|Year : 2020 | Volume
| Issue : 2 | Page : 61-65
Comparative evaluation of smear layer removal efficacy of three herbal irrigants in the apical third of the root canal: An in vitro scanning electron microscopic study
GR Ranjitha, Shreya Rajesh Mutha, R Anithakumari, R Vikram, Naveen Kumar, AK Shivekshith
Department of Conservative Dentistry and Endodontics, Vokkaligara Sangha Dental College and Hospital, Bengaluru, Karnataka, India
|Date of Submission||25-Jan-2020|
|Date of Acceptance||13-Apr-2020|
|Date of Web Publication||08-Jul-2020|
Dr. R Anithakumari
Department of Conservative Dentistry and Endodontics, Vokkaligara Sangha Dental College and Hospital, K R Road, V V Puram, Bengaluru - 560 004, Karnataka
Background: Smear layer removal from the apical third of the root canal is critical and the widely used chemical irrigants have been associated with detrimental effects. Hence, the study aimed to evaluate the smear layer removal efficacy of three herbal extracts used as endodontic irrigants in the apical third of the root canal. Materials and Methods: Forty-eight single-rooted premolars were divided into three experimental groups and a control group and were irrigated with Neem extract (Group 1), Orange peel extract (Group 2), Tulasi extract (Group 3), 5.25% sodium hypochlorite (Group 4) and their effect on smear layer removal was examined under the field-emission scanning electron microscope. Statistical Analysis: Kruskal–Wallis test followed by Mann–Whitney post-hoc analysis was done to compare the mean smear layer scores between the four groups in the apical third of the root sections. Results: The lowest mean remaining smear layer scores was shown by Neem extract (0.83), followed by sodium hypochlorite (1.08), orange peel extract (1.67), and the highest for Tulasi extract (2.42). On intergroup comparison, Neem extract showed statistically significant less smear layer scores in comparison to Orange peel extract (P = 0.006) and Tulasi (P = 0.001). Neem showed no statistically significant difference with sodium hypochlorite (P = 0.37). Conclusions: Neem extract can be considered as an efficient smear layer removal irrigant in the apical third of the root canal.
Keywords: Herbal irrigants, Neem, Orange peel extract, root canal irrigants, smear layer, Tulasi
|How to cite this article:|
Ranjitha G R, Mutha SR, Anithakumari R, Vikram R, Kumar N, Shivekshith A K. Comparative evaluation of smear layer removal efficacy of three herbal irrigants in the apical third of the root canal: An in vitro scanning electron microscopic study. SRM J Res Dent Sci 2020;11:61-5
|How to cite this URL:|
Ranjitha G R, Mutha SR, Anithakumari R, Vikram R, Kumar N, Shivekshith A K. Comparative evaluation of smear layer removal efficacy of three herbal irrigants in the apical third of the root canal: An in vitro scanning electron microscopic study. SRM J Res Dent Sci [serial online] 2020 [cited 2021 Jan 22];11:61-5. Available from: https://www.srmjrds.in/text.asp?2020/11/2/61/289167
| Introduction|| |
The key factors for successful endodontic treatment depends on the removal of microbial biofilms and the smear layer from the root canal walls., Smear layer embedding debris and microorganisms formed during the mechanical preparation is removed by the action of chemical irrigants. However, the drawbacks associated with chemical irrigants led the researchers to find a more biocompatible and dentin friendly irrigants., Natural agents such as herbal extracts are gaining significant prominence as endodontic irrigants due to their antioxidant, anti-inflammatory, immune-modulatory, antibacterial, antifungal, antiviral, antioxidant, anti-carcinogenic, and radical scavenging activity of the active constituents.,
In literature, there is no such study comparing the smear layer removal efficacy of Neem, Orange peel, and Tulasi extracts in the apical third of the root canal. The aim of this study was to evaluate the smear layer removal efficacy of these herbal irrigants in the apical third of root canal in comparison with sodium hypochlorite using scanning electron microscopic (SEM) image analysis.
| Materials and Methods|| |
Preparation of the herbal extracts
For the preparation of herbal extract, each powder was weighed (50 g) and mixed with distilled water (500 ml) and boiled at 100°C to get 50 ml of herbal extract. The prepared solutions were filtered using Whatman filter paper, and the final irrigating solutions were obtained and stored in opaque bottles.
Sample size estimation
The sample size has been estimated using the G Power software v. 184.108.40.206.
Considering the effect size to be measured (f) at 52% (based on the results of the previous literature by Sebatni MA, 2017), power of the study at 80% and the margin of the error at 5%, the total sample size needed is 48. Each group will consist of 12 samples.
(12 samples × 4 groups = 48 samples).
Forty-eight (n = 48) extracted lower premolars having a single canal and fully developed apices were selected after carefully examining the intraoral periapical radiograph. The teeth were randomly divided following a simple random method into three experimental groups and a control group of 12 samples each and stored in distilled water until use.
Access opening, working length determination using radiograph followed by cleaning and shaping was done up to apical size 30, 9% taper using Rotary Protaper gold (Densply, India). During the instrumentation, the root canals were simultaneously irrigated with 5 ml of Neem extract in Group 1, 5 ml of Orange peel extract in Group 2, 5 ml of Tulasi extract in Group 3, and 5 ml of 5.25% sodium hypochlorite in Group 4. Sterile distilled water was used as a final rinse in all the sampled teeth and was then stored at humidity. Thereafter, the teeth were decoronated at the cementoenamel junction using the diamond disc to obtain a standard root length of 10 mm. Buccal and lingual longitudinal grooves were placed and the roots were split into two halves using a chisel and mallet.
Scanning electron microscopic evaluation
The coded specimens were then mounted onto metallic stubs with carbon strip backing with the entire root canal visible and facing upward and then subsequently viewed under the SEM. The photomicrographs were obtained at ×1000 magnification using digital image analysis software and were stored appropriately for subsequent analysis. The most representative micrographs were taken of the specimen and were recorded for apical thirds. The results were then scored following Rome et al. criteria (0 = heavy smear layer, indiscernible tubular aperture, 1 = thin smear layer cover the surface outline of dentinal tubules indiscernible, tubular aperture indicated by a crack, 2 = smear layer present only in the aperture of the dentinal tubules, 3 = no smear layer, dentinal tubules open and free of debris) by the operator and tabulated.
Values obtained were subjected to statistical analysis using Statistical Package for Social Sciences (SPSS) for Windows Version 22.0 Released 2013 and IBM Corp., Armonk, NY, USA Descriptive analysis was done to express the remaining Smear layer scores in terms of mean and standard deviation. Inferential Statistics: Kruskal–Wallis Test followed by Mann–Whitney post hoc Analysis was done to compare the mean smear layer scores between four groups in the apical third of the root sections. The level of significance was set at P < 0.05.
The normality test was performed to check the distribution of data and it was observed that the data were not following normal distribution. Hence, the data analysis was performed using nonparametric tests as indicated above. Bonferroni's correction is deemed suitable for multiple comparison between groups when the data follows a normal distribution. In the present study, the scoring of the smear layer removal is done using ordinal scores and follows a nonnormal distribution; it is not wise to perform parametric tests like one-way ANOVA followed by Bonferroni's correction.
| Results|| |
None of the irrigants were able to remove the smear layer completely from the apical third of the root canal. The mean of the remaining smear layer scores was lowest for Group 1 - Neem extract (0.83), followed by Group 4 - sodium hypochlorite (1.08), Group 2 - Orange peel extract (1.67), and the highest for Group 3 - Tulasi extract (2.42) P < 0.001 [Table 1].
On intergroup comparison, Group 1 - Neem extract showed statistically significant less smear layer scores in comparison to Group 2 - Orange peel extract (P = 0.006) and Group 3 - Tulasi extract (P = 0.001). No statistically significant difference was observed between Group 1 - Neem extract and Group 4 - sodium hypochlorite (P = 0.37) [Table 2].
|Table 2: Comparison of mean between groups using Mann-Whitney post hoc analysis|
Click here to view
| Discussion|| |
Ando and Hoshino mentioned the smear layer as a potential substrate for growth and ingress of bacteria. The superficial smear layer is 1–2 μm thick and the deeper smear plug is of up to 40 μm thickness. In a study by Karagoz-Kucukay and Bayirli in 1994 smear layer has been postulated to be an avenue for leakage.
It was observed that smear was pushed up to a depth of 110 μm on using surface-active agents due to the adhesive forces and capillary action between dentinal tubules and smear layer. Various strategies have been proposed for the removal of intraradicular smear layer and chemical irrigants such as EDTA, citric acid, and maleic acid have been advocated for the same. Studies have concluded that the use of a combination of 2.5%–5% sodium hypochlorite and 10%–17% EDTA is effective in the removal of organic and inorganic debris.,
Smear layer removal within the root canal is of much importance as it aids in easy penetration of irrigants by opening up of dentinal tubules thereby improving the efficacy and reduction in time required for disinfection of canal. It further facilitates better adaptation of the obturating material and sealer to the canal walls resulting in the better apical seal. However, disinfection and smear layer removal in the apical third of root canal is critical due to a multitude of factors such as limited space, low permeability, and complex anatomical configuration.,
Many controlled clinical trials and pharmacological studies have recognized the potential of bioactive compounds in medicinal plants., The biodegradability, nontoxicity, and cost-effectiveness have made herbal medicine a popular therapeutic agent, which necessitates the need to explore, understand, and extrapolate their implication in dental practice.,,
SEM, which can detect accurate surface characteristics, was used to evaluate the presence of smear layer after using various irrigating solutions.,
In this study, Azadirachta Indica (margosa tree or Indian neem) showed the highest efficacy among all the groups [Figure 1]. A similar result was seen in a study done by Sebatni and Kumar where Neem showed the highest amount of smear layer removal in comparison to sodium hypochlorite and Orange oil. In 1992, Neem was regarded as “a tree for solving global problems” by the US National Academy of Sciences due to its efficacy in various applications. The anti-inflammatory, antibacterial, antifungal, and immunomodulatory properties make it a potential endodontic irrigant and medicament.,, The greater efficacy of Neem may be due to the various active phytoconstituents such as acid metabolites, flavonoids, isoprenoids, alkaloids, glycosides, steroids, and tannins which makes it a material of choice for root canal irrigation and also an alternative to harmful chemical irrigants such as sodium hypochlorite.
|Figure 1: Scanning electron microscopic image of root canal wall in the apical third after irrigation with neem |
Click here to view
The vitality of bacteria is impeded by inhibiting the respiratory chain due to its chemically diverse and biologically active components., It is also shown to be effective against various microorganisms found in the oral microflora such as Enterococcus faecalis and Candida albicans. As compared to sodium hypochlorite (hypochlorite accidents) Neem is less likely to cause severe harm to patients due to its high biocompatibility. Thus, it can be used as an effective herbal alternative to the more commonly used irrigant sodium hypochlorite. However, the major drawback of Neem is Nimbidin's characteristic bitter taste which has to be overcome by the addition of sweeteners.,
Orange peel extract (Citrus aurantifolia) contains 88% water, 6%–8% citric acid, 2% potassium citrate, and other substances. Owing to the citric acid as a basic constituent in C. aurantifolia, researchers have evaluated it for its possible role in the endodontic smear layer removal; though, the results were disagreeable., Flavonoids, consisting mainly of polymethoxylated flavonoids, terpenoids, such as limonene and linalool, aliphatic hydrocarbon alcohols, aldehydes such as octanal and other volatile oils make up the major phytochemical constituents of orange peel. Owing to the natural constituents, Orange peel extract does not have any harmful effect concerning the oral tissues.
In the present study, orange peel extract exhibited less amount of smear layer removal efficacy [Figure 2] compared to control group [Figure 3] and Neem which is in agreement with the study done by Bolhari et al. in which alcoholic and completed mixtures of C. aurantifolia extracts were not able to effectively remove smear layer compared with 17% EDTA. The reduced efficacy might be due to the absence of required amount of acid metabolites which is needed for the removal or it might be due to the lower penetration of irrigants into the deeper apical part due to the high surface tension of the oil-based orange peel extract irrigant.
|Figure 2: Scanning electron microscopic image of root canal wall in the apical third after irrigation with orange peel extract |
Click here to view
|Figure 3: Scanning electron microscopic image of root canal Wall in the apical third after irrigation with sodium hypochlorite|
Click here to view
Tulasi (Ocimum sanctum) due to the presence of active components such as tannins (4.6%), essential oil (up to 2%), Aerosol acid, Oleanolic acid, β-element (11%), β-caryophyllene and germacrene are being used as a medicinal plant since ancient days.,
The antimicrobial activity of Tulasi against both Gram-negative and Gram-positive microorganisms is due to their ability to form complexes with enzymes or substrates required by microorganisms for their functioning or may be related to its action on the cell membrane of the microorganisms causing protein leakage.,,
The lesser concentration of active components may be the reason for the reduced efficacy of O. sanctum as compared to other groups in this study [Figure 4]. A study conducted by Kumar et al. also concluded that Neem leaf extract was more efficient then Tulasi extract in smear layer removal.
|Figure 4: Scanning electron microscopic image of root canal wall in the apical third after irrigation with tulasi |
Click here to view
This being an in vitro study, various combinations and concentrations should be tried in animal models to obtain more efficient herbal root canal disinfectant and in vivo studies to be conducted to determine the long-term prognosis of root canal treatment.
Herbal irrigants are easily available, cost-effective, have increased shelf life, biocompatible, and lack microbial resistance which makes them potential alternative irrigating solutions.
| Conclusions|| |
Within the limitations of the presentin vitro research, it can be concluded that Neem extract can be considered as an efficient smear layer removal irrigant in the apical third of the root canal.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Gupta A, Duhan J, Tewari S, Sangwan P, Yadav A, Singh G, et al
. Comparative evaluation of antimicrobial efficacy of Syzygium aromaticum
, Ocimum sanctum
and Cinnamomum zeylanicum
plant extracts against Enterococcus faecalis
: A preliminary study. Int Endod J 2013;46:775-83.
Choudhary E, Indushekar KR, Saraf BG, Sheoran N, Sardana D, Shekhar A. Exploring the role of Morinda citrifolia
and Triphala juice in root canal irrigation: An ex vivo
study. J Conserv Dent 2018;21:443-9.
] [Full text]
Susan AC, Bharathraj AR, Praveen M, Kumar NS, Karunakaran JV. Intraradicular smear removal efficacy of Triphala as a final rinse solution in curved canals: A scanning electron microscope study. J Pharm Bioallied Sci 2019;11:S420-8.
Chhabra N, Gyanani H, Kamatagi L. Smear layer removal efficacy of combination of herbal extracts in two different ratios either alone or supplemented with sonic agitation: An in vitro
scanning electron microscope study. J Conserv Dent 2015;18:374.
] [Full text]
Sebatni MA, Kumar AA. Smear layer removal efficacy of herbal extracts used as endodontic irrigants: Anin vitro
study. Endodontology 2017;29:35-8. [Full text]
Sowjanyaa J, Thomas T, Chandana CS. Comparative evaluation of the efficacy of smear layer removal by ethylenediaminetetraacetic acid, Triphala, and German chamomile as irrigants – A scanning electron microscopy study. J Adv Pharm Edu Res 2017;7:261-71.
Kumar A, Sarthaj AS, Antony SM. Comparative evaluation of antibacterial and smear layer removal efficacy of two different herbal irrigants: Anin vitro
study. Int Healthc Res J 2018;1:350-4.
Rome WJ, Doran JE, Walker WA 3rd
. The effectiveness of Gly-oxide and sodium hypochlorite in preventing smear layer formation. J Endod 1985;11:281-8.
Ando N, Hoshino E. Predominant obligate anaerobes invading the deep layers of root canal dentin. Int Endod J 1990;23:20-7.
Karagöz-Küçükay I, Bayirli G. An apical leakage study in the presence and absence of the smear layer. Int Endod J 1994;27:87-93.
Violich DR, Chandler NP. The smear layer in endodontics – A review. Int Endod J 2010;43:2-15.
Paul ML, Mazumdar D, Niyogi A, Baranwal AK. Comparative evaluation of the efficacy of different irrigants including MTAD under SEM. J Conserv Dent 2013;16:336-41.
] [Full text]
Lottanti S, Gautschi H, Sener B, Zehnder M. Effects of ethylenediaminetetraacetic, etidronic and peracetic acid irrigation on human root dentine and the smear layer. Int Endod J 2009;42:335-43.
Ribeiro RG, Marchesan MA, Silva RG, Sousa-Neto MD, Pécora JD. Dentin permeability of the apical third in different groups of teeth. Braz Dent J 2010;21:216-9.
Ruddle CJ. Hydrodynamic disinfection: Tsunami endodontics. Dent Today 2007;26:110, 112, 114-7.
Krajczár K, Tigyi Z, Papp V, Marada G, Sára J, Tóth V. Chemomechanical preparation by hand instrumentation and by Mtwo engine-driven rotary files, an ex vivo
study. J Clin Exp Dent 2012;4:e146-50.
Pujar M, Makandar S. Herbal usage in endodontics – A review. Int J Contemp Dent 2011;2:34-7.
Gambarini G, Laszkiewicz J. A scanning electron microscopic study of debris and smear layer remaining following use of GT rotary instruments. Int Endod J 2002;35:422-7.
Lim TS, Wee TY, Choi MY, Koh WC, Sae-Lim V. Light and scanning electron microscopic evaluation of Glyde File Prep in smear layer removal. Int Endod J 2003;36:336-43.
Singh H, Kaur M, Dhillon JS, Batra M, Khurana J. Neem: A magical herb in endodontics. Stomatological Dis Sci 2017;1:50-4.
Bohora AA, Hegde VI, Kokate SH. Comparison of the antibacterial efficiency of Neem leaf extract and 2% sodium hypochlorite against E. faecalis
, C. albicans
and mixed culture – Anin vitro
study. Endodontology 2010;22:8-12.
Dutta A, Kundabala M. Comparative anti-microbial efficacy of Azadirachta indica
irrigant with standard endodontic irrigants: A preliminary study. J Conserv Dent 2014;17:133-7.
] [Full text]
Ravishankar P, Lakshmi T, Kumar AS. Ethno-botanical approach for root canal treatment-an update. J Pharma Sci Res 2011;3:1511-9.
Biswas K, Chattopadhyay I, Banerjee RK, Bandyopadhyay U. Biological activities and medicinal properties of Neem (Azadirachta indica
). Curr Sci 2002;82:1336-45.
Vinothkumar TS, Rubin MI, Balaji L, Kandaswamy D.In vitro
evaluation of five different herbal extracts as an antimicrobial endodontic irrigant using real time quantitative polymerase chain reaction. J Conserv Dent 2013;16:167-70.
] [Full text]
John P, Gopalakrishnan, Dinesh K, Romel J. Herbal root canal irrigants: A review. J Odontol Res 2015;3:9-14.
Sharifian MR, Shokouhinejad N, Monsef-Esfahani HR, Aligholi M, Amjadi M. Antimicrobial effect of Citrus aurantifolia
extract on Enterococcus faecalis
within the dentinal tubules in the presence of smear layer. J Dent Med 2011;24:148-55.
Erukainure OL, Ebuehi OA, Iqbal Chaudhary M, Mesaik MA, Shukralla A, Muhammad A, et al
. Orange peel extracts: Chemical characterization, antioxidant, antioxidative burst, and phytotoxic activities. J Diet Suppl 2016;13:585-94.
Bolhari B, Sharifian MR, Aminsobhani M, Monsef Esfehani HR, Tavakolian P. Assessing the efficacy of Citrus aurantifolia
extract on smear layer removal with scanning electron microscope. Iran Endod J 2012;7:88-97.
Oyedemi SO, Okoh AI, Mabinya LV, Pirochenva G, Afolayan AJ. The proposed -terpinene against Listeria (mechanism of bactericidal action of eugenol, α-terpineol and monocytogenes, Streptococcus pyogenes
, Proteus vulgaris
and Escherichia coli
. Afr J Biotechnol 2009;8:1280-6.
Maheshwari ML, Singh BM, Cupra R, Chien MJ. Essential oil of sacred basil (Ocimum sanctum
). Indian Perfum 1987;31:137-45.
Akiyama H, Fujii K, Yamasaki O, Oono T, Iwatsuki K. Antibacterial action of several tannins against Staphylococcus aureus
. J Antimicrob Chemother 2001;48:487-91.
[Figure 1], [Figure 2], [Figure 3], [Figure 4]
[Table 1], [Table 2]