journal of the mechanical behavior of biomedical materials 46 (2015) 83 –92

Available online at

Research Paper

The effect of polymerization mode on monomer conversion, free radical entrapment, and interaction with hydroxyapatite of commercial self-adhesive cements Paulo Henrique Perlatti D’Alpinoa,n, Marı´lia Santos Silvaa, Marcus Vinı´cius Gonc¸alves Vismarab, Vinicius Di Hipo´litoa, Alejandra Hortencia Miranda Gonza´leza, Carlos Frederico de Oliveira Graeffb a

Biomaterials Research Group, School of Dentistry, UNIAN/SP—Universidade Anhanguera de São Paulo, São Paulo, SP, Brazil b DF-FC, UNESP—Universidade Estadual Paulista, POSMAT—Programa de Pós-Graduação em Ciência e Tecnologia de Materiais, Bauru, SP, Brazil

art i cle i nfo

ab st rac t

Article history:

Objectives: This study evaluated the degree of conversion, the free radical entrapment, and

Received 19 November 2014

the chemical interaction of self-adhesive resin cements mixed with pure hydroxyapatite,

Received in revised form

as a function of the polymerization activation mode among a variety of commercial self-

15 February 2015

adhesive cements.

Accepted 19 February 2015

Materials and methods: Four cements (Embrace WetBond, MaxCem Elite, Bifix SE, and RelyX

Available online 26 February 2015

U200) were mixed, combined with hydroxyapatite, dispensed into molds, and distributed


into three groups, according to polymerization protocols: IP (photoactivation for 40 s); DP

Self-adhesive resin cement

(delayed photoactivation, 10 min self-curing plus 40 s light-activated); and CA (chemical


activation, no light exposure). Infrared (IR) spectra were obtained and monomer conversion

Chemical interaction

(%) was calculated by comparing the aliphatic-to-aromatic IR absorption peak ratio before


and after polymerization (n¼10). The free radical entrapment values of the resin cements


were characterized using Electron Paramagnetic Resonance (EPR) and the concentration of

X-ray diffraction

spins (number of spins/mass) calculated (n¼3). Values were compared using two-way ANOVA and Tukey’s post-hoc test (α¼5%). X-ray diffraction (XRD) characterized the crystallinity of hydroxyapatite as a function of the chemical interactions with the resin cements. Results: The tested parameters varied as a function of resin cement and polymerization protocol. Embrace WetBond and RelyX U200 demonstrated dependence on photoactivation (immediate or delayed), whereas MaxCem Elite exhibited dependence on the chemical activation mode. Bifix SE presented the best balance based on the parameters analyzed, irrespective of the activation protocol.

n Correspondence to: Universidade Anhanguera de São Paulo—UNIAN SP, Programa de Mestrado em Biomateriais em Odontologia, Rua Maria Cândida, 1.813 6.1 andar, Bloco G, São Paulo CEP: 02071-013, SP, Brazil. Tel.: þ55 11 29679058/þ55 11 29679077. E-mail address: [email protected] (P.H.P. D'Alpino). 1751-6161/& 2015 Elsevier Ltd. All rights reserved.


journal of the mechanical behavior of biomedical materials 46 (2015) 83 –92

Conclusions: Choice of polymerization protocol affects the degree of conversion, free radical entrapment, and the chemical interaction between hydroxyapatite and self-adhesive resin cement mixtures. & 2015 Elsevier Ltd. All rights reserved.



Resin cements are a class of dental materials of choice for bonding metal, ceramic and indirect composite restorations. These resin-based materials are generally classified as a function of their activation reaction as self-cured (chemically activated), light-cured (photoactivated), or dual-cured (a combination of both activation reaction) (Burrow et al., 1996). Based on interaction with tooth substrates, resin cements can be also classified into three categories: etch-and-rinse, self-etch, and a recently developed group of resin cements known as selfadhesive systems (SLCs) (Carvalho et al., 2004; Pegoraro et al., 2007). The main objective of the introduction of SLCs was to overcome the drawbacks of other types of cements used to join indirect restorations to tooth preparations (Radovic et al., 2008). These materials require no technique-sensitive steps, such as acid etching, priming, or bonding (De Munck et al., 2004). The adhesion strategies employed with SLCs also allow the formation of secondary reactions between the self-adhesive resin and hydroxyapatite, forming chemical bonds (De Munck et al., 2004). This innovative bonding mechanism represents an important characteristic when compared to other resin cements, which are essentially micromechanical in nature (Van Meerbeek et al., 1993). Resin-based cements are also subjected to the oral environment, in spite of the thin layer that results from seating indirect restorations. The polymerization process starts with light exposure or through a self-curing mechanism, if no light is present. Considering this clinical scenario, the polymerization protocol of resin cements is of paramount importance in the subsequent restoration performance (Svizero et al., 2013). It is claimed that the best mechanical properties are achieved when base and catalyst pastes are mixed and immediately photoactivated (Pegoraro et al., 2007). On the other hand, the chemical cure mechanism proceeds slowly and is expected to ensure polymerization in those areas where light is unable to reach (Pereira et al., 2010). As the polymerizing network develops further, the rate of radical propagation eventually becomes limited by diffusion, and the polymerization rate decelerates, providing only a limited conversion, even in the presence of unreacted monomer and free radicals (Halvorson et al., 2002). Conversely, under a delayed photoactivation condition, the cements is chemically activated at first and the polymerization reaction progresses slowly, especially in areas where the curing light is unable to reach the material (Pereira et al., 2010). In this way, a higher end conversion would be expected. Increased degree of conversion has been strongly correlated with improved mechanical properties of resin-based materials (Ferracane and Greener, 1986). Previous studies focused on the influence of different activation protocols in

conventional resin cements (Pereira et al., 2010; Svizero et al., 2013). However, studies are necessary to understand the influence of these protocols for the SLCs that present distinctive formulations, low initial pH, dual-cure polymerization, and the ability to interact with hydroxyapatite. Concerns have been expressed over the chemical composition of the self-adhesive resin cements, specifically regarding the need for balanced formulae due to the polymerization reaction occurs in an acidic environment (Di Hipolito et al., 2012; Vaz et al., 2012). This category of resin cement presents methacrylate monomers that contain phosphoric acid esters that simultaneously demineralize and infiltrate both the smear layer and the underlying dentin, providing micromechanical bonding (Gerth et al., 2006). At the same time, it is important that the pH be neutralized in order to avoid impacting the end conversion, considering the effect of both new methacrylate monomers formulation and the technology to initiate polymerization (Vaz et al., 2012). On the other hand, it has been claimed that a glass ionomer concept was added to the formulation in order to neutralize the initial low pH, which increases from 1 to 6 (Radovic et al., 2008). Thus, the comprehension of the dynamic process in which the demineralization/ monomer permeation process and the polymerization kinetics coexist in this category of material is of paramount importance. Clinically, restorative materials must set in a reasonably short time, in order to be practical, and immediate photoactivation may optimize the initial cement properties necessary to withstand clinical stresses that affect indirect restorations cemented to the remaining tooth structure (Pereira et al., 2010). It is claimed that immediate photoactivation limits the time available for polymer growth and thus inhibits the formation of a rigid polymer network (Neves et al., 2005). On the other hand, the chemical activation protocol is expected to provide the resin cement optimal mechanical properties over time, in areas where polymerization light energy is unable to reach, improving the overall polymerization of the cement at that location (Camilotti et al., 2008; Sideridou et al., 2003). However, different polymerization protocols may affect the resin cements in different ways, depending on a variety of parameters tested (Pereira et al., 2010; Svizero et al., 2013). Some resin cements depend on immediate photoactivation, whereas others seem more dependent on the chemical activation mode. In this way, it is important to evaluate this new category of dual-cure resin cements, considering among others their particular acidic characteristics. Another concern is related to the analysis of the biocompatibility of dental materials, in that the nature and amount of components released during their clinical application are most important. Monomer-release studies are commonly investigated using different solvents such as water, saliva, and organic solvents (Sideridou et al., 2003).

journal of the mechanical behavior of biomedical materials 46 (2015) 83 –92

Considering that SLCs are frequently applied to newly exposed dentin, especially in total crown preparations, and the fact that the polymerization reaction in methacrylate-based resins is radical-mediated, according to a study (Andrzejewska, 2004) to evaluate entrapped free radicals in the resin cements and their interaction with tooth substrates as a function of the activation protocol, is of clinical relevance. Drawing upon three hypotheses, this study attempts to investigate the effects of activation protocols on the degree of conversion, free radical entrapment, and the integrity of the hydroxyapatite crystal structure, when evaluating different SLCs. The following research hypotheses were tested: (1) the degree of conversion will not be negatively influenced by the polymerization activation mode and brand of resin cement; (2) the different activation mode will significantly affect the density of spins of the resin cements tested; and (3) the integrity of the hydroxyapatite (HA) crystal (as monitored in the 211 orientation) will be influenced by the brand of SLCs tested and by the polymerization activation mode used.


Materials and methods


Experimental design

In this in vitro study, the degree of conversion, free radical entrapment, and the integrity of the hydroxyapatite crystalline structure were evaluated on resin cement mixtures incorporating known amounts of hydroxyapatite, according to the following factors: (1) SLCs at four levels: Embrace WetBond, MaxCem Elite, Bifix SE, and RelyX U200; and (2) activation mode at three levels: immediate photoactivation (IP), delayed photoactivation (DP), and chemical activation (CA). The characteristics of the resin cements selected are described in Table 1. Twelve groups were categorized and treated according to the various polymerization protocols (Table 2).


Degree of conversion


Specimen preparation

Specimens were prepared by mixing 0.035 g of dry hydroxyapatite (HA) with 0.25 g of the SLCs. As a starting compound, a HA with a high degree of crystallinity and purity (Ca10(PO4)6(OH)2) was used. The SLCs were mixed according to the manufacturers’ instructions in terms of working and setting times, and then the HA was gently incorporated using manual mixing. The unpolymerized resin cement/HA mixtures were then placed into a Teflon mold (2 mm thick, 6 mm in diameter), positioned over a polyester strip. After filling the mold to excess, the material surface was covered with a Mylar strip and a glass slide and compressed to extrude excess material. The glass slide was then removed, leaving the Mylar strip. The distal end of the curing light guide was placed within 0.1 mm of the top Mylar surface. Specimens of groups IP and DP were photoactivated for 40 s on the top and bottom surfaces. Photoactivation was performed after the working time. After the photoactivation procedures (IP and DP groups), the specimens were removed from the molds and then stored in lightproof recipients for 24 h at room temperature (22 1C). The light-curing procedure was performed using a poly-wave LED


light-curing unit (Bluephase, Ivoclar Vivadent, Schaan, Liechtenstein) having an irradiance of 1200 mW/cm2 s. Prior to testing, and throughout the experiment, the light output was monitored using a handheld radiometer (Model 100, Demetron Research Corp., Danbury, CT, USA). Specimens from the chemically activated group (CA group) were kept in the molds for 24 h at room temperature in the dark prior to their removal. Ten replications were made for each test condition (n¼10).


FT-IR analysis

The specimens were placed and vertical pressed against a horizontal diamond element of an attenuated total reflectance (ATR) unit (MIRacle, PIKE Technologies, Madison, USA) of a Fourier transform infrared spectrometer (IRPrestige-21, Shimadzu, Tokyo, Japan) in order to obtain the infrared (IR) spectrum. IR spectra were collected between 1680 and 1500 cm  1 using 16 scans at 2 cm  1 resolution. The specimens were placed one at a time in the sample holder of the device, and spectra were recorded. A small amount of uncured resin cement from each material was also scanned and its spectrum was used as unpolymerized reference. The ratio between the peak absorbance value of the aliphatic (C¼C) at 1636 cm  1 to the aromatic C¼ C group absorption at 1608 cm  1 in the cured and uncured states was used to calculate the degree of conversion of aliphatic (C ¼ C) into (C–C). The results were expressed as the percentage of aliphatic C¼ C remaining after light exposure to that available in the unpolymerized condition (Rueggeberg et al., 1990). Average conversion values were calculated, and a two-way ANOVA was performed. Tukey’s post-hoc test was used as a multiple comparison test, at a pre-set alpha of 0.05.


Magnetic properties


Specimen preparation

Specimens were prepared for Electron Paramagnetic Resonance (EPR) analysis in the same way as previously described, except that the unpolymerized resin cement/HA mixtures were placed into a rectangular Teflon mold (8 mm  2 mm  2 mm). In addition, three replicas were made for each test condition (n¼3).


EPR analysis

Spectra were acquired using an EPR instrument (Magnettech Miniscope MS300, Berlin, Germany) operating at X-Band  9.43 GHz. The main EPR parameters were modulation amplitude (from 0.2 to 0.4 mT) and microwave power (0.5012 mW). The g-factors were determined by reference to the signal of the diphenylpicrylhydrazyl (DPPH) with a known value of 2.0036. The density of spins was determined by comparing the EPR spectrum of the unknown specimens (cements) to that of a sample with a known spin concentration (standard sample). Each free radical has a spin different from zero, so by measuring the spin density one is measuring the free radical density (Poole and Charles, 1983). For that standard, an amorphous silicon sample with a calibrated spin concentration of 1.13  1015 spins/ grams was used (D’Alpino et al., 2014). The number of spins in the resin cements of each experimental group was then


journal of the mechanical behavior of biomedical materials 46 (2015) 83 –92

Table 1 – Materials used in this study. Material

Lot #/expiration date

Embrace WetBond Pulpdent Corporation, Watertown, MA, USA


130711 2015-07

Co-monomers (mono-, di-, and tri-functional methacrylate monomers, Barium, glass, ytterbium trifluoride, inert minerals. Automix system

MaxCem Elite Kerr Corporation, Orange, CA, USA

5011290 2015-03

GPDM, co-monomers (mono-, di-, and trifunctional methacrylate monomers, water, acetone, and ethanol. Inert minerals and ytterbium fluoride. Automix system

Bifix SE Voco GmbH, Cuxhaven, Germany

1322421 2014-12

Bis-GMA, UDMA, Gly-DMA, phosphate monomers, initiators, stabilizers. Glass. Automix system

RelyX U200 3M ESPE, St. Paul, MN, USA

132950065 9 2014-12

Base: Methacrylate monomers containing phosphoric acid groups, methacrylate monomers, initiators, stabilizers, rheological additives Catalyst: Methacrylate monomers, alkaline fillers, silanated fillers, initiator components, stabilizers, pigments, rheological additives. Zirconia/silica fillers. Clicker delivery system

Working time (min)

Setting time (min)

Exposure duration (s)




36.6 39.0



10 - 20

69.9 59.0



10 - 20

70.0 45.0




72.0 43.0

Fillercontent W (%) V (%)

Abbreviations: Bis-GMA: bisphenol A diglycidyl ether dimethacrylate; UDMA: urethane dimethacrylate; Gly-DMA: glycerol dimethacrylate; GPDM: glycero-phosphate dimethacrylate. All information supplied by manufacturers.

Table 2 – Experimental groups and curing protocols descriptions. Polymerization mode used

Group abbreviation

Curing protocol description

Immediate Photoactivation Delayed photoactivation Chemical activation

IP (control) DP CA

Photoactivation only for 40 s 10 min delay for chemical curing, followed by photoactivation for 40 s Chemical activation only

calculated using the following formula (Poole and Charles, 1983; Warren and Fitzgerald, 1977): N¼

CT ffiffiffi p 2 PA


performed using a two-way analysis of variance (ANOVA) followed by Tukey’s post-hoc test at a pre-set alpha of 0.05.


where N is the spin concentration (in spins); C is the standard sample constant (1.13  1015); T is the temperature (in Kelvin); A is the area under the resonance absorption curve; AM is the modulation amplitude; and P is the incident microwave power. For all measurements, the microwave power and modulation amplitude were kept constant to avoid signal saturation and distortion. The identification of the free radicals was made with the assistance of software (Easyspin 4.5.1). The ESR parameters were obtained by fitting the experimental data. The spin concentration as a function of mass according to the experimental groups was calculated dividing the number of spins by the sample mass (in grams). The radicals were then calculated as a function of the resin content. The concentration in the resin portion of each product (by mass) was obtained by thermal analysis (data not shown). Statistical analysis was


Chemical interaction with hydroxyapatite


Preparation of hydroxyapatite (HA)/SLC samples

Samples were prepared for X-ray diffraction analysis (XRD) by mixing 0.070 g of dry HA powder (Ca10(PO4)6(OH)2) with 0.5 g of the freshly mixed but unset SLCs. The materials were manipulated according to the manufacturers’ instructions and were then gently mixed with HA on a glass plate until a paste-like consistency was obtained. Then, the paste was transferred and carefully spread onto an XRD specimen holder. A homogeneous layer in which the HA was uniformly distributed throughout the organic matrix of the resin cements was obtained in all cases. The specimens were then polymerized according to the activation protocols. The dry HA power was also characterized for comparative analyses. A different ratio was used here in order to fill out the specimen holder to obtain the XRD patterns.

journal of the mechanical behavior of biomedical materials 46 (2015) 83 –92


XRD analysis

XRD patterns were collected using an X-ray diffractometer (DMAX Ultimaþ Rigaku International Corporation, Tokyo, Japan) with CuKα radiation, operating at 40 kV and 20 mA. Scans were performed from 51 to 801 (2θ) at a step size of 0.021 and a scan speed of 21/min. Qualitative phase analysis was carried out by using the Joint Committee on Powder Diffraction—International Center for Diffraction Data (JCPDS–ICDD) databases by comparing the peak intensity of pure HA with that of the test specimens. In addition, a quantitative phase analysis was also performed by comparing the peak intensity of HA attributed to the 211 diffraction (highest peak of crystallinity of pure HA) before and after mixing HA with the SLCs. The results were expressed in peak intensity of the mixture resin cement/HA (in cps) and also in the percent decrease in crystallinity of HA using the strongest diffraction orientation (2 1 1) as a guide (1730 cps), in comparison to that of pure HA.




FT-IR analysis

Experimental results are displayed in Table 3. The degree of conversion ranged between 16.5% for RelyX U200 to 59.6% for MaxCem Elite, when both cements were chemically activated. When the cements were compared, significantly lower conversion averages were observed for Bifix SE when immediately photoactivated, and for RelyX U200 when chemically activated or photoactivated after 10 min (po0.05). On the contrary, MaxCem Elite presented significantly higher degree of conversion averages, irrespective of the activation protocol (po0.05). In terms of conversion, both Embrace WetBond and RelyX U200 demonstrated dependence on the photoactivation (immediate or delayed), whereas MaxCem Elite exhibited significantly higher conversion averages when chemically activated. On the other hand, Bifix SE presented significantly higher conversion averages when chemically activated and for delayed photoactivation over those seen in immediate photoactivation.


Free radical entrapment

The panels in Fig. 1 display characteristic electron paramagnetic resonance spectra of the SLCs. The EPR spectra are characterized by 9 lines. For the propagating radical g¼ 2.00370.001 and isotropic hyperfine constant a is 21.570.1 G for CH3 and 11.770.1 G for CH2.


For the allylic radical g¼ 2.00370.001 and a is 21.570.1 G related to CH2. The calculated number of spins of the experimental groups, normalized by weight (resin content), is presented in Table 4. The spin concentration was found to be between 3.6  1016 spins/g for MaxCem Elite (CA), and 1,560  1018 spins/g for RelyX U200 (IP). Embrace WetBond and RelyX U200 presented significantly higher spin concentration averages, when photoactivated immediately or after 10 min (po0.05). MaxCem Elite presented significantly higher averages of spin concentration when photoactivated after 10 min. The same was observed for Bifix SE. All of the materials tested presented significantly lower spin concentrations when the chemical activation was performed (po0.05).


XRD analysis

The panels in Fig. 2 displays the X-ray diffraction patterns of SLCs tested. In each diffractogram, the crystalline peaks of pure HA, as well as those of the resin cements/HA mixtures, are displayed. The peaks and associated planes of HA diffractograms match the crystallographic data sheet (74-0565) of calcium hydroxide phosphate, Ca10(PO4)6(OH)2, obtained from the PCPDFWIN database. In accordance with this database, the crystalline pattern has a hexagonal cell with the following parameters: a¼9.424 Å and c¼ 6.879 Å. With respect to the HA/ resin cement interactions, X-ray diffraction patterns indicate the presence of a hexagonal HA phase and an organic, amorphous phase (seen as a broad hump in the baseline) at 2θ between 101 and 301, associated with Embrace WetBond (Fig. 2A). Similar findings were observed in the diffractograms of the mixture Bifix SE/HA (Fig. 2C), irrespective of the activation protocol. The diffractograms indicating the interaction between HA and RelyX U200 (Fig. 2D) show the presence of peaks related to both Ca10(PO4)6(OH)2 and an unidentified crystalline component at 2θ¼ 17.91. In the diffractograms of the mixture of MaxCem Elite and HA, two crystalline fractions are present: one related to HA and another related to ytterbium fluoride (YbF3). These additional peaks in the diffractograms of the MaxCem Elite/HA mixture were attributed to the presence of the orthorhombic crystalline phase of YbF3. The peaks and the crystalline planes associated match the crystallographic data sheet of ytterbium fluoride (711161). Table 5 displays the results of peak intensity of the resin cement/HA mixture (in cps) and also in terms of percentage reduction in comparison to that of pure HA at (2 1 1) diffraction (1730 cps). The greatest reductions in peak intensity were obtained when Bifix SE was mixed to HA, irrespective of the activation protocol. The percentages varied from 61.8 (DP group) to 69.2 (CA group). On the other hand, the lowest reductions were

Table 3 – Percent monomer conversion (%) (sd) of products tested using various polymerization protocols. PRODUCT

Immediate photoactivation (IP)

Delayed photoactivation (DP)

Chemical activation (CA)

Embrace WetBond MaxCem Elite Bifix SE RelyX U200

32.3 52.3 27.3 28.3

31.7 52.8 39.9 25.4

27.5 59.6 40.8 16.5

(1.8) (1,5) (1.7) (3.9)

a,B b,A b,C a,B

(0.8) (3.1) (1.6) (1.0)

a,B b,A a,B a,C

(n ¼10). Groups identified using similar letters (upper case for columns and lower case for rows) are not significant (p40.05).

(3.0) (1.6) (1.8) (1.5)

b,C a,A a,B b,D


journal of the mechanical behavior of biomedical materials 46 (2015) 83 –92

obtained for RelyX U200 when photoactivated immediately (IP group) or after 10 min (DP group), in which the percentages were 42.7% and 49.7%, respectively. For the CA group, MaxCem Elite demonstrated the lowest reduction of the peak intensity (46.5%).


Fig. 1 – Characteristic X-band electron paramagnetic resonance spectra after 24 h of the self-adhesive resin cements according to polymerization activation protocol.


The results of the present study indicate that, regarding the degree of conversion, that both Embrace WetBond and RelyX U200 cements depend mostly on the photoactivation (immediate or delayed), whereas MaxCem Elite exhibits a higher dependence of the chemical activation mode. In addition, Bifix SE exhibited dependence on chemical activation and the time for photoactivation (delay of 10 min). Thus, the first research hypothesis, which anticipated that the degree of conversion of a given SLC product among the different polymerization modes would not be negatively influenced, was partially upheld by the experimental data. It can be argued that the degree of conversion significantly varied among the experimental groups and that, for some of them, the percentages were rather low. As described in the FTIR methodology, the degree of conversion of aliphatic C¼C into C–C for each specimen was calculated. The results were expressed as the percentage of aliphatic C¼C remaining after polymerization to that available in the unpolymerized condition. In this way, it would be expected that the materials were equally compared using this methodology if the resin cements presented a chemistry solely based in Bis-GMA. In analyzing the material composition (Table 1), it can be noted that the manufacturer provides no specific information. The description includes mono-, di-, and tri-functional methacrylate monomers, phosphate monomers, and others. In addition, the filler content in this type of restorative material comprises an extremely important characteristic, determining the relative viscosity of the luting cement and thus affecting polymer chain mobility and the ability of a growing species to propagate. Concerns have been previously raised as to whether the SLCs can be successfully used in clinical applications because of the low percentages of conversion in self-curing mode in the case of light attenuation (Vrochari et al., 2009). On the other hand, it is important to mention that there may be an additional glass ionomer-type reaction occurring in the RelyX product at the same time as the free radical polymerization (Nakamura et al., 2010). Thus, even if low conversion values are seen, the testing performed does not monitor other types of setting reactions that might also contribute to the overall properties, and thus the potential clinical success, of these products. The weight content of filler particles varies between 60 and 75% in SLCs (Belli et al., 2009). It is suggested that a practical limit to the amount of filler particles in resin cement formulations must be far less than the densest pack limit possible (Darwell, 2009). Considering the same resin cement volume, very high filler packing densities can be achieved with an appropriate selection of the particle size and proportions. In this way, the filler packing in resin cements differs from that in the dental composite resins, where lower filler volumes are normally used. In the present

journal of the mechanical behavior of biomedical materials 46 (2015) 83 –92


Table 4 – Mean entrapped free radical concentrations normalized by weight (sd)*, with respect to brand of self-adhesive resin cements and polymerization activation mode. PRODUCT

Immediate Photoactivation (IP)

Delayed Photoactivation (DP)

Chemical Activation (CA)

Embrace WetBond MaxCem Elite Bifix SE RelyX U200

231 (2.4) a,C 21 (0.2) b,D 494 (2.9) b,B 1560 (15.4) a,A

281 (3.1) a,C 66 (0.3) a,D 594 (1.2) a,B 1400 (14.8) a,A

63 (2.2) b,D 3.6 (0.1) c,C 318 (0.9) c,A 159 (3.2) b,B

(n ¼3). Values denoted using similar letters (upper case for columns and lower case for rows) are not significantly different (p40.05). n (  1016 spins/g).

study, the volumetric filler content varied from 39.0 to 59.0%, whereas the variation in weight content was between 39.9 and 72.0%. During polymerization, portion of the methacrylate groups involved in the formation of the cross-linked matrix remains unreacted, especially in the case of higher molecular weight monomers (D’Alpino et al., 2007). This fact, associated with high filler levels decreases polymer radical mobility, and as a consequence, there is a decrease in its reactivity: it tends to become entrapped within the viscous, cured polymer network and is incapable of causing additional chain lengthening or crosslinking. The products with higher filler loading (such as RelyX U200) react more slowly, and this fact may explain why this resin cement exhibited a significantly lower degree of monomer conversion and a dependence on the delay time prior to photoactivation (Pereira et al., 2010). Previous studies recommended immediate photoactivation for SLCs, solely based on the degree of conversion parameter (Aguiar et al., 2010; Arrais et al., 2014). In dualcured cements, both light and chemical activation procedures generate reactive free radicals by cleavage of the initiator molecules (Rueggeberg, 1999). In the propagation reaction, these radicals are able to add to the double bonds of methacrylate groups, thereby creating new radicals. This free radical polymerization reaction occurs very rapidly, until no additional monomer is able to react or the propagation reaction is stopped by termination (Truffier-Boutry et al., 2003a). The mobility of the developing polymer chains becomes progressively more restricted, due to the increase in the viscosity, the reduction in free volume, formation of microgels, and chain entanglement (Neves et al., 2005). In this way, the polymer network becomes rigid enough, its chains become essentially immobile, and the propagation of free radical reactions becomes diffusion limited, decreasing the overall conversion value (Andrzejewska, 2004). This change causes a significant percentage of the methacrylate groups (between 25 and 60%) to remain unreacted (Cook et al., 1999). In resin-based restorative materials, mostly bifunctional monomers are used and are able to react with four other monomer molecules leading to the formation of polymer chains and cross-linking. Although the network of a composite includes not only the remaining double bonds but also free radicals, which are located at functional groups that have reacted with only one methacrylate group, it is expected that cross-linked polymers present lower free radical densities. When the degree of conversion is the only parameter assessed, these radical-mediated reaction issues are not taken into consideration, because only the

percentage of the monomers that convert into polymers is calculated. EPR spectroscopy is a very useful technique for studying the structure of free radicals and for following the radical molecule through various physically or chemically modified pathways (Poole and Charles, 1983). In addition to qualitative measurements, EPR is also quantitative. The absorption of microwave energy in a sample is directly proportional to the number of spins present. The area under the EPR absorption spectrum is proportional to the number of free radicals (spins) in the sample, and the number of free radicals is proportional to the radiation energy absorbed in the sample. Thus, the radiation dose may be determined by measuring the relative concentration of radiation-generated free radicals in a given material using EPR spectroscopy (Sagstuen and Hole, 2009). Free radicals are created during the photoactivation of resinbased materials (Leprince et al., 2009). Some radicals terminate by encountering other radical species and others become trapped by vitrification of the system (Truffier-Boutry et al., 2003a). EPR spectroscopy is an analytical tool which applies to molecular systems having either an odd number of electrons (free radicals) or more unpaired electrons. Two types of trapped free radicals are detectable by EPR in polymerized composites: “propagating” and allylic radical types (Truffier-Boutry et al., 2003b). The former gives rise to an EPR spectrum with nine weak and broad peaks, and the latter (allylic) to five strong and sharp peaks. Both spectra superimpose, resulting in a “nine-line spectrum” with peaks of alternating intensities (Leprince et al., 2009). As free radicals can be trapped in a solid matrix, they are unable to recombine and the nine-line EPR spectrum persists during a period of variable length, from a couple of days to many months, depending on the storage conditions (Leprince et al., 2009). It has been previously established that the degree of conversion is related to the number of unpolymerized terminal groups in which double bonds are present (Rueggeberg, 1999). On the other hand, the conversion reaction proceeds through the formation of radicals under irradiant light, which corresponds to the EPR spectrum. In this way, an increase in the EPR signal intensity as a function of irradiation time correlates to the propagation of the conversion reaction (Leprince et al., 2009). In this way, recombination of radicals can occur, either by bimolecular termination or due to reactive diffusion, resulting in an increase of the degree of conversion was observed 24 h after the photoactivation procedure (2%) (Leprince et al., 2009). The various SLCs differ


journal of the mechanical behavior of biomedical materials 46 (2015) 83 –92

Fig. 2 – 24-h X-ray diffraction patterns of the self-adhesive resin cements: (a) Immediate photoactivation; (b) delayed photoactivation; (c) Chemical activation. n ¼crystalline peaks of ytterbium fluoride. Δ means unidentified crystalline peak at 2θ ¼17.91.

from each other in the speed of propagation of the conversion reaction, probably because of differences in the composition of the organic matrix, which is also known to influence in the degree of conversion (Rueggeberg, 1999). Tooth preparation for indirect bonded restorations (i.e., composite/ceramic inlays, onlays, and veneers) can generate significant dentin exposure (Di Hipólito et al., 2014). Considering that self-adhesive luting cements are used to cement indirect restorations in freshly cut dentin surfaces and that no adhesive system is required, the presence of trapped free radicals may cause pulpal damage, as these compounds can be released by the action of substances in the oral environment such as water or saliva (Sideridou et al., 2003). In the present study, the spectra of the resin cements follow what is found in the literature: lower-intensity EPR signals were observed, so there were significantly fewer entrapped free radicals when the chemical activation protocol was used, irrespective of the material tested. In addition, the spin concentrations varied among the materials tested. RelyX U200 presented significantly higher free radical entrapment when photoactivated immediately or after 10 min. Thus, the second hypothesis, in which it was speculated that different activation protocols would affect the density of spins or free radicals of the self-adhesive resin cements, was proven valid. The present study also evaluated the influence of SLCs on the integrity of the crystalline structure of pure hydroxyapatite, with which these materials were mixed. For all SLCs, it is presumed that free radicals created in an acidic environment demineralize the pure hydroxyapatite powder, which was somehow “altered” when mixed with the different resin cements tested. This effect was demonstrated by changes in the peak intensities in the XRD diffractograms. In this case, the integrity of the hydroxyapatite crystalline structure was evaluated. In the present study it was advocated that if the 211 value decreases, then there is a concomitant physical alteration of the HA crystal. This does not infer a chemical bonding to HA, only that the atomic arrangements of the atoms making up a hydroxyapatite unit cell have been changed. It is important to highlight that the 211 orientation represents the peak with the highest value, so changes in this orientation were more evident. The highest reductions were observed for Bifix SE/HA, irrespective of the activation protocol. On the other hand, the lowest reductions were observed in RelyX U200 when immediately photoactivated (IP group) as well as after 10 min (DP group), in which the percent reductions were 42.7% and 49.7%, respectively. For the CA group, MaxCem Elite has the lowest reduction in the 211 peak intensity (46.5%). Thus, the third hypothesis, which stated that the integrity of the hydroxyapatite crystal would be reduced by the brand of SLCs tested and by the polymerization activation mode used, was accepted. The effect of SLC in the modification of HA crystallinity differed depending on photoactivation protocol as well as on the resin cement tested. These changes in HA crystallinity are representative of the power of SLCs to demineralize remaining tooth tissues and to permeate these tissues. The acidic monomers incorporated in the SLCs are not strong enough to etch through smear layers to form a “traditional” hybrid layer along the resin-tooth interface, with the absence of deeply penetrating resin tags into the

journal of the mechanical behavior of biomedical materials 46 (2015) 83 –92


Table 5 – Quantitative analysis of intensity peak of HA attributed to the (2 1 1) diffraction after mixing HA with the selfadhesive resin cements. (One specimen tested per group). PRODUCT

Embrace WetBond MaxCem Elite Bifix SE RelyX U200

Immediate photoactivation (IP)

Delayed photoactivation (DP)

Chemical activation (CA)

Peak height (AU)

% Reduction

Peak height (AU)

% Reduction

Peak height (AU)

% Reduction

667 790 546 992

61.4 54.3 68.4 42.7

798 843 660 870

53.9 51.3 61.8 49.7

812 926 532 792

53.1 46.5 69.2 54.2

Reference: intensity peak of pure hydroxyapatite ¼ 1730 cps.

dentinal tubules (Vaz et al., 2012). In addition, it was also proven that this adhesion strategy relies not only on micromechanical retention, but also on chemical interactions between monomer acidic groups and hydroxyapatite (Gerth et al., 2006). According to the results of the present study, it can be speculated that the dynamic process, in which the acidity of the cements is progressively neutralized, as well as the polymerization process, is influenced by the SLCs and by the activation protocol. These events start immediately after mixing the base and catalytic pastes, and due to their rheological properties, the cements are able to flow according each material. Therefore, it is of paramount importance to compare the results in the light of the parameters analyzed. In general, the resin cement Bifix SE presented higher dependence on the chemical activation, in which higher conversion averages, lower free radical density, and higher reduction in HA crystallinity are observed. Although the cement MaxCem Elite also demonstrated a chemical dependence in terms of conversion efficiency and lower free radical density, this product interacted less with the HA when chemically activated. In this case, MaxCem Elite was more effective when immediately photoactivated or after a delay of 10 min. On the other hand, the cement RelyX U200, as previously discussed, showed a higher dependence on delay time prior to photoactivation in terms of conversion efficiency. Conversely, the EPR and XRD analyzes showed a higher dependence on chemical activation, considering the significantly lower spin (free radical) density and higher interaction of the resin with HA. Although a higher dependence on the chemical activation in terms of free radical concentration was observed, the cement Embrace WetBond showed higher dependence on the photoactivation, in terms of degree of conversion and higher interaction with HA. Thus, the cement Bifix SE presented the best balanced formulation based on the parameters analyzed, demonstrating higher effectiveness to demineralize hydroxyapatite, a higher degree of conversion, and lower density of free radicals, irrespective of the activation protocol. This study investigated the influence of the different activation protocols on the degree of conversion, free radical density, and changes in HA crystallinity when SLCs were mixed to pure hydroxyapatite. It was clearly demonstrated that both factors (resin cement and activation protocol) influenced the results, based on the different parameters evaluated. Clinically, restoration longevity depends on the numerous steps before a restorative process is completed. The simplification of clinical steps in using materials is critical to the success of a

restorative procedure, because many aspects need to be considered. For the development of future SLCs, studies are necessary to evaluate the longevity of indirect restorations cemented with this category of resin cement over longer evaluation times.



Within the limitations of this study, the following conclusions can be made: 1. Choice of polymerization protocols affects the degree of conversion of SLCs; 2. Entrapped, unreacted free radical density of SLCs varies as a function of the resin cement and of the activation; 3. Brand of SLC and its polymerization activation mode can alter its interaction with the crystalline structure of hydroxyapatite; and 4. Based on the parameters evaluated, one of the tested products provided more optimal characteristics than did the others: improved degree of conversion, lower density of free radicals, and higher interaction with hydroxyapatite, regardless of the activation mode.

Acknowledgements This study was partially supported by grants from CNPq nos. 479744/2010-6 and 163102/2011-2 (P.I. Paulo H. P. D’Alpino) This study was developed as partial fulfillment of the requirements of Dr. Silva’s Master degree (UNIAN—SP). The authors are grateful to Prof. Dayse Iara dos Santos (UNESP— Bauru) for the technical support in the XRD analysis.

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The effect of polymerization mode on monomer conversion, free radical entrapment, and interaction with hydroxyapatite of commercial self-adhesive cements.

This study evaluated the degree of conversion, the free radical entrapment, and the chemical interaction of self-adhesive resin cements mixed with pur...
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