrophilic monomers with the wetter, caries-affected dentin, the remaining hydrophobic resin monomers (bis-GMA) may not mixed well with this excessive amount of water. The partial demineralization of caries-affected dentin probably allows deeper infiltration of resin monomer, which resulted in the formation of a thicker hybrid layer, compared to sound dentin.10,29 However, the results of the current study did not find any correlation between the thickness of hybrid layer and the bond strength, which is in agreement with Yoshiyama et al.48 The increased silver uptake by the bonded interface of the residual dentin, compared to sound dentin, may be due to the porous nature of the caries-affected dentin, which entrapped water along the bonded interface, allowing silver ions to diffuse along the interface.21 Furthermore, this type of dentin also contains microcracks, which can act as pathways for the silver ions.10 Neither the excavation methods nor The Journal of Adhesive Dentistry
Hamama et al
the resin adhesive systems could eliminate nanoleakage, even the control sound dentin groups exhibited some silver penetration. The qualitative nanoleakage observations showed that, for the same adhesive group, there was not much variation in the amount of silver precipitation among the caries-affected dentin, following the three caries excavation methods. Silver penetration was observed within the intertubular dentin of the caries-affected dentin, following enzymebased chemomechanical caries removal, which may be due to the residual water in the interfibrillar spaces of the caries-affected dentin (Figs 2a and 2c). These residual water clusters may affect the free-radical polymerization reaction and/or form HEMA hydrogel within the hybrid layer.41 In contrast, the caries-affected dentin following Carisolv chemomechanical caries excavation exhibited high silver penetration within the intertubular dentin and on the top of the excavated dentinal surface, which may be due to its high water content,21 partial coverage with smear layer19 and entrapment of oxides molecules within the formed hybrid layer (Figs 3a and 3c).24,36 The cariesaffected dentin following rotary caries excavation showed high silver penetration on the top of the excavated dentinal surface, which may be attributed to the presence of the smear layer (Figs 4a and 4c).19 The high silver uptake of the smear layer is most likely due to the internal porosities within the smear layer, which allow silver ions to diffuse through it.45
8.
CONCLUSION
20.
Under the conditions of the present study, the chemomechanical caries excavation methods did not affect bonding of self-etching adhesives to caries-affected dentin, in comparison to rotary caries excavation method. It was concluded that the two-step MDP-containing self-etching adhesive system achieves better bonding of self-etching adhesives to caries-affected dentin than the one-step MDP-containing self-etching adhesive.
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Adebayo OA, Burrow MF, Tyas MJ, Palamara J. Effect of tooth surface preparation on the bonding of self-etching primer adhesives. Oper Dent 2012;37:137-149. Aggarwal V, Singla M, Yadav S, Yadav H. The effect of caries excavation methods on the bond strength of etch-and-rinse and self-etch adhesives to caries affected dentine. Aust Dent J 2013;58:454-460. Banerjee A, Kellow S, Mannocci F, Cook RJ, Watson TF. An in vitro evaluation of microtensile bond strengths of two adhesive bonding agents to residual dentine after caries removal using three excavation techniques. J Dent 2010;38:480-489. Bertassoni LE, Marshall GW. Papain-gel degrades intact nonmineralized type I collagen fibrils. Scanning 2009;31:253-258. Botelho Amaral FL, Martao Florio F, Bovi Ambrosano GM, Basting RT. Morphology and microtensile bond strength of adhesive systems to in situ-formed caries-affected dentin after the use of a papain-based chemomechanical gel method. Am J Dent 2011;24:13-19. Burke FM, Lynch E. Chemomechanical caries removal. J Ir Dent Assoc 1995;41:10-14. Burrow MF, Bokas J, Tanumiharja M, Tyas MJ. Microtensile bond strengths to caries-affected dentine treated with Carisolv. Aust Dent J 2003;48:110-114.
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Bussadori SK, Castro LC, Galvao AC. Papain gel: a new chemo-mechanical caries removal agent. J Clin Pediatr Dent 2005;30:115-119. Cardoso MV, de Almeida Neves A, Mine A, Coutinho E, Van Landuyt K, De Munck J, Van Meerbeek B. Current aspects on bonding effectiveness and stability in adhesive dentistry. Aust Dent J 2011;56(suppl 1): 31-44. Ceballos L, Camejo DG, Fuentes VM, Osorio R, Toledano M, Carvalho RM, Pashley DH. Microtensile bond strength of total-etch and self-etching adhesives to caries-affected dentine. J Dent 2003;31: 469-477. Chiaraputt S, Roongrujimek P, Sattabanasuk V, Panich N, Harnirattisai C, Senawongse P. Biodegradation of all-in-one self-etch adhesive systems at the resin-dentin interface. Dent Mater J 2011;30:814-826. Correa FN, Rodrigues Filho LE, Rodrigues CR. Evaluation of residual dentin after conventional and chemomechanical caries removal using SEM. J Clin Pediatr Dent 2008;32:115-120. De Oliveira MT, Di Francescantonio M, Consani S, Giannini M, Ambrosano GMB. Influence of dentin smear layer created by chemo-mechanical or bur excavation methods on adhesion of self-etching primers and a conventional adhesive. J Adhesion 2007;83:821-835. Ericson D, Kidd E, McComb D, Mjör I, Noack MJ. Minimally Invasive Dentistry--concepts and techniques in cariology. Oral Health Prev Dent 2003;1:59-72. Fukegawa D, Hayakawa S, Yoshida Y, Suzuki K, Osaka A, Van Meerbeek B. Chemical interaction of phosphoric acid ester with hydroxyapatite. J Dent Res 2006;85:941-944. Fure S, Lingstrom P. Evaluation of the chemomechanical removal of dentine caries in vivo with a new modified Carisolv gel. Clin Oral Investig 2004;8:139-144. Gianini RJ, do Amaral FL, Florio FM, Basting RT. Microtensile bond strength of etch-and-rinse and self-etch adhesive systems to demineralized dentin after the use of a papain-based chemomechanical method. Am J Dent 2010;23:23-28. Goldman M, Kronman JH. A preliminary report on a chemomechanical means of removing caries. J Am Dent Assoc 1976;93:1149-1153. Hamama HH, Yiu CK, Burrow MF, King NM. Chemical, morphological and microhardness changes of dentine after chemomechanical caries removal. Aust Dent J 2013;58:283-292. Hamama HH, Yiu CKY, Burrow MF. A new method of evaluation of fracture patterns following microtensile bond strength testing using polarized light microscopy. J Adhes Dent 2014;16:307-311. Huang X, Li L, Huang C, Du X. Effect of ethanol-wet bonding with hydrophobic adhesive on caries-affected dentine. Eur J Oral Sci 2011;119: 310-315. Kinney JH, Balooch M, Haupt DL, Jr., Marshall SJ, Marshall GW, Jr. Mineral distribution and dimensional changes in human dentin during demineralization. J Dent Res 1995;74:1179-1184. Kubo S, Li H, Burrow MF, Tyas MJ. Nanoleakage of dentin adhesive systems bonded to Carisolv-treated dentin. Oper Dent 2002;27:387-395. Kunawarote S, Nakajima M, Foxton RM, Tagami J. Effect of pretreatment with mildly acidic hypochlorous acid on adhesion to cariesaffected dentin using a self-etch adhesive. Eur J Oral Sci 2011;119: 86-92. Li H, Wang WM, Yu SL, Wen Q. Morphological and microtensile bond strength evaluation of three adhesive systems to caries-affected human dentine with chemomechanical caries removal. J Dent 2011;39: 332-339. Lohbauer U, Nikolaenko SA, Petschelt A, Frankenberger R. Resin tags do not contribute to dentin adhesion in self-etching adhesives. J Adhes Dent 2008;10:97-103. Maragakis GM, Hahn P, Hellwig E. Chemomechanical caries removal: a comprehensive review of the literature. Int Dent J 2001;51:291-299. Nakajima M, Sano H, Burrow MF, Tagami J, Yoshiyama M, Ebisu S, Ciucchi B, Russell CM, Pashley DH. Tensile bond strength and SEM evaluation of caries-affected dentin using dentin adhesives. J Dent Res 1995;74:1679-1688. Nakajima M, Sano H, Urabe I, Tagami J, Pashley DH. Bond strengths of single-bottle dentin adhesives to caries-affected dentin. Oper Dent 2000;25:2-10. Nayif MM, Shimada Y, Ichinose S, Tagami J. Nanoleakage of current self-etch adhesives bonded to artificial carious dentin. Am J Dent 2010;23:279-284. Neves Ade A, Coutinho E, Cardoso MV, de Munck J, Van Meerbeek B. Micro-tensile bond strength and interfacial characterization of an adhesive bonded to dentin prepared by contemporary caries-excavation techniques. Dent Mater 2011;27:552-562.
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Hamama et al 32. Neves Ade A, Coutinho E, De Munck J, Van Meerbeek B. Caries-removal effectiveness and minimal-invasiveness potential of caries-excavation techniques: a micro-CT investigation. J Dent 2011;39:154-162. 33. Oliveira SS, Pugach MK, Hilton JF, Watanabe LG, Marshall SJ, Marshall GW, Jr. The influence of the dentin smear layer on adhesion: a self-etching primer vs. a total-etch system. Dent Mater 2003;19: 758-767. 34. Omar H, El-Badrawy W, El-Mowafy O, Atta O, Saleem B. Microtensile bond strength of resin composite bonded to caries-affected dentin with three adhesives. Oper Dent 2007;32:24-30. 35. Perdigao J. New developments in dental adhesion. Dent Clin North Am 2007;51:333-357, viii. 36. Prasansuttiporn T, Nakajima M, Foxton RM, Tagami J. Scrubbing effect of self-etching adhesives on bond strength to NaOCl-treated dentin. J Adhes Dent 2012;14:121-127. 37. Pugach MK, Strother J, Darling CL, Fried D, Gansky SA, Marshall SJ, Marshall GW. Dentin caries zones: mineral, structure, and properties. J Dent Res 2009;88:71-76. 38. Sadek FT, Pashley DH, Ferrari M, Tay FR. Tubular occlusion optimizes bonding of hydrophobic resins to dentin. J Dent Res 2007;86:524-528. 39. Salz U, Mucke A, Zimmermann J, Tay FR, Pashley DH. pKa value and buffering capacity of acidic monomers commonly used in self-etching primers. J Adhes Dent 2006;8:143-150. 40. Scholtanus JD, Purwanta K, Dogan N, Kleverlaan CJ, Feilzer AJ. Microtensile bond strength of three simplified adhesive systems to cariesaffected dentin. J Adhes Dent 2010;12:273-278. 41. Shinoda Y, Nakajima M, Hosaka K, Otsuki M, Foxton RM, Tagami J. Effect of smear layer characteristics on dentin bonding durability of HEMAfree and HEMA-containing one-step self-etch adhesives. Dent Mater J 2011;30:501-510. 42. Sirin Karaarslan E, Yildiz E, Cebe MA, Yegin Z, Özturk B. Evaluation of micro-tensile bond strength of caries-affected human dentine after three different caries removal techniques. J Dent 2012;40:793-801. 43. Sonoda H, Banerjee A, Sherriff M, Tagami J, Watson TF. An in vitro investigation of microtensile bond strengths of two dentine adhesives to caries-affected dentine. J Dent 2005;33:335-342.
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44. Tay FR, Pashley DH. Aggressiveness of contemporary self-etching systems. I: Depth of penetration beyond dentin smear layers. Dent Mater 2001;17:296-308. 45. Tay FR, Pashley DH, Yoshiyama M. Two modes of nanoleakage expression in single-step adhesives. J Dent Res 2002;81:472-476. 46. Van Landuyt KL, Kanumilli P, De Munck J, Peumans M, Lambrechts P, Van Meerbeek B. Bond strength of a mild self-etch adhesive with and without prior acid-etching. J Dent 2006;34:77-85. 47. Van Meerbeek B, De Munck J, Yoshida Y, Inoue S, Vargas M, Vijay P, Van Landuyt K, Lambrechts P, Vanherle G. Buonocore memorial lecture. Adhesion to enamel and dentin: current status and future challenges. Oper Dent 2003;28:215-235. 48. Yoshiyama M, Tay FR, Doi J, Nishitani Y, Yamada T, Itou K, Carvalho RM, Nakajima M, Pashley DH. Bonding of self-etch and total-etch adhesives to carious dentin. J Dent Res 2002;81:556-560. 49. Yuan Y, Shimada Y, Ichinose S, Sadr A, Tagami J. Effects of dentin characteristics on interfacial nanoleakage. J Dent Res 2007;86:1001-1006. 50. Zanchi CH, Lund RG, Perrone LR, Ribeiro GA, del Pino FA, Pinto MB, Demarco FF. Microtensile bond strength of two-step etch-and-rinse adhesive systems on sound and artificial caries-affected dentin. Am J Dent 2010;23:152-156.
Clinical relevance: Chemomechanical caries removal did not affect the bonding of self-etching adhesives to caries-affected dentin. Papacárie can be used as an alternative chemomechanical caries removal agent to Carisolv.
The Journal of Adhesive Dentistry
Purpose: To evaluate the effect of enzyme-based (Papacárie) and sodium-hypochlorite–based (Carisolv) chemomechanical caries removal methods on bonding of self-etching adhesives to caries-affected dentin, in comparison to the standard rotary-instrument caries removal method. Materials and Methods: Seventy-eight carious permanent molars exhibiting frank cavitation into dentin were used. Forty-eight teeth were randomly divided into three groups, according to the caries excavation methods: (i) Papacárie, (ii) Carisolv and (iii) a round steel bur. After caries removal, each group was subdivided into two groups for two-step (Clearfil SE Bond) or one-step (Clearfil S3 Bond) self-etching adhesive application and resin composite buildups. Bonded specimens were sectioned into beams for microtensile bond strength testing. Bond strength data were analyzed using three-way ANOVA and Tukey’s test. For interfacial nanoleakage evaluation using a field-emission scanning electron microscope, caries was similarly removed from the remaining thirty carious molars, bonding was performed as for bond strength testing, and the teeth were sectioned. Results: Results of three-way ANOVA revealed that bond strength was significantly affected by “adhesive” (p < 0.001) and “dentin” (p < 0.001), but not “caries excavation methods” (p > 0.05). The bond strength of the two-step self-etching adhesive was significantly higher than that of the one-step self-etching adhesive (p < 0.001). Conversely, the bond strength of self-etching adhesives to sound dentin was significantly higher than to residual caries-affected dentin (p < 0.001). Greater silver penetration was observed in the bonded interfaces of residual caries-affected dentin and in interfaces bonded with the one-step self-etching adhesive vs those bonded with the two-step self-etching adhesive. Conclusion: Chemomechanical caries removal did not affect the bonding of self-etching adhesives to cariesaffected dentin as compared to caries excavation with rotary instruments. Keywords: chemomechanical caries removal, Carisolv, MDP, Papacárie, self-etching adhesives, caries-affected dentin. J Adhes Dent 2014; 16: 507–516. doi: 10.3290/j.jad.a33250
M
inimally invasive caries excavation techniques, including laser ablation, air abrasion, sonic abrasion, and chemomechanical caries removal, are characterized
a
Clinical Assistant Professor, Department of Operative Dentistry, Esthetic and Restorative Dentistry, Faculty of Dentistry, Mansoura University, Egypt. Ideas, hypothesis and experimental design, performed experiment in partial fulfillment of PhD degree, wrote manuscript.
b
Clinical Professor, Faculty of Dentistry, The University of Hong Kong, Prince Philip Dental Hospital, Hong Kong SAR, China. Idea, hypothesis and experimental design, proofread manuscript.
c
Professor and Chair, Biomaterials Department, Melbourne Dental School, The University of Melbourne,Victoria, Australia. Proofread manuscript.
Correspondence: Professor Cynthia Yiu, Paediatric Dentistry and Orthodontics, Faculty of Dentistry, The University of Hong Kong, Prince Philip Dental Hospital, 34 Hospital Road, Sai Ying Pun, Hong Kong SAR, China. Tel: +8522859-0251, Fax: +852-2559-3803. e-mail: [email protected]
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Submitted for publication: 03.05.14; accepted for publication: 09.10.14
by a common feature of selective removal of cariesinfected tissue, leaving the caries-affected tissues intact.14 The chemomechanical caries removal method was introduced by Goldman18 in 1967, with the first chemomechanical caries removal agent being GK-101,18 which was developed to GK101E (Caridex National Patent Dental Products; New Brunswick, NJ, USA) in 1984. The clinical application of Caridex in caries removal was limited due to the complexity of its delivery equipment and the relatively long time needed for caries excavation compared to conventional methods.6 To overcome these problems, Carisolv (Medi Team Dentalutveckling / Rubicon Life Science; Göteborg, Sweden) was introduced in 1998 and further developed in 2004.16 Carisolv is a sodium-hypochlorite-based (NaOCl) agent, which chlorinates and disrupts hydrogen bonds of the partially degraded collagen in carious dentin, facilitating its removal.27 507
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In 2005, a new chemomechanical caries removal agent ‘Papacárie’ (Formula and Acao; São Paulo, Brazil), which is a papain enzyme-based agent, was introduced by Bussadori et al.8 Papain is a proteolytic enzyme with bactericidal and anti-inflammatory actions. The precise mechanism of action of papain is unknown. It has been reported that infected carious tissues have lost the antiprotease α-1-anti-trypsin, which inhibits protein digestion in sound collagen-based tissues.8 In 2009, Bertassoni et al4 reported that papain enzyme could partially degrade intact nonmineralized type I collagen fibrils from rat tail tendon. Furthermore, papain has been shown to reduce the mechanical properties of intact mineralized dentin as a result of degradation of matrix proteoglycans, suggesting that the action of papain might be nonspecific.4 Self-etching adhesives are classified into two groups according to the number of clinical steps needed for bonding, ie, one- or two-step self-etching adhesives. In onestep self-etching adhesive systems, the acidic primer and the bonding agent are supplied in one bottle, whereas in two-step self-etching adhesive systems, the acidic primer and the bonding agent are supplied and applied separately in two steps. The self-etching adhesives are now classified into four types according to their acidity: ultramild (pH > 2.5), mild (pH 2–2.5), intermediate (pH 1–2), and strong (pH < 1).47 Bonding to caries-affected dentin is one of the most controversial topics in adhesive dentistry. Caries-affected dentin is characterized by a marked reduction in mineral content, loss of crystallinity, and alteration of the organic matrix.50 This complex substrate poses a great challenge to achieving reliable bonding. Previous studies have reported higher bond strengths of etch-and-rinse adhesives than self-etching adhesives to caries-affected dentin.10,40 Conversely, other studies have claimed that acid etching might lead to further loss of inorganic content of the caries-affected dentin.28,48 This may result in poor hybridization of the demineralized caries-affected dentin by adhesive resin.8 In contrast, Omar et al34 considered self-etching adhesives the “preferred adhesive” for bonding to the wetter, caries-affected dentin, due to its hydrophilic nature. Selecting the appropriate selfetching adhesive system compatible with caries-affected dentin raised another area of controversy, particularly with the development of one- and two-step self-etching adhesives. Several studies have evaluated the nanoleakage patterns22 and microtensile bond strengths5,7,17,25 of self-etching adhesives bonded to residual dentin following chemomechanical caries removal. Previous studies have mainly compared the effect of sodium-hypochlorite– based (NaOCl) chemomechanical caries removal agents, mainly Carisolv, on bonding to residual caries-affected dentin with the conventional rotary caries removal method. These studies reported that application of Carisolv did not compromise bonding to caries-excavated dentin.3,7,25,40,43 However, very few studies have examined the effect of enzyme-based chemomechanical caries removal agents (Papacárie or the experimental Biosolv [SFC-V and SFC508
VIII, 3M-ESPE; Seefeld, Germany] gels) on bonding to caries-affected dentin.3,31,32 Botelho Amaral et al5 evaluated the effect of bonding etch-and-rinse and self-etching adhesives to in-situ–formed, caries-affected dentin after the use of enzyme-based chemomechanical caries removal. The results of the study showed that bonding of the tested adhesives to caries-affected dentin was not affected by the application of Papacárie gel. Gianini et al17 examined the bond strength of etch-and-rinse and self-etching adhesives to demineralized dentin after the use of a papain-based chemomechanical caries removal method and showed that the papain-based caries removal method did not interfere with the bonding of the adhesives to demineralized dentin. However, the in-situ–formed demineralized dentin differed from the natural carious lesions due to the lack of vital odontoblasts. Hence, the aim of our current study was to compare the effect of enzyme-based (Papacárie) and sodium-hypochlorite–based (Carisolv) chemomechanical caries removal methods on bonding of self-etching adhesives to caries-affected dentin from extracted carious molars with the rotary caries removal method. The null hypotheses tested were that (i) chemomechanical caries removal has no effect on bonding of self-etching adhesives to caries-affected dentin and (ii) there is no difference in bond strength between the two-step self-etching (Clearfil SE Bond, Kuraray; Tokyo, Japan) and one-step self-etching (Clearfil S3 Bond, Kuraray) adhesives.
MATERIALS AND METHODS Seventy-eight permanent molars exhibiting frank moderate cavitation (severe decay, stage 5, ICDAS [International Caries Detection and Assessment System]) on the occlusal surface extending to the dentin were used. The molars were stored in 0.5% chloramine-T solution at 4°C for a period of less than one month following extraction. Forty-eight carious molars were used for microtensile bond strength testing, while the remaining thirty were used for nanoleakage evaluation. The study was approved by the local Institutional Review Board (IRB ref no: UW 11-355). Tooth Preparation Using a slow-speed diamond impreganated disk (Isomet, Buehler; Lake Bluff, IL, USA), the occlusal enamel was removed perpendicular to the long axis of the tooth to expose a flat dentin surface. The teeth were then randomly divided into three groups (n = 16) according to the caries excavation method: y Group 1: The carious cavity was treated with Papacárie (batch No 8996, Formula and Acao). The gel was left for 30 s prior to excavating the dentin using a #4 Carisolv noncutting instrument (Medi Team Dentalutveckling / Rubicon Life Science). Once the gel became cloudy, it was rinsed off with distilled water for 20 s and the process was repeated until successive applications of the gel failed to become cloudy. The Journal of Adhesive Dentistry
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Table 1
Adhesive systems used in the study
Brand name
Type
Composition
pH
Solvent
Manufacturer
Batch #
Clearfil SE
Two-step self-etching
Primer: 10-MDP, HEMA, hydrophilic dimethacrylate di-camphorquinone, N-N diethanol P-toluidine, water Bond: 10-MDP, bis-GMA, HEMA, hydrophobic dimethacrylate, di-camphorquinone, N-N diethanol P-toluidine, silanated colloidal silica
approx. 2*
Water
Kuraray Noritake Dental; Tokyo, Japan
11563
Clearfil S3
One-step self-etching
10-MDP, bis-GMA, HEMA, di-camphorquinone, ethanol, water, silanated colloidal silica
approx. 2.7**
Water, ethanol
Kuraray Noritake Dental
11113
Abbreviations:10- MDP,10-methacryloxydecyl dihydrogen phosphate; HEMA, 2-hydroxyethyl methacrylate; bis-GMA, bis-phenol diglycidyl methacrylate. *Chiaraputt et al.11 ** Botelho Amaral et al5 and Shinoda et al.41
y Group 2: The carious lesion was treated with Carisolv multimix gel (batch No. 10489, Medi Team Dentalutveckling / Rubicon Life Science), supplied as a twin syringe two-gel system. The gel was mixed automatically (Static Mixer, Medi Team Dentalutveckling / Rubicon Life Science) in the syringe tip prior to application on the carious lesion. The gel was left for 30 s prior to excavating the dentin using the #4 Carisolv noncutting instrument. Similar to Papácarie, once the gel became cloudy, it was rinsed away and the process was repeated until successive applications of the gel failed to become cloudy. y Group 3: Caries removal was performed with a lowspeed handpiece (NSK; Tochigi, Japan) at 35,000 rpm using a round steel bur (#012, Midwest, Dentsply; Surrey, UK) after application of a caries detector dye (batch No. 758AA, Kuraray) to define the carious lesion and verify the removal of “caries-infected” dentin, leaving lightly stained pink “caries-affected” dentin as per the manufacturer’s instructions. Bonding Procedures After caries removal, each group was subdivided into two groups (n = 8) according to the self-etching adhesive used. A two-step self-etching adhesive, Clearfil SE Bond (Kuraray) was used for the first group, and a one-step self-etching adhesive, Clearfil S3 (Kuraray), was used for the second group (Table 1). The adhesives were applied following the manufacturer’s instructions, followed by light curing of the adhesive for 10 s using a quartz-halogen light-curing unit (Elipar 2500, 3M ESPE; St Paul, MN, USA) with a light intensity output of 670 mW/cm2. The bonded surface was covered with a resin composite (Filtek Z250, 3M ESPE; St Paul, MN, USA). The resin composite was applied in 2-mm-thick increments and light cured for 20 s at the same light intensity. The teeth were stored in distilled water at 37°C for 24 h. Microtensile Bond Strength Testing All the bonded teeth were cut perpendicularly and vertically in the “x” and “y” directions through the bonded interface to obtain beams (0.9 mm x 0.9 mm wide; 5 to 7 mm long), each consisting of composite, adhesive, Vol 16, No 6, 2014
and dentin. At least sixteen bonded beams from each tooth were selected; eight of the bonded beams were from the flattest central region of the caries-affected dentin and the remaining from the sound dentin area of the same tooth to act as the control beams. The cross-sectional area of each specimen was measured with a pair of digital calipers (Model CD-6BS, Mitutoyo; Tokyo, Japan). The beams were attached to a Bencor Multi-T device (Danville Engineering; San Ramon, CA, USA) using a cyanoacrylate resin (Zapit, Dental Ventures of America; Corona, CA, USA) and subjected to tensile stress in a universal testing machine (Model 4444, Instron; Norwood, MA, USA) at a crosshead speed 1 mm/ min. The experimental unit of the current study was the tooth; this means that the recorded value of each group represented the mean microtensile bond strength value of the eight teeth. The number of beams which failed prior to testing was recorded for each group, and they were replaced by intact bonded beams for bond strength testing. The beams with pre-test failures were excluded from the statistical analysis. Analysis of Failure Modes All fractured beams were observed under polarized light microscopy (Nikon polarizing Microscope Eclipse LV100POL, Nikon; Tokyo, Japan) at 100X magnification to determine the mode of failure: cohesive in dentin (CD), adhesive (AD), mixed (MI), or cohesive in composite (CC).20 Representative specimens from each group were examined by field-emission scanning electron microscope (FE-SEM) (Hitachi S-3400N, Hitachi High Technologies America; Schaumburg, IL, USA) to confirm the failure mode as observed by polarized light microscopy. Nanoleakage Evaluation The remaining 30 carious molars were randomly allocated, and caries removal and bonding were performed following as for microtensile bond strength testing, except that the subgroup sample size was 5. Each bonded tooth was sectioned into halves through the cavity along the mesio-distal plane using a low-speed water-cooled diamond saw (Isomet, Buehler). The bonded specimen was coated with two layers of nail varnish within 1 mm of the bonded interface. The specimens were rehydrated 509
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Table 2 Percentages of beams with pre-test failures and microtensile bond strength of one-step and two-step self-etching adhesives to sound and residual dentin following caries excavation with Papacárie, Carisolv, or rotary instrument Caries Removal Methods Papacárie Sound dentin
Carisolv
Residual cariesexcavated dentin
Sound dentin
Rotary
Residual cariesexcavated dentin
Sound dentin
Residual cariesexcavated dentin
Beams with pre-test failures/ (percentage) Clearfil SE Bond
4 / (6.3%)
6 / (9.1%)
4 / (6.3%)
7 / (10.4%)
3 / (4.8%)
7 / (10.4%)
Clearfil S3 Bond
5 / (7.7%)
11/ (15.5%)
6 / (8.8%)
12 / (16%)
6 / (8.8%)
10 / (14.3%)
Microtensile Bond Strength (MPa) * Clearfil SE Bond
38.01a ± 2.4
31.5b ± 2.8
43.0a ±
3.4
30.8b ± 2.7
40.4a ± 4.2
26.1b ± 3.7
Clearfil S3 Bond
26.5b,c ± 1.6
23.2c ± 1.9
23.9c ± 2.1
20.1c ± 2.2
29.2b ± 4.7
17.4c ± 1.0
*Values are means ± standard deviation, n = 8 teeth/group. Multiple comparison test, all groups compared to all other groups; groups identified by different superscript letters (both in rows and columns) are significantly different at p < 0.05.
in distilled water for 10 min, immersed in 50% (W/V) ammoniacal silver nitrate solution and kept in total darkness for 24 h, and then rinsed with distilled water for 5 min. Specimens were subsequently immersed in photodeveloping solution (Kodak GBX fixer and replenishers, Kodak; Rochester, NY, USA) and exposed to fluorescent light for 8 h in order to reduce the diamine silver ions into metallic silver grains within voids along the bonded interface. Specimens were further rinsed with distilled water for 5 min. The observed surface was wet polished with 600-, 800-, 1200-, 2400-, and 4000- grit silicon carbide papers (Microcut, Buehler), followed by polishing with a polishing cloth using 6-, 3-, 1-μm diamond pastes (Diamat, Pace Technologies; Tucson, AZ, USA). Specimens were then ultrasonically cleaned, air dried for 24 h, mounted on aluminum stubs, and carbon coated. The resin/dentin interface was analyzed using FE-SEM operated in backscattered electron mode at 10 mm working distance and 12 KV accelerating voltage (ACCV). Statistical Analysis The bond strength data were analyzed using a statistical software package (SPSS, V.19, IBM; Armonk, NY, USA). Since the bond strength data were normally distributed (Kolmogorov-Smirnov test) and homoscedastic (Levine test), they were analyzed using three-way ANOVA to examine the effect of three variables (caries excavation method, adhesive, and dentin) and the interaction of the three factors on the microtensile bond strength. Tukey’s post-hoc multiple comparison test was performed to compare the means between the groups, with statistical significance set at α = 0.05. A Kruskal-Wallis test with Dunnett’s post-hoc test was used to compare the incidence of different failure modes among the different groups. A significance level of 5% was employed for all analyses. 510
RESULTS Microtensile Bond Strength Test The mean microtensile bond strengths of two-step and one-step self-etching adhesives to residual dentin following caries excavation with Papacárie, Carisolv, or rotary instruments are shown in Table 2. Results of the three-way ANOVA revealed that the microtensile bond strengths were significantly affected by the factors “adhesive” (p < 0.001) and “dentin” (p < 0.001), but not the factor “caries excavation method” (p > 0.05). The interaction of all three factors, “caries excavation method”, “adhesive”, and “dentin” was significant (p < 0.05). In general, the bond strengths of the two-step selfetching adhesive were significantly higher than those of the one-step self-etching adhesive (p < 0.001). The bond strength of self-etching adhesives to sound dentin was higher than that to residual caries-excavated dentin (p < 0.001). The method of caries removal had no effect on the bond strengths of two-step and one-step self-etching adhesives to residual dentin following caries excavation (p > 0.05). The percentage of beams with pre-test failures was higher in the residual caries-affected dentin in comparison to sound dentin areas. Moreover, it was found that the percentage of the beams with pre-test failures was higher in the one-step self-etching adhesive group than in the corresponding two-step self-etching adhesive group (Table 2). As shown in Table 3, statistical analysis (KruskalWallis test with Dunnett’s post-hoc test) revealed an increase in the percentage of cohesive failures in dentin when bonded to residual caries-excavated dentin vs sound dentin in Papacarie/S3 group, Rotary/SE, and Rotary/S3 groups. The mixed failure mode was commonly observed in the sound dentin groups. The percentage of cohesive failure in composite failure was very low. RepreThe Journal of Adhesive Dentistry
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Table 3 Failure mode analysis (Kruskal-Wallis with Dunnett’s post-hoc test) Group
Fracture mode
(%)
Statistical difference
P SE-SD
CD AD MI CC
9.38 31.25 56.25 3.13
A, B
P SE-CAD
CD AD MI CC
7.81 26.56 64.06 1.56
A, B
P S3-SD
CD AD MI CC
9.38 43.75 46.94 0.00
B
P S3-CAD
CD AD MI CC
28.13 32.81 39.06 0.00
C
C SE-SD
CD AD MI CC
6.25 43.75 46.88 3.13
B
C SE-CAD
CD AD MI CC
26.56 32.81 31.25 9.38
B, C
C S3-SD
CD AD MI CC
12.50 60.94 26.56 0.00
C
C S3-CAD
CD AD MI CC
37.50 34.38 28.13 0.00
C
R SE-SD
CD AD MI CC
4.51 17.01 75.35 3.13
A
R SE-CAD
CD AD MI CC
16.84 18.58 64.58 0.00
B
R S3-SD
CD AD MI CC
1.39 19.17 73.37 6.08
A
R S3-CAD
CD AD MI CC
19.97 26.22 53.82 0.00
B
CAD
b
CAD
CD: cohesive failure in dentin; AD: adhesive failure; MI: mixed failure; CC: cohesive failure in composite. P: Papacárie: C: Carisolv; R: rotary: S3: Clearfil S3 bond: SE: Clearfil SE bond; SC: sound dentin; CAD: cariesaffected dentin. Groups identified by different letters (left column) are significantly different at p < 0.05.
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a
Fig 1 SEM micrographs showing cohesive failure in residual caries-affected dentin following (a) NaOCl-based (Carisolv) and (b) enzyme-based (Papacárie) chemomechanical caries removal. More destruction of dentin surface was observed in Carisolv (arrow) than in Papacárie groups. CAD: caries-affected dentin.
sentative SEM micrographs from the chemomechanical caries removal groups with cohesive failure in dentin are shown in Fig 1. Nanoleakage Analysis Representative FE-SEM micrographs showing nanoleakage patterns of the bonded interfaces from the Papacárie, Carisolv, and rotary groups are shown in Figs 2 to 4. In general, the hybrid layers in the residual cariesaffected dentin groups were thicker than those in the sound dentin groups. Silver nitrate uptake was higher in the bonded interfaces of the residual caries-affected dentin groups than in the sound dentin groups. Similarly, the silver uptake at the bonded interfaces formed by the one-step self-etching adhesive was higher than those formed by the two-step self-etching adhesive. In the Papacárie caries-excavated groups, silver penetration was observed within the intertubular dentin; while 511
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Ad Ad HL
*
HL
D
D
a
b
Ad
Ad
HL HL
* c
D D
d
in the rotary caries-excavated groups (Rotary SE and Rotary S3), silver penetration was observed on the top of the excavated dentinal surface. However, both nanoleakage patterns were observed in the Carisolv caries-excavated groups (Carisolv/SE and Carisolv/S3).
DISCUSSION Previous studies which evaluated the microtensile bond strength of residual caries-affected dentin following enzyme-based chemomechanical caries removal used demineralized dentin to avoid the effect of caries lesion size, shape, depth, and surface irregularities on the results of the bond strength test.5,17 However, results obtained from demineralized dentin should be interpreted with caution, because natural carious dentin contains a wide variety of pathogenic materials, organic acids, hydrolytic enzymes, and whitlockite occluding the dentinal tubules.30 Kinney et al22 reported that artificial loss of mineral content led to collapse of the collagen fibers, which might affect resin hybridization into dentin and bond strength results. The microtensile bond strength test used in the current study considered the tooth as the test unit and calculated the mean bond strength value using the beams from each tooth. This reduced bond strength variation and permitted testing a broad area of carious lesion. The flattest portion of the residual caries-affected dentin was selected to reduce the microshear forces on the adhesive interface during microtensile bond strength testing.40 Furthermore, using the sound dentin from the same tooth as control decreases the variation of results in comparison with using another sound tooth as the control group. 512
Fig 2 Back-scattered SEM images showing the nanoleakage patterns of the Papacárie group. a: Papacárie/Clearfil S3/caries-affected dentin. b: Papacárie/ Clearfil S3/sound dentin. c: Papacárie/ Clearfil SE/caries-affected dentin. d: Papacárie/ Clearfil SE/sound dentin. The residual caries-affected dentin (CAD) (a and c) showed higher silver uptake than did sound dentin (b and d). The one-step self-etching adhesive groups (a and b) showed higher silver uptake than did the two-step self-etching adhesive groups (c and d). Silver penetration was observed within the intertubular dentin in the residual CAD following enzyme-based (Papacárie) chemomechanical caries removal (hand pointers). Ad: adhesive; HL: hybrid layer; D: dentin.
The present results showed that there was no significant difference in bond strength among the three caries excavation methods. The null hypothesis that chemomechanical caries removal has no effect on bonding of selfetching adhesives to caries-affected dentin could not be rejected. Previous studies which evaluated the morphological changes of dentin following chemomechanical caries removal reported that the excavated surface following Papacárie excavation was rough and characterized by a total absence of smear layer, with mostly patent dentinal tubules.19 In contrast, the excavated surface following Carisolv was irregular and partially covered with a smear layer with most of the dentinal tubules being patent or partially occluded.19,25 These morphological characteristics may increase the dentinal surface area for micromechanical retention of adhesive resin following caries removal. Conversely, many studies reported that the excavated surface following caries removal using burs was relatively smooth, and covered with a smear layer with occlusion of tubules.15,19,44 Some studies reported that the smear layer might act as a natural barrier preventing resin infiltration into dentin.12,25 However, Lohbauer et al26 reported that hybridization of self-etching adhesive resin with dentin mainly depended on bonding of hydrophilic monomers to the exposed collagen scaffold, and that the resin tags did not influence bond strength. Furthermore, Oliveira et al33 found that the smear layer formed following cutting with carbide burs was thin, with moderate surface roughness, and that the bond strength to dentin surfaces prepared by carbide bur was higher than to surfaces prepared by diamond bur and silicon paper. Abdebayo et al1 mentioned that application of adhesive resin with a scrubbing motion enhanced resin penetration into dentin. Therefore, the thin smear layer following caries excavation The Journal of Adhesive Dentistry
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Ad Ad
Fig 3 Back-scattered SEM images showing the nanoleakage patterns of Carisolv groups. a: Carisolv/Clearfil S3/caries-affected dentin. b: Carisolv/ Clearfil S 3/sound dentin. c: Carisolv/ Clearfil SE/caries-affected dentin. d: Carisolv/ Clearfil SE/sound dentin. The residual caries-affected dentin (a and c) showed higher silver uptake, compared to sound dentin (b and d). The one-step self-etching adhesive groups (a and b) showed higher silver uptake, compared to the two-step self-etching adhesive group (c and d). The hand pointers indicate silver penetration on the top of the excavated dentinal surface and within the intertubular dentin in the residual CAD following NaOCl-based (Carisolv) chemomechanical caries removal. Ad; adhesive, HL: hybrid layer, and D: Ad; adhesive, HL: hybrid layer, and D: dentin.
*
HL HL
* a
D
b
Ad
Ad
*
HL
HL
D
D
* c
and the scrubbing motion used in self-etching adhesive application may explain the nonsignificant difference in bond strength between the rotary and chemomechanical caries excavated groups observed in the current study. In addition to micromechanical bonding, both self-etching adhesives contained the functional monomer group MDP (10-methacryloyloxdecyl dihydrogen phosphate), which can chemically bind to hydroxyapatite.15 This chemical bonding mechanism may increase the bond strength of residual caries-affected dentin in all groups. The results of this study are in agreement with the majority of previous studies, which reported that the caries excavation method did not affect bond strength to residual caries-affected dentin.3,5,25,43 The results revealed that sound dentin showed higher bond strength than caries-affected dentin in all but the Carisolv and Papacárie one-step self-etching adhesive groups. This marked reduction in bond strength of cariesaffected dentin could be attributed to the nature of this type of dentin, which is partially demineralized, porous, and wetter, and also contains more exposed collagen fibrils than sound dentin.1,37,50 Moreover, the partial demineralization in caries-affected dentin may reduce the MDP-dentin bonding sites and affect chemical bonding of self-etching adhesives to caries-affected dentin vs sound dentin.7,46 The higher percentage of beams with pre-test failures in the one-step self-etching adhesive groups than the two-step self-etching adhesive groups confirmed the bond strength results. From a practical and clinical viewpoint, it is very difficult to limit the application of the chemomechanical gel to the carious lesion without spreading to the surrounding sound dentin. The gel may also flow to the surrounding sound dentin during the rinsing process. This may reduce Vol 16, No 6, 2014
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d
the bond strength of sound dentin in the one-step selfetching adhesive chemomechanical caries removal group, particularly Carisolv. This reduction can be explained in light of recent studies by Kunawarote et al24 and Prasansuttiporn et al,36 who reported that dentin bond strength of one-step self-etching adhesives was adversely affected by NaOCl products. The NaOCl products increased the “intrinsic” pH of dentin, even after rinsing-off with water, which buffered the acidity of the ultra-mild one-step selfetching adhesives (Clearfil S3, pH ca 2.7); however, this dentinal buffering action had a minimal effect on the mild two-step self-etching adhesives with a relatively lower pH (Clearfil SE, pH ca 2). The single adhesive layer of one-step self-etching adhesive might also be easily contaminated by the NaOCl byproducts (oxides), thereby affecting the free-radical polymerization reaction. In contrast, the two-step self-etching adhesive is covered with an additional resin layer, which may reduce the adverse effects of NaOCl by-products. The results of the current study are in agreement with De Oliveira et al,13 who reported that mild self-etching adhesives (pH > 2) could easily be buffered by the high alkalinity of Carisolv-pretreated dentin (pH ca 11). Conversely, the nanoleakage results also revealed that silver penetration was higher in Carisolv/Clearfil S3/sound dentin and Carisolv/ Clearfil S3/caries-affected dentin groups, compared to Carisolv/Clearfil SE/sound dentin and Carisolv/ Clearfil SE/ caries-affected dentin groups, respectively (Fig 3). This higher silver uptake may be attributed to the contamination of the hybrid layer by NaOCl by-products (oxide), which act as voids within the hybrid layer. The free-radical polymerization reaction of the resin adhesive may also be affected, leading to areas of uncured resin, allowing silver ions to diffuse through. 513
Hamama et al Ad Ad
*
HL HL D D
a
b
Ad
Ad
*
HL HL D
D
c
d
The results of this study showed that the two-step selfetching adhesive (Clearfil SE) had higher bond strength than the one-step self-etching adhesive (Clearfil S3). Therefore, the second hypothesis that there is no difference in bond strength between the two-step self-etching (Clearfil SE Bond) and one-step self-etching adhesives (Clearfil S3 Bond) has to be rejected. The observed lower bond strength of one-step self-etching adhesive could be due to the difference of pH between Clearfil S3 bond, which is less acidic than that of Clearfil SE primer (2.8 vs 2, respectively).35 The pH of Clearfil SE is sufficient to produce a superficial demineralization and form a uniform/shallow hybrid layer. In contrast, the demineralization caused by Clearfil S3 was not sufficient to achieve proper resin hybridization.35 The relatively higher percentages of pre-test failures in the one-step self-etching adhesive groups, compared to the two-step self-etching adhesive groups confirmed the results of the bond test. In view of the lower bond strength from Clearfil S3, use of Clearfil SE Bond is preferred clinically. The bond strength results were also supported by nanoleakage observations, which revealed higher silver impregnation in the one-step self-etching adhesive groups. This may be attributed to the ethanol solvent of Clearfil S3, which left entrapped air molecules during the volatilization process.39 Tay et al45 reported that it was more difficult to remove the last trace of water from demineralized dentin bonded with ethanol-solvated adhesive, due to the great affinity of ethanol with hydrogen ions of the residual water molecules. Furthermore, Clearfil S3 forms a relatively thin semi-permeable hybrid layer, which may lead to distinct silver penetration, in comparison to Clearfil SE.49 Tay et al44 attributed this semi-permeability of the hybrid layer to the non-uniform chain mobility structure of the hydrophilic monomers. 514
Fig 4 Back-scattered SEM images showing the nanoleakage patterns of rotary groups. a: Rotary/Clearfil S3/cariesaffected dentin. b: Rotary/Clearfil S 3/ sound dentin. c: Rotary/Clearfil SE/ caries-affected dentin. d: Rotary/Clearfil SE/sound dentin. The residual cariesaffected dentin (a and c) showed higher silver uptake, compared to sound dentin (b and d). The one-step self-etching adhesive groups (a and b) showed higher silver uptake compared to the two-step self-etching adhesive group (c and d). Silver penetration was observed on the top of the excavated dentinal surface in the residual CAD following rotary caries removal (hand pointers). Ad: adhesive; HL: hybrid layer; D: dentin.
The analysis of the fractured beams showed a high percentage of cohesive failure within the residual dentin of the chemomechanical caries removal groups. This could be due to the high degree of porosity, loss of crystallinity and partial degradation of collagen fibers, leaving a weak dentin structure (Fig 1).50 Furthermore, Pugach et al37 reported that caries-affected dentin showed marked alteration of collagen cross linking and reduction of the nanomechanical properties. The high percentage of cohesive dentin failure may also be attributed to the increased wetness of the caries-affected dentin,25 affecting the bonding efficacy of hydrophobic resin (bis-GMA).38 This was also supported by the nanoleakage results, which revealed higher silver penetration in caries-affected dentin, following both chemomechanical caries removal, when compared to the corresponding sound dentin (Figs 2 and 3). Nayif et al28 reported that despite the miscibility of hydrophilic monomers with the wetter, caries-affected dentin, the remaining hydrophobic resin monomers (bis-GMA) may not mixed well with this excessive amount of water. The partial demineralization of caries-affected dentin probably allows deeper infiltration of resin monomer, which resulted in the formation of a thicker hybrid layer, compared to sound dentin.10,29 However, the results of the current study did not find any correlation between the thickness of hybrid layer and the bond strength, which is in agreement with Yoshiyama et al.48 The increased silver uptake by the bonded interface of the residual dentin, compared to sound dentin, may be due to the porous nature of the caries-affected dentin, which entrapped water along the bonded interface, allowing silver ions to diffuse along the interface.21 Furthermore, this type of dentin also contains microcracks, which can act as pathways for the silver ions.10 Neither the excavation methods nor The Journal of Adhesive Dentistry
Hamama et al
the resin adhesive systems could eliminate nanoleakage, even the control sound dentin groups exhibited some silver penetration. The qualitative nanoleakage observations showed that, for the same adhesive group, there was not much variation in the amount of silver precipitation among the caries-affected dentin, following the three caries excavation methods. Silver penetration was observed within the intertubular dentin of the caries-affected dentin, following enzymebased chemomechanical caries removal, which may be due to the residual water in the interfibrillar spaces of the caries-affected dentin (Figs 2a and 2c). These residual water clusters may affect the free-radical polymerization reaction and/or form HEMA hydrogel within the hybrid layer.41 In contrast, the caries-affected dentin following Carisolv chemomechanical caries excavation exhibited high silver penetration within the intertubular dentin and on the top of the excavated dentinal surface, which may be due to its high water content,21 partial coverage with smear layer19 and entrapment of oxides molecules within the formed hybrid layer (Figs 3a and 3c).24,36 The cariesaffected dentin following rotary caries excavation showed high silver penetration on the top of the excavated dentinal surface, which may be attributed to the presence of the smear layer (Figs 4a and 4c).19 The high silver uptake of the smear layer is most likely due to the internal porosities within the smear layer, which allow silver ions to diffuse through it.45
8.
CONCLUSION
20.
Under the conditions of the present study, the chemomechanical caries excavation methods did not affect bonding of self-etching adhesives to caries-affected dentin, in comparison to rotary caries excavation method. It was concluded that the two-step MDP-containing self-etching adhesive system achieves better bonding of self-etching adhesives to caries-affected dentin than the one-step MDP-containing self-etching adhesive.
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Clinical relevance: Chemomechanical caries removal did not affect the bonding of self-etching adhesives to caries-affected dentin. Papacárie can be used as an alternative chemomechanical caries removal agent to Carisolv.
The Journal of Adhesive Dentistry
