|Year : 2023 | Volume
| Issue : 1 | Page : 76
Ex vivo evaluation of the efficacy of depotphoresis method in root canal disinfection
Saeed Moradi1, Siavash Moushekhian2, Reza Karazhyan3, Amir Ebrahimi4
1 Department of Endodontics, Dental Material Research Center, Faculty of Dentistry, Mashhad University of Medical Sciences, Mashhad, Iran
2 Department of Endodontics, School of Dentistry, Mashhad University of Medical Sciences, Mashhad, Iran
3 Department of Industrial Microbial Biotechnology, Research Institute for Industrial Biotechnology, ACECR, Mashhad, Iran
4 Department of Endodontics, School of Dentistry, Birjand University of Medical Sciences, Birjand, Iran
|Date of Submission||31-Jan-2022|
|Date of Acceptance||14-May-2023|
|Date of Web Publication||27-Jun-2023|
Dr. Amir Ebrahimi
Department of Endodontics, School of Dentistry, Birjand University of Medical Sciences, Birjand
Source of Support: None, Conflict of Interest: None
Background: Electrochemical disinfection of the root canal system (RCS) is introduced as an alternative to conventional irrigation. The aim of this study was to assess the efficacy of depotphoresis method in the disinfection of accessible and inaccessible RCSs.
Materials and Methods: In this comparative in vitro study disinfection of Enterococcus faecalis-infected RCS using two methods, (1) depotphoresis and (2) sodium hypochlorite (NaOCl) irrigation plus passive ultrasonic agitation (PUA) took place on 40 extracted maxillary anterior teeth. Decoronation was done with a diamond disc, and the canals were instrumented. The roots were divided into two phases: the specimens with canal obstruction and the specimens without canal obstruction. The smear layer was removed, and the specimens were infected for 21 days with E. faecalis. After disinfection procedures, bacterial samples were taken using two sterile #35 paper points, and colony-forming units (CFU) were counted. Data were analyzed statistically using the Kruskal–Wallis test, with a significance level at P < 0.05, to indicate differences between depotphoresis and NaOCl plus PUA groups.
Results: In both phases, Log CFU after depotphoresis treatment was significantly lower than NaOCl irrigation plus PUA treatment (P < 0.05).
Conclusion: Treatment with depotphoresis was significantly more effective than NaOCl irrigation plus PUA treatment.
Keywords: Disinfection, Enterococcus faecalis, iontophoresis, root canal irrigants, sodium hypochlorite
|How to cite this article:|
Moradi S, Moushekhian S, Karazhyan R, Ebrahimi A. Ex vivo evaluation of the efficacy of depotphoresis method in root canal disinfection. Dent Res J 2023;20:76
|How to cite this URL:|
Moradi S, Moushekhian S, Karazhyan R, Ebrahimi A. Ex vivo evaluation of the efficacy of depotphoresis method in root canal disinfection. Dent Res J [serial online] 2023 [cited 2023 Oct 1];20:76. Available from: https://www.drjjournal.net/text.asp?2023/20/1/76/379633
| Introduction|| |
The microbial causation of apical periodontitis has been well-documented., Dentinal tubules and apical irregularities of root canal anatomy such as deltas and isthmuses may harbor tenacious biofilms, contributing to the failure of endodontic treatment. Conventional mechanical instrumentation and irrigation with sodium hypochlorite (NaOCl) would not predictably make root canals free of bacteria.,
Calcium hydroxide (CH), as an interappointment medicament, reduces bacteria in dentinal tubules. However, a systematic review concluded that CH has limited antibacterial effectiveness, and the remnants of it can affect the sealing ability of root canal sealers.
Knappwost has introduced a technique named depotphoresis for the disinfection of root canal system (RCS) using CH with copper ions (cupral). Cupral is placed in the coronal third of the RCS, and the ions are activated and moved to the apical third of the RCS by electric current. It has been shown that depotphoresis eradicates viable bacteria in dentinal tubules to a depth of 500 μm., It has been claimed by the manufacturer of the depotphoresis system that “the electrical field transports cupral even in the mechanically not reachable apical part of the canal.”
The purpose of this study was to evaluate the efficacy of depotphoresis method in the disinfection of accessible and inaccessible RCS.
| Materials and Methods|| |
The methods of this in vitro study were approved by the Research Ethics Committee of Mashhad University of Medical Sciences (Approval ID: IR.MUMS.DENTISTRY.REC.1399.027).
Forty freshly extracted noncarious mature maxillary anterior teeth were collected and kept in saline. After placing the teeth in 5.25% NaOCl for 30 min to detach residual debris from the root surfaces, a diamond disc was used to cut the crowns perpendicular to the long axis of the teeth to have the same length of roots (16 mm). A #10 K-type file was placed into each root canal to confirm apical patency.
Then, the roots were divided into two phases: the specimens without canal obstruction and the specimens with canal obstruction.
Phase 1: The specimens without canal obstruction
The root canals were instrumented to F3 (ProTaper Universal; Dentsply Sirona, Ballaigues, Switzerland) 0.5 mm beyond the apical foramen. Irrigation with 5.25% NaOCl was performed between each file insertion. To remove the smear layer, a final irrigation sequence of 5 ml of 17% EDTA for 5 min, 5 ml of 5.25% NaOCl for 5 min, and 10 ml of normal saline was done according to Lin et al.'s study. The roots were placed in Eppendorf tubes containing 2 ml of Tryptic Soy Broth (TSB) medium and then autoclaved for 30 min at a pressure of 15 PSI and temperature of 121°C. Then, the tubes were incubated at 37°C for 24 h.
A sterile micropipette was used to aspirate TSB and deliver 1 ml of Enterococcus faecalis suspension with a standard concentration of 0.5 McFarland (1.5 × 108 colony-forming units [CFU]/ml) into the root canal and around the root so as to immerse the root in the E. faecalis suspension. Thereafter, the tubes were incubated at 37°C for 24 h, and bacterial viability was determined by CFU count. Subsequently, the specimens were incubated at 37°C for 21 days. To prohibit dehydration of the specimens during incubation, the TSB medium was refreshed every 3 days.
The roots were randomly divided into NaOCl plus passive ultrasonic agitation (PUA) group (n = 10) and depotphoresis disinfection group (n = 10). As shown in [Figure 1], each root was fixed in a 6-cm high round pot which was half-filled with 0.9% sterile saline and had two holes in its lid, one to place the root, and the other to place the lip clip of Depotphorese®-system Original II.
|Figure 1: The experimental model (a) lip clip; (b) electrode; (c) lid; (d) tooth; (e) normal saline.|
Click here to view
In the depotphoresis group, root canals were rinsed with 10 ml of sterile saline. According to the manufacturer's instructions of Depotphorese®-system Original II, one part of the cupral was mixed with nine parts of calcium hydroxide-dispersed, and 2 mm3 of the resultant mixture was placed in the coronal third of the root canal by means of a Heidemann Spatula and a plugger. The depotphoresis electrode was inserted in the coronal third of the root canal to the full length of the electrode (5 mm), and the lip clip was inserted in the pot. The electric current was established and gradually increased to 1.5 mA/min. Three times with intervals of 5 min, and each time, 1.5 mA of electric current was established. Overall, 4.5 mA of electric current was transmitted. A final rinse with 10 ml sterile saline was performed.
In the NaOCl plus PUA group, irrigation with 20 ml of 5.25% NaOCl was done in 10 min, and the ultrasonic golden tip of Ultra X (Eighteeth, Changzhou, China) was used at low power within 1 mm of the apical foramen, and agitation of the irrigant was performed for 60 s. A final rinse with 10 ml sterile saline was done.
Each canal was then instrumented for 10 s with a sterile #30 K-file to obtain dentin shavings and sampled with two sterile #35 paper points which remained in the root canal for 1 min and then were moved to Eppendorf tubes containing 1 ml of TSB medium.
After vortexing the tubes for 10 s, 0.1 ml of the 1:10 dilutions was transferred to brain-heart infusion agar plates and incubated at 37°C for 48 h, and CFU per ml was counted.
Phase 2: The specimens with canal obstruction
The root canals were instrumented to F1 (ProTaper Universal; Dentsply Sirona, Ballaigues, Switzerland) to the apical foramen. Irrigation with 5.25% NaOCl was performed between each file insertion. A weak point was created on F3 rotary file (ProTaper Universal; Dentsply Sirona, Ballaigues, Switzerland) in the area between D3 and D4 by a round high-speed bur. The weakened file was rotated in the root canal at high torque (6 N/cm) to separate the file in the canal. A radiograph was taken to confirm that the separated file is located in the middle third of the canal. Next, the apical foramen was enlarged using Gates-Glidden bur #2 to a depth of 2 mm through the retrograde direction. To remove the smear layer, a final irrigation sequence was performed similarly to the group without obstruction. The roots were placed in Eppendorf tubes containing 2 ml of TSB medium and then autoclaved for 30 min at a pressure of 15 PSI and temperature of 121°C. Then, the tubes were incubated at 37°C for 24 h. A sterile micropipette was used to aspirate TSB and deliver 1 ml of E. faecalis suspension with a standard concentration of 0.5 McFarland (1.5 × 108 CFU/ml) into the root canal and around the root so as to immerse the root in the E. faecalis suspension. After that, the tubes were incubated at 37°C for 24 h, and bacterial viability was determined by CFU count. Subsequently, the specimens were incubated at 37°C for 21 days. To prohibit dehydration of the specimens during incubation, the TSB medium was replenished every 3 days.
The roots were randomly divided into NaOCl plus PUA group (n = 10) and depotphoresis disinfection group (n = 10).
The apical third of the specimens was sealed with wax, and also a hole was made in it so as to allow the electric current to flow. Root canals assigned to the depotphoresis group were treated exactly the same as the roots without obstruction which was described previously. In NaOCl plus PUA group, root canals were treated similar to the roots without obstruction except that the ultrasonic tip was placed in the coronal third of the root canal. Thereafter, the wax covering apical foramen was removed, and the canal was instrumented for 10 s with a sterile #30 K-file from a retrograde direction to obtain dentin shavings. Sampling was performed by two sterile #35 paper points which were described previously. Other procedures were the same as the group without canal obstruction, and CFU/ml was counted.
The CFU mean of the two samples of each specimen was calculated, and a log transformation of the CFU count was done to normalize the data before statistical analysis. Data were analyzed statistically using the Kruskal–Wallis test, with a significance level at P < 0.05, to indicate differences between depotphoresis and NaOCl plus PUA groups.
| Results|| |
One of 10 depotphoresis specimens and four of 10 NaOCl plus PUA specimens showed no bacterial growth in any of their two samples. The means and standard deviations of each group are reported in [Table 1]. Using the Kruskal–Wallis test, the Log CFU of the depotphoresis group was significantly lower than NaOCl plus PUA group (P = 0.006).
|Table 1: Mean, standard deviation, lowest, highest, and median values for log colony-forming units of the specimens without canal obstruction|
Click here to view
The bacterial growth of two of the 10 specimens in the NaOCl plus PUA group was negative. The means and standard deviations of each group are reported in [Table 2]. Using the Kruskal–Wallis test, the Log CFU of the depotphoresis group was significantly lower than the NaOCl plus PUA group (P = 0.009).
|Table 2: Mean, standard deviation, lowest, highest, and median values for log colony-forming units of the specimens with canal obstruction|
Click here to view
| Discussion|| |
In either phase of the current study, depotphoresis treatment had a significantly greater reduction in Log CFU of E. faecalis than NaOCl plus PUA treatment.
Owing to the high prevalence of E. faecalis in persistent endodontic infections,,, its single-species biofilm was used in this study. The difficulty of E. faecalis biofilm eradication is supported by the studies that have shown its ability to invade dentinal tubules and form biofilms in medicated root canals. Benefits of monospecies biofilm model systems are simplicity, standardization, and control, although they are more susceptible to antimicrobial treatment than multispecies biofilms.,,,
According to similar studies,, standardization of the specimens was done by shaping the root canals 0.5 beyond the apical foramen. Removal of the smear layer was performed to get the maximum penetration of E. faecalis biofilm. Because there were no similar studies for the second phase of the present study, it seems, the best possible method that could simulate the clinical situation was adopted to allow sampling from the apical portion of the obstructed canal.
A low current electric field of about 5 mA was used to mobilize ions derived from cupral paste according to Fuss et al.'s study, which eliminated all bacteria to a depth of 500 μm in dentinal tubules. Lin et al. applied a current of 10 mA that was more effective than pure calcium hydroxide in depths of 200–500 μm. One study used a current of 15 mA that was the most effective against Gram-positive cocci. The manufacturer's instructions of Depotphorese®-systems Original II recommended 5 mA in each session of treatment.
It has been demonstrated that the maturity of the biofilm increases its resistance to NaOCl., Because E. faecalis biofilm gets mature in 14–21 days, a 3-week incubation was considered in the majority of studies.
Zou et al. noted that NaOCl penetration depth in the dentinal tubules was 77–300 μm. Haapasalo and Orstavik reported that after 3 weeks, E. faecalis caused dense infection of dentinal tubules to a depth of 400 μm, and the front of infection could reach 800–1000 μm. Furthermore, bacteria can reside in uninstrumented recesses of the main canal. 6% NaOCl eradicated 99.99% of E. faecalis biofilm bacteria in vitro; however, its anti-biofilm efficacy was lower in vivo., In the present study, 5.25% NaOCl plus PUA could not make all the root canals free of bacteria which could be related to NaOCl penetration depth, inaccessible areas of the canal, and decreased NaOCl anti-bacterial efficacy in the root canal.
Laser activation of irrigation has been shown to increase NaOCl penetration in dentinal tubules, and NaOCl irrigation plus photon-induced photoacoustic streaming was more effective than NaOCl irrigation alone in the eradication of E. faecalis biofilm. However, iontophoresis using Cu was more effective than laser therapy against both Gram-positive and Gram-negative bacteria.
It has been claimed by the manufacturer that inaccessible areas of the root canal can be disinfected by depotphoresis. Owing to fractured file conductivity, an electric current can be established in the apical portion of the canal, and the presence of the copper, calcium, and hydroxide ions can lead to disinfection. The quantity of the copper ion can be determined by spectrophotometry, which can be conducted in future studies.
Intracanal sampling technique was one of the limitations of this study as passive sampling may not detect the bacteria that reside in inaccessible areas of the root canal which can cause false-negative results. Future in vitro and in vivo studies can be conducted using newer methods of spectrophotometry like microspectroscopy, and confocal laser scanning microscopy can be used in future in vitro studies.
| Conclusion|| |
Treatment with depotphoresis was significantly more effective than NaOCl irrigation plus PUA treatment in either teeth with canal obstruction or teeth without canal obstruction.
This research has been supported by the Vice Chancellor for Research of Mashhad University of Medical Sciences, Mashhad, Iran. The authors deny any conflicts of interest related to this research.
Financial support and sponsorship
This study was supported by Mashhad University of Medical Sciences.
Conflicts of interest
The authors of this manuscript declare that they have no conflicts of interest, real or perceived, financial or non-financial in this article.
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[Table 1], [Table 2]