Physical Properties of PMMA Denture Base with Added Organoselenium as an Antifungal

Abstract

Background/Objectives: The present study investigated the effects on the mechanical and physical properties of PMMA when organoselenium was incorporated into it as antifungal at different concentrations. Methods: 141 PMMA rectangle samples were fabricated using a heat compression packing technique and assigned to 3 experimental groups (47/group): 0% organoselenium (control), 0.5% organoselenium (0.5% SE), and 1% organoselenium (1% SE). Each sample was post-processed and stored in water. A three-point bend test was performed to assess elastic modulus and flexural stress. Scanning electron microscopy (SEM) was used to examine the exterior and interior surface topography. Data were analyzed using one-way ANOVA with Tukey’s multiple comparisons test. Results: The mean flexure stress for the 0% samples was statistically significantly higher than those of the 0.5% samples and the 1% samples (p < 0.001). The mean elastic modulus for the 0% group was statistically significantly higher than those of the 0.5% group and the 1% group (p < 0.001). Under SEM, the 0.5% samples were smoother with fewer voids and irregularities. Conclusions: The incorporation of organoselenium into PMMA denture base negatively affected its physical and mechanical properties.

1. Introduction

Denture stomatitis, a common inflammatory condition that affects the mucosa in contact with the intaglio surface of the complete dentures, has been found in up to 15 to 70% of denture wearers [1,2,3,4]. It is characterized by an inflammatory and erythematous mucosal response of the denture-bearing areas [5,6], with symptoms such as pain, itching, or a burning sensation [3]. This multifactorial condition results from factors such as poor denture hygiene, poor fitness, or continuous wearing of the denture resulting in denture plaque accumulation, with Candida albicans been the predominant microbial agent associated with it [7].

To prevent denture stomatitis, previous studies have explored either coating the denture surface with various antifungal agents or incorporating them into the denture base [4,8,9,10,11,12,13,14,15,16,17,18,19]. Recently, a study found that the addition of 0.5% organoselenium (SelenBio Inc., Austin, TX, USA) into polymethylmethacrylate (PMMA) denture base inhibited C. albicans biofilm formation and growth [20]. Organoselenium is a biocidal agent that functions through the catalytic generation of superoxide radicals because of the oxidation of thiols [21,22,23,24,25]. These radicals have been shown to inhibit microbial biofilms such as Staphylococcus aureus, and recently C. albicans [20,21,22,23,24,25]. These biofilms play a significant role in the development of denture stomatitis. Our previous publication detailed the source, the characteristics, and the mechanisms of action of the organoselenium used in the present study [23]. However, it is pertinent to mention that the use of antifungal oral hygiene products or antifungals prescriptions that inhibit the growth of the fungi have been successful in the past, but patient compliance is necessary [26,27]. Thus, an agent that could be incorporated into the PMMA denture base that inhibits the growth of C. albicans, is desirable. A study completed in 2021, by Al Mojel et al., found that the incorporation and application of 0.5% and 1% organoselenium in and on heat-polymerized PMMA disks inhibited the growth of C. albicans biofilm [20]. Research studies have shown that incorporating certain compounds into PMMA denture bases can provide promising antimicrobial properties [4,8,9,13,16,17,20,25].

However, denture bases should exhibit certain material properties, such as biocompatibility (nontoxic or irritating), adequate physical properties (low solubility, low sorption of oral fluids, good thermal conductivity, and high abrasion, creep and craze resistance), and adequate mechanical properties (high flexural, transverse, and impact strength, high modulus of elasticity, long fatigue life, and low density). For this reason, care must be taken if any alterations or additions are made to denture base materials, such as PMMA, to maintain its successful capabilities [28]. If a biofilm-inhibiting denture base that retains its physical properties can be formulated, denture stomatitis could be drastically reduced in patients. However, the effect of adding organoselenium on the mechanical and physical properties of PMMA remains to be studied.

Since organoselenium has been proven to be a useful antifungal and has been shown to successfully inhibit the development and growth of C. albicans biofilm, this study aims to elucidate its effects on the physical properties of PMMA denture base material. If the antifungal works but the denture base becomes more prone to fracture or there are alterations in its structural capabilities, then the usefulness of organoselenium becomes clinically insignificant. The null hypothesis states that incorporating organoselenium into the PMMA denture base would negatively impact its physical and mechanical properties.

2. Materials and Methods

2.1. Samples Production and Experimental Grouping

PMMA samples were fabricated following ANSI/ADA specification No. 12-2002 for denture base polymers [29]. Using baseplate wax 141 (Mizzy Allcezon™; Keystone Industries, Gibbstown, NJ, USA), samples with dimensions, 10 mm width, 22 mm length, and 2 mm thickness, were created. The wax samples were flasked and invested using type III dental stone (Labstone™; Garreco Dental Products, Heber Springs, AR, USA). The wax was boiled out, leaving a mold within the flasked dental stone. A thin layer of separating medium was applied to the stone and allowed to dry. Heat-polymerized acrylic resin (Lucitone 199 powder; Dentsply Sirona, Charlotte, NC, USA) and monomer (Lucitone liquid^®; Dentsply Sirona, Charlotte, NC, USA) were mixed according to the manufacturer’s instructions. Prior to mixing with the powder, 0% (control), 0.5%, and 1.0% concentrations of organoselenium (Denteshield^®; SelenBio Inc., Austin, TX, USA) were respectively incorporated into the monomer to produce the three study groups. The organoselenium, obtained in liquid form from SelenBio Inc., Austin, TX, USA, was incorporated into the monomer by gradual continuous vigorous stirring until uniform dispersion of the organoselenium was achieved. Acrylic was packed into the mold cavity after reaching the doughy stage, followed by trial packing. After packing, processing was completed according to the manufacturer’s instructions, 90 min in water at 73 °C, followed by 30 min in boiling water at 100 °C. After processing, the edges of each sample were trimmed to remove excess acrylic, and then the samples were sonicated in an ultrasonic bath (Elmasonic P30H; Tovatech, Plano, TX, USA) with water to remove any stone debris. Following this, the samples were stored in room temperature water to avoid desiccation until used.

Following fabrication, the samples were randomly assigned into three groups, 0%, 0.5%, and 1% organoselenium, with 47 samples in each. Within each group, 22 samples were used to measure flexural stress and elastic modulus, 22 samples for microhardness, and 3 samples for scanning electron microscopic (SEM) examination.

2.2. Flexural Stress and Elastic Modulus

The flexural stress and elastic modulus of the samples were performed according to the protocols reported in previous studies [8,12]. An Instron machine (ASTM D790, Bruker’s UMT TriboLab^®, Bruker Corporation, Billerica, MA, USA) was used to perform the 3-point bend test. Twenty-two samples from each group (0%, 0.5%, 1.0% organoselenium) were used. Each sample was positioned 20 mm apart over the supports, and a 196 N load was applied progressively with the mode set at 1 mm/min crosshead speed until fracture or dismounting from the supports [8]. The flexural stress (MPa) as well as the elastic modulus (MPa) were automatically generated and recorded.

2.3. Microhardness Test

The Vickers hardness (VH) tester (Shimadzu HMV-G31DT; Shimadzu Scientific Instruments, Columbia, MD, USA) was used for the evaluation VH of the specimens as described in a previous study [30]. Each of the 22 samples from one group (0%, 0.5%, 1.0% organoselenium) was secured and placed on the microhardness tester. Three indentations were made on each sample at different sites along the diametrical line (center and left and right ends) using a Vicker’s diamond under a 50 g load applied for 15 s. Then the dimensions of each indentation were measured using the HMV-G software for HMV-G31 series (Shimadzu Scientific Instruments, Columbia, MD, USA) and averaged for each sample [30,31].

2.4. Scanning Electron Microscope Examination

Three samples from each group (0%, 0.5%, 1.0% organoselenium) were selected for this examination. A cross-section of each sample was produced to view the internal layers of the PMMA denture material. This sectioning produced two pieces of each sample. One piece was used to examine the smooth surface of the sample, while the other piece was used to examine the internal structure of the sample. The sectioned samples were then sonicated to remove any cutting debris. The samples were dried and mounted for SEM examination according to previously published procedures [32,33]. Each sample was gold-sputter-coated. The mounted samples were placed on the SEM platform, scanned, and images were captured (JCM-5700; JEOL Ltd., Peabody, MA, USA) for each sample under a magnification of ×1000. Three independent evaluators with experience in dental materials were asked to qualitatively assess porosity and surface features in the SEM images and record their findings.

2.5. Sample Size and Power Calculation

A power analysis was conducted to determine the sample size using G*Power statistical software 3.1.9.7 for Windows (Heinrich-Heine-Universität, Düsseldorf, North Rhine-Westphalia, Germany) [34]. Based on data reported in previous studies [8,11,12,14,15,30], an effect size of 0.40 (large), a 95% confidence level (α = 0.05), and 80% statistical power (1 − β = 0.80) were used as criteria to detect a significant difference between the control, 0.5%, and 1% organoselenium concentration. With these criteria, a sample size of 22 for each subgroup (0%, 0.5%, 1.0% organoselenium) was needed for the outcome measurement. However, 25 samples were used for each group to account for damage during processing.

2.6. Statistical Analysis

All data analyses were performed using Statistical Package of Social Sciences (SPSS version 23, Chicago Inc., Chicago, IL, USA). The statistical significance for each analysis was established at an alpha level of α = 0.05. Preliminary analyses were conducted to explore the dataset and to check for assumptions violations. The normality assumption was tested using the histogram, Q-Q plot and the Shapiro–Wilk test from the tests of normality table and all confirmed that the normality assumption was met for each variable at the alpha level of α = 0.05. Since more than two groups were being compared for the flexural stress, elastic modulus, and SMH, a one-way ANOVA was first used for this purpose. This was followed by a post hoc comparison using the Tukey HSD test to determine specific group means that significantly differed from each other.

3. Results

Although measures were taken to maintain the exact sizes for each sample, due to the nature of the heat compression molding technique and material, there were slight variations in sample sizes. There was a statistically significant difference [F (2, 68) = 1074.48, p < 0.001] in flexural stress among the three groups (Figure 1 and

Table 1. Descriptives statistics of the Flexural Stress of all groups.

	N	Mean ± Std. Deviation	95% Confidence Interval for Mean		Minimum	Maximum
			Lower Bound	Upper Bound
0%	25	90.9 ± 6.9	88.1	93.7	76.9	104.1
0.5%	24	34.7 ± 3.9	33.0	36.4	25.5	41.1
1%	22	35.4 ± 2.3	34.4	36.5	31.2	39.2

and

Table 2. Multiple Comparisons of the Flexural Stress of the groups (Tukey HSD).

Groups		Mean Difference (I–J)	Std. Error	Sig.	95% Confidence Interval
(I)	(J)				Lower Bound	Upper Bound
0%	0.5%	56.18872 *	1.38549	<0.001	52.8690	59.5085
	1%	55.46035 *	1.41725	<0.001	52.0645	58.8562
0.5%	0%	−56.18872 *	1.38549	<0.001	−59.5085	−52.8690
	1%	−0.72837	1.43101	0.867	−4.1572	2.7004
1%	0%	−55.46035 *	1.41725	<0.001	−58.8562	−52.0645
	0.5%	0.72837	1.43101	0.867	−2.7004	4.1572

* The mean difference is significant at the 0.05 level.

). Post hoc comparisons using the Tukey HSD test indicated that the mean flexural stress for the 0% group (90.89 ± 6.86) was statistically significantly (p < 0.001) higher than the 0.5% group (34.70 ± 3.93) and the 1% group (35.43 ± 2.33). The 0.5% and 1% groups were not statistically significantly different (Table 1 and Table 2).

There was a statistically significant difference in elastic modulus among the groups [F (2, 68) = 241.97, p < 0.001]. Post hoc comparisons using the Tukey HSD test showed that the mean elastic modulus for the 0% group (1801.78 ± 213.41) was statistically significantly (p < 0.001) higher than the 0.5% group (822.35 ± 151.50) and the 1% group (930.29 ± 130.37), as shown in Figure 2 and

Table 3. Descriptives statistics of the Elastic Modulus of all groups.

	N	Mean ± Std. Deviation	95% Confidence Interval for Mean		Minimum	Maximum
			Lower Bound	Upper Bound
0%	25	1801.8 ± 213.4	1713.7	1889.9	1356.2	2322.4
0.5%	24	822.4 ± 151.5	758.4	886.4	415.8	1097.8
1%	22	930.3 ± 130.4	872.5	988.1	696.8	1113.2

and

Table 4. Multiple Comparisons of the Elastic Modulus of the groups (Tukey HSD).

Groups		Mean Difference (I–J)	Std. Error	Sig.	95% Confidence Interval
(I)	(J)				Lower Bound	Upper Bound
0%	0.5%	979.42788 *	48.73897	<0.001	862.6452	1096.2106
	1%	871.48989 *	49.85646	<0.001	752.0296	990.9502
0.5%	0%	−979.42788 *	48.73897	<0.001	−1096.2106	−862.6452
	1%	−107.93799	50.34031	0.089	−228.5576	12.6817
1%	0%	−871.48989 *	49.85646	<0.001	−990.9502	−752.0296
	0.5%	107.93799	50.34031	0.089	−12.6817	228.5576

* The mean difference is significant at the 0.05 level.

. The 0.5% and 1% groups were not statistically significantly different.

A one-way between-subjects ANOVA was conducted to determine whether surface microhardness (SMH) significantly differed among the groups (

Table 5. Descriptives statistics of the Vicker’s hardness number (VHN) of all groups.

	N	Mean ± Std. Deviation	95% Confidence Interval for Mean		Minimum	Maximum
			Lower Bound	Upper Bound
0%	21	31.3 ± 13.3	25.2254	37.3460	17.30	66.30
0.5%	21	28.5 ± 11.7	23.2145	33.8236	12.70	49.30
1%	21	20.9 ± 9.5	16.5305	25.1743	10.20	46.60

and

Table 6. Multiple Comparisons of the Vicker’s hardness number (VHN) of the groups (Tukey HSD).

Groups		Mean Difference (I–J)	Std. Error	Sig.	95% Confidence Interval
(I)	(J)				Lower Bound	Upper Bound
0%	0.5%	2.76667	3.57772	0.721	−5.8314	11.3647
	1%	10.43333 *	3.57772	0.014	1.8353	19.0314
0.5%	0%	−2.76667	3.57772	0.721	−11.3647	5.8314
	1%	7.66667	3.57772	0.090	−0.9314	16.2647
1%	0%	−10.43333 *	3.57772	0.014	−19.0314	−1.8353
	0.5%	−7.66667	3.57772	0.090	−16.2647	0.9314

* The mean difference is significant at the 0.05 level.

). The result showed a statistically significant difference in SMH among the groups [F (2, 60) = 4.57, p = 0.014]. Post hoc comparisons using the Tukey HSD test revealed that the mean SMH for the 0% group (31.29 ± 13.31) was significantly higher than that of the 1% group (20.85 ± 9.49), but there was no significant difference between the 0% and 0.5% (28.52 ± 11.65) groups at the α level of 0.05 as shown in Figure 3 (Table 5 and Table 6). There was no significant difference between 0.5% and 1% groups. The association between SMH and modulus or flexural stress was not significant in all groups (0%, 0.5%, 1%).

When samples were examined with SEM, as seen in Figure 4 and Figure 5, there were structural differences observed in each group. Examination of the SEM micrographs of the smooth surfaces (Figure 4) showed that while samples in all groups appeared layered with different sizes of islands and elevations, the 0.5% appeared more melted together and more compact when compared to the other two groups. When reviewing the cross-sectional surface, the group that had more voids, porosities, and irregularities was the 1% samples. 0.5% had fewer irregularities, were smoother, and appeared to have fewer voids and porosities.

4. Discussion

A denture base that has antimicrobial properties would be an outstanding achievement, but it is essential that its physical and mechanical properties remain unaffected [8]. The results from previous studies have shown that it is possible to incorporate organoselenium into denture base material to inhibit the growth of Candida [20,25]. If we could help patients by incorporating organoselenium into their denture base and, in turn, decrease their possibility of experiencing denture stomatitis without adding to their responsibilities or taking away from the success of their denture, it would be ideal. Furthermore, organoselenium that is incorporated into materials does not leach out, thus no toxicity issue. This has resulted in an increased application of selenium and selenium-containing compounds in dentistry, orthopedics, and ophthalmology [20,21,22,23,24,25]. However, the effect that the incorporation of organoselenium into denture base would have on the physical properties of the denture base has not been investigated. For these reasons, the present study investigated the mechanical and physical properties of PMMA after the incorporation of organoselenium. ANSI/ADA Specification No. 12 [29] described the requirements and specifications for denture base polymers. Since the oral cavity can be a harsh environment for materials that need to withstand changing temperatures, acidities, and textures of food, there is need for a standard. The material also must be biocompatible and non-toxic.

The current study found that the addition of organoselenium had a statistically significant effect on the mechanical properties of PMMA, specifically in terms of flexural stress and elastic modulus, compared to samples that did not contain organoselenium. This negative impact raises concerns about its potential applications in PMMA. The addition of organoselenium lowered the flexural stress and elastic modulus in comparison with samples without organoselenium. Therefore, the null hypothesis of the present study was accepted. Flexural stress is when an outside source applies a force and causes a material to bend or deform, whereas elastic modulus is a material’s resistance to being deformed [8]. Both properties require that denture bases need to function well in the oral cavity, withstanding the chewing forces, parafunctional habits, and abrasive forces. The study group without organoselenium (0% concentration) displayed superior flexural stress and elastic modulus resistance under force application, suggesting that the PMMA demonstrated enhanced properties in the absence of organoselenium.

Considering the established fact that organoselenium interacts with the denture base resin through covalent bonding, which is a strong bond [21], one may wonder why it would contribute to such a dramatic reduction in mechanical properties of the denture base. However, it is pertinent to mention that whereas the setting reaction of the denture base material utilized in this study primarily follows a free radical polymerization mechanism, previous investigations into the integration of nanoparticles into denture base materials have indicated that the mechanical properties of PMMA denture material are significantly affected by their interactions with the polymeric matrix [30,35]. The covalent bonding and free radical polymerization interaction may be a contributing factor to the differences observed in flexural strength and elastic modulus when organoselenium is introduced. Additionally, critical factors such as particle concentration, morphology, and size significantly influence these mechanical properties [35]. In the study conducted by Balos et al., enhanced particle distribution within PMMA denture material was correlated with an increase in elastic modulus [36]. In contrast, the possible insufficient particle dispersion in the present study may have resulted in a decrease in the elastic modulus. Nevertheless, the covalent bonding of organoselenium with denture base resin prevents its leaching out of the denture base, thus it does not cause tissue toxicity, and it is retained within the denture base throughout the life of the denture base [21].

A study by Castro et al. revealed that flexural strength in PMMA resins containing silver nanoparticles was diminished due to poor dispersion during manual incorporation of the material [37]. Moreover, it has been documented in previous studies that the addition of specific monomers to denture base materials can compromise flexural strength, primarily due to the formation of clusters that function as impurities, thereby creating zones of stress concentration. This issue is again associated with manual mixing techniques [38,39]. The present study utilized manual incorporation methods, which may have resulted in the reduction in flexural strength noted with increasing concentrations of organoselenium. As a result, exploring alternative mixing techniques may prove advantageous for enhancing the integration of organoselenium into the polymer matrix.

The SMH was significantly higher in the group without organoselenium compared to those containing 1% organoselenium. However, there was no significant difference observed between the 0% and 0.5% samples. In a separate investigation by AlZayyat et al., it was reported that higher concentrations of SiO₂ incorporated into denture base material resulted in increased voids and porosity, which subsequently reduced its flexural strength [35]. Cross-sectional analysis in the current study revealed that the group with the highest concentration of organoselenium exhibited a greater number of voids, porosities, and surface irregularities, while the group devoid of organoselenium displayed the least. Consequently, the increases in voids and porosity associated with organoselenium incorporation likely contributed to the observed decrease in flexural strength and SMH.

In terms of physical properties as examined with SEM (Figure 4), the group with 0.5% organoselenium appeared more melted together and more compact when compared to the other two groups, which could be interpreted as smoother surface, thus potentially creating a less conducive environment for the proliferation of Candida albicans. Although the 0% group appeared less porous than the 0.5% samples, its surface texture was comparatively less smooth. These observations can be attributed to the established facts that the appearance of PMMA under SEM varies significantly depending on several factors such as its form, method of preparation, additives, and impurities, mainly because it is a non-conductive polymer that is sensitive to the electron beam [32,33,40]. Properly prepared and developed PMMA thin films generally appear smooth, although some surface roughness can be detected with higher magnification [32]. Nevertheless, the present report evaluated some mechanical properties of PMMA denture base with added Organoselenium. In the future, further studies could be conducted testing other materials such as fiber reinforced composites [41] and PEEK [42].

However, some limitations in the design of the current study were (a) maintaining a uniform sample size that can better account for the volumetric and linear shrinkage of PMMA, and (b) only three properties were investigated. As organoselenium is becoming increasingly used in dentistry, more studies are needed on incorporating it into denture material if it is to be used as an antimicrobial and still maintain physical and mechanical properties.

5. Conclusions

Within the limit of the present study, incorporation of organoselenium into polymethyl methacrylate (PMMA) denture base materials may affect their physical and mechanical properties.