The Effect of ZrO2 Nanoparticles Addition on Candida Adherence and Tensile Strength of 3D Printed Denture Base Resin

Document Type : Research Paper

Authors

1 Department of Prosthodontics, College of Dentistry, University of Baghdad, Baghdad 1417, Iraq,

2 Department of Prosthodontics, College of Dentistry, University of Baghdad, Baghdad 1417, Iraq.

10.22052/JNS.2023.02.024

Abstract

To enhance 3D printed denture base resin performance; ZrO2 nanoparticles were added to improve the biological and mechanical behavior. (110) specimens (50 dumbbell- shaped and 60 discs) were 3D printed and divided into five groups per test (n=10). The control group for each test included unreinforced 3Dprinted denture base resin, while the test groups reinforced with (1, 2, 3, and 4 %) nanoZrO2; with positive control of nystatin 1.4% for candida adherence test. Tensile strength was evaluated using universal testing machine while candida test was evaluated by spectrophotometer device through optical density verification. The study showed significant increase in antifungal activity of the 3Dprinted denture base resin after adding nano ZrO2 .The tensile strength mean was significantly higher than the control group; although the mean was decreasing with increasing the ZrO2 NPs.  The addition of ZrO2 nanoparticles increasing the antifungal activity of the 3D denture base resin, the increasing was proportional to the nanoparticles concentration. The tensile strength of the 3D denture base resin was significantly improved with 1% of ZrO2 NPs concentration among 2, 3 and 4%.

Keywords


INTRODUCTION

PMMA still the most friendly denture base material for many practitioners, in spite of its limitations of low mechanical and physical properties with the long process of fabrication [1]. Digital technology in dentistry as a whole and prosthodontics in particular has shown to have many benefits in terms of precision results of fabrication and speed of manufacturing [2-8]. However, there are still some issues that must be resolved, such as the poor mechanical properties of the base materials used in 3D printing dentures [2-5].

Despite being close to the ISO-accepted value of 65 MPa for flexural strength, 3D printed denture base materials have the lowest flexural strength and surface hardness compared to conventional and milled denture base materials. Thus, its clinical applications are constrained [5,6,8,9].

Numerous studies looked into many ways to overcome the aforementioned restrictions, modification of post-polymerization time, layer thickness, printing orientation, and the addition of nanoparticle fillers like TiO2, Al2O3, and SiO2 as metal oxide nanoparticles that appear to improve some mechanical properties of the 3D-printed denture base resin [10-13]. According to Gad et al. (2022) adding SiO2 NPs to 3D printed denture base resin improves flexural strength and impact strength without significantly affecting surface roughness [13]. Additionally, Alshaikh et al. (2022), stated that the 3D printed denture base resin was significantly increased in flexural strength, impact strength, and hardness with no appreciable changes in surface roughness after addition of ZrO2 NPs [14]. ZrO2 mimics the appearance of natural teeth and reduces peri-implant inflammatory reaction, which makes it more biocompatible than other ceramic materials like alumina. Having high strength, fracture toughness, and surface hardness, it is a biocompatible metal oxide [14-17]. Additionally, it exhibits thermal stability, corrosion resistance, antifungal and antibacterial activity against Candida albicans and Asergillus niger [17-19].E.coli and S. aureus  were used as model strains of gram-negative and gram-positive bacteria, respectively, in antibacterial activity tests of ZrO2 NPs. capable of effectively inhibiting the growth of bacterial cultures ,it was discovered that ZrO2 NPs were significantly more effective against S. aureus than E. coli, with S. aureus bacterial growth being inhibited by ZrO2 NPs to a greater than 90% degree. Scanning electron microscopy (SEM) analysis of the morphology of bacterial cells revealed that nanoparticles and nanocomposite permanently damaged the cell membrane [17,18].

ZrO2 NPs may work well as a 3D printing material reinforcement technique .To the best of the authors’ knowledge, no studies have previously examined the impact of adding ZrO2 NPs on the ability of 3D-printed resins on candida adherence and increase tensile strength [19].

Therefore, the aim of this study was to evaluate the effect of adding ZrO2 NPs to 3D-printed denture-base resins on candida adherence and on tensile strength.

 

MATERIALS AND METHODS

110 specimens were designed according to specification for each test, (60) disks of 2x10 mmfor candida test and (50) dumbbell shape specimen for tensile strength test with dimension given by (ASTM specification D-638M, 1986) (20), divided into 5 groups (n=10) according to the ZrO2 nanoparticles concentration (1, 2, 3, and4 %) by weight.

Optiprint laviva ( dentona , Germany) 3D printed denture base resin of light pink color was used with DLP open system microlay versus 385 dental printer by exporting the STL file from microform computer software program  .Pure resin was placed on mechanical mixer machine before adding the nanoparticles for 120 min; then addition of nanoparticles in mentioned concentrations and distributed into several bottles with continuous stirring in magnetic stirrer for 30 minutes at 60°C to decrease the resin viscosity, then stirred at room temperature for 8h to obtain homogenous nanocomposite for printing procedure [20]. Each layer was printed with a 50 µm layer thickness in (1.61) sec/slice in vertical Z axis following manufacturing instructions. Cleaning with isopropyl alcohol 99.9% before immersion in glycerol and placing in UV light polymerization unit for 10minutes to complete the polymerization prior to finishing the samples by removing the supports and base with low speed rotary instrument and polishing with polishing machine and cloth in a wet condition [21,22]. The whole procedure was done by one operator to insure applying same preparation conditions .The specimens immersed in distilled water 48hs at 37°C prior to testing [23].

 

Testing procedure

Candida test: sterile disks were incubated with a candida culture for 24 hours before being removed, washing with normal saline to remove any remaining candida, staining with crystal violet for 20 minutes, rewashing with normal saline, and then immersing in 3 ml of ethanol alcohol (96%) for three minutes [24-27]. The optical density was then confirmed. The tensile strength: Each specimen’s tensile strength had been evaluated using a universal testing machine. The ends of the material specimen are typically clamped on two jigs spaced apart by a specific amount, stretching the specimen as the two jigs separate until there is damage to the specimen.

Tensile strength was calculated by formula: T.S.MPa=Maximum force (N.)\ Area (mm)

 

RESULTS AND DISCUSSION

Evaluating the adherence ability of Candida albicans to 3D printed denture base resin after ZrO2 addition by OD verification, mean and standard deviation with confidence interval in Table 1, as shown; the minimum antifungal activity of 3D printed denture base resin after adding 1% nano ZrO2, and maximum value was with 4% nano ZrO2 at 95% confidence interval. Boxplot to describe the SD and median between minimum and maximum range of candida adherence test Fig. 1.

According to test of homogeneity of variance (Levene test) Table 2 and test of ANOVA Table (3) a highly significant differences (p ≤0.01) demonstrated between study groups and control group at a significant level of (0.01%).

According to the significant results, comparison between each 2 groups was decided to be evaluated by Games-Howell test. Post hoc test (Games–Howell) was selected for multiple comparisons of incorporation to compare the mean values among all study groups in Table 4.

The same for tensile test as mean and standard deviation was conducted with confidence interval at 95% in Table (5) and demonstrated in Fig. 2. The variances of tested groups for tensile strength were analyzed by Levene’s test of homogeneity in Table 6 to decide the test of multiple comparisons of the results. Comparison of means for tensile test results of the experimental groups using ANOVA in Table 7 and the result was highly significant.

With this significant result, Games-Howell test (Table 8) was selected to compare between the mean values among all study groups. SEM images of the samples surfaces at 100,000× magnification force revealed significant difference between the pure 3Dprinted resin(A) (with no addition ) that appears to have broad scattered pores with irregularity compared to the 2% (C) and 3% (D) nano ZrO2 ;while the images of the 3D resin with 2% and 3% shows the dispersion of nanoparticles within the material  to give more compact and regular surface with more diminished pores and particle size of less than 50 µm of ZrO2 NPs in (C) than (D) and this explains the ductility of the group (D) which gives the result of tensile strength Fig. 3.

Fig. 4 also shows significant differences in the surface of the pure 3Dprinted denture base resin (A) and the 2% (B),3%  (C) ZrO2 NPs at 4000 magnification force of SEM to prove the chemical reaction between the resin and the nanoparticles which was supported by the FTIR readings in Fig. 5, both (B) and (C) showed homogenous and good distribution of nanoparticles within the resin matrix with some clusters may be shown at 3% nano ZrO2.

The FTIR results showing significant difference between the pure 3D printed resin (0%), 2% nano ZrO2 3D resin and 3% nano ZrO2 3D resin especially between  ̴ 806- 636 cm̵1 range of spectra which indicate the presence of ZrO2 within the polymer of the 3D printed denture base resin ,differences between peaks of 2% and 3% ZrO2 resin as appeared at ̴ 752 cm-1 suggests the chemical reaction between the polymer resin and the nanoparticles, as the most intense peak of band for 2% NPs at ̴ 690 cm ̵ 1, while for 3% NPs at ̴ 694 cm ̵1, with similarity to some extent between the spectra of the pure 3D resin and the reinforced resin attributed to the vibration and stretching of CH3  and CH2 groups at ̴ 1716-1381 cm ̵1 bands with vibration of ester group C=O at  ̴1180-1149 cm ̵1, and this confirm the homogenous dispersion of the nanoparticles within the 3D printed resin material.

The effect of ZrO2 NPs addition on the properties of 3D printed denture base resin was testing in this study regarding antifungal activity and tensile strength; according to the results, the null hypothesis was rejected because the addition of ZrO2 NPs significantly affect the Candida albicans adherence and tensile strength. The present study showed an increase in antifungal activity of 3D printed denture base resin when ZrO2 NPs were added. DS is a condition linked to Candida albicans that frequently returns in people who wear complete dentures. An important step in the colonization and pathogenesis that results in DS is C. albicans’ adherence to the intaglio surface of a denture base [29]. It was claimed that mechanical cleaning techniques fall short of completely eliminating bacteria from denture surfaces, as a result, numerous attempts have been made to use a range of antifungal drugs to minimize C. albicans adherence and subsequent colonization on the denture base, but these treatments have shown to be ineffective and for short term [31]. Additionally, a variety of methods have been used to prevent fungal attachment to denture bases, in