Effect of pH on Sealing Ability of White Mineral Trioxide Aggregate as a Root-end Filling Material Mohammad Ali Saghiri, BSc, MSc, PhD student,* Mehrdad Lotfi, DMD, MSc,† Ali Mohammad Saghiri,‡ Sepideh Vosoughhosseini, DMD, MSc,§ Ali Fatemi, DMD,储 Vahab Shiezadeh, DMD,储 and Bahram Ranjkesh, DMD¶ Abstract The aim of the present study was to evaluate microleakage of mineral trioxide aggregate (MTA) used as a root-end filling material after its exposure to a range of acidic environments during hydration. Seventy single-rooted teeth were divided into 4 experimental and 2 control groups. All the teeth were instrumented, and their apices were resected. Root-end cavities were filled with white MTA in the experimental groups. In the control groups root-end cavities were not filled. Rootend fillings were exposed to acidic environments with pH values of 4.4, 5.4, 6.4, or 7.4 for 3 days in the experimental groups. Microleakage was evaluated by using bovine serum albumin. The evaluation was conducted at 24-hour intervals for 80 days. Data were analyzed by using one-way analysis of variance and a post hoc Tukey test. The earliest bovine serum albumin microleakage was observed in a pH value of 4.4 followed by pH values of 5.4, 6.4, and 7.4, respectively. There was a significantly longer time necessary for leakage to occur in samples stored in higher pH values (P ⬍ .000). (J Endod 2008;34:1226 –1229)
Key Words Bovine serum albumin, mineral trioxide aggregate, root-end filling material
From the *Department of Biomedical Engineering, Islamic Azad University, Science and Research Branch and Department of Dental Materials, Faculty of Dentistry, Azad University of Medical Sciences, Tehran, Iran; †Research Center for Pharmaceutical Nanotechnology and Department of Endodontics, Dental Faculty, Tabriz University (Medical Sciences), Tabriz, Iran; ‡Computer Engineering Department, Amirkabir University of Technology, Tehran, Iran; §Department of Oral and Maxillofacial Pathology, Dental Faculty and Research Center for Pharmaceutical Nanotechnology, Tabriz University (Medical Sciences), Tabriz, Iran; 储Department of Endodontics, Dental Faculty, Tabriz University (Medical Sciences), Tabriz, Iran; and ¶ Private practice, Tabriz, Iran. Address requests for reprints to Dr Mehrdad Lotfi, Tabriz University (Medical Sciences), Research Center for Pharmaceutical Nanotechnology and Department of Endodontics, Golgasht Street, Tabriz, Azarbaidjan Sharghi 5166614711, Iran. E-mail address: [email protected]
0099-2399/$0 - see front matter Copyright © 2008 American Association of Endodontists. doi:10.1016/j.joen.2008.07.017
Saghiri et al.
ndodontic surgery is necessary when conventional root canal therapy cannot treat complex anatomy, procedural misadventures, or inflammatory processes (1). In periapical surgery, some materials are used to seal the root canal contents from the periapical tissues as a root-end filling material (1). An ideal root-end filling material should be biocompatible, insoluble in tissue fluids, nonresorbable, radiopaque, dimensionally stable, and should have sufficient sealing property (2). Mineral trioxide aggregate (MTA), first introduced in 1993 (3), has many advantages that make it a suitable root-end filling material (2). In a clinical study Saunders et al. (4) demonstrated that use of MTA as a root-end filling material with a careful microsurgical technique results in a high success rate. MTA is a mixture of Portland cement, bismuth oxide, and trace amounts of other metallic oxides (5). MTA has an initial pH of 10.2, which increases to 12.5 three hours after mixing (6). In many real clinical situations, MTA is applied in an inflamed environment where a low pH exists (7). Physical and chemical properties of MTA might be influenced in a low pH environment (8). Impeded MTA setting (1) as well as reduced strength (6) and hardness (8, 9) has been reported in an acidic environment. A study revealed that mixing MTA with an acidic solution like 2% lidocaine HCl with an epinephrine concentration of 1:100,000 reduces the compressive strength of MTA in an acidic environment (10). The success of endodontic materials depends mainly on their ability to prevent leakage (11). Set MTA contains many voids in the forms of air bubbles, pores, and capillary channels (12). At MTA-dentin interface, these voids might result in leakage of bacteria and endotoxins past MTA (12). In a scanning electron microscopy analysis, a great degree of porosity was observed in MTA samples exposed to low pH environments (9). MTA gradually dissolves in the presence of synthetic tissue fluids (pH, 7.4) in root canals, and hydroxyapatite crystals nucleate and grow, filling the microscopic spaces between MTA and dentinal walls (13). Initially this seal is mechanical. Over time, a diffusion-controlled reaction between the apatite layer and dentin leads to their chemical bonding and creates a seal at MTA-dentin interface (13). Roy et al. (14) reported that the sealing ability of MTA is not affected by low pH. However, lowered physical and chemical properties of MTA and more porosity in an acidic environment lead to the speculation that MTA might not possess the same microleakage properties under different pH values. The aim of the present study was to compare microleakage of MTA as a root-end filling material in a solution at different pH values by using bovine serum albumin.
Materials and Methods Tooth Preparation Seventy single-rooted human anterior teeth (central and lateral) with straight canals and mature apices, extracted for periodontal and orthodontic reasons, were collected for the purpose of this study and were stored in phosphate-buffered saline solution until used. The surfaces of the roots were cleaned by using an ultrasonic device. The teeth with cracks and calcified canals were excluded from the study. The teeth were randomly divided into 4 experimental groups, each containing 15 teeth, and 1 negative and 1 positive control groups, each containing 5 teeth. Standard access cavities were prepared with a round diamond bur under water spray. The root
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Basic Research—Technology canals were prepared with the conventional step-back technique by using K-files (Maillefer, Ballaigues, Switzerland). The master apical file was #35. After each instrument, the root canals were irrigated with 2 mL of 2.5% NaOCl solution. Gates-Glidden drills #2 and 3 were used in the coronal third of the canals. The canals were dried with paper points. Root canals were obturated with gutta-percha (VDW, Munich, Germany) without sealer by using a cold lateral compaction technique. The apical 3 mm of the roots was resected perpendicular to the long axis of the teeth by using a diamond bur in a high-speed handpiece under continuous water spray. The apical ends of the roots were prepared with the Kis-3D microsurgical ultrasonic instrument (Spartan, Fenton, MO). A circular preparation 3 mm deep was created and rinsed with saline and dried with paper points. Microcondenser tip was checked to ensure that the tip would fit the length. The apical preparations were filled with white MTA (WMTA) (Tooth-colored Formula; Dentsply, Tulsa Dental, Tulsa, OK), which was prepared according to manufacturer’s instructions in the experimental and negative control groups. All roots were radiographed in 2 buccolingual and mesiodistal directions to ensure adequacy of root-end fillings. Apical preparations in the positive control group were left unfilled. Three wet pieces of gauze that had been soaked in butyric acid buffered at pH values of 4.4, 5.4, or 6.4 were placed in 3 separate vials. Each experimental group was placed in 1 vial for 3 days, and the acidsoaked pieces of gauze were replaced every day with fresh ones to ensure a sufficient acidic environment within the vials. One wet piece of gauze was soaked in synthetic tissue fluid (STF) that was prepared as follows: 1.7 g of KH2PO4, 11.8 g of Na2HPO4, 80.0 g of NaCl, and 2.0 g of KCl in 10 L of H2O (pH, 7.4) were placed in 1 vial, and another experimental group was placed in it for 3 days. The roots in the experimental and positive control groups were coated with 2 layers of nail varnish. The apical and coronal surfaces were left uncoated. In the negative control group the entire root surfaces were covered with 2 layers of nail varnish.
Leakage of Bovine Serum Albumin The experimental model was similar to the one used in a study carried out by Valois and Costa (15). In brief, a hole was created in the rubber stopper of a 10-mL glass vial, and the teeth were inserted through it. Resected root ends pointed upward and the crown downward. A plastic cylinder was attached over the rubber stop around the teeth (Fig. 1). The glass vial was filled with 9.5 mL of redistilled water. The cylinder was filled with 1 mL of 22% bovine serum albumin (BSA) solution (Sigma Chemical Co, St Louis, MO). The apparatus was set for all the experimental and control groups and was placed in a humidor at 37°C for 80 days. The water in the glass vial was changed and the protein solution in the reservoir was replenished daily during experiment. Protein presence was detected every day with a reagent. The number of days taken for color conversion of the protein reagent was supposed to be indicative of apical leakage. Data were analyzed with one-way analysis of variance and Tukey test at the .05 level of confidence.
Results The specimens in the positive control group showed color conversion of the protein 1 day after the experiment began (Fig 2). In the negative control group, there was no color change throughout the experiment (Fig 2). The number of days (mean ⫾ standard deviation) needed for color change at pH values of 4.4, 5.4, 6.4, and 7.4 were 8.87 ⫾ 2.75, 24.87 ⫾ 2.13, 63.80 ⫾ 12.17, and 78.53 ⫾ 5.68, respectively. The time needed for leakage to occur was significantly longer in samples stored at higher pH values (P ⬍ .000) (Fig. 3). More JOE — Volume 34, Number 10, October 2008
Figure 1. The apparatus used to test protein leakage.
Sealing Ability of White MTA as a Root-end Filling Material
Figure 2. Samples obtained from the specimens after adding reagent. Blue color change showed albumin leakage (left), and no color change showed lack of microleakage during experiment (right).
porosity was observed on the surface of MTA exposed to lower pH values in the experimental groups (Fig. 4).
Discussion In certain clinical cases, freshly mixed MTA might be applied in the presence of inflammation, and the surface of the material is exposed to an acidic environment (7). In this study, we tried to mimic a clinical situation by exposing MTA to acid-soaked pieces of gauze. In our study, MTA was exposed for no longer than 3 days to the acidic solution because we wanted to simulate the situation in which the initiating and perpetuating factors of inflammatory process are removed by appropriate treatment. Butyric acid was used in the present study to simulate the effect of by-products of anaerobic bacterial metabolism (16 –18). The pH of a human abscess has been measured as low as 5.0 (7). In this study a range of different pH values, up to neutral pH, were used to evaluate microleakage in different simulated clinical conditions.
Figure 3. Box plots of the days it took for color conversion of the protein reagent that was taken as indicative of MTA apical leakage as a root-end filling material in different pH values, which illustrate the mean ⫾ standard deviation, minimum and maximum amount of days, as well as the variance in each experimental group.
Saghiri et al.
Figure 4. Scanning electron microscopy images of specimens exposed to pH 7.4 (A) and 6.4 (B). More voids (–) can be seen on the surface of MTA exposed to the 6.4 pH (original magnification, ⫻100).
Methods that have been used to evaluate the quality of root-end filling materials made use of pigments including methylene blue and India and Pelikan ink. However, these methods have been criticized by some researchers because most dyes have a low molecular weight and can penetrate sites that protein and bacteria cannot. Most dye leakage studies have measured the degree of leakage in 1 plane, making it impossible to evaluate the total amount of leakage (19 –21). The pH and chemical reactivity of dyes might also influence the degree of dye penetration (22). The results of our study differ from the results of a study carried out by Roy et al. (14). They demonstrated that an acid environment not only does not hinder but also enhances the sealing ability of MTA. In their study, MTA was used with calcium phosphate cement (CPC) matrix, and samples were exposed to a pH value of 5 for 24 hours. In addition, the type of acid used was not stated. Chow (23) demonstrated that CPC liberates water during setting, thereby not allowing MTA to be exposed directly to the acidic environment. Tayler (24) showed that various types of acid have led to different physical and chemical effects on Portland cement. Malic acid (25) and tartaric acid (26) retard, whereas lactic acid (27) accelerates the hydration process of Portland cement. Because of the similarity of MTA and Portland cement, these effects might happen to MTA as well. MTA is a highly alkaline material (6) and has been demonstrated to release soluble fractions in short-term (12) and long-term (28) periods sufficient to maintain the pH of the surrounding environment at a high level. In addition, the pH of MTA increases to a peak of 10.39 within the first 24 hours after mixing followed by a decrease to
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Basic Research—Technology 7.72 within 390 hours. It seems that using CPC matrix, unbuffered acidic solutions, short-term exposure time of MTA to acidic solutions (24 hours), use of dye penetration method, and the type of acid used in the study by Roy et al. (14) have led to a different result. Namazikhah et al. (9) showed that a trend exists that the more acidic the solution, the more extensive the porosity of the MTA, where butyric acid is used in direct contact with MTA for 4 days during hydration. The role of voids in the form of air bubbles, pores, and capillary channels within the MTA and in the MTA-dentin interface in bacterial and endotoxin leakage has been confirmed (12). Therefore, it is not unusual to observe a relationship between the acidity of the periapical environment and microleakage of MTA when used as a root-end filling material. On the other hand, inflamed tissues are removed during periapical surgery, and the pH of tissues adjacent to root-end filling and its interface with dentin might be changed to neutral in less than 3 days. In addition, in some clinical situations like open apex nonvital teeth with periapical lesions or lateral or furcal perforations with radiolucent lesions, MTA is placed as an apical plug or sealing material directly in contact with the lesions. Therefore, MTA might be exposed to an acidic environment for a longer time. It is possible, however, that variations in the pH value of the host tissues, because of preexisting pathologic conditions at the time of MTA placement, might affect its sealing ability and jeopardize the outcome of the treatment. In conclusion, it might be advisable to use MTA with CPC in situations in which MTA comes into direct contact with the lesions, not only as a matrix to control the placement of restorative materials (29) but also as a material that is more resistant to an acidic environment.
Acknowledgments This article was based on the thesis submitted by the first author to the Faculty of Biomaterials at Azad University Science and Research Branch of Tehran, Iran, in partial fulfillment of the requirements for Master of Science degree in Biomaterial Engineering. We thank Dr Reza Saghiri for the provision of laboratory facilities in the Department of Biochemistry at the Pasteur Institute of Tehran, Iran. Also special thanks to Professor Ali Shokuhfar and Drs Mohammad Aeinehchi, Mohammad Saghiri, Hajar Afsar Ladjvardi, Morteza Daliri, and Sahar Dadvand for all of their contributions to this research effort.
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