|Year : 2014 | Volume
| Issue : 1 | Page : 8-12
Comparative evaluation of effect of three different mineral trioxide aggregate solvents on calcium content of root dentin: An in vitro study
Payal Batavia1, Vaishali Parekh1, Palak Batavia2, Paras Kothari1, Hetal Chappla1, Mayurika Dabhi1
1 Department of Conservative Dentistry and Endodontics, K. M. Shah Dental College and Hospital, Pipariya, Vadodoara, Gujarat, India
2 Department of Periodontics, College of Dental Science, Davangere, India
|Date of Web Publication||21-Jun-2014|
Department of Conservative Dentistry and Endodontics, K. M. Shah Dental College and Hospital, Pipariya, Vadodoara, Gujarat
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Aim: The aim of this study is to evaluate the calcium content of solutions of three different mineral trioxide aggregate (MTA) solvents after immersion of root dentin at different time interval. Materials and Methods: A total of 36 extracted premolars were used in the study. Teeth were sectioned 2 mm using hard tissue microtome. One section was selected from each tooth. 12 sections were then immersed in the freshly prepared 17% ethylenediaminetetraacetic acid (EDTA), 17% carbonic acid, and 10% citric acid. Calcium dissolution was checked at different time interval of 10 and 15 min and 32 h with atomic absorption spectroscopy. Results were analyzed using one-way ANOVA test and multiple comparisons were carried out by Tukey's test. Results: About 17% carbonic acid showed maximum calcium dissolution followed by 17% EDTA and 10% citric acid. There was significantly difference in result. Conclusion: About 17% carbonic acid used for MTA retrieval can cause calcium dissolution of dentin and lessen the strength.
Mineral trioxide aggregate is bioactive material used widely for different clinical use, it has major disadvantage of retrieval. There are different solvents used for the same. Those may affect the organic content of dentin.
Keywords: Calcium dissolution, carbonic acid, citric acid ethylenediaminetetraacetic acid, mineral trioxide aggregate
|How to cite this article:|
Batavia P, Parekh V, Batavia P, Kothari P, Chappla H, Dabhi M. Comparative evaluation of effect of three different mineral trioxide aggregate solvents on calcium content of root dentin: An in vitro study. J Interdiscip Dentistry 2014;4:8-12
|How to cite this URL:|
Batavia P, Parekh V, Batavia P, Kothari P, Chappla H, Dabhi M. Comparative evaluation of effect of three different mineral trioxide aggregate solvents on calcium content of root dentin: An in vitro study. J Interdiscip Dentistry [serial online] 2014 [cited 2020 Jul 15];4:8-12. Available from: http://www.jidonline.com/text.asp?2014/4/1/8/134998
| Introduction|| |
Endodontic retreatment has been defined as a procedure performed on a tooth that has received prior attempted definitive treatment resulting in a condition requiring further endodontic treatment to achieve a successful result. Retreatment is clearly indicated when a periapical lesion, clinical signs, and/or symptoms are present. , Retrieval is a prerequisite for any root canal filling material, so that it is possible to re-treat in case of failures. 
Most investigations reported low or no solubility for mineral trioxide aggregate (MTA). , The inherent disadvantages of the material are its prolonged setting time (2 h and 45 min) and its inability to be retrieved from within the root canal. , Studies have shown that the surface hardness of MTA is reduced by acids. 17% carbonic acid  was selected as one of the chemicals for disintegrating MTA and retrieve MTA. While 17% ethylenediaminetetraacetic acid (EDTA)  can affect the setting and 10% citric acid  are the common acids used for dissolving MTA. The solvents used might pose potential damage to the tooth structure's organic and inorganic proportion.
Thus, null hypothesis of this study was that the solvents (17% carbonic acid, 10% citric acid, and 17% EDTA) when used for retrieval of MTA does not affect the calcium content of root dentin.
| Materials and methods|| |
A total of 36 extracted teeth were selected and were divided into three groups. Group 1 (n = 12): 17% EDTA (AVA Chemicals Pvt. Ltd.) (freshly prepared), Group 2 (n = 12): 17% carbonic acid (Arihant chemicals Pvt. Ltd.) group (freshly prepared), Group 3 (n = 12): 10% citric acid (Arihant Pvt. Ltd.) (freshly prepared). Inclusion criteria included single rooted teeth and absence of severe curvatures on root of the tooth. While teeth with cracks and root fracture, teeth with previous restorations, teeth with anatomical variations, teeth with severe curvatures, teeth with calcifications, teeth with narrow pulp space, teeth with root caries.
Preparation of sample
Teeth were prepared after scaling [Figure 2]a and then were stored in distilled water [Figure 2]b. Root cementum was removed with low speed fine grain diamond (Mani, Dentsplky EF 11) under abundant irrigation with water [Figure 2]c. The root canals were instrumented using peso no. 4-6 mounted (Mani) on contra angle handpiece (NSK panaair). After each use of an instrument the root canals were irrigated to eliminate any possible debris remains. Instrumentation was considered completed when canal walls were visibly smooth and free of debris.
Sectioning of sample using microtome
Teeth were mounted in acrylic blocks. The crown plus apical third of the specimen were sectioned using hard tissue microtome [Figure 2]d and e (Leica 1600. Three transversal sections of 2 mm thickness were obtained from each root using hard tissue microtome. All sections were stored in distilled water.
Preparation of solution
17% EDTA, 17% carbonic acid and 10% citric acid solutions [Figure 2]f were freshly prepared by maintaining the pH of 5.1. Initially, 36 ml of solution was taken in a beaker as a blank to determine calcium levels in absence of specimen. Each specimen was immersed in each beaker containing the corresponding solution. A volume of 3 ml aliquots were extracted, at an interval of 10 min, 15 min, and 32 h, using calibrated micro pipette from each beaker, which was discarded after each extraction. These extracts were placed in hermetically sealed and labeled glass tubes. By this three extracts (at 10 min, 15 min, and 32 h) were obtained for each sample specimen.
Determination of dissolution of calcium
In these three extracts, the calcium concentration of the solution was determined by means of atomic absorption spectrophotometer (spectra AA500). Extract readings were expressed in ppm and then transformed into milligram of calcium lost per gram by applying the following formula: 
mgCa 2+ /g = ([ppmCa 2+ ]·10−3 L/mL·V)/P
ppmCa 2+ = ppm of calcium in each time period,
V = volume of solution in ml (at 10 min, V1; at 15 min, V2; and at 32 h, V3)
P = weight of solution in mg.
For the 10 min period the volume of solution (V1) was 36 ml, whereas at 15 min the volume (V2) was 33 ml because 3 ml of solution had previously been removed. Therefore, the mg of Ca 2+ corresponding to 3 ml of V1 solution has to be added to the mg of Ca 2+ obtained at 15 min. Likewise, because the volume was 30 ml at 32 h, the mg of Ca 2+ corresponding to 3 ml of V1 solution plus 3 ml of V2 solution were added to those obtained at 32 h sample.  The amount of change in calcium concentration was calculated by the help of atomic absorption spectrometry.
The statistical analysis (software used was IBM SPSS Statistics) was carried out for different groups and time interval with one-way ANOVA and then multiple comparison with Tukey's.
| Results|| |
Calcium concentration for all three freshly prepared solutions (17% EDTA, 17% carbonic acid, and 10% citric acid) was checked in different time interval of 10 min, 15 min, and 32 h. The results were expressed in mg/l. And the mean value for all the groups is given in [Table 1].
|Table 1: Mean values (mg/l) for calcium concentration for different time interval|
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The results [Table 1] showed that the maximum dissolution of calcium of about average 7.683 mg/l in 10 min, 7.968 in 15 min, and 8.430 mg/l in 32 h, from root dentin was by 17% carbonic acid. This may be due to the strong chelating effect. While 17% EDTA showed the second dissolution effect of about average of 5.218 mg/l in 10 min, 5.348 mg/l in 15 min, and 5.704 mg/l in 32 h. And 10% citric acid showed the minimum dissolution of about average of 4.032 mg/l in 10 min, 4.144 mg/l in 15 min, and 4.372 mg/l in 32 h. The results [Table 2] indicate that there was highly statistical difference between the groups. 17% carbonic acid showed the maximum dissolution followed by 17% EDTA and 10% citric acid, respectively [Figure 1].
|Figure 2: (a) Scaling of extracted tooth done. (b) Removal of cementum from outer surface. (c) Access opening and removal of pulpal tissue and irrigation with saline. (d) Hard tissue microtome. (e) Sectioning done with hard tissue microtome. (f) Sections placed in respective solutions. (g) Calcium level measured with atomic spectroscopy|
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When evaluated for different time interval 32 h had highly significant (P < 000.) difference compared to 15 min and 10 min in all the three groups [Table 3].
| Discussion|| |
The three freshly prepared solution used in this study were 17% carbonic acid, 17% EDTA, and 10% citric acid. These are the solutions generally used for the retrieval of MTA. MTA has shown to take minimum 10 and 15 min for dissolution.  The minimum time required for decalcifying effect of EDTA is 10 min and 15 min.  When MTA is exposed to MTAD (a root canal cleanser containing citric acid), its surface characteristics are altered. They also proposed that it might take 32 h for complete dissolution. Hence, we took even 32 h to check the dissolution effect on root dentin. , 15-17% EDTA is the most commonly used calcium chelator, which has inhibited the setting of MTA. The term chelate is originated from a Greek word 'chele' (crab claw). Chelation is a process that involves the uptake of multivalent positive ions by these agents. Chelating agents used in endodontics are in aid to preparation of narrow and calcified canals. In the specific case of dentin, the chelator reacts with the calcium ions in the hydroxylapatite crystals. This process can cause changes in the microstructure of the human dentin and changes in the Ca/P ratio. 
The calcium ions (Ca² + ) present in hydroxyapatite crystals are one of the main inorganic elements of dentin.  It has been reported that some chemicals used for endodontic irrigation are capable of causing alterations in the chemical composition of dentin. ,
Nygaard-Ostby (1957) used the principle of a constant solubility product to explain the demineralization of dental hard tissue by EDTA and its sodium salt.  An equilibrium is established between the saturated salt solutions and the consolidated precipitate because ions from the same time, ions from the solution are precipitated as solids. The concentration of the salt remains constant and therefore the product of concentrations of the ions in the solution at a given temperature (the solubility product) remains stable. The lyophobic substances such as dentine have mainly the mineral content as phosphate and calcium, which are soluble in water. When the disodium salt of EDTA is added to this equilibrium, calcium ions are removed from the solution.  This leads to the dissolution of further ions from dentine, so that the solubility product remains constant. Thus, chelator causes decalcification of dentine. 
The exchange of calcium from the dentine by hydrogen results in a subsequent decrease in pH. Due to the release of acid, the efficiency of EDTA decreases with time; on the other hand, the reaction of acid with hydroxyapatite affects the solubility of dentine. Chemically, two coexisting reactions can be distinguished; 
EDTAH 3− + Ca 2+ = EDTACa 2− + H
EDTAH 3− + H = EDTAH 2 2− (Perez et al. 1989).
As this reaction proceeds, acid accumulates and protonation of EDTA prevails, thus decreasing the rate of demineralization.
Morrison  described that carbonic acid (pH - 5.45) exposure significantly decreased the surface hardness of WMTA (white mineral trioxide aggregate) after day 1 and after 21 days of setting. 
The chemical reactions are as follows:
This reaction that occurs when carbonic acid is used as a solvent for WMTA is similar, but more aggressive than that reported by Holland.  They have observed that calcium hydroxide formed in MTA dissociates to calcium and hydroxyl ions when it comes in contact with tissue fluid. This calcium ion interacts with carbon dioxide in the tissues, forming calcium carbonate granulations and calcite crystals. In study conducted by Nandini et al.  the interaction of WMTA with carbonic acid was more aggressive, because the carbon dioxide released is not buffered, whereas in periapical tissues the fluids buffer the released carbon dioxide. While in our study, the carbonic acid released the highest amount of calcium ions due its chelating effect.
Citric acid, a weak organic acid, has been applied previously on root surfaces altered by periodontal disease and instrumentation in order to increase cementogenesis and to accelerate healing and regeneration of a normal periodontal attachment after flap surgery.  In endodontic research, Loel (1975) used 50% citric acid alternately with 5% NaOCl during instrumentation and found that it was an effective agent for removing necrotic tissue and preparing the dentine for subsequent sealing with endodontic sealers. Tidmarsh (1978) also reported that 50% citric acid irrigation was effective in the removal of the superficial smear layer.
To reduce the risk of section damage, cutting and precision grinding of the tooth specimens needs to be done with the teeth embedded in artificial resin. These hydrophobic materials cannot adhere to the tooth nor infiltrate it in its hydrated state. In increasing concentrations of alcohol, water is replaced by alcohol in stages. Resecting the tooth's root reduces the diffusion paths and exposure times. Microtome is a method for the preparation of thin sections for materials such as bones, minerals and teeth, and an alternative to electro-polishing and ion milling. 
Atomic absorption spectroscopy  is a spectroanalytical procedure for the quantitative determination of chemical elements employing the absorption of optical radiation (light) by free atoms in the gaseous state. The technique makes use of absorption spectrometry to assess the concentration of an analyte in a sample. In short, the electrons of the atoms in the atomizer can be promoted to higher orbitals (excited state) for a short period of time (nanoseconds) by absorbing a defined quantity of energy (radiation of a given wavelength).  This amount of energy, that is, wavelength, is specific to a particular electron transition in a particular element. In general, each wavelength corresponds to only one element, and the width of an absorption line is only of the order of a few picometers (pm), which gives the technique its elemental selectivity. Atomic absorption spectrophotometry was applied to the determination of calcium, It was shown that this technique is superior to existing methods in regard to accuracy, precision, and ease and speed of performance. 
The results of this study indicate there is significant difference between the three groups. 10% citric acid shows the least decalcification in dentin due to its less chelating effect followed by 17% EDTA and 17% carbonic acid, respectively.
| Conclusion|| |
Although MTA has revolutionized the field of endodontics with its biocompatible, hard tissue conductive, hard tissue inductive properties, the different acids, that is, 10% Citric acid, 17% EDTA and 17% carbonic acid that are used for its retrieval has proven to be harmful to the calcium content of the root dentin.
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[Figure 1], [Figure 2]
[Table 1], [Table 2], [Table 3]
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