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 Table of Contents  
Year : 2017  |  Volume : 8  |  Issue : 3  |  Page : 391-394  

Effect of different irrigating solutions on depth of penetration of sealer into dentinal tubules: A confocal microscopic study

Department of Conservative Dentistry and Endodontics, Government Dental College and Hospital, Vijayawada, Andhra Pradesh, India

Date of Web Publication14-Sep-2017

Correspondence Address:
Murali Mohan Thota
Department of Conservative Dentistry and Endodontics, Government Dental College and Hospital, Vijayawada - 520 004, Andhra Pradesh
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/ccd.ccd_114_17

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Aim: The aim of the study was to evaluate the effect of different irrigating solutions used in final irrigation on depth of sealer penetration into dentinal tubules. Materials and Methods: Thirty recently extracted, human mandibular premolar teeth with single canals were randomly divided into two groups, and one of the two irrigants was used in each group - Group A (Chitosan) and Group B (Ethylenediaminetetraacetic acid). All the teeth were obturated with gutta-percha and AH 26® sealer labeled with fluorescent dye. The teeth were sectioned at distances 2, 5, and 8 mm from the root apex. Maximum depth of sealer penetration was measured using confocal laser scanning microscopy. Statistical Analysis: Statistical analysis used One-way analysis of variance and t-test. Results: At coronal third depth, the sealer penetration was greater in ethylenediaminetetraacetic acid (EDTA) group; however, depth of sealer penetration was greater at apical third in chitosan group. Conclusion: Final irrigation with EDTA and chitosan after the use of sodium hypochlorite affected sealer penetration.

Keywords: Chitosan, ethylenediaminetetraacetic acid, sealer penetration

How to cite this article:
Thota MM, Sudha K, Malini D L, Madhavi SB. Effect of different irrigating solutions on depth of penetration of sealer into dentinal tubules: A confocal microscopic study. Contemp Clin Dent 2017;8:391-4

How to cite this URL:
Thota MM, Sudha K, Malini D L, Madhavi SB. Effect of different irrigating solutions on depth of penetration of sealer into dentinal tubules: A confocal microscopic study. Contemp Clin Dent [serial online] 2017 [cited 2020 Aug 12];8:391-4. Available from:

   Introduction Top

Creation of a sterile space is impossible in infected root canals using mechanical preparation alone due to the complex anatomy of root canal systems.[1] Almost half of the root canal walls were left unprepared during the mechanical preparation with traditional stainless steel hand instruments and current nickel–titanium instrumentation systems.[2] When dentin is planed by endodontic instruments, a smear layer forms. Smear layer containing bacteria or bacterial products might provide a reservoir of irritants. Thus complete removal of the smear layer would be consistent with elimination of irritant from the root canal.[3] This marks irrigation, an essential step in the root canal preparation.[4] There was a significantly greater pattern of reduction of bacteria when NaOCl was used as an irrigant, compared with sterile saline.[5] The most widely used irrigant for smear layer removal is ethylenediaminetetraacetic acid (EDTA). It aids in cleaning of root canal by acting on the inorganic material (calcium ions) in dentin resulting in chelation of calcium. Thereby, it promotes the decalcification of dentin. Within 5 min, a depth of 20–30 μm can be decalcified. As the concentration of EDTA is increased in lakes and rivers, because of its overuse researchers are looking for alternatives. EDTA is also considered as a pollutant as it is not normally found in the nature.

The search for better biocompatible solutions which have less detrimental effects on periapical tissues is still going on. Chitosan is a natural polysaccharide which is biocompatible, biodegradable, and bioadhesive which lacks toxicity. It chelates various metal ions in acidic conditions and hence got popular in industrial areas for removal or recovery of metal ions. It is derived from deacetylation of chitin, which is found in shrimp and crab shells (Kurita, 1998). It has become popular due to its abundance in nature and low production costs.[6] Most of the studies, using chitosan as irrigating solution, have focused on antimicrobial activity with very little literature comparing the efficacy of the same with EDTA on smear layer removal and depth of sealer penetration.

The goal of thorough canal obturation can be achieved not only by removing the root canal debris and contaminates but also by achieving high adaptability of the filling materials.[7] Sealer is used to fill spaces between the core materials and the canal walls and penetrates into dentinal tubules entombing residual bacteria. Sealers have an antibacterial effect which is even more helpful as they can penetrate into the dentinal tubules and control the bacteria located in there, if not kill them. The above reason makes it really important that the percentage of the sealer/dentin interface that is covered by the sealer and the degree of tubule penetration by the sealer is as much as possible. Sealer penetration, therefore, could serve as an indicator to know the extent of smear layer removed.[8],[9]

Based on the above information, this study was designed to know the effect of different final irrigating solutions on the maximum depth of sealer penetration into dentinal tubules at the coronal, middle, and apical areas of the root canal.

   Materials and Methods Top

Thirty recently extracted human mandibular premolar teeth with single canals were used in this study. Preoperative radiographs were obtained in the mesiodistal and buccolingual directions to interpret the presence of calcifications, root curvatures, and a number of root canals. All the teeth were decoronated at cement-enamel junction, and working length was determined with the help of #10 k-file. Chemomechanical preparation was performed in a step-back technique using K-Files (Dentsply-Maillefer, Ballaigues, Switzerland) and 2.5% NaOCl as an irrigant after the use of each file.

In the apical area, the canals were enlarged up to size #40 and stepped back to size #60. The teeth were randomly divided into two groups according to the final irrigant employed – Group A (Chitosan-Panvo organics Pvt. Ltd., Chennai, India) and Group B (EDTA – Endo-Cleanse, Roydent Dental, Johnson, TN, USA). The final irrigation sequence was as follows:

  1. Chitosan group: 5 ml of 0.2% chitosan followed by 5 ml of 2.5% NaOCl for 1 min
  2. EDTA group: 5 ml of 17% EDTA followed by 5 ml of 2.5% NaOCl for 1 min.

The root canals were finally irrigated with 5 ml of distilled water and then dried with paper points. Obturations of the canals were done with AH 26® sealer (Dentsply-DeTrey, Konstanz, Germany) and gutta-percha in lateral compaction technique. Nearly 0.2% fluorescent Rhodamine B isocyanate (Sigma– Aldrich, India) was added to AH 26® sealer for fluorescence under confocal laser microscopy. Excess gutta-percha was removed, and access cavities were sealed with Cavit, after which the teeth were stored in an incubator at 37°C and 100% humidity for 24 h to allow the sealer to set. Each tooth was sectioned horizontally at a distance of 2, 5, and 8 mm from the root apex. Sections were made at a thickness of 100 μ. The specimens were then mounted onto glass slides and examined with a Leica TCS-SPE confocal microscope (Olympus FV 100, Germany). Depth of penetration was measured from the canal wall to the point of maximum sealer penetration. Statistical analysis was performed using one-way analysis of variance (ANOVA). The level of significance was set at P < 0.05.

Statistical methods

The mean and standard deviation of depth of sealer penetration of samples were obtained for each group and comparisons were performed using one-way ANOVA and t-test.

   Results Top

Despite the results showing nonsignificant difference in depth of sealer penetration at coronal and middle third between Group A (chitosan) and Group B (EDTA), a statistically significant difference was observed at apical third (P < 0.05). In both experimental groups, the maximum depth of the sealer penetration was better in the coronal third than in the apical third of root canals [Table 1].
Table 1: Intergroup comparison of depth of sealer penetration between two groups at three different levels by Independent t-test

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   Discussion Top

The results of the present study showed that the maximum depth of the sealer penetration was better in the coronal thirds than in the apical thirds of root canals in all experimental groups [Table 1]. This might be a result of better removal of the smear layer in coronal thirds than in apical thirds of root canals. The number of dentinal tubules in the coronal third is more in number with a larger diameter than those in the apical area.[8] EDTA is effective in smear layer removal from both coronal and middle thirds but not from the apical third.[10] A 1 min application of 7% maleic acid followed by 2.5% NaOCl is an effective final irrigant for the removal of the smear layer from the root canal system in the apical third, which is a crucial area for the disinfection of the root canal system than that of 17% EDTA.[11],[12] This may be due to the higher surface tension of 17% EDTA when compared with maleic acid. Galler et al. found that irrigation with EDTA may lead to exposure of growth factors in dentinal tubules which might optimize the conditions for cellular differentiation, tissue formation, and regeneration.[8] The search for more biocompatible solutions than EDTA, aiming at reducing its harmful effect on periapical tissues continues.

Atomic absorption spectrophotometry with flame analysis of 0.2% chitosan disclosed a calcium ion concentration of 104.13 mg, with no significant difference when compared with 15% EDTA (121.80 mg). When two materials have a similar chelating effect, then the cost-effective agent with less concentration should be preferred. It is known that the efficiency of a chelating agent depends on several factors including application time, pH, concentration, and amount of the solution.[13] In addition, the relationship between the concentration of the chelating agent and the application time seems to be important since it was found that highly concentrated solutions applied for a long period, cause roughness of dentin surface.[14] To avoid these detrimental effects with the use of high concentration solution, lower concentration solution was used (0.2% chitosan) in the present study.

According to the results of the present study, the depth of sealer penetration was highest with 0.2% chitosan at apical third and showed statistically similar results with 17% EDTA at coronal and middle third [Table 2]. A volume of 0.2% chitosan solution, even in such a low concentration, was capable of removing smear layer and provides statistically similar results to those of the solutions with higher concentrations.[6] Chitosan polymer is hydrophilic which favors intimate contact with root canal dentin and is adsorbed to root canal wall. In addition, it has a large number of free hydroxyl and amino groups that make it cationic in nature that is responsible for the ionic interaction between the dentin calcium ions and the chelating agent. Moreover, in an acid medium, the amino groups present in the polymer are protonated, resulting in attraction to other molecules for adsorption to root dentin to occur and were capable of being delivered to deeper location of dentinal tubules.[15] In addition, chitin polysaccharide, the precursor to chitosan, is the most abundant substance in nature after cellulose.[16] The production cost of chitosan is considered to be low, making its use ecologically attractive. Depth of penetration of sealer is also influenced by its chemical and physical properties.
Table 2: Mean values of depth of sealer penetration in two groups at three different levels by one-way analysis of variance

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Sealer may be drawn into tubules due to the capillary action and not by hydraulic forces created during root canal filling. Among the sealers tested, AH 26® sealer appeared to have the most optimal tubular penetration and adaptation to the root canal wall. To observe the penetration under confocal laser scanning microscope, labeling the sealer with Rhodamine B is essential. According to American Dental Association specifications, the sealer labeled with 0.2% Rhodamine B did not show changes in its flow. Future studies are required to investigate in detail the physical, chemical, and biological properties of chitosan to verify its benefits and consequences to humans.

   Conclusion Top

Within the limitations of the present study, it can be concluded that the depth of penetration of sealer at the apical third is greater with chitosan when compared to EDTA and is statistically significant. There is no significant difference between EDTA and chitosan in terms of the depth of penetration of sealer at the coronal and middle thirds. Chitosan (0.2%) can be indicated as a better root canal irrigant alternative to EDTA owing to its biocompatible, biodegradable, and bioadhesive nature.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

   References Top

Siqueira JF Jr., Rôças IN, Santos SR, Lima KC, Magalhães FA, de Uzeda M, et al. Efficacy of instrumentation techniques and irrigation regimens in reducing the bacterial population within root canals. J Endod 2002;28:181-4.  Back to cited text no. 1
Suzuki S. The study of chitosan-citrate solution as a root canal irrigant: A preliminary report. Oral Hyg Health 2014;2:142.  Back to cited text no. 2
Drake DR, Wiemann AH, Rivera EM, Walton RE. Bacterial retention in canal walls in vitro: Effect of smear layer. J Endod 1994;20:78-82.  Back to cited text no. 3
Hulsmann M, Peters O, Dummer P. Mechanical preparation of root canals: Shaping goals, techniques and means. Endod Top 2005;10:30-76.  Back to cited text no. 4
Shuping GB, Orstavik D, Sigurdsson A, Trope M. Reduction of intracanal bacteria using nickel-titanium rotary instrumentation and various medications. J Endod 2000;26:751-5.  Back to cited text no. 5
Silva PV, Guedes DF, Nakadi FV, Pécora JD, Cruz-Filho AM. Chitosan: A new solution for removal of smear layer after root canal instrumentation. Int Endod J 2013;46:332-8.  Back to cited text no. 6
Ordinola-Zapata R, Bramante CM, Graeff MS, del Carpio Perochena A, Vivan RR, Camargo EJ, et al. Depth and percentage of penetration of endodontic sealers into dentinal tubules after root canal obturation using a lateral compaction technique: A confocal laser scanning microscopy study. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2009;108:450-7.  Back to cited text no. 7
Kara Tuncer A, Tuncer S. Effect of different final irrigation solutions on dentinal tubule penetration depth and percentage of root canal sealer. J Endod 2012;38:860-3.  Back to cited text no. 8
Kokkas AB, Boutsioukis AC, Vassiliadis LP, Stavrianos CK. The influence of the smear layer on dentinal tubule penetration depth by three different root canal sealers: An in vitro study. J Endod 2004;30:100-2.  Back to cited text no. 9
Prado M, Gusman H, Gomes BP, Simão RA. Scanning electron microscopic investigation of the effectiveness of phosphoric acid in smear layer removal when compared with EDTA and citric acid. J Endod 2011;37:255-8.  Back to cited text no. 10
Ballal NV, Kandian S, Mala K, Bhat KS, Acharya S. Comparison of the efficacy of maleic acid and ethylenediaminetetraacetic acid in smear layer removal from instrumented human root canal: A scanning electron microscopic study. J Endod 2009;35:1573-6.  Back to cited text no. 11
Johns D, Abraham A, Kuruvilla A, Jaganath B, Krishnegowda S, Ramachandra P. A comparative evaluation of smear layer removal by using edta, etidronic acid, and maleic acid as root canal irrigants: An in vitro scanning electron microscopic study. J Conserv Dent 2015;18:247.  Back to cited text no. 12
[PUBMED]  [Full text]  
Hülsmann M, Heckendorff M, Lennon A. Chelating agents in root canal treatment: Mode of action and indications for their use. Int Endod J 2003;36:810-30.  Back to cited text no. 13
Silva PV, Guedes DF, Pécora JD, da Cruz-Filho AM. Time-dependent effects of chitosan on dentin structures. Braz Dent J 2012;23:357-61.  Back to cited text no. 14
Darrag A. Effectiveness of different final irrigation solutions on smear layer removal in intraradicular dentin. Tanta Dent J 2014;11:93-9.  Back to cited text no. 15
Pimenta JA, Zaparolli D, Pécora JD, Cruz-Filho AM. Chitosan: Effect of a new chelating agent on the microhardness of root dentin. Braz Dent J 2012;23:212-7.  Back to cited text no. 16


  [Table 1], [Table 2]


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