Antimicrobial glass with silver for zirconia finishing.

Description

1 / 16 Antimicrobial glass with silver for zirconia finishing. Brief description.

[001] The present invention relates to a soda-lime glass of the borosilicate system with added silver that was developed to be used as a finisher on zirconia dental restorations. The present invention eliminates any undesirable darkening effect of the material, in addition, it has demonstrated greater durability, resistance, and excellent aesthetics and integration of ceramic prostheses with adjacent tissues and has antimicrobial properties. FIELD OF APPLICATION

[002] The present invention belongs to the field of Dentistry, more specifically to the field of materials for prostheses (such as ceramic prosthesis coatings) acting as a finisher on zirconia dental restorations. STATE OF THE ART

[003] Zirconia is a material with excellent mechanical properties, but although it exhibits excellent mechanical strength, its aesthetic properties are inferior to those obtained in restorations with vitreous ceramics. To meet this aesthetic demand of zirconia, a finishing glaze (enamel / varnish) has been used to provide a better finish and polish, providing characteristics similar to the natural tooth. In the context of prosthetic restorations, in addition to aesthetic and functional aspects, another very important factor that must be taken into account is the presence of pathogenic microorganisms in the oral environment. The presence of these microorganisms, combined with the patient's lack of cooperation in cleaning the prosthesis, can lead to treatment failure, regardless of how good the mechanical properties of the restorative material used are.

[004] Document CN113149438B refers to a dental enamel surface composition, a method for forming a dental enamel surface, and a dental prosthesis. The dental enamel surface composition comprises the Petition 870240109893, dated 12 / 24 / 2024, p. 9 / 42 2 / 16 The following components in mass percentage: 53 to 65% SiO2, 6 to 15% B2O3, 8 to 12% Na2O, 6 to 8% K2O, 1 to 3% U2O, 2 to 5% alkaline earth metal oxide, 5.5 to 11% AbO3 and 0.5 to 2% ZrO2.

[005] Document CN113149438B presents a product intended to finish and smooth the surface of ceramics. The authors state that they have modified the chemical proportions of products already on the market, excluding some toxic components and adding others to arrive at a less toxic and more stable product: “It is an object of the present invention to overcome the above-mentioned drawbacks of the prior art and to provide a tooth enamel composition which can be sintered at a relatively low temperature and is non-toxic”... “In addition, the prior art enamel composition includes toxic components such as barium oxide, lead oxide, etc., which may have adverse health effects.”

[006] Unlike the product presented in document CN113149438B, the glass in this patent application is a finisher that combines in a single product the finishing capabilities of zirconia, antimicrobial properties, improved strength of the material to which it is applied, and is non-cytotoxic.

[007] The product presented in document CN113149438B and the silver-impregnated antimicrobial glass for finishing zirconia are different products, with different compositions, proportions of chemical elements and temperatures. A glass is characterized by its proportions and types of oxides, as well as by the processing temperature, which determines a distinct product.

[008] It is clear that although both products are glass, and have some oxides in common, they have different proportions, chemical elements, different temperatures and are intended to solve different technical problems, thus characterizing another product.

[009] Document US11426332B2 is a granted patent and refers to a shade-coordinated porcelain veneer system for dental prostheses. The system contains two types of veneer porcelains: enamel Petition 870240109893, dated 12 / 24 / 2024, page 10 / 42 3 / 16 natural / body dentin and opaline enamel, as well as stained and glazed porcelains, with compatible CTEs that can be used for dental prosthesis veneering. The system is made of 1) a lithium disilicate-based glass-ceramic substructure formed by hot pressing or CAD / CAM machining process or 2) a YTZP zirconia-based ceramic substructure formed by CAD / CAM machining process to improve the overall aesthetics of the final prosthesis.

[010] The product of US patent 11426332B2 consists of a porcelain, a vitreous ceramic (aesthetic ceramic) developed to be applied over other ceramics, such as zirconia and disilicate. In dentistry, this outer porcelain layer is frequently used in areas of the mouth that bear greater loads, such as the posterior teeth. To ensure greater mechanical strength, more resistant, although less aesthetic, ceramics such as zirconia and disilicate are used as a base (infrastructure), and a porcelain layer is placed over them to enhance aesthetics.

[011] The US11426332B2 product was developed with the main objective of trying to solve the technical problem associated with the difficulty in harmonizing colors when one porcelain is placed over another ceramic. The US11426332B2 product applies a specific color shading to ensure that, when applied over the lithium disilicate substructure or over the YTZP zirconia substructure, both with different levels of translucency, the dental prostheses achieve color harmonization.

[012] Unlike the product in US11426332B2, the glass in this patent application is not a ceramic and is not intended to solve color problems when applied to zirconia. The products are different materials, have different compositions, different temperatures, and are intended to solve different technical problems. The glass in this patent application is a finisher for zirconia, which improves mechanical properties, is antimicrobial, and does not affect the aesthetics of the material. Therefore, the product in US11426332B2 is not relevant to the patent application in question, since it belongs to a completely different product category. Petition 870240109893, dated 12 / 24 / 2024, page 11 / 42 4 / 16

[013] Document BR 1020140227377 describes a method for preparing the surface of devices made of titanium or titanium alloys, zirconium, zirconium dioxide, alumina or zirconia / alumina compounds, medical-grade stainless steels and cobalt-based medical superalloys, which form the object of the invention; said devices being implantable in the human body or in animals and connected extracorporeal parts made of the same materials, particularly for dental and orthopedic implantology. The implantable device is thus treated: it is provided to expose at least a portion of the surface of said device to a solution comprising hydrofluoric acid, phosphoric acid, at least one surfactant substance and water.The exposure occurs over a period of time and under conditions sufficient to provide the implant surface with the desired surface roughness and the formation of self-induced surface titanium dioxide, maintaining the structural integrity of the device and without altering the centesimal measurement size. The surface thus treated is rinsed with demineralized water and ultrasound in order to avoid metallosis phenomena. In the extraosseous parts complementary to the devices to be implanted in the bone, nanometric particles of mineral trace elements smaller than 20 nm are applied, and in particular, 99.9% pure silver, having the function of self-cleaning and adhering to the surfaces treated with the method described above, without the aid of adhesives.

[014] The text of patent BR 1020140227377 describes that the patent is for a Method. The authors state that the method was developed for the purpose of preparing the surface of dental or orthopedic implants made of different materials, and to improve the effectiveness and compatibility of the implants with the human or animal body. It is therefore not a specific product but a method. The authors further state that the method would make it possible to guarantee a clean surface at the end of the treatment, which would have the advantage of facilitating the adhesion of bone tissue to the implant and avoiding contact of unfavorable metal parts or processing residues with the bone structure or tissues. Petition 870240109893, dated 12 / 24 / 2024, page 12 / 42 5 / 16 adjacent. Cleaning the surface would provide better conditions for the application of antibacterial and antimicrobial substances.

[015] In the text of BR 1020140227377 it is clear that the method itself does not possess antimicrobial properties, but the surface preparation would facilitate the subsequent application of antimicrobial substances. The focus of patent BR 1020140227377 is on surface preparation to create nano-roughness and allow better fixation to the bone, in addition to enabling the application of antimicrobial substances after the execution of the proposed method.

[016] Thus, patent BR 1020140227377, in describing a method, differs completely from the patent application for the glass of this invention, which focuses on a specific product with antimicrobial properties incorporated directly into the glass, and intended for the finishing of zirconia. This distinction reinforces the uniqueness of the invention presented in this patent application compared to the method described in BR 1020140227377. OBJECTIVES OF THE INVENTION

[017] The present patent application aims to develop a product to be used as a finishing agent on zirconia dental restorations. This product aims to improve the patient's oral health and enhance the durability, strength, aesthetics, and integration of ceramic prostheses with adjacent tissues. ADVANTAGES OF THE INVENTION

[018] The present invention has the advantage of including silver in the glass, contributing to the inhibition of the growth of microorganisms, reducing the need for antimicrobial treatments present in the oral cavity, such as Streptococcus sanguinis, Streptococcus mutans and Escherichia coli, or the premature replacement of dental restorations. The product of the invention preserves the original color of the zirconia after application, eliminating any undesirable darkening effect of the material, in addition to presenting itself as a promising and effective alternative to raise the standards of quality and durability. Petition 870240109893, dated 12 / 24 / 2024, page 13 / 42 6 / 16 in zirconia ceramic restorations. DETAILED DESCRIPTION OF THE FIGURES

[019] Figure 1 is a graph that illustrates the CFU / ml (Log10) values ​​of S. sanguinis obtained at baseline and after 6 months.

[020] Figure 2 shows photographs illustrating scanning electron microscopy (SEM) images of S. sanguinis biofilm formed on the surfaces of Glaze groups (Figure 2A) and Ag-enriched glass (Figure 2B).

[021] Figure 3 is a graph showing the colony-forming unit results obtained from each group against S. mutans.

[022] Figure 4 shows photographs illustrating SEM images of S. mutans biofilms formed on silver glass samples (Figure 4A and Figure 4B) and commercial glaze (Figure 4C and Figure D), tested initially (Figure 4A and Figure 4C) and after aging (Figure 4B and Figure 4D). A smaller number of colonies is observed on silver glass in the post-aging image (Figure 4B) compared to the initial image (Figure 4A).

[023] Figure 5 is a graph of the distribution of E. coli colony formation on 5Y-PSZ zirconia.

[024] Figure 6 shows photographs illustrating SEM images of E. coli on the specimens under analysis. Figure 6a and Figure 6b are E. coli on specimen group 5Y-C (sintered zirconia, at 10000x and 20000x magnification, respectively); Figure 6c is from group 5Y-G (commercial glaze, 2000x); Figure 6d is from 5Y-G (group with commercial glaze, 4000x); Figure 6e is from group 5Y-Cal (2000x); Figure 6f is from group 5Y-Cal (4000x); Figure 6g is from group 5Y-Ag (silver glass, 2000x) and Figure 6h is from group 5Y-Ag (silver glass, 4000x). The arrows (x) indicate the presence of E. coli cells.

[025] Figure 7 is a graph showing the cytocompatibility test. Groups with glaze (control), AgCoat (glass with silver) and Bcoat (glass with boron, control).

[026] Figure 8 shows photographs of the LIVE / DEAD test. The cytocompatibility test showed that the tested groups did not induce cell death. Petition 870240109893, dated 12 / 24 / 2024, page 14 / 42 7 / 16 DETAILED DESCRIPTION OF THE INVENTION

[027] The glass was manufactured using the sol-gel method. The sol-gel process involves the preparation of a sol, which is followed by gelation and removal of the liquid phase. Silicic acid (silica source) was obtained by passing an aqueous solution (10% w / w) of sodium silicate pentahydrate (Na2SiO3^5H2O) through an ion exchange resin (IR120, Dow Corning). After the synthesis of silicic acid (0.5 mol / l), aluminum nitrate, calcium nitrate, sodium nitrate, potassium nitrate and / or silver nitrate were added to obtain the desired mass compositions at different concentrations.

[028] The compositions were obtained by mixing the aforementioned nitrates with the cations and silicic acid solution corresponding to the amount of silica. The mixtures were placed in an oven at 100 °C for 24h and then calcined at 600 °C for 5h. At the end of this process, the resulting materials were ground and sieved (200 mesh).

[029] After obtaining the glass powder, 3Y-TZP, 4Y-PSZ, and 5Y-PSZ zirconia specimens (discs and square specimens) were sintered in a Sirona inFire HTC speed furnace, following the manufacturer's recommendations. The sintering cycle was performed as indicated by the manufacturer. At the end of the cycle, the final thicknesses of the zirconia specimens were: discs (12 x 1.2 mm) and squares (4.0 x 4.0 x 1.6 mm). The glass powder (0.20 g) was mixed separately with 5 drops of propylene glycol PA-ACS (LABSynth Produtos para Laboratórios Ltda) in a mortar. The resulting paste was applied to the polished zirconia surfaces with a brush, under manual vibration, to ensure uniform paste flow and a smooth finish. The glass-coated discs were subjected to a two-stage firing cycle: 1200°C for 20 minutes, followed by 700°C for 30 minutes, with cooling to 200°C at a rate of 25°C / min before opening the oven (InFire HTC Speed).Pilot studies were conducted to determine the temperature and holding time that would provide a smooth appearance to glass-coated specimens. Petition 870240109893, dated 12 / 24 / 2024, page 15 / 42 8 / 16

[030] The composition of the glass involves a modified soda-lime glass powder, of the SiO2-Na2O-AbO3-CaO-B2O3 system (SiO2: silicon dioxide; Na2O: sodium oxide; Al2O3: aluminum oxide; CaO: calcium oxide; B2O3: boron oxide) plus 10% silver. With the following final composition (% by weight): SiO2 (37.4); Na2O (18.0); CaO (17.6); AbOs (9.1); B2O3 (5.8); MgO (0.19); K2O (0.55); Ag (10). RESULTS OBTAINED

[031] Tests with different sintering temperatures were carried out, as well as on the surface of zirconia with different concentrations of yttrium oxide (3Y-TZP, 4Y-PSZ and 5Y-PSZ). Satisfactory aesthetic results were achieved with the following glass sintering protocol: 1200°C for 20 minutes, followed by 700°C for 30 minutes.

[032] Tests performed

[033] Evaluation of the antimicrobial action of glass on Streptococcus sanguinis biofilm applied to 4Y-PSZ zirconia:

[034] An experiment was conducted to evaluate the antimicrobial action of silver glass and commercial glaze (control) on S. sanguinis biofilm. To evaluate the antimicrobial properties of the samples, the test involved the microorganism Streptococcus sanguinis (ATCC 7073).

[035] Preparation of zirconia samples

[036] For the test, 4Y-PSZ zirconia samples were produced in the form of 4x4x1mm squares. These samples were made using a precision cutting machine equipped with a water-cooled diamond saw. Subsequently, the resulting plates were subjected to a leveling and polishing process. For the preparation of the square samples, a diamond disc was attached to a straight handpiece and a micromotor. The square samples were properly cleaned with absolute ethyl alcohol and then subjected to the sintering process.

[037] The sintered samples were divided into two distinct groups, and each group received the application of a surface treatment: commercial glaze or glass. Petition 870240109893, dated 12 / 24 / 2024, page 16 / 42 9 / 16 with silver (product of this patent), on the upper surface of these samples and were then subjected to the burning process.

[038] Colony-forming units (CFU) assay

[039] After preparing the zirconia specimens as described above, the specimens were sterilized by ultraviolet light in a laminar flow hood for 15 min per 4.0 χ 4.0 mm side. S. sanguinis were seeded onto Petri dishes with mitis salivarius (MS) agar (Synth, Diadema, Brazil). S. sanguinis were incubated for 48 hours at 37°C and 5% CO2. Subsequently, the cultured colonies were suspended in Falcon tubes filled with brain heart infusion (BHI). The suspension was maintained at 37°C and 5% CO2. After 24 h, the cultured microorganisms were centrifuged (5,000 rpm for 5 min) and the broths were discarded. The Falcon tubes were filled with sterile physiological saline solution (0.9% sodium chloride, NaCl) and centrifuged again. This washing process was performed twice. Then, the number of cells in suspension was standardized (107 cells / ml) using a spectrophotometer.The optical density and wavelength parameters used were 0.560 and 398 nm, respectively; these parameters were previously established using a standard curve for CFU versus absorbance.

[040] Specimens were placed in a 96-well plate and contaminated to obtain the homotypic biofilm, and each sample-containing well was filled with 50 µL of the standardized suspension. The streptococcus plate was maintained at 37°C and 5% CO2. After 24 h, the media were removed, the wells were washed twice with sterile 0.9% NaCl and filled with 200 µL of the specific broths. After the following 24 hours, the specimens were washed to remove weakly bound cells and placed in microtubes containing 2 mL of sterile 0.9% NaCl solution. To remove the formed biofilm, the specimens were sonicated for 15 s. The resulting suspensions were diluted 10⁻², 10⁻⁴, and 10⁻⁵ times. 5 µL aliquots were seeded in triplicate onto plates of Petition 870240109893, dated 12 / 24 / 2024, page 17 / 42 10 / 16 petri dishes with selective medium. Subsequently, colony-forming units were counted after 24 hours of incubation at 37°C and 5% CO2.

[041] The evaluation of antimicrobial activity was carried out at two periods: immediately after application of the vials and after six months of immersion in distilled water.

[042] The evaluation was carried out by counting colony forming units (CFU) and by scanning electron microscopy (SEM).

[043] The results demonstrated that after six months, the groups that received the application of silver glass showed a significant reduction in microbial growth (CFU / ml), compared to the group that received the application of commercial glaze (Table 1).

[044] Table 1 - Means and standard deviations of Colony Forming Units (CFU) of S. sanguinis from each experimental group evaluated initially and after 6 months of immersion in distilled water. Group CFU / ml (Log10) Initial CFU / ml (Log10) 6 months Glaze commercial 7.67 (0.38)Aa 7.43 (0.23)^5 Silver 7.46 (0.26)Aa 6.40 (0.65)bb

[045] The colonies were analyzed by Scanning Electron Microscopy (SEM). Through the SEM images it was possible to verify the decrease in Streptococcus sanguinis biofilm on the specimens where silver glass was applied (Figure 2).

[046] Silver-impregnated glass applied to 4Y-PSZ zirconia was effective against S. Sanguinis biofilm. The results demonstrated the long-term antimicrobial action of silver-impregnated glass (after six months of immersion in distilled water), showing that the glass was effective when applied to zirconia with a higher presence of the cubic phase (4Y-PSZ).

[047] Evaluation of the antimicrobial action of glass on Streptococcus mutans biofilm applied to 4Y-PSZ zirconia Petition 870240109893, dated 12 / 24 / 2024, page 18 / 42 11 / 16

[048] To evaluate the antimicrobial action of silver glass and commercial glaze (control) on S. mutans biofilm, the test conducted involved the microorganism Streptococcus mutans (ATCC 35688).

[049] For the test, zirconia samples were used in the form of squares measuring 4x4x1mm. The preparation of the samples and the preparation for the CFU (Colony Forming Unit) count assay was similar to that described above in the test with S. Sanguinis.

[050] For S. mutans, the surface treatment (P<0.001) and time (P = 0.004) factors showed a significant effect on CFU. The interaction between these factors was also significant (P = 0.011). Silver-coated glass demonstrated effectiveness after a six-month aging period of the specimens (Figure 3). These findings are visually presented in Figure 4 (Scanning electron microscopy), which illustrates the biofilms formed on the surfaces of the specimens. It was possible to note through SEM images that there was a significant decrease in S. mutans after six months of aging where silver-coated glass was used.

[051] Evaluation of the antimicrobial action of glass on E. coli biofilm applied to 5Y-PSZ zirconia

[052] The antimicrobial effect of silver-impregnated glass applied to 5Y-PSZ zirconia in contact with E. coli bacteria was also evaluated.

[053] For the experiment, 5Y-PSZ zirconia was cut into squares (4 χ 4 χ 1 mm) on a precision cutting machine with a diamond saw under water cooling. The samples were then leveled and polished. The squares were cleaned in absolute ethyl alcohol and sintered.

[054] After sintering, the samples were randomly divided and the groups were tested according to the surface treatment received by the zirconia: - 5Y - C (5Y zirconia - sintered only); - 5Y + G (zirconia with commercial glaze); - 5Y + Cal (zirconia with sodium lime glass); - 5Y + Ag (zirconia with Petition 870240109893, dated 12 / 24 / 2024, page 19 / 42 12 / 16 glass with silver). Sample preparation and CFU (Colony Forming Unit) assay were similar to those described in the experiments above.

[055] The results demonstrated that the group that received the application of silver glass (product of this invention) showed the lowest colony growth when compared to the other groups in the analysis (Figure 5). The quantity of E. coli cells on the surface of the specimens can be verified by SEM images (Figure 6).

[056] Evaluation of the average surface roughness of 4Y-PSZ Zirconia after application of silver-glazing

[057] 4Y-PSZ zirconia discs were rounded into cylinders (16 mm χ 14 mm). The cylinders were cut into discs (16 mm χ 1.5 mm) which were leveled and polished under water cooling in a polishing machine until they reached 1.2 mm thickness. The discs were cleaned and air-dried, and then sintered in a dental furnace at 1550°C for 2 h (heating and cooling rates: 10°C / min). After sintering, the final dimensions of the discs were 13 mm in diameter x 1.0 mm thick. The discs were randomly distributed into experimental groups according to the surface finisher: commercial glaze (control group) and silver-glazed glass (Ag).

[058] The control group discs were coated with commercial glaze paste. The glaze paste was homogenized with a spatula and applied to the polished zirconia surface with a brush. The glazed discs were fired in a dental furnace according to the manufacturer's specifications.

[059] For the discs that received the glass with silver as a finisher, the glass was mixed with 5 drops of propylene glycol in a mortar. The resulting paste was applied to the polished zirconia surfaces with a brush under manual vibration and then the discs were subjected to firing.

[060] After applying the glaze or silver glass, the discs were thus ready for the roughness test. The average roughness test of Petition 870240109893, dated 12 / 24 / 2024, page 20 / 42 13 / 16 surface test was performed by the group to evaluate the surface of zirconia after application of silver-reinforced glass compared to commercial glaze.

[061] The average roughness (Ra) and the ten-point average roughness (Rz) of the samples were evaluated using a portable contact roughness tester. The tests were performed with a cut-off of 0.80 mm, a resolution of 0.0001 µm (8 µm range), a speed of 0.5 mm / s and a total length of 4 mm (according to ISO 4287 / 1997). For each sample, three measurements were taken on the x-axis and three on the y-axis. The average of the six measurements was used for statistical analysis.

[062] The roughness results obtained from each group are described in Table 2. The surface treatment factor significantly affected Ra and Rz (both P<0.001). The glass with silver (product of this patent application) showed lower Ra and Rz values ​​compared to the commercial glaze. This demonstrates that the glass with silver, in addition to not altering the surface of the material, was also able to promote a smoother surface.

[063] Table 2 - Means (standard deviations or 95% confidence intervals) of roughness parameters obtained from each experimental group. Ra (pm)** Rz (pm)* Glaze 0.14 (0.03^ 0.61 (0.08)A Glass with Silver 0.07 (0.01 )B 0.32 (0.03)B

[064] Analysis of the translucency of 4Y-PSZ Zirconia after application of silver glass

[065] Another test was performed to verify whether the application of silver-reinforced glass over 4Y-PSZ Zirconia would affect the material's translucency. The translucency of zirconia is an important characteristic when considering its use in dental restorations such as crowns, veneers, and bridges, as it plays a fundamental role. Petition 870240109893, dated 12 / 24 / 2024, page 21 / 42 14 / 16 in achieving a natural and aesthetic appearance in restorations. Translucency is the ability of a material to allow light to pass through it, but in a diffuse way, so that the light is not completely transparent, but also not opaque. The analysis of zirconia translucency involves measuring the amount of light that passes through the material and how it propagates through it.

[066] For the tests, zirconia discs were prepared in a manner similar to that reported in the test above (roughness analysis). A spectrophotometer was used to measure the color characteristics of the discs before and after the application of the silver glass and the commercial glaze. For standardization of the measurements, the CIEL*a*b* parameters were used (Commission Internationale de l'Éclairage, where L* = luminosity axis, a* = green-red color gamut and b* = blue-yellow color gamut).

[067] Measurements were performed on black and white backgrounds under standard illumination (illuminant D65) and at a specific viewing angle of 10 degrees. Each sample was measured three times on both background types, and then an average of these three measurements was calculated to obtain information on translucency. To reduce light scattering, a small amount of glycerol (C3H8O3, refractive index: 1.47) was used between the sample and the background to minimize light dispersion. The L*a*b* results obtained on black and white backgrounds were used to calculate the translucency (TP00) of each experimental group. Simple differences (ΔTπ00) between the TP00 value of the glaze and silver glass groups were calculated. These differences were evaluated based on perceptibility and unacceptability criteria previously established by Salas et al.(2018), where differences greater than 0.62 were considered perceptible and those greater than 2.62 were considered unacceptable.

[068] Translucency was similar between groups (p = 0.299) (Table 3). Furthermore, the ΔTΡ00 of the glass with silver, compared to the commercial glaze, was below the perceptibility limit (ΔTΡ00<0.62). Petition 870240109893, dated 12 / 24 / 2024, page 22 / 42 15 / 16

[069] The translucency test result demonstrated that the glass with silver, in addition to providing an antimicrobial effect (as detailed in the tests above), did not compromise the translucency of 4Y-PSZ zirconia compared to commercial glaze.

[070] Table 3 - Means (standard deviations or 95% confidence intervals) of translucency parameters obtained from each experimental group. TPotT ΔTPoo Glaze 13.97 (0.63)A - Glass with silver 14.02 (0.88)A 0.05

[071] Cytocompatibility Assessment: WST-1 and LIVE / DEAD assays

[072] For the test, zirconia specimens were prepared in a manner similar to that described above in the colony-forming unit test. The specimens were divided into three groups according to the coating applied to the zirconia: glass with silver (product of this patent), commercial glaze (control) and another glass containing boron (control).

[073] Periodontal ligament stem cells (PDLSCs) were seeded (10³ per sample) and cultured in growth medium for 24 hours at 37°C with 5% CO₂. After that, the culture medium was replaced with fresh medium containing WST-1 reagent (Sigma Aldrich) in a 1:1 ratio. The cells were incubated for 2 ha at 37°C and absorbance was measured at 450 nm using a spectrophotometer. LIVE / DEAD reagents in a 1:1 ratio were added to the wells and incubated at 37°C for 30 min. The samples were photographed under a fluorescence microscope. Cell-free discs served as blanks. Petition 870240109893, dated 12 / 24 / 2024, page 23 / 42 16 / 16

[074] The WST-1 (Figure 7) and LIVE / DEAD (Figure 8) tests showed similar results for all groups tested, suggesting that there were no cytotoxic effects induced by the experimental coatings.

[075] The cytocompatibility test (WST-1) showed no difference between the groups tested (dashed line and * indicates the positive control for the reaction: live cells cultured in polystyrene).

[076] Considerations on technology

[077] The tests and analyses described and detailed above elucidate that the antimicrobial glass with silver (product of this patent application) emerges as a viable alternative to be used in the finishing of zirconia. These tests not only confirm its technical feasibility, but also demonstrate its innovative characteristics, as it combines in a single product antimicrobial properties, the ability to finish zirconia, and at the same time promotes better aesthetics of the material without causing darkening. In this way, it contributes to solving problems related to the durability of restorative treatments, well-being and general health of the patient. Petition 870240109893, dated 12 / 24 / 2024, page 24 / 42

Claims

1 / 1 CLAIM 1. “ANTIMICROBIAL GLASS”, manufactured by the sol-gel process, in which silicic acid is obtained by passing an aqueous solution (10% w / w) of sodium silicate pentahydrate (Na2SiO3^5H2U) through an IR120 ion exchange resin; characterized by comprising SiO2 (37.4% by weight); Na2O (18.0% by weight); CaO (17.6% by weight); AbO3 (9.1% by weight); B2O3 (5.8% by weight); MgO (0.19% by weight); K2O (0.55% by weight); Ag (10% by weight). Petition 870240109893, dated 12 / 24 / 2024, pages 33 / 42

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