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Journal of Adhesion Science and Technology
ISSN: 0169-4243 (Print) 1568-5616 (Online) Journal homepage: https://www.tandfonline.com/loi/tast20
A comparative study on monomer elution and
cytotoxicity of different adhesive restoration
materials
Tuğba Toz, Arlin Kiremitçi, Anıl Sera Çakmak, Oya Ünsal Tan, Erhan Palaska,
Menemşe Gümüşderelioğlu & Mutlu Özcan
To cite this article: Tuğba Toz, Arlin Kiremitçi, Anıl Sera Çakmak, Oya Ünsal Tan, Erhan Palaska,
Menemşe Gümüşderelioğlu & Mutlu Özcan (2017) A comparative study on monomer elution
and cytotoxicity of different adhesive restoration materials, Journal of Adhesion Science and
Technology, 31:4, 414-429, DOI: 10.1080/01694243.2016.1215768
To link to this article: https://doi.org/10.1080/01694243.2016.1215768
Published online: 01 Aug 2016.
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JOURNAL OF ADHESION SCIENCE AND TECHNOLOGY, 2017
VOL. 31, NO. 4, 414429
http://dx.doi.org/10.1080/01694243.2016.1215768
A comparative study on monomer elution and cytotoxicity of
dierent adhesive restoration materials
Tuğba Toz
a
, Arlin Kiremitçi
b
, Anıl Sera Çakmak
c,d
, Oya Ünsal Tan
e
, Erhan Palaska
e
,
Menemşe Gümüşderelioğlu
c,d
and Mutlu Özcan
f
a
Faculty of Dentistry, Department of Restorative Dentistry, School of Dentistry, Istanbul Medipol University,
Istanbul, Turkey;
b
Faculty of Dentistry, Department of Restorative Dentistry, Hacettepe University, Ankara,
Turkey;
c
Faculty of Engineering, Department of Chemical Engineering, Hacettepe University, Ankara, Turkey;
d
Faculty of Engineering, Department of Bioengineering, Hacettepe University, Ankara, Turkey;
e
Faculty of
Pharmacology, Department of Pharmaceutical Chemistry, Hacettepe University, Ankara, Turkey;
f
Dental
Materials Unit, Center for Dental and Oral Medicine, Clinic for Fixed and Removable Prosthodontics and Dental
Materials Science, University of Zurich, Zurich, Switzerland
ABSTRACT
This study evaluated monomer release and cytotoxicity of dierent
adhesive restoration materials used for dental restorations. The
extracts (1, 2, and 7days) of three types of adhesive dental restoration
materials, [Quixll (QF), Silorane Restorative (SR), and Ketac N 100
Restorative (KR)], and the adhesive resins, [XP Bond (XP), Silorane Primer
(SP), Ketac N 100 Primer (KP), and Silorane Bond (SB)] were analyzed
using high performance liquid chromatography/mass spectrometry
(HPLC-MS). The cytotoxicity levels were determined at dierent
time points (24, 48, and 72h) of cell culture using 3-(4,5-dimethyl-
2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide (MTT) assay. All
adhesive resin materials showed monomer release at varying amounts
with the highest release after 7days. The lowest amount of release
was observed in QF and the highest with KP. Bis-Phenol A (BPA) was
not detected in SP and KR that contain bisphenol-A diglycidyl ether
dimethacrylate (bis-GMA). Decamethylpenthasiloxane (D5) was not
eluted from SR. Except for SR and QF, all other adhesive restoration
materials showed dierent degrees of toxicity along with dierent
monomer release kinetics. The correlation between the monomer
release and cytotoxicity of the materials indicated that the cytotoxicity
of the materials increased with the monomer release (Spearman’s rho
correlation coecient – r). The correlation after 48h was statistically
signicant (r=−0.342, p=0.017).
Introduction
Resin-based composite materials and amalgam are both accepted as appropriate materials
for the direct restoration of class I and II cavities in restorative dentistry. However, according
to the current dental concepts, resin composites are considered as the most suitable direct
posterior lling materials since they also allow for minimal invasive restorations.[1]
KEYWORDS
Adhesion; cytotoxicity;
high-performance liquid
chromatography; L929 cell
culture; monomer release
ARTICLE HISTORY
Received 12 May 2016
Revised 7 July 2016
Accepted 12 July 2016
© 2016 Informa UK Limited, trading as Taylor & Francis Group
CONTACT Tuğba Toz ttoz@medipol.edu.tr
JOURNAL OF ADHESION SCIENCE AND TECHNOLOGY 415
e polymerizable matrix of resin materials usually contains one or more base monomers
such as bisphenol-A diglycidyl ether dimethacrylate (bis-GMA) and/or urethane dimeth-
acrylate (UDMA), diluent co-monomers such as ethylene glycol dimethacrylate (EGDMA)
and/or triethylene glycol dimethacrylate (TEGDMA), and various additives such as pho-
toinitiators (e.g. camphoroquinone), co-initiators (e.g. dimethyl-aminobenzoic-acid- ester),
polymerization inhibitors and photostabilizers (e.g. benzophenone).[2] Despite the improve-
ments in adhesive technologies, drawbacks such as polymerization shrinkage within such
materials could not be eliminated.[3] In order to overcome the shortcomings of conventional
resin-based composites, dental siloranes have been introduced that consist of a new organic
matrix.[4] Siloranes present low polymerization shrinkage due to the ring-opening oxirane
monomer, increased hydrophobicity, and thereby increased biocompatibility compared to
the conventional resin-based composites.[5]
As an alternative to resin composites, glass ionomers are also commonly used for a num-
ber of applications such as base or liner materials, as luting cements for indirect restorations,
or as temporary or permanent restorative materials for direct restorations. Glass ionomers
present several advantages of biocompatibility, low coecient of thermal expansion, the
ability to adhere to the moist tooth structure without necessitating an intermediate agent
that decreases the chair-side of the treatment procedure and anticariogenic properties due
to uoride release compared to methacrylate-based resin composites. However, they have
some drawbacks such as poor polishability and less favorable mechanical properties, surface
wear, or decreased fracture resistance over time.[6] In an attempt to overcome the disad-
vantages of glass ionomers, dierent types of glass ionomers have been developed including
resin-modied glass ionomers.[7]
e application of adhesive restoration materials in dentistry has been extensively pro-
moted over the last decade.[8] However, concerns in the dentistry over the generation
of biodegradation products have also increased and possible eects of byproducts from
composite resin matrix degradation on the function of host cells and micro-organisms are
being questioned.[9] e release of unpolymerized components from the polymerizable
resin matrix as a result of incomplete polymerization of adhesive restoration materials may
inuence the biocompatibility of the restorations.[2] Consequently, the amount of mono-
mer elution plays an important role in biocompatibility of adhesive restorative materials.
Chromatographic techniques are helpful in the analysis of compounds released from
resin-based dental llings. e components extracted from resin-modied glass ionomers,
compomers, and resin composites could be determined using gas chromatography/mass
spectrometry (MS).[10] Unfortunately, base-monomer analysis by gas chromatography is
almost impossible due to their low volatility and decomposition at higher temperature in the
injection port of the gas chromatograph. One of the few available techniques suitable for the
analysis of high-molecular-mass compounds is high-performance liquid chromatography
(HPLC), mainly coupled with mass spectrophotometry or diode array detection.[11] Several
studies have shown the elution of dierent monomers such as bis-GMA, TEGDMA, and
UDMA, from methacrylate-based resin restoration materials [12] but limited information
is available on the monomer release from silorane-based restorative materials.[12–15]
By denition, cytotoxicity of an agent is the cascade of molecular events that interferes
with macromolecular synthesis, causing unequivocal functional and structural damage
in a cell culture. e interactions of the materials and their components with the cells
at a molecular level are responsible for tissue reactions such as inammation, necrosis,
416 T. TOZ ET AL.
immunological alterations, genotoxicity, and apoptosis.[16] e in vitro cytotoxicity tests
have the advantage of easy detection of the inuence of a material on isolated cells growing
in culture plates.[17] A number of methods have been developed such as lactate dehydro-
genase assay, bromodeoxiuridine assay [18], and uorescence microscopy [19] in order to
investigate the cytotoxicity of dental resin materials. However, 3-(4,5-dimethyl-2-thiazolyl)-
2,5- diphenyl-2H tetrazolium bromide (MTT) assay is considered to be more useful to
estimate cell densities in small culture volumes and has some advantages, such as simplicity,
accuracy, reliability, and eciency.[20]
e objective of this study, therefore, was to measure the amount of monomers released
from dierent types of adhesive restoration materials with dierent monomer types using
HPLC-MS and HPLC-Ultraviolet (UV) and to assess the cytotoxicity of these materials on
L929 mouse broblast cultures at dierent time intervals. e null hypotheses tested were
that adhesive restoration materials with dierent chemistry would not show signicant
dierence in monomer elution kinetics and in cytotoxic eect on cell cultures.
Materials and methods
Specimen preparation
Disk-shaped specimens from the tested restorative materials including etch-and-rinse adhe-
sive system ‘XP’ (XP Bond, Dentsply De Trey, Germany), posterior hybrid composite ‘QF’
(Quixll, Dentsply De Trey, Germany), silorane composite ‘SR’ used with its own self-etch
adhesive system including primer ‘SP’ and bond ‘SB’ (Silorane, 3M ESPE, St. Paul, USA),
and nano-ionomer ‘KR’ applied with its own primer ‘KP’ (Ketac N 100, GC, Tokyo Japan)
were prepared (Table 1). Materials were placed in polytetrauoroethylene (Teon) molds
and processed according to the manufacturer`s instructions in a laminar ow chamber
(Bioair, Siziano, Italy). e etch-and-rinse adhesive system (XP), primer (SP and KP), and
adhesive resin (SB) disks were 5mm in diameter and 1mm in thickness and adhesive res-
toration materials (QF, SR, and KR) were 5mm in diameter and 4mm in thickness. e
adhesive restorative materials were applied in two layers, while the primers and adhesives
were applied in one layer and the surfaces of all materials were covered with a transpar-
ent strip to prevent the formation of air-inhibited surface layer. e materials were then
photopolymerized using light-emitting diode (LED, Elipar Free Light, 3M ESPE, St. paul,
USA; Light output: 1007mW/cm
2
) according to the manufacturers’ instructions (40 s for
QF, SR and KR; 20 s for XP, SP, SB, and KP). e power output of the unit was measured
with a radiometer (Cure-Rite, Dentsply-Caulk) before the placement of each restoration.
In total, 27 specimens of adhesive restoration materials and 54 specimens of primers and
adhesives were prepared.
High-performance liquid chromatography
Preparation of extracts
All specimens from material groups were divided into three subgroups (n=9 for the adhe-
sive restoration materials; n=18 for primers and adhesives) according to the extraction
period (1, 2, 7days). e extraction period was adjusted accordingly for the methyltetra-
zolium (MTT) test procedure. Immediately aer photopolymerization, the specimens in
JOURNAL OF ADHESION SCIENCE AND TECHNOLOGY 417
each subgroup were weighed (Mettler Toledo, Colombus, USA) and, except for SR, spec-
imens in all groups were placed separately into 10mL of 70% methanol/water solution
(Sigma–Aldrich, St Louis, MO USA). e extracting medium for the SR groups was pure
methanol solution as it is not soluble in methanol/water solution. e specimens were
incubated for 1, 2, and 7days at 37°C in dark. Aer the incubation period, the specimens
were removed and the extracts were analyzed using HPLC.
Table 1.The brands, types, manufacturers, batch numbers, and chemical compositions of the adhesive
restorative materials tested in this study.
Adhesive restor-
ative system
Type of the
material Manufacturer Batch number Chemical composition
Adhesive restora-
tive system
Type of the
material
Manufacturer Batch number Chemical composition
XP bond Two-step etch
and rinse
adhesive
system
Dentsply, De
Trey, Konstanz
Germany
0,707,000,223 Carboxylic acid-modified dimeth-
acrylate,phosphoric acid-modified
acrylate resin,urethane dimeth-
acrylate, TEGDMA,HEMA, butylated
benzenediol,ethyl-4-dimethylam-
inobenzoate,camphorquinone,
functionalizedamorphous silica,
t-butanol
Quixfill Methacyr-
late-based
hybrid pos-
terior resin
composite
07,030,000,799 Resin: urethane dimethacrylate
(UDMA);triethyleneglycol di-
methacrylate (TEGDMA); di- and
trimethacrylate resins; carboxylic
acid-modified dimethacrylate resin;
butylated hydroxy tolüene (BHT); UV
stabilizer, camphorquinone; ethyl-4-
dimethylaminobenzoateFillers: sila-
nated strontium aluminumsodium
fluoride phosphate silicate glass
Silorane primer Primer 3M ESPE,St. Paul,
USA
20,090,220 2-Hydroxyethyl methacrylate (HEMA)
bisphenol A diglycidyl ether
dimethacrylate (bis-GMA), water,
ethanol, phosphoric acid-meth-
acryloxy-hexylester mixture,
silane treated silica 1,6-Hexanediol
dimethacrylate,copolymer of acrylic
and itaconic acid,(Dimethylamino)
ethyl methacrylate,dl-Camphorqui-
none,phosphine oxide
Silorane bond Adhesive resin 20,090,220 Substituted dimethacrylate Silane
treated silica, triethylene glycol di-
methacrylate (TEGDMA), phosphoric
acid methacryloxy-hexylesters-
dl-Camphorquinone 1,6-Hexanediol
dimethacrylate
Silorane Silorane
restorative
20,090,220 3,4 epoxycyclohexylethylcyclopolyme-
thylsiloxane,bis-3,4-epoxycyclohex-
ylethylphenylmethylsilane, silanized
glass,yittrium floride, camphorqui-
none
Ketac N 100
Primer
Nano-ionomer
primer
3M ESPE 20,070,917 Water, HEMA, acrylic/itaconic acid
copolymer, photo-initiators
Ketac N 100 Photopolym-
erizedna-
no-ionomer
restorative
20,070,917 Paste A: silane-treated glass, si-
lane-treated ZrO2silica, silane-treat-
ed silica, PEGDMA, HEMA, bis-GMA,
TEGDMAPaste B: silane-treated
ceramic, silane-treated silica, water,
HEMA, acrylic/itaconic acidcopol-
ymer
