Oral repair membrane and preparation method thereof

Abstract

The present invention provides an oral repair membrane and a preparation method thereof, and relates to the technical field of medical repair materials. The oral repair membrane comprises a degradable repair membrane layer and a porous structure layer and a protective film layer compounded in sequence on the degradable repair membrane layer; wherein: the degradable repair membrane layer is attached to the oral wound surface for repair, and the degradable repair membrane layer is compounded by a plurality of fiber orientation sheets, and the orientation direction of the oriented fibers in the fiber orientation sheets is consistent; the porous structure layer has a relatively large pore structure, which is conducive to the flow of nutrients during the repair process; the protective film layer is a smooth and dense film layer, which does not have a pore structure and plays an isolation role. The oral repair membrane of the present application achieves a better oral repair effect through the mutual coordination of the degradable repair membrane layer, the porous structure layer and the protective film layer. Compared with the existing oral repair membranes, it has the advantages of high biocompatibility, fast tissue regeneration, good mechanical strength and excellent antibacterial properties.

Description

Technical Field

[0001] The invention relates to the technical field of medical repair materials, and in particular to an oral repair membrane and a preparation method thereof. Background Art

[0002] Oral repair membrane for soft tissue repair is used to repair shallow defects of soft tissue in the oral cavity, including cheeks, palates, tongue, gums, etc. The diseases include various common benign lesions of oral mucosa and some malignant lesions, as well as the prevention of gustatory sweating syndrome (Frey's) during parotid gland surgery.

[0003] However, existing absorbable oral repair membranes are divided into absorbable collagen membranes and absorbable polymer membranes. The absorbable collagen oral repair membrane can be absorbed by the human body and has good tissue induction regeneration function, but it is antigenic; while the absorbable polymer membrane, while providing sufficient mechanical properties, often has poor tissue adhesion at the implantation site and a long degradation cycle, which may increase the risk of infection and affect the normal growth and repair process of surrounding tissues.

[0004] Therefore, it is necessary and urgent to research and develop an oral repair membrane with better biocompatibility, mechanical strength, soft texture, good tissue adhesion, and conducive to tissue regeneration and nutrient flow during the repair process.

[0005] In view of this, the present invention is proposed. Summary of the invention

[0006] The first purpose of the present invention is to provide an oral repair membrane, which achieves a better oral repair effect through the mutual coordination of a degradable repair membrane layer, a porous structure layer and a protective film layer. Compared with the existing oral repair membranes, the oral repair membrane has the advantages of high biocompatibility, rapid tissue regeneration, good mechanical strength and excellent antibacterial performance.

[0007] The second object of the present invention is to provide a method for preparing an oral repair membrane.

[0008] In order to achieve the above-mentioned purpose of the present invention, the following technical solutions are particularly adopted:

[0009] The present invention provides an oral repair film, which comprises: a degradable repair film layer, and a porous structure layer and a protective film layer which are sequentially compounded on the degradable repair film layer;

[0010] The degradable repair film layer is prepared by compounding multiple fiber oriented sheets;

[0011] The fiber orientation sheet is formed into oriented fibers by an electrostatic spinning method, and the orientation directions of the oriented fibers in the fiber orientation sheet are consistent;

[0012] The orientation directions of the fibers in each fiber orientation sheet in the degradable repair film layer are different.

[0013] Furthermore, the oriented fibers are obtained by electrostatic spinning of degradable polyurethane or polylactic acid;

[0014] Preferably, the weight average molecular weight Mw of the degradable polyurethane is 5W to 12W;

[0015] Preferably, the weight average molecular weight Mw of the polylactic acid is 8W-15W.

[0016] Preferably, the diameter of the oriented fibers in the fiber oriented sheet is 1 nm to 20 μm, and the porosity is 20 to 80%.

[0017] Furthermore, the porous structure layer is prepared from degradable polyurethane by a casting method;

[0018] The porous structure layer has a pore structure, the porosity of the porous structure layer is 5-20%, and the pore size range of the micropores is less than 100um;

[0019] And / or, the protective film layer is prepared from degradable polyurethane by a casting method, and the protective film layer is a smooth and dense film layer without a pore structure.

[0020] Furthermore, the total thickness of the oral repair film is 0.2 to 0.6 mm, wherein:

[0021] The thickness of the degradable repair film layer is 0.02-0.5 mm, the thickness of the porous structure layer is 0.03-0.2 mm, and the thickness of the protective film layer is 0.03-0.1 mm.

[0022] Furthermore, the oral repair membrane has a tensile strength of 6-30 N / cm, an elongation at break of 80%-650%, a suture strength of 4-20 N, and a softness of <500 mN.

[0023] Furthermore, the degradable repair film layer is prepared by compounding two layers of fiber oriented sheets;

[0024] Preferably, the two-layer fiber-oriented sheet comprises a first fiber-oriented sheet and a second fiber-oriented sheet, and the first fiber-oriented sheet and the second fiber-oriented sheet have different orientation directions and are at an angle of 90°±15°.

[0025] The present invention provides a method for preparing the above-mentioned oral repair membrane, the preparation method comprising:

[0026] (A) preparing a protective film layer and a porous structure layer in sequence by a tape casting method to obtain a composite intermediate; then placing the composite intermediate on a substrate, with the protective film layer in contact with the substrate and the porous structure layer facing outward; and then preparing a degradable repair film layer on the porous structure layer by an electrospinning method to obtain a preliminary oral repair film;

[0027] (B) removing the solvent and drying the initial oral repair membrane in sequence to obtain an oral repair membrane.

[0028] Furthermore, the preparation method of the degradable repair film layer includes:

[0029] S1: dissolving the degradable polyurethane in a solvent to obtain a spinning solution;

[0030] S2: electrospinning the spinning solution to obtain a fiber orientation sheet A, then adjusting the electrospinning direction, and electrospinning on the fiber orientation sheet A to obtain a fiber orientation sheet B intersecting the orientation direction of the fiber orientation sheet A, thereby obtaining a degradable repair film layer composed of two layers of fiber orientation sheets;

[0031] S3: Repeat step S2 to obtain a degradable repair film layer composed of multiple fiber oriented sheets.

[0032] Furthermore, in step S1, the concentration of the degradable polyurethane in the spinning solution is 15-40% (w / v);

[0033] Preferably, the solvent in the spinning solution is a single solvent of N,N-dimethylformamide, a single solvent of N,N-dimethylacetamide, a mixed solvent of N,N-dimethylformamide and any one or more of tetrahydrofuran, dichloroethane or dimethyl ether, or a mixed solvent of N,N-dimethylacetamide and any one or more of tetrahydrofuran, dichloroethane or dimethyl ether;

[0034] More preferably, the solvent in the spinning solution is a mixed solvent of N,N-dimethylformamide and tetrahydrofuran, wherein the volume ratio of N,N-dimethylformamide to tetrahydrofuran is 10:1-3:7, wherein the preferred volume ratio is 10:1-1:1.

[0035] Furthermore, the electrospinning conditions in step S2 include: negative voltage: -1KV~-4KV, positive voltage: 7KV~10KV, injection speed of the nozzle: 0.1-0.4ml / min; rotation speed of the receiving device: (1000, 2500) rpm / min, distance between the nozzle and the receiving device: 0~50 (10-30) mm; translation speed of the nozzle is (0, 500) mm / min.

[0036] Compared with the prior art, the present invention has the following beneficial effects:

[0037] The oral repair film provided by the present invention mainly includes a degradable repair film layer and a porous structure layer and a protective film layer compounded on the degradable repair film layer in sequence; wherein: the degradable repair film layer is attached to the oral wound surface for repair, and the degradable repair film layer of the present application is compounded by a multi-layer fiber orientation sheet, and the orientation direction of the oriented fibers in the fiber orientation sheet is consistent. The oriented fibers with consistent orientation directions in the fiber orientation sheet of the present application are more conducive to the attachment of cells, and the cells have better mobility and growth rate on the oriented fibers, and the repair and regeneration effect is good; at the same time, the patch obtained by compounding the fiber orientation sheets with different orientation directions in the present application has better mechanical strength and is not easy to be torn. The porous structure layer has a larger pore structure, which is conducive to the flow of nutrients during the repair process. The protective film layer is a smooth and dense film layer without a pore structure. It can not only block the entry of bacteria, food residues and other substances, but also prevent the growth of fibroblasts, muscles and vascular tissue cells, etc., and play an isolation role.

[0038] Therefore, the oral repair membrane of the present application achieves a better oral repair effect through the mutual coordination of the degradable repair membrane layer, the porous structure layer and the protective membrane layer. Compared with the existing oral repair membranes, it has the advantages of high biocompatibility, rapid tissue regeneration, good mechanical strength and excellent antibacterial properties.

[0039] The preparation method of the oral repair membrane provided by the present invention comprises the following steps: firstly, a protective film layer and a porous structure layer are prepared in sequence by a tape casting method to obtain a composite intermediate; then, the composite intermediate is placed on a substrate, the protective film layer is in contact with the substrate, and the porous structure layer faces outward; then, a degradable repair membrane layer is compositely prepared on the porous structure layer by an electrostatic spinning method to obtain a preliminary oral repair membrane; and then, the preliminary oral repair membrane is sequentially subjected to solvent removal and drying treatment to obtain an oral repair membrane. The above preparation method has the technical advantages of simple processing technology and easy operation. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] In order to more clearly illustrate the specific implementation methods of the present invention or the technical solutions in the prior art, the drawings required for use in the specific implementation methods or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are some implementation methods of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0041] Figure 1 This is a 5000 times scanning electron microscope image of the fiber orientation sheet of the degradable repair membrane layer of Example 1 of the present invention;

[0042] Figure 2 This is a 500-fold scanning electron microscope image of the fiber orientation sheet of the degradable repair membrane layer of Example 1 of the present invention;

[0043] Figure 3 The cells provided in Experimental Example 1 of the present invention were co-incubated with the oral repair membrane of Example 1 of the present application for 6 days and the cell migration results on one side of the degradable repair membrane layer. DETAILED DESCRIPTION

[0044] The technical solution of the present invention will be clearly and completely described below in conjunction with the embodiments. Obviously, the described embodiments are part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0045] According to one aspect of the present invention, an oral repair film comprises: a degradable repair film layer, and a porous structure layer and a protective film layer sequentially compounded on the degradable repair film layer;

[0046] The degradable repair film layer is prepared by compounding multiple fiber oriented sheets;

[0047] The fiber orientation sheet is formed into oriented fibers by an electrostatic spinning method, and the orientation directions of the oriented fibers in the fiber orientation sheet are consistent;

[0048] The orientation directions of the fibers in each fiber orientation sheet in the degradable repair film layer are different.

[0049] The oral repair film provided by the present invention comprises a degradable repair film layer and a porous structure layer and a protective film layer which are sequentially compounded on the degradable repair film layer; wherein:

[0050] 1. The degradable repair film layer is attached to the oral wound surface for repair. The degradable repair film layer of the present application is obtained by compounding multiple fiber oriented sheets, and the oriented fibers in the fiber oriented sheets have the same orientation direction. The oriented fibers with the same orientation direction are conducive to the attachment of cells. The cells have better mobility and growth rate on the oriented fibers, and the repair and regeneration effect is good. At the same time, the patch obtained by compounding fiber oriented sheets with different orientation directions has better mechanical strength and is not easy to be torn.

[0051] 2. The porous structure layer has a larger pore structure, which is conducive to the flow of nutrients during the repair process and is beneficial to tissue repair.

[0052] 3. The protective film layer is a smooth and dense film layer without a porous structure. The protective film layer of the present application can not only block the entry of bacteria, food residues and other substances, but also prevent the growth of fibroblasts, muscles and vascular tissue cells, etc., and play an isolation role.

[0053] Therefore, the oral repair membrane of the present application achieves a better oral repair effect through the mutual coordination of the degradable repair membrane layer, the porous structure layer and the protective membrane layer. Compared with the existing oral repair membranes, it has the advantages of high biocompatibility, rapid tissue regeneration, good mechanical strength and excellent antibacterial properties.

[0054] In a preferred embodiment of the present invention, the oriented fibers are obtained by electrospinning degradable polyurethane or polylactic acid;

[0055] Preferably, the weight average molecular weight Mw of the degradable polyurethane is 5W to 12W; the weight average molecular weight Mw of the polylactic acid is 8W to 15W.

[0056] The weight average molecular weight of the degradable polyurethane may be, but not limited to, 5W, 6W, 7W, 8W, 9W, 10W, 11W or 12W, or any value between 5W and 12W. The weight average molecular weight of the polylactic acid may be, but not limited to, 8W, 9W, 10W, 11W, 12W, 13W, 14W or 15W, or any value between 8W and 15W.

[0057] Preferably, the diameter of the oriented fibers in the fiber oriented sheet is 1 nm to 20 μm, and the porosity is 20 to 80%.

[0058] It should be noted that in the repair site of oral repair membranes (such as membranes used for healing after tooth or gum surgery), multiple types of cells are involved in healing and reconstruction. They mainly include: Fibroblasts: These cells play an important role in tissue repair and reconstruction, and their diameter is usually between 10 and 30 microns. They are responsible for synthesizing collagen and other extracellular matrix components to promote wound healing. Keratinocytes: In the submucosal layer of the oral mucosa, keratinocytes are approximately 10 to 30 microns in size and are responsible for forming a protective layer of the oral mucosa. Macrophages: These cells involved in the immune response, usually between 10 and 30 microns in diameter, play a role in clearing dead cells and foreign matter, and help the healing process. Endothelial cells: If the repair area involves vascular regeneration, the diameter of endothelial cells is generally between 10 and 25 microns, and they are responsible for forming new blood vessels to supply the nutrients needed for the repair area. Satellite cells: In the surrounding neural tissue, satellite cells are usually between 10 and 15 microns in diameter and are responsible for supporting and protecting neurons.

[0059] At the same time, it has been found that the cells have better adhesion properties on fibers with a diameter smaller than their own during oral repair. When the transverse diameter of the electrospinning is controlled to be smaller than the diameter of the oral repair cells, it is beneficial for the cells to attach and grow on the electrospinning. Therefore, the diameter of the oriented fibers in the fiber orientation sheet is controlled to be 1nm to 20μm, and the porosity is 20 to 80%.

[0060] In a preferred embodiment of the present invention, the porous structure layer is prepared from degradable polyurethane by a casting method;

[0061] The porous structure layer has a pore structure, the porosity of the porous structure layer is 5-20%, and the pore size range of the micropores is less than 100um;

[0062] Preferably, the porogen of the porous structure layer is selected from any one or a combination of at least two of sodium chloride, sucrose, gelatin balls, monosodium glutamate, modified cellulose, urea, polyethylene glycol or polyvinyl pyrrolidone, preferably sodium chloride;

[0063] More preferably, the particle size of the porogen is less than 100 μm.

[0064] In a preferred embodiment of the present invention, the protective film layer is prepared from degradable polyurethane by a casting method, and the protective film layer is a smooth and dense film layer without a pore structure.

[0065] In a preferred embodiment of the present invention, the total thickness of the oral repair film is 0.2-0.6 mm, wherein the thickness of the degradable repair film layer is 0.02-0.5 mm, the thickness of the porous structure layer is 0.03-0.2 mm, and the thickness of the protective film layer is 0.03-0.1 mm.

[0066] Wherein, the total thickness of the oral repair film may be, but is not limited to, 0.2 mm, 0.3 mm, 0.4 mm, 0.5 mm or 0.6 mm, or any value between 0.2 and 0.6 mm. The thickness of the degradable repair film layer may be, but is not limited to, 0.02 mm, 0.1 mm, 0.15 mm, 0.18 mm, 0.2 mm, 0.3 mm, 0.4 mm, 0.45 mm or 0.5 mm, or any value between 0.02 and 0.5 mm. The thickness of the porous structure layer may be, but is not limited to, 0.02 mm, 0.05 mm, 0.08 mm, 0.1 mm, 0.12 mm, 0.15 mm, 0.18 mm or 0.2 mm, or any value between 0.03 and 0.2 mm. The thickness of the protective film layer may be, but is not limited to, 0.03 mm, 0.05 mm, 0.07 mm, 0.09 or 0.1 mm, or any value between 0.03 and 0.1 mm.

[0067] In a preferred embodiment of the present invention, the oral repair membrane has a tensile strength of 6-30 N / cm, an elongation at break of 80%-650%, a suture strength of 4-20 N, and a softness of <500 mN.

[0068] In a preferred embodiment of the present invention, the degradable repair film layer is prepared by compounding two layers of fiber oriented sheets;

[0069] Preferably, the two-layer fiber-oriented sheet comprises a first fiber-oriented sheet and a second fiber-oriented sheet, and the first fiber-oriented sheet and the second fiber-oriented sheet have different orientation directions and are at an angle of 90°±15°.

[0070] As a preferred embodiment, the present application has found through research that the degradable repair film layer prepared by compounding two layers of fiber-oriented sheets "wherein the orientation directions of the first fiber-oriented sheet and the second fiber-oriented sheet are different and are at an angle of 90°±15°" has a better repair effect than other multi-layered degradable repair film layers, and has good degradation controllability. In addition, the two layers of fiber-oriented sheets cross-arranged at an angle of 90°±15° also have good transverse and longitudinal mechanical properties.

[0071] Mainly manifested in:

[0072] 1. Compared with the multi-layer structure, the porosity of the degradable repair membrane layer prepared by the above two layers of fiber oriented sheets is easier to repair the combination of cells and patches, and the biocompatibility is better; at the same time, the cell migration rate is good and the cell growth rate is high.

[0073] 2. The degradable repair film prepared by two layers of fiber oriented sheets with a vertical cross structure has good mechanical strength and is not easy to be torn. Compared with the patch with multiple fiber structures arranged at other angles, it has good transverse and longitudinal mechanical properties and better strength.

[0074] 3. The thickness is thinner and has a wider applicability; the degradation is better controllable and can be effectively degraded after forming tissue engineering support, avoiding the problem of long degradation time of multi-layer structure patches affecting healing.

[0075] According to one aspect of the present invention, a method for preparing the above-mentioned oral repair membrane comprises:

[0076] (A) preparing a protective film layer and a porous structure layer in sequence by a tape casting method to obtain a composite intermediate; then placing the composite intermediate on a substrate, with the protective film layer in contact with the substrate and the porous structure layer facing outward; and then preparing a degradable repair film layer on the porous structure layer by an electrospinning method to obtain a preliminary oral repair film;

[0077] (B) removing the solvent and drying the initial oral repair membrane in sequence to obtain an oral repair membrane.

[0078] The preparation method of the oral repair membrane provided by the present invention comprises the following steps: firstly, a protective film layer and a porous structure layer are prepared in sequence by a tape casting method to obtain a composite intermediate; then, the composite intermediate is placed on a substrate, the protective film layer is in contact with the substrate, and the porous structure layer faces outward; then, a degradable repair membrane layer is compositely prepared on the porous structure layer by an electrostatic spinning method to obtain a preliminary oral repair membrane; and then, the preliminary oral repair membrane is sequentially subjected to solvent removal and drying treatment to obtain an oral repair membrane. The above preparation method has the technical advantages of simple processing technology and easy operation.

[0079] In a preferred embodiment of the present invention, the method for preparing the degradable repair film layer comprises:

[0080] S1: dissolving the degradable polyurethane in a solvent to obtain a spinning solution;

[0081] S2: electrospinning the spinning solution to obtain a fiber orientation sheet A, then adjusting the electrospinning direction, and electrospinning on the fiber orientation sheet A to obtain a fiber orientation sheet B intersecting the orientation direction of the fiber orientation sheet A, thereby obtaining a degradable repair film layer composed of two layers of fiber orientation sheets;

[0082] S3: Repeat step S2 to obtain a degradable repair film layer composed of multiple fiber oriented sheets.

[0083] In a preferred embodiment of the present invention, the concentration of the degradable polyurethane in the spinning solution in step S1 is 15-40% (w / v);

[0084] Preferably, the solvent in the spinning solution is a single solvent of N,N-dimethylformamide, a single solvent of N,N-dimethylacetamide, a mixed solvent of N,N-dimethylformamide and any one or more of tetrahydrofuran, dichloroethane or dimethyl ether, or a mixed solvent of N,N-dimethylacetamide and any one or more of tetrahydrofuran, dichloroethane or dimethyl ether;

[0085] More preferably, the solvent in the spinning solution is a mixed solvent of N,N-dimethylformamide and tetrahydrofuran, wherein the volume ratio of N,N-dimethylformamide to tetrahydrofuran is 10:1-3:7, wherein the preferred volume ratio is 10:1-1:1.

[0086] In a preferred embodiment of the present invention, the conditions for electrospinning in step S2 include: negative voltage: -1KV~-4KV, positive voltage: 7KV~10KV, injection speed of the nozzle: 0.1-0.4ml / min; rotation speed of the receiving device: (1000, 2500) rpm / min, distance between the nozzle and the receiving device: 0~50 (10-30) mm; translation speed of the nozzle is (0, 500) mm / min.

[0087] As a preferred embodiment, oriented spinning fibers can be produced under the above-mentioned electrospinning conditions, thereby realizing the preparation of fiber oriented sheets with consistent fiber orientation directions.

[0088] The technical solution of the present invention will be further described below in conjunction with embodiments.

[0089] Example 1

[0090] A method for preparing an oral repair membrane, the preparation method comprising:

[0091] 1. Preparation of protective film layer:

[0092] Polyurethane particles with a molecular weight (Mw) of 80,000 were added to tetrahydrofuran (THF) solvent at a solid-liquid ratio (w / v) of 25%, and stirred to completely dissolve. The solution was poured onto a glass plate, a coater with adjusted height was placed, and after being pushed open with an automatic coater, it was air-dried under a laminar flow hood.

[0093] 2. Preparation of porous structure layer:

[0094] Add 15% NaCl (<100um) to tetrahydrofuran (THF) solvent and stir evenly, then add polyurethane particles with a molecular weight (Mw) of 120000 at a solid-liquid ratio (w / v) of 15%, stir to completely dissolve. Pour the solution on the air-dried protective film layer, place the coater with adjusted height, push it open with an automatic coater, and air dry under a laminar flow hood; after air drying, soak it in injection water for 30 minutes, demould it, and dry it again.

[0095] 3. Preparation of degradable repair film layer:

[0096] (1) Preparation of degradable polyurethane (PU) solution:

[0097] A (20±1)% polyurethane solution was prepared by using degradable polyurethane as solute and a mixed solution of tetrahydrofuran (THF) and N,N-dimethylformamide (DMF) as solvent.

[0098] The ratio of solvents: tetrahydrofuran (THF): N,N-dimethylformamide (DMF) = 1:1.

[0099] The specific preparation operation is as follows:

[0100] 1) Add (150 mL) of tetrahydrofuran and (150) mL of N,N-dimethylformamide into a three-necked flask, turn on the stirrer, adjust the speed to (200 ± 100) r / min, and stir evenly.

[0101] 2) Weigh (60±3) g of polyurethane material and add it to a three-necked flask. Different volumes of solution can be prepared according to the amount of solution used.

[0102] 3) Stir at room temperature for more than 4 hours to completely dissolve the PU. Let stand for more than 4 hours to remove bubbles and set aside.

[0103] (2) Electrospinning:

[0104] 1) Wipe the electrospinning machine clean and ensure that the equipment is in a clean state.

[0105] 2) Fix the porous structure layer + protective film layer on the release paper, with the porous structure layer facing outward. Fix the release paper on the receiving device to ensure that the release paper is not loose.

[0106] 3) Adjust the distance between the spinneret and the receiver to 20±1cm, determine the position and clamp it.

[0107] 4) The receiver receiving speed is set to (1000±100)rpm / min; the push injection speed is set to 0.1±0.05mm / min.

[0108] 5) The translation speed is set to 450±50mm / min and the translation distance is set to 300±50mm.

[0109] 6) Use the prepared (20±1%) PU solution and slowly draw the PU solution with a disposable syringe to avoid bubbles. If bubbles are generated, they must be removed before subsequent operations. Install a 22G needle (inner diameter 0.41mm) and fix the syringe in the buckle of the electrospinning machine push pump.

[0110] The nozzle diameter range can be: 0.15-4.2mm, preferably 18G (inner diameter 0.9mm), 19G (inner diameter 0.84mm), 20G (inner diameter 0.62mm), 21G (inner diameter 0.51mm), 22G (inner diameter 0.41mm), 23G (inner diameter 0.33mm), 24G (inner diameter 0.3mm) needles. In this embodiment, a 22G needle is selected.

[0111] 7) Set the negative voltage to (-2±1)KV and the positive voltage to (7±0.5)KV. Start electrospinning and observe the spinning effect until continuous and uniform yarns are ejected. Set the electrospinning time to 30min to 50min. After the first electrospinning, rotate the release paper (with a spinning layer on the surface) 90 degrees and then fix it. Set the electrospinning time to 30min to 50min again and repeat the operation for the second electrospinning.

[0112] After the second electrospinning is completed, the initial oral repair membrane is taken off from the release paper.

[0113] Figure 1 This is a 5000x scanning electron microscope image of the fiber orientation sheet of the degradable repair membrane layer of this embodiment.

[0114] Figure 2 This is a 500-fold scanning electron microscope image of the fiber-oriented sheet of the degradable repair membrane layer of this embodiment.

[0115] 4. Post-treatment: Soak the initial oral repair membrane in water for injection until the total amount of tetrahydrofuran (THF) solvent remaining in the oral repair membrane is ≤0.1%.

[0116] 5. Drying: Dry the initial oral repair membrane until the residual moisture in the initial product is ≤ 0.5% for easy storage.

[0117] 6. Cutting: Cut off the rough edges or uneven thickness of the dried oral repair membrane. Make sure there are no bubbles, no damage, no foreign matter, and the thickness is uniform, and cut it into the required size, which is the finished oral repair membrane.

[0118] Example 2

[0119] A method for preparing an oral repair membrane, the preparation method comprising:

[0120] 1. Preparation of protective film layer: same as Example 1.

[0121] 2. Preparation of porous structure layer: same as Example 1.

[0122] 3. Preparation of degradable repair film layer:

[0123] (1) Preparation of degradable polyurethane (PU) solution: same as in Example 1.

[0124] (2) Electrospinning:

[0125] 1)~6) are the same as in Example 1.

[0126] 7) Set the negative voltage to (-2±1)KV and the positive voltage to (7±0.5)KV. Start electrospinning and observe the spinning effect until continuous and uniform yarn is ejected. Set the electrospinning time to 30min~50min. After the first electrospinning, rotate the release paper (with a spinning layer on the surface) 90 degrees and then fix it. Set the electrospinning time to 30min~50min again and repeat the operation for the second electrospinning.

[0127] After the second electrospinning is completed, the release paper (with a spinning layer on the surface) is rotated 90 degrees and then fixed to set the electrospinning time to 30 minutes to 50 minutes, and the operation is repeated for the third electrospinning.

[0128] After the third electrospinning, the release paper (with a spinning layer on the surface) was rotated 90 degrees and then fixed to set the electrospinning time to 30 min to 50 min, and the operation was repeated for the fourth electrospinning.

[0129] After the fourth electrospinning, the degradable repair film layer was removed from the release paper.

[0130] 4. Same as Example 1.

[0131] 5. Same as Example 1.

[0132] 6. Same as Example 1.

[0133] The difference between this embodiment and embodiment 1 is that the degradable repair film layer is a four-layer fiber oriented sheet structure.

[0134] Example 3

[0135] A method for preparing an oral repair membrane, the preparation method comprising:

[0136] 1. Preparation of protective film layer: same as Example 1.

[0137] 2. Preparation of porous structure layer: same as Example 1.

[0138] 3. Preparation of degradable repair film layer:

[0139] (1) Preparation of degradable polyurethane (PU) solution: same as in Example 1.

[0140] (2) Electrospinning:

[0141] 1)~6) are the same as in Example 1.

[0142] 7) Set the negative voltage to (-2±1)KV and the positive voltage to (7±0.5)KV. Start electrospinning and observe the spinning effect until continuous and uniform yarn is ejected. Set the electrospinning time to 30min~50min. After the first electrospinning, rotate the release paper (with a spinning layer on the surface) 90 degrees and then fix it. Set the electrospinning time to 30min~50min again and repeat the operation for the second electrospinning.

[0143] After the second electrospinning is completed, the release paper (with a spinning layer on the surface) is rotated 90 degrees and then fixed to set the electrospinning time to 30 minutes to 50 minutes, and the operation is repeated for the third electrospinning.

[0144] After the third electrospinning, the release paper (with a spinning layer on the surface) was rotated 90 degrees and then fixed to set the electrospinning time to 30 min to 50 min, and the operation was repeated for the fourth electrospinning.

[0145] After the fourth electrospinning, the release paper (with a spinning layer on the surface) was rotated 90 degrees and then fixed to set the electrospinning time to 30 min to 50 min, and the operation was repeated for the fifth electrospinning.

[0146] After the fifth electrospinning, the release paper (with a spinning layer on the surface) was rotated 90 degrees and then fixed to set the electrospinning time to 30 min to 50 min, and the operation was repeated for the sixth electrospinning.

[0147] After the sixth electrospinning, the electrospun membrane was removed from the release paper to obtain a degradable repair membrane layer.

[0148] 4. Same as Example 1.

[0149] 5. Same as Example 1.

[0150] 6. Same as Example 1.

[0151] The difference between this embodiment and embodiment 1 is that the degradable repair film layer is a six-layer fiber oriented sheet structure.

[0152] Example 4

[0153] A method for preparing an oral repair membrane, the preparation method comprising:

[0154] 1. Preparation of protective film layer: same as Example 1.

[0155] 2. Preparation of porous structure layer: same as Example 1.

[0156] 3. Preparation of degradable repair film layer:

[0157] (1) Preparation of degradable polyurethane (PU) solution: same as in Example 1.

[0158] (2) Electrospinning:

[0159] 1)~6) are the same as in Example 1.

[0160] 7) Set the negative voltage to (-2±1)KV and the positive voltage to (7±0.5)KV. Start electrospinning and observe the spinning effect until continuous and uniform yarn is ejected. Set the electrospinning time to 30min~50min. After the first electrospinning, rotate the release paper (with a spinning layer on the surface) 45 degrees and then fix it. Set the electrospinning time to 30min~50min again and repeat the operation for the second electrospinning.

[0161] After the second electrospinning is completed, the electrospun membrane is removed from the release paper to obtain a degradable repair membrane layer.

[0162] 4. Same as Example 1.

[0163] 5. Same as Example 1.

[0164] 6. Same as Example 1.

[0165] The difference between this embodiment and embodiment 1 is that the oriented fiber direction of the oriented fiber sheet electrospun for the first time in the degradable repair film layer crosses with the oriented fiber direction of the oriented fiber sheet electrospun for the second time at 45 degrees.

[0166] Comparative Example 1

[0167] A method for preparing an oral repair membrane, the preparation method comprising:

[0168] 1. Preparation of protective film layer: same as Example 1.

[0169] 2. Preparation of porous structure layer: same as Example 1.

[0170] 3. Preparation of degradable repair film layer:

[0171] (1) Preparation of degradable polyurethane (PU) solution: same as in Example 1.

[0172] (2) Electrospinning:

[0173] In this comparative example, except that the rotation speed of the receiving device is (500±100) rpm / min, other process parameters are the same as those of Example 1, and the produced fiber sheet does not have an oriented structure.

[0174] 4. Same as Example 1.

[0175] 5. Same as Example 1.

[0176] 6. Same as Example 1.

[0177] The difference between this embodiment and embodiment 1 is that the fibers in the degradable repair film layer do not have an oriented structure.

[0178] Comparative Example 2

[0179] A method for preparing an oral repair film. Except that the oral repair film of this comparative example "does not include a protective film layer", the selection and preparation method of the porous structure layer and the degradable repair film layer in the oral repair film are the same as those in Example 1.

[0180] Comparative Example 3

[0181] A method for preparing an oral repair film. Except that the oral repair film of this comparative example "does not include a porous structure layer", the selection of the protective film layer and the degradable repair film layer in the oral repair film and the preparation method are the same as those in Example 1.

[0182] Comparative Example 4

[0183] 1. Preparation of protective film layer: same as Example 1.

[0184] 2. Preparation of porous structure layer: same as Example 1.

[0185] 3. Preparation of degradable repair film layer:

[0186] (1) Preparation of degradable polyurethane (PU) solution: Same as Example 1.

[0187] (2) Electrospinning:

[0188] 1)~6): Same as Example 1

[0189] 7) Set the negative voltage to (-3±1)KV and the positive voltage to (9±0.5)KV. Start electrospinning and observe the spinning effect until continuous and uniform yarn is ejected. Set the electrospinning time to 30min~1h. After the first electrospinning, keep the release paper (with a spinning layer on the surface) fixed without rotating. Set the electrospinning time to 30min~1h again and repeat the operation for the second electrospinning.

[0190] After the second electrospinning is completed, the electrospun membrane is removed from the release paper to obtain the initial oral repair membrane.

[0191] 4. Same as Example 1.

[0192] 5. Same as Example 1.

[0193] 6. Same as Example 1.

[0194] The orientation directions of the two fiber orientation sheets in the degradable repair film layer of this comparative example are in the same direction.

[0195] Test Example 1

[0196] (I) Mechanical properties test:

[0197] Experimental methods:

[0198] (1) Tensile strength: First, cut the dry sample to be tested into multiple specimens with appropriate width, flat edges, parallel sides and no visible defects, then turn on the tensile testing machine, set the parameters, perform the tensile strength test, and process the data. Tensile strength (N / cm) = tension (N) / specimen width (cm).

[0199] (2) Elongation at break: First, cut the dry sample to be tested into multiple specimens with appropriate width, flat edges, parallel sides and no visible defects. Then turn on the tensile testing machine, set the parameters, perform the elongation at break test, and process the data; elongation at break (%) = (elongation at break of specimen / original length of specimen) × 100%.

[0200] (3) Suture strength: First, cut the dry sample to be tested into multiple specimens with appropriate width, flat edges, parallel sides and no visible defects. Then, use a suture thread of the selected specification to sew one end of the sample to be tested with two needles and tie a knot to fix it. Suture multiple specimens in the same way, and test the thickness of each sample with a thickness gauge. Then, use a tensile testing machine to clamp the sample to be tested and test the suture strength of the suture to obtain the test data.

[0201] (4) Tear strength: First, cut the dry sample to be tested into samples with appropriate width, flat edges, parallel sides and no visible defects. Use a cutter to cut along the center of the sample to prepare a trouser-shaped sample. The cutting edge of the cutter must be sharp and must not have curled edges or gaps. The cut is located in the center of the sample width. Use a thickness gauge to measure 3 points in the tear area of ​​the sample, and the arithmetic average is the thickness. Turn on the tensile testing machine, clamp the sample to be tested, and perform a tear strength test to obtain the test data.

[0202] (5) Bursting strength: First, cut the dry sample to be tested into multiple square specimens with appropriate width, flat edges, parallel sides and no visible defects, and use a thickness gauge to test the thickness of each sample. Then, turn on the tensile testing machine, clamp the sample to be tested, and perform the bursting strength test to obtain the test data.

[0203] Experimental results:

[0204]

[0205]

[0206] From the above results, it can be seen that the tensile strength of the oral repair membrane of the present invention reaches 6.41-30.27N / cm, the elongation at break reaches 80.78-652.16%, the suture strength reaches 4.29-20.77N, the tear strength reaches 8.54-40.53N, and the bursting strength reaches 80.23-406.45kPa. The oral repair membrane of the present invention has good mechanical properties and can well meet the mechanical property requirements of oral tissue repair for the oral repair membrane.

[0207] (II) Cell proliferation assay:

[0208] Experimental methods:

[0209] Each sample was made into a circular specimen of the same size as the bottom area of ​​a 48-well plate using a puncher. After sterilization, it was placed in a 48-well cell culture plate and immersed in MEM medium containing 10% FBS at 37°C and 60 rpm for 24 hours, with the outer layer of the specimen facing the bottom of the well.

[0210] The whole process was performed in a clean bench to ensure aseptic operation. Mouse fibroblast L-929 cells grown to the logarithmic growth phase were digested with 0.25% trypsin (containing EDTA), and the cell suspension was centrifuged (1000rpm, 5min) after digestion, the supernatant was discarded, and the cells were resuspended with MEM medium, and the cells were counted to obtain a cell suspension of 2×104 cells / mL; the soaked samples were respectively spread in 48-well plates, and 3 samples were operated in parallel in each group.

[0211] 2×10 4 300 μL of a cell suspension of 10 cells / mL was seeded into each well of a 48-well plate and cultured in a cell culture incubator (37° C., 5% CO 2 , >90% humidity).

[0212] After 24h, 3d and 6d, the original culture medium in the 48-well plate was discarded, and 30μL of MTT solution (concentration of 5mg / mL) was added to the corresponding wells of the 48-well plate, and cultured in a 37°C, 5% carbon dioxide incubator. After 4h, the supernatant was removed, and 200μL of isopropanol was added to each well to dissolve the crystals. After the crystals were completely dissolved, the sample was taken out and the liquid in the well was transferred to a 96-well plate, 100μL per well, and the absorbance value at a wavelength of 570nm was measured on an enzyme reader, and compared with the OD value of the blank group to determine whether the cells had a proliferation effect on the surface of the material and calculate its cell proliferation rate.

[0213] Experimental results:

[0214]

[0215]

[0216] It can be seen from the above results that Examples 1 to 4 have very good cell proliferation rate effects, especially the cell proliferation rate effects of the two-layer fiber oriented sheets of Examples 1 and 4 are more significant.

[0217] (III) Cell migration assay:

[0218] Experimental methods:

[0219] 1. Cut the sample into a circular shape similar to the bottom of the 48-well plate, sterilize it, and soak it in MEM culture medium containing 10% FBS at 37°C for 24 hours to fully immerse the sample.

[0220] 2. The whole process was performed in a clean bench to ensure aseptic operation. Mouse fibroblast L-929 cells grown to the logarithmic growth phase were digested with 0.25% trypsin (containing EDTA). After digestion, the cell suspension was centrifuged (1000 rpm, 5 min), the supernatant was discarded, and the cells were resuspended in MEM medium. After counting and dilution, 20×10 4The soaked samples were spread in 48-well plates, and 3 samples were processed in parallel in each group.

[0221] 3. 20×10 4 50 μL of cell suspension per well was added to three 48-well plates. The cell suspension was uniformly added to the upper left corner of the 48-well plate. After 4 hours, an appropriate amount of culture medium was added to the plate wall. The amount of culture medium should just cover the sample (without affecting the position of the cells to prevent the complete evaporation of the culture medium and affect the cell viability). After 24 hours, 1 mL of culture medium was added to each well and added to the plate wall to avoid affecting the position of the cells. The cells were cultured in a cell culture incubator for 1, 3, and 6 days respectively.

[0222] 4. Cell staining and observation: When the sampling time point is reached, the culture medium in the well plate is aspirated and rinsed twice with PBS. After rinsing, 300uL of 10ug / mL acridine orange solution is added to each well for staining. Incubate at room temperature in the dark for 15-20min. After incubation, use a fluorescence microscope (excitation filter wavelength 488nm, blocking filter wavelength 515nm) to observe the cell proliferation or migration status at the upper left corner, upper right corner, lower left corner, and lower right corner of the electrospun membrane. For experimental results, see Figure 3 :

[0223] Figure 3 The cells provided in this test example were co-incubated with the oral repair membrane of Example 1 of the present application for 6 days and the cell migration results on one side of the degradable repair membrane layer.

[0224] Analysis of cell migration experiment results: 50uL L929 cell suspension was added to the upper left of an oriented patch with a diameter of 1cm. After 6 days of incubation, it was stained with acridine orange solution. The results showed that the cells were evenly distributed on the oriented patch, and a large number of cells were distributed in the center of the patch, indicating that the cells showed good migration behavior on the oriented patch.

[0225] (IV) Degradation experiment:

[0226] Experimental methods:

[0227] (1) Preparation of enzyme buffer:

[0228] Prepare a solution containing potassium dihydrogen phosphate and disodium hydrogen phosphate with sterile double distilled water, containing 1.653g KH2PO4 and 7.744g NaH2PO4 per liter of buffer. The pH value of the buffer solution should be 7.4±0.2, and artificial saliva is added after sterilization. The salts used in the preparation are analytically pure and dried to constant weight.

[0229] The artificial saliva described in this embodiment is prepared according to the artificial saliva ratio in GB / T 18886-2019 Textile color fastness test for color fastness to saliva.

[0230] (2) Degradation test:

[0231] The sample is vacuum dried at room temperature to constant weight, and the weight of the sample is accurately measured. The sample is placed in a glass container, covered with enzyme buffer and the container is sealed. The minimum volume of the buffer should be 10 mL, and the ratio of the volume of the buffer (mL) to the mass of the sample (g) should be greater than or equal to 30:1. The sample is maintained at 37 ± 1 ° C in a constant temperature water bath or oven.

[0232] (3) Determination of degradation cycle:

[0233] For sample degradation testing, samples were taken at 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks and 13 weeks for mechanical property testing.

[0234] After wiping the liquid on the sample surface with a paper towel, load the sample into the tensile testing machine fixture and test it directly. The test rate is set to 50mm / min. The test is terminated when the sample cannot be tested for mechanical properties, and the time point for stopping the test is the degradation cycle of the sample.

[0235] Experimental results:

[0236]

[0237]

[0238] From the above results, it can be seen that the oral repair membrane prepared by the present invention completely degrades in 9 weeks to 13 weeks, which can provide sufficient strength support for oral tissue repair to meet clinical needs, and does not exist in the body for too long.

[0239] (V) Flexibility test:

[0240] Experimental methods:

[0241] The samples of Examples 1 to 4 and Comparative Examples 1 to 3 were cut into specimens of 100 mm×100 mm in size and tested according to the softness determination method in GB / T8942-2016.

[0242] Experimental results:

[0243]

[0244]

[0245] From the above results, it can be seen that the oral repair membrane prepared by the present invention is soft in texture, and the softness reaches 209.62mN-474.64mN. The smaller the softness value, the softer the oral repair membrane is, and the better the adhesion is.

[0246] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. An oral repair membrane, characterized in that: The oral repair film comprises: a degradable repair film layer and a porous structure layer and a protective film layer which are sequentially compounded on the degradable repair film layer; The degradable repair film layer is prepared by compounding multiple fiber oriented sheets; The fiber orientation sheet is formed into oriented fibers by an electrostatic spinning method, and the orientation directions of the oriented fibers in the fiber orientation sheet are consistent; The orientation directions of the fibers in each fiber orientation sheet in the degradable repair film layer are different.

2. The oral repair membrane according to claim 1, characterized in that: The oriented fibers are obtained by electrostatic spinning of degradable polyurethane or polylactic acid; Preferably, the weight average molecular weight Mw of the degradable polyurethane is 5W to 12W; Preferably, the weight average molecular weight Mw of the polylactic acid is 8W to 15W; Preferably, the diameter of the oriented fibers in the fiber oriented sheet is 1 nm to 20 μm, and the porosity is 20 to 80%.

3. The oral repair membrane according to claim 1, characterized in that: The porous structure layer is prepared from degradable polyurethane by a casting method; The porous structure layer has a pore structure, the porosity of the porous structure layer is 5-20%, and the pore size range of the micropores is less than 100um; And / or, the protective film layer is prepared from degradable polyurethane by a casting method, and the protective film layer is a smooth and dense film layer without a pore structure.

4. The oral repair membrane according to claim 1, characterized in that: The total thickness of the oral repair film is 0.2-0.6 mm, wherein: The thickness of the degradable repair film layer is 0.02-0.5 mm, the thickness of the porous structure layer is 0.03-0.2 mm, and the thickness of the protective film layer is 0.03-0.1 mm.

5. The oral repair membrane according to claim 1, characterized in that: The oral repair membrane has a tensile strength of 6-30 N / cm, a breaking elongation of 80%-650%, a suture strength of 4-20 N, and a softness of less than 500 mN.

6. The oral repair membrane according to claim 1, characterized in that: The degradable repair film layer is prepared by compounding two layers of fiber oriented sheets; Preferably, the two-layer fiber-oriented sheet comprises a first fiber-oriented sheet and a second fiber-oriented sheet, and the first fiber-oriented sheet and the second fiber-oriented sheet have different orientation directions and are at an angle of 90°±15°.

7. A method for preparing an oral repair membrane according to any one of claims 1 to 6, characterized in that: The preparation method comprises: (A) preparing a protective film layer and a porous structure layer in sequence by a tape casting method to obtain a composite intermediate; then placing the composite intermediate on a substrate, with the protective film layer in contact with the substrate and the porous structure layer facing outward; and then preparing a degradable repair film layer on the porous structure layer by an electrospinning method to obtain a preliminary oral repair film; (B) removing the solvent and drying the initial oral repair membrane in sequence to obtain an oral repair membrane.

8. The method for preparing the oral repair membrane according to claim 7, characterized in that: The preparation method of the degradable repair film layer comprises: S1: dissolving the degradable polyurethane in a solvent to obtain a spinning solution; S2: electrospinning the spinning solution to obtain a fiber orientation sheet A, then adjusting the electrospinning direction, and electrospinning on the fiber orientation sheet A to obtain a fiber orientation sheet B intersecting the orientation direction of the fiber orientation sheet A, thereby obtaining a degradable repair film layer composed of two layers of fiber orientation sheets; S3: Repeat step S2 to obtain a degradable repair film layer composed of multiple fiber oriented sheets.

9. The method for preparing the oral repair membrane according to claim 8, characterized in that: In the step S1, the concentration of the degradable polyurethane in the spinning solution is 15-40% (w / v); Preferably, the solvent in the spinning solution is a single solvent of N,N-dimethylformamide, a single solvent of N,N-dimethylacetamide, a mixed solvent of N,N-dimethylformamide and any one or more of tetrahydrofuran, dichloroethane or dimethyl ether, or a mixed solvent of N,N-dimethylacetamide and any one or more of tetrahydrofuran, dichloroethane or dimethyl ether; More preferably, the solvent in the spinning solution is a mixed solvent of N,N-dimethylformamide and tetrahydrofuran, wherein the volume ratio of N,N-dimethylformamide to tetrahydrofuran is 10:1-3:7, wherein the preferred volume ratio is 10:1-1:

1.

10. The method for preparing the oral repair membrane according to claim 8, characterized in that: The conditions for electrospinning in step S2 include: Negative voltage: -1KV~-4KV, positive voltage: 7KV~10KV, injection speed of the nozzle: 0.1-0.4ml / min; rotation speed of the receiving device: (1000, 2500) rpm / min, distance between the nozzle and the receiving device: 0~50 (10-30) mm; translation speed of the nozzle is (0, 500) mm / min.

Images