Print this page Email this page | Users Online: 720
Home About us Editorial board Search Ahead of print Current issue Archives Submit article Instructions Subscribe Contacts Login 

 Table of Contents  
Year : 2023  |  Volume : 14  |  Issue : 1  |  Page : 11-16

In vitro evaluation of apical leakage of three bioceramic materials using glucose leakage model in a simulated open apex

Department of Conservative Dentistry and Endodontics, Terna Dental College and Hospital, Navi Mumbai, Maharashtra, India

Date of Submission28-Sep-2022
Date of Decision22-Jan-2023
Date of Acceptance23-Jan-2023
Date of Web Publication18-Mar-2023

Correspondence Address:
Dr. Sonal Sahu
Department of Conservative Dentistry and Endodontics, Terna Dental College and Hospital, Sector 22, Nerul, Navi Mumbai - 400 706, Maharashtra
Login to access the Email id

Source of Support: None, Conflict of Interest: None

DOI: 10.4103/srmjrds.srmjrds_126_22

Rights and Permissions

Background: Recently, new calcium silicate-based materials have been introduced as root-end filling material which offers improved color stability and handling characteristics, while exhibiting physical and chemical properties comparable with mineral trioxide aggregate (MTA). Aim: The purpose of this study was to compare the sealability of 3 calcium silicate-based materials - MTA Plus (Prevest Denpro, India), Biodentine (Septodont, St. Maurdes Fossés, France), and TotalFill BC RRM (FKG, Brasseler, Savannah, USA) in a simulated open apex model. Materials and Methods: Thirty-two recently extracted human maxillary anterior teeth with single, straight root canals were selected and after establishing the working length, canals were prepared by Protaper Gold system up to size F3. The apical end was resected at 3 mm to simulate an open apex. Prepared roots were divided into groups according to the root repair materials – MTA (Prevest Denpro, India), Totalfill Bioceramic Root Repair Material (FKG, Brasseler, Savannah, USA), and Biodentine (Septodont, St. Maurdes Fossés, France). The apical plug of the root filling material was placed in the orthograde direction and a glucose leakage model was used to study the sealability of the materials. Results: All three materials showed more leakage at 14 days than at 7 days, however, there was no statistically significant difference between them. Conclusion: All three bioceramic materials exhibited equal amounts of apical leakage at 7 and 14 days.

Keywords: Apexification, bioceramic material, microleakage, root repair material

How to cite this article:
Sahu S, Ramachandran N, Shetty R, Salgar A, Podar R, Singh S. In vitro evaluation of apical leakage of three bioceramic materials using glucose leakage model in a simulated open apex. SRM J Res Dent Sci 2023;14:11-6

How to cite this URL:
Sahu S, Ramachandran N, Shetty R, Salgar A, Podar R, Singh S. In vitro evaluation of apical leakage of three bioceramic materials using glucose leakage model in a simulated open apex. SRM J Res Dent Sci [serial online] 2023 [cited 2023 May 31];14:11-6. Available from:

  Introduction Top

Endodontic therapy aims to completely eliminate bacteria from the root canals and prevent or treat apical periodontitis.[1] The endodontic management of teeth with open apices is not straightforward. Open apices, blunderbuss canals, and teeth with anomalies such as dens invaginatus pose a challenge during obturation due to the lack of a definite apical stop.[2] However, the outcome may be more predictable in such cases with the advent of newer biocompatible materials with better handling properties.

There are several methods of managing necrotic teeth with a wide-open apex. Some of these include a customized blunt-ended gutta-percha cone such as in the rolled cone technique[3] or the inverted cone technique wherein a bigger-sized gutta-percha cone is inserted into the canal in an “inverted” fashion or underobturating the root canal,[4] regenerative procedures using autologous platelet matrices such as platelet-rich plasma, platelet-rich fibrin, injectable plasma-rich fibrin, or even periradicular surgery.[5] However, surgery may result in an unfavorable crown-root ratio, and at the same time leave behind thin, irregular dentinal walls at the root apex, therefore precluding its use in such cases.

Conventionally, the material of choice for performing apexification in immature, nonvital teeth was calcium hydroxide. However, the long-term use of calcium hydroxide has been associated with weakening the already thin dentinal walls.[6] Reduced patient compliance owing to multiple visits, esthetic concerns, and increased risk of microleakage were some other associated disadvantages.[7] After the discovery of mineral trioxide aggregate (MTA) in 1993, there has been a paradigm shift in the treatment of necrotic teeth with open apices. However, MTA has some drawbacks, such as poor handling characteristics and potential discoloration of the teeth.[8],[9],[10] More recently, novel calcium silicate-based materials have been used owing to their better biocompatibility, sealability, and mechanical properties. In addition, these materials possess better working characteristics, color stability, shorter setting time compared to MTA, and comparable physical and chemical properties to MTA. BioDentine (Septodont, St. Maurdes Fossés, France) is one such material, among many like TotalFill BC Root Repair material (FKG, Switzerland) also known as Endosequence BC RRM (Brasseler USA, Savannah, GA, USA,), MTA Repair HP (Angelus, Brazil) iRoot BP (Innovative BioCeramix Inc., Vancouver, Canada).

BioDentine (Septodont, St. Maurdes Fossés, France) is made of tricalcium silicates, dicalcium silicates, calcium carbonate, iron oxides, zirconium oxide, and oxide fillers. It has a clinical setting time of 12 min, relatively shorter compared to MTA which can take up to 24 h to set, does not stain the tooth, releases more calcium ions than other calcium silicate materials, and has excellent biocompatibility.[11],[12] TotalFill BC RRM (FKG, Switzerland) is a novel calcium silicate-based material that is composed of calcium silicates and phosphates, calcium hydroxide, zirconium oxide, fillers, and thickening agents. It is provided in a premixed, ready-to-use container with a putty-like consistency that makes it easy to pack in root end-filling procedures. There is enhanced hydroxyapatite crystal formation due to the presence of an additional agent called calcium phosphate monobasic.[13]

While achieving a “fluid-tight seal” is still predictable in mature teeth, the sealing ability of obturating materials plays a significant role in teeth with open apices to prevent apical microleakage. Therefore, this study aimed to compare the sealability of three calcium silicate-based materials – MTA Plus (Prevest Denpro, India), Biodentine (Septodont, St. Maurdes Fossés, France), and TotalFill BC RRM (FKG, Switzerland) in a simulated open apex model.

  Materials and Methods Top

Study design

The current study was an experimental in vitro study design.

Study setting

The study was conducted in the department of conservative dentistry and endodontics of a private dental college and hospital in collaboration with metallurgical engineering and material sciences at a reputed research engineering institute.

Study population/specimens

Thirty-two human-extracted maxillary anterior teeth with single, straight canals were collected and used in this study. Teeth with multiple canals, open apices, calcification, large carious lesions, and internal or external resorption were excluded from the sample selection process. The Institutional Ethical Committee approved the study (TDC/EC/22/2020).

Sample size

Considering the mean and standard deviation (SD) values from the literature, the sample size was calculated with 80% power using the formula as


where Zα is the Z variate of alpha error, i.e., a constant with a value of 1.96, Zβ having a value of 0.84. standard deviation (SD) and mean differences, the values of which are taken from the parent article. According to the results, nine samples per group were needed to be taken for this study (n = 9).

Specimen preparation

Each specimen was standardized to a length of 15 mm with a scale and then decoronated with the help of a diamond disc under water cooling. Access was gained with a round diamond bur (Mani, India). Apical patency was confirmed and working length was established, first by visualizing a #15K file (Mani, India) tip under magnification loupes as it came out of the apical foramen, pulled back by 0.5 mm, and then verified radiographically. At the established working length, the canals were instrumented with the Protaper Gold system till-size F3 (Dentsply Maillefer, Ballaigues, Switzerland). One milliliter of 2.5% sodium hypochlorite was used as the working solution. The final irrigation protocol comprised 3 ml of 17% EDTA for 1 min, 3 ml of 2.5% sodium hypochlorite, and finally rinsing with saline.

The apical 3 mm of all the specimens were measured with a ruler, marked, and then resected with a diamond disc in a direction perpendicular to the long axis of the tooth. A #60 K-File (Dentsply Maillefer, Ballaigues, Switzerland) was used to enlarge the apical end and standardize the size of the open apex. The prepared specimens were then randomly distributed into three groups (total n = 30), according to the apical plug materials (n = 10), the canals were dried with paper points and subsequently, a 5 mm apical plug was created in all the specimens with one of each test materials. The positive control specimen (n = 1) was not obturated while the negative control specimen (n = 1) was completely coated with nail varnish except for the apical foramen. The apical plug materials were manipulated following the manufacturer's instructions. They were then divided as follows:

  • Group 1 - MTA Plus (Prevest Denpro, India)
  • Group 2 - Totalfill BC RRM (FKG, Switzerland)
  • Group 3 - Biodentine (Septodont, St. Maurdes Fossés, France).

MTA was manipulated as per the manufacturer's instructions and filled into the canals with an MTA gun (OraCraft, India). TotalFill and Biodentine were directly condensed into the canal with an amalgam carrier and hand pluggers. Radiographs were taken to confirm uniform and dense obturation. After obturation, all the specimens were covered in wet gauze for about 72 h for the complete set of materials. A glucose-leakage model was used to assess the sealability of the test materials. The model preparation was performed as described by Xu et al.[14] [Figure 1].
Figure 1: (a) Glucose leakage model used in the experiment; (b) Samples of three groups

Click here to view

}The prepared models were then placed in an incubator at 37°C at 100% humidity till they were subsequently analyzed by a spectrophotometer. To ensure no evaporation of the solution occurred, a similar model was incubated under similar conditions and weighed separately each day until further analysis was done. An aliquot of solution of 800 μL was drawn from the 5 ml plastic bottle using a micropipette and analyzed in a UV/VIS spectrophotometer (Agilent technologies), performing an assay at 480 nm. All the values obtained were plotted on a graph and mean values were deduced which were subsequently used for statistical analysis.

Statistical analysis

The statistical analysis was performed using SPSS software (BM Corp. Released 2017, IBM SPSS Statistics for Windows, Version 25.0. Armonk, NY, USA: IBM Corp.). The intergroup comparison at 7 and 14 days was performed using the ANOVA test. A paired t-test was applied for analysis in each group at 7 and 14 days. The statistical difference was established when P < 0.05.

  Results Top

All the readings obtained were plotted on a graph and mean values were deduced which were subsequently used for statistical analysis. The mean values mentioned in [Table 1] and [Table 2] are the mean values obtained from a graph that was plotted after the spectrophotometric analysis. The graph was plotted based on the wavelength at which the glucose detection is maximum (suggestive of microleakage). The maximum amount of leakage was observed in the positive control group while the negative control group showed no leakage, thereby validating the glucose leakage model used. The intergroup comparison of the three materials at 7 and 14 days is presented in [Table 1]. All three materials comparatively showed more leakage at 14 days than at 7 days; however, this was not statistically significant. In the intra-group comparison at 7 and 14 days [Table 2], there was no statistically significant difference between the three materials.
Table 1: Intergroup comparison of microleakage at 7 and 14 days

Click here to view
Table 2: Intragroup comparison of microleakage at 7 and 14 days

Click here to view

  Discussion Top

A wide variety of materials have been used for apexification. Kraiser in 1964 was the first to suggest the use of calcium hydroxide mixed with camphorated parachlorophenol (CMCP) to induce a calcific barrier. Calcium hydroxide with an alkaline pH of 12, and the ability to inactivate the lipopolysaccharide component of the cell wall[15] became the material of choice for such teeth but it had its own disadvantages.

Since the discovery of MTA in 1993, the drawbacks of calcium hydroxide have been largely addressed. Several studies have compared MTA with calcium hydroxide in the treatment of teeth with open apices.[7],[16],[17],[18] Apexification procedures with MTA took a significantly lesser number of appointments,[19] showed more predictable healing in the long term, and did not lead to coronal or radicular fracture.[18],[20] MTA since has become the material of choice for managing nonvital immature teeth. Its excellent biocompatibility, property to induce osteogenesis, cementogenesis, and the ability to form a good apical seal are particularly conducive to its use in vital pulp therapy as well as regenerative endodontic procedures. While MTA is chemically and mechanically superior to calcium hydroxide, it has disadvantages as well. Advancements in material sciences have led to the development of bioactive, biocompatible bioceramic materials with improved handling and shorter setting time that do not cause any discoloration. The availability of such materials in premixed containers with a putty-like consistency is an added advantage. Not much is known about the sealability of novel calcium silicate-based materials when compared to MTA in a simulated open-apex model.

Hence for this study, two novel calcium silicate-based materials were chosen - Biodentine and TotalFill BC RRM and compared with the time-tested MTA. Biodentine was chosen for the study as it is equally biocompatible as MTA and has shown a superior sealing ability to MTA.[21] It sets in 12 min as compared to 45 min to 4 h for MTA. Although studies are comparing MTA and Biodentine in open apex cases,[16],[22],[23],[24],[25] there has been just one study evaluating the marginal adaptation of TotalFill BC RRM, in comparison to ProRoot MTA and Biodentine.[13] TotalFill BC RRM as mentioned by the manufacturer has several applications such as in vital pulp therapy procedures, perforation repair, management of internal and external resorption, root-end filling in periradicular surgeries, and apexification. The presence of monobasic calcium phosphate in its composition and the ability to nucleate B-type carbonated apatite could result in good sealing at the material-dentin interface and promote periodontal regeneration of tissues.[25]

There are several methods of determining microleakage such as dye penetration tests, chemical tracer method, radioactive isotope method, bacteriological method, fluid filtration method, scanning electron microscopy, and confocal laser scanning microscopy. However, none of them has been regarded as the best method for evaluating the sealability of root-end filling materials due to several limitations. These include the unstable nature of tracer dyes/isotopes, inconsistent results obtained due to the complexity of the technique, and subjective analysis of the samples. A glucose leakage model as described by Xu et al.[14] was chosen as the technique of evaluation because it was possible to quantify the amount of leakage taking place over the experimental period.

Glucose, having a low-molecular weight of 180 Da, was the tracer used in the experimental setup. Glucose is considered an acceptable tracer with clinical relevance because of its small molecular size (smaller the molecule, stricter the test) and its being a source of nutrition for the bacteria. In the presence of glucose, microorganisms that persist in the canals after root canal treatment could thrive on glucose causing periapical inflammation.[14] Thus, the objective was to evaluate the presence of glucose that seeped through the obturated tooth and collected in the 5 ml glass bottle. To prevent the decomposition of glucose by bacteria, sodium azide was added as a preservative. Subsequently, the sealability of the root end filling materials was evaluated by spectrophotometry.

Spectrophotometry is a method that measures the intensity and the amount of light absorbed by a chemical substance, as a beam of light passes through a sample solution. As the spectrophotometer detects the material in the visible wavelength, peaks are formed on a graph, indicating the amount of substance in the sample, in this case, glucose.

The results of the present study show no statistical difference between the three root-end filling materials. More leakage was observed at 14 days than at 7 days in all three groups, however, at the end of 14 days, equal leakage was observed in all three groups. This is in accordance with a previous study, that showed comparable sealability of calcium silicate-based materials.[26] The present study used injectable form while the study by Juez et al. used Total Fill BC in putty form. The microleakage was evaluated at only two intervals, i.e., 7 days and 14 days which is more practical, as opposed to the study done by Juez et al. The results of a single study will be insufficient to conclude which material has superior sealability in orthograde apical plug creation. And thus, more research needs to be done in a similar direction.

No significant differences were found in the sealing ability of MTA with other materials such as biodentine, and CEM cement when evaluated by the fluid filtration method.[27],[28] However, it exhibited the least leakage as compared to Biodentine and Resin modified GIC when evaluated by a dye penetration test.[29] On the contrary, a study by Pereira et al. concluded that the sealability of White ProRoot MTA was better than TotalFill BC RRM when evaluated by the radioisotope method.[30] This could be attributed to the difference in the method of evaluation of leakage. Delikan and Aksu, in their study, found the sealing ability of BC RRM-FS significantly lower than NeoMTA. They evaluated the leakage only at a single interval (of 48 h) and under dry conditions which do not simulate the actual oral environment.[31]

  Conclusion Top

Thus, within the limitations of this study, there was no significant difference in the sealing ability of all three materials. Newer calcium silicate-based materials are comparable in their sealability to MTA but do not have long-standing clinical evidence as MTA. Therefore, further clinical research is required in this area.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

  References Top

Mamootil K, Messer HH. Penetration of dentinal tubules by endodontic sealer cements in extracted teeth and in vivo. Int Endod J 2007;40:873-81.  Back to cited text no. 1
Das T, Gupta S, Atom J, Lairenlakpam R, Gupta S, Chaudhary A. Endodontic management of blunderbuss canal with open apex and Ellis Class IV fracture using mineral trioxide aggregate and compo-post. Int J Prev Clin Dent Res 2019;6:66-8.  Back to cited text no. 2
  [Full text]  
Pollack JR. Endodontia for non-vital teeth with incompletely formed roots. Bull NJ Soc Dent Child 1967;4:2-6.  Back to cited text no. 3
Moodnick R. Clinical correlation of the development of the root apex and surrounding structures. Oral Surg Oral Med Oral Pathol 1963;16:600-7.  Back to cited text no. 4
Witherspoon DE, Ham K. One-visit apexification: Technique for inducing root-end barrier formation in apical closures. Pract Proced Aesthet Dent 2001;13:455-60.  Back to cited text no. 5
Andreasen JO, Farik B, Munksgaard EC. Long-term calcium hydroxide as a root canal dressing may increase risk of root fracture. Dent Traumatol 2002;18:134-7.  Back to cited text no. 6
Holden DT, Schwartz SA, Kirkpatrick TC, Schindler WG. Clinical outcomes of artificial root-end barriers with mineral trioxide aggregate in teeth with immature apices. J Endod 2008;34:812-7.  Back to cited text no. 7
Torabinejad M, Parirokh M. Mineral trioxide aggregate: A comprehensive literature review – Part II: Leakage and biocompatibility investigations. J Endod 2010;36:190-02.  Back to cited text no. 8
Parirokh M, Torabinejad M. Mineral trioxide aggregate: A comprehensive literature review – Part I: Chemical, physical, and antibacterial properties. J Endod 2010;36:16-27.  Back to cited text no. 9
Parirokh M, Torabinejad M. Mineral trioxide aggregate: A comprehensive literature review – Part III: Clinical applications, drawbacks, and mechanism of action. J Endod 2010;36:400-13.  Back to cited text no. 10
Grech L, Mallia B, Camilleri J. Characterization of set intermediate restorative material, biodentine, bioaggregate and a prototype calcium silicate cement for use as root-end filling materials. Int Endod J 2013;46:632-41.  Back to cited text no. 11
López-García S, Myong-Hyun B, Lozano A, García-Bernal D, Forner L, Llena C, et al. Cytocompatibility, bioactivity potential, and ion release of three premixed calcium silicate-based sealers. Clin Oral Investig 2020;24:1749-59.  Back to cited text no. 12
Lertmalapong P, Jantarat J, Srisatjaluk RL, Komoltri C. Bacterial leakage and marginal adaptation of various bioceramics as apical plug in open apex model. J Investig Clin Dent 2019;10:e12371.  Back to cited text no. 13
Xu Q, Fan MW, Fan B, Cheung GS, Hu HL. A new quantitative method using glucose for analysis of endodontic leakage. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2005;99:107-11.  Back to cited text no. 14
Marinho AC, To TT, Darveau RP, Gomes BP. Detection and function of lipopolysaccharide and its purified lipid A after treatment with auxiliary chemical substances and calcium hydroxide dressings used in root canal treatment. Int Endod J 2018;51:1118-29.  Back to cited text no. 15
Sogukpinar A, Arikan V. Comparative evaluation of four endodontic biomaterials and calcium hydroxide regarding their effect on fracture resistance of simulated immature teeth. Eur J Paediatr Dent 2020;21:23-8.  Back to cited text no. 16
Lin JC, Lu JX, Zeng Q, Zhao W, Li WQ, Ling JQ. Comparison of mineral trioxide aggregate and calcium hydroxide for apexification of immature permanent teeth: A systematic review and meta-analysis. J Formos Med Assoc 2016;115:523-30.  Back to cited text no. 17
Kandemir Demirci G, Kaval ME, Güneri P, Çalışkan MK. Treatment of immature teeth with nonvital pulps in adults: A prospective comparative clinical study comparing MTA with Ca(OH)(2). Int Endod J 2020;53:5-18.  Back to cited text no. 18
Panda P, Mishra L, Govind S, Panda S, Lapinska B. Clinical outcome and comparison of regenerative and apexification intervention in young immature necrotic teeth-A systematic review and meta-analysis. J Clin Med 2022;11:3909.  Back to cited text no. 19
Mente J, Hage N, Pfefferle T, Koch MJ, Dreyhaupt J, Staehle HJ, et al. Mineral trioxide aggregate apical plugs in teeth with open apical foramina: A retrospective analysis of treatment outcome. J Endod 2009;35:1354-8.  Back to cited text no. 20
Tang JJ, Shen ZS, Qin W, Lin Z. A comparison of the sealing abilities between Biodentine and MTA as root-end filling materials and their effects on bone healing in dogs after periradicular surgery. J Appl Oral Sci 2019;27:e20180693.  Back to cited text no. 21
Ürkmez EŞ, Pınar Erdem A. Bioactivity evaluation of calcium silicate-based endodontic materials used for apexification. Aust Endod J 2020;46:60-7.  Back to cited text no. 22
Cechella B, de Almeida J, Kuntze M, Felippe W. Analysis of sealing ability of endodontic cements apical plugs. J Clin Exp Dent 2018;10:e146-50.  Back to cited text no. 23
Refaei P, Jahromi MZ, Moughari AA. Comparison of the microleakage of mineral trioxide aggregate, calcium-enriched mixture cement, and Biodentine orthograde apical plug. Dent Res J (Isfahan) 2020;17:66-72.  Back to cited text no. 24
Zamparini F, Siboni F, Prati C, Taddei P, Gandolfi MG. Properties of calcium silicate-monobasic calcium phosphate materials for endodontics containing tantalum pentoxide and zirconium oxide. Clin Oral Investig 2019;23:445-57.  Back to cited text no. 25
Juez M, Ballester ML, Berástegui E. In vitro comparison of apical microleakage by spectrophotometry in simulated apexification using white mineral trioxide aggregate, total fill bioceramic root repair material, and bio dentine. J Conserv Dent 2019;22:237-40.  Back to cited text no. 26
[PUBMED]  [Full text]  
Tabrizizade M, Asadi Y, Sooratgar A, Moradi S, Sooratgar H, Ayatollahi F. Sealing ability of mineral trioxide aggregate and calcium-enriched mixture cement as apical barriers with different obturation techniques. Iran Endod J 2014;9:261-5.  Back to cited text no. 27
Mousavi SA, Khademi A, Soltani P, Shahnaseri S, Poorghorban M. Comparison of sealing ability of ProRoot mineral trioxide aggregate, biodentine, and ortho mineral trioxide aggregate for canal obturation by the fluid infiltration technique. Dent Res J (Isfahan) 2018;15:307-12.  Back to cited text no. 28
Nikoloudaki G, Kontogiannis T, Meliou H, Kerezoudis N. A comparative in-vitro study of sealing ability of four different materials used in furcation perforation. Open J Stomatol 2014;4:402-11.  Back to cited text no. 29
Pereira IR, Carvalho C, Paulo S, Martinho JP, Coelho AS, Paula AB, et al. Apical sealing ability of two calcium silicate-based sealers using a radioactive isotope method: An in vitro apexification model. Materials (Basel) 2021;14:6456.  Back to cited text no. 30
Delikan E, Aksu S. Comparison of the sealing ability of apical plug materials in simulated open apices: An in vitro study. J Oral Res Rev 2020;12:70-5.  Back to cited text no. 31
  [Full text]  


  [Figure 1]

  [Table 1], [Table 2]


Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
Access Statistics
Email Alert *
Add to My List *
* Registration required (free)

  In this article
Materials and Me...
Article Figures
Article Tables

 Article Access Statistics
    PDF Downloaded71    
    Comments [Add]    

Recommend this journal