KR100564366B1 - Nonwoven nanofibrous membranes for guided tissue regeneration and their fabrication method - Google Patents

Abstract

The present invention relates to a tissue regeneration shielding membrane and a method for manufacturing the same, and more particularly, the present invention relates to a tissue regeneration shielding membrane and a method for producing the non-woven fabric of nanofibers prepared from a mixture of natural and synthetic polymers.

The tissue regeneration shielding membrane according to the present invention has a certain strength, biocompatibility and biodegradability, and when the drug is added to maintain the sustained release system of the drug. In addition, the tissue regeneration shielding film according to the present invention can control the thickness of the shielding membrane by adjusting the thickness of the nanofibers and the degree of nanofibers accumulation when forming the nonwoven fabric, and can also control the pore size of the porous structure to use the shielding membrane It can be used depending on the conditions. In addition, since the nanofibers constituting the tissue regeneration shielding membrane according to the present invention can be produced from a mixture of a natural polymer and a synthetic polymer at a time, it can be produced simply without a lamination process.

Shielding film, nanofiber, nonwoven fabric, chitosan, lactic acid

Description

TECHNICAL FIELD The shielding membrane for tissue regeneration using a nanofiber type nonwoven fabric and a method of manufacturing the same {NONWOVEN NANOFIBROUS MEMBRANES FOR GUIDED TISSUE REGENERATION AND THEIR FABRICATION METHOD}             

1 is a scanning electron micrograph of the surface of the tissue regeneration shielding membrane (Example 1) according to the present invention,

2 is a scanning electron micrograph of the surface of the tissue regeneration shielding film (Example 2) according to the present invention,

3 is a graph showing the concentration of tetracycline released from the tissue regeneration shielding membrane according to the elapsed time according to the present invention.

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a tissue regeneration shielding membrane and a method of manufacturing the same, and more particularly, to a tissue regeneration shielding membrane in which nanofibers prepared from a mixture of a natural polymer and a synthetic polymer are in the form of a nonwoven fabric.

As a method of regenerating alveolar bone damaged by periodontal disease, there is a method of inducing solid by filling the damaged area by autografting. Another method is to use the bones of human or animal from which immunogenicity has been removed as an artificial bone substitute or to use commercially available hydroxide apatite.

In recent years, attempts have been made to improve the healing of damaged periodontal tissues by introducing artificial membranes into tissues, improve restoration of complete periodontal tissues, improve bone graft outcomes, and induce the production of new alveolar bone. The shield used in this technique isolates and blocks the connective tissue around the damaged area, creating new alveolar bone and periodontal ligament tissue to facilitate regeneration of the periodontal tissue. In other words, blocking the damaged area with other surroundings to prevent the gingival fibroblasts to invade, so that the cells of the bone and periodontal ligament regeneration in the tissue is not disturbed and the new periodontal tissue is regenerated.

Early screening studies used non-degradable materials such as polytetrafluoroethylene, cellulose acetate, silicone rubber or polyurethane. However, the shielding membrane made of non-degradable material requires secondary surgery to remove the shielding membrane again after the periodontal bone is formed, and in this process, unnecessary inflammation or tissue necrosis may occur, abscesses to the neoplastic tissue, and the epithelial downtake Proliferation (epithelial down growth), periodontal pocket formation, inflammation may occur.

Recently, studies using biodegradable polymers such as polyester and collagen have been reported. The use of biodegradable barriers has been reported to eliminate the need for reoperations to remove the barriers and to show no significant difference in tissue regeneration compared to shields made of non-degradable materials. However, even in the case of applying a shielding membrane manufactured using biodegradable material in clinical practice, it does not have sufficient strength and thus cannot maintain a certain shape, and does not secure a space for tissue growth and prevents secondary inflammation caused by the material. There is a problem that occurs.

Therefore, the shielding membrane should have the strength and structure to maintain the space for the periodontal bone growth, and should be compatible with the bone cells when applied to the periodontal bone injury site to induce the attachment and proliferation of the bone cells. It should be porous to facilitate the supply of moisture.

As part of this research, a shielding membrane produced by applying a biodegradable polymer selected from drugs, homopolymers of lactic acid, copolymers of lactic acid and glycolic acid, or mixtures thereof to a mesh made of polyglycolic acid has been reported (registered patent 0180585). The shielding membrane is tensioned by a polymer solution containing a biodegradable polymer is applied to the polyglycolic acid mesh and then precipitated by the evaporation of the solvent, the precipitated polymer is attached to the polyglycolic acid mesh in the surroundings to prevent movement It is to be produced, so that the micro-pores are formed in the space between the mesh. However, in order to facilitate the regeneration of the periodontal tissue when applying the shielding membrane, it is necessary to isolate the periodontal tissue and the connective tissue, so it is necessary to adjust the pore size of the micropores formed in the shielding membrane.

In addition, a membrane for inducing tissue regeneration using a chitosan which is a natural polymer and a biodegradable polymer which is a synthetic polymer has been reported (see Patent Publication No. 2003-2224). Specifically, a biodegradable polymer solution is applied to a nonwoven fabric made of chitosan to form a polymer membrane, and a nonwoven fabric made of chitosan is laminated thereon, followed by compression to prepare a shielding membrane. The nonwoven fabric made of biodegradable polymer in the shielding membrane is provided with micropores to provide conditions for growing periodontal bone, and the nonwoven fabric is sequentially laminated to improve mechanical strength. However, the shielding film is manufactured according to the step of preparing a nonwoven fabric made of chitosan, applying a biodegradable polymer solution on the nonwoven fabric to form a polymer film, and laminating a nonwoven fabric made of chitosan on the polymer film. There is a problem with this complexity.

The present invention is designed to solve the above problems, the object of the present invention has a certain strength and biocompatibility and biodegradability, easy to control the pore size of the micropores, and can be produced by a simple manufacturing process and It is to provide a shielding film containing a synthetic polymer and a method of manufacturing the same.

The present invention provides a mask for regenerating tissue in which a nanofiber made from a mixture of a natural polymer and a synthetic polymer is in the form of a nonwoven fabric.

Hereinafter, the present invention will be described in detail.

The present invention relates to a mixture of synthetic polymers selected from the group consisting of natural polymers selected from the group consisting of chitosan, collagen and alginic acid and lactic acid, copolymers of lactic acid and glycoic acid, homopolymers of glycoic acid, and mixtures thereof. Provided is a membrane for tissue regeneration in which the nanofibers produced are in the form of a nonwoven fabric.

In the present invention, the natural polymer constituting the tissue regeneration shielding membrane is selected from the group consisting of chitosan, collagen and alginic acid having a molecular weight of 80,000 to 150,000, and preferably used as chitosan or collagen. In particular, chitosan is known to be similar to the structure of glycosaminoglycan, so it is excellent in biocompatibility and has a wound repair effect by activating a fibrogenic mediator. In addition, chitosan contains an amine group in the structure, thereby neutralizing the acid of polyesters such as lactic acid and glycoic acid, thereby reducing the inflammatory response and tissue toxicity that can be locally generated by the acid. In addition, it is possible to maintain the mechanical properties when manufacturing the nanofibers to increase the stability of the nanofibers, to maintain a constant voids and form even when manufactured in the form of non-woven fabric, and to withstand the pressure to the damage site sufficiently have.

As a solvent capable of dissolving the natural polymer, 1,1,1,3,3,3-hexafluoropropyl alcohol may be used, and in addition to this, the viscosity may be adjusted by adding methylene chloride or acetone. It doesn't happen. Chitosan in the natural polymer is a mixed solvent of 1,1,1,3,3,3-hexafluoropropyl alcohol and methylene chloride, and is preferably added at 0.5 to 2% with respect to the solvent, and collagen is 1, Preference is given to adding 8-12% using 1,1,3,3,3-hexafluoropropyl alcohol.

In the present invention, the synthetic polymer constituting the shielding membrane for tissue regeneration is selected from the group consisting of a homopolymer of lactic acid having a molecular weight of 50,000 to 300,000, a copolymer of lactic acid and glycoic acid, a homopolymer of glycoic acid, and a mixture thereof. Preferably a homopolymer of lactic acid (poly lactic acid) is used. In particular, the homopolymer of lactic acid is a biodegradable synthetic polymer licensed by the US Food and Drug Administration, which is stable in vivo and capable of practical clinical application. In addition, there is crystallinity in the structure of the homopolymer of lactic acid, so that it is possible to maintain stability even when prepared in the form of nanofibers by electrospinning, and even in an aqueous solution of body temperature, pores are not reduced or the shape is not changed.

As a solvent capable of dissolving the synthetic polymer, 1,1,1,3,3,3-hexafluoropropyl alcohol, acetone, chloroform, methylene chloride and a mixed solvent thereof may be used, but are not limited thereto. Synthetic polymer may be added in 5 to 10% by weight based on the solvent.

The tissue regeneration shielding membrane according to the present invention allows the natural polymer and the synthetic polymer to be mixed in a weight ratio of 1: 4 to 4: 1, and preferably in a weight ratio of 1: 2.

Nanofibers constituting the tissue regeneration shielding membrane according to the present invention can be prepared by electrospinning the mixture containing the natural polymer and synthetic polymer in an electrospinner.

Specifically, in step 3 to prepare a nanofiber using the spinning solution electrospinner.

In the present invention, as the electrospinner, a DH High Voltage Generator (model name: CPS-40K03VIT) sold by Chungpa EMT Co., Ltd. (Korea) may be used. At this time, it is possible to adjust the shape to be produced according to the spinning conditions, such as the concentration of the spinning solution and the spinning distance, in this step to produce nanofibers by optimally adjusting the conditions of the electrospinner. The condition of the electrospinner is preferably 10 to 20 cm in radiation distance and 15 to 25 V in voltage.

As shown in FIGS. 1 and 2, the membrane for regenerating tissue according to the present invention has a nanofiber having a thickness of 10 nm or more in the form of a nonwoven fabric, and includes a porous structure having a pore size of 2 to 10 μm formed of nanofibers. Doing. Such a porous structure may block the invasive connective tissue cells, which are regeneration inhibitory cells of 10 to 30 μm in size. Therefore, the porous structure according to the present invention can induce tissue regeneration while blocking regeneration inhibitory cells, thereby securing a selective cell growth induction function of the shielding membrane. In particular, the tissue regeneration shielding membrane made of a nonwoven fabric of nanofibers prepared from a mixture comprising a homopolymer of chitosan and lactic acid can maintain a certain strength and maintain a constant pore size due to its properties.

In addition, the tissue regeneration shielding film according to the present invention can control the thickness of the shielding membrane by adjusting the thickness of the nanofibers and the degree of nanofibers accumulation when forming the nonwoven fabric, and can also control the pore size of the porous structure to use the shielding membrane It can be used depending on the conditions. The thickness of the shielding film is preferably 0.1 to 5 mm, and the size of the voids is preferably 2 to 10 μm, but is not limited thereto.

Specifically, the nanofibers can control the thickness of the nanofibers by controlling the diameter of the inlet of the electrospinner from which the mixture is radiated, the rate at which it is released, the voltage and electric field, the properties of the polymer to be added, and the composition and concentration of the mixture. Preferably, the thickness of the nanofibers is 0.001 to 10 μm, but is not limited thereto.

In addition, the degree of accumulation of the nanofibers can be controlled by adjusting the accumulation time and the voltage distance to adjust the accumulation degree and the voids.

In addition, the tissue regeneration shielding membrane according to the present invention is made of a nanofiber from a mixture comprising a biodegradable and biocompatible natural polymer and a synthetic polymer made of a non-woven membrane, especially after the nanofiber is manufactured It can be made into nonwoven form without treatment. Therefore, after forming the basic skeleton which comprises a shielding film, it can manufacture easily, without the need to apply | coat with a biodegradable polymer. And, since the nanofibers can be produced from a mixture of a natural polymer and a synthetic polymer at a time, the tissue regeneration shielding film according to the present invention is made of a non-woven fabric made of a natural polymer, and then coated with a synthetic polymer, the non-woven fabric made of a natural polymer again It can be produced simply without the need for lamination.

In addition, when comparing the tensile strength of RESOLUT® or biofix® that is commercially used with the tissue regeneration barrier according to the present invention, the tissue regeneration barrier according to the present invention showed similar or higher tensile strength.

The tissue regeneration shielding membrane according to the present invention may further include a drug, a growth factor or a ceramic in addition to the natural polymer and the synthetic polymer.

The drug may include an antibiotic for reducing the inflammatory response or a drug for treating periodontal disease. The drug for the treatment of periodontal disease is selected from the group consisting of mephenamic acid, ibuprofen, flubiprofen, indomethacin, naproxen, metronidazole, tetrasiacline, minocycline, oxytetracycline and mixtures thereof. The drug for treating periodontal disease may be previously dispersed in a mixture of the natural polymer and the synthetic polymer or encapsulated in an emulsion to be radiated during electrospinning to be included in the shielding membrane, or the shielding membrane may be prepared first, and then Soaking in the solution containing the drug may be included in the shielding membrane, but is not limited thereto.

The membrane for regenerating tissue according to the present invention prepared by further adding the drug may maintain the sustained release system of the drug.

The growth factor is selected from the group consisting of platelet-derived growth factor, insulin-like growth factor, epithelial growth factor, tumor growth factor or mixtures thereof. The growth factor is added in an amount of 5 to 20% by weight based on the total polymer weight of the natural polymer and the synthetic polymer.

The ceramic may be used hydroxyapatite, tricalcium phosphate (calcium phosphate) or calcium metaphosphate (calcium metaphosphate). The ceramic is added to enhance the ex vivo matrix component and / or mechanical strength and bone tissue regeneration effect to enhance biocompatibility. In particular, the hydroxide apatite chemically and crystallographically has properties similar to that of inorganic components in bones or teeth, and thus has an advantage of excellent adhesion and stability with surrounding bones or tissues when implanted in the human body. Therefore, the apatite hydroxide was slowly released from the shielding film prepared by further adding apatite hydroxide to stabilize the bone growth.

The present invention also provides a method for producing the tissue regeneration shielding membrane. Specifically, the method for producing a tissue regeneration shielding membrane according to the present invention is a homopolymer of lactic acid and a natural polymer selected from the group consisting of chitosan, collagen and alginic acid, a copolymer of lactic acid and glycoic acid, a homopolymer of glycoic acid and their Electro-spinning a mixed polymer solution containing a synthetic polymer selected from the group consisting of mixtures to produce nanofibers, and manufacturing the nanofibers in the form of a nonwoven fabric.

The mixed polymer solution preferably comprises a solvent, 0.5-10% of the natural polymer of the solvent and 5-10% of the synthetic polymer of the solvent. In addition, the above-described drugs, growth factors, ceramics or mixtures thereof may be further included.

The solvent is selected from the group consisting of 1,1,1,3,3,3-hexafluoropropyl alcohol, methylene chloride, acetone, chloroform and mixtures thereof.

Electrospinning the mixed polymer solution to make nanofibers includes making a polymer mixed solution and filling and spinning the mixed solution.

In the manufacturing of the nanofibers in the form of nonwoven fabric, in the process of spinning the nanofibers, a step of preparing a shielding membrane having a predetermined thickness of 0.3 to 0.5 mm by spinning a mixed polymer solution on the current collector plate using an electrospinner for a predetermined time or more is remaining solvent. Removing the step.

Hereinafter, the present invention will be described in more detail based on the following examples. The following examples are only intended to illustrate preferred embodiments of the present invention, but the scope of the present invention is not limited by the following examples.

<Example 1>

Chitosan solution was prepared by dissolving chitosan in a 2: 1 (volume ratio) solution of 1,1,1,3,3,3-hexafluoropropyl alcohol and methylene chloride. A homopolymer solution of lactic acid at a concentration of 10% was prepared by dissolving a homopolymer of lactic acid with a molecular weight of 100,000 in the solution. The mixed polymer solution was prepared by mixing the prepared chitosan solution and the lactic acid homopolymer solution in a ratio of 1: 1, 1: 2, 1: 4, 2: 1, or 4: 1. . The prepared mixed polymer solution was filled in a reservoir, and nanofibers were prepared by electrospinning. For the electrospinning method, an electrospinning machine (DH High Voltage Generator, CPS-40K03VIT) was used, and the inlet diameter of the electrospinner was 0.5 mm, the discharge rate of the solution was 0.5 ml / min, the voltage was 20 V, and the distance of the electric field was 20 cm. It was. By spinning the solution on the current collector for a predetermined time or more to prepare a nanofiber nonwoven fabric of 0.2mm-0.5mm thickness.

The surface of the tissue regeneration shielding membrane in which the weight ratio of the homopolymer of chitosan and lactic acid in the prepared tissue regeneration shielding membrane was 1: 2 was observed with a scanning electron microscope, and the results are shown in FIG. 1. As shown in FIG. 1, it can be seen that the nanofibers formed a non-woven membrane, and the porous structure can be confirmed with the nanofibers. The pore size of the porous structure was 2 to 10 μm. The pore size was measured by observing a photograph of a differential scanning microscope.

<Example 2>

After dispersing tetracycline at a concentration of 5% in a mixed polymer solution in which the weight ratio of the homopolymer of chitosan and lactic acid prepared in Example 1 was 1: 2, the same procedure as described above was performed to prepare a tissue regeneration shielding membrane.

The surface of the prepared tissue regeneration shielding membrane was observed under a scanning electron microscope, and the results are shown in FIG. 2. As shown in FIG. 2, it can be seen that the nanofibers formed a non-woven membrane, and the porous structure formed of the nanofibers can be confirmed.

<Example 3>

An acid soluble collagen obtained from the skin of a young cow was dissolved in a 1,1,1,3,3,3-hexafluoropropyl alcohol solution to prepare a collagen solution at a concentration of 10% by weight. After adding the collagen solution at a concentration of 8% by weight to a mixed polymer solution having a weight ratio of the homopolymer of chitosan and lactic acid of 1: 2 in Example 1, the tissue regeneration shielding membrane was prepared in the same manner as in Example 1. Prepared.

<Example 4>

20 wt% of sintered apatite hydroxide (diameter in diameter) was added to the mixed polymer solution in which the weight ratio of the chitosan and lactic acid homopolymer in Example 1 was 1: 2, and then in the same manner as in Example 1. A shield for tissue regeneration was prepared.

Experimental Example 1 Measurement of Mechanical Strength of Tissue Regeneration Shielding Membrane

Tensile strength was measured to determine the mechanical strength of the tissue regeneration shielding membrane according to the present invention. Tensile strength of Instron (Instron 8511; Instron corp., USA) was used, 500N load cell was used at a rate of 1 mm / minute, the sample was used for the tissue regeneration shielding membrane prepared in Examples 1 and 4 Was used. The results are shown in Table 1.

In addition, as a comparative example, commercially available RESOLULT® or biofix used in the clinic was used, and tensile strength was measured in the same manner as described above. The results are shown in Table 1.

Tensile Strength (MPa) Example 1 10.936 ± 0.846 Example 4 15.919 ± 0.728 RESOLULT 11.494 ± 0.267 biofix 14.772 ± 0.696

As shown in Table 1, the tissue regeneration shielding membrane prepared in Examples 1 and 4 according to the present invention had a similar or higher tensile strength than RESOLULT® or biofix®. In particular, the tissue regeneration shielding membrane of Example 4 prepared by adding apatite hydroxide showed very high tensile strength.

Experimental Example 2 Drug Release Experiment

In order to observe the release of the drug over time in the tissue regeneration shielding membrane prepared by the addition of the drug was performed as follows.

The tissue regeneration shielding membrane prepared in Example 2 was placed in 0.1 M phosphate buffer (PBS, pH 7.4) and maintained at 37 ° C. and 15 rpm. The release liquid was taken at regular time intervals to measure the tetracycline released from the shielding membrane. The concentration of the released tetracycline was calculated by standard assay after measuring the absorbance at 274 nm using an ultraviolet absorbance photometer. The results are shown in FIG.

As shown in Figure 3, tetracycline initially released the drug quickly, it can be seen that the drug is continuously released for 8 weeks in a constant amount to maintain a constant drug dose.

As described above, the tissue regeneration shielding membrane according to the present invention has a certain strength, biocompatibility and biodegradability, and can be maintained as a sustained release system of the drug when prepared by adding the drug. In addition, the shielding membrane for tissue regeneration according to the present invention can adjust the thickness of the nanofibers when the thickness of the nanofibers and the non-woven fabric is formed to control the thickness of the shielding membrane, it is possible to control the pore size of the porous structure conditions for using the shielding membrane It can be changed depending on use. In addition, since the nanofibers constituting the tissue regeneration shielding membrane according to the present invention can be produced from a mixture of a natural polymer and a synthetic polymer at a time, it can be produced simply without a lamination process.

Claims (13)

1 or 2 natural polymers selected from the group consisting of chitosan, collagen and alginic acid, a homopolymer of lactic acid, a copolymer of lactic acid and glycoic acid, a homopolymer of glycoic acid, and a mixture thereof A shield membrane for tissue regeneration in which a nanofiber made from a mixture of synthetic polymers of a species is in the form of a nonwoven fabric. The method of claim 1, wherein the natural polymer and the synthetic polymer are mixed in a weight ratio of 4: 1 to 1: 4, the tissue regeneration shielding membrane. The method of claim 1, wherein the shielding membrane is a tissue regeneration shield, characterized in that it comprises a porous structure formed of nanofibers. The membrane for regenerating tissue of claim 3, wherein the pore size of the porous structure is 2 to 10 μm. The shielding film for tissue regeneration according to claim 1, wherein the shielding film has a thickness of 0.1 to 5 mm. The thickness of said nanofibers is 0.001-10 micrometers, The tissue regeneration shielding film of Claim 1 characterized by the above-mentioned. The shield membrane of claim 1, wherein the mixture of the natural polymer and the synthetic polymer further comprises a drug, a growth factor, or a ceramic. 8. The drug according to claim 7, wherein the drug is one selected from the group consisting of mefenamic acid, ibuprofen, flubiprofen, indomethacin, naproxen, metronidazole, tetrasiacline, minocycline, oxytetracycline, and mixtures thereof. A shielding film for tissue regeneration. The barrier membrane for tissue regeneration according to claim 7, wherein the growth factor is one selected from the group consisting of platelet-derived growth factors, insulin-like growth factors, epidermal growth factors, tumor growth factors, and mixtures thereof. 8. The barrier membrane for tissue regeneration according to claim 7, wherein the ceramic is apatite hydroxide, tricalcium phosphate or calcium metaphosphate. One or two natural polymers selected from the group consisting of chitosan, collagen and alginic acid and a homopolymer of lactic acid, a copolymer of lactic acid and glycoic acid, a homopolymer of glycoic acid and a mixture thereof Method of producing a tissue regeneration shielding film comprising electrospinning a mixed polymer solution containing a synthetic polymer of the nanofibers, and manufacturing the nanofibers in the form of a nonwoven fabric. 12. The method of claim 11, wherein the mixed polymer solution is a solution containing a solvent, 0.5 to 2% of chitosan in the solvent, 8 to 12% of the collagen polymer in the solvent, and 10 to 20% of the synthetic polymer in the solvent. A method for producing a tissue regeneration shielding membrane. 13. The tissue according to claim 12, wherein the solvent is one selected from the group consisting of 1,1,1,3,3,3-hexafluoropropyl alcohol, methylene chloride, acetone, chloroform and mixtures thereof. Method of manufacturing a shielding film for regeneration.

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