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Year : 2007  |  Volume : 10  |  Issue : 3  |  Page : 99-103
An invitro evaluation of calcium hydroxide root canal sealers and its effect on six microorganisms

Department of Conservative Dentistry, Ragas Dental College and Hospitals, Uttandi, Chennai -600 119, India

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A study was done to quantify the release of calcium ions from calcium hydroxide containing root canal sealers and their pH at various time intervals and the antimicrobial action. pH was measured using the pHmeter and the calcium amount using the atomic absorption spectrometer. Six strains of microorganisms were used to evaluate the antimicrobial action of the sealers using Agar diffusion test. Seal apex showed the maximum release of calcium ion and pH level at 30 days but with no antibacterial effect.

How to cite this article:
Miglani R, Shankar, Indira R, Ramachandran S. An invitro evaluation of calcium hydroxide root canal sealers and its effect on six microorganisms. J Conserv Dent 2007;10:99-103

How to cite this URL:
Miglani R, Shankar, Indira R, Ramachandran S. An invitro evaluation of calcium hydroxide root canal sealers and its effect on six microorganisms. J Conserv Dent [serial online] 2007 [cited 2023 Jun 4];10:99-103. Available from:

   Introduction Top

One of the major goals of root canal treatment is the elimination of bacteria from the root canal, which is normally achieved by mechanical instrumentation combined with various irrigating solutions along with anti-bacterial dressing of the canal during inter-appointments. Complete elimination of the microorganisms from within root canal system and its effective obturation are considered to be major factors in the success of endodontic treatment [1] . Bystrom and Sunquist (1981,83,85) have observed that bacteria, which survive instrumentation and irrigation, rapidly increase in numbers in the empty canals in the period between appointments [2] . Sunquist (1993) has recommended the use of anti-bacterial medicaments between appointments during root canal treatment of non-vital teeth calcium hydroxide is considered to be the material of choice as an intra-canal medicament because of its superior activity and reduced cytotoxicity to the periradicular tissues [2] .

An ideal root canal sealer in addition to its ideal "sealing" properties, should leave the tooth in the most biologically inert condition possible and must prevent reinfection and growth of any microorganisms remaining in the canal, thereby favoring periapical tissue repair. The therapeutic effect of calcium hydroxide is due to the constant and continous dissociation of calcium and hydroxyl ions. It has been postulated that the high pH level of calcium hydroxide in the root canal promotes a state of alkalinity in adjacent tissues, a condition that favors repair of periradicular tissues. The aim of the present study was to evaluate the efficacy of four commercially available calcium hydroxide root canal sealers by comparing their calcium ion release, pH and antimicrobial action against the microorganisms found commonly in the root canal.

   Materials and Methods Top

The four calcium hydroxide sealers evaluated were Apexit(Ivoclar, Vivadent), Sealapex (Kerr, Sybron, USA), Endoflos(sanlor &Cia), Dentalis (Diadent, Canada). 2cm long plastic tubes of 4 mm internal diameter were used to condense the sealers.l0 specimen were prepared for each sealer group following the manufacturer instructions for manipulation. After the materials were allowed to set, the plastic tubes were immersed in clean tubes containing 10ml of deionized water and were sealed tightly with rubber cork. The sealer material kept in the test tubes were evaluated for pH and calcium ion release at periods of 24hr, 48hr, 7days and 30 days.

After 24hrs, the plastic tubes immersed in the deionized water were removed and immersed again in fresh solution of deionized water. This fresh solution was analyzed for pH with the help of pHmeter. The same procedure was repeated after 48hrs, 7days and 30 days for each specimen for pH evaluation. The tested solution was then diluted with 10%. EDTA and the amount of calcium in each sealer material was analyzed with atomic absorption spectrometer at a concentration of 0.025, 0.05, 0.1 and 0.3mg/100ml, at the end of 24hrs, 48hrs, 7 days and 30days. The samples were diluted with 10% EDTA to prevent the possible interference of alkaline phosphatase and metals.

Agar plate diffusion method was used to evaluate the antimicrobial effect of the sealers. The microbial strains used were Staphylococcus aureus Scientific Name Search  (ATCC#10556),  Streptococcus mutans Scientific Name Search 175),  Enterococcus faecalis Scientific Name Search s Institute, Chennai, India), Candida albicans (Kings Institute, Chennai, India)  Escherichia More Details coli (Kings Institute, Chennai, India), Pseudomonas aureus (Kings Institute, Chennai, India). These lypholized bacteria were rehydrated and their purity and viability were confirmed in blood agar. They were inoculated on the plates containing blood agar and incubated at 37° C in an atmosphere of 10% C02 for 48 hrs. From these cultures bacterial suspensions were prepared in a sterile isotonic saline solution to obtain turbidity, compatible with macFarland scale.

Preparation of Agar plates

Brain heart infusion agar powder of 0.52 gms was dissolved in 100ml of sterile saline solution according to manufacturers recommendation in a closed glass beaker. After the solution was autoclaved and cooled, it was poured on 6 petridishes to an equal thickness and allowed to solidify. 150ml of the inocula was then spread out evenly with sterile cotton swab on the agar plates. Wells of 4mm depth and diameter were made in each agar plate with a sterile stainless steel cork bore. 4 wells were made in each plate and the four root canal sealers were manipulated in accordance with manufacturers instructions and packed into the wells in two increments. The plates were preincubated for one hour at room temperature, followed by incubation at 37°C for 48 hrs. The diameters of the zones of microbial inhibition, if any, were measured and the four sealers were evaluated for antimicrobial effect.

   Results Top

The values were recorded and statistically analyzed using oneway - ANOVA and multiple comparisons was done with post hoc tests.

Calcium ion release was more in Sealapex at 24 hrs, 48 hrs and 30 days time interval. This was statistically significant (p<0.05). At 7 days Sealapex and Dentalis has similar calcium ion release [Table 1].

With respect to pH, Sealapex has the highest values from 24hrs to 30 days, differing significantly with Dentalis and Endoflos (p<0.05). Apexit showed almost consistent pH throughout the period [Table 2].

Antimicrobial results showed that Endoflos had the maximum inhibition zone of 15mm in comparison with Dentalis, Apexit and Sealapex [Table 3].

   Discussions Top

Root canal sealers are used in root canal with combination of core filling materials, such as gutta­percha or silver points. At one time it was thought that sealer played a secondary role by simply cementing the core filling material into the canal .However, it is now appreciated that the sealer has a primary role in sealing the canal by obliterating the irregularities between the canal wall and the core materials. All modern obturating techniques make use of sealer to enhance a fluid tight seal of the pulp space [7] .Calcium hydroxide is the most commonly used intracanal medicament and also available as a therapeutic sealer in different compositions. These sealers show varied antimicrobial action to various microorganisms detected in the root canal flora and this action is attributed to their calcium ion release and change in pH. Hence, this study was done to compare the efficacy of four commonly available calcium hydroxide sealers.

Standardized Plastic tubes open at both the ends of 2cm long and 4mm internal diameter were used in the study to insert the sealer material after mixing and also for easy immersion and removal of the specimen at different time intervals from the deionised water. Plastic tubes were used as it would not have any effect either on the deionised water or on the sealer. The tubes were open in both the ends which helped to condense the sealers without any voids and later helped in pH measurements.

Tagger et al in a study on calcium and hydroxyl ions found that, the hydroxyl ion release from the sealers (Seal apex, CRCS and Hermetic) followed a course similar to that of calcium ions and this resulted in high pH changes causing quick disintegration of the sealer [10],[12] . He also suggested that this solubility is the cause for increased antimicrobial activity favoring the apical healing process.

The results of this study has shown that Sealapex had the highest calcium release at 24hr, 48hrs , 7days and 30 days with significant difference at 30 days between Apexit and Endoflos. These results were consistent with the result observed by Durate Marcro Antonio Hungararo et al 2001 [3] . It can be attributed that in Endoflos and Dentalis the free eugenol of the sealer would have captured calcium ions and made them unavailable to the surrounding medium. Indeed, if the sole source of hydroxyl in a material is from hydrolysis of calcium hydroxide, then calcium ions should have also been released. However, it is important to know whether their release is hampered because of being bound more strongly to eugenol or because calcium is more difficult to detect and measure precisely [13] .

Leonardo et al reported that among the sealing materials studied, Sealapex was one that best permitted the deposition of mineralized tissue at the apical level. This is because of its greater solubility which leads to release of calcium ions into the tissue, favoring the apical repair [8] .

The elevated pH of calcium hydroxide with values reaching 12.6 is due to the greater liberation of hydroxyl ions, which are capable of altering the integrity of bacterial cytoplasmic membrane [6] . The release of hydroxyl ions from the sealers depends on the hydrophilic nature of its matrix and other substances added to its formulations [4] .

Results of the present study show that Sealapex has the highest pH values, which is attributed to its high solubility and disintegration, thereby releasing the hydroxyl ions. Apexit also showed consistent pH throughout the period and the pH values were comparatively less in Dentalis and Endoflos. This can be attributed to the free eugenol molecule that hampers the release of the hydroxyl ions.

Anti bacterial effect of calcium hydroxide is attributed to its high pH. According to McComb and Ericson [9] , the antibacterial activity of calcium hydroxide was influenced by both the total concentration of hydroxyl ions and the rate of their release.

Agar diffusion test used in the study to evaluate the antimicrobial activity of calcium hydroxide sealers against the facultative anaerobes showed that Endoflos F.S had the maximum zone of inhibition of 15mm, followed by Dentalis, Apexit and Sealapex. This antimicrobial activity of Endoflos RS and Dentalis can be attributed to the presence of iodoform in their components rather than the calcium ions. Saggar v and Chandra et al have shown that the iodoform zincoxide-eugenol sealers were effective for both the aerobic ad anaerobic bacteria of the root canals of the teeth [11] .

Studies have demonstrated that calcium hydroxide is superior to camphorated paramonochlorophenol in effectiveness on anaerobic bacteria. However, its effect are highly dependent on the availability of hydroxyl ions in solution, culture media, tissue fluids and dentin, that can prevent calcium hydroxide activity by causing a pH drop. Sealapex and Apexit having high pH and calcium ion release did not show good antibacterial action, which is attributed to its low calcium diffusibility in Agar [5] .

Further studies are required to evaluate more types of calcium hydroxide sealers and to see the effect of their pH and calcium release against more microorganisms found in the root canal flora.

   Conclusion Top

Under the limitations of this present study it can be concluded that seal apex obtained the highest calcium and hydroxyl release (pH) especially after 30 days, followed by Apexit, Dentalis and Endoflos F.S.

Endoflos F.S. showed maximum zone of inhibition towards all the microorganisms and also towards candida albicans, which is due to the presence of lodoform in the composition.

   References Top

1.BarbosaA.M. Carlos, Gonclaves B.Reginaldo, F.Jose Siqueia Jr, and Uzeda Milton De Evaluation of the antibacterial activities of Calcium hydroxide, Chlorhexidine and camphorated para monochlorophenol as intracanal medicament A clinical and laboratory study. J Endod.1997;23(5):297-300.  Back to cited text no. 1    
2.Beltes G.P, Pissiotis Eleftheris, Koulaouziduo Elisabeth and Kortsaris H.Alexander. In vitro release of hydroxyl ions from six types of calcium hydroxide nonsetting pastes. J Endod. 1997;23(7):413-415.  Back to cited text no. 2    
3.Dutante Hungato Macro Antonio, Demarch Anna Claudia Cardoso. Evaluation of pH and calcium ion release of three root canal sealers. J Endod.2000;26:389-390.  Back to cited text no. 3    
4.Economides Nikolaos, Koulaouzidou A Elizabeth, Beltes Panagiotis and Korsaris H.Alexander. In vitro release of hydroxyl ions from calcium hydroxide gutta percha points. J Endod.1999;25(7)481-482.  Back to cited text no. 4    
5.Estrelo carlos, Cristina Fabiana Pimenta, Izabel Yoko lto and Lili Luschke Bammann. In vitro determination of Direct Antimicrobial effect of calcium hydroxide. J Endod.1998;24(1):15-17.  Back to cited text no. 5    
6.Estrela Carlos,Sydney Biltzkow Gilson, Bammann Luescke Lili. Mechanism of action of calcium and hydroxyl ions of calcium hydroxide on tissue and bacteria. Braz Dent J.1995; 6(2):85-90.  Back to cited text no. 6    
7.Harty's Endodontics in clinical practice 4 th edition  Back to cited text no. 7    
8.Leonardo MR, Silva LA, Utrilla LS, Assed S, Ether SS.Calcium hydroxide root canal dressing: Histopahological evaluation of periapical repair at different time periods. Braz Dent J.2002;13(1):17-22.  Back to cited text no. 8    
9.Mickel K.Andre and E.Ronald Wright. Growth inhibition of Streptococcus anginosus (milleri) by three calcium hydroxide sealers and one zinc oxide eugenol sealer. J Endod.1999;25(1):34-37.  Back to cited text no. 9    
10.Rehman et al. Calcium ion diffusion from calcium hydroxide containing materials in endodontically treated teeth. An invitro study. Int Endod J. 1996;29:271-279.  Back to cited text no. 10    
11.Sagger V, Chandra S,Jaiswal JN and Singh M. Antimicrobial efficacy of iodoformized zinc oxide-eugenol sealer on micro-organisms of root canal. Journal of Indian Soc Pedod Prev Dent, 1991 Mar;9:1-3.  Back to cited text no. 11    
12.Silva Bezerra Da L.A.,Leonardo M.R, Silva Da R. S et al. Calcium hydroxide root canal sealers evaluation of pH, calcium concentration and conductivity. Int Endod J.1991;30:205-209.  Back to cited text no. 12    
13.Zmener Osvaldo et al. Biocompatibility of two calcium hydroxide based endodontic sealers. A quantitative study in the subcutaneous connective tissue of rat. J Endod. 1998;14: 229-235.  Back to cited text no. 13    

Correspondence Address:
Revathi Miglani
Department of Conservative Dentistry, Ragas Dental College and Hospitals, Uttandi, Chennai -600 119
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/0972-0707.42269

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  [Table 1], [Table 2], [Table 3]

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