Evaluation of debris and smear layer removal efficacy of different irrigating solutions using the C-RCC irrigation system: An in vitro SEM study

Abstract

Background:

Conventional use of sodium hypochlorite (NaOCl) and ethylenediaminetetraacetate (EDTA) for debris and smear layer removal may weaken dentin and prolong clinical procedures, prompting a search for safer, more efficient, and multifunctional irrigants. Thus, the present study aimed to evaluate debris and smear layer removal efficacy of different irrigating solutions using C-RCC irrigator.

Materials and Methods:

Eighty extracted single-rooted mandibular premolars were decoronated, instrumented with ProTaper Universal files, and randomly allocated into four groups (n = 20): Group I-NaOCl-EDTA, Group II-NaOCl-Neem extract, Group III-Superoxidised water, and Group IV-Normal saline. Irrigation was performed using C-RCC irrigator. Samples were longitudinally sectioned and examined under scanning electron microscopic at × 500 and × 1000 magnification. Hulsmann’s scoring criteria were followed, and data were analyzed with Kruskal–Wallis and post hoc Dunn tests.

Results:

All irrigants except saline showed varying cleaning abilities. NaOCl–EDTA (debris = 1, smear layer = 1.25) and superoxidized water (debris = 1.2, smear layer = 1.45) demonstrated similar efficacy with statistically insignificant difference, although values were slightly higher for NaOCl–EDTA and both were significantly more effective than NaOCl–neem extract and saline. Coronal thirds showed better cleanliness than the middle and apical thirds.

Conclusion:

NaOCl–EDTA remained the most effective regimen. However, superoxidised water showed promise as a single, efficient irrigant.

INTRODUCTION

Root canal therapy is performed with objective of disinfecting root canal system by combining mechanical instrumentation and chemical irrigation, which is subsequently completed by three-dimensional obturation.[1,2]

Debris and smear layer are the unavoidable byproducts formed after cleaning and shaping of the root canal. The smear layer is composed of organic and inorganic constituents. Organic matter comprises remnants of pulp tissue (necrotic or vital), odontoblastic processes, microorganisms, blood cells, dentinal collagen and glycosaminoglycans, whereas inorganic matter is largely composed of dentinal debris. Smear layer retention may harbor microorganisms, compromise disinfection, and adversely affect sealer adhesion, thereby necessitating the use of a chemical irrigant.[3]

Sodium hypochlorite (NaOCl), the only irrigant capable of dissolving tissue, is effective against the organic portion of the smear layer only. Its inability to eliminate inorganic constituents requires its combination with ethylenediaminetetraacetic acid (EDTA), which chelates calcium ions and removes the inorganic portion of dentin.[4,5] Nevertheless, prolonged application of this combination leads to dentin erosion and alterations in dentinal microstructure by disrupting the organic–inorganic balance.[6]

Increasing interest in biocompatible alternatives has encouraged the exploration of plant-based irrigants. Neem (Azadirachta indica) contains several bioactive compounds such as nimbin, nimbolide, and azadirachtin, having potential smear layer removal ability and antimicrobial properties with fewer reported adverse effects, making it a promising herbal substitute.[7]

The requirement for multiple irrigants and an intermediate neutralizing step renders the NaOCl–EDTA protocol time-consuming and technique-sensitive, highlighting the need for a single irrigant with comparable efficacy and fewer drawbacks. Superoxidized water, which is created by electrolyzing a diluted sodium chloride solution, has demonstrated promise as a single irrigant that may remove smear layers and debris, cutting down on procedure time and effort.[6,8]

Conventional needle-syringe irrigation often struggles to deliver and distribute irrigants effectively throughout the complex root canal system. In addition, controlling and standardizing the flow rate of irrigant can be challenging.[9]

Coxo’s root canal irrigation and suction system (C-RCC irrigator), developed by Dr. Mandar Pimprikar, is a novel irrigation device that allows simultaneous delivery of irrigants and evacuation of waste fluid using positive and negative pressure through its two specialized tips. It uses a continuous warm activated irrigations and suction of irrigant, causing flushing of the root canal efficiently. The tip A, which applies positive pressure, removes debris from the coronal and middle third, whereas tip B, which applies negative pressure remove nano debris from the apical third of the canal. A key advantage of this system is the continuous delivery of irrigants at a preset and constant flow rate of 5ml/min, which improves standardization and reduces operator’s fatigue during irrigation. Consequently, it may enhance the overall efficacy of root canal irrigation.[10]

Thus, the present study aimed to evaluate the debris and smear layer removal efficacy of different irrigating solutions using the C-RCC irrigation system under a scanning electron microscope.

MATERIALS AND METHODS

Preparation of neem leaf extract

Hundred grams of shade-dried and finely powdered neem leaves were subjected to Soxhlet extraction at 60°C using 1000 ml of ethanol [Figure 1a].[11] The obtained solution was filtered with Whatman No. 42 filter paper [Figure 1b]. The solution was concentrated using rotary evaporator and redissolved in 100 ml dimethyl sulfoxide [Figure 1c].[12] The prepared neem leaf extract was then stored in an amber-colored container at 4°C until use.

Figure 1.

Methodology, (a) Soxhlet extraction, (b) Filtration, (c) Neem leaf extract, (d) Buccal and lingual grooves without perforating main canal, (e) Cleaning and shaping till size F3 using Protaper Universal rotary instrument, (f) CRCC irrigator, (g) Irrigation with CRCC irrigator during cleaning and shaping using Tip A, (h) Irrigation with CRCC irrigator after cleaning and shaping using Tip B

Sample selection

Eighty freshly extracted human mandibular first premolars with single, straight canals and mature apices were selected. Teeth with cracks, root caries, previous endodontic treatment, resorption, or calcified canals were excluded from the study. The samples were disinfected using 10% formalin solution and stored in 0.1% thymol solution until use.

A low-speed diamond disc was used for decoronation of sample teeth, providing a uniform 12mm root length. The working length was determined by advancing #15 K-file until its tip appeared at the apical foramen and reducing the measured length by 1 mm. Thereafter, a glide path was established with a #20 K-file.

Buccal and lingual vertical grooves were made with a diamond disc without perforating the root canal [Figure 1d]. The samples were then mounted in silicon-based impression material for stabilization. Root canals were prepared with ProTaper Universal rotary files (Dentsply, Switzerland) up to size F3, in accordance with the manufacturer’s instructions [Figure 1e].

The samples were randomly assigned to four groups (n = 20) based on the irrigation protocol:

• Group I: 5.25% NaOCl (SafeEndo, India) - 17% EDTA (SafeEndo, India) (Positive control)

• Group II: 5.25% NaOCl-Neem leaf extract

• Group III: Superoxidised water (OXUM, Alkem Labs, India)

• Group IV: Normal saline (Vision Parentral, India) (Negative control).

All irrigation procedures were performed using the C-RCC irrigator (COXO, China) [Figure 1f].

Irrigation protocol

Irrigation was carried out using 5ml of the assigned solution for 1 min. Initial irrigation was done using tip A positioned in the middle third of the canal [Figure 1g]. After completion of the biomechanical preparation, irrigation was done using tip B placed till the working length [Figure 1h]. For Groups I and II, NaOCl was used during cleaning and shaping, followed by the respective secondary irrigant after preparation. In Groups III and IV, the same irrigating solution was used throughout. A final rinse was carried out in all groups using 5ml distilled water for 1 min to eliminate any remaining irrigant.

Scanning electron microscopic preparation and evaluation

The samples were retrieved from the silicon material; the roots were split longitudinally using an orthodontic wire cutter, and either the mesial or distal half was selected. The samples were dehydrated through a series of increasing ethanol concentrations and labeled A, B, and C to represent the apical, middle, and coronal thirds at 3mm, 6mm, and 9mm from the root apex, respectively. Each sample was mounted on aluminum stubs, gold-sputtered, and examined under SEM at ×500 and ×1000 magnifications for debris and smear layer evaluation, respectively [Figure 2]. Scoring was performed by two blinded investigators (kappa score of 0.83) according to the Hulsmann criteria (1997).[13]

Figure 2.

Scanning electron microscopic analysis. Reference images for remaining debris (×500) and smear layer (×1000), (a) Group I (5.25% sodium hypochlorite [NaOCl] with 17% EDTA), (b) Group II (5.25% NaOCl with neem leaf extract), (c) Group III (Superoxidised water), (d) Group IV (normal saline)

Scores for residual debris

• Clean canal walls; only a few small debris particles

• Few small agglomerations of debris

• Moderate debris; <50% of the surface covered

• Substantial debris; >50% of the surface covered

• Complete or nearly complete coverage by debris.

Scores for residual smear layer

1. No smear layer; dentinal tubules open

2. Small amount of smear layer; some dentinal tubules open

3. Homogeneous smear layer covering most of the surface; few tubules open

4. Homogeneous smear layer covering the surface; no open tubules

5. Heavy, nonhomogeneous smear layer covering the surface.

Statistical analysis

Data wwere analyzed using: Kruskal–Wallis test for overall intergroup comparison and Dunn’s post hoc test for pairwise comparisons. Significance was set at P < 0.05.

RESULTS

All groups except normal saline demonstrated effective debris and smear layer removal [Table 1]. NaOCl–EDTA and superoxidized water demonstrated the highest efficacy, with statistically insignificant difference between them (P > 0.05), although NaOCl–EDTA showed slightly higher values. This was followed by NaOCl–neem extract, while normal saline exhibited the lowest effectiveness [Table 2]. Across the entire canal length, the coronal third showed better cleanliness than middle and apical thirds.

Table 1.

Descriptive statistics for debris and smear layer scores for the study groups at three levels of cross-sections

Groups (n=20)	Sections	Debris				Smear layer
		Mean±SD	Median	Upper quartile	Lower quartile	Mean±SD	Median	Upper quartile	Lower quartile
Group I (NaOCl-EDTA)	Coronal	1.00±0.00	1.00	1.00	1.00	1.25±0.44	1.00	1.00	1.75
	Middle	1.20±0.41	1.00	1.00	1.00	1.30±0.47	1.00	1.00	2.00
	Apical	1.25±0.44	1.00	1.00	1.75	1.80±0.62	2.00	1.00	2.00
Group II (NaOCl-Neem extract)	Coronal	1.50±0.83	1.00	1.00	2.00	2.20±0.52	2.00	2.00	2.75
	Middle	1.50±0.76	1.00	1.00	2.00	2.50±0.51	2.50	2.00	3.00
	Apical	1.70±0.87	1.50	1.00	2.00	2.65±0.49	3.00	2.00	3.00
Group III (Superoxidised water)	Coronal	1.20±0.41	1.00	1.00	1.00	1.45±0.51	1.00	1.00	2.00
	Middle	1.20±0.41	1.00	1.00	1.00	1.45±0.51	1.00	1.00	2.00
	Apical	1.45±0.51	1.00	1.00	2.00	1.55±0.51	2.00	1.00	2.00
Group IV (Normal saline)	Coronal	2.60±0.60	3.00	2.00	3.00	4.00±0.00	4.00	4.00	4.00
	Middle	2.75±0.79	3.00	2.00	3.00	4.00±0.00	4.00	4.00	4.00
	Apical	3.05±1.00	3.00	2.00	4.00	4.15±0.37	4.00	4.00	4.00

n: Number of values per study group, SD: Standard deviation, EDTA: Ethylenediaminetetraacetate, NaOCl: Sodium hypochlorite

Table 2.

Mean rank difference between the study groups for the debris and smear layer scores in the coronal, middle and apical thirds along with post hoc comparisons

Dependent variable	I Groups	J Groups	Debris		Smear layer
			Rank sum difference (I−J)	P	Rank sum difference (I−J)	P
Coronal	Group I	Group II	−13.50	0.2079 (NS)	−23.38	0.005 (HS)
		Group III	−6.700	>0.9999 (NS)	−5.500	>0.9999 (NS)
		Group IV	−41.80	<0.0001 (HS)	−49.63	<0.0001 (HS)
	Group II	Group III	6.800	>0.9999 (NS)	17.88	0.063 (NS)
		Group IV	−28.30	<0.0001 (HS)	−26.25	0.001 (HS)
	Group III	Group IV	−35.10	<0.0001 (HS)	−44.13	<0.0001 (HS)
Middle	Group I	Group II	−7.88	0.2079 (NS)	−26.25	0.0012 (HS)
		Group III	0.00	>0.9999 (NS)	−3.75	>0.9999 (NS)
		Group IV	−36.93	<0.0001 (HS)	−50.00	<0.0001 (HS)
	Group II	Group III	7.88	>0.9999 (NS)	22.50	0.0085 (HS)
		Group IV	−29.05	<0.0001 (HS)	−23.75	0.0046 (HS)
	Group III	Group IV	−36.93	<0.0001 (HS)	−46.25	<0.0001 (HS)
Apical	Group I	Group II	−11.10	0.6200 (NS)	−19.13	0.041 (S)
		Group III	−6.500	>0.9999 (NS)	5.63	>0.999 (NS)
		Group IV	−37.90	<0.0001 (HS)	−44.50	<0.0001 (HS)
	Group II	Group III	4.600	>0.9999 (NS)	24.75	0.003 (HS)
		Group IV	−26.80	0.0005 (HS)	−25.38	0.002 (HS)
	Group III	Group IV	−31.40	<0.0001 (HS)	−50.13	<0.0001 (HS)

NS: Not statistically significant at P>0.05, S: Statistically significant at P≤0.05, HS: Highly statistically significant at P<0.01

DISCUSSION

Effective debris and smear layer elimination is essential for facilitating root canal disinfection.[14] Among various available irrigants, 5.25% NaOCl remains the gold standard owing to its ability to dissolve organic tissue and antimicrobial activity.[15] However, NaOCl alone cannot eliminate inorganic components; therefore, it is often used in combination with 17% EDTA, which effectively removes inorganic material within 1 min.[16]

Prolonged contact with NaOCl–EDTA can weaken dentin by decreasing its elastic modulus and flexural strength, along with potential cellular damage that may compromise long-term tooth prognosis.[6] These limitations underscore the importance of safer and more biocompatible alternatives, with herbal irrigants representing a promising option.

Setia et al. reported superior smear layer removal efficacy of neem extract compared with other tested herbal irrigants including propolis and orange oil, an effect attributed to its bioactive constituents such as flavonoids, alkaloids, glycosides, and tannins.[7] Based on these findings, neem leaf extract was selected for present study.

Previous studies have demonstrated that herbal extracts exhibit limited organic tissue dissolution, even with prolonged exposure.[17] Therefore, NaOCl remains necessary for effective tissue dissolution and antibacterial action. Bhavsar et al. suggested the combined use of NaOCl and neem leaf extract, to evaluate their complementary effects in removing debris and smear layer, which formed the basis for the irrigation protocol used in this study.[18]

The sequential use of multiple irrigants can result in interactions, making an intermediate rinse with a neutral solution necessary, thereby increasing treatment time and procedural complexities. Therefore, the development of a single irrigant capable of providing both organic tissue dissolution and chelating action, while being biocompatible, would serve as a convenient and efficient alternative to the conventional dual-solution protocol.[19]

Superoxidized water had been proposed as a single irrigating solution that can serve the aforementioned purpose. Thus, it can be a potential alternative for the conventional combination, NaOCl-EDTA.[6] This is also supported by Murugesan et al.[4] and Qing et al.[20] Moreover, it causes less dentin erosion and thus exhibits the combined advantages of NaOCl-EDTA without adversely affecting dentin microhardness.[20] Thus, superoxidised water was selected in present study.

The COXO root canal irrigation and suction system (CRCC) irrigator was employed in this study to enhance debridement within complex root canal anatomies. It provides machine-assisted irrigation with simultaneous delivery and suction of the irrigant, alongside adjustable parameters such as volume and working time. This approach enhances irrigant penetration to the working length and lateral canals, improving overall cleaning efficacy compared with conventional syringe irrigation. By facilitating continuous fluid exchange and more effective contact with canal walls, the CRCC system may further optimize the performance of various irrigants in endodontic procedures.[10]

It creates positive and negative pressure inside the canal with its two specially designed tips (tip A and tip B) to be used during and after biomechanical preparations, respectively. Each tip features a dual-needle configuration, with one needle for irrigant delivery and the other for waste fluid evacuation.[10]

Positive pressure irrigation effectively cleans and disinfects up to the middle third of the canal, primarily removing macro debris, whereas negative pressure irrigation reaches the apical third, efficiently eliminating micro and nano debris.[10]

SEM was employed for analysis owing to its high-resolution visualization of canal surfaces and dentinal tubules, enabling reliable assessment of debris and smear layer removal.[21,22]

Group I and Group III showed the highest cleaning efficacy with no significant difference between them, suggesting that superoxidised water can efficiently eliminate debris and smear layer similar to NaOCl-EDTA. This could be attributed to the combined action of hypochlorite, hydroxyl, and chloride ions present in the superoxidised water, which contributed to the tissue dissolution property, detergent action, and inorganic smear layer removal, respectively, and led to effective removal of both debris and smear layer.[1,23] These findings are consistent with earlier reports.[6,8]

Group II showed moderate root canal cleaning efficacy, which may be attributed to the comparatively lower chelating capacity of neem extract when compared with EDTA.[16,24] Limited literature exists comparing the effects of neem extract and superoxidised water.

Superior cleaning in the coronal and middle thirds may be attributed to wider canal dimensions facilitating irrigant penetration.[25] The apical third remains challenging because of its narrow anatomy and complex morphology, which restricts effective irrigant delivery.[26]

Limitations

Owing to its in vitro design, this study has inherent limitations and the results may not entirely represent clinical conditions where biological factors such as blood and tissue remnants can influence irrigant efficacy. In addition, curved canals present greater irrigation challenges. The use of single-rooted premolars with relatively wider canals allowed deeper penetration of irrigation tips; therefore, the results may not be directly applicable to posterior teeth with narrower and more complex canal anatomy, potentially reducing irrigant penetration and effectiveness.

CONCLUSION

Considering the limitations of this investigation, effective root canal cleaning was observed with all tested irrigants other than normal saline. Superoxidized water demonstrated similar efficacy to NaOCl–EDTA, suggesting its potential as safe, efficient, and convenient irrigating solution.

Furthermore, the use of C-RCC irrigator enhanced irrigant distribution and improved apical cleaning, thus emphasizing the use of irrigant delivery systems alongside irrigating solutions. The findings of this study align with the ongoing global trend toward safer, biocompatible, and simplified irrigation protocols in endodontics. Validation of these outcomes under clinical conditions requires additional in vivo research.

Conflicts of interest

There are no conflicts of interest.

Funding Statement

Nil.