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ORIGINAL ARTICLE
Year : 2017  |  Volume : 6  |  Issue : 2  |  Page : 98-102

An in-vitro assessment of the residual antimicrobial effects and surface alterations of gutta-percha disinfected with four different solutions


Department of Conservative Dentistry, Chhattisgarh Dental College and Research Institute, Chhattisgarh, India

Date of Web Publication13-Jun-2017

Correspondence Address:
Shweta Bandhe
Chhattisgarh Dental College and Research Institute, Campus, Girls Hostel, Sundra, Rajnandgaoun - 491 441, Chhattisgarh
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/JDRNTRUHS.JDRNTRUHS_102_16

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  Abstract 

Context: To evaluate the residual antimicrobial effects and surface alterations of gutta-percha disinfected with four different solutions.
Materials and Methods: Eighty gutta-percha cones (ISO #40) were randomly assigned to four experimental groups; different types of disinfecting solutions were used for each, namely sodium hypochlorite (NaOCl), chlorhexidine (CHX), aloe vera, and propolis (n = 20 for each group). The cones were immersed in each solution for 5 minutes and 10 minutes; thereafter they were dried. Agar diffusion assay was performed to assess the zone of inhibition. Scanning electron microscope analysis was conducted on additional 80 disinfected cones.
Statistical Analysis Used: The data was analyzed using ANOVA and Bonferroni multiple comparison test and Man–Whitney U test.
Results: Statistical significance was observed among the four groups (P < 0.05). Group IV displayed no statistical significance at 5 minutes and 10 minutes exposure, whereas groups I, II, and III displayed statistical significance at both exposures. Based on the above-mentioned results, gutta-percha when immersed in propolis for 5 minutes and 10 minutes had shown better residual antibacterial action as compared to other test solutions. No surface alterations were detected on gutta-percha cones during the selected time intervals.
Conclusion: Propolis followed by aloe vera possesses superior antibacterial efficacy as compared to sodium hypochlorite and chlorhexidine and does not cause any surface alterations to gutta-percha.

Keywords: Aloe Vera, chlorhexidine, gutta-percha cones, propolis


How to cite this article:
Nair R, Bandhe S. An in-vitro assessment of the residual antimicrobial effects and surface alterations of gutta-percha disinfected with four different solutions. J NTR Univ Health Sci 2017;6:98-102

How to cite this URL:
Nair R, Bandhe S. An in-vitro assessment of the residual antimicrobial effects and surface alterations of gutta-percha disinfected with four different solutions. J NTR Univ Health Sci [serial online] 2017 [cited 2017 Nov 22];6:98-102. Available from: http://www.jdrntruhs.org/text.asp?2017/6/2/98/207999




  Introduction Top


Success of endodontic therapy depends on the complete cleaning and shaping of the root canal, disinfection of the canal, and obturation of the canal space. However, despite the use of aseptic techniques during these procedures, infection may recur.[1] One of the viable causes behind such recurrence may be the use of contaminated gutta-percha.[1]

Introduced for over a span of 100 years into the field of endodontics, gutta-percha still continues to be the most commonly used filling material today. They are biocompatible, dimensionally stable, radiopaque, and thermoplastic.[2] It can be anticipated that if manufactured under aseptic conditions, they might get contaminated by aerosols, by handling, or by improper storage.[1]

Studies have shown that staphylococcus genus has been found to be the most common microorganism that contaminates the gutta-percha cones in their boxes while handling with gloves. Hence, the need for disinfection of gutta-percha is a prerequisite prior to obturation.[1] However, due to its thermoplastic nature, conventional methods of moist or dry heat sterilizations cannot be used. A rapid chair side chemical disinfection is indicated.[3]

Studies about gutta-percha disinfection have evaluated several chemically cold disinfection solutions such as polyvinyl pyrolidone-iodine, ethyl alcohol, sodium hypochlorite, hydrogen peroxide, glutaraldehyde, and chlorhexidine (CHX). Among which, immersing gutta-percha in 5.25% sodium hypochlorite (NaOCl) for 1 minute is a good standard method for rapid sterilization.[2] CHX has also been found to be an effective gutta-percha disinfectant and it shows no surface alteration.

In the recent years, herbal medicines have gained popularity because of their natural origin, ease of availability, efficacy, safety, and meagre side effects.[4] Propolis is a brownish resinous substance collected by bees, mainly from plants. It is a potent antimicrobial, antioxidant, and anti-inflammatory agent.[5] Recently, it has attracted much attention due to its antibacterial and antifungal activities. Aloe vera (Aloe Barbadensis Miller) is a cactus-like plant that grows in hot and dry climates belonging to family Liliaceae. This medicinal herb possesses potent antibacterial, antifungal, antiviral, and antioxidant properties.[6]

Therefore, the purpose of this study was to evaluate the antimicrobial effects and surface alterations of gutta-percha when disinfected with 2% CHX, 5.25% NaOCl, 20% aloe vera, and 10% propolis against Enterococcus faecalis.


  Materials and Methods Top


A total of 80 gutta-percha cones (ISO #40, Dentsply, Maillefer), previously sterilized by ethylene oxide were selected for study.

We dissolved 20 g of dry extract of aloe vera in 100 ml of distilled water to prepare a 20% solution. Propolis (10%) was prepared by dissolving 10 g of propolis dry extract in 100 ml of glycerine. The samples were randomly assigned to four experimental groups with 20 samples in each group according to disinfecting solutions; namely, CHX (Neelkanth Health Care Ltd. Dental Division, India), NaOCl (Neelkanth Health Care Ltd. Dental Division, India), Aloe vera (Herbs and Crops Overseas Ahmedabad, Gujrat), propolis (Hi-Tech Natural Products Ltd. Delhi), and time of exposure to each substance:

  • Group 1: (n = 20) Ten cones were immersed in 2% CHX solution for 5 minutes and 10 cones for 10 minutes
  • Group 2: (n = 20) Ten cones were immersed in 5.25% NaOCl for 5 minutes and 10 cones for 10 minutes
  • Group 3: (n = 20) Ten cones were immersed in 20% aloe vera for 5 minutes and 10 cones for 10 minutes
  • Group 4: (n = 20) Ten cones were immersed in 10% propolis for 5 minutes and 10 cones for 10 minutes.


After immersion, the gutta-percha cones were removed from the disinfecting solutions, washed with distilled water, and dried using sterile gauze. Each cone was added to the surface of the media for microbiologic testing.

Microbiologic testing

The species of microorganism used in this study was E. faecalis ATCC 29212. The microorganism E. faecalis was grown in cultured media and inoculated into test tubes containing 10 ml of brain heart infusion media (High Media Laboratory Private Ltd. Mumbai) and grown overnight. E. faecalis suspension density was maintained at 0.5 scale Mc Farland (108 CFU). Following this, the microorganism was inoculated into previously set Muller Hinton agar plates (High Media Laboratory Private Limited, Dombivali, Thane, Mumbai). Each cone was transferred to the test tubes containing 3 ml of each solution being tested to stand for the various experimental periods. After removal from the solutions, they were washed, dried, and then added to the surface of the media.

The plates were kept for 2 hours at room temperature and then incubated at 37°C under appropriate gaseous conditions, for 24 hours. The zones of inhibition of microbial growth around the cones were measured and recorded after the incubation period [Figure 1].
Figure 1: Agar diffusion assay

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Scanning electron microscope (SEM) evaluation

Additional 80 disinfected cones had been selected and subjected to same disinfecting protocol, which was subjected for microbiologic study. Ten cones were selected as the control group. Sample of each group was subjected to scanning electron microscope (SEM) (ZEISS, EVO 18, National Institute of Technology, Raipur, Chhattisgarh) to observe surface alterations.

Statistical analysis

The data was analyzed using ANOVA and Bonferroni multiple comparison test and Man Whitney U test by using analysis software SPSS 17.0. The significance level for all the tests was set to P< 0.05.


  Results Top


The antimicrobial efficacy was assessed by the presence of zones of inhibition. When the four solutions were compared to each other in terms of time intervals, statistical significance was seen (P < 0.05) [Table 1].
Table 1: Diameter to zone of inhibition in millimeters

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Group IV (propolis) showed better antimicrobial efficacy at both 5 minutes and 10 minutes exposure against E. faecalis with 8 mm zone of inhibition. This was followed by group III (aloe vera), which was on par with propolis after 5 minutes of exposure but its efficacy decreased after 10 minutes where the zone of inhibitions were 8 mm and 6 mm, respectively.

The 2% CHX was slightly more effective (significantly) than group I, while 5.25% NaOCl was the least effective at 5 minutes and 10 minutes of immersion. (P < 0.05). None of the tested solutions showed any surface alterations on gutta-percha during selected time interval [Figure 2].
Figure 2: The SEM evaluation of guttapercha cone exposed to different solutions in different time intervals

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  Discussion Top


The surface of gutta-percha cones involves the presence of surface irregularities that are filled by root canal sealers. These gaps and deep irregularities create a large interface between the root canal wall and gutta-percha, allowing the leakage of molecules that serve as nutrients for the microorganisms present in the root canal system.[7]

Although the number of these organisms is quite low at the time of packaging, the number of organisms increase during routine clinical practice. Once exposed to dental chair side clinical condition, gutta-percha becomes a potential source for contamination by cocci, rods, and yeast. Incorrect handling of gutta-percha cones during the retreatment or checking the fit of the master cone before final rinse may lead to the contamination of the cones.[1] Studies have revealed the presence of microorganisms in 5–19% of freshly opened gutta-percha packs.[1] This was verified by Gomes et al.[8]E. faecalis being the most resistant intracanal pathogen in failed root canal, serves as a gold standard to represent other possible microorganism. Therefore, E. faecalis was selected for this study.[1]

NaOCl is widely used at different concentrations for the sterilization of gutta-percha against possible contamination and its activity is correlated to its concentration.[7] Senia et al. (1975) and Siqueria et al. (1998) reported that 5.25% NaOCl has been routinely used as an effective chemical for rapid decontamination of GP cones which kills the micro-organisms in 1 minute of exposure.[9],[10] Vianna et al. too reported that 5.25% NaOCl eliminates most organisms within 15 seconds.[7] Studies have also shown that 5.25% NaOCl solution is a strong oxidizing agent that causes deterioration of gutta-percha. Crystal formation was also identified after rapid sterilization with 2.5 and 5.25% NaOCl.[3] Valois et al. detected surface alterations even at a short period of exposure to different concentrations of NaOCl.[11] These alterations were not found in the present study during the SEM evaluation.

CHX 2% is a broad spectrum antimicrobial agent with substantively and relatively low toxicity.[1] Studies have shown that CHX is efficient only after 5 minutes. Chandrappa et al. in their study concluded that 5.25% NaOCl and 2% CHX completely eliminated the Staphyloccocus aureus within 5 minutes but was unable to completely eliminate E. faecalis even after 5 minutes. Therefore, the time interval was taken as 5 minutes and 10 minutes.[1] The clinical importance of CHX on gutta-percha cones have been attributed to being related to its immediate release inside the root canal, during the obturation time.[3]

Gomes et al. (2001) and Vianna et al. (2004) reported that 2% CHX and 5.25% NaOCl have similar antimicrobial performance.[8] Valois et al. reported that 2% CHX solution did not cause deterioration of gutta-percha cone structure.[11] This was in accordance to our study where 2% CHX was seen slightly better than 5.25% NaOCl after 5 minutes and 10 minutes.

Recently, the medicinal property of different herbs have been exploited in the field of endodontics.[11] Oncag et al. (2008), Awawdeh et al. (2009), Athiban et al. (2012)[12],[13],[14] had conducted studies to demonstrate the efficiency of propolis and aloe vera as intracanal medicaments, yet very few studies have been conducted on sterilization of gutta-percha and its effect on the topography of gutta-percha.[14]

Propolis a natural antibiotic is a resinous yellow brown to dark brown substance that honey bees collect from tree buds, shrubs, or other botanical sources.[11] It also possesses good antioxidant and anti-inflammatory activity. The main chemical elements present in propolis are flavonoids, phenolic, and various aromatic compounds.[4]

Aloe vera, an ancient medicinal herb that contains around 75 active ingredients including vitamins, enzymes, sugars, minerals, lignin, saponins, salicylic acid, and amino acids, which have antioxidant, antiviral, and antibacterial properties.[15] Numerous studies on aloe vera are being done to demonstrate these properties. In dentistry, it is also used in cases of apthous ulcers, lichen planus, alveolar osteitis.[15]

Athiban et al. (2012) concluded the efficacy of aloe vera as decontaminant of gutta-percha cones for a short duration. Its antimicrobial activity is due to the presence of organic compounds p-coumaric acid, ascorbic acid, pyrocatechol, and cinnamic acid.[14] In this study we found that aloe vera showed effective antibacterial activity against E. faecalis when compared to 2% CHX and 5.25% NaOCl but this decreased with time.

In this study, propolis was found to be most effective against micro-organism as compared to all groups. Kujumgiev et al. reported the antimicrobial action of propolis to be due to flavonoids and esters of phenolic acids.[5] Its efficacy remained the same at the exposure time 5 minutes and 10 minutes, whereas the zone of inhibition (8 mm) remained the same. While aloe vera, initially, on exposure for 5 minutes has a zone of inhibition on par with propolis but at 10 minutes exposure, its efficacy seems to have reduced.

When subjected to SEM, the presence of deposits was seen over the cones after disinfection with 5.25% NaOCl with 5 minutes and 10 minutes exposure. Disinfection of gutta-percha with aloe vera also shows presence of some deposits on the surface of gutta-percha. However, none of the solutions showed significant alterations in the surface of gutta-percha after 5 minutes and 10 minutes.


  Conclusion Top


Under the limitation of this study, it can be concluded that propolis was more effective than aloe vera; both can be effectively used to decontaminate the gutta-percha as a source of rapid sterilization. None of the solutions produced any irregularities on the surface of gutta-percha.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
  References Top

1.
Chandrappa MM, Mundathodu N, Srinivasan R, Nasreen F, Kavitha P, Shetty A. Disinfection of gutta-percha cones using three reagents and their residual effects. J Conserv Dent 2014;17:571-4.  Back to cited text no. 1
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2.
Sahinkesen G, Oktay EA, Er O, Kocak MM, Kilic A. Evaluation of residual antimicrobial effects and surface changes of gutta-percha disinfected with different solutions. J Contemp Dent Pract 2011;12:47-51.  Back to cited text no. 2
    
3.
Gomes BP, Berber VB, Montagner F, Sena NT, Zaia AA, Ferraz CC, et al. Residual effects and surface alterations in disinfected guttapercha and resilon cones. J Endod 2006;33:948-51.  Back to cited text no. 3
    
4.
Ravishankar P, Lakshmi T, Kumar A. Ethno botanical approach for root canal treatment – An update. Journal of pharmaceutical science and research.  Back to cited text no. 4
    
5.
Garg. Comparison of antimicrobial efficacy of Propolis, morindacitrifolia, azadirachtaindica, triphala, green tea polyphenols and 5.25% sodium hypochlorite against enterococcus faecalis biofilm. J Saudi Endod 2014;4:122-7.  Back to cited text no. 5
    
6.
Sureshchandra B, Kumar AJ. Antimicrobial efficacy of aloe vera extract on resistant antimicrobial strains in endodontics. Endodontology.  Back to cited text no. 6
    
7.
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.  Back to cited text no. 7
    
8.
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.  Back to cited text no. 8
    
9.
Senia ES, Marraro RV, Mitchell JL, Lewis AG, Thomas L. Rapid sterilization of gutta-percha cones with 5.25% sodium hypochlorite. J Endod 1975;1:136-40.  Back to cited text no. 9
    
10.
Siqueira JF Jr, da Silva CH, Cerqueira M das D, Lopes HP, de Uzeda M. Effectiveness of four chemical solutions in eliminating bacillus subtilis spores on gutta-percha cones. J Endod and Dent Trumatol 1998;14:124-6.  Back to cited text no. 10
    
11.
Valois CRA., 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.  Back to cited text no. 11
    
12.
Oncag O, Cogulu D, Uzel A, Sorkun K. Efficacy of Propolis as an intracanal medicament against Enterococcus faecalis. Gen Dent 2008;54:319-22.  Back to cited text no. 12
    
13.
Awawdeh L, Al-Beitawi M, Hammad M. Effectiveness of Propolis and calcium hydroxide as a short-term intracanal medicament against Enterococcus faecalis: A laboratory study. Aust Endod J 2009;35:52-8.  Back to cited text no. 13
    
14.
Athiban PP, Borthakur BJ, Ganesan S, Swathika B. Evaluation of antimicrobial efficacy of aloevera and its effectiveness in decontaminating gutta percha cones. J Conserv Dent 2012;15:3:246-8.  Back to cited text no. 14
    
15.
Tayal E. Current perspectives on use of aloe vera in dentistry. Eur J Med Plants 2014;12:1408-19.  Back to cited text no. 15
    


    Figures

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    Tables

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