2022, Number 1
LIBS analysis of the cleaning performed by a Nd:Yag laser on used Nickel Titanium instruments
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ABSTRACT
Introduction: used endodontic instruments retain dentin debris which represents a biological risk that affects their sterilization. Objective: to assess the cleaning of NiTi endodontic instruments treated with an Nd:Yag laser using the laser-induced plasma spectroscopy (LIBS) technique. Materials and methods: ProTaper Next X2 instruments (Dentsply Maillefer®, Ballaigues, Switzerland) and WaveOne Gold Primary (Dentsply Maillefer®, Ballaigues, Switzerland) were evaluated under a stereoscopic microscope (20X) in 2 stages: 1) used instruments, 2) instruments cleaned with ultrasonics or Nd:Yag laser. In both stages, the presence and extent of dentin debris stained with Van Gieson solution were recorded. Subsequently, the elements contained in the dentin debris found in used NiTi instruments and treated with different cleaning methods were analyzed with the LIBS technique. Results: the Nd:Yag laser technique presented more cleaning capacity (p < 0.0001). The LIBS analysis did not show spectra of elements contained in the biological debris in the instruments treated with both methods. Conclusions: the Nd:Yag laser is an effective method for cleaning NiTi instruments by removing the elements contained in the dentin debris from its surface.INTRODUCTION
Cleaning the root canal system is one of the crucial steps for endodontic shaping and obturation.1 Different types of nickel-titanium (NiTi) rotary instruments are used to fulfill this operative purpose.2-4 During instrumentation of the root canal system, a large amount of dentin debris is retained and adheres to the active surface of these instruments;5,6 it contains a high amount of microorganisms and residues of vital or necrotic pulp tissue.5 Dentin debris is a biological material that can be transported to other patients if the instruments are not properly sterilized.7-9 Often, NiTi instruments used for the biomechanical preparation of root canals are reused in other patients. Therefore, the removal of dentin debris retained in the active part of NiTi instruments represents a significant challenge during the cleaning, disinfection, and sterilization of these instruments. In this regard, Popovic J et al.5 reported the presence of 96% of residual biological debris in used endodontic instruments subjected to different cleaning protocols.
Endodontic instrument cleaning methods are varied and include mainly procedures such as pre-wetting in enzymatic solution, cleaning of the active part of the instruments with a sponge, instruments subjected to cleaning employing ultrasonic tanks,10-12 and sonochemical reactors that intensify the cavitation effect in ultrasonic tanks.9 However, these instrument cleaning methods are not able to eliminate prion proteins that are theoretically capable of transmitting spongiform encephalopathy-type diseases because these proteins are resistant to the high temperatures generated during sterilization procedures.13,14 Recently, the Nd:Yag laser was introduced as a method to remove dentin debris caused by root canal instrumentation.15 It has also been reported that the Nd:Yag laser is capable of removing dental calculus.16 The Er:Yag laser has also proven to be a useful technique for the removal of dentin debris from instrumented root canals.17
Based on these studies, it is assumed that it is possible to achieve thorough cleaning of NiTi instruments by applying Nd:Yag laser to remove the dentin debris adhered to the active surface of these instruments. The presence of biological debris on endodontic instruments can be recorded using Laser Induced Breakdown Spectroscopy (LIBS). This technique can determine the constituent elements of a sample in a fast, non-invasive, and real-time manner that may be useful in the dental field.18,19 Therefore, the objective of the study was to evaluate through the LIBS technique the surface cleanliness of NiTi endodontic instruments treated with Nd:Yag laser.
MATERIALS AND METHODS
New rotary instruments ProTaper Next X2 (Dentsply Maillefer®, Ballaigues, Switzerland) and WaveOne Gold Primary (Dentsply Maillefer®, Ballaigues, Switzerland) were used for the in vivo biomechanical preparation of 6 mesial roots of lower first molars with irreversible pulpitis. The instruments were assessed under stereo microscopy in 2 stages: stage 1) used in clinical conditions and 2) subjected to ultrasonic or Nd:Yag laser cleaning. The dirty instruments were immersed for 3 minutes in a Van Gieson solution (Sigma-Aldrich, St. Louis, MO, USA) to stain the collagen contained in the debris (red and orange color). Once the instruments were stained, they were randomly assigned to different cleaning methods: Group A: Three ProTaper Next X2 instruments that received ultrasound cleaning. Group B: Six ProTaper Next X2 instruments that received Nd:Yag laser cleaning. Group C: Three WaveOne Gold Primary instruments that received ultrasound cleaning. Group D: Six WaveOne Gold Primary instruments that received Nd:Yag laser cleaning. The four facets of each instrument were observed under stereo microscopy (Leica EZ4D, Singapore) at 20X in sections of 5 mm per face.
On each instrument, the basal dentin debris record was obtained using the method established by Linsuwanont et al.20 The category for classifying dentin debris was: SD (stained debris) red or orange pigmented particles on the instrument surface, F (organic film) a thin unstructured layer on some part of the instrument surface and generally stained red, UD (unstained debris) fine particles not showing red or orange staining and C (clean surface). The evaluation of the extent of Stained Debris was recorded as 0 (none), 1 (film only), 2 (light) particles scattered widely over the instrument groove, 3 (moderate) numerous particles with areas of continuous coverage on the surface, 4 (severe) areas of the instruments in which the grooves are covered by debris to their full depth. Two endodontic specialists previously calibrated in the evaluation of dentin debris measurement performed the records (Kappa=0.86, good concordance strength). Once the basal records were obtained, the instruments immediately received the cleaning protocols indicated above.
Ultrasound cleaning. The dirty instruments were inserted 10 times in a sponge soaked in 0.12% chlorhexidine. Subsequently, they were placed in a container containing enzymatic solution (Zymex-Sultan Healthcare) for 30 minutes and afterward, in an ultrasonic tank (Biosonic, Coltene UC50D) for 15 min. Finally, they were rinsed under running water and left to dry at room temperature.
Nd:Yag laser cleaning. The NiTi instruments were fixed in the sample holders in a vertical position. Forty shots were manually performed with Nd:Yag laser (Bralax Laser Labs, S de RL) 250 mJ, 3Hz, at a distance of 6 cm along the active part of each face of the instrument for 2 min.
Once the cleaning protocols were completed, the instruments were again subjected to a microscopic recording of dentin debris in the same way as that performed for the measurement of the dirty instruments.
LIBS analysis. Using the LIBS technique, the elements calcium (Ca), magnesium (Mg), sodium (Na), fluorine (F), and phosphorus (P) were identified in a debris sample at different wavelengths under the obtained spectral lines. The LIBS technique identified the elements contained on the surface of new instruments (control), used and treated with ultrasound or Nd: Yag laser. An Nd: YAG laser equipped with a passive Cr: YAG Q: Switch was used as the excitation source. The laser emits in the wavelength of 1064 nm laser shots, each containing trains of up to 4 pulses with maximum energy of 80 mJ. The laser light was focused on the sample with a lens at a distance of 10 cm. The laser caused the formation of a plasma introduced in an optical fiber of 200 μm diameter thus conducting the radiation to an OceanOptics spectrometer (model HR2000), with a spectral range from 200 to 1100 nm, a spectral resolution of 1.5 nm and integration time of 3.8 ms. Spectra Suite software was used for the detection and capture of spectra. The elements of interest were visually identified by obtaining a normalized spectrum. The Merge Graph application was used to individually analyze specific regions of the spectral characterization of both biological debris and NiTi instruments. For the analysis of the identified elements, the database published by the National Institute of Standards and Technology (NIST)21 was used as a reference.
The Mann Whitney Wilcoxon U test was used to compare the stained debris and debris extension between both cleaning methods. An alpha value of 0.05 was used in the IBM SPSS Statistics® 23.0 program.
RESULTS
The instruments used under clinical conditions presented stained dentin debris on more than 90% of their active surface (Table 1). In both brands of instruments, a greater amount of stained debris was observed in the Nd:Yag laser group (p < 0.05) (Figure 1). The results of the cleaning achieved by each method are shown in Table 2 and the extent of debris in Table 3. Under microscopic observation, the Nd:Yag laser technique completely removed the dentin debris from the surface in both instrument brands (p < 0.0001). Ultrasonic cleaning left a higher percentage of organic film on both types of instruments (p < 0.05). The extension of debris in the used WaveOne Gold Primary instruments assigned to both cleaning techniques did not show significant differences (p = 0.58) (Table 4). The Protaper Next X2 instruments treated with ultrasound presented less extension of debris (p < 0.001) while those treated with laser did not present any debris film. Ultrasonic cleaning identified instruments with organic film. The Nd: Yag laser cleaning technique presented a greater cleaning capacity of the NiTi instruments (p < 0.0001).
Similar spectra were observed in new NiTi instruments treated with ultrasound and Nd-YAG laser. In none of the analyzed spectra Ca, Mg and Na elements characteristic of the presence of dentin debris were found on the surface of new ProTaper Next X2 and WaveOne Gold Primary instruments cleaned with ultrasound or Nd: Yag laser (Figures 2 and 3).
DISCUSSION
In endodontic practice, endodontic instruments are generally reused, being of utmost importance that they are completely clean before sterilization. Smith et al.22 reported that disinfection methods for treating contaminated endodontic instruments under clinical conditions fail to completely remove the biological material adhered to these instruments. Our results indicate that used NiTi instruments retain dentin debris with potential biological risk according to the staining shown by the Van Gieson solution.20 Endodontic instruments containing contaminated biological debris can act as a vehicle for the transmission of various diseases such as that acquired through the Creutzfeldt-Jakob prion protein, a type of spongiform encephalopathy that has no treatment and can be fatal.23,24
The present study found that Nd:Yag laser irradiation is capable of removing biological debris from the surfaces of rotary NiTi instruments used under clinical conditions. We found that the Nd:Yag laser completely removed the debris adhering to the surface of endodontic instruments like the reported efficacy of the Nd: Yag laser in the removal of dental calculus.16 The way in which the removal of debris from the surface of endodontic instruments is achieved is similar to that observed in the removal of dental calculus. When the laser hits a surface, a thermal degradation process is established on this surface, where the energy density applied to the sample, in this case, the surface of a NiTi instrument, raises the temperature of this surface causing a micro explosion and vaporization that detaches the material adhered to the instrument. The laser has particular effects on an irradiated sample and depends strongly on the absorption of the sample at the irradiated wavelength. If the absorption of laser light by the organic tissue is very strong, the energy is deposited near the surface, and vaporization is confined to a surface layer of the instrument.
We also found that the use of the LIBS technique as a method for testing dentin debris clearance is a technique that allows the elemental composition to be analyzed by laser ablation of material layers.19,25 It was observed that the WaveOne Gold Primary instruments lost their characteristic golden color. This may be due to thermal changes suffered by the alloy derived from the heat generated on the surface and the ablation process. In the microscopic evaluation, it was observed that the ultrasound did not achieve a complete cleaning of the instruments. Popovic et al.5 reported that manual cleaning, the use of a detergent, and ultrasound waves are factors that favor the cleaning of endodontic instruments. Although these authors did not report a complete cleaning of the instruments treated with ultrasound, our results coincide in that the use of ultrasound does not completely remove the organic material under microscopic inspection. However, in our study, the LIBS technique was not able to find elements contained in the debris of ultrasound-treated instruments. This situation can be explained since obtaining samples for analysis by LIBS technique is microscopic, generating plasma from a crater (50 μm approximately) on the examined surface that may not be representative of the condition present in the entire sample. Linsuwanont et al.20 concluded that the total removal of organic debris from endodontic instruments is possible using a combination of physical and chemical procedures, but meticulous care in its removal is necessary. Our results confirm the need for a detailed application of the endodontic instrument cleaning process in order to provide the correct conditions for guaranteeing effectiveness during the sterilization process.
CONCLUSIONS
NiTi endodontic instruments used in clinical conditions retain a significant amount of biological debris that may represent a biohazard and affect effective sterilization. The Nd: Yag laser (250 mJ, 3Hz, at 6 cm from the sample) is a technique that removes the biological debris and the elements contained in the debris from the surface of used endodontic instruments, making it a viable alternative to the use of ultrasound cleaning. The LIBS technique is a useful method for detecting constituent elements of dentin debris and NiTi instruments.
REFERENCES
Montero-Miralles P, Estévez-Luaña R, DeGregorio-González C, Valencia-dePablo O, Jaramillo DE, Cisneros-Cabello R. Effectiveness of Nd:YAG Laser on the elimination of debris and Smear Layer. A comparative study with two different irrigation solution: EDTA and QMix® in addition to NaOCl. J Clin Exp Dent. 2018; 10 (1): 70-74. doi: 10.4317/jced.54395.
AFFILIATIONS
1 Egresada del Posgrado en Endodoncia. Facultad de Odontología de la Universidad Autónoma de Tamaulipas.
2 Profesor de tiempo completo. Facultad de Odontología de la Universidad Autónoma de Tamaulipas.
CORRESPONDENCE
Rogelio Oliver Parra. E-mail: roliver@docentes.uat.edu.mxReceived: Junio 2020. Accepted: Abril 2021.