Assessments of antibacterial and physico-mechanical properties for dental materials with chemically anchored quaternary ammonium moieties: thiol-ene-methacrylate vs. conventional methacrylate system.

DENTAL-2490; No. of Pages 18

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Available online at www.sciencedirect.com

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Assessments of antibacterial and physico-mechanical properties for dental materials with chemically anchored quaternary ammonium moieties: Thiol–ene–methacrylate vs. conventional methacrylate system Saeed Beigi Burujeny, Mohammad Atai, Hamid Yeganeh ∗ Iran Polymer and Petrochemical Institute, PO Box 14965-115, Tehran 1497713115, Iran

a r t i c l e

i n f o

a b s t r a c t

Article history:

Objective. Fabrication of low shrinkage stress and strain dental resins containing highly avail-

Received 3 June 2014

able immobilized bactericidal moieties has been reported. The goal of this study is producing

Received in revised form

dental restorative materials with long-last antibacterial activity and reduced secondary

9 November 2014

caries.

Accepted 16 December 2014

It is anticipated that antibacterial properties of quaternary ammonium moieties chem-

Available online xxx

ically immobilized in the backbone of dental resins is directly depended on accessibility

Keywords:

monomers polymerized in a ternary thiol–ene–methacrylate system were compared with

Dental resins

corresponding classical methacrylate system against Streptococcus mutans (an oral bacteria

of these functions. In the present study the antibacterial effect of a series of antibacterial

Thiol–ene

Strain). Physical and mechanical properties of dental materials obtained from these two

Antibacterial

systems were also evaluated and compared.

Shrinkage strain and stress

Methods. The viscosities of the resin matrixes were measured on a MCR 300 rheometer.

Quaternary ammonium moieties

Degree of conversion (DC%) of monomers was measured using FTIR spectroscopy. The shrinkage-strain of photocured resins was measured using the bonded-disk technique. A universal testing machine combined with a stress measurement device was utilized to measure the polymerization-induced shrinkage stress. Viscoelastic properties of the samples were also determined by dynamic mechanical thermal analysis (DMTA). Assessment of antibacterial properties was performed through agar diffusion test (AD) to confirm non-release behavior of chemically anchored moieties. Quantitative assay of antibacterial activity was evaluated through direct contact test (DCT) against S. mutans. Direct contact cytotoxicity assay with fibroblast cell line L-929 was also performed to find more insight regarding cytotoxicity of the antibacterial matrixes. The data were analyzed and compared by ANOVA and Tukey HSD tests (significance level = 0.05). Results.

Neat

methacrylate

systems

had

significantly

higher

viscosity

than

thiol–ene–methacrylate analogous. The degree of conversion of methacrylate moieties in thiol–ene–methacrylate system was improved in comparison to conventional methacrylate system. Shrinkage stress and strain of thiol–ene–methacrylate system was

∗

Corresponding author. Tel.: +98 21 48662447; fax: +98 21 44580021. E-mail address: [email protected] (H. Yeganeh).

http://dx.doi.org/10.1016/j.dental.2014.12.014 0109-5641/© 2014 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved. Please cite this article in press as: Beigi Burujeny S, et al. Assessments of antibacterial and physico-mechanical properties for dental materials with chemically anchored quaternary ammonium moieties: Thiol–ene–methacrylate vs. conventional methacrylate system. Dent Mater (2015), http://dx.doi.org/10.1016/j.dental.2014.12.014


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lower than the neat methacrylate system. The thiol–ene-methacrylate systems show increased homogeneity and decreased glass transition temperature (Tg ) and crosslink density (c ) in comparison to the neat methacrylate-based resins. The incorporated monofuctional quaternized monomer reduces degree of conversion, shrinkage stress and crosslink density of matrix. The results showed significant improvement in antibacterial activity and cytocompatibility of dental materials obtained from thiol–ene polymerization system. Significance. It was shown that with proper control of monomers molar ratio, significant improvement in antibacterial activity and cytocompatibility as well as acceptable mechanical properties can be attained for dental resins prepared through the application of thiol–ene polymerization methodology. © 2014 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

1.

Introduction

Secondary caries, which are the main reason of dental composites failure, have been mostly attributed to the plaque accumulation adjacent to the marginal, surface and bulk cracks of dental composites [1,2]. Photopolymerization induced shrinkage strain and stress are the main reasons of crack propagation in dental matrixes [3,4]. In fact, occurrence of shrinkage strain during photopolymerization of monomers is a result of conversion of intermolecular van der Waals distances of monomers to the covalent bond length [2,5]. The shrinkage stress is another drawback occurs during photopolymerization of dental resins. This phenomenon depends on different factors in a complex manner. These factors may include monomer viscosity, shrinkage strain and photopolymerization rate [6,7]. However, shrinkage stress has mainly arisen as a consequence of propagation of shrinkage strain which occurred under confinement after gelation [8]. Many works have been carried out to reduce polymerization induced-shrinkage strain and stress [7,9–12]. Thiol–ene based dental restorative materials are currently attracted much interest as a versatile novel system for overcoming the limitation of conventional methacrylate resins [13–18]. The step growth addition mechanism of thiol–ene system result in delayed gelation and consequently significant reduction in shrinkage stress as compared with conventional methacrylate analogous [13,16–19]. Lower shrinkage stress and enhanced fracture toughness of the thiol–ene system causes significant reduction in crack development in dental composites [2,20]. On the other hand, the preparation of antibacterial restorative dental materials has attracted a great deal of attention in order to prevent secondary caries [21–23]. Antibacterial activity in dental restorative materials can be provided by incorporation of biocides like silver and zinc metals as well as organic compounds such as chlorhexidine (CHX) and quaternary ammonium salts (QAS) [24–28]. The gradual release of these antibacterial compounds may result in short-lasting antibacterial activity, reduction in mechanical properties of the dental composites and toxic side effects on the surrounding soft tissues. Therefore, preparations of dental materials with immobilized bactericidal moieties have currently attracted more attention for overcoming the deficiencies of

systems based on releasing bactericides. While these compounds inhabit activity of contacted bacteria, the active agents chemically bonded to the matrix and will not leach out [29–31]. For preparation of this category of materials, copolymerization of quaternary ammonium salt containing monomers (QASM) with proper multifunctional monomers have widely been considered. [32–38]. The success of this approach directly depends on effective collision of negatively charged bacterial cell surface with the positive charge of QAS moieties; therefore, accessibility of these functions has prime importance. In the present work, we have been trying to show that with the proper formulation of thiol–ene–methacrylate system, it is possible to tune accessibility of chemically embedded QAS moieties and reach to improved antibacterial effectiveness with reduced shrinkage strain and stress in cured matrix. We have shown that the thiol–ene polymerization method enable the preparation of matrixes with the higher possible loading of active monomer due to higher conversion of whole starting components. As well, higher degree of freedom for matrix components due to presence of flexible thiol–ether linkages will provide a higher chance for blooming and exposing of reactive groups on the surface of final networks. Different formulations based on ternary thiol–ene–methacrylate and neat methacrylate monomers containing QASM were prepared. Physical, mechanical and antibacterial activity as well as cytocompatibility of these systems was assessed and compared.

2.

Material and methods

2.1.

Materials

Camphorquinone (CQ), N,N-dimethylaminoethyl methacrylate (DMAEMA) and N,N-diethylaminoethyl methacrylate (DEAEMA) were purchased from Merck (Germany). Tetrahydrofuran (THF) was purchased from Merck (Germany) and dried via distillation over sodium wire. 2,2-Bis-(2-hydroxy3-methacryloxypropoxy)phenyl propane (Bis-GMA) and triethyleneglycol dimethacrylate (TEGDMA) were kindly donated by Evonik (Germany) and used as received. Pentaerythritol tetra (3-mercaptopropionate) (PETMP) was supplied by Aldrich (Germany). Benzyl chloride and octyl bromide were supplied by Merck (Germany). Urethane tetra allyl ether monomer (UTAE), as ene monomer was synthesized by condensation

Please cite this article in press as: Beigi Burujeny S, et al. Assessments of antibacterial and physico-mechanical properties for dental materials with chemically anchored quaternary ammonium moieties: Thiol–ene–methacrylate vs. conventional methacrylate system. Dent Mater (2015), http://dx.doi.org/10.1016/j.dental.2014.12.014


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three-necked round-bottomed flask equipped with a condenser, oil bath and magnetic stirrer. The mixture was stirred at 40–45 ◦ C for 4–5 h and then the resulting white crystals were filtered and washed several times with dry diethyl ether. The hygroscopic product was dried under vacuum at ambient temperature. Octyl bromide quaternized dimethylaminoethyl methacrylate (DMAEMA-OB) was synthesized according to the procedure reported in [32]. DMAEMA (10.7 ml, 0.064 mol) was added to octyl bromide (5.53 ml, 0.032 mol) in the presence of a small amount of hydroquinone as an inhibitor. The mixture was stirred over night at 50 ◦ C. The white powder was collected after several washings with dry diethyl ether. The product was dried under vacuum at room temperature. Synthetic routes for the preparation of different QASM monomers are depicted in Scheme 2.

2.3. Preparation of the matrix resin of dental restorative materials (methacrylate and thiol–ene–methacrylate formulations) All formulations of materials used in this study are shown in Table 1. Combination of CQ (0.5 wt.%) and DMAEMA (0.5 wt.%) were used as visible light activating photoinitiator system in all formulations. The weight ratio of methacrylate part of formulations (BisGMA and TEGDMA) was kept constant at 70 to 30, respectively. The mole ratio of thiol to ene functional groups was also kept constant at 1:1. The photopolymerization reactions were carried out by irradiation of the resin mixture with a visible light source (450–500 nm, 550 mW/cm2 , Optilux 501, Kerr, USA). The structure of networks formed through thiol–ene– methacrylate photopolymerization is depicted in Scheme 3.

2.4.

Scheme 1 – Chemical structure of monomers utilized in this study along with their abbreviations.

The viscosity of the resin matrixes was measured on a MCR 300 rheometer (Anton Paar GmbH, Austria) with a cone and plate geometry (25 mm diameter) and a separation of 0.5 mm between the plates. The measurements were performed at 27 ◦ C over the shear rate range of 0.1 to 1000 s−1 .

2.5. reaction of isophorone diisocyanate (IPDI) and trimethylolpropane diallyl ether (DAE) according to the procedure reported in our previous article [2]. The chemical structures of monomers are shown in Scheme 1.

2.2. Synthesis of quaternary ammonium salt monomer (QASM) Benzyl chloride quaternized dimethylaminoethyl methacrylate (DMAEMA-BC) and Benzyl chloride quaternized diethylaminoethyl methacrylate (DEAEMA-BC) were synthesized according to the procedure reported in [34]. Briefly, DMAEMA or DEAEMA (5.35 ml, 0.032 mol), benzyl chloride (4 ml,0.035 mol), a small amount of hydroquinone and dichloromethane (15 ml) as solvent were charged into a 100 ml

Viscosity

Degree of conversion

The degree of conversion of methacrylate functions was followed using FTIR spectroscopy (EQUINOX 55, Bruker, Germany). The thin resin specimens were placed between two polyethylene films to prevent oxygen inhibition during photopolymerization and photopolymerized using the light curing unit. DC was evaluated by comparing the absorbance spectrum of uncured methacrylate double bond (peak at 1638 cm−1 ) before and after 100 s curing of the specimen [2,5]. The spectrum of aromatic carbon–carbon double bond (peak at 1608 cm−1 ) was used as internal reference. The degree of conversion was then calculated as follows:

DC% =

1−

1636/1608 cm−1

peak area after curing

(1636/1608 cm−1 ) peak area before curing

× 100%



4


Scheme 2 – Synthetic routes for the preparation of QASM.

The equivalent degree of conversion of methacrylate functions at the maximum shrinkage rate was evaluated by assuming that there is a linear correlation between degree of conversion and shrinkage strain [5]. It was calculated through following equation:

DC%MS

VSMS DC%T = VST

where DC%MS is equivalent degree of conversion at the maximum shrinkage rate, VSMS is the shrinkage strain at the maximum shrinkage rate, DC%T is the total degree of conversion at the end of light irradiation (100 s) and VST is the total shrinkage strain at the end of light irradiation (100 s).

2.6.

Shrinkage strain

The shrinkage-strain was measured using the bonded-disk technique with a 3 mm thick glass base plate [39]. The specimens were photopolymerized for 100 s using the light source with an irradiance of circa 550 mW/cm2 . The shrinkage-strain rate was calculated by numerical differentiation of shrinkage strain data with respect to time. The shrinkage strain

rate curves were deconvoluted using Origin software (Ver. 6, Originlab Corp., Northapton, MA, USA).

2.7.

Shrinkage stress

A universal testing machine (STM-20, Santam, Iran) combined with a shrinkage measurement device and vertical visiblelight source (Optilux 501, Kerr USA) was utilized to measure the polymerization-induced shrinkage stress [40]. The tensile force generated by the polymerization shrinkage during the photocuring reaction of the resin was recorded as a function of irradiation time. Shrinkage stress was calculated by dividing the shrinkage force by the cylindrical cross-section area of the rod (diameter: d = 10 ± 0.05 mm; length: l = 50 ± 0.05 mm). The specimens were irradiated for 100 s and the stress profile was monitored for an additional 1 min. The maximum shrinkage stress value was taken from the plateau at the end of shrinkage stress/time curve.

2.8.

Viscoelastic properties

Viscoelastic properties of the specimens were determined by dynamic mechanical analysis (DMA) using a Triton

Table 1 – Different Formulations of dental resins. Sample code TM55-BC TM37-BC M-BC TM55 TM37 M TM55-OB TM37-OB M-OB TM55-EBC TM37-EBC M-EBC

Thiol/ene (phr) 50 30 – 50 30 – 50 30 – 50 30 –

BisGMA/TEGDMA (phr) 50 70 100 50 70 100 50 70 100 50 70 100

DMAEMA–BC (phr) 20 20 20 – – – – – – – – –

DEAEMA–BC (phr) – – – – – – – – – 20 20 20

DMAEMA–OB (phr) – – – – – – 20 20 20 – – –



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Scheme 3 – The structure of network formed through thiol–ene–methacrylate photopolymerization reaction.

instrument (model Tritec 2000, England) in bending mode over the temperature range from −100 to 200 ◦ C at a heating rate of 5 ◦ C min−1 and frequency of 1 Hz. The storage modulus, loss modulus, and loss tangent were recorded as a function of temperature. Glass transition temperature (Tg ) was taken as the maximum of loss tangent. The crosslink density (c ) of cured resins was calculated using DMA data from rubber elasticity theory according to the following equation [19,41,42]: ϑc =

E 3RT

where E designates storage modulus at rubbery plateau region. R and T stand for gas constant and absolute temperature, respectively. Tg1/2width was measured as the half peak width of tan ı curves.

2.9.

Bulk hydrophilicity

Disc shape specimens with 7 mm diameter and 1 mm thickness were fabricated through light curing for 100 s on each side. The prepared specimens were transferred to a drying oven maintained at 37 ◦ C until a constant mass (W0 ) was reached. They subsequently immersed in distilled water and incubated at 37 ◦ C to release unpolymerized monomers. After 72 h, they were removed and stored in drying oven until a constant weight (Wd ) was obtained. The specimens were then placed back in distillated water and periodically were picked up and reweighed (Wt ) after removing of surface moisture with

tissue paper [43]. This procedure was continued for 24 h. The water uptake (%) was determined using the following equations: Watere uptake% =

2.10.

Wt − Wd × 100 Wd

Antibacterial assay

2.10.1. Agar diffusion test None-release behavior of chemically anchored quaternary ammonium salt containing polymers was evaluated through qualitative agar diffusion test (AD). In this test, disc shaped specimens (1 cm diameter) were exposed to S. aureus (ATCC 25923) and E. coli bacteria (ATCC 25922) to observe the inhibition zone around and under the specimens. The specimens were placed on an LB agar plate (Himedia) which previously seeded with 1.0 × 106 colony forming units (CFU) of bacteria for 5–10 min. After incubation at 37 ◦ C for 18–24 h, inhibition zone for bacterial growth was determined visually [44].

2.10.2. Direct contact test For quantitative measurement of antibacterial activity, direct contact test (DCT) was evaluated against Streptococcus mutans (ATCC 35668). The disc shaped samples (1 cm diameter) were incubated in 37 ◦ C sterilized artificial saliva for 24 h. Ten microliters of overnight cultured S. mutan suspension (approximately 106 CFU) were then placed on the samples in 24-well


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micro titer plate and the plate was incubated for 1 h in a humid anaerobic atmosphere at 37 ◦ C. During the incubation period, the suspension liquid, brain–heart infusion (BHI), evaporated and a thin layer of bacteria was maintained in direct contact with the samples. BHI (1 ml) was then added to each well. The ingredients were mixed through gentle pipetting for 2 min. The plate was incubated for 4 h in a humid anaerobic atmosphere at 37 ◦ C. A serial tenfold dilution was made with BHI broth and each suspension was placed on blood agar plates and incubated for 48 h. The CFU of each plate was then counted [24,45]. A blank sample from each formulation with no QAS moiety was tested as negative control. Results were expressed as log reduction in CFU according to reduction in CFU = the following equation:Log Log10

CFU(control cured resin) CFU(antibacterial cured resin)

Fig. 1 – Viscosities of M, TM37 and TM55 as a function of shear rate.

,

1 log reduction means a 10 fold or 90% reduction in CFUs.

2.11.

Cytotoxicity

The possible cytotoxicity of resins was evaluated on mouse L929 fibroblast cells. The photocured samples were sterilized by incubation at 120 ◦ C for 15 min before test. Mouse fibroblast cells (1 × 104 cells per well) were incubated in a 24-well plate containing RPMI-1640 medium/10% fetal FBS at 37 ◦ C under 5% CO2 atmosphere for 24 h. Then, the samples were placed in the center of each well, and incubated in the same conditions for 48 h. A growth medium, containing cells but no resin was tested as negative control. The morphological changes indicating cytotoxicity and cell growth characteristics of attached cells were recorded using a Nikon TMS inverted optical microscope equipped with a Sony DSC-W7 camera [44].

2.12.

Statistical analysis

The results were analyzed and compared using one-way ANOVA and the Tukey test at the significance level of 0.05.

3.

Results

The structures of monomers utilized for the preparation of dental restorative matrixes are shown in Scheme 1. A urethane bond containing tetrafunctional ene monomer was synthesized and characterized as described in our previous report [2] and used along with methacrylate monomers. Three different quaternary ammonium salt containing monomers were also prepared through alkylation reaction of dialkylaminoethyl methacrylate with either benzyl chloride or octyl bromide (Scheme 2). Different formulations of ternary thiol–ene-methacrylate system, used in the study, are shown in Table 1. A mixture of CQ (0.5 wt.%) and BD (0.5 wt.%) was used as visible light activated photoinitiator in all formulations. The weight ratio of methacrylate part of formulations (BisGMA and TEGDMA) was kept constant at 70 to 30, respectively. But the weight ratio of methacrylate to thiol–ene monomers were altered as tabulated in Table 1. To evaluate the effects of QASM on physical, mechanical and biological properties of final dental restorative materials, each of these functional monomers (20 wt.%) were added to both ternary thiol–ene–methacrylate and methacrylate systems.

The monomer mixtures and resulting networks (Scheme 3) were subject to different studies.

3.1.

Viscosity

Figs. 1 and 2 represent the viscosity of different resin mixtures (TM55, TM37, M, TM55-BC, TM37-BC and M-BC) as a function of shear rate. It was observed that the neat methacrylate systems had significantly higher viscosities than thiol–ene–methacrylate analogous. Increasing thiol–ene content slightly reduced the viscosity of resin mixture. But, addition of DMAEMA-BC to monomer mixtures increased the viscosity of the resins (Figs. 2 and 3).

3.2.


Table 2 represents the degree of conversion of methacrylate groups (DC %). All of thiol–ene based networks showed higher DC% and DC%MS in comparison to the neat methacrylate systems (P < 0.05). Also DC% and DC%MS were improved as the content of thiol–ene moieties increased (P < 0.05). Incorporation of DMAEMA-BC into resin mixtures decreased the DC% and DC%MS . Fig. 4 shows the variation of DC% and shrinkage strain versus methacrylate content. Incorporation of thiol–ene moieties into resin mixture noticeably increased methacrylate DC%, even though resulted in lower shrinkage strain.

Fig. 2 – Viscosities of M-BC, TM37-BC and TM55-BC as a function of shear rate.


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Table 2 – Maximum shrinkage strain rate (%/S), time at a maximum shrinkage strain rate (S), volume shrinkage at maximum shrinkage strain rate %, total volume shrinkage %, equivalent conversion at maximum shrinkage strain rate%, total degree of conversion and water uptake% of specimens1,2 . Sample

Maximum shrinkage strain rate (%/S)

Time at maximum shrinkage strain rate (S)

Volume shrinkage at Maximum shrinkage strain rate %

Total volume shrinkage %

TM55 TM37 M TM55-BC TM37-BC M-BC

0.11 (0.012)a 0.23 (0.01)b 1.33 (0.03)d 0.16(0.02)e 0.27(0.01)f 1.12(0.05)g

30.3 (2.6)a 17.2 (1.2)b 2.7 (0.3)c 14.2 (1.3)d 10.2 (0.6)e 2.5 (0.3)c

2.50 (0.29)a,c 2.70 (0.21)a 2.58 (0.12)a 1.70 (0.11)b 2.10 (0.13)c 2.20 (0.20)c

6.01 (0.23)a 6.75 (0.19)a 7.52 (0.24)b 5.70 (0.13)c 6.50 (0.20)a 7.55 (0.11)b

1

2

DC%

84.72 (1.16)a 80.17 (2.40)a 63.30 (4.61)b 82.58 (2.20)a 74.77 (0.30)c 59.87 (2.42)b

Equivalent conversion at maximum shrinkage strain rate%

Water uptake%

37.02 (1.60)a 33.01 (2.35)b 21.74 (1.45)c,e 25.49 (1.05)d,e 24.24 (2.29)e 17.50 (1.64)f

1.23 (0.34)a 1.23 (0.19)a 1.30 (0.14)a 8.72 (0.46)b 5.43 (0.15)c 5.53 (0.46)c

According to analysis of variances (P ≤ 0.05) the difference between quantities with similar superscripts (a, b, c, d, e, f and g) is not significant for data of each column. For data of each column, standard deviation value is shown in parenthesis.

strain rate parameters are indicated in these curves. These values are measured for different formulations of studied resins and corresponding data are tabulated in Table 2. The curves related to variation of shrinkage strain rate versus irradiation time for samples with and without thiol–ene and DMAEMA-BC moieties are presented and compared in Fig. 6a–d. With introducing thiol–ene moieties into the neat methacrylate formulation, the maxima of shrinkage strain rate decreased and shifted to higher irradiation time (Fig. 6a). Meanwhile, shrinkage strain value decreased with increasing thiol–ene content (Fig. 6a ). Also, higher shrinkage rate maximum at lower irradiation time were recorded for thiol–ene Fig. 3 – Comparison of viscosity between QAS-incorporated resins and neat resins as a function of BiSGMA/TEGDMA content.

3.3.

Shrinkage strain

Fig. 5a shows the curves of shrinkage strain and shrinkage strain rate versus irradiation time for a typical methacrylate based dental resin mixture. Total shrinkage strain, maximum shrinkage strain rate, time at maximum shrinkage strain rate and shrinkage strain at the maximum shrinkage

Fig. 4 – DC% and volume shrinkage versus methacrylate content for TM55-BC, TM37-BC and M-BC resins.

Fig. 5 – Typical Shrinkage strain curve and shrinkage strain rate curve for M specimen (a), Typical Shrinkage stress curve for M specimen (b).



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Fig. 6 – Characteristic shrinkage strain rate curves for specimens, TM55,TM37 and M (a), TM55-BC,TM37-BC and M-BC (b), TM55 and TM55-BC (c), TM37 and TM37-BC (d), M-BC and M (e), Shrinkage strain curve for specimens, TM55,TM37 and M (a ), TM55-BC,TM37-BC and M-BC (b ), TM55 and TM55-BC (c ), TM37 and TM37-BC (d ), M-BC and M (e ), Shrinkage stress curve for specimens, TM55,TM37 and M (a ), TM55-BC,TM37-BC and M-BC (b ), TM55 and TM55-BC (c ), TM37 and TM37-BC (d ), M-BC and M (e ).

based samples containing DMAEMA-BC monomer (Fig. 6b). In contrast to thiol–ene based formulations, incorporation of DMAEMA-BC monomer in the neat methacrylate system led to lower value of shrinkage strain rate maxima, without significant difference in time needed to reach to maximum shrinkage strain rate (Fig. 6e). To find better perspective

regarding mode of reactions in photopolymerization of thiol–ene containing system, variation of shrinkage strain rate during irradiation time was studied for different formulations and the curve of thiol–ene based formulations were deconvoluted using Origin software. The curves of thiol–ene containing systems showed a shoulder at the beginning of



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higher induction time in comparison to the neat methacrylate analogous (P < 0.05). Within thiol–ene–methacrylate based networks those containing more thiol–ene components exhibited lower shrinkage stress and higher induction time. Induction time and total shrinkage stress of samples are decreased upon addition of DMAEMA-BC monomer to formulations (Table 3 and Fig. 6c –e ). However, there was no significant difference in induction time between M and M-BC specimens (P > 0.05). The ratio of shrinkage strain before the gel point to the total shrinkage strain (Shrinkage strain(gel) /Shrinkage strain(total) ) was calculated for DMAEMABC containing system. Gelation time was adopted from the initial part of shrinkage stress versus irradiation time curve (Fig. 6a –d ) and Shrinkage strain(gel) /Shrinkage strain(total) parameter was calculated from shrinkage strain versus irradiation time curve (Fig. 6a –d ). Shrinkage strain(gel) /Shrinkage strain(total) parameter as well as shrinkage stress values were plotted against methacrylate content of each system and shown in Fig. 9. [19].

3.5.

Fig. 7 – Shrinkage strain rate of thiol–ene–methacrylate and pure methacrylate specimens: (a) TM55, TM37 and M; (b) TM55-BC,TM37-BC and M-BC.

photopolymerization time (Fig. 7). Also, shrinkage strain rate curve of thiol–ene–methacrylate based systems deconvoluted into two Gaussian components (Fig. 8).

3.4.

Shrinkage stress

The maximum shrinkage stress and induction time needed for shrinkage stress buildup was calculated from curves of shrinkage stress versus time variation of irradiation (Figs. 5b and 6a –d ) and extracted data were collected in Table 3. All thiol–ene based samples showed lower shrinkage stress and

Table 3 – Induction time and total shrinkage stress adopted from shrinkage stress curve1,2 . Code TM55-BC TM37-BC M-BC TM55 TM37 M 1

2

Induction time (S) 15.48(0.502)a 9.19(0.35)b 2.14(0.16)c 26.97(0.92)d 12.78(0.65)e 1.97(0.23)c


Fig. 10 shows the storage modulus and loss tangent versus temperature for different formulations. The parameters including Tg , Tg1/2width , c and storage modulus of samples were extracted from DMA curves and collected in Table 4. Networks made through thiol–ene–methacrylate system showed lower Tg and c in comparison to neat methacrylate analogous. Increasing thiol–ene content was also decreased these values. The Tg1/2width for thiol–ene–methacrylate samples was lower than that for neat methacrylate analogous, which confirmed higher homogeneity of thiol–ene based networks. The homogeneity of system increased as contribution of thiol–ene content increased. Incorporation of DMAEMA-BC monomer into networks caused reduction of crosslink density, while, showed higher storage modulus in the glassy state.

3.6.

Bulk hydrophilicity

The water uptake percent of different formulations with and without DMAEMA-BC monomer is collected in Table 2. This value is almost similar for pure methacrylate and thiol–ene based system without DMAEMA-BC (P > 0.05). However, upon introduction of DMAEMA-BC monomer significant difference was observed for water uptake value of these resins in comparison to those without DMAEMA-BC containing

Total shrinkage stress (MPa) 0.08(0.01)a 0.17(0.02)b 0.31(0.01)c 0.11(0.02)d 0.19(0.02)b 0.36(0.01)e

According to analysis of variances (P ≤ 0.05) the difference between quantities with similar superscripts (a, b, c, d and e) is not significant for data of each column. For data of each column, standard deviation value is shown in parenthesis.

Table 4 – Data obtained from DMTA of different specimens. Code

TM55-BC TM37-BC M-BC TM55 TM37 M

Storage modulus (GPa)

Crosslink density (mol/L)

2.15 3.17 3.11 1.57 2.03 2.17

0.62 1.57 2.61 0.98 2.70 4.97

Tg (◦ C)

81.9 94.4 128.5 68.7 91.5 171.3

Tg1/2 (◦ C)

44.5 48.7 74.5 26.3 40.6 63.2



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Fig. 8 – Shrinkage strain rate curve deconvoluted into two Gaussian components. Dash line represents the methacrylate homopolymerization stage and round dot line corresponds to the thiol–ene polymerization reactions: (a) TM55; (b) TM37; (c) TM55-BC (d), TM37-BC.

monomer (P < 0.05). Especially, the sample TM55-BC with the highest contribution of thiol–ene component showed the highest amount of water uptake. But there was no significant difference between the behavior of TM37-BC and M-BC (P > 0.05).

3.7.

Antibacterial assays

3.7.1.

Agar diffusion test

antibacterial activity and reduced the number of bacterial colony more efficiently than neat methacrylate system. However, no significant difference was observed between TM-37 and M based samples (P > 0.05).

Representative images showing behavior of TM55-BC sample against both gram positive (S. aureus) and gram negative (E. coli) bacteria are depicted in Fig. 11. According to this figure, no inhibition zone was detected around the sample and no bacterial growth was observed on the area under the specimen. Meanwhile, no bacterial colonies attached to sample surface were detected.

3.7.2.

Direct contact test

Fig. 12 shows the respective plate images of DCT assay and the result of Log reduction in CFU are collected in Fig. 13. As it represents, sample TM55-BC showed the highest

Fig. 9 – The ratio of the shrinkage strain which arises prior to gelation relative to the total shrinkage strain (Triangle symbols) and shrinkage stress (square symbols) vs. methacrylate content for TM55-BC, TM37-BC and M-BC resins.




11

Fig. 10 – DMA curves for cured samples: Tan delta of samples without DMAEMA-BC monomer (a) and based on DMAEMA-BC monomer (c) and storage modulus of samples without DMAEMA-BC monomer (b) and based on DMAEMA-BC monomer (d).

3.8.

Cytotoxicity

Since cytocompatibility of antibacterial dental resins is as important as their biocidal activity, their behavior against

fibroblast cells was evaluated through microscopic investigation of cells morphology in vicinity of samples. Fig. 14 shows the optical microscopic images of L929 cells after incubation with samples. The results were compared with tissue culture

Fig. 11 – Results of agar diffusion test for TM55-BC sample against E. coli (a and b), S. aureus (c and d) bacteria. Please cite this article in press as: Beigi Burujeny S, et al. Assessments of antibacterial and physico-mechanical properties for dental materials with chemically anchored quaternary ammonium moieties: Thiol–ene–methacrylate vs. conventional methacrylate system. Dent Mater (2015), http://dx.doi.org/10.1016/j.dental.2014.12.014


12


Fig. 12 – Respective plate images of direct contact test assay against Streptococcus mutans for DMAEMA-BC incorporated specimens and blank samples: (a) TM55-BC, (b) TM55, (c) TM37-BC, (d) TM37, (e) M-BC and (f) M.

plate as negative control. As shown in Fig. 14 none of samples exhibit cytotoxicity response.

4.

Discussion

4.1.

Viscosity

Since the initial viscosity of dental resins has considerable effect on some important parameters such as DC%, shrinkage

strain and stress, the initial viscosity of the neat methacrylate system and thiol–ene–methacrylate systems with and without antibacterial monomer was measured as a function of shear rate (Figs. 1 and 2). Generally, the viscosity and flow property of the monomer mixture depend on the intermolecular interactions, free volume and molecular weight of components. The high viscosity of BisGMA relates to powerful intermolecular hydrogen bonding of hydroxyl groups present in its backbone. Reduction of overall viscosity via the addition of thiol–ene




Fig. 13 – Log reduction in CFU of the photopolymerized antibacterial specimen.

components can be related to the lowering of intermolecular hydrogen bonding between BisGMA molecules. Addition of DMAEMA-BC caused a considerable increase in viscosity for both systems (Figs. 2 and 3). This observation can be attributed to ionic interaction between QAS moieties.

4.2.


The progress of polymerization reaction and network formation was evaluated by measurement of methacrylate group conversion (DC%). Improvement of DC% in the

13

presence of thiol–ene moieties is attributed to the lower initial viscosity of monomer mixture. Furthermore, radical mediated step growth polymerization mechanism of thiol–ene leads to formation of flexible thioether linkages and longer pregelation step. Therefore, the delayed vitrification point is expected for thiol–ene containing systems [2]. In fact, after reaching the vitrification point, the movements of growing radicals are restricted considerably. Propagation of radicals become diffusion controlled and combination of these events can lead to an overall reduction of DC%[5]. For thiol–ene based systems with prolonged pregelation step, all of these events will be postponed and higher overall DC% and DC%MS is attained. For better illustration, the variation of DC% and shrinkage strain versus methacrylate content is shown in Fig. 4. Although thiol–ene systems had higher DC%, lower shrinkage strain was recorded for them. This observation confirms the lower shrinkage strain factor associated with each double bond in thiol–ene systems (12–15 cm3 /mol) in comparison to shrinkage strain factor of neat methacrylate (22–23 cm3 /mol) per each reacted double bond [13,16]. The significantly higher shrinkage strain factor for the neat methacrylate system is in part related to the homopolymerization behavior of methacrylate moieties. Incorporation of DMAEMA-BC into resin mixture decreased DC% and DC%MS of methacrylate functions. This observation is attributed to the increase of reaction medium viscosity in the presence of DMAEMA-BC. Viscosity buildup will restrict availability of monomers for participating in photopolymerization reaction. This phenomenon will shorten the pregelation step and consequently the vitrification step is reached more quickly at a

Fig. 14 – Optical microscopy of L-929 cells in direct contact with the samples: negative control (a), TM55-BC(b), TM37-BC (c), M-BC (d). Please cite this article in press as: Beigi Burujeny S, et al. Assessments of antibacterial and physico-mechanical properties for dental materials with chemically anchored quaternary ammonium moieties: Thiol–ene–methacrylate vs. conventional methacrylate system. Dent Mater (2015), http://dx.doi.org/10.1016/j.dental.2014.12.014


14


lower degree of conversion for DMAEMA-BC containing system.

4.3.

Shrinkage strain

As it is shown in Table 2 and Fig. 6(a ), shrinkage strain of the specimens decreases with increasing the thiol–ene content and the highest value is recorded for the specimens containing neat methacrylate monomers. This observation is attributed to the thiol–ene photopolymerization mechanism. Thiol–ene photopolymerization proceed via a radical mediated stepgrowth polymerization mechanism in which propagation and chain transfer take place alternatively.[14–16,20,46]. In an ideal thiol–ene photopolymerization each double bond reacts with just one thiol-containing monomer. However, in common chain-growth radical photopolymerization, two monomers are generally coupled to each double bond [13,16,41,46]. This difference implies less contraction of Van der Wasls distances between (macro)molecules as the polymer forms. This phenomenon leads to the reduced density difference between monomer and polymer in thiol–ene polymerization in comparison to common methacrylate polymerization [16]. Furthermore, the longer length of the carbon–sulfur bond ˚ in comparison to the carbon–carbon bond length (1.81–2.55 A) ˚ leads to increase the distance between crosslink(1.20–1.54 A) ing sites. The combination of aforementioned events results in reduced shrinkage strain of thiol–ene based system in comparison to the neat methacrylate ones. On the other hand, there is a direct connection between the development of shrinkage strain during polymerization reaction and consumption of the double bonds of monomers. As a result, this factor can be considered as an indicator for polymerization reaction progress [5]. Therefore, derivative of shrinkage strain values versus irradiation time (shrinkage strain rate) can be used for evaluation of photopolymerization rate. According to Fig. 6a–d, all curves show a rapid increase in the shrinkage strain rate at the earliest stage of light curing followed by a dramatic decrease in shrinkage strain rate. These events are representative of auto-acceleration and auto-deceleration stages of polymerization reaction respectively. Auto-acceleration stage occurs after the gelation point when termination reaction becomes diffusion controlled. This phenomenon has led to an increase in the propagation rate. By increasing the viscosity of reaction mixture, the propagation reaction also becomes diffusioncontrolled and the rate of reaction decreases considerably after vitrification stage [47]. This decrease in the shrinkage strain rate is called auto-deceleration effect [5]. The maximum of shrinkage–strain rate curve, which is an indicator of the polymerization rate, is accompanied by vitrification stage and time at the maximum shrinkage strain rate represents the vitrification time. The decrease observed in maximum shrinkage strain rate in the presence of thiol–ene components is attributed to the polymerization kinetics of these systems. According to the polymerization kinetics of thiol–allyl ether–methacrylate systems the propagation rate constant of methacrylate homopolymerization reaction is higher than both chain transfer rate constant to thiol functional groups and propagation rate constant for the cross polymerization reaction of methacrylate and allyl ether monomers [48]. However, at the initial stage of photopolymerization

reaction with high concentration of methacrylate functional groups, the reaction rate for methacrylate homopolymerization is much higher not only than the rate of methacrylate radical chain transfer to thiol moieties, but also than the methacrylate–allyl ether cross coupling reactions [41,48]. In the following stage and after significant depletion of methacrylate functions, an abrupt increase is observed in thiol–ally ether polymerization rate. As a result, polymerization of thiol–allyl ether–methacrylate system can be divided into two distinct stages including methacrylate homopolymerization and simultaneous chain transfer to thiol moieties followed by thiol–allyl ether step growth polymerization reaction as the second stage [41,48]. Inspection of shrinkage strain rate behavior of thiol–ene containing specimens, confirms two distinct photopolymerization mechanisms of ternary thiol–ene–methacrylate system (Fig. 7). The curves related to thiol–ene–methacrylate based systems consists a broad peak with a shoulder at about 3–4 s and a maximum at about 20–30 s. The first part of the curve (solid line, up to 4 s) with a rapid increase in the shrinkage strain rate represents the methacrylate homopolymerization reaction which occurs at the initial photopolymerization stage and the second part of the curves (dotted line, 4 to 30 s) with a lower increase in shrinkage strain rate correspond to the thiol–ene polymerization reactions. As a result, increasing thiol–ene content of formulations leads to higher contribution of thiol–ene reaction (second stage) and consequent lower final maximum shrinkage strain rate during photopolymerization reaction. For better illustration, the shrinkage strain rate curves were deconvoluted using Origin software (Fig. 8). Each curve of thiol–ene–methacrylate based systems was split into two Gaussian peaks, which confirms two distinct photopolymerization mechanisms in thiol–allyl ether–methacrylate systems. The first peak at about 3–4 s (dash line) represents methacrylate moieties homopolymerization reaction and the second peak at about 20–30 s (round dotted line) corresponds to the thiol–allyl ether polymerization reactions. Incorporation of DMAEMA-BC monomer in thiol–ene based system had no significant effect on photopolymerization stages and two distinct peaks were observed by deconvolution. As indicated in Table 2 for thiol–ene–methacrylate based systems (TM55-BC and TM73-BC), the maximum shrinkage strain rate of polymerization reaction increases in the presence of DMAEMA-BC monomer (Fig. 6c and d). This observation is attributed to the extra concentration of methacrylate functions arising from DMAEMA-BC monomer. However, incorporation of this monomer to the neat methacrylate based dental resins (M), causes a reduction in maximum shrinkage strain rate (Fig. 6e). DMAEMA-BC is a monofunctional methacrylate monomer with lower reactivity than difunctional methacrylate monomers (BisGMA/TEGDMA), but higher reactivity than allyl ether functions of ene monomer [48]. Therefore the copolymerization of DMAEMA-BC monofunctional monomer with difunctional methacrylate monomers results in reduced maximum shrinkage strain rate. However, incorporation of DMAEMA-BC monomer in thiol–ene based systems will increase the concentration of methacrylate functions and increases maximum shrinkage strain rate. Investigation of Fig. 6a–d and Table 2 data reveals that more time needs to reach to maximum of shrinkage strain rate for




a system consisting higher contribution of thiol–ene components. This phenomenon can be attributed to the reaction medium viscosity. In fact, not only the presence of thiol–ene moieties decreases the initial viscosity of the neat methacrylate mixture (Fig. 1), but also, the buildup of viscosity occurs more slowly during the progress of thiol–ene photopolymerization reactions through step-growth mechanism. On the other hand, the radical mediated step-growth polymerization mechanism of thiol–ene systems results in formation of relatively low molecular oligomeric species in the initial stage of photopolymerization. The overall consequence of this phenomenon is a low viscosity of reaction medium during polymerization. As a consequence, lower viscosity of medium consisting thiol–ene components facilitates propagation of macroradicals and consequently leads to delayed vitrification point [5,49]. Table 2 also reveals the shift of maximum shrinkage strain rate to lower polymerization time by incorporation of DMAEMA-BC monomer. This observation can be attributed to ionic interactions between the ionic center of DMAEMABC moieties which increases the viscosity of reaction medium (Figs. 2 and 3). Therefore, in this formulation, vitrification point attained at shorter time due to diffusion-controlled nature of termination reactions [5].

4.4.

Shrinkage stress

For studied formulations, there is a short time lag between the beginning of light curing and the first stress recorded (Figs. 5b and 6a –d ). This induction time indicates photopolymerization-induced gelation. After this period a two-stage rapid linear increase in shrinkage stress with different slopes was observed followed by a very slow increase of stress when the lamp was switched off [40]. All thiol–ene based samples showed lower shrinkage stress and higher induction time in comparison to the neat methacrylate analogous. This phenomenon attributed to the delayed gelation of thiol–ene system, arising from step growth polymerization nature of this reaction [18]. The amount of shrinkage strain occurs prior to gelation stage, which has considerable contribution in the total shrinkage strain of thiol–ene system, can be accommodated by movement of segments rather than residual stress buildup [6]. Since shrinkage stress is a consequence of the shrinkage strain occurring under confinement and restricted polymer network after gelation, then, lower shrinkage strain and delayed gelation of thiol–ene methacrylate systems result in a dramatic reduction of photopolymerization shrinkage stress [16]. For better illustration, the ratio of shrinkage strain before the gel point to the total shrinkage strain (Shrinkage strain(gel) /Shrinkage strain(total) ) was calculated. This parameter as well as shrinkage stress values were plotted against methacrylate content of each system (Fig. 9). All of the methacrylate-thiol–ene systems exhibited higher shrinkage strain(gel) /Shrinkage strain(total) relative to the neat methacrylate formulation, which results in lower final shrinkage stress in the former system. On the other hand, incorporation of DMAEMA-BC monomer in thiol–ene–methacrylate system increases the rate of shrinkage stress buildup. In contrast, higher shrinkage stress rate is recorded for M specimen in comparison to M-BC system. This observation can be attributed to the viscosity

15

of reaction medium and the photopolymerization rate of systems. Strong electrostatic interactions between ions of DMAEMA-BC moieties are responsible for higher viscosity and shorter pregelation step during photopolymerization reaction of DMAEMA-BC containing systems. Shorter pregelation step decreases the induction time and increases the rate of shrinkage stress buildup in thiol–ene systems. On the other hand, the high viscosity of DMAEMA-BC containing system leads to lower total conversion and consequently, reduces the maximum shrinkage stress. For neat methacrylate system lower reactivity of monofunctional DMAEMA-BC monomer is responsible for the lower rate of shrinkage stress buildup.

4.5.


The thiol–ene–methacrylate systems (TM55 and TM37) show increased homogeneity and decreased Tg and c in comparison to pure methacrylate-based resins (M). This behavior is attributed to the polymerization mechanism of this system. As it was described previously, this mode of reaction leads to increased distance between cross-linking sites and consequently decreased cross-linking density is obtained for thiol–ene based systems. Also, the lower Tg of the thiol–ene–methacrylate network relative to neat methacrylate system is the consequence of the flexible thioether linkages formation and lower overall cross-linking density [41]. Conventional methacrylate systems, on the other hand, have a heterogeneous structure which is resulted from nonuniform distribution of densely crosslinked parts within the loosely crosslinked section. This heterogeneous structure is a consequence of agglomeration of the high crosslinked microgels which dispersed in the unreacted monomers[50]. Increasing thiol–ene content decreased Tg and c values. The homogeneity of system increased as contribution of thiol–ene content increased. This observation is attributed to the reduced methacrylate chain length due to the occurrence of more chain transfer to thiol functions during methacrylate homopolymerization. The decrease in methacrylate chain length not only leads to reduced Tg , but also results in decreased crosslink density while enhancing homogeneity of network [18,41]. The viscoelastic properties of the specimens are influenced by incorporation of DMAEMA-BC monomer too. The incorporated monomfuctional DMAEMA-BC monomer reduces the crosslink density of the matrix. Consequently, the storage modulus value at the rubbery plateau region is lowered for DMAEMA-BC-containing resins in comparison to neat ones. However, DMAEMA-BC-containing resins have higher storage modulus value at glassy region. This is attributed to the formation of new ionic intermolecular interactions owing to the presence of ions in DMAEMA-BC that can compensate the effect of lower crosslink density [51].

4.6.

Bulk hydrophilicity

Antibacterial activity of dental resins with non-releasing chemically bonded bactericidal, extremely depends on close contact between active agents on the surface of cured resins with surrounding bacteria. The amount of absorbed water in final cured resin is a determining factor that can influence on the accessibility of QAS functions embedded into



16


the resin structure. Water can act as an internal plasticizer and reduce the intermolecular interaction of chain segments. Therefore, the degree of freedom for immobilized QAS functions increases and these groups find more chances to expose at the surface of cured resins. The absorbed water was measured as a means to access the hydrophilicity of the samples after 24 h. The sample TM55-BC with the highest contribution of thiol–ene component showed the highest amount of water uptake. Generally, this observation is attributed to the higher accessibility of the quaternary ammonium moieties in thiol–ene reach system and consequently, the higher chance for salvation of ions in moist environment. It is worth to mention that for such events reaching to a certain degree of freedom is essential. It seems this criterion was met for TM55-BC sample.

4.7.

Antibacterial assay

4.7.1.

Agar diffusion

In order to elucidate the effect of matrix homogeneity and accessibility of immobilized bactericidal functions on the antibacterial effect of cured dental resin, the antibacterial activity of prepared resins were measured and compared with each other. At first, for confirming none-releasing behavior of active ingredient, the qualitative antibacterial activity of TM55-BC was evaluated against two common gram positive and gram negative bacterial strains i.e. S. aureus and E. coli, respectively. According to Fig. 11, no inhibition zone was detected around the sample and no bacterial growth was observed on the area under the specimen. Meanwhile, no bacterial colonies attached to sample surface were detected. These observations validated non-realizing the behavior of chemically anchored groups.

4.7.2.

Direct contact test

To find a quantitative evidence regarding possible differences in the behavior of thiol–ene–methacrylate and pure methacrylate systems against bacteria, direct contact test was performed against a dental bacterial strain, S. mutans (Figs. 12 and 13). The result of this test is a strong means of bactericidal activity of immobilized functions. As it was expected the thiol–ene rich system (TM55-BC), showed the highest value of log reduction. In fact this sample could kill 93% of CFU of bacteria. This finding confirms higher accessibility of active immobilized QAS moieties of cured resin. The result of this test is in complete agreement with other recorded data including higher water uptake%, lower Tg and c and more homogeneity of network. According to Fig. 12 there is no significant difference between the antibacterial activity of TM37-BC and M-BC resins (P > 0.05). Although there is some difference between Tg , c and network homogeneity of these samples, there is no significant difference between their water uptake%. It seems the amount of absorbed water has considerable influence in bactericidal activity of this category of material. To further confirm that the observed bactericidal activity is associated with structural characteristics of prepared material, two other antibacterial monomers (DMAEMA-OB and DEAEMA-BC) were prepared and incorporated into the same formulations. Again the same behavior was recorded (Fig. 13). Therefore, it is concluded that regardless of the structure of

antibacterial moieties, the systems with suitable homogeneity and water uptake% have the most favor accessibility of reactive bactericidal functions.

4.8.

Cytotoxicity of samples

Cytotoxicity are known as a big challenge for most cationic carriers, as they can disturb the membrane integrity of cell, decrease the metabolic activity or activate the intracellular signal transduction pathways after interaction with cells [52]. It was reported that high concentration of positively charged groups in polymers resulted in cytotoxic effect [53–55]. However, it is also well recognized that low concentration of charged QAS incorporated in polymers can be helpful for improving the material biocompatibility and rendered them to be safer for clinical applications [56,57,34,58]. Therefore, adjusting the concentration of QAS groups in final networks is critical for maintaining proper cytocompatibility as well as suitable bactericidal activity. In this regard, cytocompatibility of dental resins was evaluated against fibroblast cells through microscopic investigation of cell morphology in vicinity of samples. The results showed that cells kept their spindle shapes and no cell lysis was observed. In fact, none of the samples appeared to give off any toxic or inhibitory leachates, since cells grew to confluence in the vicinity of all samples. Therefore, based on qualitative cell morphology inspection, the level of cytocompatibility of samples with embedded the chosen concentration of embedded QAS containing monomer is acceptable.

5.

Conclusion

In summary, the effect of structural characteristics of the resin matrix on bactericidal and physico-mechanical properties of final crosslinked networks prepared through a ternary thiol–allyl ether–methacrylate system was studied and compared with conventional methacrylate analogous. Thiol–ene containing systems produced more homogeneous structure with lower shrinkage stress and strain in comparison with the classical methacrylate system. Inspection of shrinkage strain rate behavior of specimens, confirms two distinct photopolymerization mechanisms of ternary thiol–allyl ether–methacrylate system with regardless existence of QASM. Photopolymerization of QASM Incorporated thiol–ene rich resins (TM55 based samples) produced low shrinkage, cytocompatible,homogeneous network with adequate water uptake and freedom degree which provided bactericidal functions high available on the matrix surface without sacrificing of the physico-mechanical properties of the cured resin.

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Effect of a novel quaternary ammonium methacrylate polymer (QAMP) on adhesion and antibacterial properties of dental adhesives.

Effect of charge density of bonding agent containing a new quaternary ammonium methacrylate on antibacterial and bonding properties.

Preparation of antibacterial and radio-opaque dental resin with new polymerizable quaternary ammonium monomer.

Antibacterial activity of gemini quaternary ammonium salts.

Antimicrobial polymeric materials with quaternary ammonium and phosphonium salts.

Quaternary ammonium poly(diethylaminoethyl methacrylate) possessing antimicrobial activity.

Antibacterial, physicochemical and mechanical properties of endodontic sealers containing quaternary ammonium polyethylenimine nanoparticles.

Synthesis and Antibacterial Activity of Quaternary Ammonium 4-Deoxypyridoxine Derivatives.

Three-dimensional biofilm properties on dental bonding agent with varying quaternary ammonium charge densities.

Antibacterial activity of bone cement containing quaternary ammonium polyethyleneimine nanoparticles.

Dental materials with antibiofilm properties.

The Use of Quaternary Ammonium to Combat Dental Caries.

Preparation and characterization of quaternary ammonium chitosan hydrogel with significant antibacterial activity.

Performance properties and antibacterial activity of crosslinked films of quaternary ammonium modified starch and poly(vinyl alcohol).

Cytocompatibility and antibacterial properties of capping materials.

quaternary ammonium modified hyperbranched polyglycerol.

Synthesis and characterization of antibacterial polyurethane coatings from quaternary ammonium salts functionalized soybean oil based polyols.

Subcutaneous implants for assessments of dental materials with emphasis on periodontal dressings.

Antimicrobial and Hemolytic Studies of a Series of Polycations Bearing Quaternary Ammonium Moieties: Structural and Topological Effects.

Antibacterial activity of a modified unfilled resin containing a novel polymerizable quaternary ammonium salt MAE-HB.

Aqueous quaternary ammonium compounds and rabies treatment.

In vitro evaluation of antibacterial effect of AH Plus incorporated with quaternary ammonium epoxy silicate against Enterococcus faecalis.

Cellulose-Chitosan-Keratin Composite Materials: Synthesis, Immunological and Antibacterial Properties.

Interaction of ochratoxin A with quaternary ammonium beta-cyclodextrin.