Elution of residual monomers from dental composite materials

Transcription

1 E.B. Tuna*, O. Aktoren*, Y. Oshida**, K. Gencay* *Istanbul University, Faculty of Dentistry Dept of Pedodontics, Istanbul, Turkey **Syracuse University L.C.Smith College of Engineering Dept of Mechanical and Aerospace Engineering, New York, USA Elution of residual monomers from dental composite materials ABSTRACT Aim This study was designed to determine the type and amount of the monomers leached from the different particle sizes of the composite materials. Methods Three different disk sizes (2, 4, 6 mm) prepared for each material group (Filtek Flow, Filtek A110, Filtek P60 and Filtek ) were polymerised by LED and halogen light; the specimens were then placed in artificial saliva. The monomer release in 30 min and 24 hrs from the specimens was analyzed in HPLC calibrated for the monomer extracts before. Results TEGDMA release was detected in all material groups after 30 min and after 24 hrs. BisGMA and BisEMA were not determined in any groups and UDMA was detected only in Filtek. Significant differences in release of TEGDMA and UDMA were obtained between the different sizes of discs. Significantly high amount of TEGDMA and UDMA monomer release was obtained in LED than Halogen groups. Lower amount of monomer release was obtained in species of 30 min than 24 hrs. Conclusion Data has revealed that the monomer release could be detected significantly high from the composite materials polymerized by a lower output curing light device; and higher elution of monomers was determined as the composite thickness has increased. Therefore, the clinical applications of composite materials and the type of curing units have very important effects on the success of restorations and in the decrease of potential side effects. Keywords: Resin composites; Monomers; High performance liquid chromatography (HPLC); TEGDMA; Elution. Introduction During the last few decades, the enthusiasm is great for the tooth-colored resin composites which have been considered as potential substitutes to mercury-containing silver amalgam fillings [Goldberg, 2008]. Light-curing resin composites have been proposed as an alternative to amalgam, ceramic and gold restorations due to advantages including aesthetic and toxicologic 110 considerations [Geurtsen, 1998; Polydorou et al., 2007; Yap et al., 2004]. The elution of components, residual monomers and degradation products from composites has a potential impact both on the biocompatibility and the properties of the materials [Geurtsen, 1998; Spahl et al., 1998]. A high amount of leachable monomers could cause local and systemic mutagenic and estrogenic effects and could also lead to the development of poor mechanical properties such as wear resistance, hardness and tendency to discoloration. Furthermore, the residual monomers gaining access to the pulp via dentinal tubules in cases of insufficient cavity lining could cause pulpal reactions [Munksgaard et al., 2000; Spahl et al., 1998], and unbound monomers are also excellent substrates for the cariogenic bacteria, which contributes to the development of secondary caries [Goldberg, 2008]. Therefore, a higher degree of conversion is desired to minimise the amount of leachable monomers from resin composites. Polymerisation of composite materials with different curing lights and the factors affecting the degree of monomer conversion have been widely investigated. It has been reported that the amount of leachable monomer strongly depends on the light intensity, the wavelengths of photoinitiator system and the duration of the irradiation. The concentrations, types and mixture of photoinitiators, co-initiators, stabilizers, inhibitors and types and proportions of monomers have also been stated as the factors affecting the process of monomer conversion [Muller et al., 1997; Munksgaard et al., 2000; Tanaka et al., 1991]. The mechanisms of cytotoxicity are related firstly to the short-term release of free monomers occurring during the monomer polymer conversion [Goldberg, 2008]. The presence of residual methacrylate monomer and segmental carbon-carbon double bond unsaturation has been blamed for prolonged and increased elution of toxic reactants [Inoue and Hayashi, 1982]. The quantity and composition of the eluted substances are key factors for the potential mutagenic effects of resin composites [Noda et al., 1999]. The purpose of this study is to assess the qualitative and quantitative analysis of extractable residual monomers elutable from various dental composites after halogen and LED (Light Emitting Diodes) curing at different storage times. Materials and methods Composite materials Four light-curing composite resins (3M Dental Products, St. Paul, MN, USA) were chosen for this study. The materials, shades, batch numbers, filler content and load and type of monomers are specified in Table I. Specimen preparation For each composite material were prepared 5 specimens 2, 4, and 6 mm large, and 4 mm thick by cutting them with stainless steel disks. The composites were then placed in the molds using the incremental technique (2 mm), and were polymerised

2 RESIDUAL MONOMERS FROM COMPOSITES Product Brand name Type Shade Filler load description(% wt) Average filler size Filtek Flow Flowable composite A3 Bis-GMA (10-20%), TEGDMA (10-20%), zirconia µm filler (50-70%), silica and dicholorodimethysilane reaction prodeuct (4-9%),dimethacrylate polymer (< 5%) A110 Microfilled composite A3D Bis-GMA (%5), TEGDMA (%5), silika, µm prepolimerize doldurucu P60 Hybrid posterior composite A3 Bis-GMA (<5 %), TEGDMA (< 5%), UDMA µm (3-10%), Bis-EMA( 5-10%), Silane treated ceramic (75-85 %) Filtek Nanofilled composite A3B Bis-GMA (5-15%) TEGDMA (< 5%), nm UDMA (5-15%), Bis-EMA( 1-10%) Silane treated ceramic (65-75%), silan treated silica (5-10%) TABLE 1 - The types and the compositions of the products used in the study. from one side either by LED or halogen light curing. During polymerization all specimens were covered by transparent plastic matrix strips in order to prevent the formation of an oxygen-inhibited surface layer. Polymerization method and storage Polymerization was performed by Halogen-Visilux II and LED-Elipar Freelight I (3M Dental Products, St. Paul, MN, USA) curing lights emitting a light intensity of 500 mw/cm 2 and 400 mw/cm 2, respectively, as measured with a curing radiometer (Model 100, Demetron, Danbury, CT, USA). The curing times for the sample of different thickness are listed in Table 2. Immediately after polymerization, each specimen was removed from the mold and immersed in artificial saliva for 30 minutes and 24 hours. After storage time, the specimens were removed from the storage medium and analyzed in HPLC calibrated for TEGDMA, UDMA, BisGMA and BisEMA monomer extracts. HPLC analysis The analysis was performed by HPLC with a HPLC pump (Perkin Elmer Series 200 Pump), a tunable detector (Perkin Elmer 785 A UV/VIS detector), PE Nelson- Network Chromotograph (Interface NCI 900) with a 20 µl sample loop, a reversed phase C18 analytical column (250 x 4 mm with 5 µ Hypersil PEP 300) at room temperature. The mobile phase was CH3CN (acetonitrile) and H2O at a flow rate of 1 ml/min and detection was performed at a wavelength of 204 nm. The injection volume was 50 µl loop. Known concentrations of TEGDMA (Sigma, 250 ml, Lot 122K3755), Bis-GMA (Sigma Batch # 084K0023) and UDMA (Sigma Aldrich, ) were run as reference standards. The linear fittings of the calibration curves were calculated for the chromatographic peaks at the corresponding retention time (RT) versus monomer concentration. The relationship of concentration with absorbance was plotted using areas under the peak calculated and expressed as a mean (SD) µg/mm 2 (Fig. 1). Statistical analysis The statistical analysis of the results for the amount of eluted monomer for each different group of data were performed by Kruskal Wallis, Dunn s Multiple Comparison and Mann-Whitney-U tests using GraphPad Prisma (V3.0) software at significance level Results The retention times of HPLC peaks of the standard solutions of TEGDMA, UDMA, BisGMA and BisEMA were found as 8.75, 18.73, and min., respectively. Based on the retention time of the monomers, release of TEGDMA was detected in all material groups and UDMA was detected only in the group of Filtek. The monomers eluted ranged from µg/mm 2 for TEGDMA and µg/mm 2 for UDMA. BisGMA and BisEMA were not detected in any of the specimens. The mean values of TEGDMA (µg/mm 2 ) released in 30 min and 24 hrs and the values related to disc diameters are presented in Table 3. The findings have shown that higher monomer release was detected on wider surface area; and lower amount of monomer release was obtained in all species of 30 min than 24 hrs. Significant differences were found in the groups of A110, P60 and cured by LED or Halogen and stored for 30 min or 24 hrs. The released TEGDMA and UDMA monomer values were increased by the storage time in all groups (Table 3). The paired comparisons and significances found between the composite materials in different curing types and storage times for TEGDMA were identified in Table 4. No significant differences in TEGDMA release were Product Layer Curing LED/Halogen thickness (mm) time (sec) Curing device Filtek Flow Standard Filtek A Standard Filtek P Standard Filtek Standard TABLE 2 - Concentration and absorbance peak standard values for TEGDMA and UDMA. 111

3 TUNA E. B. ET AL. FIG. 1 - Concentration and absorbance peak standard values for TEGDMA and UDMA. Material Light curing unit Storage time 2 mm 4 mm 6 mm KW p Flow LED 30 min ± ± ± h ± ± ± Halogen 30 min ± ± ± h ± ± ± A110 LED 30 min ± ± ± h ± ± ± Halogen 30 min ± ± ± * 24 h ± ± ± * P60 LED 30 min ± ± ± h ± ± ± * Halogen 30 min ± ± ± * 24 h ± ± ± LED 30 min ± ± ± * 24 h ± ± ± Halogen 30 min ± ± ± * 24 h ± ± ± *p<0.05 TABLE 3 - The mean TEGDMA values (µg/mm2) (±SD) in each group and the values related to the disc diameters. 30 minutes 24 hours LED Halogen LED Halogen Diameter Diameter Diameter Diameter Materials 2 mm 4 mm 6 mm 2 mm 4 mm 6 mm 2 mm 4 mm 6 mm 2 mm 4 mm 6 mm Flow P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 A110 Flow P > 0.05 P > 0.05 P > 0.05 P > 0.05 P < 0.01 P > 0.05 P > 0.05 P < 0.05 P < 0.01 P > 0.05 P > 0.05 P < 0.01 P60 Flow P > 0.05 P < 0.01 P < 0.01 P > 0.05 P < 0.05 P < P < 0.05 P < 0.01 P < 0.05 P > 0.05 P <0.05 P < 0.05 A110 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P60 A110 P < 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P < 0.05 P < 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P60 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 P > 0.05 Statistical evaluation by Dunn s Multiple Comparison test (p<0.05; p<0.01; p<0.001). Bold values refer to significant differences between the material groups TABLE 4 - The values related to the material types for TEGDMA. 112

4 RESIDUAL MONOMERS FROM COMPOSITES determined between the groups of Flow/A110, A110/P60, and P60/. Elution of TEGDMA from Filtek Flow was found significantly higher than Filtek in disc groups of 4 and 6 mm. The values related to the types of light curing for TEGDMA showed no significant differences in Filtek Flow groups of 2, 4, and 6 mm cured by Halogen or LED; and some significant differences were noted between the other groups (Table 5). UDMA monomer in groups of showed no significances related to types of light curing. Discussion Conventional halogen light-curing units were introduced in the 1970 s and soon became a standard in composite polymerisation [Christensen, 2002; Filipov and Vladimirov, 2006]. The LED offers an extremely long and relatively constant and stable emission of energy stream of the blue spectrum of the visible light. Although thermal emission, depth of cure, degree of conversion and polymerisation efficiency associated with LED and halogen lights have been widely investigated, studies reporting the elution of leachable components from the composites cured with these lights/curing modes are generally limited [Filipov and Vladimirov, 2006; Mills et al., 2002]. The filler type of the composites and the changes in the light energy value per unit area could affect the quality of polymerisation. In this study too, the intensities and exposure time of the light source, the distance between the tip of light source and mold disc have been observed as the factors affecting the low level of monomer-polymer conversion. The released unpolymerized monomers determined in the saliva are the result of inefficient polymerisation. The mechanical and biological properties of dental composites are highly influenced by their monomer to polymer conversion. It has been known that monomer conversion is never complete and the degree of conversion 30 min 24 hrs Material LED/Halogen LED/Halogen Flow 2 mm mm mm A110 2 mm 0.016* 0.05* 4 mm * 6 mm 0.032* 0.016* P60 2 mm mm 0.032* mm * 2 mm mm 0.05* mm 0.036* Statistical evaluation by Mann-Whitney-U test (*p<0.05). TABLE 5 - The values related to the types of light curing for TEGDMA. varies between 35-77% [Ferracane, 1994; Filipov and Vladimirov, 2006; Spahl et al., 1998; Yap et al., 2004]. Under the conditions of this in vitro study, lower amount of monomer release was obtained in species of 30 min immersion than 24 hrs. It has been determined that the all composite materials polymerised by lower output curing light device could cause more TEGDMA release and this device seems insufficient in preventing the high release of monomers. The elution of monomers from the composite resin materials could be dependent on both the material and the curing unit. However, several factors contribute to process of elution from dental composites. According to Ferracane the following play a role [Ferracane, 1990]: the amount of elutable species which is directly related to the extent of the polymerization reaction and number of unreacted monomers; the chemistry of solvent; the size and the chemical composition of the composite materials. Smaller molecules/ions have enhanced mobility and will therefore be eluted faster than larger molecules. TEGDMA molecules being smaller (lower molecular weight) are therefore eluted at a faster rate than larger (higher molecular weight) BisGMA molecules [Thompson et al., 1982]. In parallel, TEGDMA was detected in this study as a main monomer in all material groups and UDMA release only from one material. BisGMA did not leach into the aqueous media. This is consistent with other studies which have also found that TEGDMA is the major monomer eluted from composite resins [Geurtsen, 1998; Moharamzadeh et al., 2007; Tanaka et al., 1991]. In particular, the cytotoxicity of TEGDMA may be of great clinical interest. This relatively hydrophilic comonomer is released by many resin-based materials in considerable amounts [Geurtsen, 1998; Geurtsen, 2000; Spahl et al., 1998]. Due to its high cytotoxic potency, this substance may significantly contribute to adverse local and systemic effects. It may also serve as substrate for polyethylene glycol degrading micro-organisms proliferating within the marginal gap between a filling and cavity wall, causing secondary caries as well as pulp irritations due to production of microbial cytotoxic substances [Qvist, 1990; Spahl et al., 1998]. The liberation of TEGDMA from resin restorations, therefore, should be minimised or prevented [Geurtsen, 2000]. In most studies, release of monomers has been measured as the total release after a certain period of time [Geurtsen, 1998]. Leaching of monomers from a polymeric material in the mouth might be higher than that measured in water, due to enzymatic breakdown of the monomer [Larsen and Munksgaard, 1991], which increases the rate of diffusion. In this study polymerised samples were kept in artificial saliva for simulating mouth environment with higher solubility than water; and the co-monomer TEGDMA with relatively small molecules were determined in all tested material samples. The results of this study support the hypothesis that selected light curing type and their irradiance (light intensity) affect the degree of conversion and monomer release. The power and intensity of the light have effects on the amount of released unpolymerised monomer 113

5 TUNA E. B. ET AL. residue from the composite restorations. In addition, as the composite thickness are increased, the resin restorations need to achieve the optimal levels of degree of conversion. Furthermore, it was determined that the storage times of the composite materials also could reflect more monomer release due to the cumulative accumulation. Conclusion The data of this study demonstrated that various particle and surface sizes of the composite materials, storage times, and types of light curing have effects on monomer release. Thus, higher monomer release could be determined as the surface area and the storage times are increased; and light curing types and intensity could affect on monomer release. Therefore, side potential effects could be prevented and minimised by the optimal polymerisation techniques. In addition, all ingredients of a dental composite should be declared by the manufacturer. Acknowledgements This study was supported by The Research Support Unit of Istanbul University as the project no: T-198/ The corresponding author thanks 3M ESPE, Turkey for supplying the composite materials. References Christensen G. The curing-light dilemma. J Am Dent Assoc 2002;133: Ferracane JL, Condon JR. Rate of elution of leachable components from composite. Dent Mater 1990;6: Ferracane JL. Elution of leachable components from composites. J Oral Rehabil 1994;21: Filipov IA, Vladimirov SB. Residual monomer in a composite resin after lightcuring with different sources, light intensities and spectra of radiation. Braz Dent J 2006;17(1): Geurtsen W. Substances released from dental resin composites and glass ionomer cements. Eur J Oral Sci 1998;106: Geurtsen W. Biocompatibility of resin-modified filling materials. Crit Rev Oral Biol Med 2000;11(3): Goldberg M. In vitro and in vivo studies on the toxicity of dental resin components: a review. Clin Oral Invest 2008;12:1-8. Inoue I, Hayashi I. Residual monomer (Bis-GMA) of composite resins. J Oral Rehab 1982;9: Larsen IB, Munksgaard EC. Effect of human saliva on surface degradation of composite resins. Scand J Dent Res 1991;99: Mills R, Uhl A, Jandt K. Optical power outputs, spectra and dental composite depths of cure, obtained with blue light emitting diode (LED) and halogen light cutting units (LCUs). Br Dent J 2002;193: Moharamzadeh K, Van Noort R, Brook IM, Scutt AM. HPLC analysis of components released from dental composites with different resin compositions using different extraction media. J Mater Sci Mater Med 2007;18(1): Muller H, Olsson S, Soderholm KJ. The effect of comonomer composition, silane heating and filler type on aqueous TEGDMA leachability in model resin composites. Eur J Oral Sci 1997;105: Munksgaard EC, Peutzfeldt A, Asmussen E. Elution of TEGDMA and Bis-GMA from a resin and resin composite cured with halogen or plasma light. Eur J Oral Sci 2000;108: Noda M, Komatsu H, Sano H. HPLC analysis of dental resin composites components. J Biomed Mater Res 1999;47(3): Polydorou O, Trittler R, Hellwig E, Kümmerer K. Elution of monomers from two conventional dental composite materials. Dent Mater 2007;23(12): Qvist, V. Resin restorations: leakage, bacteria, pulp. Endod Dent Traumatol 1993;9:127. Soderholm KJM. Filler leachability during water storage of six composite materials. Scand J Dent Res 1990;98: Spahl W, Budzikiewicz H, GeurtsenW. Determination of leachable components from four commercial dental composites by gas and liquid chromatography/mass spectrometry. J Dent 1998;26: Tanaka K, Taira M, Wakasa K, Yamaki M. Residual monomers (TEGDMA and Bis-GMA) of a set visible-light-cured dental composite resin when immersed in water. J Oral Rehabil 1991;18: Thompson LR, Miller EG, Bowles WH. Leaching of unpolymerized materials from orthodontic bonding resin. J Dent Res 1982;61(8): Yap AU, Han VTS,Soh MS, Siow. Elution of leachable components from composites after LED and Halogen light irradiation. Oper Dent 2004;29(4):