Introduction

The evolving field of prosthodontics is synonymous with the constant search for superior materials that not only mimic natural teeth in form and function but also resist microbial colonization and biofilm forming. Heat-cured acrylic resins have been widely employed in the field of dentistry, especially in the fabrication of implant-supported hybrid prostheses and complete dentures. Specifically, heat-cured acrylic resins, particularly the Procryla and Imicryl brands, remain the benchmark for prosthetic base materials due to their aesthetics, ease of manipulation, and compatibility (Shim et al. 2020). However, bacterial adhesion remains a concern, possibly leading to peri-implantitis, prosthetic stomatitis, and other biofilm-related complications (Paranhos et al. 2007). As the American Academy of Implant Dentistry (AAID) reports, the prevalence of dental implants in the USA is substantial, with 3 million citizens possessing implants and an additional 500,000 joining this cohort annually (de Avila et al. 2020). This widespread adoption underscores the importance of addressing bacterial adhesion, as even with a high success rate of up to 98%, the industry faces a 20% rate of infection-related implant loss within 5–10 years post-placement (de Avila et al. 2020). While oral bacteria have the capability to bind to diverse surfaces through various adhesive mechanisms, the material properties can significantly dictate the extent and type of bacterial colonization. Hence, understanding and optimizing these material properties are crucial in modern prosthetic and implant solutions (de Avila et al. 2020).

Different denture cleaning products, antifungal medications, and the addition of antifungal agents to denture base materials are some of the therapeutic modalities that have been developed to stop fungal growth on denture bases. But employing denture cleaning solutions might cause the base material of the denture to deteriorate and provide a rougher surface, which makes the denture more vulnerable to Candida adherence. Also, the dilution impact of saliva, the physical cleaning action of the oral muscles, and the dependence on patient compliance are major causes of therapeutic failure of antifungal drugs (Karci et al. 2019).

The incorporation of antimicrobial agents, particularly zirconia nanoparticles, into heat-polymerisable acrylic resins has been proposed as a potential game changer. The natural properties of zirconia, when grafted into acrylic resins, may act as a deterrent against bacterial adhesion and thus prolong the life of prostheses (Puri et al. 2008; Ajay et al. 2019).

The growing consumption of vegan beverages, such as almond milk and water kefir, presents potential implications for dental materials due to their lower pH. Although the effects of acidic beverages on prosthetic surfaces are well-established, the influence of plant-based alternatives on bacterial adhesion is insufficiently examined (Sumner and Burbridge 2020). Exploring how these beverages interact with zirconia-modified resins could provide critical insights for enhancing the durability and performance of dental prosthetics in contemporary dietary contexts.

The shift in consumers’ dietary preferences in recent years may potentially impact dental health (Sumner and Burbridge 2020). On the other hand, the growing trend of veganism has brought various beverages such as almond milk and water kefir to the forefront. These are not only consumed but also occasionally interact with prosthetic surfaces. The implications of this frequent interaction, especially with regard to bacterial adherence, are still relatively unexplored territory (Saeed et al. 2020; Gad et al. 2022). Such beverages, especially when combined with antimicrobial agents such as zirconia, can modulate bacterial adhesion by altering the surface properties of acrylic resins.

This study seeks to investigate bacterial adhesion on heat-cured acrylic resins enhanced with zirconia nanoparticles in the context of their exposure to various vegan beverages. Employing SEM and FTIR analyses over a defined period, the research aims to elucidate the intricate microbial interactions and possible variations in adhesion patterns on the modified resin surfaces. The null hypothesis posits that no significant differences in bacterial adhesion will be observed across the different zirconia-modified resin groups or among the vegan beverages tested.

Materials and methods

In this research, the bacterial adhesion on two brands of heat-cured acrylic resins, Imicryl (Imident, Konya, Turkey) and Procryla (President Dental GmbH, Munchen, Germany), was examined, with a particular focus on the modified versions of Procryla, which incorporated 1 wt% and 3 wt% zirconia (Aconia, Chengdu Besmile Biotechnology Co., Chengdu, Sichuan, China) nanoparticles. The study comprised 192 specimens, categorized into groups of unmodified Procryla (n = 48), Procryla 1Z (1% Zirconia) (n = 48), Procryla 3Z (3% Zirconia) (n = 48), and Imicryl (n = 48). The specimens were fabricated using silicone molds to create wax templates of 10 × 2 mm dimensions. These templates were embedded in plaster within flasks, where the wax was subsequently melted and removed. The cavities formed were filled with the respective acrylic resin mixtures as per manufacturer specifications. The flasks were pressed and cured in a pressure pot at 100 °C for 30 min, allowing for the polymerization of the acrylic resins adopted according to the manufacturer’s protocol. For the zirconia-modified specimens, the required amounts of zirconia nanoparticles (1 wt% and 3 wt%) were first accurately weighed using a precision scale and manually integrated into the polymer component. Subsequently, the liquid monomer was incorporated, and the entire mixture was thoroughly blended to achieve a uniform dispersion of the nanoparticles, thereby ensuring the correct zirconia concentrations prior to molding and curing. Notably, the Imicryl group was not modified with zirconia nanoparticles; this unmodified group, together with the unmodified Procryla, was included to serve as a control. This design enabled a direct comparison between modified and unmodified specimens within the same resin category, as well as a comparative evaluation between the two acrylic resin brands.

Post-curing, all specimens were polished with 600, 800, 1000, and 1200 grit sandpapers to achieve a smooth surface. The specimens were then individually immersed in 50 mL of one of four solutions—distilled water, mineral water (Beypazari, Beypazari Karakoca Dogal Maden Suyu Company, Ankara, Turkey), almond milk, and water kefir (Bir Sey, Bir Sey Gida Imalat Ltd. Sti., Antalya, Turkey). To prevent spoilage of the almond milk and water kefir and to maintain consistent experimental conditions, all specimens were stored at + 4 °C, with the immersion medium replaced daily. The specimens were immersed for the time point of initial, 1 day, and 14 days. This immersion aimed to simulate different oral environmental conditions. In a previous in vitro study, it was reported that 1 day of immersion corresponds to 1 month of in vivo exposure. In our study, the immersion periods of 1, 7, and 14 days were selected to simulate approximately 1 month, 7 months, and 1 year of clinical exposure, respectively (Aboushahba et al. 2020). In the study, specimens were grouped according to additive ratio and vegan beverage names. These are shown in Table S1.

The study’s primary goal was to quantify bacterial colonization on each specimen’s surface at the end of the immersion periods. Additionally, morphological and surface properties of chemical interactions of bacterial adhesion were examined by instrumental analysis techniques using scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR) methods. Zeiss brand EVO50 model SEM device (Carl Zeiss AG, Oberkochen, Germany) and SHIMADZU IR TRACER 100 brand model FTIR device (Shimadzu Corporation, Kyoto, Japan) were used for these analyzes. This research offers significant contributions to understanding the interplay between dental material composition, environmental factors, and bacterial colonization, which is crucial for advancing dental material technologies and improving oral health outcomes.

Supply and preparation of beverages

Distilled water

Prior to usage, distilled water was subjected to the process of boiling and subsequent cooling.

Mineral water

The acquisition of mineral water was made from the Burdur market (Beypazari, Beypazari Karakoca Dogal Maden Suyu Company).

Almond milk

Almond milk was produced using unprocessed almonds. During the initial phase, the almonds underwent a soaking process in water for a duration of 24 h. Subsequently, the shells were removed and the interior almonds were processed using a blender. The almond extract obtained was further filtered using a cloth to yield almond milk. Almond milk was prepared daily.

Water kefir

The sugar solution with a concentration of 4% was subjected to boiling at a temperature of 90 °C for 3 min, followed by subsequent cooling to a temperature of 25 °C. Following the introduction of water kefir grains, constituting approximately 4% of the total volume, into a solution of sugar water, the mixture was subjected to a fermentation process lasting 24 h, with the ambient temperature maintained at 25 °C. The water kefir grains were extracted using a sterile plastic strainer, and subsequently, the resulting beverage was put into a sterile jar. Water kefir was prepared daily.

Total aerobic mesophilic bacteria count

Following the washing of each specimen with sterile clean water, specimens were extracted from the upper layer via the swap procedure and subsequently transferred into tubes containing 9 mL of sterile peptone water. Subsequently, dilution specimens were meticulously produced to ensure a colony count ranging from 30 to 300, hence allowing an accurate assessment of the overall quantity of aerobic bacteria by the utilization of the bulk culture technique with Plate Count Agar. The specimens underwent incubation at a temperature of 37 °C for a duration of 3 days, and calculations have been performed with the common logarithm (Shimizu et al. 2005).

SEM analysis

After all the specimens obtained were washed and dried from the solutions, SEM analysis was performed by coating gold with a thickness of 5 nm. All images were obtained at 1000 × magnification so that they could be interpreted under the same conditions.

FTIR analysis

After all the specimens obtained were washed and dried from the solutions, FTIR analysis was performed between 400 and 4500 nm wavelength.

Statistic analysis

Data analysis was conducted with the IBM SPSS Statistics software, version 24.0 (IBM Corp.). Repeated measures ANOVA was performed to analyze the effects of groups, vegan beverages, and time followed by Tukey’s multiple comparison test to compare the bacterial adhesion of all the groups and beverages at different time points (α = 0.001). Repeated measures ANOVA and Tukey’s tests were performed to compare different CFUs/mL groups’ results.

Results

The repeated measures ANOVA was performed using IBM SPSS Statistics for Windows, Version 24.0 (IBM Corp., Armonk, NY, USA) indicated a significant difference for the interactions of heat-cured denture base resin groups, vegan beverages, times, and bacterial adhesion (P < 0.001, F: 1276,013) (Table S2).

Statistical analysis revealed that the difference between the number of bacterial colonies growing on the surfaces of heat-cured acrylic denture base resin specimens with different concentrations of zirconia (1 wt% and 3 wt%) and control discs without added zirconia was statistically significant (P < 0.001). Initially, in the distilled water immersion, the bacterial adhesion was lowest in the Procryla1Z group (3.05 ± 0.03) (P < 0.001). As the percentage of zirconia increased, the number of colonies increased (3.16 ± 0.03 for Procryla3Z) (P < 0.001). In the specimen groups kept in mineral water beverage for one day, the bacterial adhesion of the Imicryl group (3.94 ± 0.04) was found to be low, while in the specimen groups kept in mineral water after 14th day, the colony number of the Procryla3Z group was found to be the lowest (3.30 ± 0.01) (P < 0.001). In the specimen groups kept in almond milk, the lowest colony number was observed in the groups without zirconia. On day 1, the number of colonies of the Imicryl group was 5.11 ± 0.2 in the specimens kept in almond milk, while the number of colonies of the Procryla group kept in almond milk for 14 days was 5.00 ± 0.02 and the lowest (P < 0.001). Similarly, in the specimen groups kept in water kefir for 1 and 14 days, the colony number of the groups without zirconia was found to be lower, while the colony number of the groups with zirconia was found to be the highest (P < 0.001). The colony number of Procryla 3Z specimens kept in water kefir for 1 and 14 days (5.50 ± 0.02 and 5.80 ± 0.03) was found to be the highest (P < 0.001) (Fig. S1) (Table S3).

SEM results

Images of all material groups produced were obtained by scanning electron microscopy at 1000 × magnification.

Images of the Imicryl resin group are shown in Fig. 1, images of the Unmodified Procyla resin group are shown in Fig. 2, images of the Procryla 1% Zirconia resin group are shown in Fig. 3, and SEM images of the Procryla 3% Zirconia resin group are shown in Fig. 4.

Fig. 1
figure 1

Unmodified Imicryl resin SEM image

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Fig. 2
figure 2

Unmodified Procyla resin SEM image

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Fig. 3
figure 3

Procryla 1% Zirconia SEM image

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Fig. 4
figure 4

Procryla 3% Zirconia SEM image

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As can be seen in Fig. 1, bacterial adhesions from the images of the Imicryl resin group differed according to the environment. The most adhesive specimen is seen to be in the water kefir and almond milk group. Mineral water and distilled water have the lowest bacterial involvement. This situation is parallel to the SPSS results. In Fig. 2, when viewed according to the SEM images of the undoped 1% zirconia doped resins, the bacterial accumulation on the resin surface showed differences in the samples in this group. These differences vary according to the addition of zirconia nanoparticle and ambient conditions. It is seen to be more adhesive in distilled water environment compared to undoped resin. Figure 3 shows a parallel to these situations. In all resin groups, only 1% zirconia nanoparticle added heat-cured acrylic resins do not show the bead texture in the main structure of the resin. In this case, it can be said that the rate of 1% Zr nanoparticle prevents the deformation of the resin.

FTIR results

In this section, the FTIR results of all specimens kept in four different ambient conditions in the study were compared with each other. The FTIR spectra of Imicryl resin, Unmodified Procryla, 1% zirconia added Procryla resins and 3% zirconia added Procryla resins are shown in Figs. 5, 6, 7, 8, and 9, respectively. The FTIR spectra of all material groups produced correspond to the characteristic peak of the resin. The peaks of the FTIR spectrum of the undoped resin are shown in Fig. 5 at 466, 480, 557, 599, 675, 1120, 1635, 2918, 3302, and 3855 cm−1, respectively. Looking at the graphs in general terms, it is seen that three main absorption peaks emerge in the spectra of resins. These were mainly the peak of the C-H aromatic band seen at 675 cm−1, the C = C band of the methacrylate group at 1636 cm−1, and the H = O absorption peak of the hydroxyl ester group at about 3345 cm−1. In Fig. 5, vibrations between 710 and 1490 were seen in Imicryl resins as seen in the spectrum of milk kefir. The aliphatic C = C absorbance peak at 1637 cm−1 wavelength on the FTIR spectrum showed slight shifts in the spectra (Igci and Ozel Demiralp 2020). In acrylic resin in the FTIR spectrum, as shown in Fig. 5, a sharp portion of the C = O peak was observed at 1719 cm−1. Differences in the peaks of I0K, P0M, P1M, and P3A at 1140 cm−1 may be due to C–O–C stress vibrations (Cheung et al. 2011). The stretch at 1237 cm−1 is attributed to the C–O modes. 2948 cm−1 may be due to the increased vibration of C–H bond stress vibrations. The FTIR test graph provides strong evidence that all experimental groups did not undergo chemical changes except for the C = C band in the methacrylate group, in which residue monomer was not seen in samples that significantly reduced (Movasaghi et al. 2008).

Fig. 5
figure 5

Unmodified Procyla resin FTIR analysis

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Fig. 6
figure 6

Unmodified Imicryl resin FTIR analysis

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Fig. 7
figure 7

Unmodified Procyla resin FTIR analysis

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Fig. 8
figure 8

Procryla 1% Zirconia FTIR analysis

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Fig. 9
figure 9

Procryla 3% Zirconia FTIR analysis

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Discussion

Heat-cured acrylic-based resin materials have been used in dentistry for many years for the production of denture bases and hybrid implant overdentures. These materials play an important role in restoring aesthetic and masticatory functions, thus improving the quality of life of edentulous patients. However, a major drawback of these acrylic materials is their susceptibility to bacterial adhesion (Gupta et al. 2017).

The human oral cavity is a diverse microbial ecosystem with more than 1000 species of bacteria, mainly from the phyla Firmicutes, Actinobacteria, Proteobacteria, Fusobacteria, Bacteroidetes, and Spirochaetes (Sterzenbach et al. 2020). These microorganisms, influenced by factors such as oral health status and environmental conditions such as diet and gingival fluid, form complex biofilms both on teeth and mucosal surfaces and on the surfaces of dental materials such as implants, crowns, and prostheses. Bacterial adhesion to these surfaces can lead to tissue inflammation, causing conditions such as mucositis and peri-implantitis, and can also promote biofilm formation around dental restorations, leading to secondary caries (Sterzenbach et al. 2020; Sedghi et al. 2021).

To overcome these challenges, the incorporation of various nanoparticles into acrylic materials has been proposed. This approach aims to reduce cytotoxic effects and improve antifungal and bacterial adhesion properties of prosthetic base materials (Puspitasari et al. 2023). In our study, zirconia nanoparticles were added to heat-cured acrylic-based resins at concentrations of 1% and 3% by weight. The effect of this modification on bacterial adhesion was evaluated using various beverages (such as distilled water, mineral water, almond milk, and water kefir) at different time intervals (baseline, day 1 and day 14).

Our null hypothesis posited that the integration of zirconia nanoparticles into heat-cured acrylic resins, at concentrations of 1% and 3% by weight, and the exposure to different vegan beverages would not significantly alter the bacterial adhesion properties of these dental materials. Contrary to this hypothesis, our findings elucidate a complex interplay between the nanoparticle concentration, the type of beverage, and the resultant bacterial adhesion, thereby necessitating a rejection of the null hypothesis.

Different microorganisms commonly present in various foods and beverages can form biofilms on teeth, gums, and dental materials. The extent of biofilm formation, particularly that driven by Streptococcus mutans, may vary depending on interactions with other bacterial communities. Zirconia nanoparticles exhibit broad-spectrum bactericidal properties, effectively targeting both Gram-positive and Gram-negative oral bacteria, in addition to demonstrating antifungal efficacy against fungal spores (Pérez-Tanoira et al. 2016; Garcia et al. 2021).

Our study demonstrated that the incorporation of zirconia nanoparticles into heat‐cured acrylic resins significantly influenced bacterial adhesion as quantified in mg/mL. In distilled water, the group modified with 1 wt% zirconia exhibited bacterial adhesion levels of approximately 3.05 ± 0.03 log cfu/mL, indicating a reduction in microbial load compared to unmodified specimens. In contrast, in nutrient‐rich environments such as almond milk and water kefir, the incorporation of zirconia was associated with higher bacterial adhesion levels. For example, the Procryla group modified with 3 wt% zirconia showed bacterial adhesion values reaching up to 5.80 ± 0.03 log cfu/mL in water kefir after 14 days—reflecting an increase of nearly 90% relative to the values observed in distilled water. As the zirconia concentration increased in our study, bacterial adhesion correspondingly rose. Notably, the adhesion levels observed in specimens immersed in almond milk were comparable to those in water kefir. This phenomenon is likely attributable to the distinct bacterial communities inherent in these beverages, which can influence biofilm formation by modulating cellular motility, nutrient composition, and microbial load. Additionally, a study incorporating titanium, aluminum, vanadium, and yttria-stabilized zirconia demonstrated that the S. epidermidis strain p33 and S. aureus P4 exhibited significant adhesion on these modified surfaces (Pérez-Tanoira et al. 2016). These numerical findings align with observations from similar studies in the literature. Puspitasari et al. Puspitasari et al. (2023) reported that the integration of ZnO nanoparticles into acrylic resin resulted in a reduction in mucin adhesion from 20.59 ± 0.85 mg/mL in control specimens to 18.07 ± 0.80 mg/mL in optimally modified samples, suggesting an approximately 10% decrease in adhesion at effective nanoparticle concentrations. Similarly, Altarazi et al. (2023) found that incorporating TiO2 nanoparticles at low concentrations reduced the fungal biomass by 20–30% compared to controls, whereas higher nanoparticle concentrations led to an unexpected increase in microbial colonization—likely due to nanoparticle agglomeration and consequent alterations in surface free energy. The surface structure of zirconium oxide diminishes bacterial adherence relative to other materials, attributable to its electrical conductivity (Pérez-Tanoira et al. 2016).

A study investigated the antifungal effect of a nanocomposite against Candida albicans. The fungus was cultured in Sabouraud dextrose broth at 37 °C for 24 h. Afterward, the suspension’s optical density was adjusted for consistency. This suspension was then diluted to a specific cell concentration. Nanocomposite discs were prepared and placed in a 6-well plate. Each well was filled with the fungal suspension and incubated for 24 h. Post-incubation, the samples were washed and ultrasonically treated to detach fungal cells. The fungal solutions were then plated on agar and incubated again. Fungal colonies were counted, with results expressed as colony-forming units per milliliter. The study found that increased TiO2 (0.10, 0.25, and 0.50 wt%) in the nanocomposites significantly reduced fungal growth (Altarazi et al. 2023).

In our study, parallel to the research as mentioned above, we observed that adding 1% by weight zirconia to the specimens reduced bacterial adhesion in groups immersed in distilled water and 3 wt% zirconia in those immersed in mineral water. However, in groups exposed to other beverages, the incorporation of 1 wt% or 3 wt% zirconia increased bacterial adhesion.

Contrary to expectations, an increase in zirconia concentration was not associated with a further reduction in bacterial colonization. This phenomenon may be attributed to the complex interaction of the surface properties of the materials, in particular their surface free energy and the acidity of the beverages consumed. The changing surface free energy of dental materials with varying zirconia concentrations affects bacterial adhesion patterns. In addition, the acidity of beverages, an important environmental factor, plays an important role in modulating bacterial adhesion to these surfaces. Thus, while the incorporation of zirconium at low concentrations proved beneficial in reducing bacterial adhesion in distilled water media, its effectiveness did not increase linearly with increasing concentrations, underlining the complex nature of bacterial interactions with modified dental material surfaces (Sterzenbach et al. 2020).

There are studies identifying the addition of titanium dioxide nanoparticles in the range of 0.2–2.5 wt% as the most effective concentration for antibacterial and antifungal properties. However, it has been shown that exceeding a certain titanium dioxide concentration threshold can negatively affect some mechanical properties of the nanocomposite material. This negative effect is assumed to be due to the fact that higher nanoparticle content alters the intrinsic molecular structure of the polymerized material, acting as an impurity and potentially affecting the polymerization reaction. Consequently, an excessive amount of nanoparticles may lead to further leaching of unreacted monomers and TiO2NPs (titanium dioxide nanoparticles), resulting in instability of the printed samples and less desirable properties. Therefore, while TiO2NPs are effective in reducing bacterial adhesion, their concentration needs to be carefully balanced to maintain the mechanical integrity of the material (Karci et al. 2019; Altarazi et al. 2023).

In a recent study, zirconia nanoparticles (ZrO2NP) (1%, 2.5%, and 5%) modified groups showed significant antifungal effects and reduced bacterial adhesion compared to the control, especially as ZrO2NP concentration increased. Notable exceptions in antifungal effectiveness were observed between the 2.5% and 5% groups on day 14 (T1), and initially and after 5000 thermal cycles (T0 and T2) in the 1% group. Additionally, no significant effect was seen initially and on day 14 (T0 and T1) in the 5% group. While disk diffusion and filtration paper methods indicated no inhibition zones for ZrO2NP groups, the study confirmed ZrO2NPs’ antifungal properties and their sustained activity in heat-polymerized acrylic resin denture base materials after simulating 1 year of clinical use, suggesting long-term efficacy (Hamid et al. 2021).

Various atomic planes exhibit different surface energies and the surface energy of nanostructures is primarily influenced by the density of dangling bonds present on their surface. The difference in surface energy is crucial in determining the antifungal and antibacterial activity of nanoparticles. It can also be hypothesized that ZrO2NPs have identical surface geometries, but with various shapes, various active planes may exhibit different antimicrobial effects. Another explanation for the antifungal and antibacterial activity of ZrO2NPs is the presence of some nanoparticles on the surfaces of the samples, which may lead to direct contact between the nanoparticles and C. albicans and bacteria. This could interrupt cell function and actively inhibit the growth of fungal and bacterial strains (Hamid et al. 2021).

The findings of this study highlight the significance of beverage selection in relation to bacterial adhesion on heat-polymerized acrylic resins reinforced with zirconia. Specifically, the study draws attention to the consumption of water, mineral water, almond milk, and water kefir and their potential interactions with denture materials. From a bacterial adhesion standpoint, these interactions are crucial (Ebida et al. 2019).

Water, generally considered safe, does not typically contribute to increased bacterial adhesion or denture deterioration. However, mineral water’s acidic nature can enhance bacterial colonization on the denture surface, leading to increased risk of oral infections and necessitating strict denture hygiene post-consumption to mitigate these risks. Almond milk, while a healthier option, may still leave residues that could promote bacterial growth on dentures if not thoroughly cleaned afterward. This necessitates careful cleaning practices to prevent residue buildup, which can serve as a breeding ground for bacteria over time (Sutula et al. 2012).

In our study, the samples were immersed in distilled water, mineral water, almond milk, and water kefir without the use of artificial saliva for 14 days. At the end of this period, bacterial adhesion was measured, but the specific bacterial species present were not identified. Furthermore, aspects such as surface angle, surface roughness, and hydrophobicity of the surface were not investigated in the zirconia (ZrO2) added groups. Future research could usefully incorporate these parameters to provide a more comprehensive understanding of the interaction of the material in various environments. This would offer a deeper insight into the potential of ZrO2 modified materials in dental applications, especially in simulating conditions closer to the natural oral environment (Alla et al. 2017; Ebida et al. 2019).

Within the limitations of this study, incorporating zirconia nanoparticles into heat-cured acrylic denture base resins demonstrated significant effects on bacterial adhesion, depending on zirconia concentration and the immersion medium. In neutral environments like distilled water, a 1 wt% zirconia addition effectively reduced bacterial colonization, while higher concentrations (3 wt%) showed diminishing antibacterial benefits. In mineral water, the 3 wt% zirconia group exhibited enhanced long-term antibacterial properties after 14 days. However, in nutrient-rich environments such as almond milk and water kefir, the addition of zirconia appeared to promote bacterial growth, with the highest adhesion observed in the 3 wt% zirconia group.

In conclusion, a 1 wt% zirconia concentration may be optimal for reducing bacterial adhesion in neutral or mineral-rich environments without compromising resin integrity. However, in nutrient-rich conditions, zirconia may facilitate bacterial growth, underscoring the need for careful consideration of zirconia content based on specific oral conditions.