Abstract

Objective: Antibacterial effect of several root canal sealers were compared which have different chemical structures. Materials And Methods: Enterococcus faecalis (EF) incubated into brain-heart infusion agar at 37 C for 24 h. 20 mg of used root canal sealers were impregnated blank antimicrobial discs which were Endomethasone N, AH Plus, MTA Fillapex and Sealapex. Each plates were separated four quadrants and root canal sealers impregnated antimicrobial discs were inserted into their quadrants. Inhibition zones were measured with eye-witness on the plates by well-educated specialist after 48 h. Results: Endomethasone N have made higher inhibiton zones than other root canal sealers. AH Plus has larger inhibition zone than MTA Fillapex and Sealapex. Sealapex has less antibacterial effect on E. Faecalis. Conclusion: The results demostrated that eugenol based root canal sealers have significant antibacterial effects.

1. Introduction

The main cause of endodontically unsuccessful root canal treatment is the continuation of intraradicular or secondary infections. ¹ One of the goals of endodontic treatment is to completely remove the microorganism from the root canal cavity or at least to the levels appropriate for periradicular tissue health. However, many studies have reported that bacteria are present in dentinal tubules and cement, even after treatment. ² Permanence of microorganisms, ineffective irrigation, lack of most of the surfaces in root canal shaping, and anatomical variations may result from incomplete or inadequate chemomechanical preparation. ³ Compared to microorganisms in teeth that failed endodontic treatment and untreated teeth, it has been shown that there are significantly different species. ⁴ The Gram-positive bacteria, such as Streptococci, Lactobacilli, Staphylococci, Enterococcus faecalis, Olsenella uli, Parvimonas micra, Pseudoramibacter alactolyticus, Propionibacterium spp., can be found in root canals of inadequate endodontic treatment. ⁵ Enterococcus faecalis are present in at least one third of the dental canals with persisted periapical lesions. ⁶ As it is unknown that the microorganism is completely eliminated in endodontic treatment, the antimicrobial effect of root canal sealers can help to remove residual microorganisms that are not impaired by the chemomechanical preparation of the root canal system. ⁷

Root canal sealers with adequate antimicrobial activity are used to help minimize or inhibit microorganism development and to restore apical and periapical tissues. ⁸. Today, many commercial root canal sealers vary in their chemical structure and physicochemical properties. ⁹. Compared to the antimicrobial of root canal sealers, those containing eugenol showed better results than those containing calcium hydroxide and resin based ones. ¹⁰ Agar diffusion test is one of the most commonly used techniques to evaluate the antibacterial effect of root canal sealants. ⁸

The purpose of this study is to investigate the antimicrobial activity of root canal sealers zinc-oxide eugenol based, calcium hydroxide based, MTA-based salicylate resin and epoxide-amine based resin on E. faecalis.

2. Material and Methods

All microbiological analyzes were performed under aseptic conditions in a laminar flow chamber. (FTLFU, Fitateknik, Istanbul, Turkey). Standard Enterococcus faecalis strain (ATCC 29212, American Type Culture Collection, Manassas, USA) has been used for reliable antibacterial activity assessment. The microorganisms were cultivated in brain-heart infusion broth (Brain Heart Infusion Broth Cat. No: 53286, Sigma-Aldrich, Taufkirschen, Germany) at 37°C for 18 h. Bacterial suspension of %0.85 saline solution was designed to suit the turbidity equivalent of 0.5 McFarland standard tube. Brain Heart Infusion (BHI) agar were spread with 0.1 mL of the bacterial suspension. BHI agar was prepared as follows respectively; 1) dehydrated medium is melted by heating in distilled water at 52.0 g / L, sterilized in an autoclave at 121 oC for 15 minutes and poured 12.5 mL in sterile Petri dishes, its pH is 7.4 ± 0.2 at 25°C after sterilization; (2) The prepared medium is clear and sometimes slightly opalescent (tempered, turns on) and brown; (3) BHI agar media can be prepared by adding Agar-agar (Merck 1.01613, Merck, Darmstadt, Germany) to Brain Heart Infusion medium (Merck 1.10493, Merck, Darmstadt, Germany) at 15 g / L.

Afterwards, each medium was divided into 4 equal parts on the petri dish with an indelible pen. Used materials are shown in Table 1. A pit was opened in the middle of each quarter with a diameter of 6 mm and a depth of 4 mm, and this area was filled with experimental materials. One petri dish was left empty and the other used dental materials to find out if there was contamination. All plates were maintained at room temperature for 2 h for prediffusion of the materials and then incubated at 37°C for 48 h under aerobic conditions. Inhibition zones were measured with eye-witness on the plates by well-educated specialist wiyh milimetric ruler after 48 h. The size of the inhibition zone was calculated as follows: size of inhibition zone = (diameter of halo – diameter of specimen) x ½ All the assays were conducted in triplicate and the results were recorded in terms of the average diameter of inhibition zone. (Figure 1) The Mann-Whitney’s U test, in the SPSS statistical software package (SPSS, version 20.0, IBM, Armonk, NY, USA) was used for the statistical analysis.

3. Results

The diameters of the inhibition zones on agar plates with agar diffusion testing technique provided by each material are shown in Table 2.

Endomethasone N developed a maximum inhibition region of 19 mm and there was a statistically significant difference in the size of the inhibition regions between the other root canal sealers. (p<0.05).

In our study, Sealapex was the root canal sealer that showed the least effect on E. faecalis with the mean of 1.33 mm inhibition zone. No statistically significant difference was found between MTA Fillapex and Sealapex. In some petri dishes, MTA Fillapex and Sealapex did not form an inhibition zone.

4. Discussion

Many researchers have blamed many types of anaerobic and facultative anaerobic bacteria for unsuccessful root canal cases. Therefore, the elimination or inhibition of bacteria from the root canal systems is very important for the success of endodontic treatment. ^{11, 12} While several changes have been made in recent years, the latest endodontic chemo-mechanical disinfection protocols can not guarantee complete disinfection of the root canal complex. ¹³

Many types of anaerobic bacteria such as Staphylococcus aureus, Fusobacterium nucleatum, Streptococcus anginosus are found in failed root canal treatment. ¹⁴ Nevertheless, Enterococcus faecalis is typically the microorganism associated with the etiology of chronic periradicular lesions. ¹⁵ E. faecalis can survive in difficult conditions. It can do this thanks to its ability to create biofilms. This feature makes it resistant to antimicrobial agents, phagocytosis and antibodies. ¹⁶ Because of these features, E. faecalis was chosen in our study.

After the root canal is prepared, chemical irrigation is applied to remove the biofilm layer and eliminate the remaining microorganisms. The root canal sealers used after all these procedures must have a long-lasting antimicrobial properties to remove the residual microorganisms and to avoid them from being re-colonized. Root canal sealers, however, should be biocompatible and dimensionally stable. ¹⁷ Agar diffusion test is one of the most popular and simple methods for studying the antimicrobial properties of root canal sealers. There are, however, certain drawbacks, such as the failure to standardize the amount of bacteria placed, the inadequate culture medium, the agar viscosity, the plate storage state and the solubility and diffusion properties of both the test substance and the media. ¹⁸ Since water soluble materials are used in our study, agar diffusion test was performed. One reason for the difference in results suggests that the solubility of the materials.

In our study, zinc-oxide eugenol-based Endomethasone N was found to be more effective than other groups. As seen in other studies on antimicrobial activity, antibacterial abilities are probably due to the presence of zinc oxide and eugenol in their composition. ^{19, 20} AH Plus, which includes epoxypropane and formaldehyde, has shown better performance than other resin-containing root canal sealers. In our study, less inhibition zones seen in MTA Fillapex and Sealapex may be due to the low solubility of these root canal sealers. Because the most critical criteria for the agar diffusion test is solubility. ¹⁸ Although Cobankara et al ¹⁸ said otherwise, in some studies, it has been shown that long setting time affects antimicrobial activity.

Its high zinc-oxide content makes Endomethasone N a step forward. The solubility of zinc-oxide and eugenol provided a better antimicrobial activity. It is an important advantage that AH Plus releases formaldehyde during setting time. ²¹ In our study, since Sealapex completed the setting period in 15-60 minutes, its antimicrobial properties were limited. ⁸ Morgental et al ²² showed that MTA Fillapex showed antimicrobial properties before setting and this decreased over time. Although MTA Fillapex showed a certain antimicrobial feature in our study, it remained behind the groups containing eugenol and formaldehyde.

5. Conclusion

Endomethasone N, containing zinc-oxide and eugenol, was found to be more effective on E. faecalis. As a result, the antimicrobial activity is affected by the composition and chemical properties of root canal sealers.

References