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 Table of Contents  
ORIGINAL RESEARCH
Year : 2020  |  Volume : 12  |  Issue : 1  |  Page : 74-79

Compare and evaluate the antibacterial efficacy of sodium hypochlorite and Calendula officinalis against Streptococcus mutans as a root canal irrigating solution: An in vivo study


Department of Conservative Dentistry and Endodntics, Maratha Mandal’s Nathajirao G Halgekar Institute of Dental Sciences and Research Centre, Belagaum, Karnataka, India

Date of Submission18-Jun-2019
Date of Decision01-Oct-2019
Date of Acceptance07-Oct-2019
Date of Web Publication25-Feb-2020

Correspondence Address:
Dr. Viraj S Yalgi
Department of Conservative Dentistry and Endodntics, Maratha Mandal’s Nathajirao G Halgekar Institute of Dental Sciences and Research Centre, Belagaum, Karnataka.
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/jioh.jioh_164_19

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  Abstract 

Aim: Elimination of microorganisms should be of priority in the treatment of any endodontic pathogenic process. The prevalence of microorganisms within the root canal will not cause treatment failures but the absence of microorganisms will certainly lead to success. Materials and Methods: In vivo study was conducted to compare the antibacterial effectiveness of sodium hypochlorite and Calendula officinalis against Streptococcus mutans as root canal irrigants. A total of 40 subjects (older than 18 years of age) were selected with single-rooted teeth, necrotic pulp, and chronic apical periodontitis. Access preparation was carried out, canal was negotiated and sample was taken with paper points before biomechanical preparation. After cleaning and shaping with ProTaper files, irrigation was carried out with sodium hypochlorite and C. officinalis, and was then neutralized with saline and then second sample was taken. Both samples were sent for culturing in pre-reduced medium. Mitis salivarius agar was used for S. mutans, and then calculation of the number of colony forming units per milliliter reduced transport fluid was carried out. Results: The results of the study showed good antibacterial potential for both the medicaments used. Conclusion: Sodium hypochlorite remains the gold standard for bacterial elimination in root canal therapy; however, C. officinalis also has shown promising results and may require further investigation.

Keywords: Antibacterial, Calendula Officinalis, Chronic Apical Periodontitis, Mitis Salivarius Agar, Sodium Hypochlorite, Streptococcus Mutans


How to cite this article:
Yalgi VS, Bhat KG. Compare and evaluate the antibacterial efficacy of sodium hypochlorite and Calendula officinalis against Streptococcus mutans as a root canal irrigating solution: An in vivo study. J Int Oral Health 2020;12:74-9

How to cite this URL:
Yalgi VS, Bhat KG. Compare and evaluate the antibacterial efficacy of sodium hypochlorite and Calendula officinalis against Streptococcus mutans as a root canal irrigating solution: An in vivo study. J Int Oral Health [serial online] 2020 [cited 2020 Apr 3];12:74-9. Available from: http://www.jioh.org/text.asp?2020/12/1/74/279214




  Introduction Top


Most purulent orofacial infections are of odontogenic origin.[1] The dynamics of root canal system infections has been studied along the years. Considerable progress has led in clarifying the etiopathogenesis of endodontic infections due to the development of anaerobic techniques, which has shown their polymicrobial nature and prevalence of obligate anaerobia bacteria.[2] It is established that periapical lesion is directly related to the bacteria and their by-products present in the root canal.[3] Bacteria may still be detected in the root canal system after chemo-mechanical preparation. Their numbers may rapidly increase in the empty canals in between appointments.[4] An important objective of root canal treatment is to clean the root canal system thoroughly free of microbiota and debris. This mainly revolves around a process called chemo-mechanical preparation.[5] Numerous endodontic irrigants have been proposed for clinical use, of which sodium hypochlorite is one of the most preferred irrigants because of its excellent nonspecific proteolysis and antimicrobial activity.[6] The undesirable properties of sodium hypochlorite include unpleasant taste toxicity and its inability to remove the smear layer by itself. Also many studies have shown the limited antimicrobial effectiveness of sodium hypochlorite in vivo, which is also a major drawback.[5] All these drawbacks along with resistant strains being reported have prompted to look for alternatives. Media coverage of homoeopathy over the past few years has tended to concentrate on the very negative position taken by skeptics, whereas possible benefits of homoeopathy are ignored. Homoeopathy is a system of medicine that is based on the principle of like cures like. These medicines can be used in both low and ultramolecular dilutions, and both clinical experience and evidence suggest that they are still effective.[7]Calendula officinalis contains phytochemicals such as flavonoids, saponins, carotenoids, triterpenoids, and tannins, which possess antibacterial activity.[8] Bissa and Bohra[9] studied the antibacterial potential of C. officinalis and found good activity. So the aim of this study was to compare the antibacterial activity of sodium hypochlorite and C. officinalis as root canal irrigating solution.


  Materials and Methods Top


The Ethical Committee, Maratha Mandal’s NGH Institute of Dental Sciences & Research Centre, Belgaum with approval no:MM/BDS/MDS/2017-2018/25 dated 16/12/2017. An in vivo randomized control prospective study was carried out to compare the antibacterial effectiveness of sodium hypochlorite and C. officinalis against Streptococcus mutans as root canal irrigants.

The study cohort was obtained from the Department of Conservative Dentistry and Endodontics, and the samples were analyzed at the Department of Microbiology and Immunology at Maratha Mandal’s Nathajirao G. Halgekar Institute of Dental Sciences and Research Centre, Belagavi, Karnataka, India. Subjects were followed for six months.

Inclusion and exclusion criteria: Subjects with necrotic pulp and chronic apical periodontitis were included in the study. All the subjects satisfied the exclusion criteria listed as following: Subjects who were undergoing antibiotic treatment, immunosuppressant medications, medically compromised subjects such as those infected with human immunodeficiency virus (HIV), with conditions such as acute abscess, osteomyelitis, and malignant lesions, prevailing oral infections such as Herpes Simplex Virus (HSV) type II, Candida albicans, tuberculosis, hematologic factors (bleeding or clotting disorders), idiopathic thrombocytopenic purpura, coagulation defects, subjects with a known history of valvular heart disease, fractured teeth with trauma, teeth with open apices, irreversible pulpitis, grossly decayed teeth, teeth with large cystic lesions, and teeth that have undergone previous endodontic therapy were excluded from the study.

Method of collection of data (including sampling procedure, if any): A total of 40 subjects (older than 18 years of age) were obtained. All the 40 subjects with single-rooted teeth with necrotic pulp and chronic apical periodontitis, confirmed by clinical and radiological examination, were included. The subjects satisfying the clinical requirements were presented with the details of the study and requested to sign informed consents. Ethical committee approval was taken. All the subjects with necrotic pulp and chronic apical periodontitis were selected. The subjects had not undergone any antibiotic therapy at least three months before. The subjects were randomly divided into two groups with 20 subjects in each group.

In vivo study procedure: A rubber dam was used to isolate the tooth. Disinfection of the tooth was carried out using 3% H2O2 [Figure 1]. It was neutralized by sodium thiosulfate. Access cavity was prepared by using two sterile burs, and first caries excavation was conducted with one bur, and access to the root canal was carried out with another sterile bur. Working length determination was performed by using apex locator [Figure 2]. Then samples were taken with paper points before biomechanical preparation [Figure 3]. After cleaning and shaping with ProTaper till file size F3, irrigation with sodium hypochlorite and C. officinalis was carried out [Figure 4] as with the following groups:
Figure 1: Rubber dam application and disinfection with hydrogen peroxide

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Figure 2: Working length determination

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Figure 3: First sample before biomechanical preparation

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Figure 4: Use of the compound as irrigant

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  • Group 1: Irrigating with sodium hypochlorite


  • Group 2: Irrigating using C. officinalis


  • Then sample was taken by inserting the paper point 1mm short of the apex and keeping for 1min, which was then removed and sent in a pre-reduced medium for culturing [Figure 5]. The null hypothesis was that C. officinalis as root canal irrigating solution is not effective.
    Figure 5: Second sample after irrigation with the compound

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    Microbiological examination: The contents of the vials were dispersed and homogenized by using a vortex mixer. A small aliquot was pipetted from the neat solution for preparing a smear. Each sample was serially diluted and inoculated onto agar media. After appropriate incubation, the number of colonies was counted and the growth was analyzed. The number of colony forming units (CFUs) per milliliter of reduced transport fluid (RTF) was calculated for each sample. For S. mutans, Mitis salivarius agar was used, and incubation was carried out in CO2 jar for two days. The strains of bacteria were identified microscopically and also by gram staining, micromorphology, growth characters, and biochemical reactions [Figure 6].
    Figure 6: Colony forming units for Streptococcus mutans

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    Statistical analysis: The results were statistically analyzed by independent sample t test for between-group comparison and paired t test for within-group comparison. The software used was Statistical Package for the Social Science Program by IBM, SPSS Software Version 26, India.


      Results Top


    Results of the study showed good antibacterial potential for both the medicaments used. With sodium hypochlorite, there was no growth of organisms except in sample no. 13, but there was a marked reduction in the number of CFUs [Table 1]. C. officinalis also showed promising results wherein there was no growth in majority of the samples except sample numbers 2, 7, 13, 15, and 18, but there was reduction in the number of CFUs [Table 2]. The P value of 0.05 was taken as a cutoff value.
    Table 1: Group 1 preoperative and after irrigation samples for sodium hypochlorite

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    Table 2: Group 2 preoperative and after irrigation samples for Calendula officinalis

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    Interpretation: No statistically significant difference was observed between Group 1 and Group 2 for pretreatment comparison as well as between Group 1 and Group 2 for posttreatment comparison [Table 3]. There was a significant reduction in CFUs when pretreatment values were compared with posttreatment values in Group 1 as well as when pretreatment values were compared with posttreatment values in Group 2 [Table 4]. Hence, null hypothesis was accepted for between-group comparison as the P value was not significant [Table 3], and alternative hypothesis was accepted for within-group comparison wherein the P value was significant [Table 4].
    Table 3: Between-group comparison of Group 1 and Group 2 by independent samples t test

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    Table 4: Within-group comparison between the pre- and posttreatment values (in 103 colony forming units) by paired t test

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      Discussion Top


    Lots of authors considered microorganisms as primary etiologic agents in endodontic diseases.[10] The key challenges for effective disinfection in endodontics are complexities of the root canal and the structure of dentin. The flow of irrigants, their penetration and exchange within the root canal space, and the forces produced by them occur under irrigation dynamics.[11] In this study, two medicaments were used as intracanal irrigants. Preoperative samples for Group 1 showed growth of S. mutans in all the samples. However, the second sample after thorough biomechanical preparation and irrigation with 5.25% sodium hypochlorite showed elimination of S. mutans in all the samples except in one, but there was a marked reduction in the number of CFUs. Sodium hypochlorite is bactericidal and proteolytic, and it has been used as an endodontic irrigant as early as 1920.[6] Maciel et al.[12] conducted a study to review the importance of the clinical use and outcome of sodium hypochlorite in endodontic treatment of canals, and concluded that it had great fundamental properties and could not be disregarded. It also has the ability to partially neutralize the necrotic tissues or any antigenic or microbial component left in the root canal space.[13] Studies have found that sodium hypochlorite remains stable for 23 months and 10 weeks at different concentrations. However, current literature states that for effective root canal disinfection and antimicrobial efficacy, fresh sodium hypochlorite solution is prepared.[14] The advantages of using herbs as antimicrobial agents are minimal side effects, cost-effective better patient tolerance, and lastly, renewable in nature.[15] Preoperative samples of Group 2 showed colonies of S. mutans in all the samples. After through biomechanical preparation and the use of C. officinalis as irriganting solution there was complete elimination of S. mutans in majority of samples. However five samples, showed persistence of S. mutans, but with reduction in the number of CFUs.[16] As revealed by many studies, the presence of terpene alcohols and terpene lactones of the essential oil in C. officinalis shows antimicrobial activities. It is also interesting to note that the action of extracts of C. officinalis is nontoxic.[17] In a study, Eslami et al. showed that gram-positive bacteria such as Streptococcus and Staphylococcus were sensitive to C. officinalis.[18] Calendula extract heals wounds and is an antiseptic and in addition improves blood flow to the affected area.[19]

    Limitations of study: The limitations of this study were small sample design and the limited literature available on the use of C. officinalis in endodontics.

    Future scope: However, owing to the presence of various phytochemicals with significant antibacterial potential in C. officinalis, experimental studies are needed to determine the quality and extent of effect of the aforementioned substances on pathogenic microorganisms.

    The use of plant extracts as well as other alternative forms of medical treatments was enjoying great popularity in the late 1990s. Sodium hypochlorite remains the gold standard for bacterial elimination in root canal therapy. However, with C. officinalis, further scientific investigation can be carried out to elucidate its antimicrobial effectiveness and mechanisms inside the root canal system as well as the need to associate it with other medicaments.

    Acknowledgement

    We are grateful and acknowledge the support received for the study from the Department of Conservative Dentistry and Endodontics and Department of Microbiology and immunology, Maratha Mandal’s Nathajirao G. Halgekar Institute of Dental Sciences and Research Centre, Belagavi, Karnataka, India.

    Financial support and sponsorship

    Nil.

    Conflicts of interest

    There are no conflicts of interest.



     
      References Top

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    Kuriyama T, Karasawa T, Nakagawa K, Yamamoto E, Nakamura S. Bacteriology and antimicrobial susceptibility of gram-positive cocci isolated from pus specimens of orofacial odontogenic infections. Oral Microbiol Immunol 2002;17:132-5.  Back to cited text no. 1
        
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    Lana MA, Ribeiro-Sobrinho AP, Stehling R, Garcia GD, Silva BK, Hamdan JS, et al. Microorganisms isolated from root canals presenting necrotic pulp and their drug susceptibility in vitro. Oral Microbiol Immunol 2001;16:100-5.  Back to cited text no. 2
        
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    Kakehoshi S, Stanly HR, Fitzgerald RJ. The effect of surgical exposures of dental pulps in germ free and conventional laboratory rats. Oral Surg Oral Med Oral Pathol Radiol Endod 1965;20:340-9.  Back to cited text no. 3
        
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    Eames S, Darby P. Homeopathy and its ethical use in dentistry. Br Dent J 2011;210:299-301.  Back to cited text no. 7
        
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    Pazhohideh Z, Mohammadi S, Bahrami N, Mojab F, Abedi P, Maraghi E. The effect of Calendula officinalis versus metronidazole on bacterial vaginosis in women: A double-blind randomized controlled trial. J Adv Pharm Technol Res 2018;9:15-9.  Back to cited text no. 8
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    Lin YH, Mickel AK, Chogle S. Effectiveness of selected materials against Enterococcus faecalis: Part 3. The antibacterial effect of calcium hydroxide and chlorhexidine on Enterococcus faecalis. J Endod 2003;29:565-6.  Back to cited text no. 10
        
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    Abuhaimed T, Ensanya A, Abou N. Sodium hypochlorite irrigation and its effect on bond strength to dentin. Biomed Res Int 2017;17:1-8.  Back to cited text no. 13
        
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    Fabian TM, Walker SE. Stability of sodium hypochlorite solutions. Am J Hosp Pharm 1982;39:1016-7.  Back to cited text no. 14
        
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    Saxena D, Saha SG, Saha MK, Dubey S, Khatri M. An in vitro evaluation of antimicrobial activity of five herbal extracts and comparison of their activity with 2.5% sodium hypochlorite against Enterococcus faecalis. Indian J Dent Res 2015;26:524-7.  Back to cited text no. 15
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    Uribe-Fentanes LK, Soriano-Padilla F, Pérez-Frutos JR, Veras-Hernández MA. [Action of Calendula officinalis essence on bone preservation after the extraction]. Rev Med Inst Mex Seguro Soc 2018;56:98-105.  Back to cited text no. 16
        
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    Roopashree T, Raman D, Shobha RH. Antibacterial activity of antipsoriatic herbs Cassia tora, Momordica charantia and Calendula officinalis. Nat Pro 2008;1:20-8.  Back to cited text no. 17
        
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    Eslami G, Taheri S, Ayatollahi SA, Mohazzabiyyeh RZ. Effects of Calendula officinalis extract on bacteria isolated. Res Med. 2011;34:214-218.  Back to cited text no. 18
        
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        Figures

      [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6]
     
     
        Tables

      [Table 1], [Table 2], [Table 3], [Table 4]



     

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