This study investigates the antibacterial effects of erbium chromium laser at 2780 nm, silver nanoparticles, and erbium chromium along with silver nanoparticles on
Silver nanoparticles have various medical applications due to their antibacterial and anti-inflammatory properties. The size of these particles ranges from 10 to 100 nm and they have antibacterial effects against Gram-negative and positive bacteria and also multidrug-resistant bacteria [
The root canal diseases are associated with pain, inflammation, and discomfort of the patient. On the other hand,
The following materials were used: diamond disk (DFS Diamon Co., Germany) for removing the crown, K-files (Mani Inc., Japan), and rotary files (V-Taper Gold, Shanghai Fanta Dental, China) for preparation of the root canals. Normal saline (sodium chloride, 0.9%) (Shahid ghazi Co., Tabriz, Iran), EDTA 17% (Maraboon, Iran), and 5% sodium hypochlorite (Paxan Co., Tehran, Iran) for irrigation of root canals. Eppendorf tube (Amin Co., Iran) and Acryle (Acropars Co., Iran) for fixing the roots. Standard strain of
The study was performed on 90 newly extracted single-canal human teeth and their single-canal was confirmed by radiography. The teeth were collected from comprehensive health centers. Sampling of single-canal anterior teeth was performed randomly based on the following inclusion criteria: teeth without any developmental defects, caries, and cracks or fractures. Moreover, nonaged teeth were selected to ensure that there is no dentin sclerosis and the calcified dental canal in the samples. Dental radiographs were taken to ensure that the selected teeth are single channel before operation, and teeth with curved roots were excluded.
The samples were classified randomly into different treatment groups. In order to prepare the samples, their roots were cut at approximately 15 mm of apex using a diamond disk, and its working length was adjusted at 1 mm shorter than the apical foramen (14 mm). The root canals were first prepared up to No. 20 using manual K-files (Mani Inc, Japan), and 5% sodium hypochlorite (Paxan Co., Tehran, Iran) then prepared by rotary files (V-Taper Gold, Shanghai Fanta Dental, China) to obtain the original file. During the use of rotary files, the canals were washed with normal saline (sodium chloride, 0.9%) and 5% sodium hypochlorite. After preparation with files, the canals were washed with 1 mL of17% EDTA (Maraboon, Iran), 5 mL of normal saline, and 1 mL of 5% sodium hypochlorite and kept for 5 min in order to remove smear layer. Eventually, all canals were washed with 5 mL of normal saline. Afterwards, the roots of the teeth were fixed into a 2 mL microtube with acrylic (Acropars Co., Iran) and incubated at 121°C for 15 minutes [
To prepare silver nanoparticle suspension, pomegranate skin was dried in an oven at 50°C for 2 days, and to prepare the hydroalcoholic extract, 50 g of the pomegranate skin was soaked in 500 mL of alcohol (70%) at room temperature for 72 hours. The extract of pomegranate powder was then extracted using Whitman No. 40 filter papers. In order to separate the fine suspended particles from the extract, a centrifuge was used. A certain amount of silver nitrate and the extract were dissolved in 200 mL of water to prepare the desired concentrations (100 and 200 ppm). The color of the solution changed from colorless to gray, which indicated the successful synthesis of AgPPENPs [
To examine potential toxicity of formulation, RMPI medium in 5% CO2 of atmosphere with the temperature of 37°C was used and fibroblast cell line L929 was cultured within. The cultured cells were exposed to different concentration of PPE and AgPPENPs (100, 200, 300, 400, and 500
The standard strain of
Group 1: cleaning with 5% sodium hypochlorite: in this group, the root canal of the teeth was cleaned routinely with injecting 5 mL of 5% sodium hypochlorite solution (Snow, Paxan Co., Tehran, Iran) in apical third of canal (one or two mm from the foramen) using a 30-G insulin syringe and kept in the canal for 5 min. Then, 1 mL of 5% sodium thiosulfate solution (Samchun Pure Chemical, Korea) was injected into neutralize sodium hypochlorite solution and kept in the canal.
Group 2: cleaning with silver nanoparticles: the root canals were washed with 5 mL of the prepared silver nanoparticle suspension with a concentration of 100 ppm (with mean particle size of 20 nm). For this reason, an insulin syringe (Supa, Iran) was utilized and kept in the canal for 5 min.
Group 3: using 2780 nm erbium chromium laser: the root canals of the teeth were cleaned using erbium chromium laser (Biolase, Waterlase, USA) at a wavelength of 2780 nm, 0.0% of water, 15% of air, and 75 watts of power. The power density was set at 2.5 watt/cm2 and energy density was equal to 25 J/cm2. The laser head with a diameter of 200
Group 4: simultaneous use of erbium chromium laser and silver nanoparticles: in this group, the root canals were cleaned with 5 mL of the synthetized silver nanoparticles at a concentration of 100 ppm (with mean particle size of 20 nm) using an insulin syringe (Supa, Iran). After 5 min, the erbium chromium laser with a wavelength of 2780 nm was applied for 30 sec, and this operation was conducted three times with an interval of 15 sec between each irradiation similar to the group.
Group 5: negative control group (normal saline): root canals were washed with 5 mL normal saline using insulin syringe.
To standardize the bacterial prototype after the extraction of TSB and washing the canal with 5 ml normal saline, root canals were filled with sterile saline solution using a 30-G syringe, and dentin was scraped from inside the canals using a #40 Hedstrom file (Mani Inc, Tochigi, Japan). A #40 sterile paper point (Gapadent Co, Hamburg, Germany) was placed inside the canals for 60 seconds and was then transferred into a sterile microtube containing 1 mL Tween 80 solution (Samchun Pure Chemical, Korea) and vortexed for 20 seconds to isolate the bacterial biofilm. One hundred dilution was made from the first sample and cultured on blood agar medium and the plates were incubated at 37°C for 24 h and colony counts were reported according to CFU/mL (8).
To standardize all groups, the canals were washed with 5 mL of normal saline and the solution was kept in the root canal for 30 sec. Then, the sampling was performed in accordance with the primary sampling method and colony counts were reported according to CFU/mL.
Accordingly, SPSS software (version 21) was utilized to analyze the data. For this purpose, the central dispersion indices for the number of live bacterial colonies were calculated and reported before and after the intervention. The decrease in bacterial colonies after the intervention in different groups was compared by Kruskal–Wallis nonparametric test. Moreover, changes in bacterial colonies before and after the intervention were recorded using Wilcoxon signed-rank test. Given the fact that the data were not in line with the normal distribution and presence of dispersed data in the colonies, nonparametric analyses were performed in the study. Type I error rate was set at 0.05 (
The mean of zeta potential for the nanoparticles at pH = 7 was −28 mV.
TEM image results showed that the mean diameter of NPs was 20 nm (Figure
Transmission electron microscopy (TEM) image of the AgPPENPs (scale bar is 50 nm).
Figure
X-ray diffraction (XRD) pattern of AgPPENPs.
To assess the potential toxicity of synthesized nanoparticles, cell viability assay was applied. The results pointed that AgPPENPs could decrease cell viability in the concentration of 400 and 500
Cells viability of treatments with different concentrations of AgNPs-PPE and PPE after 48 hrs.
According to the results of Wilcoxon test, there was a significant decrease in the number of bacterial colonies after the intervention (
Descriptive values of
Groups | Time | Minimum | Maximum | RCC % | |
---|---|---|---|---|---|
Sodium hypochlorite | Before | 103 × 12 | 103 × 27 | 100 | 0.001> |
After | 0 | 0 | |||
Silver nanoparticle | Before | 103 × 5 | 103 × 60 | 83.15 | 0.001> |
After | 0 | 103 × 8 | |||
Erbium chromium laser | Before | 103 × 11 | 103 × 36 | 47.90 | 0.001> |
After | 103 × 3 | 103 × 30 | |||
Silver nanoparticles and erbium chromium lasers | Before | 103 × 4 | 103 × 49 | 64.72 | 0.001> |
After | 0 | 103 × 13 | |||
Negative control (washing with normal saline) | Before | 103 × 3 | 103 × 58 | 49 | 0.005> |
After | 103 | 103 × 41 |
Bacterial culturing before and after intervention. (a) Negative control, a1 before a2 after. (b) Hypochlorite sodium, b1 before b2 after. (c) Nanosilvers, c1 before c2 after. (d) Erbium chromium laser, d1 before d2 after. (e) Nanosilvers and erbium chromium laser, e1 before e2 after.
Kruskal–Wallis test was used to compare the performance of different interventions with respect to the reduction percentage of bacterial colonies based on the mean values of the reduction of colony count (RCC). The results of this test showed that there was a significant difference between the reductions of bacterial colonies between different groups (
Based on this result, Mann–Whitney test was used to compare the two groups. In most cases, the type I error rate
Results of Mann–Whitney test with Bonferroni adjustment for comparing the two groups.
Groups | RCC mean |
---|---|
Sodium hypochlorite, negative control group | 0.001 |
Erbium chromium laser, negative control group | 0.943 |
Silver nanoparticles, negative control group | 0.001 |
Erbium chromium laser and silver nanoparticles, negative control group | 0.102 |
Silver nanoparticles, sodium hypochlorite | 0.001 |
Erbium chromium laser, sodium hypochlorite | 0.001 |
Erbium chromium laser and silver nanoparticles, sodium hypochlorite | 0.001 |
Silver nanoparticles, erbium chromium laser | 0.001 |
Silver nanoparticles, erbium chromium lasers and silver nanoparticles | 0.009 |
Silver nanoparticles and erbium chromium laser, erbium chromium laser | 0.152 |
Results of Mann–Whitney test with Bonferroni adjustment showed that there was a significant difference between the sodium hypochlorite and silver nanoparticle groups with negative control group (
This study evaluated the antimicrobial effect of erbium chromium laser, silver nanoparticles, and their simultaneous applications on
In Kasić et al.’s [
Due to the high absorption of erbium laser radiations by water and hydroxyapatite, and thus the reduction of the laser thermal effect, lasers are more efficient in the dried canals. In general, lasers such as erbium chromium laser can kill bacteria through their thermal effects. Therefore, the moisture can act as a reducing agent and subsequently reduces the lethal effect of lasers on bacteria [
According to the results of Wanda Gordon et al.’s study, the use of Er, Cr: YSGG laser at 240 nm and an exposure time of 240 sec showed higher efficiency than sodium hypochlorite in dried root canal, which resulted in 99.7% reduction in bacterial count [
In Eldeniz et al.’s study, sodium hypochlorite was used in root canal for 15 min, while in the present study this detergent was kept in canal for 5 min. Given the toxic effects of sodium hypochlorite, it has been suggested to be used for short contact times in dental applications. In Eldeniz et al.’s [
The antimicrobial property of sodium hypochlorite depends on the concentration of unconjugated hypochlorous acid in the solution. The hypochlorous acid interacts with sulfhydryl groups in bacterial enzymes. By preventing the activity of these essential enzymes, their basic metabolic reactions are disrupted and results in bacterial death [
In Charannya et al.’s study, the efficiency of silver nanoparticles in the mentioned study was lower than the present study, which could be attributed to the use of low concentrations [
There was a contradiction regarding the use of silver nanoparticles in cleaning root canals between our findings and the results of We et al.’s study, because the results of Wu et al.’s study showed that sodium hypochlorite 2% and 98.2% reduce the number of
The results implied that the application of erbium chromium reduced the number of bacterial colonies and there was a difference between before and after laser treatment. But they were not as effective as sodium hypochlorite and did not significantly reduce the number of
The data can be accessible to the interested researchers by the corresponding authors on reasonable request.
The authors declare that they have no conflicts of interest.
This study has been adapted from a M.Sc. thesis at Hamadan University of Medical Sciences (project no. 9510146067).