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 Table of Contents  
RESEARCH
Year : 2022  |  Volume : 22  |  Issue : 4  |  Page : 368-376

Effect of silver nanoparticles on wettability, anti-fungal effect, flexural strength, and color stability of injection-molded heat-cured polymethylmethacrylate in human saliva


Department of Prosthodontics, SRM Dental College, Chennai, Tamil Nadu, India

Date of Submission30-Dec-2021
Date of Decision23-May-2022
Date of Acceptance17-Jun-2022
Date of Web Publication03-Oct-2022

Correspondence Address:
J Brintha Jei
Department of Prosthodontics, SRM Dental College, Chennai, Tamil Nadu
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/jips.jips_574_21

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  Abstract 


Aim: Aim of this study was to evaluate the effect of silver nanoparticles incorporated injection molded heat-cured polymethylmethacrylate resin on wettability, anti-fungal effect, flexural strength and colour stability in human saliva.
Settings and Design: An In-Vitro study with In -Vivo parameter
Materials and Methods: Rectangular and circular stainless-steel dies were fabricated according to ISO standardization 20795-1:2018 and ADA specification number 12. A total of 144 samples were prepared and divided into 4 groups with thirty-six samples in each group. Each of the 4 groups were subdivided into 3 subgroups based on concentration of silver nanoparticles as 0% in subgroup A, 0.05% in subgroup B and 0.2% in subgroup C. Group 1 samples evaluated wettability, they were assessed at 0, 7, 90 and 180 days after immersing in human saliva using goniometer. Group 2 samples evaluated antifungal effect, they were assessed against Candida albicans in Muller hinton agar plate enriched with 2% glucose. Group 3 samples evaluated flexural strength, they were assessed by using universal testing machine. Group 4 samples evaluated colour stability, they were assessed using UV spectrophotometer at 0, 3 and 7 days after immersing in human saliva.
Statistical Analysis Used: One-way ANOVA and Post- Hoc Tukey test were used to evaluate the significant differences in the mean values of the groups.
Results: Subgroup C samples with 0.2% Ag nanoparticles had better wettability, maximum antifungal property, highest flexural strength and good colour stability followed by subgroup B and subgroup A samples.
Conclusion: Injection molded denture base resin incorporated with 0.2% Ag nanoparticles could be used clinically as a denture base material for completely and partially edentulous patients.

Keywords: Antifungal effect, color stability, flexural strength, injection-molded resins, silver nanoparticles, wettability


How to cite this article:
Vaiyshnavi W, Jei J B, Kumar B M. Effect of silver nanoparticles on wettability, anti-fungal effect, flexural strength, and color stability of injection-molded heat-cured polymethylmethacrylate in human saliva. J Indian Prosthodont Soc 2022;22:368-76

How to cite this URL:
Vaiyshnavi W, Jei J B, Kumar B M. Effect of silver nanoparticles on wettability, anti-fungal effect, flexural strength, and color stability of injection-molded heat-cured polymethylmethacrylate in human saliva. J Indian Prosthodont Soc [serial online] 2022 [cited 2022 Dec 7];22:368-76. Available from: https://www.j-ips.org/text.asp?2022/22/4/368/357804




  Introduction Top


Denture base resins used for fabricating removable prosthesis are polymethylmethacrylate (PMMA) was introduced in 1937.[1] Denture base resin has good esthetics, excellent biocompatibility, reliability, dimensional stability, absence of taste, odor, teeth adhesion, insolubility in body fluids, relative ease of manipulation, color stability, and low thermal conductivity.[2] The disadvantages are residual monomer toxicity and its effect on the oral tissues, the microbial colony with moderate-to-low mechanical properties that are susceptible to distortion and discoloration. The generation of cracks in denture base leads to fracture, which can also act as a point of entry for various bacteria.[3] High-impact injection molded PMMA was emerged in 1942 and provided better dimensional stability, wear strength, better deflection, and water sorption than conventional and reinforced PMMA.[4]

Wettability is affected by denture base resin to saliva contact. The degree of wetting is evaluated as the contact angle formed between liquid and solid. Human saliva has an important role to play in wettability with denture base resin. Denture base resins have influence toward the adhesion of candidal species. Nanoparticle incorporation exhibited enhanced mechanical, electrical, magnetic, and optical properties when compared with conventional PMMA. Various nanoparticles were incorporated in previous studies, but silver nanoparticles gained considerable attention because of its unique physical, biological, and anti-bacterial properties against Gram-positive and Gram-negative bacteria.[5] Flexural strength of denture base resin is one of the most important mechanical properties. The conventional PMMA has moderate flexural strength. By the addition of reinforcers flexural strength can be improved drastically. PMMA has a disadvantage of discoloration due to intrinsic and extrinsic staining from diet and habits. The use of high-impact injection-molded PMMA improves the physical properties of the denture base. This study evaluates the effect on wettability, anti-fungal effect, flexural strength, and color stability of injection molded heat-cured PMMA in human saliva with varying concentrations of silver nanoparticles.


  Materials and Methods Top


The study was approved by the Institutional Review Board (SRMDC/IRB/2019/MDS/No.202). The aim of this study was to evaluate the wettability, antifungal effect, flexural strength, and color stability of high-impact injection-molded PMMA reinforced with varying concentrations of silver nanoparticles in human saliva. The overall sample size was estimated to be 144. The samples were divided into four groups were and samples per group were 36. Each of four groups was again divided into 3 based on silver nanoparticle concentration. Rectangular stainless steel master die was fabricated according to ISO 20795-1:2018 with dimensions of 65 mm × 40 mm × 5 mm for evaluating flexural strength [Figure 1]. Circular stainless steel master die was fabricated according to ADA specification number 12 with dimensions 50 ± 1 mm × 1.0 ± 0.5 mm for evaluating wettability, antifungal effect, and color stability [Figure 2]. The samples were fabricated using this master die with high-impact injection-molded PMMA reinforced with silver nanoparticles at various concentrations [Figure 3]. All the samples were finished and polished using acrylic trimmers and sandpapers. The samples for flexural strength were cut into four strips of equal size measuring 65 mm × 10 mm × 5 mm [Figure 4].
Figure 1: Wax pattern for Group 1, 2 and 4

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Figure 2: Wax pattern for Group 3

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Figure 3: Specimens for Group 1, 2 and 4

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Figure 4: Specimens for Group 3

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Collection of unstimulated saliva

Unstimulated saliva from edentulous patients was collected by passive drooling method into a sterile container. Ten milliliters of the patient's saliva has been collected and was immediately stored in the freezer at 4°C to prevent bacterial growth and to prevent further degradation of salivary molecules. These salivary samples were used without further treatment for evaluating wettability and color stability.

Sample distribution

Group 1: Wettability

  • Subgroup A-12 samples – 0% Ag nanoparticles (control group)
  • Subgroup B-12 samples – 0.05% of Ag nanoparticles
  • Subgroup C-12 samples – 0.2% of Ag nanoparticles.


Group 2: Antifungal effect

  • Subgroup A-12 samples – 0% Ag nanoparticles (control group)
  • Subgroup B-12 samples – 0.05% of Ag nanoparticles
  • Subgroup C-12 samples – 0.2% of Ag nanoparticles.


Group 3: Flexural strength

  • Subgroup A-12 samples – 0% Ag nanoparticles (control group)
  • Subgroup B-12 samples – 0.05% of Ag nanoparticles
  • Subgroup C-12 samples – 0.2% of Ag nanoparticles.


Group 4: Color stability

  • Subgroup A-12 samples – 0% Ag nanoparticles (control group)
  • Subgroup B-12 samples – 0.05% of Ag nanoparticles
  • Subgroup C-12 samples – 0.2% of Ag nanoparticles.


Test for wettability

The control and reinforced group samples were placed in human saliva (collected in vails) under optimum conditions for 0, 7, 90, and 180 days and were tested for wettability using goniometer. Goniometer has a CCD camera that captured and recorded the image of a droplet of test liquid which was placed onto the surface of the specimen using microsyringe and the image processed to determine the contact angle in two different positions and average was calculated.

Test for antifungal effect

The antifungal activity was evaluated using the Kirby-Bauer disk diffusion method against Candida albicans. Fabricated control and reinforced group samples were placed on Mueller–Hinton agar plates in petri dish incorporated with 2% glucose and were inoculated with the microorganisms at 37°C for 24 h. The antifungal activity of the control and reinforced specimens were evaluated by measuring the formation of the inhibition zone in millimeters around the samples after 24 h and were statistically analyzed.

Test for flexural strength

The control and reinforced group samples were staged on Universal Testing Machine under 3-point loading for the evaluation of flexural strength. Load was being applied at the mid-point of the samples with crosshead speed of 5 mm/min. The maximal load before fracture was measured.

Flexural strength was calculated using the formula. M = 3WI/2bd2 where, M = flexural strength (MPa), W = fracture load (N), I = test span distance between support points in mm, b = width of specimen (mm), and d = thickness of the specimen (mm)

Test for color stability

Fabricated control and reinforced group samples were immersed in human saliva collected from healthy individuals and stored for 7 days and tested using spectrophotometer. The evaluation was done before immersion, on the 3rd day and on the 7th day after immersion in human saliva. Distilled water was used as a negative control medium. Images were taken under northern daylight on a clear day and color measurements were performed according to the Commission Internationale del'Eclairage L* a* b* uniform color scale. Measurements were taken on 3 different occasions and the mean values of L* (brightness), a* (red-green proportion), and b* (yellow–blue proportion) were calculated. The “corrected” L*, a*, b* values of each specimen were recorded as the baseline color readings before immersion in solution (T0-Dry). The posttreatment digital images of test specimens were obtained and analyzed to determine L*, a*, b* values of each specimen as mentioned previously. The total color change and ΔE of each test specimen were calculated using the equation. ΔE values ≤3.7 were considered to be visually imperceptible as well as clinically acceptable.


  Results Top


This study was done to investigate the effect of silver nanoparticles on wettability, anti-fungal effect, flexural strength, and color stability of injection-molded heat-cured PMMA in human saliva.

Wettability

The one-way ANOVA values for between and within the subgroup's comparison of the Group 1 samples are listed in [Table 1]. The sum of square value, the mean square value, and F value were found to be 123.656, 38.815, and 27.830 for the 0 day, for the samples stored for the 7th day the sum of square value, the mean square value and F value were found to be 298.669, 97.279, and 30.834, respectively. The sum of the square value, the mean square value, and F value for the samples stored for the 90th day were found to be 139.377, 3.345, and 0.832, respectively. The sum of square value, the mean square value and F value for the samples stored for the 180th day were found to be 234.607, 78.987, and 34.014, respectively. The significant difference was seen only on the 90th day with a value of 0.444 while other samples between and within the subgroups remained 0.
Table 1: One-way ANOVA descriptive analysis between and within the subgroups for wettability (Group 1)

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[Table 2] shows Tukey post hoc honestly significant difference (HSD) multiple comparison values among the subgroups for wettability of Group 1 samples. The highest mean difference for subgroup A was found to be when compared with C and the value was 3.7758. The highest mean difference for subgroup B was when compared with C and the value was 1.2775. The highest mean difference for subgroup C was 1.2775. The standard error remained the same for all the subgroups.
Table 2: Tukey post hoc honest significant difference multiple comparison values within the subgroups for the wettability (Group 1)

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Anti-fungal effect

[Table 3] shows the Tukey post hoc HSD multiple comparison values for the antifungal effect of Group 2 samples. The highest mean difference for subgroup A was with the value of −22.667 when compared with B. Highest mean difference for subgroup B was with the value of 22.667 when compared with A. Highest mean difference for subgroup C was with the value of 27.417 when compared with A. The standard error remained the same for all the subgroups.
Table 3: Tukey post hoc honest significant difference multiple comparison values for antifungal test (Group 2)

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Flexural strength

[Table 4] shows Tukey post hoc HSD multiple comparison values for flexural strength of Group 3 samples. The highest mean difference for subgroup A was −6.91833 when compared with subgroup B. Highest mean difference for subgroup B was 6.91833 when compared with. The highest mean difference for subgroup C was 9.99833 when compared with subgroup A. The standard error remained the same for all the subgroups.
Table 4: Tukey post hoc honest significant difference multiple comparison values for flexural strength (Group 3)

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Color stability

The one-way ANOVA values for between and within the subgroups comparison of Group 4 samples were listed in [Table 5]. The total sum of squares on the 0th, 3rd, and 7th days were 0.308, 0.310, and 0.318, respectively. The mean square value and F value on the 0 day were found to be 0.154 and 399549.453, respectively, and on the 3rd day, the mean square value and F value were found to be 0.155, and 1185201.538, respectively. The mean square value and F value for the samples stored for the 7th day were found to be 0.159, and 1147167.962, respectively.
Table 5: One-way ANOVA between and within the groups for color stability (Group 4)

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[Table 6] shows the Tukey post hoc HSD multiple comparison values for Group IV samples. The highest mean difference on the 0th day was 0.2024583 for subgroup C when compared with subgroup A. The highest mean difference on the 3rd day was 0.2028417 for subgroup C when compared with subgroup A. Highest mean difference on the 7th day was 0.2056250 for subgroup C when compared with subgroup A.
Table 6: Tukey post hoc honest significant difference multiple comparison values for color stability (Group 4)

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[Graph 1] represented the wettability values for the Group 1 samples. On the 0th day, 7th day, 90th day, and 180th-day subgroup A had maximum wettability, followed by subgroup B and subgroup C. Overall wettability was increased in all subgroups on all 4 days.



[Graph 2] represented the values for the antifungal property of the Group 2 samples and it was found that subgroup C had the highest antifungal activity, followed by subgroup B and the least or nil antifungal effect was seen for subgroup A.



[Graph 3] represented the values for the flexural strength of the Group 3 samples and it was found that subgroup C had the highest flexural strength, followed by subgroup B and the least flexural strength was seen in subgroup A.



The [Graph 4] represented the color stability values for the Group 4 samples. On the 0th day, all the three subgroups had similar values. On the 3rd day, all the three subgroups had similar values. On the 7th day, subgroup C had the highest value and subgroups A and B had similar values. Overall, the was an increase in color change in all three subgroups on all the 3 days.




  Discussion Top


High-impact injection-molded PMMA was emerged in 1942 and provided better dimensional stability, wear strength, better deflection, and water-sorption than conventional heat-cured PMMA.[6] Comparing the properties of conventional denture base materials and injection molded materials, Parvizi et al. concluded that high-impact injection-molded PMMA had the best dimensional accuracy among conventional heat cure PMMA, high -impact injection-molded PMMA, and nylon injection-molded PMMA.[4] Although high-impact injection-molded PMMA is better than conventional heat-cured PMMA, it also suffers such demands as fracture of the prosthesis, water sorption on long-term usage, denture stomatitis due to candidal adhesion, and color changes. Metal reinforcements can be used in high-impact injection-molded PMMA in a similar way that was carried out in conventional heat-cured PMMA. Such metal reinforcers used are nanoparticles.

Nanotechnology is a recently emerging field with extensive research in the characteristics of various nanoparticles that aid in dentistry as part of its treatment. Various nanoparticles were used as reinforcers in denture base materials. Silver nanoparticles were used for extensive experimentation in dentistry due to its optical property and antimicrobial effect. Few studies incorporated silver nanoparticles in conventional denture base materials and evaluated their properties. Sodagar et al. had reinforced silver nanoparticles in conventional heat-cured PMMA and showed improved results in terms of verifying the flexural strength.[7]

Apart from the evaluation of the flexural strength of silver nanoparticles reinforced high impact injection-molded PMMA, a few other properties of interest in this study were wettability, antifungal effect, and color stability. The wetting properties of the denture and the palatal mucosa occur through the adhesive forces (saliva) at the two interfaces which affect denture retention. Contact angle hysteresis is influenced by surface heterogeneity, surface roughness, surface deformation, and chemical contamination of water while rinsing.[8] Jaiswal et al. studied the wettability of conventional heat-cured PMMA in various artificial saliva and concluded all the artificial saliva had better wetting properties than distilled water.[9] The study by Farcasiu and Păuna compared the wettability of conventional heat-cured PMMA and injection-molded PMMA in natural and artificial saliva and concluded that injection-molded PMMA had the best wettability.[10] Zissis et al. verified the acrylic and nylon denture bases and concluded by stating that, the high-impact heat-polymerized PMMA denture base resin showed the best wettability with the least advancing and receding contact angle values.[11] He also mentioned that physical and mechanical properties would change when reinforcing materials were added. Hence, by adding silver nanoparticles to high-impact injection molding PMMA, there might be change in their wetting properties of the denture base resin.

The antifungal property of silver nanoparticles is highly appreciated in medical and dental fields. This can be used in denture bases to prevent the occurrence of denture stomatitis, which is considered because of the microbial biofilm adhesion on the porous denture material and accompanied by defective cleansing by saliva and poor tongue movements. The study by Aslanimehr et al. compared candidal adhesion in conventional heat-cured PMMA and injection-molded PMMA and concluded that there was significantly lesser adhesion of candida species in the surface of the denture in injection-molded PMMA due to less porous structure of the denture base resin.[12] Other ways to reduce candidal adhesion are the use of antifungal medications incorporated in denture base material to prevent the growth of C. albicans on the surface of the prostheses. The local drug delivery system directly delivers the drug at the site of infection decreases the risk of systemic side effects. A number of effective antifungal agents had been used, either topically or systemically, for the management of oral candidiasis. Amphotericin B, nystatin, various nanoparticles are common topical antifungal agents, whereas azoles such as fluconazole and ketoconazole are available for systemic antifungal treatment. The effect of conventional heat-cured denture base resin containing nano silver on C. albicans adhesion and biofilm formation had been reported by Wady et al. and concluded that silver nanoparticles had lesser candidal adhesion indicating its good antifungal property.[13] The study by Suganya et al. concluded a positive antifungal effect on silver nanoparticles reinforced conventional PMMA.[14]

The most important mechanical property of a denture base is flexural strength, and resistance to withstand masticatory forces. Reinforcers like nanoparticles change their spatial arrangement of molecules and get incorporated within the structure and improve its strength. The increased masticatory force beyond a certain limit leads to the formation of small cracks resulting in fracture of the prosthesis. High-impact injection-molded PMMA denture material has better mechanical properties than conventional heat-cured PMMA. Hamanaka et al. found that injection-molded thermoplastic denture base resins had better flexural strength compared to conventional heat cure PMMA.[15] Vallittu et al. concluded that higher flexural strength was seen for the injection-molding technique compared to the conventional method.[16] Nogueira et al. concluded that injection-molded PMMA had better dimensional accuracy compared to conventional heat cure PMMA.[17] Studies reinforcing injection-molded PMMA with fiber particles improved its properties. Various other reinforcing materials used were cyanoacrylate, metal wire, fibers, and woven glass. A study by Karacaer et al. compared injection-molded PMMA and convention heat-cured PMMA after reinforcing with E-glass fibers and concluded that reinforced injection-molded PMMA had better impact strength, transverse strength, and elastic modulus.[18] Flexural strength can also be increased in high-impact injection-molded PMMA by metal reinforcements like in conventional heat-cured PMMA.

Color stability of denture base reinforced with nanoparticles is critical for the aesthetics of long-term restorations and had been previously studied in vitro for a variety of denture base materials. While denture base materials are vulnerable to water sorption and solubility, they can absorb or loose soluble components in the liquids, depending on their composition of saliva and this is the reason why the degradation occurs in materials leading to discoloration.[19] Color changes in denture base materials may be due to exposure to oral fluids, beverages, and denture cleaners. Color stability of provisional restoration using PMMA and bis-acrylic-based materials was evaluated by Gujjari et al., and concluded that PMMA was more color stable than bis-acrylic composite-based resin using ultraviolet spectrophotometer.[20] The study by Goiato et al., evaluated color stability and flexural strength of ocular prosthesis after reinforcing with ZnO2, TiO2, and Ba2SO4.[21] They concluded that color of the prosthesis was affected by concentration and type of material reinforced. In their study, TiO2 had the best color stability and acceptable strength. Bohra et al. stated that conventional heat-cured resin had better stability than cold-cured resin.[22] The study by Shah et al. showed no color change in both nylon and acrylic PMMA denture base resin using conventional and injection molded techniques after immersion in denture cleansers.[23] In this study, silver nanoparticles were incorporated in various concentrations (0.05% and 0.2%) and evaluated for color stability on 0, 3, and 7 days. The procedure for evaluating color stability followed in this study was similar to the study by Gujjari et al.

Therefore, wettability, antifungal effect, flexural strength, and color stability were evaluated in this study after incorporating 0.05% and 0.2% silver nanoparticles in high-impact injection-molded PMMA and compared with the control group. The results showed increased wettability, positive antifungal effect, improved flexural strength, and negligible color changes in the denture base. Further scope of this study can be verified by clinical application and evaluation of the physical and mechanical properties for future use in the field of prosthodontics. Furthermore, varying the concentration of silver nanoparticles and the use of other nanoparticles may influence the results of the study.


  Conclusion Top


Within the limitations of the study following conclusions were made:

  • Subgroup C had slightly improved contact angle values than subgroup A and subgroup B throughout the storage periods, indicating that the incorporation of 0.2% of silver nanoparticles had significant changes in their wettability
  • The maximum zone of inhibition was seen in subgroup C with the mean value of 27.42, followed by subgroup B with a mean value of 22.67 and the least antifungal effect in subgroup A, indicating that 0.2% of silver nanoparticles improved their antifungal effect
  • High flexural strength was seen in subgroup C with the mean value of 130.0833, followed by subgroup B with the mean value of 127.0033 and the lowest flexural strength in subgroup A with the mean value of 120.085, indicating that reinforcement of silver nanoparticles with 0.2% showed a significant increase in flexural strength
  • No change in color was observed in all the groups during the storage periods. This indicates that silver nanoparticles in 0.05% and 0.2% concentrations do not influence the color of the samples
  • On comparing the various concentrations of Ag nanoparticles, subgroup C samples made with 0.2% Ag NP showed the highest wettability, antifungal effect, and flexural strength while the color stability remained almost unchanged, followed by subgroup B and subgroup A (control group) which had lesser values and least effective. Hence, it was concluded that high-impact injection-molded PMMA incorporated with 0.2% silver nanoparticles provided improved physical and mechanical properties in human saliva.


Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
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Aslanimehr M, Rezvani S, Mahmoudi A, Moosavi N. Comparison of Candida albicans adherence to conventional acrylic denture base materials and injection molding acrylic materials. J Dent (Shiraz) 2017;18:61-4.  Back to cited text no. 12
    
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Gujjari AK, Bhatnagar VM, Basavaraju RM. Colour stability and flexural strength of poly (methyl methacrylate) and bis-acrylic composite based provisional crown and bridge auto-polymerizing resins exposed to beverages and food dye: An in vitro study. Indian J Dent Res 2013;24:172-7.  Back to cited text no. 20
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[PUBMED]  [Full text]  


    Figures

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    Tables

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