|Year : 2018 | Volume
| Issue : 1 | Page : 9-14
Comparative evaluation of different gutta-percha disinfecting agents: A microbiological study
Singh Shailja, Chandra Ramesh, Saxena Anubha, Siddiqui Shazia
Department of Conservative Dentistry and Endodontics, Career Post Graduate Institute of Dental Sciences and Hospital, Lucknow, Uttar Pradesh, India
|Date of Web Publication||4-Jun-2018|
Dr. Singh Shailja
Department of Conservative Dentistry and Endodontics, Career Post Graduate Institute of Dental Sciences and Hospital, Lucknow, Uttar Pradesh
Source of Support: None, Conflict of Interest: None
Aim: The purpose of this study was to evaluate the effectiveness of chemical agents and herbal alternatives such as Aleo vera, Neem, and Amla for rapid decontamination of GP cones.
Materials and Methods: The reference strains of E. Faecalis were obtained, to know the efficacy of different agents aginst this microbe. Two experimental groups used in this study comprises of Herbal agents and chemical agents. The extracts of the Herbal agents which included amla, aloe vera, neem were prepared whereas the chemical agents were commercially available. Chemical agents • Group 1% – 5% Sodium hypochlorite • Group 2% – 2% Glutaraldehyde • Group 3% – 2% CHX. Herbal agents • Group 4 – Amla • Group 5 – Aloe vera • Group 6 – Neem. The zone of inhibition of the different herbal and chemical agents were checked in two different media.
Results: The result of the study showed that CHX was effective in disinfection of GP cones for 1 min followed by glutaraldehyde and sodium hypochlorite. Whereas, Amla was the most effective herbal extract in disinfecting GP followed by Aloe vera and Neem.
Conclusion: t CHX and glutaraldehyde are efficient chemical disinfectants. Herbal agents also hold a promising future as a medium for disinfection, in which Amla gave the best results within a short duration of time.
Keywords: Aleo vera, Amla, gutta percha disinfectant, herbal disinfectant, Neem
|How to cite this article:|
Shailja S, Ramesh C, Anubha S, Shazia S. Comparative evaluation of different gutta-percha disinfecting agents: A microbiological study. Endodontology 2018;30:9-14
| Introduction|| |
Microorganisms are major etiological agents in pulpal and periapical disease., The major goals of endodontic therapy are the elimination of microorganisms from the root canal system and the prevention of subsequent reinfection. In clinical practice, the dentist is occasionally come across with a problem of infection that occurs after obturation of root canal space. The endodontic treatment involves an initial mechanical phase of the instrumentation of root canal space; the chemical phase which involves antimicrobial solutions as irrigation agents and a final phase of obturation of root canal space with a suitable obturating material.
Gutta-percha (GP) at present is the most common obturating material used for filling root canal space. They are biocompatible dimensionally stable radiopaque and thermoplastic and can be removed easily from root canal space. According to some authors as GP cones are produced under the aseptic condition and possess strong antimicrobial properties owing to its zinc oxide component, and they have been used directly from their sterile packages, so the need for decontamination for GP cones is unnecessary. However, the risk of GP cones contamination by glove handling and other physical source during its storage process can also be not ruled out.
Unlike most instruments used in the endodontic treatment, GP cones cannot be sterilized by conventional autoclaving or in a hot-air oven; therefore, they require rapid decontamination before use to maintain the aseptic condition required for successful endodontic therapy., Various chemical agents have been proposed as GP cones disinfectant, including sodium hypochlorite,,,,,, glutaraldehyde,,, alcohol, iodine compounds, and hydrogen peroxide.
The appropriate disinfecting agent should be effective in killing different bacterial species. The objective of this study is to evaluate the effectiveness of a herbal alternative, Aloe vera and Amla for rapid decontamination of GP cones.
Aloe barbadensis mill, is a short succulent herb, it has strong antibacterial, antifungal, and antiviral properties., The antimicrobial effects have been attributed to the plant's natural anthraquinones. Neem has an established antibacterial, antifungal, and antiviral effect. Neem also has anti-inflammatory, antiseptic, astringent, and analgesic properties. Hence, it can be effective in decontaminating GP cones as well.
Therefore, the objective of the present study was to evaluate the antimicrobial properties of three commonly used chemical agents, namely 5% sodium hypochlorite, 2% chlorhexidine (CHX), and 2% glutaraldehyde, and three herbal agents, namely Amla, Aloe vera, and Neem, against the most commonly isolated microorganism from the root canal, that is, Enterococcus faecalis.
E. Faecalis is considered to be the most common Gutta Percha Contaminant, hence in this study the reference strains of E. Faecalis were obtained, t know the efficacy of different agents aginst this microbe.
Two experimental groups used in this study comprises of Herbal agents and chemical agents.
The extracts of the Herbal agents which included amla, aloe vera, neem were prepared whereas the chemical agents were commercially available.
- Group 1% – 5% Sodium hypochlorite
- Group 2% – 2% Glutaraldehyde
- Group 3% – 2% CHX.
- Group 4 – Amla
- Group 5 – Aloe vera
- Group 6 – Neem.
The antibacterial activity of the chemical and herbal agents were tested against E. faecalis using two different media, blood agar (BA) and Mueller-Hinton agar media. The strains of E. faecalis were cultured overnight in brain–heart infusion broth, and then, the culture was streaked on a plate of BA and Mueller-Hinton agar media. Three wells of 5 mm × 5 mm measure were made for chemical and herbal agents on the plates of both the media. About 0.1 ml of the agents were delivered into the well using a micropipette. They were then incubated at 37°C for 24 h; these were then monitored for clear zones around the agents. The antibacterial activity was assessed by the diameter of the inhibition zone.
The data were summarized as mean ± standard deviation. The groups were compared by two-way analysis of variance and the significance of mean difference within (inter), and between (intra) the groups was done by Tukey's honestly significant difference post hoc test after ascertaining normality by Shapiro–Wilk's test and homogeneity of variance by Levene's test. A two-tailed (α = 2) P < 0.05 (P< 0.05) was considered statistically significant.
| Results and Observations|| |
The present study compares the different disinfecting agents for GP cones. Total 72 samples, 36 for Muller-Hinton agar (MHA) medium and 36 for BA medium were selected. Both medium were then treated with sodium hypochlorite (Group 1), glutaraldehyde (Group 2), CHX (Group 3), Amla (Group 4), Aloe vera (Group 5), and Neem (Group 6) each with six samples. The outcome measure of the study was zone of inhibition against E. faecalis measured in millimeter (mm).
The zone of inhibition of different herbal and chemical disinfectants against E. Faecalis in the Muller hinton Agar Media and Blood Agar is depicted in [Figure 1]a, [Figure 1]b, and [Figure 2]a, [Figure 2]b. Whereas the zone of inhibition is summarized in [Table 1].
|Table 1: Zone of inhibition (mean±standard deviation) of two medium and six groups|
Click here to view
|Figure 1: (a) Zone of inhibition obtained in Muller-Hinton agar medium with herbal disinfectant (b) Zone of inhibition obtained in Muller-Hinton agar medium with chemical disinfectant|
Click here to view
|Figure 2: (a) Zone of inhibition obtained in blood agar medium with herbal disinfectant (b) Zone of inhibition obtained in blood agar medium with chemical disinfectant|
Click here to view
The comparison between the zones of inhibition in different media can be done with Graphs depicted in [Graph 1]. [Table 1] and [Figure 1]a, [Figure 1]b and [Figure 2]a, [Figure 2]b showed that the mean zone of inhibition in groups of MHA medium was higher than the BA medium except Group 1 and Group 2. Further, in MHA medium, it was highest in Group 3 and least in Group 5 [Table 1] and [Graph 2]. In contrast, in BA medium it was highest in Group 2 and least in Group 5 [Table 1] and [Graph 3].
For each medium, comparing the mean zone of inhibition between the groups, Tukey's test showed significantly (P< 0.001) different zone of inhibition between the groups in both medium except Group 1 and Group 5, Group 1 and Group 6, Group 2 and Group 3, and Group 5 and Group 6 in MHA medium; and Group 1 and Group 4, Group 2 and Group 3, and Group 5 and Group 6 in BA medium [Table 2].
|Table 2: For each medium, comparison (P value) of mean difference in zone of inhibition between the groups by Tukey's test|
Click here to view
Similarly, for each group, comparing the mean zone of inhibition between the medium, Tukey's test showed similar (P > 0.05) zone of inhibition between the medium at all groups except Group 1 [Table 1] and [Graph 1]. In Group 1, the mean zone of inhibition of BA medium was found significantly different and higher as compared to MHA medium.
The analysis found CHX (Group 3) of MHA media the best disinfecting agent for GP cones.
| Discussion|| |
Disinfection of GP cones before to use as an obturating material is so important for a successful root canal treatment. Obturation is the final and one of the important steps in the root canal treatment. Hence, the materials used for obturation should also be properly disinfected to prevent the persistence of microbes. These cones can be easily get contaminated due to its improper storage or even when they exposed in clinics as well as during handling. Even though the fact that, some authors have said that there is no need to use disinfectant solutions on cones because it may change the properties of GP. In above study, cones contaminated with well-defined microbial populations were sterilized in different antimicrobial solutions. Due to its thermoplastic properties of cones, it become impossible to sterilize them by heat as it would result in their deformation. Therefore, the cones were sterilized with ethylene oxide. Several other chemical solutions have been proposed for a rapid decontamination of GP cones.
E. faecalis is found to be the main cause of endodontic failures as this is one of the most resistant bacteria which greatly adapt with the endodontic system. Hence, this bacteria is chosen in this study including sodium hypochlorite, glutaraldehyde, CHX as chemical disinfectant; Amla, Aleo vera, and Neem as herbal disinfectant.
Sodium hypochlorite is used at varying concentration, and it is most commonly used disinfectant. It has antibacterial as well as sporicidal effect, the mechanism of action involve the liberation of active chlorine which is an oxidizing agent and help to inactivate the bacterial enzyme., Gomes et al. found that the time required for sodium hypochlorite to eliminate the microbes was inversely proportional to concentration of the solution used. The sodium hypochlorite (NaOCl) of 5.25%, can easily eliminate Candida albicans and E. faecalis within 45 s, whereas 2.5% NaOCl takes 10 min for the complete elimination of these microbes. That's why, NaOCl can be used effectively for the disinfection of GP cones. It is very important to wipe off GP with gauze piece after disinfecting with sodium hypochlorite because at concentrations of 5.25% NaOCl can be a reason of having a large quantity of chloride crystals on the GP cone and also might cause GP deterioration as well as loss of its elasticity, which could hamper the obturation quality and can also impair the seal of the root canal. In case of having lower concentration of NaOCl can take more time to disinfect the GP.
CHX is a cationic bisbiguanide with broad-spectrum antibacterial properties. The CHX molecule reacts on the bacterial cell surface (with negatively charged groups), causing an irreversible loss of cytoplasmic components. Gomes et al. stated that 2% CHX liquid took <30 s to completely eliminate E. faecalis from contaminated GP cones. Whereas, in other study, 2% CHX solution was found more effective from 15 s to 2 h in direct contact with infected GP cones.
Glutaraldehyde has been used as chemical sterilizer or high-level disinfectant. Aqueous solution of 2% glutaraldehyde shows a broad spectrum of activity as well as fast destruction rate against the majority of microorganisms. They are capable of destroying bacteria in <1 min of exposure. Spores can be wiped out completely within 3 h or less.
Recently, some herbal alternatives are also being suggested as disinfectants for GP cones. In this study, we have used Amla, Aloe vera, and Neem as a herbal alternative for GP cones disinfection.
Aloe vera has a popular about its antimicrobial activity credited to many compounds which are now precisely acknowledged as p-Coumaric acid, ascorbic acid, pyrocatechol, and cinnamic acid. It has been used from previous years for the treatment of a many type of diseases ranging from peptic ulcers to its use in cosmetic purpose. Aloe vera gel has been found to be effective in decontamination of GP cones within 1 min.
In other study, the extracts of Neem leaf showed significant antibacterial activity against. However, the extracts of Neem leaf shows inhibitory activities of were both organism – as well as solvent-dependent. Its extracts reduced the growth of Gram-positive as well as Gram-negative bacterial species. It also shows an anti-adherence property by modifying bacterial adhesion, and it became more helpful to limit the colonization by microorganisms within the root canals.
Emblica officinalis (Amla) is extensively used in the Indian system of medicine and also believed that it increases body's defense mechanism against many diseases. Amla can protect cells against free-radical damage done by many microbes and also give antioxidant protection. Amla has antibacterial and astringent properties that it becomes more helpful for the prevention of many type of infections and also in the healing of ulcers.
The presence of so many phytochemical components such as alkaloids, tannins, terpenoids, flavonoids, different type of polyphenols, Vitamin C, gallic acid, pectin, and quercetin would have added to better disinfection action. Shoko et al. confirmed that phenolics were the very important compounds that is used against bacteria, whereas gallic acid is known to as the most active substance for inhibition many bacteria. The inhibitory effect of phenolic compounds is due to its adsorption on the cell membranes and its interaction with different enzymes, substrate, and metal ion.
For the evaluation of the efficacy of these disinfecting solutions, these solutions were cultured in BA media and Muller-Hinton media for 24 h at 37°C. The antimicrobial efficacy was assessed by the presence of zones of inhibition. E. faecalis showed maximum zone of inhibition with CHX in chemical agents and Amla among the herbal agents.
The result of the study showed that CHX was effective in disinfection of GP cones for 1 min followed by glutaraldehyde and sodium hypochlorite.
Whereas, Amla was the most effective herbal extract in disinfecting GP followed by Aloe vera and Neem.
| Conclusion|| |
Within the limits of this study, it can be concluded that CHX and glutaraldehyde are efficient chemical disinfectants. Herbal agents also hold a promising future as a medium for disinfection, in which Amla gave the best results within a short duration of time.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Sen BH, Piskin B, Demirci T. Observation of bacteria and fungi in infected root canals and dentinal tubules by SEM. Endod Dent Traumatol 1995;11:6-9.
Foschi F, Cavrini F, Montebugnoli L, Stashenko P, Sambri V, Prati C, et al.
Detection of bacteria in endodontic samples by polymerase chain reaction assays and association with defined clinical signs in Italian patients. Oral Microbiol Immunol 2005;20:289-95.
Dummer PM. Root canal filling. In: Pitt Fort TR, editor. Hartys Endodontic in Clinical Practice. 4th
ed. Oxford: Wright; 1997. p. 123-53.
Moorer WR, Genet JM. Antibacterial activity of gutta-percha cones attributed to the zinc oxide component. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 1982;53:508-17.
Stabholz A, Stabholz A, Friedman S, Heling I, Sela MN. Efficiency of different chemical agents in decontamination of gutta-percha cones. Int Endod J 1987;20:211-6.
Linke HA, Chohayeb AA. Effective surface sterilization of gutta-percha points. Oral Surg Oral Med Oral Pathol 1983;55:73-7.
Cardoso CL, Kotaka CR, Redmerski R, Guilhermetti M, Queiroz AF. Rapid decontamination of gutta-percha cones with sodium hypochlorite. J Endod 1999;25:498-501.
Cardoso CL, Redmerski R, Bittencourt NL, Kotaka CR. Effectiveness of different chemical agents in rapid decontamination of gutta percha cones. Braz J Microbiol 2000;31:72-5.
da Motta PG, de Figueiredo CB, Maltos SM, Nicoli JR, Ribeiro Sobrinho AP, Maltos KL, et al.
Efficacy of chemical sterilization and storage conditions of gutta-percha cones. Int Endod J 2001;34:435-9.
Valois CR, Silva LP, Azevedo RB. Effects of 2% chlorhexidine and 5.25% sodium hypochlorite on gutta-percha cones studied by atomic force microscopy. Int Endod J 2005;38:425-9.
Zalp N, Okte Z, Ocelik B. The sterilization of gutta-percha cones with sodium hypochlorite and gluteraldehyde. J Endod 2006;32:1202-4.
Royal MJ, Williamson AE, Drake DR. Comparison of 5.25% sodium hypochlorite, MTAD and 2% chlorhexidine in the rapid disinfection of polycaprolactone-based root canal filling material. J Endod 2001;27:172-4.
Ramasubramanian TS, Sivakumar VT, Thirumalai AV. Antimicrobial activity of Aloe vera
(L) Burm. F. against pathogenic microorganisms. J Bio Sci Res 2010;4:251-8.
Arunkumar S, Muthuselvam M. Analysis of phytochemical constituents and antimicrobial activities of Aloe vera
L. against clinical pathogens. World J Agric Sci 2009;5:572-6.
Athiban PP, Borthakur BJ, Ganesan S, Swathika B. Evaluation of antimicrobial efficacy of Aloe vera
and its effectiveness in decontaminating gutta percha cones. J Conserv Dent 2012;15:246-8. [Full text]
Natasha F, Dutta K, Mollah AM. Antimicrobial and decontamination efficacy of neem, Aloe vera
and neem + Aloe vera
in gutta percha cones using Escherichia coli
and Staphylococcus aureus
as contaminants. Asian J Microbiol Biotechnol Environ Sci 2015;17:915-7.
Polaquini SR, Svidzinski TI, Kemmelmeier C, Gasparetto A. Effect of aqueous extract from neem (Azadirachta indica
A. Juss) on hydrophobicity, biofilm formation and adhesion in composite resin by Candida albicans
. Arch Oral Biol 2006;51:482-90.
Bloomfield SF, Miles GA. The antibacterial properties of sodium dichloroisocyanurate and sodium hypochlorite formulations. J Appl Bacteriol 1979;46:65-73.
Dychdala GR. Chlorine and chlorine compounds. In: Block SS, editor. Disinfection, Sterilization, and Preservation. 4th
ed. Philadelphia: Lea & Febiger; 1991. p. 133-5.
Gomes BP, Vianna ME, Matsumoto CU, Rossi Vde P, Zaia AA, Ferraz CC, et al.
Disinfection of gutta-percha cones with chlorhexidine and sodium hypochlorite. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2005;100:512-7.
Stepanović S, Antić N, Dakić I, Svabić-Vlahović M.In vitro
antimicrobial activity of propolis and synergism between propolis and antimicrobial drugs. Microbiol Res 2003;158:353-7.
Murray PR, Kobayashi GS, Pfaller MA, Rosenthal KS. Medical Microbiology. 2nd
ed. St. Louis: Mosby; 1994.
Borick PM. Chemical sterilizers (chemosterilizers). Adv Appl Microbiol 1968;10:291-312.
Lawrence R, Tripathi P, Jeyakumar E. Isolation, purification and evaluation of antibacterial agents from Aloe vera
. Braz J Microbiol 2009;40:906-15.
Rajasekaran C, Meignanam E, Vijayakumar V, Kalaivani T, Ramya S, Premkumar N, et al
. Investigations on antibacterial activity of leaf extracts of Azadirachta indica
A. Juss (Meliaceae): A traditional medicinal plant of India. Ethnobot Leaflets 2008;12:1213-7.
Botelho M, Santos AD, Martins J, Carvalho C, Paz M, Azenha C, et al
. Efficacy of a mouthrinse based on leaves of neem in the treatment of patients with chronic gingivitis. J Med Plants Res 2008;2:341-6.
Singh E, Sharma S, Pareek A, Dwivedi J, Yadav S, Sharma S. Phytochemistry, traditional uses and cancer chemopreventive activity of Amla (Phyllanthus emblica
): The sustainer. J Appl Pharm Sci 2011;2:176-83.
Shoko T, Soichi T, Megumi MM, Eri F, Jun K, Michiko W. Isolation and identification of an antibacterial compound from grape and its application to foods. Nippon Nogeikagaku Kaishi 1999;73:125-8.
Duman AD, Ozgen M, Dayisoylu KS, Erbil N, Durgac C. Antimicrobial activity of six pomegranate (Punica granatum
L.) varieties and their relation to some of their pomological and phytonutrient characteristics. Molecules 2009;14:1808-17.
[Figure 1], [Figure 2]
[Table 1], [Table 2]