Method for testing adhesive force of tooth or tooth-like material interface and application of method

Abstract

The invention discloses a method for testing the adhesive force of a tooth or tooth-like material interface and application. The method comprises the following steps: carrying out scratch test on a to-be-tested surface layer by using linear loading force in a specified range by adopting a micro-adhesion tester or an atomic force microscope; scratches are divided into three stages, in the first stage, linear loading force is used for detecting height changes in the whole process, in the second stage, scraping is slight and discontinuous, obvious furrows do not appear, in the third stage, obvious furrows appear, but the width of the furrows is narrow, and obvious base body scratches cannot be observed in the centers of the furrows; calculating a sudden change point load of the to-be-tested surface as an index for evaluating the adhesive force; the linear loading force in the specified range is the force from the minimum force for breaking the to-be-tested surface to the force for puncturing the to-be-tested surface without forming obvious scratches on the substrate. According to the method, the adhesive force of the in-vitro biological model can be tested, the comparability of different research data is ensured, a unified in-vitro test standard can be formulated, and a foundation is laid for promoting industry standardization.

Description

Technical Field

[0001] This application relates to the field of tooth or tooth model testing technology, and in particular to a method for testing the interfacial adhesion of teeth or tooth-like materials and its application. Background Technology

[0002] In the field of oral medicine, to study the effects of oral hygiene products such as toothpaste, it is often necessary to construct various in vitro biological models using tooth-like materials, such as single-species biofilms, plaque, soft deposits, tartar, or similar mimics. Through research on plaque, tartar, soft deposits, and tartar, it is now possible to effectively simulate the actual oral environment in vitro. For example, using artificial saliva and under conditions such as oral temperature, structures such as plaque, tartar, soft deposits, and tartar can be simulated on the surface of tooth models. These in vitro biological models can be used for various research in oral medicine.

[0003] However, the adhesion data of existing in vitro biological models are unknown compared to the adhesion in the real human oral cavity. In other words, although biological models such as dental plaque can be constructed in vitro, there is no quantitative testing technology to determine whether the adhesion properties of these in vitro biological models can effectively reflect the actual adhesion of dental plaque on human teeth. This results in existing in vitro test results being unable to provide a basis for judging their correlation with clinical outcomes, leading to distortions in the evaluation of the anti-adhesion performance of dental materials.

[0004] Conventional methods for adhesion testing include peel tests, such as attaching a test column to the coating with adhesive, stretching it vertically with a tensile testing machine until separation, and recording the tensile force per unit area to characterize the coating's adhesion; alternatively, peeling the coating at a specific angle, such as 180° or 90°, and measuring the required peel force; or applying force vertically until the coating separates and measuring the maximum tensile stress. However, these existing adhesion testing methods are not applicable to adhesion testing of in vitro biological models of teeth or tooth-like materials; this results in a lack of unified standards for constructing in vitro biological models.

[0005] Therefore, how to effectively test the adhesion of tooth or tooth-like material interfaces remains a key research focus and challenge in this field. Summary of the Invention

[0006] The purpose of this application is to provide a new method for testing the interfacial adhesion of teeth or tooth-like materials and its application.

[0007] To achieve the above objectives, this application adopts the following technical solution:

[0008] The first aspect of this application discloses a method for testing the interfacial adhesion of teeth or tooth-like materials, including using a micro-adhesion force tester or atomic force microscope to perform a scratch test on the surface layer to be tested using a linear loading force within a specified range; the scratch is divided into three stages according to the changes in the scratch; the first stage uses a linear loading force within a specified range to detect the height change throughout the process; the second stage is a slight and discontinuous scratch without obvious furrows; the third stage has obvious furrows, but the furrows are narrow and no obvious scratches formed by the friction between the mating material and the substrate can be observed in the center of the furrows.

[0009] The abrupt change point load that breaks down the surface layer under test is calculated using the following formula and used as an indicator to evaluate the adhesion of the surface layer under test.

[0010] The load at the abrupt change point = the profile height of the third stage ÷ the total profile height of the first stage × the maximum load of the linear loading force within the specified range;

[0011] A linear loading force within a specified range is defined, with a minimum loading force capable of breaking through the surface layer under test and a maximum loading force capable of penetrating the surface layer under test without causing significant scratches to the teeth or tooth-like materials. In this application, the abrupt change point load is the minimum force required to penetrate the surface layer under test, ensuring effective penetration of the surface layer under test.

[0012] It should be noted that this application creatively employs a micro-adhesion force tester or atomic force microscope to test the adhesion force on the surface of teeth or tooth-like materials, and uses the abrupt change load that breaks down the test surface layer as an indicator to evaluate the adhesion force of the test surface layer, thus developing a new method for adhesion force testing of in vitro biological models. The adhesion force test data obtained by the testing method of this application show excellent consistency with the test data of real human teeth, bridging the gap between in vitro biological model test data and clinical data. The testing method of this application can quantitatively analyze the adhesion performance of in vitro biological models, which is conducive to the formulation of unified in vitro testing standards, promotes industry standardization, and ensures the comparability of data from different studies.

[0013] In one implementation of this application, the linear loading force within a specified range is 0.03-0.5N.

[0014] In one implementation of this application, the teeth are human teeth or other animal teeth extracted from the body.

[0015] It is understood that the test method of this application can be used for adhesion testing on tooth surfaces, including but not limited to human teeth, and is equally applicable to the teeth of other animals.

[0016] In one implementation of this application, the tooth-like material includes at least one of the following: materials that form teeth or other arbitrary shapes with hydroxyapatite as the main component, resin materials, metal alloy materials, bioactive glass materials, ceramic materials, and polymer materials.

[0017] In one implementation of this application, the ceramic material includes at least one of zirconia ceramic and lithium disilicate glass ceramic.

[0018] In one implementation of this application, the polymer material includes at least one of polyetheretherketone and polymethyl methacrylate.

[0019] In one implementation of this application, the surface layer to be tested is dental plaque, soft deposits, tartar, or a similar simulant.

[0020] In one implementation of this application, the surface layer to be tested is plaque, stains, soft deposits, tartar, or calculus naturally present on the extracted teeth of a person or animal.

[0021] In one implementation of this application, the surface layer to be tested is plaque, soft deposits, dental tartar, or dental calculus constructed through in vitro culture, or a simulated coating prepared from water-based paint or coating.

[0022] It should be noted that many paints or resin-based coatings on the market claim to be able to act as dental plaque mimics; therefore, the testing method of this application can also be used to test the adhesion of simulated coatings prepared from water-based paints or coatings, and the simulation effect can be quantitatively evaluated through the testing method of this application.

[0023] In one implementation of this application, the plaque constructed by in vitro culture is a plaque biofilm constructed by in vitro cultured bacteria, or a plaque biofilm simulated by sodium carboxymethyl cellulose and / or hydroxyethyl cellulose.

[0024] In one implementation of this application, the bacteria cultured in vitro include at least one of Streptococcus mutans, Streptococcus sanguinis, and Actinomycetes.

[0025] In one implementation of this application, the dental calculus constructed in vitro includes a chemical deposition mimicry formed from at least one of hydroxyapatite, octacalcium phosphate, calcium carbonate, and collagen.

[0026] In one implementation of this application, the waterborne paint or coating includes at least one of waterborne acrylic resin, waterborne matting agent, waterborne cobalt blue paste, waterborne toluene diethyl cyanate, and waterborne propylene glycol methyl ether acetate.

[0027] The second aspect of this application discloses the application of the method for testing the interfacial adhesion of teeth or tooth-like materials in predicting the anti-plaque durability of a particular material surface in the oral environment.

[0028] The third aspect of this application discloses the application of the method for testing the interfacial adhesion of teeth or tooth-like materials in the design of anti-adhesion materials.

[0029] In one implementation of this application, the application of the method in the design of anti-adhesion materials specifically includes: using the method of this application to test the adhesion force on the surface of the material to be tested; using machine learning to analyze the nonlinear relationship between the obtained adhesion force and the surface physicochemical properties of the material to be tested; thereby guiding the design of anti-adhesion materials.

[0030] The fourth aspect of this application discloses a method for designing anti-adhesion materials, which includes testing the adhesion force on the surface of the material to be tested using the method of this application for testing the interfacial adhesion force of tooth or tooth-like materials, and using machine learning to analyze the nonlinear relationship between the obtained adhesion force and the surface physicochemical properties of the material to be tested, thereby guiding the design of anti-adhesion materials.

[0031] In this application, surface physicochemical properties include surface energy and / or surface topology.

[0032] The fifth aspect of this application discloses a method for testing the adhesion of dental plaque biofilm, comprising the following steps:

[0033] The probe is brought into contact with the dental plaque biofilm to be tested, and a load within a specified range is applied to the probe, which slides from the surface of the dental plaque biofilm at a preset speed.

[0034] The peak adhesion force required for plaque biofilm separation is measured by a force sensor integrated with a probe, and the change in the contact area with the detection interface is measured in real time by an optical system.

[0035] The specified load range is the minimum force required to break the surface of the dental plaque biofilm to be tested, up to the force required to penetrate the surface of the mat and substrate to which the dental plaque biofilm is attached, but without causing obvious scratches to the mat and substrate.

[0036] It should be noted that the plaque biofilm adhesion testing method of this application is actually the application of the tooth or tooth-like material interface adhesion testing method of this application in the plaque adhesion testing; this application not only quantitatively tests the adhesion of plaque, but also uses a high-resolution microscopic optical system for visual evaluation.

[0037] It should also be noted that the adhesion test method for dental plaque biofilm in this application is based on the following principle: a precisely controlled micron-sized probe makes controlled contact with the dental plaque biofilm, applies a slight load within a certain range, and slides vertically upward from the plaque surface at a constant speed to separate it; the peak tensile force (adhesion force) required for separation is measured by an integrated force sensor, and the change in the interface contact area is observed and measured in real time by a high-resolution microscopic optical system, thereby providing a quantitative and visual evaluation of the interfacial micro-adhesion strength of dental plaque.

[0038] In one implementation of this application, the substrate is hydroxyapatite; therefore, the load within a specified range, i.e., the linear loading force within a specified range, is specifically the minimum force required to break the surface of the dental plaque biofilm to be tested, up to the force required to penetrate the surface of the hydroxyapatite sheet without causing obvious scratches to the surface of the hydroxyapatite sheet.

[0039] In one implementation of this application, the dental plaque adhesion force testing method specifically uses the MAT-2 micro adhesion force tester to test the dental plaque adhesion force. The MAT-2 micro adhesion force tester is equipped with a capacitive mechanical sensor with a test range of 5mN-1N and a maximum resolution of 0.015mN. It also has a high-magnification microscope system for the lower sample to observe and measure changes in the interface contact area.

[0040] It should be noted that, in one implementation of this application, the specific dental plaque adhesion force is measured using the MAT-2 micro-adhesion force tester. The instrument is equipped with a capacitive mechanical sensor with a testing range of 5mN-1N, and includes a high-powered microscope system for observing the contact area. It is understood that, under the same inventive concept, other devices with similar functions may be used, and no specific limitations are made here.

[0041] In one implementation of this application, the load within the specified range is 0.03-0.5N.

[0042] In one implementation of this application, the loading rate of the load within a specified range is 0.1-10 N / min, for example, 0.94 N / min.

[0043] In one implementation of this application, the preset speed is 1μm / min-1mm / min, for example, 400μm / min.

[0044] In one implementation of this application, the probe slides a distance of 0.5-5 mm, for example, 2 mm, on the surface of dental plaque biofilm.

[0045] The sixth aspect of this application discloses the application of the method of this application for testing the interfacial adhesion of teeth or tooth-like materials, or the method of this application for testing the adhesion of dental plaque biofilms, in the preparation of in vitro biological models for performance evaluation of oral care devices.

[0046] Due to the adoption of the above technical solutions, the beneficial effects of this application are as follows:

[0047] The method for testing the interfacial adhesion of teeth or tooth-like materials in this application can test the surface adhesion of in vitro biological models, and the test results are in good agreement with the actual surface adhesion data of teeth, bridging the gap between in vitro biological model test data and clinical data. The method of this application can quantitatively analyze the surface adhesion performance of in vitro biological models, which not only ensures the comparability of data from different studies, but also facilitates the development of unified in vitro testing standards, laying the foundation for promoting industry standardization and normalization. Attached Figure Description

[0048] Figure 1 These are photographs of extracted human teeth from embodiments of this application;

[0049] Figure 2 This is a photograph of an extracted human tooth, cut into small pieces and embedded in an epoxy resin cylinder, after polishing, according to an embodiment of this application.

[0050] Figures 3 to 5 These are the results of three tests on the extracted human teeth in the embodiments of this application. The left image shows the scratch test results, and the right image shows the corresponding microscopic observation results.

[0051] Figures 6 to 8 These are the results of three tests on HAP sample 1 in this embodiment of the application. The upper figure shows the scratch test results, and the lower figure shows the corresponding microscopic observation results.

[0052] Figures 9 to 11 These are the results of three tests on HAP sample 2 in this embodiment of the application. The upper figure shows the scratch test results, and the lower figure shows the corresponding microscopic observation results.

[0053] Figure 12 These are images of optical microscope data analysis from embodiments of this application. Detailed Implementation

[0054] The adhesion data of existing in vitro biological models (such as single-species biofilms, dental plaque, soft plaque, dental calculus, or similar simulated materials) are unknown compared to the adhesion in the real human oral cavity. This makes it impossible to make a basis for judging the correlation between existing in vitro test results and clinical effects, resulting in distortion in the evaluation of the anti-adhesion performance of dental materials.

[0055] Therefore, this application creatively develops a new method for testing the interfacial adhesion of teeth or tooth-like materials, including using a micro-adhesion force tester or atomic force microscope to perform a scratch test on the surface layer under test using a linear loading force within a specified range; the scratch is divided into three stages according to the changes in the scratch. In the first stage, the height change is detected throughout the process using a linear loading force within a specified range. In the second stage, there is slight and discontinuous scratching without obvious furrows. In the third stage, obvious furrows appear, but the furrows are narrow, and no obvious scratches formed by the friction between the mating material and the substrate can be observed in the center of the furrows.

[0056] The abrupt change point load that breaks down the surface layer under test is calculated using the following formula and used as an indicator to evaluate the adhesion of the surface layer under test.

[0057] The load at the abrupt change point = the profile height of the third stage ÷ the total profile height of the first stage × the maximum load of the linear loading force within the specified range;

[0058] A linear loading force within a specified range, wherein the minimum loading force is the minimum force that can break through the surface layer under test, and the maximum loading force is the loading force that can penetrate the surface layer under test without causing obvious scratches to the teeth or tooth-like materials.

[0059] The innovations of this application include: 1. Establishing the first method for evaluating the micro-adhesion of dental or tooth-like material interfaces in the oral cavity, bridging the data gap between the laboratory and clinical settings. 2. Contributing to industry standardization. Currently, many paints and resin-based coatings on the market claim to act as plaque mimics, but without any supporting evidence. The testing method in this application can quantitatively evaluate these plaque mimics, thereby promoting the standardization of plaque mimics or other similar materials. 3. Correlating the micro-adhesion data obtained from this application with macro-clinical results (plaque index, periodontal index, etc.) to construct predictive mathematical models; for example, using the adhesion threshold measured by AFM to predict the anti-plaque durability of a specific material surface in the oral environment. 4. Utilizing machine learning to analyze the nonlinear relationship between the adhesion force tested in this application and surface physicochemical properties (such as surface energy and topology), thereby guiding the design of anti-adhesion materials.

[0060] The testing method described in this application has the following advantages: 1. Controllability and standardization. 2. Suitable for high-throughput screening of appropriate materials (e.g., the development of materials that resist oral bacterial adhesion), providing a reference for the construction of in vitro biological models. 3. Visualization and quantification of the dynamic process; the structure of the test interface is observed using a microscope, surface profilometer, or white light interferometer; adhesion force is measured using an atomic force microscope (AFM) or microadhesion analyzer; adhesion strength is quantified through shear force and peel force tests, and an adhesion force database is established. 4. Promotes a standardized testing system, facilitating the development of unified in vitro testing standards, promoting industry norms, and ensuring the comparability of data from different studies.

[0061] The interfaces applicable to the testing method in this application include:

[0062] Tooth or tooth-like materials: human extracted teeth (including enamel and dentin), animal extracted teeth (including enamel and dentin), teeth or teeth of any shape with hydroxyapatite as the main component, various types of resins, various metal alloys, bioactive glass, ceramics (including but not limited to zirconia ceramics and lithium disilicate glass ceramics), and polymer materials (Peek, PMMA).

[0063] The test method of this application can test surface layers including dental plaque, soft deposits, tartar, or similar simulants: plaque, stains, soft deposits, tartar, and tartar naturally present on human or animal teeth; plaque constructed through in vitro culture (including, but not limited to, plaque biofilms formed by Streptococcus mutans, Streptococcus sanguinis, Actinomyces, etc., and, in addition, plaque biofilms simulated by sodium carboxymethyl cellulose and hydroxyethyl cellulose); soft deposits, tartar, and tartar (including, but not limited to, chemical deposits simulated by hydroxyapatite, octacalcium phosphate, calcium carbonate, and collagen); and simulated coatings of water-based paints or coatings (including, but not limited to, water-based acrylic resins, water-based matting agents, water-based cobalt blue pastes, water-based toluene diethyl cyanate, and water-based propylene glycol methyl ether acetate).

[0064] This application also relates to the evaluation of the differences in the plaque biofilm removal effects of tested electric toothbrushes using a bacterial strain-carrier method. Using common oral mutagenesis streptococci as the research tool, the bacteria were treated and exposed to hydroxyapatite sheets to construct a plaque biofilm model. Different electric toothbrushes were used as cleaning tools to brush the surface of the plaque biofilm model. The brushing time, direction, and pressure were fixed during the test. Quantitative analysis of the plaque biofilm before and after brushing was performed, and the plaque removal rate was calculated by measuring the absorbance of the samples before and after brushing. Finally, the removal effect of the electric toothbrush was evaluated based on the removal rate.

[0065] The specific evaluation methods utilize the following equipment, reagents, and consumables:

[0066] Human hand brushing motion simulation device: This device can realize the function of brushing motion on the surface of teeth or similar tooth materials, and can apply a corresponding level of contact pressure.

[0067] Plaque content analysis equipment: ELISA reader (manufacturer: Nanjing Detie, model: HBS-ScanX) or other color quantitative analysis equipment, using a wavelength of 575nm.

[0068] Micro-volume liquid mixing equipment: Micro-oscillator (manufacturer: Jintan Dadi, model: MM-1), operating speed is 0-1500rpm.

[0069] reagents and consumables

[0070] — Hydroxyapatite flakes (Manufacturer: Sichuan Baiameng Bioactive Materials Co., Ltd., Specifications: Φ12±0.5*2mm)

[0071] —Glacial acetic acid (Manufacturer: Maclean, CAS: 64-19-7, Concentration used: 33% (v / v))

[0072] —Phosphate-buffered saline (PBS) (Manufacturer: Boster Biological, Ingredients: Potassium dihydrogen phosphate, Sodium chloride, Disodium hydrogen phosphate, pH: 7.2-7.4, Concentration used: 0.1 mol / L)

[0073] —Ultrapure water (prepared in the laboratory, meeting the requirements of GB / T 6682-2008 for water specifications and test methods in analytical laboratories, meeting the requirements for Class III water)

[0074] The experimental conditions for this evaluation method, including the experimental procedures for evaluating the effectiveness of dental plaque biofilm removal, can all be carried out at room temperature, as detailed below:

[0075] Temperature: 18-26℃;

[0076] Humidity: 45-65%.

[0077] The operational steps of this evaluation method include:

[0078] 1. Test Grouping

[0079] Table 1 Test Groups

[0080]

[0081] 2. Dental plaque biofilm culture:

[0082] Oral flora are mostly divided into periodontal pathogens and cariogenic bacteria. Among cariogenic bacteria, *Streptococcus mutans* has strong adhesion and is easy to culture. By changing the culture conditions, a better dental plaque biofilm can be obtained. In the oral cavity, *Streptococcus mutans* adheres to and aggregates on the acquired membrane carrier formed by saliva. It uses sucrose in the oral cavity to form a large amount of extracellular polysaccharides. *Streptococcus mutans* binds to the extracellular polysaccharides and adheres to the tooth surface. At the same time, *Streptococcus mutans* releases a large amount of extracellular substances, which participate in the formation of biofilm.

[0083] This culture method involves activating and culturing *Streptococcus mutans* in a brain and heart infusion medium to obtain a bacterial enrichment broth. Hydroxyapatite tablets are then immersed in artificial saliva for culture to obtain a carrier with an acquired membrane. The activated *Streptococcus mutans* broth, the carrier with the acquired membrane, and a brain and heart infusion medium containing sucrose are co-cultured, allowing *Streptococcus mutans* to adhere to the carrier containing the acquired membrane and form a dental plaque biofilm model. The dental plaque biofilm model is collected, washed to remove excess airborne bacteria and culture medium, and stained with a dye to obtain a dental plaque biofilm model for evaluating the cleaning effect of an electric toothbrush.

[0084] 3. Evaluation of the cleaning effect of dental plaque biofilm

[0085] 1) Electric toothbrushes must be tested by brushing while fully charged.

[0086] 2) Pressure Calibration Process: First, turn the electric toothbrush off. Fix the blank hydroxyapatite sheet carrier onto the 3D printed part. Fix the electric toothbrush to the human brushing motion simulation device by adjusting the electric toothbrush fixing component, so that the electric toothbrush does not move in the horizontal and vertical directions. Adjust the contact area between the hydroxyapatite sheet and the electric toothbrush (adjust the pressure by moving the height of the 3D printed part up and down). Turn on the human brushing motion simulation device (set the brushing speed to 4mm / s) and make the electric toothbrush make a linear reciprocating motion. Generally, the middle position of the 3D printed part is used as the pressure calibration position. The electric toothbrush moves forward to the same position and in the same direction each time. After the pressure value stabilizes at 200±20g, the formal test can begin.

[0087] 3) Place the hydroxyapatite sheets in a forced-air drying oven and dry at 37℃ for about 10-30 minutes (if the ambient humidity is around 45%, the drying time is about 10 minutes; if the ambient humidity is around 65%, the drying time is about 25 minutes). Fix the three hydroxyapatite sheet carriers with dental plaque biofilm on their surfaces in the same group vertically upwards in a straight line inside the groove of the 3D printed part. Add a blank hydroxyapatite sheet on each side of the three parallel hydroxyapatite sheets to buffer the deformation caused by the bristles. Before testing, immerse the electric toothbrush head in a 10mL beaker filled with ultrapure water for 10 seconds, with the base of the bristles flush with the water surface. After immersion, gently wipe the brush head with a non-woven cloth to ensure that no water droplets are hanging from the bristles. Three hydroxyapatite tablets were repeatedly used under the same conditions to simulate the action of brushing teeth with a human hand. The electric toothbrush moved in a single linear motion over the 3D printed part. One complete test process was completed from the start of brushing the first blank hydroxyapatite tablet to the completion of brushing the last blank hydroxyapatite tablet. After the brushing was completed, the brushed hydroxyapatite tablets were transferred to 2mL of PBS for temporary storage. After all tests were completed, the hydroxyapatite tablets were placed in a new 24-well plate, and water was added to the well to clean them, simulating the human brushing process. The cleaning was repeated 3 times, 2mL each time.

[0088] 4) Use the same brushing process for each electric toothbrush, and replace the brush head before each brushing test.

[0089] 4. Quantitative analysis of dental plaque biofilm

[0090] After brushing, all hydroxyapatite tablets were placed in a drying oven at 37°C for approximately 20-30 minutes. Once the surface was completely dry, the brushed samples from each group were placed in a new 24-well plate. 1 mL of 33% glacial acetic acid was added to each well. The plate was then placed on a shaker. Before shaking, the bottom of the plate was secured with sealing film or tape. Shaking was performed at room temperature for 60 minutes at a speed below 500 rpm. The endpoint was determined by the complete elution of the bacterial film from the surface of the hydroxyapatite tablets. Complete elution was considered achieved when the color of the hydroxyapatite tablets no longer changed after elution. After shaking, the solution from each well or tube was transferred to a new 2 mL centrifuge tube and centrifuged at 11000-15000 rpm for 5 minutes. 200 μL or 100 μL of glacial acetic acid-gentian violet solution was then transferred from each centrifuge tube to a new 96-well plate. Three wells were measured in parallel per tube. The absorbance was measured at 575 nm using a microplate reader.

[0091] 5. Result Calculation: Formula for Calculating Dental Plaque Biofilm Removal Rate

[0092]

[0093] In the formula:

[0094] Before OD: Negative control group, absorbance value before any brushing test.

[0095] After OD: Sample group, using the absorbance values ​​of the sample after processing.

[0096] 6. Statistical Analysis of Results

[0097] It is recommended to use GraphPad Prism or SPSS for graph analysis, and the results are expressed as Mean ± SD. Comparisons between groups were performed using t-tests, and all statistical analyses were two-tailed. P < 0.05 was considered statistically significant, and P < 0.01 was considered highly statistically significant; the smaller the P value, the more significant the difference.

[0098] 7. Result Determination

[0099] Each batch of experiments must include a blank control group, a negative control group, and a sample group. The groups set up in the experiment must meet the following requirements:

[0100] Compared with the blank control group, the absorbance value of hydroxyapatite tablets in the negative control group was significantly increased (P<0.05), which is statistically significant, proving that the experimental modeling conditions were effective.

[0101] The absorbance values ​​for the blank control group and the negative control group are as follows:

[0102] When the test solution volume was 200 μL, the OD575 nm of the blank control group was 0.3–1.1, and that of the negative control group was 3.0–3.9.

[0103] When the test solution volume was 100 μL, the OD575 nm of the blank control group was 0.05–0.5, and the OD575 nm of the negative control group was 1.5–2.1.

[0104] The recommended values ​​are approximately 15 μm in thickness for the dental plaque biofilm and approximately 0.24–0.26 N for plaque adhesion.

[0105] The stability of the test must meet the following requirements:

[0106] In batch experiments, the standard deviation (SD) and average value of absorbance between replicates in each group are calculated, and the coefficient of variation (CV) is calculated. The CV value is ≤20%.

[0107]

[0108] In the formula:

[0109] OD standard deviation – standard deviation of absorbance value at 575nm

[0110] Average OD value – the average absorbance value at 575 nm

[0111] The validity of a sample test must meet the following criteria:

[0112] Compared with the negative control group, the absorbance value of the sample group was significantly lower (P<0.05), indicating a statistically significant difference. This suggests that the sample group can effectively remove dental plaque.

[0113] In some implementations, the test subject can be other oral care devices such as water flossers or interdental brushes, used to evaluate the cleaning effect on the test subject.

[0114] The present application will be further described in detail below through specific embodiments. The following embodiments are only for further illustration of the present application and should not be construed as limiting the present application.

[0115] Unless otherwise specified, the technical means used in the embodiments are conventional means well known to those skilled in the art.

[0116] Example

[0117] I. Plaque Adhesion Test of Exposed Human Teeth

[0118] 1. Sample to be tested

[0119] This example uses extracted human teeth with dental plaque as the test sample for adhesion testing. A photograph of the test sample is shown below. Figure 1 As shown.

[0120] 2. Sample processing

[0121] Tooth samples with plaque growth were cut into small pieces and embedded in epoxy resin cylinders using an epoxy resin casting method. The tooth surfaces were then polished with 1000-grit sandpaper to create a plane that fully exposes the plaque. The polished samples looked like... Figure 2 As shown. Figure 2 In the image, the black part in the middle represents dental plaque.

[0122] Meanwhile, using an optical microscope to observe the side of the cut tooth sample, a thick transition layer region was found between the tooth sample and the test surface, with the thickest layer exceeding 300 μm.

[0123] 3. Test methods and instruments used

[0124] The adhesion testing method for dental plaque biofilm in this example includes:

[0125] The probe is brought into contact with the dental plaque biofilm to be tested, and a load within a specified range is applied to the probe, which slides from the surface of the dental plaque biofilm at a preset speed.

[0126] The peak adhesion force required for plaque biofilm separation is measured by a force sensor integrated with the probe, and the change in the contact area between the probe and the detection interface is measured in real time by an optical system.

[0127] The specified load range is the minimum force required to break the surface of the dental plaque biofilm to be tested, up to the force required to penetrate the surface of the mat and substrate to which the dental plaque biofilm is attached, but without causing obvious scratches to the mat and substrate.

[0128] Because the plaque coverage area was small and uneven in this experiment, the scratch length was changed to 0.5 mm.

[0129] Instrument used: MAT-2 micro-adhesion force tester. The MAT-2 micro-adhesion force tester is equipped with a capacitive mechanical sensor with a test range of 5mN-1N. It also has a high-magnification microscope system for the sample to observe and measure changes in the interface contact area.

[0130] Indenter: A hemispherical diamond indenter with a radius of 50 μm (0.785010^-4 cm) 2 )

[0131] Sliding distance: 0.5mm

[0132] Loading force range: 0.03~0.5N

[0133] Sliding speed: 1000 μm / min; Loading rate: 0.94 N / min

[0134] Scratch procedure: A linear loading force of 0.03–0.5 N is used to perform a scratch test on the test surface. The force selection range requires that the peak value can penetrate the test surface. The scratches show a stage change and are divided into three stages. The first stage is a full height change of 0.03–0.5 N. The second stage is a slight and discontinuous scratch without obvious furrows. The third stage has obvious furrows, but the furrow width is narrow, and no obvious scratches formed by the friction between the substrate and the target material can be observed in the center of the furrow.

[0135] Evaluation method: The abrupt change load that breaks down the test surface layer is calculated according to the following formula as an index for evaluating the adhesion of the test surface layer.

[0136] The load at the mutation point = the profile height of the third stage ÷ the total profile height of the first stage × the maximum load of the linear loading force within the specified range.

[0137] 4. Surface test results

[0138] A suitable region was selected for testing the sample, with each sample tested three times. No abrupt change point appeared when testing in one region, suggesting a possible connection to substrate changes; therefore, a second region was selected. Three scratch tests were performed in the second region, and the results were compared with microscopic images. Microscopic examination confirmed that the scratch width suddenly increased after the load exceeded the abrupt change point, which was positively correlated with the decrease in scratch depth. The results of the three tests and microscopic observations are as follows: Figures 3 to 5 As shown.

[0139] 1) First test

[0140] The test results and microscopic observation results are as follows: Figure 3 As shown.

[0141] The load at the mutation point = (325.3 / 687.3) * 0.5 N = 0.24 N

[0142] 2) Second test

[0143] The test results and microscopic observation results are as follows: Figure 4 As shown.

[0144] The load at the mutation point = (351.8 / 695.8) * 0.5 N = 0.25 N

[0145] 3) Third test

[0146] The test results and microscopic observation results are as follows: Figure 5 As shown.

[0147] The load at the mutation point = (314.7 / 682.7) * 0.5 N = 0.23 N

[0148] The results of the three tests showed that the mutation point load was between 0.23 and 0.25 N, which was similar to the results of the subsequent tests on the hydroxyapatite flake samples.

[0149] II. Plaque Adhesion of HAP

[0150] 1. Sample to be tested

[0151] The HAP test sample in this example, namely the hydroxyapatite flake sample, was prepared by referring to patent application 202210689560.7, which describes an in vitro dental plaque biofilm model and method for evaluating product efficacy. An in vitro HAP dental plaque model was established according to the method in the patent application for adhesion testing.

[0152] 2. Testing Methods

[0153] The same equipment used for extracted human teeth was employed for testing, and the specific parameters are as follows:

[0154] Sliding distance: 2mm

[0155] Loading force range: 0.03~0.5N

[0156] Sliding speed: 400 μm / min; Loading rate: 0.94 N / min

[0157] Scratch procedure: A linear loading force of 0.03–0.5 N is used to perform a scratch test on the test surface. The force selection range requires that the peak value can penetrate the test surface. The scratches show a stage change and are divided into three stages. The first stage is a full height change of 0.03–0.5 N. The second stage is a slight and discontinuous scratch without obvious furrows. The third stage has obvious furrows, but the furrow width is narrow, and no obvious scratches formed by the friction between the substrate and the target material can be observed in the center of the furrow.

[0158] Evaluation method: The abrupt change load that breaks down the test surface layer is calculated according to the following formula as an index for evaluating the adhesion of the test surface layer.

[0159] The load at the mutation point = the profile height of the third stage ÷ the total profile height of the first stage × the maximum load of the linear loading force within the specified range.

[0160] 3. Test Results

[0161] Two parallel samples were tested, with each sample tested three times. The result of the first test for sample 1 is as follows: Figure 6 As shown, the mutation point load is 0.25 N; the second test results for sample 1 are as follows. Figure 7 As shown, the mutation point load is 0.26 N; the third test result for sample 1 is as follows. Figure 8As shown, the mutation point load is 0.26 N; the first test results for sample 2 are as follows. Figure 9 As shown, the mutation point load is 0.26 N; the second test results for sample 2 are as follows. Figure 10 As shown, the mutation point load is 0.29 N; the third test result for sample 2 is as follows. Figure 11 As shown, the load at the mutation point is 0.24 N.

[0162] 3. Optical Microscopy Data Analysis

[0163] Optical microscopy data analysis of the scratches from the third test of Sample 2 showed that, with increasing load, the scratch depth abruptly changed at two locations: 0.1–0.13 N and 0.24–0.29 N. It is speculated that the damage mechanism of the scratches changed near these two abrupt change points. The optical microscopy data also revealed that the scratches gradually changed from discontinuous to continuous, exhibiting a cutting morphology, and eventually the entire coating peeled off. Therefore, the coating adhesion can be evaluated using the load corresponding to the two abrupt change points.

[0164] like Figure 12 As shown, the total length of the scratch is 2.1 mm, with abrupt changes at 0.59 mm and 1.04 mm, corresponding to stresses of 0.59 / 2.1*0.5 = 0.14 N and 1.04 / 2.1*0.5 = 0.24 N. Slight and discontinuous scratching occurs within the range of 0.03–0.14 N, without obvious grooves, suggesting the scratches primarily occur on the surface. Obvious grooves appear between 0.14 and 0.25 N, but the grooves are narrow, and no obvious scratches from the substrate are observed at the center of the grooves, which can be attributed to the elastic recovery of the surface. Above 0.24 N, the grooves suddenly widen, and obvious scratches appear at the center, suggesting the coating has been penetrated, exposing the hydroxyapatite substrate (3.1 * 10⁻⁵ N / cm²). 2 ).

[0165] 4. Correlation analysis

[0166] Correlation analysis was performed on the plaque adhesion test results of extracted human teeth and the plaque adhesion test results of HAP (Hyperplasia of the Otolaryngology-Adhesive-Polymer). The results showed that the correlation coefficient between the two sets of data was 1, proving that there was a significant correlation between the two sets of results. This indicates that the adhesion test data obtained by the adhesion test method in this example has a good consistency with the test data of real human teeth, bridging the gap between in vitro biological model test data and clinical data.

[0167] The above description, in conjunction with specific embodiments, provides a further detailed explanation of this application and should not be construed as limiting the specific implementation of this application to these descriptions. Those skilled in the art to which this application pertains can make several simple deductions or substitutions without departing from the concept of this application.

Claims

1. A method for testing the interfacial adhesion of teeth or tooth-like materials, characterized in that: This includes using a micro-adhesion force tester or atomic force microscope to perform scratch tests on the surface layer under test using a linear loading force within a specified range; the scratches are divided into three stages based on the changes in the scratches. In the first stage, the height change is detected throughout the entire process using a linear loading force within a specified range. In the second stage, there is slight and discontinuous scratching without obvious furrows. In the third stage, obvious furrows appear, but the furrows are narrow, and no obvious scratches formed by the friction between the mating material and the substrate can be observed in the center of the furrows. The abrupt change point load that breaks down the surface layer under test is calculated using the following formula and used as an indicator to evaluate the adhesion of the surface layer under test. The load at the abrupt change point = the profile height of the third stage ÷ the total profile height of the first stage × the maximum load of the linear loading force within the specified range; The specified range of linear loading forces has a minimum loading force that can break through the surface layer under test, and a maximum loading force that can penetrate the surface layer under test without causing obvious scratches to the teeth or tooth-like materials.

2. The method according to claim 1, characterized in that: The specified linear loading force is 0.03-0.5N.

3. The method according to claim 1, characterized in that: The teeth mentioned are human teeth or other animal teeth extracted from the body.

4. The method according to claim 1, characterized in that: The aforementioned dental materials include at least one of the following: materials that form teeth or other arbitrary shapes with hydroxyapatite as the main component, resin materials, metal alloy materials, bioactive glass materials, ceramic materials, and polymer materials.

5. The method according to claim 4, characterized in that: The ceramic material includes at least one of zirconia ceramic and lithium disilicate glass ceramic.

6. The method according to claim 4, characterized in that: The polymer material includes at least one of polyetheretherketone and polymethyl methacrylate.

7. The method according to claim 1, characterized in that: The surface layer to be tested is dental plaque, soft deposits, tartar, or similar simulants.

8. The method according to claim 7, characterized in that: The surface layer to be tested is plaque, stains, soft deposits, tartar, or calculus that is present on the extracted teeth of humans or animals.

9. The method according to claim 7, characterized in that: The surface layer to be tested is plaque, soft deposits, dental tartar, or dental calculus constructed through in vitro culture, or a simulated coating prepared from water-based paint or coating.

10. The method according to claim 9, characterized in that: The plaques constructed in vitro are plaque biofilms constructed from bacteria cultured in vitro, or plaque biofilms simulated by sodium carboxymethyl cellulose and / or hydroxyethyl cellulose.

11. The method according to claim 10, characterized in that: The in vitro cultured bacteria include at least one of Streptococcus mutans, Streptococcus sanguinis, and Actinomycetes.

12. The method according to claim 9, characterized in that: The dental calculus constructed in vitro includes chemical deposition mimics formed from at least one of hydroxyapatite, octacalcium phosphate, calcium carbonate, and collagen.

13. The method according to claim 9, characterized in that: The water-based paint or coating includes at least one of water-based acrylic resin, water-based matting agent, water-based cobalt blue paste, water-based toluene diethyl cyanate, and water-based propylene glycol methyl ether acetate.

14. The application of the method of any one of claims 1-13 in predicting the antiplaque durability of a particular material surface in an oral environment.

15. The application of the method according to any one of claims 1-13 in the design of anti-adhesion materials.

16. The application according to claim 15, characterized in that: This includes testing the adhesion force on the surface of a material under test using the method described in any one of claims 1-13, and using machine learning to analyze the nonlinear relationship between the obtained adhesion force and the surface physicochemical properties of the material under test, thereby guiding the design of anti-adhesion materials.

17. The application according to claim 16, characterized in that: The surface physicochemical properties include surface energy and / or surface topology.

18. A method for designing anti-adhesion materials, characterized in that: This includes testing the adhesion force on the surface of a material under test using the method described in any one of claims 1-13, and using machine learning to analyze the nonlinear relationship between the obtained adhesion force and the surface physicochemical properties of the material under test, thereby guiding the design of anti-adhesion materials.

19. The method according to claim 18, characterized in that: The surface physicochemical properties include surface energy and / or surface topology.

20. A method for testing the adhesion of dental plaque biofilm, characterized in that: The method includes, The probe is brought into contact with the dental plaque biofilm to be tested, and a load within a specified range is applied to the probe, which slides from the surface of the dental plaque biofilm at a preset speed. The peak adhesion force required for plaque biofilm separation is measured by a force sensor integrated with the probe, and the change in the contact area between the probe and the detection interface is measured in real time by an optical system. The specified load range is the minimum force required to break the surface of the dental plaque biofilm to be tested, up to the force required to penetrate the surface of the mat and substrate to which the dental plaque biofilm is attached, but without causing obvious scratches to the mat and substrate.

21. The adhesion force testing method according to claim 20, characterized in that: The matrix is ​​hydroxyapatite; The specified load range, i.e. the linear loading force within the specified range, is specifically the minimum force required to break through the surface of the dental plaque biofilm to be tested, up to the force required to penetrate the surface of the hydroxyapatite sheet without causing obvious scratches to the surface of the hydroxyapatite sheet.

22. The adhesion test method according to claim 20, characterized in that: This includes using the MAT-2 micro-adhesion tester to test dental plaque adhesion. The MAT-2 micro-adhesion force tester is equipped with a capacitive mechanical sensor with a testing range of 5mN-1N. It also features a high-magnification microscope system for the lower sample, used to observe and measure changes in the interface contact area.

23. The adhesion force testing method according to claim 20, characterized in that: The specified load range is 0.03-0.5N.

24. The adhesion test method according to claim 23, characterized in that: The loading rate of the load within the specified range is 0.1-10 N / min.

25. The adhesion force testing method according to claim 20, characterized in that: The preset speed is 1μm / min-1mm / min.

26. The adhesion force testing method according to claim 20, characterized in that: The probe slides a distance of 0.5-5 mm on the surface of the dental plaque biofilm.

27. The use of the method according to any one of claims 1-13 or the adhesion testing method according to any one of claims 20-26 in the preparation of an in vitro biological model for performance evaluation of oral care devices.

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