Evaluation of the physicochemical properties of silicone-and epoxy resin-based root canal sealers

To assess the physicochemical properties of AH Plus, GuttaFlow 2, GuttaFlow BioSeal, and MM Seal, five samples of each root canal sealer were evaluated to determine their setting time (ST), dimensional change (DC), solubility (SL), flow (FL), and radiopacity (RD) according to American National Standards Institute/American Dental Association (ANSI/ADA) Specification 57. The distilled and deionized water obtained from the SL test were subjected to atomic absorption spectrometry to observe the presence of Ca2+, K+, and Na+ ions. Statistical analysis was performed by using one-way ANOVA and Tukey–Kramer tests (p < 0.05). The following results were obtained: ST (min) (AH Plus 463.6 ± 13.22; GuttaFlow 2 24.35 ± 2.78; GuttaFlow Bioseal 17.4 ± 0.55; MM Seal 47.60 ± 4.39), DC (%) (AH Plus 0.06 ± 0.12; GuttaFlow 2 −26.06 ± 1.24; GuttaFlow Bioseal 2.10 ± 1.47; MM Seal 8.47 ± 2.41), SL (%) (AH Plus 0.41 ± 0.21; GuttaFlow 2 5.13 ± 4.11; GuttaFlow Bioseal 3.03 ± 1.05; MM Seal 0.94 ± 0.17), FL (mm) (AH Plus 36.42 ± 0.40; GuttaFlow 2 36.44 ± 0.05; GuttaFlow Bioseal 35.4 ± 0.03; MM Seal 52.75 ± 0.60), and RD (mmAl) (AH Plus 7.52 ± 1.59; GuttaFlow 2 6.85 ± 0.14; GuttaFlow Bioseal 7.02 ± 0.18; MM Seal 3.32 ± 0.90). ST, DC, SL, FL, and RD showed statistical differences among the root canal sealers (p < 0.05). As AH Plus showed the lowest DC and SL values (p < 0.05), the findings indicate that this sample is the only sealer conforming to ANSI/ADA standards.


Introduction
The goal of endodontic treatment is to promote proper cleaning, shaping, and obturation of the root canal system (RCS); these are interdependent procedures that must be carefully performed so that successful treatment can be achieved 1 .Three-dimensional sealing is necessary in the filling of root canals to prevent bacterial microleakage 2 , thereby avoiding possible reinfection, and create a favorable environment for the repair of periapical tissues. 1,2,3lthough gutta-percha combined with the sealer AH Plus remain the most widely used materials in the filling procedure; 1,3,4,5,6,7,8,9 numerous techniques and materials with different physicochemical and biological properties have also been developed. 2,3,8,9,10oot canal sealers can be classified according to their composition as zinc oxide and eugenol-based sealers; sealers containing calcium hydroxide; glass ionomer, epoxy resin, and methacrylate resin-based sealers; or bioceramic and silicone-based sealers 1,3,[7][8][9] .
In 2012, GuttaFlow 2 (Coltene/Whaledent, Langenau, Germany) was launched in the market to replace the sealer GuttaFlow.This sealer was formulated with polydimethylsiloxane, gutta-percha particulates (smaller than 30 μm), and nano silver particles. 14,17,18GuttaFlow 2 is supplied in pre-mixed syringes or capsules to improve the consistency of the mixture and facilitate its use; according to its manufacturer, this sealer has an SL close to zero.
GuttaFlow Bioseal sealer (Coltene/Whaledent) was subsequently introduced to provide tissue repair along with obturation.It has the same formulation as the GuttaFlow sealer but also includes calcium silicate, which, upon contact with biological tissues, releases natural repair constituents and aids in the regeneration of periapical tissues. 17,18,19o analyze the physicochemical properties of root canal sealers, the American National Standards Institute/American Dental Association (ANSI/ADA) approved Specification 57, 20 which proposed the carrying out FL, film thickness, setting time (ST), RD, ST, and dimensional change (DC) tests on dental sealers.In 2007, Carvalho-Junior et al., 4 considering the standards determined by Specification 57, suggested reducing the volume of the sealer needed for the SL and dimensional alteration tests to 80%.They also suggested complementary evaluation of the resulting liquid by atomic absorption spectrometry to analyze the ions released in the ST test. 4s the development of new materials, such as silicone-based sealers and their various compositions, offers the prospect of improving the quality of the resulting fillings, the objective of the present work is to evaluate the physicochemical properties of GuttaFlow 2 and GuttaFlow Bioseal (silicone-based sealers) as well as MM Seal and AH Plus (epoxy resinbased sealers) according to ANSI/ADA Specification 57 with the modifications proposed by Carvalho-Júnior et al. 4 Here the ST, DC, SL, FL, and RD of the sealers were studied

Methodology
To accomplish this study, the following endodontic sealers were used: GuttaFlow 2, GuttaFlow Bioseal, MM Seal, and AH Plus.The tested materials were manipulated according to the manufacturer's instructions.The environment in which the experiments were performed featured a temperature of 23 ± 2°C and relative air humidity (RH) of 95% ± 5%, as recommended by ANSI/ADA Specification 57.The methodology used was proposed by the ANSI/ ADA and followed by several authors, including Resende et al., 21 Flores et al., 3 and Borges et al. 7 Detailed information (material, manufacturer and composition) of the tested sealers is shown in Table 1.

Setting time
Five plaster of Paris cast rings with an internal diameter of 10 mm and a thickness of 2 mm were prepared.The external borders of the molds were fixed with wax on a glass plate.The sealers were manipulated according to the manufacturer's instructions, and the molds were filled with the material and transferred to a chamber with 95% RH and temperature of 37°C.About 150 ± 10 s after the start of mixing, a Gilmore-type needle with a mass of 100 ± 0.5 g and a flat end of 2.0 ± 0.1 mm in diameter was carefully lowered vertically into the horizontal surface of each sample.The needle tip was cleaned and probing was repeated until the indentations formed ceased to be visible.If the results differed by more than ±5%, the test was repeated.

Dimensional change
Five Teflon (polytetrafluroethylene, DuPont, HABIA, Knivsta, Sweden) molds prepared for the production of 3.58 mm-high cylindrical test bodies measuring 3 mm in diameter were placed on a glass plate wrapped with a fine cellophane sheet.The molds were filled with a slight excess of freshly mixed sealer, and a microscope slide, also wrapped in cellophane, was pressed onto the upper surface of the mold.The assembled group was kept firmly joined by a C-shaped clamp, transferred to an incubator (37°C, 95% RH), and left to stand for a period corresponding to three times the ST.Afterward, the flat end of the molds containing the samples were ground with 600 grit wet sandpaper.The samples were removed from the mold, measured with a digital caliper, stored in a 50 mL vessel containing 2.24 mL of deionized distilled water, and kept in an incubator (37°C, 95% RH) for 30 days.The sample was then removed from the container, blotted dry with absorbent paper, and measured again for length.The percentage of the dimensional alterations was calculated using the formula: [(L 30 -L) / L] × 100 where L 30 is the length of the sample after 30 days of storage and L is the initial length of the sample.

Solubility
For the SL tests, 10 circular 1.5 mm-thick Teflon molds measuring 7.75 mm in inner diameter were filled with freshly mixed sealer.Each mold was supported by a larger glass plate and covered with a cellophane sheet.An impermeable nylon thread was placed inside the material, and another glass plate, also covered with cellophane film, was positioned on the mold and pressed manually in such a way that the plates touched the entire mold in a uniform manner.The assembly was placed in an incubator (37°C, 95% RH) and left to stand for a period corresponding to three times the ST.As soon as the samples were removed from the mold, they were weighed three times each (HM-200, A&D Engineering, Inc., Bradford, MA, USA), and the mean reading was recorded.The samples were suspended by nylon thread and placed two-by-two inside a plastic vessel with a wide opening containing 7.5 mL of deionized distilled water, taking care to avoid any contact between them and the inner surface of the container.The containers were sealed and left for 7 days in an incubator (37°C, 95% RH).Afterward, the samples were removed from the containers, rinsed with deionized distilled water, blotted dry with absorbent paper, and placed in a dehumidifier for 24 h.They were then weighed again.The weight loss of each sample (initial mass minus final mass) was expressed as a percentage of the original mass (m% = m i -m f ) and taken as the SL of the sealer.A volume of 7.5 mL of distilled water was taken from each sample and poured into a clean and dry porcelain crucible.Each crucible was put into a muffle furnace and burned at 550°C.The resulting ash was dissolved in 10 mL of concentrated nitric acid (Merck KGaA, Darmstadt, Germany) using a glass stick.Thereafter, the samples were poured into 50 mL volumetric flasks, and the volume was made up with ultrapure deionized water (MilliQ, Millipore, Billerica, MA, USA).The solutions were sprayed into an atomic absorption spectrophotometer (Perkin Elmer, Uberlingen, Germany) to verify the presence of Ca 2+ , Na + , and K + ions.The arithmetic mean of three replicates for each specimen was recorded and considered as the result, expressed as mg/L.

Flow
For the FL tests, 0.5 mL of the sealer was manipulated.About 3 minutes after the start of mixing, the sealer was placed in the center of a glass plate (40 mm × 40 mm × 3 mm) with the aid of a graduated syringe.Then, a second glass plate weighing 120 g was placed onto the sample.After 10 minutes, the largest and smallest diameters of the samples were measured using a digital caliper.In this test, the difference between the larger and smaller diameters should not exceed 1 mm, and the sealers should be uniform.

Radiopacity
Five acrylic plates (2.2 cm × 4.5 cm × 1 mm) containing four wells measuring 1 mm in depth and 5 mm in diameter were prepared and placed over a glass plate covered by a cellophane sheet.Each well was filled with one of the sealers, following a sequence according to the ST of the material so that the samples were ready for radiographic evaluation immediately after the final setting of all materials.To avoid the formation of bubbles, the freshly mixed sealer was introduced into the wells using a syringe.Another glass plate covered with cellophane was placed on top of the samples until complete setting, and any excess sealer was removed.Each plate was kept in an incubator (37°C, 95% RH) for a period corresponding to three times the ST.
At the time of radiographic exposure, each of the acrylic plates containing the sealers was positioned alongside another acrylic plate (1.3 cm × 4.5 cm × 1 mm) containing an aluminum step wedge made of 1100 alloy with the thickness varying from 1 mm to 10 mm in uniform steps of 1 mm each (Margraf Dental MFG Inc., Jenkintown, PA, USA).This set of acrylic plates was placed in front of a phosphor plate next to the aluminum step wedge, and a digital radiograph was taken (Digora TM system; Soredex Orion Corporation, Helsinki, Finland).Radiographic images were obtained using a Spectro 70X X-ray machine (Dabi Atlante, Ribeirão Preto, SP, Brazil) at 70 kVp and 8 mA.The object-to-focus distance was 30 cm, and the exposure time was 0.2 s.Exposed imaging plates of the test samples were immediately scanned after exposure (Digora TM Scanner) and analyzed using Digora TM for Windows 5.1 software.
Digora TM software provides the radiographic density determination (densitometric analysis) or, in other words, the RD of a given material through its gray levels (mmAl).Thus, a 2 mm 2 area (44.5 × 44.5 px 2 ) was standardized and used for each specimen in the radiographic images of the sealers.

Statistical analysis
Five specimens from each group were tested and the means were statistically compared with the aid of SPSS Statistics 17.0 software (LEAD Technologies, Inc., Chicago, IL, USA).The Kolmogorov-Smirnov test showed that the results were consistent with a normal distribution curve; thus, parametric statistical analysis was possible (one-way ANOVA and post hoc Tukey-Kramer test), and the null hypothesis was set as 5%.

Results
Table 2 shows the analysis results of the physicalchemical properties of the root canal sealers.

Setting time
Among the sealers tested, AH Plus presented the highest mean values, and the difference in results between this sealer and those of the other sealers was statistically significant (p < 0.05) (Table 2).According to ANSI/ADA Specification 57, all root canal filling materials should not have an ST greater than 10% of the time determined by the manufacturer.Therefore, GuttaFlow Bioseal, MM Seal, and AH Plus conform to the recommended specifications.

Dimensional change
Among the sealers tested, GuttaFlow 2, GuttaFlow Bioseal, and MM Seal sealers showed the highest mean values, and statistically significant differences were observed among groups.AH Plus presented the lowest mean value, and this difference was statistically significant when compared with the values of the other sealers (p < 0.05) (Table 2).ANSI/ADA Specification 57 recommends that sealers should not exceed 1% contraction or 0.1% expansion.

Solubility
GuttaFlow 2 and GuttaFlow Bioseal presented the highest mean values and standard deviations in the SL test, and the results of these sealers were statistically similar to each other and statistically different when compared with those of AH Plus, which presented the lowest averages (p < 0.05) (Table 2).ANSI/ADA Specification 57 stipulates that an ideal endodontic sealer should not lose more than 3% of its mass when its SL is tested.
The amounts of metal ions released into the liquid resulting from immersion of the SL test samples are listed in Table 3.The data in Table 3 show that GuttaFlow 2 and GuttaFlow Bioseal present higher calcium release rates than either MM Seal or and AH Plus and that GuttaFlow Bioseal is the only sealer with a high sodium release rate.All of the sealers showed low rates of potassium release.

Flow
All of the sealers presented similarly intermediate values, with no statistically significant difference among them (p > 0.05) (Table 2).ANSI/ADA Specification 57 states that sealers should not have a difference greater than 1 mm between the largest and smallest diameters of the flowed material.

Radiopacity
The radiographic densities of the sealers were obtained in gray scale.MM Seal showed the lowest mean values and standard deviations, and differences between the results of this and other groups were statistically different (p < 0.05) (Table 2).According to ANSI/ADA Specification 57, all root canal filling materials must have an RD greater than or equal to 3 mmAl.All of the sealers presented an RD greater than 3 mmAl.  of the calcium silicate present in GuttaFlow Bioseal may result in the greatest accumulation of water between the chains of this polymer. 19,30he results of AH Plus and MM Seal are in agreement with those of Bodrumlu et al., 15 Resende et al., 21 Bodrumlu et al., 5 Marin-Bauza et al., 6 Poggio et al., 16 Flores et al., 3 Sonntag et al., 31 and Prüllage et al., 27 which suggests that their low SL may be related to their composition and polymerization reactions.When the two pastes are mixed, the diepoxy and polyamine compounds react, forming a covalent reaction.Amine groups promote changes in the ST, density, and morphology of the epoxy resins.Each amine group can react with an epoxy group, resulting in the formation of a strong and compact cross-linked polymer, which promotes low levels of SL. 3,7 Complementary analysis of the solutions by means of atomic absorption spectrometry showed the significant release of calcium ions from all of the evaluated sealers, as well as the release of sodium ions from GuttaFlow Bioseal (732.97 ± 124.32).The high rate of sodium release from this sealer is due to the addition of bioactive glass ceramics in its composition.The material is mainly composed of sodium oxide, which stimulates the formation of mineralized tissue. 32,33,34,35,36Calcium and sodium ion release increases the pH of the dental environment and prohibits bacterial viability. 36n the FL test, all of the sealers presented a diameter greater than 20 mm as determined by ANSI/ADA.The flowabilities of GuttaFlow 2 (36.44 ± 0.05 mm) and GuttaFlow BioSeal (35.40 ± 0.03 mm) can be attributed to the reaction between the polydimethylsiloxane, silicone oil, and paraffin they contain, as well as the pressure exerted by the glass plate during the test, once these sealers have thixotropic capacity under compression, which allows the fluidity and greater penetrability of these sealers. 3,14 17 18 37,38he FL test results of MM Seal (52.75 ± 0.60 mm) and AH Plus (36.42 ± 0.40 mm) can be attributed to the presence of thickening agents and polymerization activators, including ethylene glycol salicylate, aerosil, the adamantylamine, and N,N-dibenzyl-5-oxanonanediamine-1,9 in their composition, which promotes the thixotropy of the internal structure of the sealers during the polymerization reaction and increases their fluidity. 1,3,5,6,7,8,9,13,14,15,16,21 21,39 The RDs of GutaFlow 2 (6.85 ± 0.14 mmAl) and GutaFlow Bioseal (7.02 ± 0.18 mmAl) are related to the presence of radiopacifiers such zirconium dioxide and nano silver particles in their formulation, 3,14,7,18 while those of MM Seal (3.32 ± 0.90 mmAl) and AH Plus (7.52 ± 1.59 mmAl) are due to the presence of zirconium oxide, iron oxide, and calcium tungstate in their composition.1,3,4,8,9,14,24,40 The results found in the present study are in accordance with those of other studies, which found similar radiographic densities for AH Plus 3,4,6,21,26,27,31,39 and for GuttaFlow 2 and GuttaFlow Bioseal.3,14,17,18

Conclusion
According to the methodology employed and the results obtained in this study, AH Plus can be concluded to be the only sealer that fulfills ANSI/ ADA specifications.New studies should be carried out to better interpret the physicochemical properties of endodontic sealers and provide more support to researchers and clinicians attempting to develop an ideal sealer.

Table 1 .
Composition of the root canal sealers and their manufacturers.

Table 3 .
Metal solubility (mg/L) of root canal sealers determined by atomic absorption spectrometry.