Hydration Characterization of Two Generations of MTA-Based Root Canal Sealers

Abstract

Since the setting characterization of a root canal sealer has an impact on its biological behavior of final obturation, this study evaluated the setting characterization of mineral trioxide aggregate MTA-Fillapex versus MTA-Bioseal compared with epoxy resin (Adseal) root canal sealers. Freshly mixed sealer was inserted into the mold (n = 10). The initial and final setting times were evaluated using a Vicat needle and were then statistically analyzed by one-way ANOVA at p < 0.05. The raw pastes and the stages of the setting reaction were analyzed using Fourier Transform Infrared (FTIR) Spectroscopy. The phase compositions were evaluated using X-ray diffraction (XRD). A significant and fast setting time was recorded by Adseal (4.7 ± 0.46 h) followed by MTA-Bioseal (11.4 ± 1.34 h) at p < 0.001. The MTA-Fillapex did not set completely in three months. The FTIR and XRD of both MTA-Bioseal and Adseal detected bands of the polymerized phases, while those of MTA-Fillapex detected partial polymerization with a low percentage of polymerized silica. MTA-Bioseal and Adseal met the ISO standards for setting times. However, MTA-Fillapex did not fulfill the ideal requirement for the sealer. Although the raw pastes of both MTA-sealers had nearly similar compositions, they behaved differently during the hydration reaction. MTA-Bioseal set completely, while MTA-Fillapex was not completely set.

1. Introduction

Due to the lack of gutta-percha adhesiveness, root canal sealers are regularly used to achieve three dimensional obturation and fill minute spaces between the gutta-percha cones and canal walls, providing a fluid-tight seal [1]. Different types of sealers have been developed possessing different compositions to meet the criteria of Grossman described in 1988 [2]. Since the development of mineral trioxide aggregate (MTA), it has been considered a gold-standard biocompatible material with high numbers of interesting clinical applications in endodontic treatment [3]. Based on the cohesive strength, biological behavior and osteogenic potential of MTA [4,5], MTA-based root canal sealers were introduced to offer all of these advantages. MTA is mainly composed of tri-calcium silicate (C₃S), di-calcium silicate (C₂S), and bismuth oxide [5,6]. It is characterized by alkaline pH [5] that creates a suitable environment for periapical healing [7].

The setting time and setting reaction have an impact on the performance of root canal sealers and, in turn, on the quality of root canal obturation. According to Grossman, the sealers must have ample working time to allow sufficient time for condensation procedures. On the other hand, an extended setting time is an undesirable property when considering the inflammatory reaction and cytotoxic effect of the adjacent periapical tissues [8,9].

MTA-Fillapex was first developed by Angelus^® (Londrina/Parana/Brazil) in 2010 [10]. It is available in a syringe with an automixed tip. According to the manufacturer, it is composed of 40% MTA (C₃S, C₂S, tri-calcium oxide, and tri-calcium aluminate), fumed silica, titanium dioxide, natural resin, salicylate resin, and bismuth trioxide. It is characterized by alkaline pH due to the formation of calcium hydroxide as a by-product of its hydration reaction [6]. It provides some biological properties such as antibacterial activity [4,11] and bioactivity [12]. However, the conventional MTA-cement had an extended setting time [5]. According to manufacturer’s guidelines, MTA-Fillapex has a setting time of 130 ± 10 min [10]. One study showed a prolonged working time, initial setting time ≈ 2.27 ± 0.06 h, and final setting time ≈ 4.55 ± 0.05 h. However, it remained partially set in a previous study even for one month [11,13]. In this context, manufacturers compete to produce modified generations to improve the physical and chemical properties of MTA-based sealers.

Recently, a new generation of MTA-based sealer was launched by ITENA Clinical, (Paris, France). The manufacturer claims it has a suitable working time and short setting time ≈ 2 h [14]. There is no further information about its setting characterization other than that given in the manufacturer’s reports. The setting characterization of the sealer has an impact on its handling and quality as well as the biological behavior of the final obturation [15].

The current study aimed to evaluate the setting characterization of the first (MTA-Fillapex) and the recent (MTA-Bioseal) generations compared with epoxy resin (Adseal) sealers. The null hypothesis was that there would be no significant difference between the three root canal sealers.

2. Materials and Methods

The MTA-Bioseal (ITENA Clinical, Paris, France) and MTA-Fillapex (Angelus, Londrina, Brazil) were evaluated. Adseal (META Biomed Co., Chungbuk, Korea) was used and served as a control.

2.1. Setting Time

Based on the manufacturers’ instructions, each sealer was mixed and inserted into a polyethylene mold of 10 mm diameter and 3 mm height (n = 10). The procedure was performed according to ISO 6876 and ADA specification # 57 [16,17]. The molds containing the mixed sealers were packed between two glass plates and kept in incubator at 37 °C and 100% humidity. After 30 min, periodic penetration of a Vicat needle was performed every 15 min. At room temperature (23 ± 1 °C), the Vicat needle (Jin-Ching-Her, New Taipei City, Taiwan), of 10 mm diameter, 50 mm length and 100 gm weight, was applied to the surface of the mixed sealer. The initial setting time was recorded when there was decrease in needle penetration as it was recorded among the millimeter scale presented on the device. The final setting time was recorded when there was no obvious indentation on the sealer surface. The Vicat initial and final setting times were recorded in this way. The procedure continued until the final setting of the sealers. However, for the sealer that failed to set, the procedure was repeated every day for three months.

2.2. Setting Reaction Based on Fourier Transform Infrared (FTIR) Spectroscopy

The raw base and catalyst pastes of each sealer were firstly evaluated by Fourier Transform Infrared Spectroscopy (FT/IR-6100, Jasco Int Co, Tokyo, Japan). Fresh mix was prepared from each sealer and was chemically analyzed by FTIR spectroscopy. Readings were obtained immediately, after 4 h, 24 h, and after complete setting, according to the data obtained from vicat setting time. The spectra were collected at 4000–400 cm⁻¹ wave number and 1 cm⁻¹ resolution. To verify the data, three scans were carried out for each sample.

2.3. Phase Analysis by X-ray Diffraction (XRD)

After complete setting, the discs of each sealer were milled into a fine powder and analyzed by X-ray diffraction (XRD, Empyrean, Analytical 2010, Eindhoven, The Netherlands, UK) to determine the phase composition after the setting reaction.

2.4. Statistical Analysis

After confirmation of a normal distribution, the data of setting time were statistically analyzed by one-way ANOVA and post-hoc Tukey HSD tests using SPSS software (Version 16.0; SPSS, Inc, Chicago, IL, USA) at a significance level of 5% to compare the tested materials.

3. Results

3.1. Setting Time

The initial setting of MTA-Fillapex was significantly prolonged (29 ± 0.74 h) versus the other two sealers (p < 0.001), with no significant difference between MTA-Bioseal (1.3 ± 0.13 h) and Adseal (1.3 ± 0.12 h) (p = 0.99). Regarding the final setting time, the Adseal exhibited significantly faster setting (4.7 ± 0.46 h) followed by MTA-Bioseal (11.4 ± 1.34 h) at p < 0.001 (

Table 1. Mean ± standard deviation values of initial and final setting times of all investigated root canal sealers.

		MTA-Bioseal	MTA-Fillapex	Adseal	p Value
Setting time (hours)	Initial	1.3 ± 0.13	29 ± 0.74 *	1.3 ± 0.12	<0.001
	Final	11.4 ± 1.34	NA *	4.7 ± 0.46

The asterisk (*) indicates a significant difference at p < 0.001.

). Although MTA-Bioseal showed no indention at this time, it was still fragile and became hard after 72 days. The MTA-Fillapex was not set completely, and the Vicat needle still marked the material surface three months later.

3.2. Setting Reaction by FTIR

FTIR analysis showed nearly similar compositions of the raw pastes of both MTA-based sealers at varying intensities (Figure 1A,B). Their spectra detected bands of calcium hydroxide (Ca(OH)₂ ≈ 3642 cm⁻¹ [18,19,20,21,22], OH ≈ 3300 cm⁻¹ [22], methyl (C-H) at 2980–2870 cm⁻¹, carbonyl group (C=O) at 1700–1500 cm⁻¹ and 1320–1300 cm⁻¹, and aromatic C-O bond (≈1210 cm⁻¹) of salicylate [21,23,24,25] with low intensity in MTA-Fillapex. Bands of carbonate (C-O) at 1460–1420, 1240–1200, and ≈700 cm⁻¹ [18,20,22,23,24,25,26], sulphate band ≈ 1155 cm⁻¹ [22] (in Bioseal), titanium oxide at 1092 cm⁻¹ [27] (in Bioseal), v₃SiO₄ of tricalcium silicate (C₃S, alite) at 1086 cm⁻¹ [21], dicalcium silicate (C₂S, belite) at 868 cm⁻¹ [20], SiO₄ at 859 cm⁻¹, v₄SiO₄ at 800–700 cm⁻¹ [22], v₂SiO₄ of C₃S ≈ 522 cm⁻¹ [20], v₁SiO₄ of C₂S ≈ 452 cm⁻¹ [21], phosphate bands ≈ 1040–1030 cm⁻¹ [18,23,28,29], aluminum oxide (Al-O-Si) ≈ 750 cm⁻¹ [25,30,31], and Ca-O of tricalcium aluminate ≈ 420 cm⁻¹ (in Fillapex) [32] were also detected.

The spectra of the mixed MTA-sealer showed a dip of calcium hydroxide, amide C=O, C-H, and C₃S (≈860 cm⁻¹) bands synchronized with a hump of carbonate and silicate bands. The dip/hump of band intensity was a gradual progression with time. In the spectra of MTA-Bioseal, v₃v₂SiO₄ bands (at 940 and 573–515 cm⁻¹) gradually shifted to higher wave numbers (≈950 and 592–577 cm⁻¹, respectively). Furthermore, the O-Si-O bands at 795 and 440 cm⁻¹ assigned to calcium silicate hydrate (CSH) [22] became more intense. In MTA-Bioseal spectra, the sulphate band ≈ 1155 cm⁻¹ [22] gradually dipped over time. No further changes were detected after 24 h. In the spectra of MTA-Fillapex, the bands related to CSH ≈ 950, 795, and 440 cm⁻¹ failed to be detected up to three months (Figure 1B).

The spectra of raw pastes of Adseal detected the bands of OH stretching at 3342 cm⁻¹, C-H at 2980–2800 and ≈1340 cm⁻¹ [21,33,34], carbonyl group (C=O) at 1670 and 1602 cm⁻¹ [23], C=C of the benzene ring at 1518 cm⁻¹ [35], CH₃ group ≈ 1450 cm⁻¹, phosphate of v₃PO₄ at 1100–1000 cm⁻¹, v₄PO₄ at 600 and 560 cm⁻¹ and v₂PO₄ ≈ 490 cm⁻¹ [33], CH₃ of epoxoid ring at 740–700 cm⁻¹ [36,37], and Si-O at 790 cm⁻¹ [38]. The spectra of the immediate mix showed humping of the C=O bands at 1672 and 1508 cm⁻¹, C-C at 1298 cm⁻¹, CH₂ at 1248 and 790 cm⁻¹, and CH at 740–700 cm⁻¹ [37] of the epoxoid ring [34,36,37]. The bands of aromatic rings at 829 and 755 cm⁻¹ humped and shifted to higher wave numbers [34]. There was dipping in the band intensity of C-O-C and C-O of the anhydrous epoxoid ring at 916 and 863 cm⁻¹, respectively [35,36,39] that gradually decreased in intensity in the spectra of 4 h and disappeared in the spectra of 24 h. There were no further changes after 24 h, except a slight dipping of v₃PO₄ at 900 cm⁻¹ (Figure 1C).

3.3. Phase Composition Determined by X-ray Diffraction (XRD) Analysis

The XRD patterns of the tested sealers were obtained at 0–60° 2θ with varying intensities (Figure 2). MTA-Bioseal specimens showed peaks of dicalcium silicate (Ca₂O₄S, Entry # 96-210-3317), calcium carbonate (Calcite, CCaO₃, Entry # 96-900-0967), calcium hydroxide (CaH₂O₂, Portlandite, Entry # 96-100-8781), calcium silicate (Hatruite, Ca₉O₁₅Si₃, Entry# 96-900-9266), titanium oxide (O₂Ti, Entry # 96-810-4260), aluminum phosphate (AlO₄P, Entry # 96-201-2012), and tricalcium silicate (Alite, Ca₃O₅Si, Entry # 96-901-6126) (Figure 2A).

MTA-Fillapex specimens showed peaks of tricalcium silicate (Alite, Ca₃(Si O₄)O, Entry # 96-154-0705), bismuth oxide (Bismite, Bi₂O₃. Entry # 96-101-0005), calcium silicate (Hatrurite, Ca₉O₁₅Si₃, Entry # 96-900-9266), calcium hydroxide (Portalandite, CaH₂O₂, Entry # 96-100-1769), calcium aluminate (Aluminium calcium 2:1, Al₂Ca, Entry # 96-101-0451) (Figure 2B).

Adseal specimens showed peaks of tricalcium phosphate (Ca₃O₈P₂, Entry # 96-210-6195), silicon oxide (Quartz, O₂Si, Entry # 96-500-0036), and bismuth oxide (Bismite, Bi₂O₃, Entry #96-101-0005) (Figure 2C).

4. Discussion

The initial setting time indicates the beginning of sealer solidification that was associated with a decrease in the penetration depth of the Vicat needle [40], while the final setting time indicates the complete hardness of the sealer without obvious indentation at the sealer surface [12]. According to the favorable requirements described by Grossman, the root canal sealer must have a slow initial setting time to promote sufficient working time [2]; however, a prolonged final setting time directly correlates with the degree of cytotoxic reaction [12]. The present study evaluated the setting characterization of the first (MTA-Fillapex) versus recent (MTA-Bioseal) generations of MTA-based sealers and compared them with an epoxy resin (Adseal) sealer. The current result revealed that the three tested sealers had different setting characterizations; therefore, the null hypothesis was rejected.

This study revealed a significant prolonged initial setting time exhibited by MTA-Fillapex (29 ± 0.74 h) with no significant difference between MTA-Bioseal and Adseal (≈1.3 ± 0.12 h at p = 0.99). Finally, Adseal exhibited significantly fast setting (4.7 ± 0.46 h). MTA-Bioseal showed no indentation after 11.4 ± 1.34 h, but remained fragile until 72 h. The samples of MTA-Fillapex were not completely set for three months. The current study is the first publication to evaluate the setting characterization of MTA-Bioseal. It recorded a setting time that exceeded that claimed by the manufacturer with a 23-min working time and a setting time within two hours. This finding may be attributed to the nature of MTA-based materials, as they take a prolonged time for setting [5,6]. According to ISO 6876 for testing the root canal sealers, it did not exceed 10% longer than that claimed by the manufacturer [16]. Sealers that exhibit a prolonged setting time may promote periapical irritation and inflammatory reaction if exposed to periapical tissues [12].

MTA-Fillapex and Adseal have been extensively investigated [13,41,42,43]. The fast setting of Adseal (≤2 h) was consistent with previous studies [13,44], but there was a conflict with the previous results related to MTA-Fillapex. Some studies recorded a short setting time similar to that of Adseal [41,42] or faster than AH plus [43]; however, it failed to completely set in other studies [11,13]. Although Vitti et al. [43] determined a final setting time of MTA-Fillapex (≈4.55 h), similar to that described by the manufacturer (130 ± 10 min) [10], other studies by Lee et al., 2017 [13] and Benezra et al., 2017 [11] suggested that it did not set indefinitely.

The setting times of the sealer are proposed to depend on their composition, temperature, and humidity [45]; it was suggested that calcium silicate sealers could not set when stored dry [46]. The MTA-Fillapex is a premixed injectable paste of hydraulic calcium silicate sealer, containing a water-free thickening vehicle [12]. For its hydration reaction, it needs water to permit final setting and maximum hardness. According to Lee et al., 2017 [13], a humid incubator condition is not sufficient for a complete setting of such hydraulic sealers (MTA-Fillapex). However, in a clinical situation, when such a sealer is inserted in the root canal, the dentinal tubules are filled with fluid that enhances its hydration reaction and complete setting. This fact can explain the failure of MTA-Fillapex setting during the environment of the current study.

The constituents of the sealer affect and regulate its setting time and the hydration reaction. Salicylate resin was recently added to a root canal sealer to enhance its antibacterial properties [47]. All three tested sealers contain calcium particles and salicylate resin to promote a favorable biological response [42] and allow their polymerization. As described by the manufacturers, both MTA-based sealers are composed of calcium silicate (C₂S and C₃S), salicylate resin, and bismuth oxide [10,14], while Adseal is composed of epoxy resin, calcium phosphate, calcium oxide zirconium oxide, bismuth subcarbonate, and ethylene glycol salicylate [13]. The content of salicylate resins seemed to affect the setting time. After the mixing of the base and catalyst paste, MTA reacts with salicylate forming anionic polymer [43]. The raw FTIR spectra of MTA-Fillapex showed low intense salicylate bands that may be responsible for its prolonged setting times.

In general, MTA is a hydrophilic material, composed of tri- and di- calcium silicate [Alite (C₃S) and belite (C₂S)] that react with water to form calcium silicate hydrate (CSH) as a polymerized (cured) calcium silicate phase [45]. At the end of the setting reaction, the curing stage of MTA depends on the formation of CSH that is responsible for the material hardening and strength [6]. The presence of a high amount of unreacted alite and/or belite indicates the incomplete setting of the material. FTIR spectroscopy can offer reliable information to evaluate the chemical structures [48,49]. FTIR analysis explains the process of hydration reaction until set, which was confirmed by XRD analysis. In the current study, FTIR analysis revealed similar compositions of the raw pastes of both MTA-based sealers. Their spectra showed bands of salicylate resin, calcium hydroxide and/or carbonate, calcium silicate (C₃S, C₂S), phosphate (vPO₄), and tricalcium aluminate (Figure 1). The finding related to MTA-Fillapex is similar to that in previous studies [13,50]. The FTIR spectra of its raw pastes determined the low intense C-O bond (≈1210 cm⁻¹) of salicylate (catalyste) that remained prominent in spectra after three months. It seems to not be involved in the setting reaction and results in a prolonged setting time. Regarding MTA-Bioseal, there was no more information other than that described by the manufacturer [14]. When the MTA-sealer is mixed in the presence of a humid environment and during the carbonation process, the hydration reaction is characterized by the formation of Ca(OH)₂ and calcium carbonates (CaCO₃) as primary by-products [51] that are responsible for its antimicrobial activity and biocompatibility [4,11]. Calcite is added to the cement for the hydration reaction and improves the mechanical properties of the sealer [52]. The carbonated bands dipped as the hydration reaction progressed, indicating their consumption during the hydration reaction.

During the hydration reaction, immediately after the mixing of the pastes of the base and catalyst, the bands of C₃S (≈850 and 590 cm⁻¹) or C₂S (≈470 cm⁻¹) were dipped/humped and were replaced by the band of polymerized calcium silicate and the formation of a CSH phase at 440 cm⁻¹ [22,26]. In addition, as the hydration reaction progressed, the unhydrated calcium silicate (v₃SiO₄) band ≈ 940 cm⁻¹ was shifted to a higher wave number ≈950 cm⁻¹ (Figure 1) [22,51] synchronized with a dip/hump of v₃SiO₄ and v₂SiO₄ (Alite) ≈ 522 and 450 cm⁻¹, respectively, and the prominent of a sharp band at 1083 cm⁻¹. This is indicative of silica polymerization and growth of CSH as a final cured by-product when the MTA-based sealer undergoes complete setting [22,26]. Concerning the FTIR spectra of MTA-Bioseal, no further changes were detected after 24 h, which confirmed that the setting time did not exceed 24 h. In the spectra of MTA-Fillapex, the bands related to CSH ≈ 950, 795, and 440 cm⁻¹ failed to be detected up to three months, indicating its incomplete setting. Although the formation of CSH was detected, the sealer surface may have contained some of unreacted particles of C₂S and/or C₃S or limited polymerized calcium silicate (CxS). This feature was true for MTA-Bioseal, but it failed to be detected in the spectra of hardened MTA-Fillapex with detection of the high intense alite band at 1086 cm⁻¹ [21], which is indicative of incomplete setting of the sealer. The complete setting reaction is determined by the increased proportion of cured (set) CSH in the form of Hatruite with respect to unreacted alite (C₃S) and/or belite (C₂S). This finding was confirmed by the phase composition detected in XRD analysis. The MTA-Bioseal spectra revealed a high proportion of Ca(OH)₂ and Hatruite, indicating complete setting. However, MTA-Fillapex XRD spectra showed a higher proportion of C₃S and low proportion of Hatruite indicating less of a polymerization process. In a previous study, MTA-Fillapex showed only Hatruite, bismuth oxide, and silicon oxide [46]. The XRD spectra of the final setting of Adseal also confirmed the composition of cured resin as they revealed calcium phosphate, silicon oxide, and bismuth oxide. This confirmed recent previous findings, where calcium, phosphorous, silicon, and bismuth elements were detected by EDX analysis [53]. There were no marked changes in the FTIR spectra of MTA-Bioseal and Adseal after 24 h. This finding is indicative of the complete sealer set within 24 h.

The curing process of the epoxy resin sealer was characterized by the dipping of the calcium hydroxide, amide C=O, C-H, and C₃S (≈860 cm⁻¹) bands synchronized with the hump of the carbonate and silicate bands. The dip/hump of the band intensities gradually progressed over time until 24 h. The same finding was previously described [33]. During the curing process of epoxy resin, the FTIR band intensity of C-O-C and C-O of the anhydrous epoxoid ring at 917 and 863 cm⁻¹ decreased, and the band intensity at 1672 cm⁻¹ increased [35,39,54], indicating complete resin polymerization and setting. There were no more changes in the spectra after four hours, supporting the finding of the final setting time for Adseal at 4.7 ± 0.46 h.

5. Conclusions

Regarding the setting time, both MTA-Bioseal and Adseal complied with the ISO standards; however, MTA-Fillapex did not fulfill the ideal requirement described by Grossman. MTA-Fillapex presented an extended initial setting time and did not completely set up to three months. Although the raw pastes of both MTA-sealers had a similar composition, they behaved differently during the hydration reaction. A dip/hump of band intensity was detected in the FTIR spectra of MTA-Bioseal and Adseal up to 24 h indicating the final setting time and complete polymerization of silicate phases. However, the spectra of MTA-Fillapex showed a limited polymerized silicate phase indicating a partial polymerization process. Since the premixed MTA-Fillapex root canal sealer contains water-free vehicle, during the clinical situation, a sufficient moisture environment is essential for the complete setting of such a sealer.