| Abstract|| |
Aim: The purpose of the study was to evaluate and compare the sealing ability of mineral trioxide aggregate (MTA) with three different methods.
Materials and Methods: Forty single canal teeth were decoronated, and root canals were enlarged to simulate immature apex. The samples were randomly divided into Group M D = MTA-angelus mixed with distilled water and Group M C = MTA-angelus mixed with 2% chlorhexidine, and apical seal was recorded with glucose penetration method, fluid filtration method, and dye penetration methods and compared.
Results: The three methods of evaluation resulted differently. The glucose penetration method showed that M D sealed better than M C , but difference was statistically insignificant (P > 0.05). The fluid filtration method resulted that Group M C was statistically insignificant superior to Group M D (P > 0.05). The dye penetration method showed that Group M C sealed statistically better than Group M D .
Conclusion: No correlation was found among the results obtained with the three methods of evaluation. Addition of chlorhexidine enhanced the sealing ability of MTA according to the fluid filtration test and dye leakage while according to the glucose penetration test, chlorhexidine did not enhance the sealing ability of MTA. This study showed that relying on the results of apical sealing by only method can be misleading.
Keywords: Apical plug, dye penetration, fluid filtration, glucose penetration, mineral trioxide aggregate
|How to cite this article:|
Nikhil V, Jha P, Suri NK. Effect of methods of evaluation on sealing ability of mineral trioxide aggregate apical plug. J Conserv Dent 2016;19:231-4
|How to cite this URL:|
Nikhil V, Jha P, Suri NK. Effect of methods of evaluation on sealing ability of mineral trioxide aggregate apical plug. J Conserv Dent [serial online] 2016 [cited 2019 Nov 17];19:231-4. Available from: http://www.jcd.org.in/text.asp?2016/19/3/231/181938
| Introduction|| |
Open apex necessitates the use of an apical barrier to stop the egress of obturating material into the periapical tissues. A hard tissue barrier formation after achieving the disinfection of infected root canal with copious irrigation 0.5% NaOCl and Ca (OH) 2 is often carried out. Traditionally, Ca (OH) 2 was used to promote the formation of an apical barrier which may exceed to a period as long as 5-19 months  but the long-term exposure of root dentin to Ca (OH) 2 may cause it more prone for fracture. , In addition to the risk of fractures between appointment, possibility of recontamination by the dislocation of temporary filling, is additional problem associated.  Mineral trioxide aggregate (MTA), which is a powder aggregate containing a mineral oxide, is highly alkaline (pH 12.5 when set) thus antimicrobial and promotes hard tissue deposition, biocompatible, and has good sealing ability.  In addition, it has ability to set in the presence of moisture and blood. Andreasen et al. advocated the use of MTA for apexification over calcium hydroxide. Holt et al. advocated the use of 2% chlorhexidine gluconate instead of distilled water to enhance the antimicrobial activity of MTA.
Although substantivity and evidence are available for chlorhexidine, apart from dye penetration method, no other microleakage evaluation method so far has been used to determine whether the addition of 2% chlorhexidine will enhance the sealing ability of MTA.
Therefore, the objective of this study was to evaluate, in vitro, the apical sealing ability of MTA combined with either distilled water or 2% chlorhexidine solution, in simulated immature teeth, using glucose penetration, fluid filtration, and dye penetration methods.
| Materials and methods|| |
Forty single canal extracted teeth free of cracks, caries, and resorption with canal curvature between 0 and 10° and single apical foramen were collected, cleaned, and stored in 1% thymol solution. The length of each root was standardized to 17 mm by removing the coronal portion (3 mm coronal to the cementoenamel junction) and apical portions. Root canal instrumentation was performed using rotary ProTaper Files (Dentsply-Maillefer, Konstanz, Switzerland) till F4 as per manufacturer's instructions. For the simulation of teeth with immature apices, Peeso reamers between #1 and #6 were introduced in the root canals, and a #6 Peeso reamer was allowed to protrude 1 mm beyond the apex. The root canals were irrigated using 2 mL of 5.25% sodium hypochlorite after each file and a final flush with 5 mL of 17% EDTA to remove the smear layer. Finally, the root canals were flushed with distilled water and dried using paper points (Diadent, Diadent Group, International, Burnaby, BC, Canada). Ca (OH) 2 paste (Pulpdent, Watertown, MA, USA), used as an intracanal medicament, was introduced in the root canals using lentulo spirals (Dentsply-Maillefer, Konstanz, Switzerland). Orifices of the root canals were sealed with a temporary restorative material (Cavit, 3M, ESPE, USA). All samples were incubated for 7 days at 37°C under 100% humidity. After 7 days, the calcium hydroxide was retrieved using 17% EDTA in combination with ultrasonic agitation for one minute as recommended by Nandini et al. A final rinse with distilled water was performed. The samples were randomly divided into two groups on the basis of liquid mixed with MTA:
The powder to solution ratio used was 3:1 to achieve putty consistency. Four mm thick apical plug was placed, using hand pluggers. The density and the length of the plug were confirmed by radiographs. Any excess extruded from the apical portion was wiped away with a moist cotton pellet. A sterile cotton pellet moistened with sterile water was placed over the canal orifice, and the openings were sealed temporarily. The apices of the samples were covered with a wet cotton pellet, and the samples were kept for 12 h at 37°C under 100% humidity. All the external surface of sample except for 1 mm around the apical plug and canal orifice were covered with sticky wax. Temporary filling and cotton pellet were removed. The apical leakage was evaluated with different evaluation methods.
- Group M D : MTA mixed with distilled water
- Group M C : MTA mixed with 2% chlorhexidine.
Glucose penetration test
Samples were individually inserted into an Eppendorf tube (1.5 ml) with the apical 7 mm protruding through the end. The coronal part of each root was glued to the end of an Eppendorf vial using cyanoacrylate. A hole was created in the cap of the Eppendorf vial through which a plastic tube of at least 15 mm long was connected. A seal was obtained using cyanoacrylate glue. The lower portion of the Eppendorf was inserted into a glass vial containing distilled water in such a way that 3 mm of the root apex was immersed in the water. The distilled water was used here to inhibit the proliferation of microorganisms that might decompose glucose. The tracer used in this study was a 1 mol/L glucose solution (pH = 7.0/density = 1.09 × 103 g/L/viscosity = 1.18 × 10 -3 Pa/s at 37°C). Glucose has a low molecular weight of 180 Da and is hydrophilic and chemically stable. About 0.75 mL of the glucose solution was injected into the Eppendorf vial from the plastic tube. The upper part of the plastic tube containing glucose solution was connected to a pressure source to create a headspace pressure of 0.5 kg/cm 2 for 2 h. A 10 µL aliquot of the solution was drawn from the glass vial using a micropipette and then analyzed using a glucose kit in a vitros 250 dry acid analyzer. After analyzing all the 40 samples, they were rinsed with distilled water for the next technique.
Fluid filtration method
The apparatus consisted of the two-neck bottle with openings: One for gas (oxygen) and other for a micropipette with a 3-way tube (internal diameter 1 mm).The oxygen cylinder and micropipette were connected to the two-neck bottle with glass tubes. The glass tube connecting micropipette was immersed in distilled water placed in the two-neck bottle while the other glass tube connecting oxygen cylinder was fixed above the fluid level. The 3-way tube provided attachment for the tooth sample and two syringes. Test samples were mounted on a fluid transport model one at a time. Coronal end of each root was then connected to the filtration apparatus by cyanoacrylate. All pipettes, syringes, and the glass tubes of the device were filled with distilled water. Using a syringe, water was sucked back into the open end of the glass capillary, and an air bubble was created. A headspace pressure of 0.07 kg/cm 2 was supplied from the inlet side, through the micropipette to the coronal end of the canal, through the voids along the filling, thus displacing the air bubble in the capillary tube. After 3 h the movement of an air bubble was recorded at 2 min intervals for 8 min (2 min, 4 min, 6 min, and 8 min) for every sample. The linear movement of an air bubble in the micropipette was measured with a digital vernier caliper.
The samples were then removed from the fluid transport device and stored in 0.9% saline in between the study. After completion of fluid filtration test, the samples were washed with distilled water for the next technique.
Dye penetration method
The samples were completely coated with sticky wax except for the apical 1 mm of the resected root surface, where dye penetration was needed. Samples were placed in 0.2% rhodamine B for 24 h at 37°C and 100% relative humidity. After 24 h, samples were rinsed under running water for 5 min and were allowed to dry. After removal of the external layers of wax, the roots were divided into two equal halves along the long axis, in buccolingual direction with a double-sided diamond disc. The dye penetration was measured under a stereomicroscope at ×10 magnification.
| Results|| |
The mean values and standard deviation were calculated from the data collected [Table 1] and statistical analysis was performed using Student's t-test. Results of glucose penetration test showed more leakage in Group M C as compared to Group M D but the difference between was statistically insignificant (P > 0.05) [Table 2]. Fluid filtration test results showed less leakage in Group M C compare to M D although the difference was not statistically significant (P > 0.05). In dye leakage test, Group M C showed less leakage as compared to Group M D ; however, the difference was statistically significant (P < 0.05).
|Table 1: Mean and standard deviation of leakage values in groups evaluated with different methods |
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| Discussion|| |
MTA was developed at Loma Linda University in 1993. It consists of tricalcium silicate, tricalcium aluminate, tricalcium oxide, silicate oxide, and other mineral oxides forming a hydrophilic powder which sets in the presence of water. It is ranked with good sealability results in several studies. ,,,,
The antibacterial properties of MTA have been extensively evaluated. Several investigations reported that MTA has limited antimicrobial effect against some microorganisms. ,,, An investigation on facultative and strict anaerobic bacteria showed that MTA has an antibacterial effect on some facultative bacteria and no effect on any species of strict anaerobes.  In an another antimicrobial study on MTA and Portland cement against Staphylococcus aureus, Enterococcus faecalis, Pseudomonas aeruginosa, Bacillus subtilis, Candida albicans, and a mixture of these bacterial and fungal species, both materials exhibited diffusion in agar without inhibition of microbial growth. 
When antibacterial effects of different concentrations of gray MTA and white MTA were compared against E. faecalis and Staphylococcus sanguis, the former material required lower concentrations to produce the same antibacterial effect against each of the bacteria tested. In addition, E. faecalis requires a higher MTA concentration for growth inhibition than S. Sanguis. 
2% chlorhexidine is a disinfecting agent that is effective against various microorganisms, including E. faecalis. According to Stowe et al.,  MTA shows improved antimicrobial properties when mixed with 0.12% chlorhexidine instead of distilled water. MTA can be substituted with chlorhexidine for water when used as a retrofilling material, provided its sealing ability, and biocompatibility is confirmed.  Arruda et al. showed that there was no statistically significant difference in sealing ability of MTA when mixed with distilled water and chlorhexidine. Furthermore, Shahi et al.  did not observe differences in the sealing ability of gray and white MTA mixed with water or 0.12% chlorhexidine. They suggested that mixing MTA with chlorhexidine does not affect the sealing ability of the material. Holt et al. tested the effect of the addition of chlorhexidine to MTA on compressive strength and concluded that it could be recommended in areas exposed to minimal compressive forces thus it was used as apical barrier in this study.
Various techniques have been advocated for detection and evaluation of leakage around filling material such as dye penetration, , scanning electron microscope,  electrochemical,  bacterial penetration method,  fluid filtration, , and glucose penetration model  . In this study, MTA when mixed with chlorhexidine showed better sealing ability as compared to MTA mixed with distilled water, except in glucose penetration test, in which MTA mixed with distilled water showed better results as compared to MTA mixed with chlorhexidine.
MTA does not bond to dentin, interaction of the released calcium, and hydroxyl ions of MTA with a phosphate-containing synthetic body fluid results in the formation of apatite-like interfacial deposits. These deposits fill up any gaps induced during the material shrinkage phase and improve the frictional resistance of MTA to the root canal walls. The formation of this nonbonding, gap-filling apatite deposits probably also accounts for the seal of MTA. ,
The limitations of this study were that the applied pressure in glucose leakage model and fluid filtration model could not be maintained constant for all the samples and in dye leakage test, the samples were sectioned in one plane, which does not provide circumferential assessment for the leakage. Furthermore, the time periods of storage during tests were different.
| Conclusion|| |
From this study, it can be concluded that:
Further in vitro studies with larger sample size and in vivo studies must be carried out to evaluate not only the sealing ability but also several other mechanical and physical properties of such compounds, before its clinical use.
- MTA mixed with chlorhexidine showed superior sealing as compared to MTA mixed with distilled water with exception of glucose penetration test, in which MTA mixed with distilled water showed better results
- The three methods: Glucose penetration, fluid filtration, and dye penetration showed no agreement among each other.
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Conflicts of interest
There are no conflicts of interest.
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[Table 1], [Table 2]