R E S E A R C H
Background. The authors conducted a
The effect of saliva
contamination on
enamel and dentin
using a self-etching
adhesive
RICHARD D. TOWNSEND, D.M.D.;
WILLIAM J. DUNN, D.D.S.
lmost 50 years ago, Buonocore1,2 observed
A how phosphoric acid improved adhesion of
paint to metal surfaces in industry and
applied this concept to the dental profession.
That discovery and the development of
bisphenol glycidyldemethacrylate resin by Bowen3 in the
early 1960s led to the evolution of the first dentin adhe-
sive systems. Failure rates for the early dentin adhesive
systems were high.4-6 It was not until the early 1990s
that significant advances in dentin bonding were made
with the development of multistep
dentin adhesive systems. These fourth-
Self-etching generation adhesive systems typically
adhesives are used conditioners, primers or both to
not as pretreat dentin to make the hydrophilic
dentin substrate more receptive to
tolerant of
bonding. The final step in this compli-
contamination cated bonding scheme was the applica-
of enamel tion of a low-viscosity adhesive resin
surfaces as that copolymerized with the primed
they are of dentin surface, creating a hybrid layer
contamination that was amenable to bonding with
restorative composite material.7
of dentin
To reduce the number of steps
surfaces. involved in fourth-generation bonding,
researchers and manufacturers devel-
oped more simplified systems by combining the etchant
and primer, or the primer and adhesive. These fifth-
generation adhesive systems demonstrated similar bond
strengths to dentin in vitro8,9 and in vivo,10,11 but their
claims of significantly reduced application times may
Copyright ©2004 American Dental Association. All rights reserved.
ABSTRACT
study to evaluate the effect of
saliva contamination on A D A
J
enamel and dentin before
or after application of a
✷

✷
N
CON
self-etching adhesive.
IO
Methods. The authors
T
T
A
N
I
C
divided 120 extracted U
IN U
A G ED
3
molars into enamel and RT
I CLE
dentin groups of 60 specimens
each, and then further divided each group
into three groups of 20 specimens each.
Resin-based composite cylinders were
bonded to tooth specimens that were uncon-
taminated (control) or were contaminated
with saliva either before or after application
of the adhesive and then loaded to the
failure point. The authors calculated the
mean shear bond strength for each group
and subjected data to a one-way analysis of
variance, or ANOVA, to test for differences
among groups.
Results. Mean shear bond strength was
16.7 megapascals (± 3.0 standard deviation,
or SD) for the dentin control group, 14.8
MPa (± 2.3 SD) for dentin contaminated
with saliva before adhesive application,
15.9 MPa (± 3.8 SD) for dentin contami-
nated with saliva after adhesive applica-
tion, 19.1 MPa (± 4.8 SD) for the enamel
control group, 12.3 MPa (± 4.4 SD) for
enamel contaminated with saliva before
adhesive application, and 13.1 MPa (± 5.5
SD) for enamel contaminated with saliva
after adhesive application. One-way
ANOVA detected no difference in the dentin
groups, but the authors found a significant
difference among enamel groups. Uncon-
taminated enamel had higher bond
strengths than either of the contaminated
enamel groups.
Conclusions. Saliva contamination did
not affect the dentin shear bond strength of
the
the self-etching adhesive. Saliva did have a
detrimental effect on enamel bond strength.
Clinical Implications. Newer genera-
tions of adhesives appear to be more tol-
erant of saliva contamination, but proper
isolation still should be accomplished until
further research proves otherwise.
JADA, Vol. 135, July 2004 895
R E S E A R C H

have been premature. Self-etching primer sys- and Garcia-Godoy,20 Vargas and colleagues21 and
tems generally were regarded as the “sixth” gen- Hitmi and colleagues22 were unable to demon-
eration, but these systems still required the sepa- strate statistically significant decreases in shear
rate application of an adhesive. bond strength to tooth surfaces contaminated
To simplify things further, researchers devel- with saliva when testing modern adhesive sys-
oped all-inclusive adhesive systems. These dentin- tems that incorporated a primer in the adhesive.
bonding agents incorporate etchant, primer and el-Kalla23 also reported that salivary contamina-
adhesive into one bottle. There is some disagree- tion did not affect the formation of the hybrid
ment on the classification of these adhesive sys- layer. These in vitro results suggest that modern
tems, and it should be noted that the nomencla- adhesive systems may be more forgiving of con-
ture is dictated by the manufacturer who releases tamination on dentin and enamel than were ear-
the products to the dental market and not by sci- lier separate-bottle systems.
entific categorization. A better classification The dental profession uses all of the adhesive
system would separate adhesives into total-etch system categories. Depending on the adhesive
and self-etch categories. Total-etch systems then system used, bonding resin-based composite to
would be subdivided into separate-bottle systems, tooth structure can involve multiple steps, and the
in which the primer and adhesive are separate, operative field could become contaminated during
and one-bottle systems that combine the primer any of these steps. Some studies have reported
and adhesive. These total-etch systems require that salivary contamination could adversely affect
that both the enamel and dentin be the bond strength to dentin, but
etched with acid before placement only when contamination occurred
of the adhesive. Self-etching adhe- Depending on the after adhesive application23 or light
20
sives would be subdivided into self- adhesive system used, curing. Because self-etching adhe-
etching primers that require a sepa- sives incorporate etchant, primer
bonding resin-based
rate application of adhesive and and adhesive into one application,
all-in-one self-etching adhesives composite to tooth the time points at which salivary
that combine etchant, primer and structure can involve contamination could occur are
adhesive. multiple steps, and reduced. There are few studies eval-
With the addition of a glu- the operative field uating whether saliva contamina-
taraldehyde disinfectant, a “sev- could become tion affects the enamel and dentin
enth” generation surfaced in 2003. bond strength of self-etching adhe-
contaminated during
However, Settembrini and col- sive systems.
leagues12 demonstrated that acid any of these steps. We conducted this study to inves-
etching alone appears to suffice as tigate whether saliva contamination
a disinfectant. at various time points has an effect
Before the introduction of hydrophilic primers, on the shear bond strength of an all-in-one self-
the bonding of resin-based composite to etched etching adhesive system used on enamel and
enamel involved the application of a hydrophobic dentin.
resin, which required meticulous attention to the
operative field. Proper isolation was necessary to MATERIALS AND METHODS
prevent unwanted contamination of the tooth sur- We collected 120 recently extracted human
face that could significantly decrease the bonding molars that were free from restorations or caries,
efficacy of resin to enamel.13,14 As early as 1963, and we stored them in a solution of 0.5 percent
Buonocore2 concluded that a dry tooth surface chloramine-T and refrigerated them until we used
was essential for achieving good adhesion because them in the study. No more than six months
etched tooth surfaces readily absorb salivary con- elapsed between the extraction of the molars and
stituents, thus reducing their surface energy and their use in the study.
rendering them less favorable to bonding. This We randomly divided the teeth into two groups
finding has been supported by other for enamel and dentin specimen preparation. For
researchers.14-18 the dentin group, we ground the occlusal surfaces
The concept of salivary contamination’s of the teeth serially with 180-, 320- and 600-grit
decreasing bond strength is not universally silicon carbide paper on a polishing lathe under
accepted, however. Fritz and colleagues,19 el-Kalla running water until all islands of enamel were

896 JADA, Vol. 135, July 2004

Copyright ©2004 American Dental Association. All rights reserved.
 R E S E A R C H

just removed. We then embedded the specimens Mass.) with a crosshead speed of 0.5 mm per
in dental mounting stone inside acrylic molds to minute until we noted failure. We calculated
expose a flat dentin surface for bonding. mean shear bond strength for all groups and sub-
For the enamel group, we ground the teeth jected the data to a one-way analysis of variance,
serially with 180-, 320- and 600-grit silicon car- or ANOVA, to determine if a difference existed
bide paper until a flat enamel surface of at least among the groups. We used Tukey’s Honestly Sig-
10 square millimeters was prepared. We nificant Difference post hoc test to identify the
embedded the specimens in dental mounting differences. We considered all statistical analyses
stone inside acrylic molds to expose a flat enamel to be significant at the P < .05 level.
surface for bonding. We kept all specimens moist
before bonding. RESULTS
We further separated the dentin and enamel Results of the one-way ANOVA comparing the
groups into three additional subgroups each. The shear bond strength of resin-based composite to
experimental groups were as follows: dentin and enamel are presented in Tables 1 and
ddentin group 1: no saliva contamination 2. We found no evidence to suggest a difference
(control); among any of the three dentin groups (P = .16).
ddentin group 2: saliva contamination before Among the enamel groups, we found that the non-
application of self-etching adhesive system; contaminated enamel control group demonstrated
ddentin group 3: saliva contamination after higher bond strengths than either of the groups
application of self-etching adhesive that were contaminated with
system, but before light curing; saliva. This difference was signifi-
denamel group 1: no saliva The noncontaminated cant (P < .05).
contamination (control); enamel control group
denamel group 2: saliva contami- DISCUSSION
demonstrated higher
nation before application of self- The advantage of self-etching adhe-
etching adhesive system; bond strengths than sive systems is that all three basic
denamel group 3: saliva contami- either of the groups steps (etching, priming and
nation after application of self- that were applying adhesive) occur simulta-
etching adhesive system, but before contaminated neously. The benefits of simplicity
light curing. with saliva. and time savings are intuitive, but
The principal investigator (R.T.) an added advantage is that there
collected his fresh whole saliva. are fewer time points during place-
After contaminating each tooth specimen, he ment when contamination can occur. Self-etching
gently air-thinned the saliva with an air-water adhesive systems simultaneously demineralize
syringe, leaving the specimens visibly moist with dentin while infiltrating it with monomers to the
saliva. He did not rinse or blot dry the specimens. same depth, and they then are polymerized in
We placed the self-etching adhesive system on situ.24 Simultaneous demineralization-infiltration
the tooth specimens according to the manufac- might improve the marginal seal and adhesion to
turer’s recommendations. To bond a cylinder of dentin; when etching and priming are accom-
resin-based composite to each specimen, we plished in separate steps, some regions of de-
secured a circular, split polyethylene mold (inner mineralized dentin may not be penetrated by the
diameter: 3.0 mm; height: 3.0 mm) to the flat resin primer. Self-etching primer systems do not
tooth surface. We filled the mold with a light- require the primer to be rinsed off after applica-
curable resin-based composite in two equal tion. What happens to the acid still is not under-
increments. We polymerized each increment stood completely, but it is thought to be neutral-
separately for 40 seconds with a curing light. To ized at some point by hydroxyl ions that are
ensure a minimum intensity of at least 500 milli- released from the hydroxyapatite during deminer-
watts per square centimeter, we used a hand-held alization.24 The high buffering ability of the
curing radiometer to test the light output after dentin should limit further demineralization of
each specimen was handled. After storing the hydroxyapatite.
tooth specimens in water at 37 C for 48 hours, we The components of the adhesive system we
subjected them to shear force in a universal used in our study are listed in Table 3. Bonding
testing machine (model 1541s, Instron, Canton, agent A contains phosphoric acid and methacry-

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Copyright ©2004 American Dental Association. All rights reserved.
R E S E A R C H

TABLE 1 The results of our study

on salivary contamination—
MEAN SHEAR BOND STRENGTHS OF DENTIN GROUPS. which agree with the find-
DENTIN GROUP NO. OF TEETH MEAN SBS (MPa)* STANDARD DEVIATION ings of Hitmi and
Group 1: 20 16.7† ± 3.0
colleagues,22 Fritz and
No Saliva colleagues,19 Vargas and
Contamination
(Control)
colleagues21 and el-Kalla
and Garcia-Godoy20—
Group 2: Saliva 20 14.8† ± 2.3
Contamination
demonstrated that the
Before newer generations of adhe-
Application of
Self-etching
sive systems that use
Adhesive System hydrophilic primers may be
Group 3: Saliva 20 15.9† ± 3.8
less sensitive to salivary
Contamination contamination of prepared
After Application
of Self-etching
tooth surfaces than pre-
Adhesive System vious generations of
* SBS: Shear bond strength, MPa: megapascals.
adhesive systems. Ideal
† Groups are not statistically different from each other (P > .05). operative dentistry still
mandates the use of rubber
TABLE 2 dams, but certain instances
may preclude their use.
MEAN SHEAR BOND STRENGTHS OF ENAMEL GROUPS. Unfortunately, an
ENAMEL GROUP NO. OF TEETH MEAN SBS (MPa)* STANDARD DEVIATION increasing number of dental
Group 1: No Saliva 20 19.1 ± 4.8
adhesive manufacturers
Contamination state in their instructions
(Control)
that the operating field rou-
Group 2: Saliva tinely may be isolated with
Contamination 20 12.3† ± 4.4
Before Application
cotton rolls.19 The results of
of Self-etching our study suggest that self-
Adhesive System
etching adhesive systems
Group 3: Saliva are more tolerant of sali-
Contamination
After Application 20 13.1† ± 5.5
vary contamination on
of Self-etching dentin surfaces. This, how-
Adhesive System
ever, does not imply that
* SBS: Shear bond strength, MPa: megapascals. proper isolation is unneces-
† Groups are not statistically different from each other (P > .05).
sary, as there will be
late monomers. Bonding agent B contains more instances in which saliva has contaminated the
methacrylate monomers, water and filler parti- field despite the operator’s best efforts and obser-
cles. This particular self-etching adhesive system vations. Previous investigators have examined
incorporates a bifunctional acid monomer (MAC- the effect of saliva after it was blotted dry or
10, J. Morita USA, Irvine, Calif.) into the adhe- rinsed off.19-21,23,25 This implies that the operators
sive (Figure). This bifunctional molecule essen- were aware of saliva contamination. Because
tially is a hydrophilic acid monomer with a modern adhesive dentistry uses a wet, total-etch
methacrylate group on the hydrophobic portion of approach, salivary contamination may go unno-
the molecule and two carboxylic acid groups on ticed because the tooth surface already is wet. In
the hydrophilic portion. The carboxylic acid por- our study, we examined saliva-contaminated
tion of the molecule aids in dentin bonding. The specimens that were neither blotted dry nor
methacrylate portion of the molecule copolymer- rinsed to investigate the effects of an “unnoticed”
izes with other methacrylic resin monomers in saliva-contaminated surface.
establishing the bond between the adhesive and There are several explanations for the toler-
the resin-based composite restorative material. ance of self-etching adhesives to salivary contami-
An additional phosphoric acid monomer was nation. If contamination occurs after etching but
added to facilitate self-etching. before adhesive application, the tooth surface

898 JADA, Vol. 135, July 2004

Copyright ©2004 American Dental Association. All rights reserved.
 R E S E A R C H

already should be moist with water to TABLE 3

prevent collagen collapse in the dem-
ineralized dentin. This moist tooth sur- COMPONENTS OF THE SELF-ETCHING
face may be an important factor in pre- ADHESIVE.
venting salivary proteins from BONDING COMPONENTS PERCENTAGE
penetrating and blocking openings in AGENT*

the collagen mesh in dentin and pores A Methacrylate 100

monomers (includes
in enamel. Further, the hydrophilicity two functional acid
of the self-etching products may allow monomer groups,
MAC-10 [J. Morita
for their diffusion through the salivary USA, Irvine, Calif.],
20
film. which contain car-
boxylic acid, and a
Similarly, if the tooth surface is con- phosphoric acid
taminated with saliva after the applica- monomer)†
tion of the adhesive, but before light B Methacrylate 74
curing, there appears to be no signifi- monomers
cant effect on bond strength. There is, Water 10
however, evidence that if salivary con-
Fluoroaluminosilicate 16
tamination occurs after light curing, glass filler
significant decreases in bond strength
Dye sensitize Trace amounts
may occur.19 Fritz and colleagues19
* Bonding agent A contains phosphoric acid and methacrylate monomers.
hypothesized that if saliva contamina- Bonding agent B contains more methacrylate monomers, water and filler particles.
tion occurred after the adhesive already † Individual percentages of the acid groups are proprietary.
was polymerized and the operator
intervened by rinsing and air drying the
preparation or by applying additional COOH
adhesive after rinsing, a collapsed zone
CH3
of resin-deprived collagen could result.
They assumed that not all of the resin CH2 = C – C – O — (CH2)10 CH
in the interstices of the collagen mesh
will be polymerized. The addition of
O
more resin likely would not penetrate COOH
this altered collagen surface completely.
This assumes that some adhesive would Figure. The multifunctional acid monomer.
not fill the collagen mesh completely,
permitting its collapse after subsequent rinsing, primarily of an inorganic mineral matrix that
drying or application of additional resin. Theoreti- contains little moisture, whereas dentin consists
cally, a self-etching primer system would dem- of a mineral-impregnated organic matrix and
ineralize dentin and infiltrate it simultaneously fluid-filled tubules. Because of the high mineral
with monomers that are polymerized in situ so content of enamel, acid etching results in the for-
that no gaps would be left between the primed mation of micropores, which increases the
surface and the organic dentin surface.26 Another enamel’s surface energy for bonding and allows
theory for decreased bond strength that occurs for micromechanical retention.27 In dentin, disso-
when surfaces are contaminated with saliva after lution of the inorganic constituents results in
light curing is the adsorption of glycoproteins to exposure of the organic collagen matrix, which
the poorly polymerized, air-inhibited adhesive must be kept moist to prevent collapse.7
surface. These glycoproteins may act as a barrier These inherent differences in enamel and
that prevents complete wetting with the next dentin led to the understanding and development
increment of resin, preventing adequate of the total-etch, wet-bonding technique. Some
copolymerization.19 research has indicated that newer, hydrophilic
Results from our study suggest that self- adhesives are tolerant of saliva contamination on
etching adhesives are not as tolerant of contami- enamel,19,20 but our results demonstrated that the
nation of enamel surfaces as they are of contami- bond strength still was significantly higher in
nation of dentin surfaces. Enamel consists uncontaminated enamel specimens than in the

JADA, Vol. 135, July 2004 899

Copyright ©2004 American Dental Association. All rights reserved.
R E S E A R C H
contaminated enamel specimens (Table 2). In
similar fashion, Powers and colleagues25 deter-
mined that in vitro bond strengths to enamel
were sensitive to the effects of saliva. Re-etching
improved the bond strength, except for enamel
contaminated by plasma. In contrast, Fritz and
colleagues19 determined that the adverse effects of
salivary contamination of etched enamel were
reduced greatly if the saliva was rinsed off or
blotted dry. They also determined that any con-
tamination of the already cured adhesive layer
seriously compromised the bond—regardless of
how the contaminant was removed or altered—
and that the entire bonding procedure should be
repeated to ensure adequate adhesion.
The long-term effects of saliva contamination
are not known. It is conceivable that contamina-
tion could result in nanoleakage in the hybrid
layer itself. Nanoleakage is thought to be the
result of incomplete resin infiltration of deminer-
alized dentin, which leaves exposed collagen fibers
and results in nanopores in the hybrid layer.28
These nanopores are large enough to allow fluid
entry into the hybrid layer, but small enough to
prevent invasion by microorganisms. Nanoleakage
is a laboratory discovery, and its long-term effects
on the longevity of the bond are unknown.29 There
is, however, concern that the exposed collagen
fibrils will hydrolyze over time, leading to micro-
leakage and failure of the restoration in the long
term.Traditionally, tooth structure has been re-
etched to compensate for contamination of tooth
structure. Research by Perdigão and Lopes30
demonstrated that reapplication of acid etchant on
tooth structure demineralizes deeper into dentin
than a single application for the same amount of
time. This suggests that re-etching dentin could
result in increased risks of nanoleakage in the
hybrid layer or poor hybridization due to the col-
lapse of the collagen fibers.
Although a simple-to-use, all-in-one adhesive
system is appealing, no long-term data exist on
the clinical performance of self-etching adhesive
systems or the effects of salivary contamination.
Therefore, we recommend caution in interpreting
these in vitro results, as they cannot necessarily
be extrapolated to the clinical situation. Further
clinical studies are required, and use of proper
isolation techniques still is highly encouraged.
CONCLUSIONS
In this in vitro investigation, we did not find any
evidence to suggest a difference in mean shear
900 JADA, Vol. 135, July 2004
Copyright ©2004 American Dental Association. All rights reserved.
bond strength
between dentin
specimens con-
taminated with
saliva before or
after application
of a self-etching Dr. Townsend is the
chief, Dental
Dr. Dunn is the
director of research
adhesive system Operations, Royal Air and biomaterials,
and uncontami- Force Upwood, Ramsey,
England.
Lackland Air Force
Base, Texas. Address
nated dentin reprint requests to
Dr. Dunn at 9406
specimens. However, enamel Tranquil Park Dr.,
specimens contaminated with San Antonio, Texas,
e-mail “William.dunn@
saliva displayed a significant lackland.af.mil”.
decrease in mean shear bond
strength compared with uncon-
taminated enamel specimens. Although the shear
bond strength to dentin was not significantly
affected, the long-term effects of saliva contami-
nation on the integrity of the hybrid layer are not
known. Incomplete hybridization between the
resin adhesive and dentin likely will lead to
future failure of the restoration. Therefore, rou-
tine use of isolation techniques still is highly
encouraged. ■
The views expressed in this article are those of the authors and do
not reflect the official policy of the Department of Defense or other
departments of the United States government.
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