Influence of Degree of Conversion on the

Biocompatibility of Different Composites In Vivo

Rogrio Lacerda dos Santosa / Gisa Aiane de Morais Sampaiob / Fabiola Galbiatti de Carvalhoc /
Matheus Melo Pithond / Gymenna Maria Tenrio Gunese / Polliana Muniz Alvesf
Purpose: To evaluate the relationship between biocompatibility and degree of monomer conversion of composites used to bond brackets to enamel, porcelain, resin, or metal surfaces at different time intervals.
Materials and Methods: Twenty-four male Wistar rats were used, divided into 4 groups (n = 6) as follows: group
C (control, polyethylene), group TCC (Transbond Color Change), group QC (Quick-Cure), and group EB (Eagle
Bond). These substances were inserted into subcutaneous tissue. The events of inflammatory infiltrate, edema,
necrosis, granulation tissue, multinuclear giant cells, young fibroblasts, and collagen formation were analyzed.
The degree of conversion was evaluated by the Fourier method using infrared spectroscopy. Biocompatibility
and degree of conversion were statistically analyzed using the Kruskal-Wallis and Dunn tests, and ANOVA and
Tukeys test, respectively (p < 0.05).
Results: The composites caused a small amount of inflammatory infiltrate, edema, and granulation tissue at
all experimental time intervals, showing a gradual reduction over time (p > 0.05). Group TCC showed the highest amount of fibroblasts and EB the smallest at the time interval of 15 days (p = 0.035). Group TCC showed
the highest amount of collagen fibers and EB the smallest throughout the experiment; there was a significant
difference in terms of collagen fibers between groups QC and EB, which differed from the control at 7 days
(p = 0.006), and between groups EB and TCC (p = 0.018) at 30 days. Monomer conversion ranged from 64.1%
in group EB at 7 days to 85.3% in group TCC at 30 days.
Conclusion: Transbond Color Change composite showed a higher degree of conversion and a better healing process compared to Eagle Bond composite at 15 and 30 days. Quick-cure composite demonstrated a better degree
of conversion and healing process than that of Eagle Bond, but this was not statistically significantly different.
Keywords: composites, biocompatibility, adhesive dentistry.
J Adhes Dent 2014; 16: 1520.
doi: 10.3290/j.jad.a29704

Professor, Department of Orthodontics and Pediatric Dentistry, Federal University of Campina Grande, Patos, Paraba, Brazil. Idea, experimental design,
wrote manuscript, performed statistical evaluation.

Student of Dentistry, Department of Orthodontics and Pediatric Dentistry,

Federal University of Campina Grande, Patos, Paraba, Brazil. Performed
experiments, wrote manuscript.

Professor, Department of Orthodontics and Pediatric Dentistry, Federal University of Campina Grande, Patos, Paraba, Brazil. Performed experiments,
proofread manuscript.

Professor, Department of Orthodontics, State University of Sudoeste da

Bahia, Jequi, Bahia, Brazil. Performed experiments, contributed substantially to discussion.

Professor, Department of Restorative Dentistry, Federal University of

Campina Grande, Patos, Paraba, Brazil. Contributed substantially to discussion, proofread manuscript.

Professor, Department of Pathology, State University of Paraba, Campina

Grande, Paraba, Brazil. Performed histological tests, consulted on statistical
evaluation.

Correspondence: Rogrio Lacerda dos Santos, Federal University of Campina

Grande (UFCG), Center for Health and Technology Rural (CSTR), Av. dos Universitrios, s/n, Rodovia Patos/Teixeira, Km1, Santa Ceclia, CEP: 58700-970,
Patos, Paraba, Brazil. Tel: +55-83-9977-7100. e-mail: lacerdaorto@hotmail.com
or lacerdaorto@bol.com.br

Vol 16, No 1, 2014

Submitted for publication: 17.10.12; accepted for publication: 19.03.13

nsufficient polymerization of the resin matrix may result

in reduction in the mechanical properties and clinical
performance of the resin, such as poor bond strength,
increase in water absorption rates, solubility, and degradation.4,25 In addition, leachable components from
the resins, such as bisphenol A-diglycidyl dimethacrylate
(bis-GMA) and triethyleneglycol dimethacrylate (TEG-DMA),
found in a wide range of composites, have been shown to
have a defined cytotoxic effect.16,17 Due to the proximity
of orthodontic appliances, similar cytotoxicity could occur
in the gingiva and other oral tissues,9 as well as in the
alveolar bone if an orthodontic appliance is adhesively
bonded there after surgery to expose an unerupted tooth.
The behavior of these composite resins is directly linked
to their chemical formulation and the release of 25% to
45% of monomers left unconverted after polymerization5
when using the conventional irradiation method.7,29 Therefore, residual monomers11 may trigger limited to moderate or even severe inflammatory reactions,20 in addition
to having a direct influence on the physical, mechanical,
and biological properties of the material.6,21
15

dos Santos et al

Table 1

Composition of the composites tested

Groups

Composites

Composition*

Manufacturer

Lot no.

TCC

Transbond Plus Color

Change adhesive

Bis-GMA (5%-10%), bis-EMA (15%-20%), TEG-DMA (5%10%), quartz/silica (70%-80%), camphorquinone

3M Unitek; Monrovia,
CA, USA

N172806

QC

Quick-Cure

Bis-GMA (2%-10%), TEG-DMA (5%-10%), silica (55%-90%),

sodium fluoride (<2%), camphorquinone

Reliance; Itasca, IL, USA

116861

EB

Eagle Bond

Bis-GMA (1%-10%), bis-EMA (10%-20%), TEG-DMA (5%10%), quartz/silica (60%-80%), camphorquinone

American Orthodontics;
Sheboygon, WI, USA

A8623

*Bis-GMA, bisphenol A diglycidylmethacrylate; bis-EMA, bisphenol A bis(2-hydroxethyl ether) dimethacrylate; TEG-DMA, triethyleneglycol dimethacrylate

Table 2

Distribution of groups and day of sacrifice of rats

Control

Transbond Plus

Quick-Cure

Eagle Bond

Rats

Samples

Rats

Samples

Rats

Samples

Rats

Samples

7 days

15 days

30 days

Total

12

12

12

12

Assessment of the biological behavior of orthodontic

materials has been performed.12,20 However, there is a
lack of studies that directly relate the degree of monomer
conversion and the inflammatory and healing events associated with composites in vivo. Thus, the aim of the
present study was to evaluate the influence of the degree
of conversion on the biocompatibility of composites used
to bond brackets to enamel, porcelain, resins, or metal
surfaces at different time intervals.

MATERIALS AND METHODS

Animal Model and Experimental Groups
For this study, 24 adult male Wistar rats were used, with
a mean weight of 250 g, belonging to the vivarium of the
Federal University of Campina Grande, UACB/UFCG. The
animals were divided into 4 experimental groups (6 rats
per group): group C (control, polyethylene tube), group
TCC (Transbond Plus Color Change adhesive, 3M Unitek;
Monrovia, CA, USA), group QC (Quick-Cure, Reliance;
Itasca, IL, USA), and group EB (Eagle Bond, American Orthodontics; Sheboygon, WI, USA) (Table 1).
Each rat was to receive 2 implanted tubes (6 rats per
group, two implants per rat = 12 samples per group,
Table 2). The tubes 1.5 mm inner diameter 5 mm
long, made of polyethylene (nontoxic Scalp Vein 19G)
were fabricated and previously kept in 70% alcohol
for 120 min, washed with deionized water, and finally
autoclaved at a temperature of 110C for 20 min before
use as inoculation vehicles for the tested materials.
The composites were introduced via syringe (Centrix;
16

Shelton, CT, USA) into the openings at the ends of the

tubes supported on a glass slide at one end and a small
glass slide at the other to flatten the material. After this,
they were light polymerized for 40 s using an LED appliance (Radii, SDI; Baywater, Victoria, Australia) fixed on
a rod to guarantee that the distance to the specimens
remained constant, using a light intensity of 1000 mw/
cm2, regularly calibrated with a radiometer (Model 100,
Demetron Research; Danbury, CT, USA). After the composites were polymerized, the tubes were implanted in
the animals as follows.
The rats were anesthetized with an intraperitoneal injection of sodium thiopental (50 mg/kg) (Cristlia; Campinas, SP, Brazil). After this, trichotomy was performed in
the dorsal region of each animal, using razor blades to
eliminate the hair from a 4-cm2 area. The animal experiment was approved by the Ethics Committee on Animal
Research of the Academic Unit of Biologic Sciences,
CSTR\UFCG, Protocol CEP/No.082011.
For antisepsis of the operative field, 4% chlorhexidine
gluconate was used.20 On the midline, equidistant between the tail base to the head of the animal, two incisions approximately 8 mm long were made using a No.15
scalpel blade (Embramac; Itapira, SP, Brazil) adapted to
a scalpel handle. With the aid of blunt-tipped scissors
(Duflex, SS White: Rio de Janeiro, RJ, Brazil), the subcutaneous tissue was laterally parted to make a tunnel in
the lateral direction, forming two surgical recesses, each
approximately 18 mm deep. After two tubes per rat were
implanted, the surgical recesses were sutured with a 4.0
suture needle with thread (Ethicon, Jonhson & Jonhson;
So Jos dos Campos, SP, Brazil).
The Journal of Adhesive Dentistry

dos Santos et al

The animals were kept in cages and fed with balanced

rations food and water ad libitum according to the principles
of the Canadian Council on Animal Care.3 After time intervals of 7, 15, and 30 days, the animals were anesthetized
to obtain excisional biopsies of the implant area, including sufficient normal surrounding tissue. Finally, the rats
were sacrificed by the cervical dislocation technique after
having been sedated with sodium thiopental (50 mg/kg)
(Cristlia).

followed by Dunns test to determine the differences

among the groups (p < 0.05), because the data were not
normally distributed. The parametric data of the degree
of materials conversion were analyzed using ANOVA followed by Tukeys test (p < 0.05) to determine whether
there were statistical differences between the groups.

Biocompatibility
Posthumous samples were taken and submitted to
fixation in 4% formaldehyde (Milony solution) for 24h.
Then the polyethylene tubes containing composite
were carefully removed from the biopsied specimen
from the lateral incision to central region with the aid
of a blade No. 15 (Embramac) adapted to the scalpel
handle. Subsequently, the specimens were embedded in paraffin to obtain serial histological slices 6
m thick, and stained with hematoxylin and eosin. The
inflammatory reaction induced by the composites was
evaluated by a blind examiner using a light microscope
(BX40, Olympus; Hamburg, Germany) at 100, 200,
and 400X magnifications. The examiner was calibrated
before data analysis (kappa = 0.7). For each biopsy
taken, five representative sections of the histological
condition of the tissue adjacent to the implanted materials were evaluated.
The cellular events, ie, the presence of inflammatory
infiltrate, edema, necrosis, granulation tissue, multinuclear giant cells, young fibroblasts, and collagen, were
given points according to the following scores: 1 absent;
2 scarce, 3 moderate, and 4 intense.

Biocompatibility
All the groups presented a small amount of inflammatory infiltrate, circulatory alterations (edema), and
granulation tissue at all experimental time intervals,
showing gradual reduction with time, without statistically
significant differences among them (p > 0.05). Throughout the 30-day observation period, the composite resins
did not induce tissue degeneration (necrosis) around
or within the surgical recesses in which they were implanted. There was rarely any finding of multinucleated
giant cells (Figs 1A and 1B) in response to the nonpersistence of severe inflammatory infiltrate, and good
tolerance of the organism to the materials (Table 3).
When considering the healing process with regard to
young fibroblasts (Fig 1C), these were statistically significantly more numerous in group TCC than in group EB
(p = 0.035) at the time interval of 15 days. With regard
to the presence of collagen fibers, groups QC and EB
demonstrated significantly slower repair compared to the
control group (p = 0.006) at the time interval of 7 days.
This persisted in group EB after 30 days, which was statistically significantly different than group TCC (p = 0.018)
(Fig1D) (Table 3).
After 30 days, the control and experimental groups
showed a chronic inflammatory process characterized by
discrete events of mononuclear infiltrate, formation of
granulomas, and formation of young fibroblasts and collagen around the tubes.

Degree of Conversion
In a separate test series, standardized test samples
were prepared that measured 5 mm in diameter and
1.5 mm in thickness as follows: Stainless steel bipartite
matrices were placed on a glass slide, the composite
resins were injected into them using a syringe (Centrix) and flattened with a small glass slide, followed by
polymerization. A total of 45 test samples (n = 5 per
group) were stored in artificial saliva at 37C in lightproof boxes to prevent additional exposure to light.
At 7, 15, and 30 days after curing and storage, each
specimen was ground to obtain resin-composite powder,
which was subsequently mixed with potassium bromide
(KBr) at a ratio (by weight) of 1:10. This powder was
placed in a tablet maker under a pressure of approximately 8 tons. A spectrophotometer (Bomen-MB-102;
Dawson, Yukon, Canada) was used to evaluate the infrared spectrum measurements using the Fourier transformation method, to determine the degree of monomer
conversion in percent.
Statistical Analysis
To evaluate biocompatibility, the cellular events of inflammatory infiltrate, edema, necrosis, granulation tissue,
multinuclear giant cells, young fibroblasts, and collagen,
were submitted to the Kruskall-Wallis nonparametric test,
Vol 16, No 1, 2014

RESULTS

Degree of Conversion
Monomer conversion of the composites increased progressively up to the 30th day, with the lowest conversion
of 64.1% in group EB at 7 days and the highest of 85.3%
in group TCC at 30 days. There were no statistically significant differences between the composites at 7 and 15
days (p > 0.05). The composite Eagle Bond showed the
lowest conversion values during the experiment, which
was statistically significantly different from Transbond
(p = 0.006) at 30 days (Table 4).

DISCUSSION
The biocompatibility of the orthodontic bonding composites is an important concern, because they are frequently bonded close to or in contact with the gingival
tissues and/or alveolar bone in transsurgical bonding
for the purpose of performing tooth traction,17 and may
remain in direct contract with the tissues for months or
even years,15 possibly generating tissue damage.1,26
17

dos Santos et al
CV

PT

GC

GC

BV

CV

BV

GC

Fig 1 Photomicrographs of histological samples. PT: area of polyethylene

tube implant. A) 7 days after implantation, control group: granulation reaction area with congested vessels (CV)
and scarce multinucleated giant cells
(GC) (HE, 100X magnification; scale:
100 m). B) 7 days after implantation, group EB: moderate granulation
reaction with small caliber blood vessels (BV) and the presence of multinucleated giant cells (GC) (HE, 100X
magnification; scale: 100 m). C) 15
days after implantation, group QC:
recess surrounded by fibroblast proliferation (FP) and collagen fiber bundles
(CFB) (HE, 100X magnification; scale:
100 m). D) 30 days after implantation, group TCC: recess surrounded by
proliferation of young fibroblasts and
deposition of collagen fibers (DCF)
in the midst of a granulation reaction, scarce inflammatory cells could
still be seen (HE, 400X magnification;
scale: 25 m).

BV

CV
PT

PT

FP
CFB
DFB

Table 3 Mean of scores attributed to composites and control group after 7, 15 and 30 days, for the 7 events evaluated
Event

Time

Groups

TCC

QC

EB

12.50
10.00
10.00

13.75
11.25
10.00

Inflammatory Infiltrate

7 days
15 days
30 days

14.75
12.50
10.00

Edema

7 days
15 days
30 days

6.25
7.50
5.00

10.00a
5.00b
7.50ab

Necrosis

7 days
15 days
30 days

5.00
5.00
5.00

Granulation tissue

7 days
15 days
30 days

Giant cells

C
12.50
10.00
10.00

0.591
0.374
1.000

10.00a
5.00b
5.00ab

6.25
5.00
5.00

0.102
0.237
0.237

5.00
5.00
5.00

5.00
5.00
5.00

5.00
5.00
5.00

1.000
1.000
1.000

13.75
11.25
11.25

15.00
13.75
12.50

15.00
11.25
12.50

18.75a
7.50b
8.75ab

0.289
0.125
0.457

7 days
15 days
30 days

5.00
5.00
5.00

5.00
8.75
5.00

7.50
5.00
5.00

6.25
6.25
5.00

0.237
0.057
1.000

Young fibroblasts

7 days
15 days
30 days

8.75a
18.75Ab
17.50ab

5.00a
15.00ABb
15.00b

7.50a
10.00Bab
17.50b

6.25a
15.00ABb
15.00b

0.089
0.035
0.443

Collagen

7 days
15 days
30 days

8.75ABa
15.00ab
18.75Ab

5.00Aa
12.50ab
16.25ABb

5.00Aa
11.25b
11.25Bb

10.00Ba
15.00b
15.00ABb

0.006
0.185
0.018

These values represent the mean of scores of the sum of the five representative histological sections of the evaluated tissue. Thus, when all five representative sections of the evaluated tissue showed the same histological condition, the scores 1-absent, 2-scarce, 3-moderate, and 4-intense represent
1-absent (5.00), 2-scarce (10.00), 3-moderate (15.00), and 4-intense (20.00). Means followed by different letters express statistically significant difference
(p < 0.05) according to non-parametric Kruskal-Wallis Test, followed by Dunns multiple comparisons test, represented by: a,b (in columns, comparison
between times for each event evaluated) and A,B (in rows, comparison between composites for each time).

18

The Journal of Adhesive Dentistry

dos Santos et al

In this study, biocompatibility was evaluated in terms

of inflammatory phenomena.12,20 In studies with the
purpose of evaluating inflammatory and healing events
caused by materials,12,19 polyethelyne tubes have been
used as the control, as this is considered a substance
that is harmless to epithelial and conjunctive tissues,12
and was therefore used in the present study.
The main cause of the cytotoxic effects of resin composites is the release of unpolymerized residual monomers.1 According to the literature,8,17,20 resin components such as the methacrylate monomers TEG-DMA,
bis-GMA, UDMA, and HEMA present characteristics of
cytotoxicity and may cause cellular damage.1,16,17 In spite
of light-activated adhesives having been shown to be less
cytotoxic than the chemically activated types due to the
smaller quantity of monomers released,1,15 TEG-DMA, bisGMA, and UDMA are hydrophobic monomers frequently
associated with HEMA, and HEMA may facilitate the diffusion of these monomers into tissues, which increases the
hydrophilic character of the material. Under these conditions, they may reach the cells and cause damage.13,23
The action of bisphenol A, the bis-GMA monomer precursor, is known to cause estrogenicity.18 Moreover, the
monomers might cause soft-tissue irritation and favor
bacterial growth around a restoration.14
In this study, the events immediately after implantation
of the composite resins reflect an intense inflammatory
response due to the surgical procedures. After 7 days,
the inflammatory reaction is more organized and due to
the tested material, not to the surgical procedure,20 which
justifies an evaluation as of the 7th day, as established
in this experiment.
In the present study, little inflammatory infiltrate was
observed in any of the groups, without statistically significant differences among them. Inflammatory infiltrate
decreased throughout the study. Inflammatory infiltrate,
edema, necrosis, granulation tissue, and giant cells demonstrated no significant difference between the materials
and control group, indicating that the composite resins
were well tolerated by the organism. On the 7th day, the
composites Transbond Color Change adhesive (TCC) and
Eagle Bond (EB) induced a slightly greater inflammatory
reaction than did Quick-cure (QC), an observation supported by the lower degree of monomer conversion found
at this time point. Furthermore, bis-EMA monomers have
shown a cytotoxic effect analogous to that of TEG-DMA,10
which together may influence the inflammatory potential of
these composites.17 In this line of research, the findings
of Walther et al27 support the idea that cytoxic effects of
resin components depend on time and their concentration.
The intensity of the inflammatory process is related
to the cytotoxic character of the material. The decrease
of inflammatory intensity relies on the control of the
host defense system, which organizes itself to limit the
aggressive action from the compounds existing in the
composites; this process is usually associated with local edema, an extravasation of liquid for the cellular
interstitial space of surrounding tissues.11,12,20 In this
study, there was evidence of small dispersed edematous
areas surrounded by cell proliferation and young fibroVol 16, No 1, 2014

Table 4 Mean values and standard deviation (SD) of

the degree of conversion (%) of composites after 7,
15, and 30 days
Times

Groups
TCC

7 days
15 days
30 days

67.1 (3.4)a
76.0

(2.1)ab

85.3

(3.0)Ab

QC
71.7 (2.9)a
74.2

(6.2)ab

83.0

(4.1)ABb

EB
64.1 (3.7)a

0.093

71.3

(2.6)ab

0.283

78.1

(1.9)Bb

0.006

Means followed by different letters express statistically significant differences (p < 0.05) according to ANOVA and Tukeys post-hoc test represented by: a,b (in columns, comparison between times) and A,B (in rows,
comparison between composites for each time).

blasts, with subsequent deposition of collagen fibers

and a reduction in the number of blood vessels observed
from days 15 to 30. There were rare findings of multinucleated giant cells in the period from 7 to 15 days, and
these were frequently linked to the formation of granulomas due to the polyethylene and/or test composite.
Consequently, these phenomena followed by a repair
process in which the increased number of fibroblasts
and collagen fibers replace the areas of inflammatory
response, edema, and necrosis20 are described as a
favorable tissue response as regards the biological compatibility of the composites. On the other hand, necrosis
(tissue death caused by factors that lead to irreversible
cell damage and consequent cell death11,20) was not
observed in this study.
The gradual reduction in inflammatory reactions and the
increase of the healing process at 30 days corroborates the
findings of other studies12,20 and is related to the pattern
of monomer conversion into polymers and consequent2,6
release of residual monomers30 in the first 4 weeks.4,20
Monomer conversion may influence both the inflammatory
and healing events, and in this context, the composites
Quick-cure (QC) and Eagle Bond (EB) showed fewer signs
of healing, such as young fibroblasts and collagen fibers,
at the time interval of 7 days. This suggests slower repair
of the tissues exposed to these composites, still seen at
30 days for Eagle Bond, which had a lower conversion of
monomers at 30 days than did the other composites.
The group TCC showed lower monomer conversion than
did group QC at 7 days, but this was not statistically significantly different. In the same period, there was a tendency for faster repair (fibroblasts and collagen) in group
TCC than in group QC, suggesting that this response may
be linked to the type of unconverted monomer released
and its action in tissues.
Among several methods, Fourier transformation has
been supported as appropriate and has been widely used
as a reliable method for showing the conversion of monomers of resin-based materials.24,29 In order to evaluate
the conversion of monomers, the composite disks were
ground into powder and remade into disks with KBr. KBr
is a pure salt (transparent), undetectable by infrared spectrometry; thus, when mixed with the test material, no
spectral line appears.28
19

dos Santos et al

Other authors24 have shown general conversion values

ranging from 56% to 68%. In the present study, the values
reached from 64.1% to 85.3%, close to those found by
Corekci et al.4 The degree of conversion is dependent on the
intensity of the light source used. Usumez et al24 suggested
polymerizing for 10 to 15 s using a fast halogen lamp (850
mW/cm2) or 20 s using an LED (400 mW/cm2) in order to
obtain a similar degree of conversion among tested orthodontic composites. In this study, an LED system with a light
intensity of 1000 mW/cm2 was used for 40 s. The higher
monomer conversion observed with this light source may be
attributed to the greater light energy used on the material.
This is in agreement with the findings by Silikas et al,22 who
demonstrated with different light energy densities from the
same light source that when light energy was decreased,
the degree of conversion diminished considerably.
Comparing the inflammatory and healing phenomena
of the experimental groups with the control, it was found
that Transbond Color Change, Quick Cure and Eagle Bond
demonstrated similar biological behavior after 30 days,
with the formation of chronic inflammation, young fibroblasts, and collagen fibers.

9.

CONCLUSION

19.

Transbond Color Change composite showed a higher

degree of conversion and a better healing process as
demonstrated by the greater quantity of fibroblasts and
collagen compared to Eagle Bond composite at 15 and
30 days. Quick-cure composite demonstrated better degree of conversion and healing process than that of Eagle
Bond, but this was not statistically significantly different.

20.

10.

11.

12.

13.
14.

15.

16.

17.
18.

21.

22.

23.

ACKNOWLEDGMENTS
The authors thank the National Council for Scientific and Technological Development-CNPq for financial support (N.471372/2011)
and granting the PIBIC scholarship for this study.

24.

25.

26.

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Clinical relevance: Based on our histological results in

an animal model and degree of monomer conversion
of 3 orthodontic composite resins tested, Transbond
Color Change adhesive followed by Quick-cure showed
a higher degree of conversion and a better healing process compared to Eagle Bond.

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