Cleaning ability of reciprocation and rotation motions in conjunction with either continuous or sequential chelation: An SEM study

Abstract

Context:

Recent advancements have introduced a debate over the most effective combination of file kinematics and irrigation timing. While continuous rotation and reciprocation motions each offer distinct advantages in debris extrusion and file safety, their synergistic effect with different chelation protocols remains a critical area of investigation.

Aims:

The aim was to evaluate the cleaning performance of reciprocation and continuous motions in combination with continuous versus sequential chelation irrigation protocols.

Subjects and Methods:

Sixty single-rooted single-canalled teeth were prepared and divided into two groups according to the type of file used (n = 30). Group A: one curve (OC) file system (rotation motion) and Group B: one RECI (OR) file system (reciprocation motion). Prepared teeth were then subdivided into three subgroups according to the irrigation protocol used (n = 10): Subgroup 1: saline, Subgroup 2: sequential chelation (sodium hypochlorite [NaOCl] + ethylenediaminetetraacetic acid), and Subgroup 3: continuous chelation (NaOCl/etidronic acid). Samples were observed using an environmental scanning electron microscope to measure the percentage of opened dentinal tubules.

Statistical Analysis Used:

Data were analyzed using one-way analysis of variance followed by Tukey’s post hoc test for pairwise comparisons between the different groups and thirds.

Results:

In all the groups, the coronal third had significantly cleaner dentinal tubules (P < 0.001). The difference between the two files was not statistically significant (P = 0.643). Sequential chelation showed the highest mean values of the opened dentinal tubules in conjunction with OC and OR (7.89 ± 2.95 and 8.95 ± 5.14, respectively). With continuous chelation, the mean values of the opened dentinal tubules in conjunction with OR were significantly higher than with OC (4.22 ± 3.21 and 2.39 ± 1.19, respectively).

Conclusions:

The cleaning abilities of the OC and OR file systems are comparable. Sequential chelation resulted in cleaner and more open dentinal tubules regardless of the file system used. Continuous chelation with reciprocation yielded cleaner canals than with rotation motion.

INTRODUCTION

The presence of bacteria and their byproducts is widely recognized as the major factor in the formation of pulp and periapical lesions. Therefore, a primary objective of endodontic treatment is the elimination of microorganisms from the root canal. This is achieved through effective chemomechanical preparation, which involves cleaning and shaping the root canal.[1] Irrigants play a crucial role during root canal procedures. They act as lubricants, help remove necrotic pulp tissue and debris, and assist in eliminating microorganisms and their byproducts.[2] Sodium hypochlorite (NaOCl) is the most used irrigant for root canal disinfection due to its several benefits. It has an antibacterial effect and can effectively destroy bacterial biofilm. In addition, it has proteolytic action with the ability to dissolve pulp tissue remnants.[1,2] However, the inorganic fraction of the smear layer and the hard-tissue debris produced during instrumentation cannot be effectively removed by NaOCl alone. Therefore, it is recommended to use ethylenediaminetetraacetic acid (EDTA) and NaOCl alternately. However, combining EDTA with NaOCl eliminates the active chlorine in NaOCl, and excessive use of EDTA leads to dentine erosion. Hence, it is advisable to use EDTA as a final irrigant before obturation.[3] Etidronic acid, also known as 1-hydroxyethylidene-1,1-biphosphonate (HEBP), is compatible with NaOCl and exhibits mild chelating properties. Thus, the concept of “continuous chelation” was proposed, which involves using NaOCl in combination with etidronic acid. Mixing HEBP with NaOCl allows continuous chelation, offering a simpler and more time-efficient irrigation protocol than alternating between separate solutions.[4]

A variety of instruments and techniques have been developed to shape root canals effectively while minimizing the risk of iatrogenic errors. Nickel–titanium (NiTi) instruments, with high taper, cyclic fatigue resistance, and flexibility, can help achieve meeting these biomechanical objectives.[5] One Curve (OC) is a single-file system that is used in continuous rotation. One RECI (OR) is a single-use reciprocating file for root canal shaping.[6]

To our knowledge, none of the previous studies investigated the effect of the OR on the cleaning ability of root canals. Thus, the aim of this study was to evaluate the cleaning ability of two heat-treated controlled memory NiTi single-file systems using reciprocation or rotation motions while irrigating with HEBP and NaOCl simultaneously versus NaOCl irrigating solutions using an environmental scanning electron microscope (ESEM).

The null hypothesis was that there would be no significant difference between using HEBP and NaOCl simultaneously versus NaOCl separately as root canal irrigant regarding the smear layer removal after instrumentation with either the OC or OR file systems.

SUBJECTS AND METHODS

Sample size calculation

Sample size determination was conducted using G*Power version 3.1.9.2 (Universität Kiel, Kiel, Germany). Based on data from a previous study, a two-sided power analysis was performed to detect differences between the groups.[7] Using an effect size of f = 0.56, an alpha level of 0.05, and a beta level of 0.05 (power = 95%), the required sample size was calculated to be 60 specimens for the two main instrumentation groups. Furthermore, the two groups were subdivided equally into three subgroups (10/subgroup) according to the chelation protocol. This research was conducted in accordance with the Preferred Reporting Items for Laboratory Studies in Endodontology (PRILE) guidelines, and the study design is illustrated in the associated PRILE flowchart[8] [Figure 1].

Figure 1.

The study design according to the Preferred Reporting Items for Laboratory Studies in Endodontology guidelines for ethical approval

Sample selection

The study protocol was approved by the Ethical Committee of the Faculty of Dentistry, Suez Canal University, Egypt (approval no. 650/2023). All procedures followed relevant guidelines and regulations. Sixty single-rooted mandibular premolars with a single canal were obtained from the university’s department of oral surgery. The teeth had been extracted for periodontal or orthodontic reasons. Permission was obtained from the department to collect the teeth. Since the teeth were collected retrospectively and without any direct involvement with patients, informed consent was not required. The teeth were cleaned using an ultrasonic scaler and examined under a surgical operating microscope (OMS 2350, Zumax, China) to exclude fractures, caries, calcifications, resorptions, and cracks. Preoperative buccolingual and mesiodistal radiographs confirmed the presence of a single canal. Inclusion criteria required straight roots with Vertucci type I canals, complete root formation, and the absence of cracks, calcifications, extensive caries, or internal or external resorption. Teeth with a root curvature between 0° and 10°, as measured by Schneider’s method, were included in the study.[9] Teeth with open apices, root caries, or a history of root canal treatment were excluded. All specimens were stored at room temperature in fresh physiologic saline, which was replaced daily.[10]

Before decoronation, canal patency was confirmed by introducing a size #10 K-file until its tip appeared at the apical foramen under a surgical operating microscope. Working length was established by subtracting 1 mm from this measurement. Each tooth was then decoronated at the cementoenamel junction using an IsoMet saw under continuous water cooling, and the root length was standardized to 12 mm.

Randomization and blind allocation

One of the co-authors, who was not involved in the steps of instrumentation, performed the blind allocation of the samples. This allocation was completed by Microsoft Excel randomization, which was completed by placing papers in an opaque envelope to avoid any bias.

Root canal instrumentation

The roots were randomly divided into two equal groups (n = 30) according to the type of file used.

The groups were as follows:

Group A: OC file system (Micromega, Besancon, France) (rotation motion)

Group B: OR file system (Micromega, Besancon, France) (reciprocation motion).

In Groups A and B, the glide path was done using size #10 K-file. Root canals were then instrumented using the One Flare file (Micromega, Besancon, France) for coronal flaring at a speed of 250–400 rpm and a maximum torque of 3 N.cm with the electrical endomotor (Eighteeth, Changzhou, China). Later, manual instrumentation was performed using file sizes #10, #15, and #20, which were utilized to create a glide path. Irrigation, according to the subgroup, was performed using a 30G side-vented needle (Fanta Co., Shenzhen, China) for 20 s between the two consecutive files. The final apical preparation was done using a single-file system of OC with a tip size of #35 (.04) using rotary motion with a speed of 300–450 rpm and a maximum torque of 2.5 N.cm with the electrical endomotor. The final apical preparation was done using a single-file system of OR with a tip size of #35 (.04) using reciprocating motion, with a reciprocation angle of 170° counterclockwise and 60° clockwise with an electrical endomotor in a crown-down manner according to the manufacturer’s recommendations. During root canal preparation, K-file size #10 was used to check the apical patency among each instrumentation. After using each file, the canals were irrigated according to the subgroup with a 30G side-vented needle for 20 s. The needle was positioned into the canal until a small resistance was felt. The same process was repeated for all the Groups; Following instrumentation, both groups were subdivided into three subgroups, differentiated by their irrigation protocol (Figure 1). Subgroup 1 received 10 ml of saline. Subgroup 2 received sequential chelation, consisting of 1 ml of 2.5% NaOCl irrigation between every two files, followed by a final 1-minute rinse with 5 ml of 17% EDTA. Saline was employed as an intermediate rinse between the applications of NaOCl and EDTA, and a final rinse was performed with 2 ml of saline. In Subgroup 3, irrigation with continuous chelation using 10 ml of 9% HEBP (Aquapharm, Maharashtra, India) and 2.5 NaOCl was used simultaneously throughout the preparation, and the final rinse was performed with 2 ml of saline.

Sample preparation

The teeth were longitudinally grooved on the buccal and lingual surfaces using a diamond disc, allowing minimal penetration into the canal space to avoid creating artificial debris. Each root was then split into two halves with a chisel, and the half displaying the clearest view of the coronal, middle, and apical thirds was selected for evaluation under the ESEM (FEI Quanta 3D 200i, Oregon, USA).[11]

Environmental scanning electron microscope evaluation

The samples were mounted on standard-diameter aluminum stubs using carbon double-sided adhesive tape. Imaging was performed with an ESEM (FEI Quanta 3D 200i) at 30 kV accelerating voltage, 1 nm resolution, and ×2000. Representative images were selected for analysis. A calibrated, blinded examiner evaluated the root canal walls. ImageJ software (NIH, Bethesda, MD, USA) was used to quantify the open dentinal tubule area. The total image area (μm²) was measured automatically, and the percentage of opened dentinal tubules was calculated using the formula:

Open dentinal tubules % = (Total area of opened dentinal tubules [μm²])/(Total image area [μm²]) × 100.[11]

Statistical analysis

Statistical analysis was performed using SPSS (version 25,IBM Corp., Armonk, NY: USA). Data normality was verified using the Shapiro–Wilk test, confirming a parametric distribution. Group and canal third comparisons were conducted using one-way analysis of variance with Tukey’s post hoc test for pairwise comparisons. P ≤ 0.05 was considered statistically significant, and P ≤ 0.001 was considered highly significant.

RESULTS

For both OC and OR systems, there were highly significant differences for the irrigants, where EDTA groups had the cleanest dentinal tubules, followed by HEBP groups (P < 0.001). Regarding thirds, for all the subgroups, the coronal represented the highest, cleanest dentinal tubules, followed by the middle, and the apical third was the least in all the subgroups. The saline groups failed to show any cleaned, opened dentinal tubules [Table 1 and Figure 2].

Table 1.

Mean±standard deviation and comparison of the percentage of opened dentinal tubules between the three thirds (coronal, middle, and apical) and different irrigants for the two studied files

Group	Third irrigant	Coronal	Middle	Apical	Overall (%)	P α
One curve	Saline	0±0C,a	0±0C,a	0±0B,a	0±0C	-
	Sequential chelation (NaOCl+EDTA)	10.83±4.72A,a	9.26±5.07A,a	3.57±2.9A,b	7.89±2.95A	<0.001 (HS)
	Continuous chelation (NaOCl + HEBP)	4.01±2.81B,a	2.22±1.29B,b	0.93±0.78B,b	2.39±1.19B	<0.001 (HS)
P β		<0.001 (HS)	<0.001 (HS)	<0.001 (HS)	<0.001 (HS)
One RECI	Saline	0±0C,a	0±0C,a	0±0B,a	0±0C	-
	Sequential chelation (NaOCl + EDTA)	13.92±6.88A,a	8.67±5.44A,b	4.26±4.22A,c	8.95±5.14A	<0.001 (HS)
	Continuous chelation (NaOCl + HEBP)	6.98±5.6B,a	4.54±3.57B,b	1.13±0.74B,c	4.22±3.21B	<0.001 (HS)
P β		<0.001 (HS)	<0.001 (HS)	<0.001 (HS)	<0.001 (HS)

^αOverall P value for comparisons among canal thirds (ANOVA), ^βOverall P value for comparisons among irrigants (ANOVA). Lowercase superscripts indicate differences among the thirds within the same group; uppercase superscripts indicate differences among irrigants within the same group (Tukey’s post hoc test). Means with different superscripts differ significantly at P≤0.05. HS: Highly significant (P≤0.001), S: Significant (P≤0.05), NS: Not significant (P>0.05), EDTA: Ethylenediaminetetraacetic acid, NaOCl: Sodium hypochlorite, HEBP: 1-hydroxyethylidene-1,1-bisphosphonate

Figure 2.

Representative environmental scanning electron microscope images showing dentinal tubules in (A) coronal, (B) middle, and (C) apical thirds for all main groups and subgroups

Comparing both the groups (OC and OR), no significant difference was found when EDTA was used (P = 0.543). On the other hand, when HEBP was used, OR showed a significant difference in the cleaned open dentinal tubules (P = 0.019). In addition, OR showed increased cleanliness of the opened dentinal tubules compared to OC in all the subgroups in all thirds except for the middle third when EDTA was used, where OC showed insignificant cleanliness of the dentinal tubules (P = 0.715). When using HEBP, the One RECI system provided significantly better dentinal tubule cleanliness than One Curve in the coronal, middle, and apical-middle thirds, with no significant difference observed in the apical third. Overall, a comparison between One RECI (OR) and One Curve (OC) across all irrigants showed no significant difference in cleaning effectiveness (P = 0.643) [Table 2].

Table 2.

Mean±standard deviation and comparison of the percentage of opened dentinal tubules between the two studied rotary system files at the three thirds under different irrigants

Irrigant	Group third	One curve	One RECI	P*
Saline	Coronal	0±0	0±0	-
	Middle	0±0	0±0	-
	Apical	0±0	0±0	-
	Overall (%)	0±0	0±0	-
Sequential chelation	Coronal	10.83±4.72	13.92±6.88	0.096 (NS)
(NaOCl + EDTA)	Middle	9.26±5.07	8.67±5.44	0.715 (NS)
	Apical	3.57±2.9	4.26±4.22	0.415 (NS)
	Overall (%)	7.89±2.95	8.95±5.14	0.543 (NS)
Continuous chelation	Coronal	4.01±2.81	6.98±5.6	0.036 (Significant)
(NaOCl + HEBP)	Middle	2.22±1.29	4.54±3.57	0.008 (Significant)
	Apical	0.93±0.78	1.13±0.74	0.390 (NS)
	Overall (%)	2.39±1.19	4.22±3.21	0.019 (Significant)
Overall (%)		3.42±3.80	4.39±5.02	0.643 (NS)

Overall, the P value reflects the comparison between the two rotary file systems (Independent t-test). *denotes statistical significance, HS: Highly significant (P≤0.001), S: Statistically significant (P≤0.05), NS: Not significant (P>0.05), EDTA: Ethylenediaminetetraacetic acid, NaOCl: Sodium hypochlorite, HEBP: 1-hydroxyethylidene-1,1-bisphosphonate

DISCUSSION

Irrigation is a critical component of root canal disinfection, as it eliminates microorganisms, dissolves organic and inorganic debris, removes accumulated canal residues, and clears the smear layer produced during mechanical instrumentation.[12,13]

Therefore, this study assessed the cleaning ability of two heat-treated controlled memory NiTi single-file systems using either reciprocation or rotation motions while irrigating with HEBP and NaOCl simultaneously versus NaOCl irrigating solution and a final rinse of 17% EDTA using ESEM.

The most popular irrigant, NaOCl, was selected due to its broad antimicrobial action and its alkaline pH, which allows it to dissolve organic tissue. NaOCl comes in a range of concentrations from 0.5% to 6%. Higher concentrations of NaOCl cause dentin erosion, collagen degradation, and a decrease in dentin microhardness, elastic modulus, and flexural strength.[14] As 2.5% NaOCl is less cytotoxic than 5.25% NaOCl, it was chosen in the current study. Furthermore, it has been demonstrated that the antibacterial efficiency of 2.5% and 5.25% NaOCl concentrations is equivalent.[15]

The success of treatment depends on the irrigation needle depth. Thus, the irrigation was done in the present study using a 30G side-vented needle positioned 2 mm from the working length. Straight, single-rooted human mandibular premolars, Vertucci’s type I root canal classification with oval canals and mature apices was used in this study to avoid any anatomical variation and ensure standardization. Teeth were prepared up to size 35, and the root canals were standardized to be straight or to have a curvature of <20. This ensures the needle depth and penetration of irrigants to the apical one-third in comparison to curved roots. It was shown that EDTA decreased both the antibacterial and tissue-dissolving properties of NaOCl and that frequent use of EDTA causes erosion of the root canal dentin, decalcification of the peritubular dentin, and dissolution of the exposed organic matrix. In addition, when EDTA was combined with NaOCl, it eliminated active chlorine. That is why the decalcifying HEBP was chosen to preserve NaOCl’s potent oxidizing activity and chlorine content.[16] In addition to the importance of the chemical action of irrigation, shaping the root canals is of paramount importance. It prevents and treats periapical diseases and produces an easily irrigated funnel that removes germs, toxins, and organic waste from the root canal. However, to prevent iatrogenic harm to the root canal system, this must be done cautiously with NiTi instruments that are flexible, exhibit cyclic fatigue resistance, and have a high taper.[17] OC was chosen in this study as it is a heat-treated file, which allows controlled memory and prebending of the wire to shape root canals. Thus, it preserves the root canal anatomy. It also has great cutting efficiency and centering ability, as it features a distinct cross section all the way around the blade.[18]

In our study, OR single-file system was employed in a reciprocating motion. It was chosen because it has a unique variable asymmetric cross section and C-wire heat treatment technology, which provides it with exceptional flexibility, cutting efficiency, and controlled memory that permits the file to prebend. Furthermore, its reciprocating action is safe, comfortable, and resistant to cyclic fatigue. Using this minimally invasive file in a reciprocating motion respects root canal anatomy and protects pericervical dentin. Both the OC and the OR that were used in our study had the same size (35) and taper (04).

In this study, the image analysis for ESEM images was processed using ImageJ software. A noteworthy methodological advantage in this study was the use of Environmental SEM (ESEM), which allowed for the observation of the root canal wall in its natural, hydrated state. Unlike conventional SEM, which requires aggressive dehydration and sputter coating, ESEM maintains a gaseous environment that prevents the shrinkage of the collagen matrix. This was critical for accurately assessing the cleaning ability of the tested filing systems and irrigation protocols as it ensured that the patency of the dentinal tubules was not an artifact of sample preparation, but rather a true reflection of the treatment’s efficacy.The null hypothesis was partly rejected, as there was a significant difference between the groups and subgroups regarding thirds. The highest mean percentage of opened dentinal tubules was in the coronal third, followed by the middle third, and the lowest was in the apical third in both reciprocating and rotating motion file systems.

The results of the current study were in accordance with many previous studies. These results might be attributed to the reduced dentin permeability and the sclerotic nature of the apical region. Furthermore, the formation of vapor lock obstructs the flow and effectiveness of irrigating solutions. In addition, it should be noted that the number and surface area of dentinal tubules decrease as one moves apically. Irrigants might have difficulty reaching the apical third due to its smaller diameter compared to the coronal and middle thirds.[19]

While in all the subgroups irrigated with saline, the mean percentage of open dentinal tubules in the saline was 0%. This result demonstrated the ineffectiveness of saline to remove the smear layer and open the dentinal tubules. Several previous studies have been conducted to evaluate the smear layer removal using saline as the control group. These results might be because of the poor efficacy of saline in dissolving both organic and inorganic debris. The present study is consistent with many past studies.[20]

When using OC and OR in all thirds, the highest mean percentage of opened dentinal tubules was with EDTA, followed by HEBP, and the lowest was with saline.

The results of the present study were consistent with several previous studies, as they found that EDTA was more effective in smear layer removal and proved to be more potent than HEBP. This might be due to the higher pH of HEBP compared to EDTA, as well as the fact that EDTA forms a more stable complex than HEBP; thus, it dissolves and binds to more calcium than HEBP. The opened tubular regions and the dissolved calcium had a strong connection. Added to that, sequential chelation resulted in increased proteolytic activity, which helps break down proteins in the smear layer and dissolve organic components, such as tissue remnants, by interacting with their protein structure. Furthermore, Dual Rinse HEDP increased the surface tension of NaOCl, which may limit the solution’s ability to penetrate the root canal isthmuses, fins, and dentinal tubules. This makes it less efficient than the NaOCl and EDTA in smear layer removal.[21,22]

Conversely, SEM images from several previous studies showed that the continuous chelation protocol appeared to be as effective as, or more effective than, the traditional sequential chelation protocol. As it maintains the cleanliness of root canals, removes the smear layer and hard-tissue debris, and reduces the torsional load on rotary files, This might be due to the higher concentration of HEBP, as they used 18%, which enhances HEBP’s ability to remove the smear layer by eliminating phosphate, which exposes the collagen matrix and increases the amide III/phosphate ratio. Nevertheless, 18% HEBP caused moderate erosion at the coronal level of radicular dentin. This compromises the integrity of the tooth structure. Unlike the 9% HEBP, which did not cause any noticeable erosion at any level, it is a more favorable option for preserving dentin.[1,17,20,21,23]

According to our results for both chelating systems, EDTA and HEBP, in all thirds, OR produced the highest mean percentage of opened dentinal tubules with HEBP. While OC achieved the lowest mean percentage of opened dentinal tubules.

Our results were in accordance with several previous studies. This might be attributed to the fact that the steady rotating motion has minimal impact on the inner curvature of the canal, while it enlarges the canal exterior, especially in the apical third. As a result, the reciprocating motion leads to higher cleaning efficiency. Added to that, the reciprocating motion offers several benefits for root canal cleaning, such as reduced torsional and flexural stresses, better canal centering, and a reduction in taper lock within the root canal. Notably, the design of the reciprocating instrument working part, which along with its cutting efficiency, enhances the agitation of the irrigation fluid. Furthermore, the cross-sectional design of the instruments, specifically the positive rake angles of reciprocating files, allows the blade to cut dentin chips that curl away from the edge, whereas negative rake angles cause the blade to scratch the dentin surface. Besides, HEBP facilitates smear layer removal by eliminating phosphate, a process that exposes the collagen matrix and increases the amide III/phosphate ratio, and reduces the torsional load on rotary files.[21]

As an overall comparison, the difference between the two files was not statistically significant. Our study agreed with a previous one where they found that particularly in the middle and coronal thirds of the canal, there were no statistically significant differences in smear layer removal between both the file systems.[24]

Our findings differed from those of a previous study,[25] which reported that reciprocating instruments were less effective than continuous motion instruments in cleaning the root canal walls, removing the smear layer from dentin, and completely opening the dentinal tubules. This could be the result of the use of a continuous motion full-sequence file system instead of a single reciprocating motion file system. The continuous motion files facilitate the upward removal of debris along the file flutes. In contrast, the backward motion of the reciprocating files compacts the debris against the dentinal walls, pushing it into lateral canals and beyond the apex. This prevents the blade from cutting into the dentin as effectively.[26]

One of the limitations of the current study was the small sample size, which might not provide sufficient statistical power to detect significant differences between the groups. That is why it is recommended to increase the sample size in further studies. In addition, extra studies comparing sequential and continuous chelation using activation methods are needed to get more clinical implications. Similarly, more research is required to compare different kinematics, single- versus full-sequence file systems.

CONCLUSIONS

None of the tested files nor chelation techniques produced totally clean and open dentinal tubules. The coronal one-third showed the cleanest opened dentinal tubules. Changing kinematics did not affect the cleanliness of the root canal. Both continuous and sequential chelation with reciprocation resulted in clean dentinal tubules. Continuous chelation with reciprocation yielded cleaner canals than with rotation motion.

Conflicts of interest

There are no conflicts of interest.

Funding Statement

Nil.