CN112618802A - Fluorine-containing antibacterial invisible appliance for improving enamel demineralization and preparation method thereof - Google Patents
Fluorine-containing antibacterial invisible appliance for improving enamel demineralization and preparation method thereof Download PDFInfo
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- CN112618802A CN112618802A CN202011424135.2A CN202011424135A CN112618802A CN 112618802 A CN112618802 A CN 112618802A CN 202011424135 A CN202011424135 A CN 202011424135A CN 112618802 A CN112618802 A CN 112618802A
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- fluorine
- solution
- antibacterial
- lysozyme
- appliance
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- 230000000844 anti-bacterial effect Effects 0.000 title claims abstract description 72
- 229910052731 fluorine Inorganic materials 0.000 title claims abstract description 67
- 239000011737 fluorine Substances 0.000 title claims abstract description 67
- YCKRFDGAMUMZLT-UHFFFAOYSA-N Fluorine atom Chemical compound [F] YCKRFDGAMUMZLT-UHFFFAOYSA-N 0.000 title claims abstract description 65
- 230000002328 demineralizing effect Effects 0.000 title claims abstract description 35
- 238000005115 demineralization Methods 0.000 title claims abstract description 32
- 210000003298 dental enamel Anatomy 0.000 title claims abstract description 20
- 238000002360 preparation method Methods 0.000 title claims abstract description 17
- 102000016943 Muramidase Human genes 0.000 claims abstract description 38
- 108010014251 Muramidase Proteins 0.000 claims abstract description 38
- 108010062010 N-Acetylmuramoyl-L-alanine Amidase Proteins 0.000 claims abstract description 38
- 239000004325 lysozyme Substances 0.000 claims abstract description 38
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- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 claims abstract description 30
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- WUKWITHWXAAZEY-UHFFFAOYSA-L calcium difluoride Chemical group [F-].[F-].[Ca+2] WUKWITHWXAAZEY-UHFFFAOYSA-L 0.000 claims description 2
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- 238000000034 method Methods 0.000 abstract description 20
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- 239000010410 layer Substances 0.000 description 29
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 15
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 12
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- 238000000338 in vitro Methods 0.000 description 8
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- -1 fluorine ions Chemical class 0.000 description 3
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- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 229920002385 Sodium hyaluronate Polymers 0.000 description 2
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
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- 229940010747 sodium hyaluronate Drugs 0.000 description 2
- YWIVKILSMZOHHF-QJZPQSOGSA-N sodium;(2s,3s,4s,5r,6r)-6-[(2s,3r,4r,5s,6r)-3-acetamido-2-[(2s,3s,4r,5r,6r)-6-[(2r,3r,4r,5s,6r)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2- Chemical compound [Na+].CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 YWIVKILSMZOHHF-QJZPQSOGSA-N 0.000 description 2
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- 108010058846 Ovalbumin Proteins 0.000 description 1
- 229910010443 TiO2-xNx Inorganic materials 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 239000000783 alginic acid Substances 0.000 description 1
- 229920000615 alginic acid Polymers 0.000 description 1
- 229960001126 alginic acid Drugs 0.000 description 1
- 235000010443 alginic acid Nutrition 0.000 description 1
- 150000004781 alginic acids Chemical class 0.000 description 1
- 230000022972 amelogenesis Effects 0.000 description 1
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- DBLXOVFQHHSKRC-UHFFFAOYSA-N ethanesulfonic acid;2-piperazin-1-ylethanol Chemical compound CCS(O)(=O)=O.OCCN1CCNCC1 DBLXOVFQHHSKRC-UHFFFAOYSA-N 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
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Abstract
The invention discloses a fluorine-containing antibacterial invisible appliance for improving enamel demineralization and a preparation method thereof. According to the invention, lysozyme and fluorine-containing hyaluronic acid are fixed by a layer-by-layer self-assembly method, so that the fluorine-containing antibacterial invisible appliance with lysozyme antibacterial performance and fluorine remineralization promoting performance is obtained, and the lysozyme and the hyaluronic acid have good biocompatibility, low cost and high efficiency. The fluorine-containing antibacterial appliance prepared by the invention can promote the remineralization of enamel, can also inhibit the occurrence and development of gingivitis, caries, pulposis and periodontitis related to bacteria, has simple, high-efficiency and pollution-free preparation process and wide clinical application prospect.
Description
Technical Field
The invention belongs to the technical field of material modification and oral medicine. More particularly, relates to a fluorine-containing antibacterial invisible appliance for improving enamel demineralization and a preparation method thereof.
Background
The invisible orthodontic appliance has become one of the mainstream orthodontic means because of the advantages of self-taking, wearing, easy cleaning, beautiful appearance, comfort, etc. However, with the expansion of the population of patients wearing the invisible orthodontic appliance, complications caused by the invisible orthodontic appliance are gradually exposed, for example, the invisible orthodontic appliance has large tooth surface area and long wearing time, and the microenvironment in the oral cavity can be influenced, so that the abundance of the flora is remarkably increased; plaque accumulation can also be caused by acid erosion to the tooth surface and rough surfaces around the attachments in the attachment process of the invisible appliance; when oral hygiene is poorly controlled, plaque accumulates in large amounts in the relatively confined space formed by the appliance and the tooth surface, thereby increasing the risk of gingivitis, enamel demineralization, and even pulposis and periodontal disease, especially in young people with poor self-restraint and self-oral health awareness. Studies have shown that enamel demineralization occurs at greater than 60% in adolescent patients who lack health guidance and oral hygiene interventions after wearing an invisible appliance.
So far, many orthodontic appliances with antibacterial effect have been reported to be used for preventing and treating related diseases caused by bacteria in the orthodontic process, for example, chinese patent application 201110192472.8 discloses a method for attaching a nitrogen-doped TiO2-xNx film on the surface of a metal bracket by a radio frequency magnetron sputtering method, but the method has high reaction temperature and is difficult to popularize in invisible orthodontic appliances; chinese patent application 201911357625.2 provides a method for manufacturing silver-doped titanium dioxide nano-antibacterial composite coating on an invisible appliance, but the used magnetron sputtering ion plating equipment is expensive and complicated in procedure, and due to the introduction of inorganic nano-materials, the good biocompatibility of the appliance is difficult to ensure; although the antibacterial invisible appliance formed by tertiary ammonification and in-situ quaternization provided by the Chinese patent application 202010062403.4 avoids the influence on biological safety caused by the release of an antibacterial agent into human metabolism, the grafting group is still an exogenous substance and has a gap with ideal biocompatibility, and the manufacturing process involves various chemical reactions and has more complex steps. However, the manufacturing method of the antibacterial appliance product disclosed above has the disadvantages of expensive equipment, complex operation, high cost and the like, and the adopted antibacterial raw material is inorganic metal and a compound thereof with poor biocompatibility. In addition, the above technology only considers the antibacterial problem and does not relate to promoting the enamel mineralization layer, and the function is single, so that the requirements of consumers cannot be fully met. Therefore, it is urgently needed to design an invisible orthodontic appliance which is simple and convenient to manufacture, good in biological safety, durable and stable in antibacterial effect and capable of promoting remineralization of demineralized tooth surfaces.
Disclosure of Invention
The invention aims to provide a fluorine-containing antibacterial invisible appliance for improving enamel demineralization for patients susceptible to caries. The fluorine-containing antibacterial invisible appliance can effectively release fluorine ions, promote remineralization of demineralized tooth surfaces, has good and stable antibacterial performance, and can be used for improving enamel demineralization level and preventing and treating bacterial plaque related diseases in the invisible appliance process.
The invention aims to provide a fluorine-containing antibacterial invisible appliance for improving enamel demineralization.
The invention also aims to provide a preparation method of the fluorine-containing antibacterial invisible appliance for improving enamel demineralization.
The above purpose of the invention is realized by the following technical scheme:
a fluorine-containing antibacterial invisible appliance for improving enamel demineralization is prepared by the following steps:
s1, soaking a substrate in a mixed solution containing a reducing agent and lysozyme to obtain a substrate with phase-transition lysozyme on the surface, and washing and drying the substrate;
s2, soaking the substrate with the phase transition lysozyme on the surface in negative biomacromolecule solution containing fluoride, washing and drying;
s3, soaking the obtained substrate in a lysozyme solution, washing and drying, and then soaking in a negative biomacromolecule solution containing fluoride;
s4, repeating the operation of the step S3 for 3-7 times, and drying to obtain the fluorine-containing antibacterial invisible appliance;
wherein, in the negatively charged biomacromolecule solution obtained in the steps S2 and S3, the concentration of the fluoride is 0.5-2 mg/L.
Lysozyme is the basis of human innate immunity, has high biocompatibility and can participate in maintaining the steady state of the oral environment, negative biomacromolecules and fluoride are widely used in clinic, and fluoride can not only prevent enamel demineralization but also promote remineralization. According to the invention, the fluorine-containing biomacromolecules with negative charges and the lysozyme with positive charges can be stacked on the phase-change lysozyme layer by layer through electrostatic acting force, and the method can fix natural antibacterial component lysozyme on one hand and fix the negative charge biomacromolecules capable of effectively releasing fluoride ions on the other hand, so that the fluorine-containing antibacterial invisible appliance with the function of improving tooth surface demineralization is obtained.
The method for obtaining a substrate having a phase transition lysozyme on the surface in the above step S1 is a method well known to those skilled in the art, and a detailed method is disclosed in, for example, patent application CN 105039953A.
Preferably, the pH of the mixed solution in the step S1 is 6-8, and the soaking temperature is 15-30 ℃.
Preferably, the concentration of the reducing agent in the step S1 is 30-60 mmol/L.
More preferably, the reducing agent concentration in step S1 is 50 mmol/L.
Preferably, the concentration of the lysozyme in the step S1 is 2-20 mg/mL.
More preferably, the concentration of lysozyme in step S1 is 10 mg/mL.
The phase-change lysozyme can provide abundant positive charges and C-H bonds, and is favorable for further combining with negative biomacromolecule solution containing fluoride.
Preferably, in the negatively charged biomacromolecule solution of steps S2 and S3, the fluoride is calcium fluoride or sodium fluoride.
More preferably, in the negatively charged biomacromolecule solution of steps S2 and S3, the fluoride is sodium fluoride.
Because the fluorine concentration is too high and harmful to the health of human bodies, the concentration of the fluoride in the negatively charged biomacromolecule solution in the step S2 and the step S3 is 0.5-2 mg/L.
Preferably, the concentration of the fluoride in the negatively charged biomacromolecule solution obtained in the steps S2 and S3 is 1.0-2.0 mg/L.
The preparation method of the negative charge biomacromolecule solution containing fluoride comprises the following steps: adding 0.2% glacial acetic acid solution into the negatively charged biomacromolecule powder, stirring until the negatively charged biomacromolecule powder is completely dissolved to obtain a negatively charged biomacromolecule solution, adding fluoride powder until the powder is completely dissolved, and finally preparing the negatively charged biomacromolecule solution containing fluoride. The inventor researches and discovers that hyaluronic acid has good biocompatibility and can effectively load fluoride, and other negative-charged biomacromolecules, such as alginic acid, have high viscosity in a prepared aqueous solution, and are easy to form hydrogel so as to increase the preparation difficulty; ovalbumin, while well biocompatible, cannot be used to load or release fluorine. Therefore, in the solution of negatively charged biological macromolecules in steps S2 and S3 of the present invention, the negatively charged biological macromolecules are preferably hyaluronic acid.
Preferably, in the negatively charged biomacromolecule solution obtained in the steps S2 and S3, the concentration of the negatively charged biomacromolecule solution is 0.5-2 mg/mL.
Because the antibacterial adhesive capacity of the hydrophilic surface is stronger than that of the hydrophobic surface, the invention increases the amino (-NH) on the surface of the substrate by introducing hyaluronic acid and lysozyme on the surface of the substrate2) And carboxyl (-COOH), thereby improving the hydrophilicity of the base material and improving the antibacterial effect of the fluorine-containing antibacterial invisible appliance.
Preferably, the concentration of the lysozyme solution in the step S3 is 0.5-2 mg/mL.
Considering the balance between hydrophilicity and operation complexity, it is preferable that the operation of step S3 is repeated 3 times at step S4, the number of times being related to the number of layers formed, having an influence on the performance of releasing fluorine. The inventor researches and discovers that 3 bilayers formed by assembling lysozyme and fluorine-containing compound negatively charged biomacromolecules after 3 times, and the outermost layer is the fluorine-containing antibacterial appliance containing the fluorine-containing compound negatively charged biomacromolecules, so that the fluorine releasing effect is the best.
Compared with the prior art, the invention has the following beneficial effects:
the invention provides a fluorine-containing antibacterial invisible appliance for improving enamel demineralization, which can effectively release fluorine ions and promote remineralization of demineralized tooth surfaces, and has good and stable antibacterial performance, good biocompatibility and ideal appearance. The phase-transition lysozyme is used as a base layer, the lysozyme and the fluorine-containing hyaluronic acid are fixed on the surface of the base body through a layer-by-layer stacking method and electrostatic acting force, the fluorine-containing antibacterial invisible appliance with good stable antibacterial and effective fluorine release effects is obtained, and the fluorine-containing antibacterial invisible appliance is simple in preparation process, cheap and easily available in raw materials, few in required varieties, low in cost, high in efficiency and the like. Therefore, the fluorine-containing antibacterial invisible appliance can improve the demineralization level of enamel, can also prevent and treat oral diseases related to bacteria such as gingivitis, caries, pulposis, periodontal disease and the like in the orthodontic treatment process, has simple and easy preparation process, high efficiency and no pollution, and has wide clinical application prospect.
Drawings
FIG. 1 is a process diagram for preparing the fluorine-containing antibacterial invisible appliance of the invention;
FIG. 2 is a graph showing the results of a long-term antibacterial test of the fluorine-containing antibacterial invisible orthosis of the present invention;
FIG. 3 is a graph showing the results of the remineralization promotion test of the fluorine-containing antibacterial invisible appliance of the present invention.
Detailed Description
The invention is further described with reference to the drawings and the following detailed description, which are not intended to limit the invention in any way. Reagents, methods and apparatus used in the present invention are conventional in the art unless otherwise indicated.
Unless otherwise indicated, reagents and materials used in the following examples are commercially available.
Example 1
1. Preparation of thermoplastic polyurethane surface phase transition lysozyme film
At room temperature, a 10mM 4-hydroxyethylpiperazine ethanesulfonic acid (HEPES) solution having a pH of 7.4 was prepared as a solvent, then a 10mg/ml lysozyme solution (10mM HEPES solution, solution pH7.4) and a 50mM tris (2-carboxyethyl) phosphine (TCEP) solution (10mM HEPES solution, solution pH6.2) were prepared, respectively, and equal volumes of lysozyme and TCEP solution were mixed uniformly to prepare phase-transition lysozyme (PTL). Soaking Thermoplastic Polyurethane (TPU) in a phase-change lysozyme solution, incubating for 2 hours in a humid environment, and washing the surface of the TPU substrate with sterile deionized water to wash away residual substances, N2And (4) drying, namely, marking the obtained sample as TPU-PTL, placing the TPU-PTL in a sterile culture dish, and sealing the TPU-PTL with a preservative film for later use.
2. Preparation of fluorine-containing hyaluronic acid and lysozyme solution
Preparation of Hyaluronic Acid (HA) solution: pouring 0.69mL of glacial acetic acid and 19.31mL of deionized water into a beaker, magnetically stirring to prepare a 0.2% glacial acetic acid solution, putting 0.06g of sodium hyaluronate powder into the prepared solution, and magnetically stirring until the sodium hyaluronate powder is completely dissolved to prepare a 1mg/mL Hyaluronic Acid (HA) solution.
Preparation of a solution of fluorine-containing hyaluronic acid (HA/NaF): the method comprises the steps of weighing three parts of sodium fluoride (NaF) powder with different masses by an electronic balance, dividing the prepared 1g/L hyaluronic acid solution into three groups, taking the hyaluronic acid solution as a solvent, adding the sodium fluoride powder with different masses into a beaker under the stirring of a magnetic stirrer until the powder is completely dissolved, and finally preparing three groups of fluorine-containing hyaluronic acid (HA/NaF) solutions with the concentrations of 0.5mg/L, 1mg/L and 2 mg/L.
Preparation of lysozyme solution: the electronic balance weighs lysozyme powder, adds deionized water to prepare a lysozyme solution with the concentration of 1mg/mL, and adds 0.05M sodium chloride to maintain the ionic strength.
3. Preparation of fluorine-containing antibacterial appliance
The TPU-PTL prepared previously has a layer of film with stable positive charges, and the film is used as a starting basis for layer-by-layer self-assembly. At room temperature, a sample TPU-PTL is immersed in a sodium fluoride hyaluronic acid solution (HA/NaF) with the concentration of 0.5mg/L for 30min, a layer of film with negative charges is adsorbed on the surface, the sample is taken out, washed by acetic acid, dried in vacuum, then immersed in a lysozyme solution (LY) with the concentration of 1mg/mL for 30min, a layer of film with positive charges is adsorbed on the surface again, and washed by sterile deionized water. LY and HA/NaF are respectively used as a single layer, LY-HA/NaF is used as a double layer, the operation is circulated for 3 times, the multi-layer polyelectrolyte composite coating consisting of three double layers is obtained, wherein HA/NaF is arranged on the outermost layer, and the fluorine-containing antibacterial appliance can be obtained after drying.
Example 2
The TPU-PTL prepared in example 1 was used as the starting basis for the layer-by-layer self-assembly. At room temperature, a sample TPU-PTL is immersed in a sodium fluoride hyaluronic acid solution (HA/NaF) with the concentration of 1mg/L for 30min, a layer of film with negative charges is adsorbed on the surface, the sample is taken out, washed by acetic acid, dried in vacuum, then placed in a 1mg/mL lysozyme solution (LY) for immersion for 30min, a layer of film with positive charges is adsorbed on the surface again, and washed by sterile deionized water. LY and HA/NaF are respectively used as a single layer, LY-HA/NaF is used as a double layer, the operation is circulated for 3 times, the multi-layer polyelectrolyte composite coating consisting of three double layers is obtained, wherein HA/NaF is arranged on the outermost layer, and the fluorine-containing antibacterial appliance can be obtained after drying.
Example 3
The TPU-PTL prepared in example 1 was used as the starting basis for the layer-by-layer self-assembly. At room temperature, a sample TPU-PTL is immersed in a sodium fluoride hyaluronic acid solution (HA/NaF) with the concentration of 2mg/L for 30min, a layer of film with negative charges is adsorbed on the surface, the sample is taken out, washed by acetic acid, dried in vacuum, then placed in a lysozyme solution (LY) with the concentration of 1mg/mL for soaking for 30min, a layer of film with positive charges is adsorbed on the surface again, and washed by sterile deionized water. LY and HA/NaF are respectively used as a single layer, LY-HA/NaF is used as a double layer, the operation is circulated for 3 times, the multi-layer polyelectrolyte composite coating consisting of three double layers is obtained, wherein HA/NaF is arranged on the outermost layer, and the fluorine-containing antibacterial appliance can be obtained after drying.
Experimental example 1 Long-lasting antibacterial verification
1. Preparing a culture medium: two clean glass bottles with a maximum scale of 400mL are taken as containers and prepared according to the concentration of Agar (Agar) of 20 g/L. 8g of agar was weighed and poured into a clean glass bottle containing 400mL of deionized water, and stirred with a clean glass rod until the powder was completely dissolved, and the solution was transparent. Sterilizing the prepared culture medium with high temperature and high pressure steam (120 deg.C, 1 hr), cooling to room temperature, sealing with sealing glue, and storing in 4 deg.C refrigerator.
2. Establishing a staphylococcus aureus model: taking out the staphylococcus aureus strain stored at-20 ℃, recovering, and placing in a constant temperature cabinet at 37 ℃ for 24 h. Centrifuging the recovered Staphylococcus aureus liquid (4 deg.C, 10000rpm, 10min), washing the precipitate with sterile physiological saline for 2 times, and adjusting the concentration of the suspension to about 5 × 107～5x108CFU/mL. 1mL of each bacterial suspension was added to 50 vials containing artificial saliva and randomly divided into two groups of 25 each.
Establishing an antibacterial sample model: 25 fluorine-containing antibacterial invisible appliances prepared in example 1 and 25 common invisible appliances were divided into two groups and immersed in small conical bottles containing bacterial suspension artificial saliva, respectively, and then placed in a 37 ℃ incubator, and samples of the immersion liquid were taken out at predetermined time points (1, 3, 5, 7, and 14 days) and collected.
3. And (3) determination of antibacterial rate: sucking 0.1mL of the soak solution sample into a first culture dish by using a pipette, then diluting by 10 times by using a small test tube, taking 0.1mL of the soak solution sample into a second culture dish after dilution, and sequentially diluting until reaching a fourth culture dish. The other groups are repeated in sequence.
The formula for calculating the antibacterial rate is as follows: r ═ B-A)/(B × 100%
Wherein R represents the antibacterial rate, A represents the average recovered colony number (CFU/mL) in the fluorine-containing antibacterial invisible orthodontic appliance soaking solution, and B represents the average recovered colony number (CFU/mL) in the common invisible orthodontic appliance soaking solution. And averaging the antibacterial rate measured by each group, and drawing an antibacterial rate curve.
The experimental result is shown in fig. 2, the fluorine-containing antibacterial invisible appliance of the invention keeps higher antibacterial rate in 14 days in a simulated oral environment, the antibacterial rate to a 14-day node is not less than 95%, and the antibacterial rate and the durability of the fluorine-containing antibacterial invisible appliance are obviously superior to those of a common appliance, so that the fluorine-containing antibacterial invisible appliance of the invention has excellent long-acting antibacterial performance.
Experimental example 2 verification of Remineralization promoting Capacity
1. Preparing an isolated tooth: 25 healthy premolars (normal enamel development, no defect, no crack) were collected from 12-28 years old patients who had been removed by oral surgery for orthodontic treatment. Removing surface residual tissue, polishing, washing with flowing water, storing in physiological saline, and storing in refrigerator at 4 deg.C. The random classification was four groups:
group A: without any treatment, the sample is used as a sample before demineralization;
group B: manufacturing a demineralization model, and taking the demineralization model as a demineralized sample without carrying out remineralization treatment;
group C: manufacturing a demineralization model, fixing the demineralization model on a common invisible orthodontic appliance, and performing in-vitro demineralization circulation;
group D: a demineralization model is manufactured and is fixed on the fluorine-containing antibacterial invisible orthodontic appliance prepared in the embodiment 1, and in-vitro demineralization circulation is carried out;
group E: a demineralization model is manufactured and is fixed on the fluorine-containing antibacterial invisible orthodontic appliance prepared in the embodiment 2, and in-vitro demineralization circulation is carried out;
and F group: a demineralization model was prepared, and the model was fixed to the fluorine-containing antibacterial invisible orthosis prepared in example 3, and subjected to an in vitro demineralization cycle.
2. Manufacturing a demineralization model: the method for adhering the accessories of 25 isolated tooth simulated intraoral invisible appliances in B, C, D, E, F groups is adopted, acid etching is carried out on the buccal surface of the tooth by using acid etchant, adhesive is coated to adhere the accessories to the center of the buccal surface of the tooth, and redundant adhesive around the accessories is removed. Soaking the sample in the demineralizing liquid for 1 hour every day, carrying out demineralizing in a constant-temperature water bath box for 2 weeks, changing the demineralizing liquid every day, removing the bracket and the adhesive on the in-vitro tooth after 2 weeks, and polishing for later use.
3. And (3) washing the demineralized in-vitro teeth with deionized water, cleaning and drying cheek surfaces of the teeth, wrapping and fixing the same positions of the inner side surfaces of 5 fluorine-containing antibacterial invisible appliances with D, E, F groups of fluorine-containing antibacterial invisible appliances and 5 common invisible appliances with sterile films by using 5 in-vitro teeth and D, E, F groups of fluorine-containing antibacterial invisible appliances, and then putting the in-vitro teeth back into the artificial saliva. The fixed samples were soaked in artificial demineralization solution daily for 1 hour, simulating daily demineralization time in the oral environment under normal conditions, and then rinsed with deionized water and placed back into artificial saliva. The above process was carried out in a 37 ℃ thermostated water bath, wherein artificial saliva and artificial demineralization were changed daily. The circulation is carried out for 14 days.
4. Enamel hardness measurement: microhardness instruments measure microhardness values of all sample buccal enamel. And taking the average value of the three-point measurement values as the hardness value result of the sample by measuring the diagonal length of the indentation and converting the diagonal length into the hardness value.
The experimental results are shown in fig. 3, the microhardness values of the group a are the highest, the microhardness values of the group B are the lowest, and the microhardness values of the group D (example 1), the group E (example 2) and the group F (example 3) are obviously higher than those of the group C (common invisible appliance). The fluorine-containing antibacterial invisible appliance of the groups D, E and F can effectively remineralize demineralized enamel and increase the microscopic hardness value of the enamel surface compared with the common invisible appliance of the group C. The microhardness value of the F group is higher than that of the D group and the E group, which shows that the mineralization promoting effect of the fluorine-containing antibacterial appliance has certain fluorine concentration dependence.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
Claims (10)
1. A fluorine-containing antibacterial invisible appliance for improving enamel demineralization is characterized in that the preparation method comprises the following steps:
s1, soaking a substrate in a mixed solution containing a reducing agent and lysozyme to obtain a substrate with phase-transition lysozyme on the surface, and washing and drying the substrate;
s2, soaking the substrate with the phase transition lysozyme on the surface in negative biomacromolecule solution containing fluoride, washing and drying;
s3, soaking the obtained substrate in a lysozyme solution, washing and drying, and then soaking in a negative biomacromolecule solution containing fluoride;
s4, repeating the operation of the step S3 for 3-7 times, and drying to obtain the fluorine-containing antibacterial invisible appliance;
wherein, in the negative biomacromolecule solution obtained in the step S2 and the step S3, the concentration of the fluoride is 0.5-2.0 mg/L.
2. The fluorine-containing antibacterial invisible appliance of claim 1, wherein the concentration of fluoride in the negatively charged biomacromolecule solution in steps S2 and S3 is 1.0-2.0 mg/L.
3. The fluorine-containing antimicrobial invisible appliance of claim 1, wherein in the negatively charged biomacromolecule solution of the steps S2 and S3, the fluoride is calcium fluoride or sodium fluoride.
4. The fluorine-containing antimicrobial invisible appliance of claim 1, wherein in the negatively charged biomacromolecule solution of steps S2 and S3, the negatively charged biomacromolecule is hyaluronic acid.
5. The fluorine-containing antibacterial invisible appliance of claim 1, wherein in the negatively charged biomacromolecule solution in steps S2 and S3, the concentration of the negatively charged biomacromolecule solution is 0.5-2 mg/mL.
6. The fluorine-containing antibacterial invisible appliance of claim 1, wherein the concentration of the lysozyme solution in the step S3 is 0.5-2 mg/mL.
7. The fluorine-containing antibacterial invisible appliance of claim 1, wherein the pH of the mixed solution in the step S1 is 6-8.
8. The fluorine-containing antibacterial invisible appliance of claim 1, wherein the concentration of the reducing agent in the mixed solution in the step S1 is 30-60 mmol/L.
9. The fluorine-containing antibacterial invisible appliance of claim 1, wherein the concentration of lysozyme in the mixed solution in the step S1 is 2-20 mg/mL.
10. The fluorine-containing antimicrobial invisible appliance of claim 1, wherein the step S4 is repeated 3 times in the step S3.
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