Scanning Electron Microscopic and Dye Penetration Evaluation of Hand Instrumentation Techniques on Formation of Smear Layer in Root Canal Preparation: An In vitro Study.

AIMS AND
OBJECTIVES: The present study was undertaken for evaluating various hand instrumentation techniques with different instruments in the formation of smear layer.
MATERIALS AND METHODS: One hundred and seventy-five extracted mandibular molars were collected and were cut at cementoenamel junction. Afterward, the distal roots were separated. Working length was determined, and roots were divided into five equal groups with 35 roots in each group as follows: Group A: K-type files with conventional step-back technique, Group B: Canal Master U instrument with clockwise-counterclockwise rotation, Group C: Flex-R® files with clockwise-counterclockwise rotation, Group D: FlexoFiles® with conventional step-back technique, and Group E: Nickel titanium files with conventional step-back technique. Biomechanical preparation was done according to their respective groups. The teeth in various groups were scanned under scanning electron microscope and scoring was done. All the results were analyzed by SPSS software version 17.0.
RESULTS: Scanning electron microscopic results indicated that apparently there seems to be a significant difference in the scoring of the smear layer between each group at all the three levels, but the difference was statistically nonsignificant. Microleakage was present in all the samples. Nonsignificant results were obtained while comparing the microleakage in between different the groups. However, microleakage was highest in Group D.
CONCLUSION: The amount of smear layer is maximum at the apical third and lesser in the middle third followed by the coronal third in all the groups. All the samples showed microleakage to a varying extent. Copyright:

INTRODUCTION

Treatment in dentistry is aimed not only at the eradication of the disease and alleviation of pain but also toward the restoration of the oral system to a healthy state. This is possible only if an attempt is made to minimize the probable etiologic factor which could result in treatment failure.[123] Unknown and unrecognized for years, the smear layer has become a force to be reckoned with during the last decade. The smear layer is thought to modify dentin permeability, reduce the sealer adhesion, and alter root canal obturation so much that it might serve as an index of an instrument's capacity to instrument and to mill the dentin surface.[45]

The cleanliness of the root canal wall after preparation has been investigated by various authors using hand instrumentation or automated devices. All of them have found that completely clean root canal wall could not be achieved without any of the different techniques and devices investigated.[678] Hence, under the light of the above-mentioned data, the present study was undertaken for evaluating various hand instrumentation techniques with differ instruments in the formation of smear layer. The aim was to further establish whether the amount of smear layer had any role to play in the amount of microleakage associated with each type of instruments after canals were obturated with gutta percha.

MATERIALS AND METHODS

The present study was conducted with the aim of assessing hand instrument techniques on the formation of smear layer in root canal preparation by scanning electron microscope and dye penetration. One hundred and seventy-five extracted mandibular molars were collected and were cut at cementoenamel junction. Afterward, the distal roots were separated. Only those teeth were selected which had a straight distal canal. If the apical foramen was open to over # 20 K-file, the tooth was rejected. The teeth were cleaned and stored in normal saline for 1 week.

Working length determination

Contents of the root canal were removed by conventional method. The length of the distal canal was measured with # 15 K-file, with the stopper placed in contact with a coronal landmark and tip of the file flushed with the anatomic apex of the root. These roots were divided into five equal groups with 35 roots in each and they were grouped according to the type of instrument and technique used for the preparation of canals, as shown in Table 1.

Table 1

Division of the roots according to study groups

Group	Instrument type
Group A	K-type files with conventional step-back technique
Group B	Canal Master U instrument with clockwise-counterclockwise rotation
Group C	Flex-R files with clockwise-counterclockwise rotation
Group D	FlexoFiles with conventional step-back technique
Group E	NiTi files with conventional step-back technique

Biomechanical preparation

Group A

K-type file with conventional step-back technique was used in the preparation of root canals. Canals were instrumental at the apex up to #35. From #35 onward, each file was shortened by 1 mm in progressive sequence up to #50 during the biomechanical preparation. 5.25% solution of sodium hypochlorite was used as an intracanal irrigant, and the tooth was held in wet gauze so as to prevent drying of the root. After preparation, the roots were stored in distilled water.

Group B

Manual Canal Master U instrument with Canal Master instrumentation technique was used to prepare the root canals. Every manual Canal Master is designed to work in clockwise rotation of 120°. Apical preparation was done till #35. From 35 onward, each Canal Master instrument was shortened by 1 mm in sequential order till size 50. Intracanal irrigant was used in the manner as in Group A.

Group C

Flex-R® file with balanced force technique was used to prepare the root canals. Instrumentation was done by placement cutting and removal of each file using only rotary motions. Placement was accomplished using clockwise rotation of 180° and light apical pressure. Cutting was accomplished using counterclockwise rotation 120° or more, and apical pressure was adjusted to match file's strength. Cleaning or debris removal was accomplished using one to two noncutting no-pressure or slight outward-pull clockwise rotations so as to engage all debris in the fluids. Cleaning was accomplished only after the desire working length had been reached and maintained with counterclockwise rotation.

Group D

The canals were prepared with FlexoFiles® in the conventional step-back technique as was done in Group A.

Group E

Nickel titanium (NiTi) files with conventional step-back technique were used as in Group A.

The teeth in various groups were scanned under scanning electron microscope and scoring was done. Photomicrographs of representative samples were taken as reference standard for the scoring.

Scoring:

0 – No smear layer with all tubules opened

1 – Little smear layer with >50% tubules opened

2 – Moderate smear layer, with <50% tubules opened

3 – Heavy smear layer with outline of tubules obliterated.

All the results were analyzed using SPSS software version 16.0 (IBM, Armonk, New York). Chi-square test and analysis of variance for one-way classified data were used for evaluation of level of significance.

RESULTS

In order to study the association between the five groups and the gradings made at apical level, Chi-square test of association among attributes was performed and coefficient of contingency was computed [Figure 1]. It was found that the computed value of Chi-square was 6.52 at 8 degrees of freedom, whereas the critical value was 15.51 at 5% and 20.09 at 1%. Since, the computed value did not exceed the critical value, the difference between the groups was insignificant [Tables 2 and 3]. The results between all the groups were insignificant [Table 4]. At the apical level, all groups showed only scores 2 and 3, though the percentage of Grade 2 was maximum in Group C, that is, 60% and minimum in Group D, that is, 20% and percentage of score 3 was minimum in Group E, that is, 30% and maximum in Group A, that is, 70% [Table 5]. The mean value of microleakage was the highest (3.493) in Group D, whereas on the other extreme, the minimum value was 2.932 in Group E. The 95% confidence interval for means of rest of the group can be interpreted similarly [Table 6]. Value of mean squares between groups turned out to be 1.298, whereas that within groups was 1.334; consequently, the value of variance ratio as computed from the available data turned out to be only 0.973, which was considerably less than the critical value of 2.46 (5% probability level) of variance ratio at the corresponding number of degrees of freedom [Table 7].

Figure 1

(a) Score 0 – smear layer at the coronal third, (b) score 1 – smear layer at the middle third, (c) score 2 – smear layer at the apical third, (d) score 3 – smear layer at the apical third

Table 2

Statistical evaluation of scanning electron microscope study of smear layer at apical level

Group	Grade				Total
	0	1	2	3
A	0	0	3	7	10
B	0	0	4	6	10
C	0	0	6	4	10
D	0	0	2	8	10
E	0	0	7	3	10
Total	0	0	22	28	50
χ 2	6.98 (nonsignificant)
Degree of freedom	4
Coefficient of contingency	0.3500

Table 3

Statistical evaluation of scanning electron microscopic study of smear layer at middle level

Group	Grade				Total
	0	1	2	3
A	1	1	8	0	10
B	1	1	8	0	10
C	2	2	6	0	10
D	0	4	6	0	10
E	2	3	5	0	10
Total	6	11	33	0	50
χ 2	6.52 (nonsignificant)
Degree of freedom	8
Coefficient of contingency	0.3395

Table 4

Statistical evaluation of scanning electron microscopic study of smear layer at coronal level

Group	Grade				Total
	0	1	2	3
A	2	6	2	0	10
B	3	4	3	0	10
C	4	5	1	0	10
D	4	5	1	0	10
E	4	4	2	0	10
Total	17	24	9	0	50
χ 2	3.08 (nonsignificant)
Degree of freedom	8
Coefficient of contingency	0.2409

Table 5

Percentages of scoring at all the three levels in all the five groups

Group	Apical (%)				Middle (%)				Coronal (%)
	0	1	2	3	0	1	2	3	0	1	2	3
A	0	0	30	70	0	80	10	10	20	60	20	0
B	0	0	40	60	0	80	10	10	30	40	30	0
C	0	0	60	40	0	60	20	20	10	50	40	0
D	0	0	20	80	0	60	40	0	40	50	10	0
E	0	0	70	30	0	50	30	20	40	40	20	0

Table 6

The mean value of apical microleakage along with standard deviation and coefficient of variation of all the five groups

Group	Number of observations	Mean	SD	CV	95% CI for mean
A	25	3.388	1.033	30.5	2.953-3.823
B	24	3.301	0.946	28.7	2.893-3.709
C	25	3.076	1.329	43.2	2.515-3.635
D	25	3.493	1.230	35.2	2.975-4.012
E	24	2.932	1.063	36.3	2.473-3.391

SD: Standard deviation, CV: Coefficient of variation, CI: Confidence interval

Table 7

Analysis of variance for one-way classified data

Source of variation	Degree of freedom	Sum of squares	Mean squares	Variance ratio (F)
				Computed	Critical
					5%	1%
Between groups	4	5.1918	1.2979	0.973	2.46	3.49
Within groups	118	157.4041	1.3339	-	-	-

DISCUSSION

FlexoFiles introduced by Maillefer in 1981 are twisted instruments that have a triangular cross-section. The working end has 29 spirals. The angle between the cutting flutes and long axis is 30° at the tip region and 45° at the working end. When used in a rotary reaming motion, the FlexoFiles displays the highest cutting efficiency of all instruments.[89101112] Nevertheless, it has been observed that enlargement of severely curved canals may lead to undesirable changes in canal shape.[13141516]

More flexible systems were developed with the passage of time. NiTi, known as NiTi files, were introduced for endodontic use.[151617]

In the present study, the root canals were prepared with different instrumentation techniques and their effectiveness was evaluated by scoring of smear layer in the coronal, middle, and apical region, ×1000 magnification. The scoring of photomicrographs of all the five groups at the three different levels is given in Table 1. These results were put to statistical analysis, and it was found that there was no statistically significant difference between all the groups [Tables 2–4]. These findings are in concurrence with those of Prati et al.,[18] who also got similar results when they used K-files, Ergoflex, Canal Master, and Flex-R instruments. Reddy et al., in another study, observed that Manual NiTi files produced significantly less smear layer and debris compared to rotary ProTaper instruments.[19] Similarly, apparent difference was observed in the scoring between K-type files and Flex-R files, which is concurrence with Wu and Wesselink[20] who revealed significant difference in cleanliness between Flex-R and K-files in terms of debris. They also revealed that the amount of debris remaining in the apical portion is more than that in the middle and coronal portion regardless of the technique performed.

In our study, we obturated the teeth using lateral condensation with zinc oxide eugenol as a sealer. Lateral condensation was chosen because of insignificant volumetric changes after obturation, to produce apical seal and a three-dimensional filling, and due to long history of use. Zinc oxide eugenol sealer was used as it has adequate flow characteristic ability to chelate with gutta percha and has good tissue tolerance, antimicrobial property, and nonstaining characteristic.[12131421]

Our study revealed microleakage in all samples, but Karagöz-Küçükay and Bayirli[22] in their study found 78.9% samples with no leakage and 21.1% samples with leakage. The difference in the results may be due to the use of Calcibiotic root canal sealer sealer and thermoplasticized gutta percha used in their study.

The result of our study are in concurrence with the results of a study conducted by Economides et al.[23] who assessed the microleakage after obturating the teeth with gutta percha and Roth 811, a zinc oxide eugenol-based sealer, in the presence of smear layer. The efficacy may depend completely on the operator and how meticulously he/she is using each instrument. In another study conducted by Zand et al., the authors compared the efficacy of NiTi Flex K-file instruments and rotary FlexMaster and RaCe instruments in root canal preparation and concluded that NiTi Flex K-files resulted in significantly more smear layer (P < 0.05) compared to RaCe and FlexMaster instruments only in the apical third of the canal.[24]

CONCLUSION

In the present study, smear layer is produced by all the hand instrumentation techniques after irrigation with 5.23% sodium hypochlorite. In addition, the amount of smear layer is maximum at the apical third and lesser in the middle third followed by the coronal third in all the groups. All the samples showed microleakage to a varying extent.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.