|Year : 2022 | Volume
| Issue : 3 | Page : 338-346
Comparative assessment of the efficacy of low concentration bleaching agents using quantitative light induced fluorescence in removing stains: An In vitro study
Aiswarya Balakrishnan, Nandlal Bhojraj, Raghavendra Shanbhog, KP Ashwini
Department of Pediatric and Preventive Dentistry, JSS Dental College, Mysuru, Karnataka, India
|Date of Submission||02-Jul-2022|
|Date of Decision||16-Sep-2022|
|Date of Acceptance||17-Sep-2022|
|Date of Web Publication||18-Oct-2022|
Department of Pediatrics and Preventive Dentistry, JSS Dental College, Mysuru, Karnataka
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background: Tooth discoloration has become a common esthetic problem in recent years. Removal of stains by bleaching is well-documented. Low concentration home bleaching products are available in market in different forms and concentrations. Aim: The aim of this study is to evaluate and compare the efficacy of low concentration commercially available home bleaching products (whitening strip, gel, and mouthwash) in removing stains and whitening the tooth using clinical and digital methods. Materials and Methods: Sixty permanent enamel samples mounted in an acrylic block were artificially stained and randomly divided into four groups. Negative control, 15 % Carbamide peroxide gel group, 2% Hydrogen 16 peroxide mouthwash group and 6% Hydrogen peroxide strip group respectively. The samples were bleached with respective agents according to the manufacturer's instructions. The efficacy on 7th and 14th day was evaluated clinically (SGU change), photographically (ΔE), and using quantitative light-induced fluorescence (ΔF). The data were analyzed using paired t-test and analysis of variance. Results: Postbleaching, 6% hydrogen peroxide strips and 15% carbamide peroxide gel showed maximum improvement (ΔΔF – 15.73 and 11.89, ΔE – 19.8 and 18.9, respectively) when compared to 2% hydrogen peroxide mouthwash and negative control group (ΔΔF – 9.68 and 6.59, ΔE – 15.04 and 9.44, respectively). The difference was statistically significant (P = 0.001). Conclusion: 6% hydrogen peroxide strips and 15% carbamide peroxide gel showed maximum improvement in stain removal and tooth whitening however, the strips showed better efficacy than the gel. Strips have the added advantage of lesser contact period, less salivary dilution, and no gingival contact. Therefore, strips can be a better alternative for gels and mouthwashes.
Keywords: Home bleaching agents, quantitative light-induced fluorescence, whitening gel, whitening mouthwash, whitening strips
|How to cite this article:|
Balakrishnan A, Bhojraj N, Shanbhog R, Ashwini K P. Comparative assessment of the efficacy of low concentration bleaching agents using quantitative light induced fluorescence in removing stains: An In vitro study. J Indian Soc Pedod Prev Dent 2022;40:338-46
|How to cite this URL:|
Balakrishnan A, Bhojraj N, Shanbhog R, Ashwini K P. Comparative assessment of the efficacy of low concentration bleaching agents using quantitative light induced fluorescence in removing stains: An In vitro study. J Indian Soc Pedod Prev Dent [serial online] 2022 [cited 2022 Dec 3];40:338-46. Available from: http://www.jisppd.com/text.asp?2022/40/3/338/358838
| Introduction|| |
Esthetic dentistry has received increased attention in recent years, especially due to the fact that people are more concerned about the esthetic appearance of their smile. Dental esthetics is said to have a significant psychological impact on quality of life even in children, adolescents, and young adults.,,
The prolonged color durability of stain molecules on the tooth enamel is considered one of the significant factors included in esthetic requirements. Tooth discoloration can be influenced by a combination of intrinsic and extrinsic factors. Intrinsic stains are related to enamel and dentin properties, while extrinsic stains are associated with the deposition of either food or beverage stains on the tooth surface.
A number of methods are available to improve the color of teeth such as whitening toothpastes, professional stain removal, enamel microabrasion, vital tooth bleaching, nonvital tooth bleaching, crowns, and veneers. Of these, there is more demand for noninvasive dental procedures aiming to improve the tooth color, and dentist-supervised home-use tooth bleaching with custom trays is the most common bleaching procedure dispensed by dentists to their patients.
However, in the current past, many other over-the-counter (OTC) products with low levels of carbamide or hydrogen peroxide are widely available to consumers at pharmacies, supermarkets, and over the Internet. They are available in different concentrations and forms such as gels, rinses, gums, dentifrices, whitening strips, or paint-on films.,, The results obtained from these self-applied bleaching treatments may not be as good as those offered by dentist-guided treatments but they have some benefits like lesser side effects and can be used at consumer's convenience. Moreover, unlike tray-based gel system where a customized tray is to be fabricated by a professional, other products such as rinses, gums, dentifrices, whitening strips, or paint-on films do not require any special armamentarium for its application and does not have hours of wear time which makes it more user friendly than others. Also, above-said products act as an alternative to conventional treatment for tooth discoloration with lower cost than traditional professional-prescribed/guided products.
There is an increased accessibility of OTC whitening agents with very little data in the literature regarding their effectiveness. Furthermore, there is a lack of clinical trials that provide substantial scientific background regarding the efficacy of whitening mouthwashes and whitening strips against tray-based home bleaching gel.
Hence, the present study was planned with the aim to evaluate and compare the efficacy of three 67 commercially available low concentration bleaching agents in removing stains from tooth enamel.
| Materials and Methods|| |
Study design and location
The present randomized group design experimental in vitro study was conducted in the departmental research unit. This experiment was approved by the Institutional Ethical Committee (Protocol no. 59/2019). Written informed consent was obtained from the subjects from whom the tooth samples were collected.
Sample size calculation
The sample size was calculated based on previous study results in which the mean difference of ΔE values between the groups was found to be 16 with SD ± 4 using 80% power and 95% confidence interval; sample size was calculated to be 15 in each group. The study included three interventional groups and one control, thus having four groups and was completed in a span of 2 months.
For this study, 100 human premolar teeth extracted for orthodontic reasons with their initial shade greater than or equal to B2 (Vita Classical guide-VITA Zahnfabrik, Bad Säckingen, Germany) were collected, cleaned, and stored in distilled water at 4°C till further use. Teeth were examined under microscope and with QLF for the defects. Sixty premolars free from caries, cracks, restorations, or other surface defects were finally selected for the study.
The collected teeth were decoronated at the cement enamel junction, and the crown was sectioned mesiodistally into two equal halves using a diamond disc (Diatech CH- 9435, Swiss Dental Instruments, Lot: 9605) mounted on a slow-speed Micromotor Straight Handpiece (SH-E, NSK, Nakanishi Inc., Shimohinata, Kanuma, Japan) under continuous irrigation with deionized water. From sectioned samples, 60 buccal halves were used for the study. For mounting the samples, a customized jig was prepared using acrylic sheets having slots of 2 cm × 3 cm. The jig consisted of 10 slots; 5 on each side of dimension 2 cm × 3 cm. Cold cure acrylic was placed within after application of separating media using sprinkle on method and then tooth positioning groove made using pre-fabricated acrylic cut out. The tooth was positioned in this groove with the buccal surface facing outward and remaining cold cure placed to obtain a block of dimension 2 cm × 2.5 cm × 1 cm. The horizontal orientation of the groove was controlled using the acrylic cut out along with double-ended adhesive paper. The tooth was positioned in a manner so that only the buccal surface was exposed and the cut surface and was embedded into the acrylic block. The outer 200 μm of surface enamel was removed using a fine emery grit of 400, 800, 1000, and 2000 in a serial manner to obtain a flat enamel surface free of surface contaminants. The samples were then polished with a slurry of pumice. Following this, a polyvinyl-colored adhesive tape of 2 mm × 2 mm dimension was placed on the surface of samples and they were coated with an acid-resistant transparent nail varnish (Lakme, India). After setting of the varnish, the adhesive tape was removed leaving a window of 2 × 2 mm on the enamel surface (Area of interest [AOI]). The AOI was cleaned with a micro brush under a low magnification microscope.
Staining of samples
The artificial staining protocol used for the study was followed as recommended by Vejai Vekaash et al. The AOI of all the prepared samples was etched using 35% phosphoric acid gel for 15 s and rinsed for 30 s using distilled water. This was carried out to encourage the uptake of stain into the sample. For staining tea solution was used. The tea solution was prepared by boiling 100 mL of distilled water with 2 g of tea (Brooke bond, Red label) in it for 5 min and filtered using gauze. The solution was cooled to room temperature. Each sample was then immersed in 30 mL of tea solution and incubated at 37°C for 24 h. Later, samples were washed thoroughly and stored at 37°C and 100% humidity.
Randomization and stratification
The stained samples were randomly divided into four groups. Group I – Negative Control-Distilled water, Group II – 15% carbamide peroxide gel, Group III – 2% hydrogen peroxide mouthwash, and Group IV – 6% hydrogen peroxide strips. The samples were number coded for easy identification. The composition of the interventional agents is elaborated in [Table 1].
Group 1 samples were immersed in 2ml distilled water for 2 min twice daily for 14 days. For Group II samples opalescent PF home bleaching gel containing 15% carbamide peroxide was applied using a customized tray. Customized tray was prepared using soft thin tray material using a vacuum molding machine (Jaypee Ashvac, India). This was fabricated to standardize the amount of gel applied for each sample. Impressions of the samples were taken using alginate and casts were poured with dental stone. At the AOI, 0.5 mm of block out resin was applied to create a relief area in the tray. A liquid separator was applied on all surfaces and allowed to dry. Moreover, customized tray was fabricated using a vacuum molding machine [Figure 1]. After preparing the tray, a drop of gel was placed over the relief area (0.5 mm) of the tray and the tray was placed on the samples and stored in a humid environment for 5 h in the incubator. The samples were then washed and stored in artificial saliva. This was done once daily for 14 days. In Group III, the samples were immersed in 2 ml of optic white mouth wash containing 2% hydrogen peroxide for 2 min under constant agitation using a dental laboratory vibrator (Major Vibrator c-71 Confident Dental Equipment Limited). This was done twice daily for 14 days. After the application of mouthwash, the samples were washed and stored in artificial saliva. For Group 4, Crest 3D professional effect whitening strips containing 6% hydrogen peroxide were cut into 2 × 2 mm dimension and were placed over the AOI of the sample for 30 min in a humid environment and were kept in the incubator. After 30 min, the strips were removed manually, washed, and stored in artificial saliva. This was done twice daily for 14 days.
|Figure 1: (a) Customized tray fabricated for gel application (b) application of the gel to the tray (c) sample fitted in the tray|
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The efficacy of stain removal at baseline and postintervention 7th and 14th days was assessed using Clinical (shade Guide) and Digital (QLF and Photographic) methods. Two trained examiners performed the evaluation. The interexaminer reproducibility of shade scoring was assessed by the kappa statistics. The k value for interexaminer reproducibility was 0.87.
Clinical evaluation was performed using Classical Vita shade guide (Vita Zahnfabrik, Germany). The shade tabs were arranged in a sequence suggested by the manufacturer, and each shade was assigned a numeric value ranging from 1 to 16 (B1, A1, B2, D2, A2, C1, C2, D4, A3, D3, B3, A3.5, B4, C3, A4, C4). Shade selection was performed at AOI of each sample visually with reference to shade guide with shade mapping technique as illustrated in [Figure 2]. The shade mapping technique is used in esthetic dentistry where the tooth surface is divided into 3 or 9 segments and each region is matched independently. The Shade obtained for all samples after staining was darker than the darkest shade (C4) available in the shade guide. Therefore, C4 was given for all samples at baseline. Once shade selection was done at all intervals shade guide unit (SGU) change which indicates the improvement in shade from baseline to post bleaching was computed.
|Figure 2: The marked area was used as AOI for shade selection using Vita shade guide, for obtaining CIELAB values automatically generated by a software (Image J) and for QLF analysis. QLE: Quantitative light-induced fluorescence, AOI: Area of interest|
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Quantitative light-induced fluorescence TM evaluation
QLF™ images were captured by a single trained examiner using a QLF-D Biluminator™ device (Inspektor Research Systems BV, The Netherlands) with the help of a customized jig to ensure standardized position of samples. The images were captured under class 1 ASA darkroom conditions. Images were 168 taken with the following specifications; for fluorescent images ISO speed of 1600, an aperture value of 7.1 and shutter speed of 1/20 s and for the white-light images ISO speed of 1600, an aperture value of 13 and shutter speed of 1/30 s. The distance and the angle between the samples and QLF camera were maintained at 1 cm and 90° respectively using the customized jig. The fluorescent images were then analyzed using the QLF software (QA2 v 1.26, Inspektor Research Systems BV, Amsterdam, The Netherlands) to obtain (ΔF) which is the percentage loss of fluorescence in the stained area when compared to the unstained areas. The percentage change in ΔF was calculated from baseline to 7th day 175 and 14th day respectively to compare the reduction in stain and were depicted as ΔΔF.
For photographic evaluation, the white light images obtained from QLF were used. The CIELAB values were measured from white-light images with the help of standard image analysis software (Image J version 1.47, National Institutes of Health, USA). The CIELAB space is three dimensional and covers the entire range of human color perception where L values represented the lightness ranging from 0 (black) to 100 (white), and the a and b values representing the redness-to-greenness and yellowness-to-blueness axes, respectively. The stained area was selected within the same AOI by one calibrated examiner to obtain the L*a*b values. Tooth color difference (ΔE) refers to the color distance within a color space. The ΔE values were measured on each of the three principal axes of the CIELAB color space. The following formula was used to calculate color differences to obtain tooth color difference (ΔE) at 7th and 14th day intervals. ΔE = [(L After bleaching – L After staining) 2+ (a After bleaching – a After staining) 2+ (b After bleaching – b After staining) 2] ½.
Data were tabulated and statistical analysis was done using SPSS software v 23.0 (SPSS Inc., IBM, 190 Chicago, IL, USA). Intragroup comparison was done using One-way analysis of variance with Tukey's Post-hoc. Group comparison of means at two different time intervals was done by independent t-test.
| Results|| |
Intragroup comparison of ΔF
Significant reduction of ΔF was observed from baseline to 7th and 14th days in all the groups including the negative control group [Table 2]. This reduction was maximum for 6% hydrogen peroxide strips followed by 15% carbamide peroxide, 2% hydrogen peroxide mouth wash when compared to negative control. This difference is statistically significant (P = 0.001).
|Table 2: Mean and standard deviation of ΔF values at given time intervals for different groups|
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Intragroup comparison of bleaching efficacy
[Table 3] demonstrates that the improvement showed by all the groups from baseline to 7th day and 14th day, respectively, in terms of ΔΔF, ΔE values, and SGU change.
|Table 3: Intra-group comparison of ΔΔF, ΔE and shade guide unit change obtained for different groups|
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6% hydrogen peroxide strips and 15% carbamide peroxide gel showed a significant improvement in the initial 7 days only and very minimal change was seen in the following 7 days whereas 2% hydrogen peroxide mouth wash showed minimal improvement change in the initial 7 days but a significant improvement was observed by 14th day. This indicates that 2% hydrogen peroxide mouth wash requires the usage of longer time period to show significant improvement.
Intergroup comparison of bleaching efficacy
[Table 4] demonstrates the intergroup comparison of ΔΔF, ΔE, and SGU values of different interventional agents at 7th day and 14th day intervals.
|Table 4: Inter group comparison between negative control and interventional agents|
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Similar results were obtained from all three different methods. On 7th day, 6% hydrogen peroxide strips showed maximum improvement followed by 15% carbamide peroxide Gel, 2% hydrogen peroxide mouthwash and least for negative control group. This difference was statistically significant for ΔΔF, ΔE, and SGU values. Similar results were obtained on 14th day as well. However, Tukey's B post hoc showed that on 7th and 14th days the ΔΔF values of 15% carbamide peroxide gel and 2% hydrogen peroxide mouthwash were comparable.
[Figure 3], [Figure 4], [Figure 5], [Figure 6] shows the gradual removal of stains and shade improvement of respective groups from baseline to the 14th day of bleaching.
|Figure 3: Negative control (a) white light images (b) fluorescent images (i) baseline (ii) 7th day (iii) 14th day|
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|Figure 4: Fifteen percent carbamide peroxide gel (a) white light images (b) fluorescent images (i) baseline (ii) 7th day (iii) 14th day|
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|Figure 5: Two percent hydrogen peroxide mouthwash (a) white light images (b) fluorescent images (i) baseline (ii) 7th day (iii) 14th day|
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|Figure 6: Six percent hydrogen peroxide strips (a) white light images (b) fluorescent images (i) baseline (ii) 7th day (iii) 14th day|
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| Discussion|| |
The growing interest in the treatment of tooth discoloration is reflected in the ample number of commercially available tooth whitening products. A few among these are over-the-counter products that do not require the involvement of a dental professional in their application. These systems use stock trays, strips, and mouthwashes other means for delivering peroxide. A variety of products are constantly entering the market. They differ in terms of peroxide concentration, product formulation, delivery method, contact time, and other factors, all of which can affect individual's tolerability and response.,
Gel form is one of the oldest methods used where lower concentration of hydrogen peroxide or carbamide peroxide is used as active agent. A recent meta-analysis reported that tray-delivered carbamide peroxide gels showed a slightly better whitening efficacy with lesser side effects such as gingival irritation and hyper-sensitivity than hydrogen peroxide-based products. Hence, 15% carbamide peroxide gel was selected for the present study. Compared to other OTC products, the need to visit a dentist for a custom fabricated tray is an additional requirement for this product. Strips and mouth washes are relatively newer products. Strips deliver a controlled and relatively low dose of bleaching active to the anterior dentition only. Strips are available generally at 6%–10% hydrogen peroxide concentrations. The lowest available concentration was selected for the study. Mouth rinses are popular due to their ease of use. They are usually composed of low HP concentration (1%2%). Even though theoretically, it is expected that due to lesser concentration, liquid consistency, and shorter contact period bleaching efficacy of mouthwash can be lesser than strips or tray-based gel only limited studies were found in literature where it was evaluated and compared to other OTC products. Thus, in the present study, we have compared three different commercially available home bleaching systems the 3D Crest professional effects whitening strips (Crest 3D, USA) containing 6% hydrogen peroxide, opalescent PF 15% home bleaching gel (Ultradent, USA) containing 15% carbamide peroxide and optic white mouth wash (Colgate-Palmolive, USA) for the efficacy of removing stains and whitening the tooth.
A variety of artificial staining protocols can be found in the literature producing mild-to-severe degree of staining.,,, Extensive staining requires appropriate professional care with a higher concentration of bleaching agents. For the present study, moderate staining was adequate. Hence, a protocol that generated an appropriate degree of stain was selected.
Shade guide comparison, digital image analysis, or combination of them is commonly used to detect color change. However, visual assessment and longitudinal monitoring are subjective and vary according to the observer, shade guide used, and the viewing environment., Traditionally used methods for measuring color change such as reflectance colorimetry and spectrophotometry were originally meant for industrial use and hence can be costly, not easily available, and unable to use as a chairside method. Furthermore, its application is limited to larger samples and hence cannot be used in dentistry as we deal with smaller teeth samples.,,
Hence, in this study, a noninvasive diagnostic equipment, QLF was used for this purpose. Although QLF was developed originally for detecting and quantifying white spot lesions, it also has the ability to quantify stain uptake and removal., The stain produced on the surface of the tooth resembles demineralized lesions under QLF imaging. In 2001, Amaechi and Higham demonstrated the use of QLF to quantify the gradual color change of stained teeth followed by application of whitening agent. Adeyemi et al. in a study in 2010 reported a novel method using QLF, enabling its use for assessing and quantifying tooth surface staining and whitening. He compared the ΔF values obtained from QLF with the ΔE values obtained from spectrophotometer and concluded that QLF had a high correlation with spectrophotometry for identifying and monitoring the progression and removal of tooth stains. Later, in another study by Adeyemi et al. they showed a high correlation between ΔQ computed from QLF and ΔE obtained from digital image analysis. Lee et al. in their study in 2019 used white light images recorded from QLF to obtain ΔE using Image J software and found the ΔE and ΔΔF (change in ΔF) obtained from blue light images of QLF showed a statistically significant correlation.
Even though QLF has been used to evaluate the staining and whitening,,,,,, none of the previous studies have assessed low concentration home bleaching products under QLF. This study aimed to contribute to the literature by quantifying the gradual change in color of stained teeth following the application of commercially available home use tooth whitening agents using QLF. In the present study, QLF™-D Biluminator (Inspektor Systems, Amsterdams) was used. QLF raises the visual contrast between sound and pathogenic tissue. Hence, in order to maintain sound tooth structure, a 2 * 2 window (AOI) was created on the surface of samples. ΔF was obtained from fluorescent images which is the measure of percentage loss of fluorescence with respect to that of surrounding sound tooth structure. This value is related to the lesion depth.
Photographic analysis was used to measure the tooth color difference (ΔE). ΔE values are measured on each of the three principal axes of the CIE L a b color space using Image J software from white light images at AOI. Also, for visual shade assessment a classical Vita shade was used. These data were expressed as SGU change. The measurement of tooth color (ΔE) with the help of digital image analysis and expression of the color changes in terms of CIELab values to evaluate the efficiency of bleaching products over time has been successfully used., Visual tooth color determination by comparison with standard shade guides is the most commonly used method in dentistry and a standard Vita shade guide is the most commonly used shade guide. Shade selection was done by comparing the shades at AOI of each sample visually to the shade guide and a value was given as done in shade mapping technique. This technique has been used here as it was necessary to maintain sound tooth structure for QLF analysis. In the present study, we used shade guide analysis based on a study by Suleiman et al. where shade guide was used for shade determination after staining but this method was found inaccurate as shade obtained after staining was darker than any shade available in the guide. Hence digital image analysis can be considered a better option for color analysis following staining.
Results acquired from QLF, photographic, and shade guide analysis was similar. The obtained results can be directly attributed to the concentration of bleaching agent. The null hypothesis of this was rejected as the present study results showed a significant difference in the efficacy of the bleaching products used. A better efficacy was showed by 6% hydrogen peroxide strip. Apart from the concentration, flexible adhesive polyethylene strip can hold peroxide close to the tooth surface avoiding any salivary dilution. Thus, this system has shown to produce better result even with shorter contact period. Tray-based system utilizes 15% CP which is equal to ~5% HP that can be delivered on a customized bleaching tray and must be worn for several hours. Because of comparatively lesser concentration and possible salivary dilution factor, tray-based gels require a prolonged contact period. Furthermore, maximum reduction of stain in the 6% hydrogen peroxide strip and 15% carbamide peroxide gel has occurred in the initial 7 days only. From 7th day to 14th day the reduction is minimal indicating that these 2 systems can produce appreciable improvement within a span of 1 week use only. While the mouthwash group becomes more effective after 7th day, therefore, it would be advisable to use mouthwash for longer span for good results. Further studies are required to find the number of days to be used for this whitening mouthwash to give the best results. The results based on ΔE and SGU change obtained using 6% hydrogen peroxide strips group in the present study is similar to that conducted by Swift et al., where he concluded that teeth became lighter and less yellow when whitening strips were used twice daily for 30 min. In addition, in the present study, the efficacy of 6% hydrogen peroxide strips group was slightly better than 15% carbamide peroxide gel group based on the ΔE values and SGU change even though there was no statistical difference present. A similar result was obtained by Karpinia et al.
The following can be considered as the limitations of the present study,
- The stain uptake and removal have been observed using visual and digital means of color assessment with Vita Shade guide and QLF, respectively, this was not compared with a gold standard method like chromatographic analysis with a spectrophotometer
- Since, the present study was conducted in vitro, in vivo usage of bleaching agents might show variation due to the presence of contributing factors. Hence, further clinical studies would be required on the same.
| Conclusion|| |
Considering the limitations of the current study, it can be concluded that:
- Six percent hydrogen peroxide strips showed maximum improvement followed by 15% carbamide peroxide gel, 2% hydrogen peroxide mouth wash group when compared to the negative control group
- Six percent hydrogen peroxide strips and 15% carbamide peroxide gel showed significant improvement in the initial 7 days, only minimal change was only seen in the following 7 days whereas 2% hydrogen peroxide mouth wash group showed minimal improvement change in the initial 7 days but a significant improvement was observed by 14th day.
The authors would like to express their gratitude to JSS Dental College and Hospital for the support.
Financial support and sponsorship
JSS AHER Research Grants/Financial assistance DCH/ACG/09/2021-22/111.
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6]
[Table 1], [Table 2], [Table 3], [Table 4]