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Basic ResearchTechnology

Effect of Ultrasonic Activation of Irrigants

on Smear Layer Removal
Tamer F. Schmidt, DDS, MSc,* Cleonice S. Teixeira, DDS, MSc, PhD,*
Mara C.S. Felippe, DDS, MSc, PhD,* Wilson T. Felippe, DDS, MSc, PhD,*
David H. Pashley, DMD, PhD, and Eduardo A. Bortoluzzi, DDS, MSc, PhD*

Introduction: The objective of this study was to eval-
uate the efficacy of passive ultrasonic irrigation (PUI)
with 17% EDTA and 1% NaOCl solutions on smear
T he removal of smear layer produced after root canal instrumentation has been rec-
ommended because its presence can have deleterious effects on the endodontic
treatment. The presence of bacteria and their by-products and necrotic residue in end-
layer removal. Methods: Root canal preparations of odontic smear layers compromise the disinfection process (1). In addition, smear layer
32 human teeth were performed with the ProTaper sys- decreases dentin permeability, interfering with diffusion of antimicrobial agents from
tem. Next, they were longitudinally fractured to permit irrigants and intracanal medications into root dentin (2, 3). Smear layers also block
quantitation of smear layer creation from the cervical, tubular entry of endodontic sealers and act as a barrier between obturation
middle, and apical thirds of the roots by using scanning materials and canal walls, compromising root canal sealing and increasing chances
electron microscopy. After reassembling the fractured of reinfection (4).
tooth halves, they were divided into 4 groups according The alternating use of EDTA and sodium hypochlorite (NaOCl) solutions is used to
to different final irrigation protocols: group1, EDTA + remove the inorganic and organic portions of the smear layer, respectively (5). To be
NaOCl; group 2, EDTA with PUI + NaOCl; group 3, effective, the solutions must come into contact with root canal walls (6). Irrigation
EDTA + NaOCl with PUI; and group 4, EDTA + NaOCl, methods that use syringe and needle have been shown to be incapable of reaching diffi-
both with PUI. After irrigation, the tooth halves were cult access areas such as apical and isthmus regions. Thus, the activation of irrigating
separated to permit imaging the same areas by scanning solutions applied by various methods has been proposed to enhance their action and
electron microscopy, and a percentage of opened penetration (7, 8). Studies have shown that ultrasonic activation of irrigating solutions
dentinal tubules in irrigated areas as a percent of the to- as passive ultrasonic irrigation (PUI) promotes better removal of the smear layer in the
tal area was obtained. The results were submitted to apical region and isthmus regions (911).
Kruskal-Wallis, analysis of variance, and Bonferroni However, despite the publication of many articles on smear layer removal, there is
tests (a = 0.05). Results: The cervical third of the sam- no well-established protocol for PUI (11). In addition, the scoring of dentin only after
ples from all groups showed higher percentage of smear final irrigation and qualitative analysis of smear layer removal by scores have been re-
layer removal and open dentinal tubule areas, followed ported to be technical failures (1214).
by the middle and apical thirds. Among the irrigation Because of these technical problems, it is necessary to establish final irrigation
groups, there were statistically significant differences protocols for PUI. For this reason, this study evaluated longitudinally and quantitatively
in cervical third between group 2 and group 4 samples, the efficacy of PUI on 17% EDTA and 1% NaOCl solutions to remove the smear layer.
with the highest and lowest percentage of smear layer
removal, respectively. Conclusions: PUI by using 1% Materials and Methods
NaOCl and ultrasonic tip placed within 1 mm of the The protocol used in the present study was approved by the Human Research
apical foramen did not show higher efficacy in smear Ethics Committee, Federal University of Santa Catarina. Thirty-two human premolars
layer removal compared with conventional irrigation. with single straight or slightly curved canals and fully formed roots were collected. Ra-
(J Endod 2015;41:13591363) diographs were taken to confirm straight single canal and the canal space size. Those
teeth were extracted from young adult patients between 13 and 17 years old for ortho-
Key Words dontic reasons. After accessing the root canal, the tooth length was obtained by intro-
Irrigation, passive ultrasonic irrigation, SEM, smear layer ducing #10 K-file (Dentsply Maillefer, Ballaigues, Switzerland) into the canal to the
point of displaying its tip at the apical foramen. The working length (WL) was obtained
by subtracting 1 mm from the tooth length.
The apical region of each root was covered with a layer of heavy condensation
silicone impression material (Zetaplus; Zhermack, Polesine Badia, Italy) to avoid

From the *Endodontic Division, Department of Dentistry, Federal University of Santa Catarina, Florianopolis, Santa Catarina, Brazil; and Department of Oral Biology,
College of Dental Medicine, Georgia Regents University, Augusta, Georgia.
Address requests for reprints to Dr Eduardo A. Bortoluzzi, Department of Dentistry, Center for Health Sciences, Campus Reitor Jo~ao David Ferreira Lima, Florianopolis,
Santa Catarina, Brazil 88040-900. E-mail address: eduardo.bortoluzzi@gmail.com
0099-2399/$ - see front matter
Copyright 2015 American Association of Endodontists.

JOE Volume 41, Number 8, August 2015 Smear Layer Ultrasonic Removal Evaluation 1359
Basic ResearchTechnology
extravasation of irrigating solutions and simulate the clinical condition taining anhydrous silica for the same period to remove any moisture.
of the presence of periapical tissues (15). Without any coating or additional preparation, the samples were
Root canal preparations were performed by the same operator evaluated by SEM operated at low vacuum (JCM-6390LV; JEOL, Pea-
with rotary instruments by using nickel-titanium ProTaper Universal body, MA).
files (Dentsply Maillefer) up to an F4 file. At each file change, the canals After locating the cross-shape markings on the canals, the most
were irrigated with 2 mL 1% NaOCl by using a 5-mL syringe and NaviTip well-defined area completely covered by smear layer in each of the
30-gauge tip (Ultradent Products Inc, South Jordan, UT) calibrated to thirds was chosen at 100 (Fig. 1B). Another image was obtained at
stop 2 mm from the WL, with back-and-forth movements of 23 mm. 500, with its edges coinciding to the limits of the marks (Fig. 1C).
Simultaneously, suction was accomplished by using a metal cannula. Then a third image was obtained at 1000 without changing the posi-
Apical patency was maintained at each change of instrument by inserting tion of the sample in SEM (Fig. 1E). In total, 9 images were obtained per
#10 K-file up to the apical foramen. At the end of the procedure, canals sample before irrigation, 3 images for each third. These initial images
were irrigated with 3 mL distilled water and dried with absorbent paper were used to evaluate the condition of the root canal walls before the
points. final irrigation.
A gutta-percha cone was introduced into the canal, and longitudi- Next, the halves of each tooth were placed back together, and the
nal grooves were made on the buccal and lingual external surfaces of grooves previously created for cleavage were filled with resin (Topdam;
each tooth by using diamond double-sided disks with a diameter of FGM, Joinville, Brazil) to stabilize the parts. The reassembled tooth root
22 mm and a thickness of 0.1 mm (ref. 7020; KG Sorensen, Cotia, was inserted into heavy condensation silicone impression material to
Brazil) operated at low speed until the presence of the pink gutta- increase stability and prevent leakage of the solutions used in the final
percha cone was seen, thereby avoiding accidental contamination irrigation protocols.
and invasion of the canal by cutting debris. A small cotton pellet was Thirty-two teeth were randomly divided into 4 groups (n = 8)
placed in the access opening to prevent entry of cutting debris. according to the final irrigation protocol used (Table 1).
After the teeth were cleaved with the aid of a chisel, 1 of the 2 halves All canals were irrigated with the same techniques used during the
was selected for preirrigation evaluation by scanning electron micro- chemical-mechanical preparation. PUI was performed with a specific
scopy (SEM). Three external markings were made on this half with a tip without cutting power, with apical diameter #20, taper .01 (Irrisonic
fine-tip pen on the external root surface, perpendicular to the long E1; Helse, Santa Rosa de Viterbo, Brazil) calibrated to 1 mm short of WL,
axis, to divide it into cervical, middle, and apical thirds of the same activated by ultrasound (JetSonic; Gnatus, Ribeir~ao Preto, Brazil) at a
length. The markings served as references to make 3 grooves in the ca- power of 20% indicated by the manufacturer, avoiding contact with
nal wall, delimiting the root into thirds (cervical, middle, and apical). the walls of the root canal. The canals of all groups received 3 mL
The grooves were created by using double-sided diamond micro-disks, EDTA for 3 minutes and 3 mL NaOCl for 3 minutes.
7 mm in diameter and 0.1 mm in thickness, deep enough to be satisfac- At the end of the procedure, the canals were irrigated with 3 mL
torily viewed by SEM. A no. 11 scalpel blade was used to create a new distilled water to remove possible salt residues from the irrigation so-
mark approximately 5 mm in length on the axial grooves. Thus, an im- lutions. Then they were dried with paper points.
age similar to a cross could be visualized on the root canal wall of each After the experiment, the teeth were separated into their 2 halves
of the thirds (Fig. 1A). and were dried, coated with gold, and analyzed by conventional SEM
The fractured tooth samples were kept in an incubator at 37 C for (high vacuum). New images were obtained from the same preselected
48 hours. Then the samples were placed in a vacuum desiccator con- and pre-photographed areas, following the methodology described

Figure 1. (A) Marks to determinate the evaluation area. Black arrows: grooves perpendicular to long axis of root canal made with diamond disk. White arrows:
marks made with scalpel blade in direction of long axis of the canal. (B) Cross at the center of the canal wall. LL, lower left area; LR, lower right area; UL, upper left
area; UR, upper right area. (C) Image obtained from apical third of the root, after root canal preparation showing the smear layer formed (original magnification,
500). Arrows indicate marks made on root canal wall. (D) Image obtained after final irrigation. Note marks (arrows) that enabled to re-evaluate the same area of
the image (original magnification, 500). (E) Image of smear layer taken at 1000. (F) The 1000 image showing effect of final irrigation. (G) Image processing
by Image J software (cervical third). Dark areas correspond to open dentinal tubules. (H) Identification of open dentin tubules (in red) by Image J software.

1360 Schmidt et al. JOE Volume 41, Number 8, August 2015

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TABLE 1. Experimental Groups and Irrigation Protocols
EDTA 17% NaOCl
Groups v t tPUI v t v t tPUI v t
1: Conventional 3 3 3 3
2: PUI EDTA 1.5 1 1 1.5 1 3 3
3: PUI NaOCl 3 3 1.5 1 1 1.5 1
4: PUI EDTA + PUI NaOCl 1.5 1 1 1.5 1 1.5 1 1 1.5 1
t, solution action time (min); tPUI, PUI action time (min); v, volume solution (mL).

above (Fig. 1D and F). The images taken at 1000 were evaluated by Furthermore, the methodology applied in most studies of smear layer
using Image J version 1.47 to enable identification and expression of the removal has been questioned (14). This SEM study was designed to
percentage of area of open dentinal tubules in relation to the total area quantitatively and longitudinally evaluate the efficacy of PUI on removing
of the analyzed image (Fig. 1G and H). the smear layer from cervical, middle, and apical thirds of instrumented
root canals.
Statistical Analysis To replicate clinical conditions, a closed apical system was created
The Kolmogorov-Smirnov test was used to analyze the normality of to simulate clinical conditions because the presence of periapical tis-
the continuous variables, and the Levene test was used to analyze the sues and possible entrapment of air in the apical region can hinder
homogeneity of variances among the groups. The combinations of vari- the penetration of solutions in this region (15).
ables group and canal thirds were statistically shown to have a To ensure that enough space was allowed for adequate flow of irri-
normal distribution (P > .05). Because of the significant differences gating solutions to the WL, the canal was prepared up to ProTaper F4,
among the variances of the groups, a nonparametric analysis of variance with an apical diameter #40 (16).
test (Kruskal-Wallis) was used (P = .05). The analysis of variance was One percent NaOCl was chosen in this study because when it is
then used for multiple comparisons, with Bonferroni correction, to associated with the use of EDTA, it has been shown to be effective in
isolate the differences, which reduced the P value to .005. the removal of smear layers (5, 9, 17). Moreover, the 1% NaOCl
solution showed the same antibacterial effect of higher
concentrations (18, 19). Its use can minimize the adverse effects of
Results accidental extrusion (20) and deleterious changes in the structural
The average  standard deviations of the percentages of open properties of the dentin (21).
dentinal tubule areas in relation to the total area of the image are sum- Techniques that promote activation of irrigating solutions have
marized in Table 2. In all groups, the highest percentage of opened been used to allow better dispersion and penetration, enhancing its ef-
dentinal tubule areas was found in the cervical third, followed by the fects (7, 8). Among those techniques, PUI has been the subject of
middle and apical thirds. several studies (711). The term PUI (passive ultrasonic irrigation)
When the thirds were compared, the results of different groups is considered inappropriate by some researchers because it is
were similar (P > .05) except for the cervical third (P < .001), where impossible to prevent the ultrasonically activated instrument from
a greater percentage of open tubule areas was observed in samples of touching the canal walls. The term is used to differentiate ultrasonic
group 2 compared with group 4 (P = .02). irrigation procedures from ultrasonic root canal preparation that is
When the average percentages of open tubule areas in all thirds of aimed at cutting or shaping dentin walls (11). In this study, a
all groups were compared, a significant difference was observed only smooth-edge tip with no cutting power was used, with an apical diam-
among groups 2 and 4 (P = .018). eter corresponding to #20 instruments, to avoid undesirable morpho-
logic changes in already prepared canal walls (22).
Discussion In studies that used PUI with EDTA followed by NaOCl to remove
Although PUI use has been reported to optimize smear layer the smear layer, there is no consensus on which solution should be ul-
removal (9, 10), there is no set protocol for quantity of irrigating trasonically activated. Some investigators used EDTA (17, 23), only
solution, working time, or choice of solution to be activated. NaOCl (24), or both (9, 25). Furthermore, there is no comparison

TABLE 2. Average (A) and Standard Deviations (SD) of Open Dentinal Tubule Areas in Relation to Total Area of the Image, According to the Groups and Canal
Cervical Medium Apical Total
Groups A SD A SD A SD A SD
A,B a A
1: Conventional 21.14 3.40 12.51 6.16 3.00 1.75 12.22 8.56a,b
2: PUI EDTA 25.22* 3.23A 15.70 8.48a 5.26 4.17A 15.39* 9.98a
3: PUI NaOCl 20.12 6.21A,B 13.65 6.75a 5.76 2.86A 13.17 8.01a,b
4: PUI EDTA + PUI NaOCl 18.60* 2.81B 9.06 7.33a 5.02 2.19A 10.90* 7.35b
Total 21.27 4.66A,B 12.73 7.29a 4.76 2.94A 12.92 8.56a,b

A, average; SD, standard deviation.

Groups identified by different letters in each vertical column are significantly different (P < .05).
*Values with statistically significant difference (P < .05).

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in the same study between the effects of PUI in each solution separately According to the results of the current study, it can be concluded
or together, as was done on this study. that by using 1% NaOCl and ultrasonic tip placed within 1 mm of the
Likewise, activation times of PUI, which vary from 20 seconds apical foramen, PUI did not show higher efficacy in smear layer removal
to 5 minutes, can be found in the literature (2628). Activation compared with conventional irrigation.
time in this study was 1 minute because it is sufficient to remove
the smear layer from the apical region, and it has been used in Acknowledgments
other studies (9, 17, 23).
This research was conducted with support from the Central
The majority of studies on the removal of smear layer are per-
Laboratory of Electron Microscopy from Federal University of Santa
formed by using conventional SEM, which means that it requires high
Catarina (LCME-UFSC).
vacuum and metalized specimen surfaces to allow visualization of the
The authors deny any conflicts of interest related to this study.
area to be evaluated. This type of analysis allows only acquisition of
post-treatment images in a single moment in the study, which are ac-
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