Effect of Artificial Aging Regimens on the Performance of

Self-Etching adhesives
Mustafa Ülker,1 Mutlu Özcan,2 Abdulkadir S
! engün,3 Füsun Özer,4 Sema Belli5
1
Department of Conservative Dentistry, Faculty of Dentistry, Erciyes University, Kayseri, Turkey

2
Center for Dental and Oral Medicine, Clinic for Fixed and Removable Prosthodontics and Dental Materials Science,
University of Zürich, Head of Dental Materials Unit, Zürich, Switzerland
3
Department of Conservative Dentistry, Faculty of Dentistry, Selçuk University, Konya, Turkey
4
Department of Preventive and Restorative Sciences, University of Pennsylvania School of Dental Medicine, Philadelphia,
Pennsylvania
5
Department of Endodontics, Faculty of Dentistry, Selçuk University, Konya, Turkey

Received 7 January 2009; revised 4 October 2009; accepted 17 October 2009

Published online 20 January 2010 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jbm.b.31572

Abstract: This in vitro study was performed to compare the microtensile bond strengths
(MTBS) of current self-etching adhesives to dentin and to evaluate the effects of artificial aging
[(thermocycling (TC) and/or mechanic loading (ML)] on MTBS and on nanoleakage of self-
etching adhesives. Two-step (AdheSE Bond, Clearfil Protect Bond, Clearfil SE Bond,
Optibond Self-Etch) and one-step (Hybrid Bond, G-bond, Clearfil Tri-S Bond, and Adper
Prompt L-Pop) self-etching adhesives were tested. Resin composite build-ups were created,
and the specimens were subjected to 104 TC, 105 ML, and 104/105 TC/ML. Non-aged
specimens served as controls. In the control group, no significant differences were found
among the MTBS of the one-step self-etching adhesives and among those of three two-step self-
etching adhesives (AdheSE Bond, Clearfil Protect Bond, and Clearfil SE Bond) (p > 0.05). The
MTBS of AdheSE Bond and Clearfil Protect Bond were higher than were those of all one-step
self-etching adhesives and than those of Optibond Self-Etch. The MTBS of Clearfil SE Bond
was higher than were those of two one-step self-etching adhesives (Adper Prompt L-Pop, G-
bond) (p < 0.05). Compared with the non-aged controls, TC did not decrease (p > 0.05), but
ML and TC/ML significantly decreased the MTBS of the adhesives tested (p < 0.05). Two-step
self-etching adhesives tended to fail more cohesively in dentin. Transmission electron
microscopy revealed different nanoleakage patterns in the adhesive and hybrid layers of all
adhesives examined, and signs of additional silver-filled water channels were more readily
detectable after TC/ML. ' 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 93B:
175–184, 2010

Keywords: adhesion; degredation; composite/hard tissue; fatigue; microscopy

INTRODUCTION introduced for use in dentistry.2 One-step self-etching adhe-

In dental procedures involving bonding, separate etching
and water-rinsing phases can be omitted with use of acidic
monomer-containing self-etching adhesive systems, making
the application of adhesives less technique-sensitive for
clinicians.1 Therefore, a number of self-etching adhesives
involving either one- or two-step application have been
Correspondence to: M. Ülker (e-mail: mulker@selcuk.edu.tr)
Contract grant sponsor: Scientific Research Project Centre of Selçuk University
(BAP)
' 2010 Wiley Periodicals, Inc.
sives are also known as ‘‘all-in-one adhesives,’’ and reduce
the time required for bonding procedures.
Two-step self-etching adhesives create only a submicron
hybrid layer.1–3 However, they usually have bond strengths
equivalent to those obtained with total-etch adhesives.1,2,4,5
In addition to micromechanical interlocking through
hybridization, specific functional monomers of mild or in-
termediate two-step self-etching adhesives were shown to
interact chemically with residual hydroxyapatite crystals
that remain available in the submicron hybrid layer.6 De
Munck et al.4 and Sensi et al.7 reported that one-step self-
etching adhesives resulted in lower dentin bond strength
175
176 ÜLKER ET AL.
then did two-step self-etching adhesives. However, Abo
et al.8 reported comparable dentin bond strengths for one-
and two-step self-etching adhesives. Currently, only limited
data are available regarding the dentin bonding perform-
ance of several one or two-step self-etching adhesives.
In mixing self-etching adhesives, water is essential as an
ionization medium to enable self-etching activity. Even after
the completion of the ionization reaction, residual water may
remain in the hybrid or adhesive layer. One-step self-etching
adhesives are highly hydrophilic and act as semipermeable
membranes, allowing the passage of fluids.9 Nanoleakage at
the resin/dentin interface with some self-etching adhesives has
been attributed to incomplete water removal.10 As deminerali-
zation and resin infiltration occur simultaneously, self-etching
adhesives are less likely to show a discrepancy between the
depth of demineralization and the depth of resin infiltration.11
Although manufacturers claim optimized compositions, little
information is available regarding nanoleakage at the resin/
dentin interface of newly introduced self-etching adhesives,
especially after aging simulation that may help to estimate the
longevity of such adhesives.
Well-conducted randomized controlled clinical trials are
considered the standard for the evaluation of adhesive sys-
tems.12 However, considerable time and resources are
needed for such trials, and confounding factors make it dif-
ficult to discriminate among commercial adhesives. On the
other hand, adhesive systems change rapidly. Therefore, an
easy and rapid means is necessary to estimate the clinical
outcome of adhesives in a realistic manner. By simulating
the conditions of the oral cavity, such as the thermal
changes and chewing forces, in laboratory settings, the nat-
ural aging process can be mimicked to a certain extent. In
this context, the use of mechanical loading and/or thermo-
cycling (TC) in vitro has been considered a useful method
for simulating in vivo challenges.13-17
The objectives of this study were as follows: (1) to
compare the microtensile bond strengths (MTBS) between
one- and two-step self-etching adhesives to dentin based
on the hypothesis that dentin bond strengths of one- and
two-step self-etching adhesives may be similar; (2) to
investigate the effects of artificial aging (TC and/or me-
chanical loading [ML]) on the MTBS of self-etching adhe-
sives based on the hypothesis that artificial aging may
decrease the dentin bond strength; and (3) to evaluate the
effects of aging with TC/ML on nanoleakage of self-etch-
ing adhesives.
MATERIALS AND METHODS
Specimen Preparation
A total of 160 human third molars were used for the
MTBS tests. After removing the soft tissue remnants, the
teeth were stored in 0.01% thymol aqueous solution at 48C
and were used within 3 months following extraction. The
occlusal third of each crown was removed perpendicular to
the long axis with a low-speed diamond saw with water
cooling. Flat dentin surfaces were prepared and polished
with 400-grit silicon carbide paper to create bonding surfa-
ces to be covered with smear layers,18 and the bonding pro-
cedures were then carried out.
Four two-step and four one-step self-etching adhesives
were used in this study. The composition and manufac-
turers of the materials used are listed in Table I. Adhesive
resins were applied according to each manufacturer’s
instructions and then built up with a hybrid resin composite
(Clearfil AP-X; Kuraray, Japan) in two layers to a height
of 4 mm. Adhesives and resin composites were polymer-
ized with a LED polymerization unit (Elipar LED Freelight
II; 3M ESPE AG, Seefeld, Germany) at a constant distance
of 2 mm from the surface. Adhesives were polymerized
according to each manufacturer’s instructions, and then
each resin composite layer was polymerized for 40 s. The
light intensity was 1007 mW/cm2, verified by a radiometer
after every 10 specimens (Model 100; Kerr, Orange
County, CA). After polymerization, the specimens were
stored in distilled water at 378C for 24 h.
Twenty specimens were prepared for each adhesive and
then randomly distributed into four test groups (n 5 5).
Group 1: Control group (no thermocycling or mechanical
loading); Group 2: Thermocycling group (TC) [104 thermo-
cycling (Nova, Konya, Turkey) was performed at 5–558C
(628C); dwell time: 30 s]; Group 3: Mechanical loading
group (ML) [105 mechanical loading was performed (50 N;
0.5 Hz)]; Group 4: Thermomechanical loading group (TC/
ML) [104 thermocycling and 105 mechanical loading]. Me-
chanical loading of specimens was performed after thermo-
cycling.
A chewing simulator (Vega Chewing Simulator; Nova,
Konya, Turkey) was designed and custom-made for me-
chanical loading where the specimens were fixed. The cen-
ter of the composite resin was occluded against a stainless
steel metal jig with a rounded end that acted as the antago-
nist (diameter: 5 mm). Mechanical load was achieved with
consistent loading of 50 N at a frequency of 0.5 Hz in dis-
tilled water. The total time spent for aging a tooth–compos-
ite specimen with TC was 2 weeks, aging with ML was
performed for 2 days, and aging with TC/ML was per-
formed for 16 days.
After the artificial aging procedures, each tooth was sec-
tioned with a slow-speed saw (Isomet; Buehler Ltd., Lake
Bluff, IL) under water cooling into multiple 1 3 1 mm
beams using the ‘‘non-trimming’’ version of the microten-
sile test. When the external beams from the periphery were
excluded, five beams were obtained from the central part of
each tooth–composite specimen. Twenty-five beams were
obtained for each test condition by adhesive (5 3 5) group.
Twenty of these 25 beams were randomly selected for
MTBS tests (n 5 20). The remaining five beams were used
for TEM analysis. Beams from the peripheral dentin were
not included in the study, and pre-test failures were not
included in the statistical analysis.
Journal of Biomedical Materials Research Part B: Applied Biomaterials
 EFFECT OF AGING ON SELF-ETCHING ADHESIVES 177

TABLE I. Brand Names, Composition, Manufacturers and Batch Numbers of Self-Etching Adhesives

Brand Name Composition Manufacturer Batch No.

One-step Self-etching Adhesives
Clearfil Tri-S Bond MDP, Bis-GMA, HEMA, Hydrophobic dimetacrylate, Kuraray, Kurashiki, 41111
Camphorquinone, Ethyl alcohol, Silanated colloidal silica Japan
Hybrid Bond Liquid: 4-META, Polyfunctional acrylate, Monomethacrylate, Sun Medical, Shiga, KL 1
%40 Acetone, Water, Camphoroquinone, Stabilizers Microbrush Japan
activators: Sodium p-toluensulphate, Aromatic amine
G-Bond 4-META, Phosphoric ester monomer, UDMA, Silica filler, GC, Tokyo, Japan 0408241
Photo initiator, Acetone, Water
Adper Prompt L-pop Liquid 1: Methacrylated phosphoric esters, Bis-GMA, 3M ESPE AG, Seefeld, 206168
Camphorquinone, Stabilizers Germany
Liquid 2: Water, HEMA, Polyalkenoic acid, Stabilizers
Two-step Self-etching Adhesives
AdheSE Bond Dimethacrylate, Phosphonic acid acrylate, Stabilizers, Initiators, Ivoclar Vivadent, Schaan, Test Pack
Water Liechtenstein
HEMA, Dimethacrylate, Silicone dioxide, Stabilizers, Initiators
Clearfil Protect Bond MDPB, MDP, HEMA, Hydrophilic dimethacrylate, Water, Kuraray, Kurashiki, Japan 41114
Initiators
MDP, Bis-GMA, HEMA, Hydrophobic dimethacrylate,
Camphorquinone N,N- Diethanol p-toluidine, Silanated
colloidal silica Sodium fluoride
Clearfil SE Bond MDP, HEMA, Hydrophilic dimethacrylate, Camphorquinone, Kuraray, Kurashiki, Japan 413277
N,N- Diethanol p-toluidine, Water
MDP, Bis-GMA, HEMA, Hydrophobic dimethacrylate,
Camphorquinone N,N- Diethanol p-toluidine, Silanated
colloidal silica
Optibond Self-etch Alkyl dimethacrylate resins, Barium aluminoborosilicate glass, SDS KERR, California 41410833
Silicon dioxide, Sodium hexa-fluorosilicate, Ethyl alcohol
Alkyl dimethacrylate resins, Ethyl alcohol, Water, stabilizers and
activators

Beams were fixed to a universal testing machine (Micro-
tensile Tester; Bisco, Schaumburg, IL) using cyanoacrylate
glue (Model Repair II Pink; Dentsply-Sankin, Otawara, Ja-
pan) and stressed at a cross-head speed of 1 mm/min until
failure. The cross-sectional area at the site of failure was
measured to the nearest 0.01 mm with a digital caliper
(Model CD-6BS; Mitutoyo, Tokyo, Japan), and the micro-
tensile bond strength was calculated from this measurement
and expressed in MPa. Failure modes were evaluated at
340 magnification with a stereoscopic microscope and
classified as follows: a, adhesive failure between dentin and
resin; c, cohesive failure in the resin; d, cohesive failure in
dentin; m, mixed failure (including adhesive failure
between dentin and resin and cohesive failure of resin or
dentin).4,19
Transmission Electron Microscopy
For transmission electron microscopy (TEM) observation,
one beam was randomly selected among the remaining five
beams obtained from the central parts of the teeth from
Group 1 and Group 4 for each self-etching adhesive. These
were varnish coated, except for 1 mm along the bonded
interfaces. To avoid desiccation of the beams, varnish coat-
ing was finished as quickly as possible. The beams were
then immersed in a tracer solution consisting of 50 wt %
ammoniac silver nitrate for 24 h according to the nanoleak-
age evaluation protocol described by Tay and coworkers.20
The silver-impregnated beams were rinsed with distilled
water and placed in a photodeveloping solution for 8 h
under a fluorescent light to reduce the diamine silver ion
complexes into metallic silver grains. The beams were then
dehydrated in an ascending ethanol series (50–100%) and
embedded in epoxy resin (Serva, Heidelberg, Germany)
according to the TEM processing protocol described by
Tay and coworkers.21 After the beams were embedded in
epoxy resin, sections 90–100 nm in thickness were pre-
pared with an ultramicrotome (Ultracut S; Leica, Vienna,
Austria) and a diamond knife (Diatome, Bienne, Switzer-
land) and examined without additional staining using TEM
(Philips EM208S; Philips, Eindhoven, The Netherlands)
operating at 80 kV. TEM observations were used as illus-
trative to support the investigation.
Statistical Analysis
Mean MTBS of each group were analyzed using two-way
ANOVA to evaluate the effects of the two experimental
factors, that is, the type of adhesive and the aging regimen
(TC, ML, and TC/ML) (Statistix 8.0 for Windows; Analyti-

Journal of Biomedical Materials Research Part B: Applied Biomaterials

178 ÜLKER ET AL.

TABLE II. Mean (6Standard Deviation—SD) of Microtensile Bond Strength Data (MPa). Different Small Superscript Letters Indicate
Significant Differences in the Column and Capital Superscript Letters in the Row

Control MTBS Thermocycling Mechanical Loading Thermomechanical

(6SD) MTBS (6SD) MTBS (6SD) Loading MTBS (6SD)
Two-step Self-etch
AdheSE Bond 42.32 6 8.52Aa 39.57 6 7.43ABa 34.14 6 9.76BCa 26.64 6 11.25Ca
Clearfil Protect Bond 38.18 6 5.75Aa 34.40 6 4.95Aa 27.53 6 5.04Ba 22.94 6 4.72Ca
Clearfil SE Bond 35.76 6 9.28Aab 36.88 6 9.07Aa 25.66 6 5.42Bab 25.88 6 5.07Ba
Optibond Self-etch 28.83 6 6.04Abc 25.63 6 6.64Ab 18.91 6 6.07Bcd 15.70 6 2.78Cc
One-step Self-etch
Clearfil Tri-S Bond 27.24 6 7.79Abc 24.38 6 7.31ABb 20.93 6 6.20Bbc 20.60 6 5.81Bab
Hybrid Bond 26.40 6 9.57Abc 23.16 6 5.69ABb 18.79 6 6.30Bcd 19.69 6 6.27Bab
G-Bond 22.85 6 4.81Ac 20.29 6 6.35ABbc 15.58 6 3.68Bcd 16.35 6 4.40Bbc
Adper Prompt L-Pop 21.68 6 9.49Ac 15.05 6 5.66ABc 13.26 6 4.41Bd 13.09 6 5.99Bc

cal Software, Inc, Tallahassee, FL). Post hoc comparisons MTBS of self-etching adhesives, with the exception of
were performed using Tamhane’s T2 test. Statistical signifi- Clearfil Protect Bond and Optibond Self-Etch (p \ 0.05).
cance was set in advance at p 5 0.05.

RESULTS
Microtensile Bond Strength Test
Some pre-testing failures were recorded for the one-step
self-etching adhesives after ML and TC/ML (1/2/2/5 after
ML and 2/2/3/6 after TC/ML for Clearfil Tri-S Bond,
Hybrid Bond, G-Bond, and Adper Prompt L-Pop, respec-
tively). All pre-testing failures occurred during specimen
processing.
MTBS results were significantly affected by both the ad-
hesive type (p \ 0.05) and the aging conditions (p \
0.05). Interaction terms were also significant (p 5 0.001).
The mean MTBS results, standard deviations, and signifi-
cant differences between groups are presented in Table II.
In the control group, no significant differences were
observed among the MTBS results of three of the two-step
self-etching adhesives (AdheSE Bond, Clearfil Protect
Bond, and Clearfil SE Bond) (p [ 0.05). The mean MTBS
of one-step self-etching adhesives also showed no signifi-
cant differences (p [ 0.05). AdheSE Bond and Clearfil
Protect Bond showed higher bond strengths than did all
one-step self-etching adhesives. These two-step self-etching
adhesives also showed higher bond strength than did Opti-
bond Self-Etch adhesive. Although the bond strength of
Clearfil SE Bond was higher than those of G-bond and
Adper Prompt L-Pop (p \ 0.05), it was not significantly
different from those of other self-etching adhesives tested
(p [ 0.05). The mean bond strength of two-step self-etch-
ing Optibond Self-Etch was not significantly different from
those of one-step adhesives (p [ 0.05).
In comparison to the control group, TC did not decrease
the mean MTBS of self-etching adhesives (p [ 0.05). On
the other hand, ML and TC/ML significantly decreased the
mean MTBS of self-etching adhesives (p \ 0.05). Com-
pared with the ML, TC/ML did not further decrease the
Failure-Type Analysis
For all adhesives, the most frequent pattern of failure was
adhesive failure in both aged and non-aged groups (Table
III). In the non-aged control group, two-step self-etching
adhesives tended to show more cohesive failure in dentin.
However, this type of failure decreased gradually after
aging. After TC/ML, only Clearfil SE Bond showed cohe-
sive failure in dentin (10%). None of the one-step self-etch-
ing adhesives presented cohesive failures in dentin after
either ML or TC/ML.
TEM Analysis
TEM revealed the reticular mode of nanoleakage expres-
sion along the resin/dentin interfaces of all specimens. In
addition, a spotted mode of nanoleakage expression was
also seen consisting of isolated silver grains that were dis-
persed randomly throughout both the adhesive and the
hybrid layers [Figs. 1(b–c), 2(b–c), 3(a)].
Two-step self-etching adhesives exhibited similar nano-
leakage patterns in the control [Figs. 1(a–d)]and TC/ML
groups [Figs. 2(a–d)]. However, signs of additional silver-
filled water channels (water trees) were more detectable af-
ter TC/ML; most of these originated from the surface of
the hybrid layer, extended to the adhesive layers, and were
associated with silver-filled void-like structures [Figs. 2(b–
d)].
Reticular modes of nanoleakage expression were
detected at the resin/dentin interfaces of the one-step self-
etching adhesives. In the control groups, some of these
modes of nanoleakage expression were more bulky [Figs.
3(b–c)]. However, these were reduced in size after TC/ML,
and silver-filled water channels become more detectable
[Figs. (4a–d)].

Journal of Biomedical Materials Research Part B: Applied Biomaterials

 EFFECT OF AGING ON SELF-ETCHING ADHESIVES 179

TABLE III. Distribution of Failure Types Per Group in Percentage Based on the Light Microscope Observations; a 5 Adhesive Failure;
m 5 Mixed Failure; c 5 Cohesive Failure in the Resin; d 5 Cohesive Failure in dentin; n 5 20

Thermomechanical
Control Thermocycling Mechanical Loading Loading
a m c d a m c d a m c d a m c d
Two-step Self-etching
AdheSE Bond 50 15 5 30 60 15 10 15 50 40 0 5 85 15 0 0
Clearfil Protect Bond 65 15 0 20 65 15 5 15 65 25 0 10 80 20 0 0
Clearfil SE Bond 70 20 0 10 60 16 5 20 70 20 5 5 70 20 0 10
Optibond Self-etch 80 15 0 5 85 10 0 5 80 15 0 5 95 5 0 0
One-step Self-etching
Clearfil Tri-S Bond 95 5 0 0 100 0 0 0 95 5 0 0 80 20 0 0
Hybrid Bond 84 16 0 0 90 10 0 0 85 15 0 0 95 5 0 0
G-Bond 85 15 0 0 80 15 0 5 95 5 0 0 90 10 0 0
Adper Prompt L-Pop 95 5 0 0 90 10 0 0 90 10 0 0 95 5 0 0

DISCUSSION interface after a period of clinical use, we attempted to

Under in vivo conditions, normal daily functioning, thermal
stress, mal occlusion, habitual bruxism, and trauma impose
stresses throughout the tooth and the restorative system,
which may affect and destroy the adhesive bond. Therefore,
to predict changes in bond strength and the resin/dentin
simulate the conditions of the oral cavity (functional
stresses (fatigue) and thermal changes) in a laboratory
study.
Our in vitro study was performed to predict changes in
bond strength and quality of resin/dentin interface of self-
etching adhesives after at least 1 year of in vivo clinical

Figure 1. TEM micrographs of the resin-dentin interfaces in the control groups (a) AdheSE Bond,
(b) Clearfil Protect Bond, (c) Clearfil SE Bond, and (d) Optibond Self-Etch. Note that these two-step
self-etching adhesives created 0.5–1 lm-thick hybrid layers (H) that contained reticular mode (open
arrow head) and some additional spotted mode (arrows) of nanoleakage that were identified as sil-
ver deposits in hybrid layer. The spotted mode of nanoleakage expression consisted of isolated sil-
ver grains were dispersed also into adhesive layer as in the hybrid layer. A: Adhesive; C:
Composite; D: Dentin; H: Hybrid layer; DT: Dentin tubule.

Journal of Biomedical Materials Research Part B: Applied Biomaterials

180 ÜLKER ET AL.

Figure 2. TEM micrographs of the resin-dentin interfaces after TC/ML [104 Thermocycling and 105
Mechanical Loading] in (a) AdheSE Bond, (b) Clearfil Protect Bond, (c) Clearfil SE Bond and (d)
Optibond Self-Etch. Reticular mode (open arrow head) and some spotted mode (arrows) of nano-
leakage can be identified in the hybrid layer. In addition, water trees (pointers) most of which origi-
nated from the surface of the hybrid layer and extending to adhesive layer are detectable after TC/
ML (b-d). A: Adhesive; C: Composite; D: Dentin; H: Hybrid layer; DT: Dentin tubule.

use. Restorations were predicted to undergo 1,000,000
active stress cycles in 20 years.22 In this study, mechanical
loading of 100,000 cycles was performed with a frequency
of 0.5 Hz, which is close to the chewing cycle in vivo.23
Higher frequencies (1–60 Hz) used in previous studies24,25
may minimize laboratory working time, but they may result
in internal heating of the specimens.26 Several factors, such
as the type of tooth, age, and gender, make it difficult to
simulate in vivo chewing forces. In clinical situations, these
factors interfere with the force employed. Nevertheless,
simulation of the mean force is necessary for further com-
parison between studies. Similar to some previous studies
using chewing simulators, a force of 50 N was chosen to
simulate the average constant load during mastica-
tion.14,15,17,27
Contradictory results have been reported regarding the
influence of mechanical loading on the bond strength of
dental adhesives. The results of this study indicated that
ML can markedly affect the dentin bond strength of self-
etching adhesives. The reduced incidence of cohesive fail-
ure in dentin with two-step adhesives after ML was prob-
ably due the adverse effect of ML on dentin bond strength.
Osorio et al.28 reported reduced MTBS of a self-etching ad-
hesive applied to flat dehydrated dentin after ML (5000
cycles, 90 N). In contrast, Bedran-de Castro et al.15,16
reported that mechanical loading did not influence the
MTBS of a total-etch adhesive system at cervical margins
of Class II restorations (100,000 cycles, 50 N) or its shear
bond strength to flat dentin (50,000 cycles, 80 N). The
varying results may be due to the adhesive system used,
cavity shape, and differences in both the force and the
number of cycles applied to specimens.
As our aim was to predict changes in bond strength at
the resin/dentin interface after 1 year of in vivo clinical
use, specimens were subjected to 10,000 thermocycles,
which was reported to correspond to approximately 1 year
of in vivo functioning.29 TC allows bonded specimens to
be subjected to extreme temperatures, similar to the situa-
tion in the oral cavity. During TC, repetitive contraction–
expansion stresses are generated at the resin/dentin inter-
face due to higher contraction–expansion coefficient of the
restorative material than of the teeth, which may eventually
result in crack propagation along the resin/dentin inter-
face.30 In this study, the number of cycles applied was rela-
tively high, but the dentin bond strengths of the self-
etching adhesives were unaffected. Large composite cylin-
ders and dentin around the bonded interface showed good
resistance against thermal stresses and the hydrolyzing

Journal of Biomedical Materials Research Part B: Applied Biomaterials

 EFFECT OF AGING ON SELF-ETCHING ADHESIVES 181

Figure 3. TEM micrographs of the resin-dentin interfaces in the control groups (a) Clearfil Tri-S
Bond, (b) Hybrid Bond, (c) G-bond, and (d) Adper Prompt L-pop. One-step self-etching adhesives
created 1–1.5 lm-thick hybrid layers (H)-except Adper Prompt L-Pop (d) exhibiting 3–4 lm-thick
hybrid layer- that contained reticular mode (open arrow head) of nanoleakage expressions some of
which are more bulk in size (black arrow in b and c). Some additional spotted modes (arrows) of
nanoleakage expressions are present in adhesive layer. Note the water trees (pointer) only at the
resin-dentin interface of Clearfil Tri-S Bond (a). A: Adhesive; C: Composite; D: Dentin; H: Hybrid
layer; DT: Dentin tubule.

effect of hot water.30,31 A number of authors also reported
that TC had no effect on bond strength,15-17 but some stud-
ies indicated that TC decreases the bond strength of adhe-
sives.30,32,33 This may be related to variations in the TC
regimens used among studies. It can also be argued that
due to the poor heat conductivity of the tooth, the central
part of the adhesive was not subjected to thermal stress to
the same extent as the peripheral part was in TC. In this
study, the peripheral beams were not used. We anticipated
that the marginal beams would be affected to a greater
extent by TC, and this should be verified in future studies.
The stresses caused by ML were considered to have a
greater effect than those caused by TC alone because aging
with TC/ML did not result in further decreases in the
MTBS of most self-etching adhesives compared with aging
with ML alone. Stresses during ML were presumably accu-
mulated at the resin/dentin interface leading to the forma-
tion of micro-gaps between the adhesive and dentin
surface, or they may have caused plastic deformations in
the adhesive layer itself, ultimately decreasing the MTBS
of the self-etching adhesives. Recently, Frankenber et al.13
reported that increasing the amount of TC/ML stress leads
to a reduction in the MTBS of self-etching and total-etch
adhesives. Bedran-de-Castro et al.15 also reported that the
MTBS of a total-etch adhesive at the cervical margins of
Class II resin restorations decreased when 100,000 load cy-
cling (50 N) was applied together with 2000 thermal cy-
cling. Our results support these findings, although the
adhesives tested were not identical.
In controls and all aging groups, three of the two-step
self-etching adhesives (AdheSE Bond, Clearfil Protect
Bond, and Clearfil SE Bond) showed similar higher MTBS.
Clearfil SE Bond is a well-established and well-docu-
mented self-etching adhesive that contains 10-methacryloy-
loxydecyl dihydrogen phosphate (10-MDP) as a functional
monomer. The monomer 10-MDP can readily adhere to re-
sidual hydroxyapatite in the hybrid layer, and this bond
was reported to be very stable.6,34 In addition to the self-
etching effect on dentin, the specific functional monomer
10-MDP has additional chemical bonding efficacy. This
seems to contribute to the adhesive potential of Clearfil SE
Bond. The chemical formulation of Clearfil Protect Bond is
very similar to that of Clearfil SE Bond, with the exception
of the antibacterial monomer (MDPB) content. The self-

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182 ÜLKER ET AL.

Figure 4. TEM micrographs of the resin-dentin interfaces after TC/ML in (a) Clearfil Tri-S Bond, (b)
Hybrid Bond, (c) G-bond and (d) Adper Prompt L-pop. Apart from the presence of nanoleakage
(closed arrow) in the hybrid layer (H), note the water trees (pointer) along the surface of the hybrid
layer and within the bulk of the adhesive layer (a,b,c,d). A: Adhesive; D: Dentin; H: Hybrid layer.

etching capacity of AdheSE Bond is based on phosphonic
acid acrylates. This adhesive is more acidic than are Clear-
fil SE Bond and Clearfil Protect Bond, but it still does not
dissolve all hydroxyapatite crystals in the hybrid layer.1,2,4
A chemical interaction may occur between the functional
monomer of AdheSE Bond and residual hydroxyapatite
crystals, as observed for some functional monomers.6,34
Our results were not in agreement with those of other
groups who reported higher bond strengths with Optibond
Self-Etch.4,7,35 The poorer results and less frequent cohe-
sive failures in dentin obtained with Optibond Self-Etch
may be due to the composition of the adhesive. In contrast
to AdheSE Bond, Clearfil SE Bond, and Clearfil Protect
Bond, the bonding resin of Optibond Self-Etch is not sol-
vent-free. This may induce some technique sensitivity, as
the solvent must be properly removed by air drying. There-
fore, factors such as application conditions may influence
the bond strength results.
With one exception (Optibond Self-Etch), the mean
bond strengths were higher for two-step self-etching adhe-
sives than for one-step self-etching adhesives. Similar find-
ings were reported previously.4,36,37 The lower dentin bond
strength results with one-step self-etching adhesives have
been attributed to phase separation between hydrophilic
and hydrophobic ingredients of one-step self-etching adhe-
sives38 or their highly hydrophilic and permeable nature.39
The low bond strengths observed with one-step self-etching
adhesives may indicate that a single-component material
cannot yet fulfill all the requirements for the production of
an effective adhesive layer. Furthermore, the time allowed
for the chemical reactions appears to be markedly reduced
with these adhesives. It is very difficult to remove water
from the adhesive resin of one-step self-etching adhesives,
and even if removed, water from dentin may diffuse into
the adhesive layer again. Water in the adhesive layer may
soften the polymers and thus decrease the adhesion
capacity.37 Failure-type evaluation confirmed this finding;
cohesive failure type in dentin was not observed with any
of the one-step self-etching adhesives. This type of failure
could be interpreted as an indicator that the adhesive
strength has exceeded the cohesive strength of the dentin.
Therefore, higher MTBS values are often accompanied by
increased frequency of cohesive failures in dentin.
Adper Prompt L-Pop was introduced for dental use earlier
than the other one-step self-etching adhesives examined.
Although the differences were not statistically significant,
this adhesive showed the lowest bond strengths among all
aging and control groups. Adper Prompt L-Pop is the succes-
sor of Prompt L-Pop, a typical strong self-etching adhesive.
The main difference between this and the older version is
that the amount of non-acidic methacrylates has been
increased to enhance viscosity (according to the technical in-

Journal of Biomedical Materials Research Part B: Applied Biomaterials

 EFFECT OF AGING ON SELF-ETCHING ADHESIVES
formation obtained from 3M ESPE). This may help to pro-
vide a sufficient amount of primer on the dentin surface.
However, an in vitro study suggested that the adhesive layer
formed by Adper Prompt L-Pop may contain zones with no
adhesive layer.4 In addition, the hybrid layer formed by
Adper Prompt L-Pop was thinner on areas where the adhe-
sive layer was absent.4 The low bond strength obtained with
Adper Prompt L-Pop may be attributable to the insufficient
adhesive penetration to dentin.
The relationships between nanoleakage and TC or ML
have been examined, and previous studies indicated no
effect on nanoleakage of adhesives.40-42 Li et al.40 reported
that 5000 mechanical load cycling (90 N) did not increase
nanoleakage. Therefore, we evaluated the nanoleakage at
the resin/dentin interface after specimens were subjected to
both TC and ML to simulate a worst-case scenario. Under
our experimental conditions, self-etching adhesives were
not able to prevent nanoleakage. Water trees that were
observed very rarely in the adhesive or hybrid layers of
control groups were identified after TC/ML, confirming the
hypothesis that water trees may be a potential mechanism
for degradation of resin–dentin bonds.20 These water trees
(silver-filled void-like structures) could have been formed
by water sorption through the adhesives enhanced by me-
chanical loading and thermocycling. Not only the decreased
MTBS results but also the supportive nanoleakage patterns
observed in this study confirmed the hypothesis that aging
may act as a mechanism for degradation of resin–dentin
bonds.
Considering the MTBS results and failure modes, the
two-step self-etching adhesives showed better performance
than did the one-step self-etching adhesives. As the bond
strength and nanoleakage expression of self-etching adhe-
sives were affected by artificial aging, simulation of ther-
mal changes and chewing forces in vitro may be essential
to better evaluate and understand the performance of dental
adhesives. Based on the results of this study, from a clini-
cal viewpoint, all adhesives should be expected to deterio-
rate in terms of adhesion even after short-term use.
This study was based on the work performed by M. Ulker for
fulfillment of the degree of Philosophy, University of Selçuk, Tur-
key. The TEM analysis of this study was supported by the Uni-
versity Medical Center Groningen, University of Groningen,
Department of Dentistry and Dental Hygiene, Clinical Dental Bio-
materials, Groningen, The Netherlands.
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