Laminated structure and method for producing the same

Abstract

To provide a novel laminated structure in which a non-dielectric substrate and a sheet-form fiber layer are integrally laminated.SOLUTION: A voltage is applied to the back side of a non-dielectric substrate to polarize the non-dielectric substrate, thereby charging the surface of the non-dielectric substrate with a first charge. While a voltage is applied to a fiber-forming material solution, fibers bearing a second charge are ejected by electrospinning with the fiber-forming material solution directed toward the surface of the non-dielectric substrate. Through electrostatic attraction, a laminated structure is produced where the non-dielectric substrate and a sheet-form fiber layer are integrally laminated.SELECTED DRAWING: Figure 2

Description

[Technical Field]

[0001] The present invention relates to a laminated structure that can be used as a novel medical material, a substance adsorption film, etc., and a method for producing the same. [Background technology]

[0002] Electrospinning is a well-known method for producing nanofibers. Electrospinning involves filling a syringe with a raw material solution, applying a voltage between a nozzle and a site where the nanofibers are deposited and collected (also known as a collector), drawing the raw material solution through the nozzle to form nanofibers, and producing the nanofibers on the collector (Patent Documents 1, 2, and 3).

[0003] In the field of regenerative medicine, a method has been implemented in which tissue defects are covered with wound treatment dressings made by incorporating tissue formation factors into a matrix resin of biocompatible polymers such as polylactic acid or polylactic acid-glycolic acid copolymers. Biocompatible polymers are widely used as tissue regeneration materials, and electrospinning is used as a method for producing fibers of such biocompatible polymers (Non-Patent Document 1).

[0004] Nanofibers spun by electrospinning are expected to be promising for engineering materials such as various tissue regeneration scaffolds, wound dressings, and anti-adhesion membranes.

[0005] The structure of nanofibers spun by electrospinning resembles the collagen structure of the extracellular matrix (ECM) of biological tissues (a three-dimensional network of collagen nanofibers with diameters of 50 to 500 nm). Furthermore, nanofibers spun by electrospinning possess useful properties for tissue regeneration, such as precise shape characteristics (three-dimensional porosity, nanoscale size, and orientation), encapsulation and localized slow release of growth factors, and surface functionalization (introduction of functional groups, etc.) (Patent Document 4).

[0006] Electrospinning is also used to produce gelatin fibers. For example, a mixed solution of hexafluoroisopropanol and water is an excellent solvent for producing gelatin fibers by electrospinning, and it has been reported that when gelatin is dissolved in this mixed solution and electrospun, gelatin fibers with a uniform fiber diameter on the order of μm can be obtained (Patent Document 5).

[0007] Specifically, the syringe with a nozzle is filled with gelatin solution, the raw material solution. The collector is grounded. When the high-voltage power supply is turned on, high voltage is applied to the nozzle. This high voltage induces and accumulates an electric charge in the gelatin solution flowing through the nozzle. After being sprayed from the nozzle, the gelatin solution particles become positively charged and repel each other. This repulsive force counters the surface tension of the gelatin solution, and when the charge threshold is exceeded, the gelatin solution becomes a charged mist. Because the surface area of ​​this charged mist is very large relative to its volume, the solvent evaporates efficiently. Furthermore, the decrease in volume increases the charge density, causing the gelatin solution to break up into charged micro-mists. Because a high voltage is applied to the nozzle and the collector is grounded, a strong electric field is formed between the nozzle and the collector. The charged micro-mists repel each other and move toward the collector due to the formed electric field. Along the way, the solvent evaporates and is collected on the collector as gelatin fibers.

[0008] In this way, the collector is merely used as a component for collecting the fibers, and the idea of ​​using the collector in the electrospinning method as an integral part of the fibers has not existed in the past. [Prior art documents] [Patent documents]

[0009] [Patent Document 1] Special Publication No. 2013-503661 [Patent Document 2] Special Publication No. 2013-519805 [Patent Document 3] Japanese Patent Application Laid-Open No. 2012-72514 [Patent Document 4] Patent No. 5855783 [Patent Document 5] Japanese Patent Application Laid-Open No. 2007-138364 [Non-patent literature]

[0010] [Non-Patent Document 1] In vivo and in vitro evaluation of flexible, cottonwool-like nanocomposite as bone substitute material for complex defects Acta Biomaterialia 5 2009 Summary of the Invention [Problem to be solved by the invention]

[0011] An object of the present invention is to provide a novel laminated structure using an electrospinning method. [Means for solving the problem]

[0012] The laminated structure according to the present invention comprises: Dielectrics containing non-metallic or polymeric materials The conductive substrate and the sheet-like fiber layer are laminated together. [Effects of the Invention]

[0013] According to the present invention, Temptation A novel laminated structure is obtained in which the conductor substrate and the sheet-like fiber layer are integrally laminated. [Brief explanation of the drawings]

[0014] [Figure 1] 1 is a photograph of a specific example of a laminated structure of the present invention. [Figure 2] 1 is a diagram showing an outline of a manufacturing apparatus for a laminated structure of the present invention. [Figure 3] FIG. 1 is a diagram showing an outline of a manufacturing apparatus for a laminated structure of the present invention to which an ionizer is attached. [Figure 4] Photographs of laminate structures, where (A) is a comparative example, (B) is a laminate structure according to the present invention produced without using an ionizer, and (C) is a laminate structure according to the present invention produced with an ionizer. [Figure 5] 1A and 1B are SEM images of gelatin fibers, where (A) is a comparative example, (B) is example 1, and (C) is example 3. [Figure 6] This figure shows the degree of neutralization of the amount of positive charge in the sheet-like fiber layer when the ionizer irradiation time interval is changed, where (A) is every 1 min of spinning, (B) is every 2 min of spinning, and (C) is every 3 min of spinning. DETAILED DESCRIPTION OF THE INVENTION

[0015] Hereinafter, embodiments of the present invention will be described in detail with reference to the accompanying drawings. However, these embodiments are intended to facilitate understanding of the principles of the present invention, and the scope of the present invention is not limited to the following embodiments. Other embodiments in which a person skilled in the art appropriately replaces the configuration of the following embodiments are also included in the scope of the present invention.

[0016] As shown in FIG. 1, the laminated structure according to the present invention comprises: Temptation The conductor substrate and the sheet-like fiber layer are laminated together. Note that in the present invention, "laminated" does not only mean that one sheet-like fiber layer is laminated, but also includes that multiple layers are laminated.

[0017] The polymeric material of the fibers forming the sheet-like fiber layer is not particularly limited and may be, for example, collagen, gelatin, polylactic acid, polyglycolic acid, lactic-glycolic acid copolymer, polycaprolactone, polyglycerol sebacic acid, polyhydroxyalkanoic acid, polybutylene succinate, polystyrene, polycarbonate, polyacrylic acid, polymethyl acrylate, polyvinyl chloride, polyethylene terephthalate, cellulose, polyamide, polyurethane, or polyvinyl alcohol. Preferably, the polymeric material is a biocompatible polymeric material, more preferably gelatin.

[0018] The sheet-like fiber layer is a layered structure in which nanofibers or microfibers are formed into a sheet shape.

[0019] In the present invention, the polarized Temptation Since the sheet-like fiber layer is laminated integrally with the electric body substrate, the density of the sheet-like fiber layer is high. In other words, the fibers are densely packed together to form a sheet, and the sheet Temptation It has good adhesion to the electrical substrate and is integrated without the use of adhesives (i.e., without an adhesive layer).

[0020] The fibers constituting the sheet-like fiber layer preferably have an average fiber diameter of 1 nm to 400 μm, more preferably 10 nm to 200 μm. If the average fiber diameter of the fibers is within this range, for example, when the laminated structure is used as a medical material, cells can easily invade through the sheet-like fiber layer.

[0021] The thickness of the sheet-like fiber layer is not particularly limited, but is, for example, 1 μm to 1 mm, and preferably 5 μm to 500 μm.

[0022] Temptation The conductor substrate is not particularly limited, and various materials such as non-metallic materials and polymeric materials can be used. TemptationThe material of the conductor substrate may be the same as or different from the material of the sheet-like fiber layer.

[0023] Temptation The conductive substrate may be, for example, a porous body. The porous body is not particularly limited, and may be, for example, a gelatin sponge.

[0024] One specific example of the laminated structure according to the present invention is a laminated structure in which a gelatin sponge and a sheet-like gelatin nonwoven fabric layer are laminated together, and is shown in FIG.

[0025] The gelatin sponge has a porosity of, for example, 30 to 85% and a pore diameter of 30 to 600 μm.In this gelatin sponge, the pores are interconnected.

[0026] The method for producing a gelatin sponge is not particularly limited. For example, first, gelatin is weighed and transferred to a container, and purified water is added to form a gelatin solution. Next, the gelatin solution is transferred to a tube and left in a refrigerator to gel the gelatin. The gelled gelatin solution is freeze-dried, and after drying is complete, the gelatin sponge is removed from the tube and thermally crosslinked. This completes the production of a gelatin sponge.

[0027] The laminated structure according to the present invention can be produced by electrospinning as described below.

[0028] As mentioned above, the collector is merely a target for depositing fibers, and there was no idea of ​​using the collector of the electrospinning method as a medical material. However, the substrate used for medical materials is Temptation Therefore, it is difficult to use the substrate and the fiber as a single unit by simply applying the electrospinning method.

[0029] Therefore, as a result of intensive research, the present inventors have found that TemptationBy polarizing the conductive substrate and applying it to the collector in the electrospinning method, Temptation The present invention was completed based on the new finding that a laminated structure can be obtained in which a conductor substrate and a sheet-like fiber layer are integrally laminated. Temptation By applying a voltage to the backside of the dielectric substrate, Temptation The dielectric substrate is polarized to Temptation The surface of the dielectric substrate is charged with a first charge, and the fiber-forming material solution is immersed in the above-mentioned solution while a voltage is applied to the fiber-forming material solution. Temptation The fibers charged with the second charge are ejected by electrospinning toward the surface of the electric substrate, and the above-mentioned Temptation A laminated structure is produced in which the conductor substrate and the sheet-like fiber layer are laminated together.

[0030] Temptation The polarization means for polarizing the conductor substrate is not particularly limited, and examples thereof include: Temptation Anything can be used as long as a charge polarization electric field is applied between the rear surface and the front surface of the electric substrate and charge polarization occurs under the action of this charge polarization electric field.

[0031] The manufacturing apparatus for a laminated structure according to this embodiment is outlined in Fig. 2. Specifically, the apparatus includes a syringe containing a fiber-forming material solution, a syringe pump for pushing out the solution of raw materials and a solvent in the syringe, a solution supply unit for supplying the fiber-forming material solution, a nozzle having a spinneret, and a spinning nozzle having a first charge on the front surface thereof due to polarization caused by application of a voltage to the rear surface thereof. Temptation A dielectric substrate and a nozzle Temptation and a high-voltage power supply that applies a high voltage between the conductive substrate and the substrate. Note that not all of the components are shown in Fig. 2 in order to make the present invention easier to understand.

[0032] In the present invention, Temptation When the amount of the second electric charge in the sheet-like fiber layer laminated on the surface of the electric substrate becomes high, it is preferable to neutralize the amount of the second electric charge by an ionizer. TemptationAs fibers continue to accumulate on the surface of the dielectric substrate, repulsive forces (forces that repel each other) occur between the fibers, but by suppressing (reducing) these repulsive forces through static elimination, it is possible to keep the thickness and pore size small. In other words, neutralization with an ionizer makes it possible to increase the thickness of a high-density sheet-like fiber layer.

[0033] In such a case, as shown in FIG. Temptation Irradiating the surface of the conductive substrate with neutralizing ions, Temptation An ionizer is provided as a static eliminator that neutralizes the second amount of charge in the sheet-like fiber layer laminated on the surface of the electric substrate.

[0034] When the spinneret is charged with a positive high voltage, a negative ion generator is used to emit negative ions. Conversely, when the spinneret is charged with a negative high voltage, a positive ion generator is used to emit positive ions.

[0035] Nozzle and Temptation When no high voltage is applied between the nozzle and the conductive substrate, the fiber-forming material solution remains at the tip of the spinneret at the tip of the nozzle due to surface tension.

[0036] Spinneret and Temptation When a high voltage of, for example, several kV to 30 kV is applied between the spinneret and the substrate, the fiber-forming material solution at the tip of the spinneret is positively charged, and the solution is charged to the opposite polarity (or earth potential). Temptation It is attracted by the surface of the dielectric substrate.

[0037] As mentioned above, Temptation When positively charged fibers are deposited on the surface of the dielectric substrate, the positive charge does not disappear completely and remains. TemptationFibers deposited on the conductive substrate repel each other with newly deposited fibers. In such cases, an ionizer, which is a static electricity removal device, is installed, and negative ions are irradiated to neutralize the positive charge of the fibers, preventing repulsion between charges of the same polarity, resulting in a thick sheet-like fiber layer. In addition, as an effect of the ionizer, when charges accumulate during spinning, the ionizer can be used to remove the charges, Temptation The polarization state of the dielectric substrate is reset to return to the initial spinning state, allowing for continuous spinning.

[0038] The use of the laminate structure according to the present invention is not particularly limited, and it can be used, for example, as a medical material, specifically, a wound treatment dressing, a scaffold for regenerative medicine, etc. Furthermore, the use of the laminate structure according to the present invention is not limited to medical materials, and it can also be applied, for example, to substance adsorption membranes, filters, battery separators, etc.

[0039] The laminated structure in which the gelatin sponge and the sheet-like gelatin nonwoven fabric layer are integrally laminated is suitably used, for example, as a scaffold for tissue regeneration, for wound healing, for scar treatment, or for preventing scar and keloid formation. When used for wound treatment, it is applied to the injured part of the body.

[0040] When applied, for example, the sheet-shaped gelatin nonwoven fabric layer is applied toward the injured area. This allows the sheet-shaped gelatin nonwoven fabric layer to come into contact with the injured area, and the fibers that make up the nonwoven fabric promote the adhesion, migration, and proliferation of cells supplied by the body. The injured area is moistened by moistening fluids such as blood flow and body fluids. When applied to such an injured area, the gelatin sponge becomes rich, promoting healing of the injured area. [Example]

[0041] 《 Temptation Manufacturing of electrical substrates The gelatin sponge was prepared as follows. First, gelatin was weighed and transferred to a container, and purified water was added to prepare a 2 w / v% gelatin solution. The gelatin solution was then transferred to a tube and left in a refrigerator at 4°C for 1 hour to gel the gelatin. The gelled gelatin solution was frozen at -25°C for 24 hours and then freeze-dried. After drying, the gelatin sponge was removed from the tube and thermally crosslinked at 140°C for 48 hours. This produced the gelatin sponge.

[0042] <<Manufacturing of laminated structures>> As shown generally in Figure 3, the laminated structure manufacturing apparatus of this example was configured to include a syringe containing a 7 w / v % gelatin solution, a syringe pump that extrudes the gelatin solution, a solution supply unit that supplies the gelatin solution, a nozzle with a spinneret, a gelatin sponge whose surface is negatively charged by polarization caused by application of a positive voltage to the back side, a high-voltage power supply that applies a high voltage between the nozzle and the gelatin sponge, and an ionizer, which is an electrostatic removal device that irradiates the surface of the gelatin sponge with discharging ions to neutralize the positive charge of the sheet-like fiber layer laminated on the surface of the gelatin sponge.

[0043] The solvent used for the gelatin solution was hexafluoroisopropanol. The voltage applied between the spinneret and the gelatin sponge was 6-7 kV. The flow rate of the gelatin solution extruded from the syringe pump was 20 μL / min. The distance between the spinneret and the gelatin sponge was 80 mm.

[0044] In Example 1, a laminated structure was fabricated without operating the ionizer. In Example 2, a laminated structure was fabricated with the ionizer operating. The ionizer irradiation time interval was set at 1 min for each spinning period, and the ionizer irradiation time was fixed at 10 s. In Example 3, a sheet-like nonwoven fabric layer was fabricated using a PLLA solution instead of a gelatin solution. Specifically, PLLA was dissolved in a solution containing 1,3-dioxolane (DOL) and HFIP (DOL:HFIP in a wt% ratio of 70:30) to prepare an 8 wt% PLLA solution. To further refine the fibers, 5 wt% benzyltriethylammonium chloride was added to the solution. This solution was placed in a syringe, and a laminated structure was fabricated using the above-mentioned apparatus under the same conditions as in Example 2. This resulted in the fabrication of a sheet-like PLLA nonwoven fabric layer with a thickness of 170 ± 29 nm on a gelatin sponge. As a comparative example, a laminate was formed under the same conditions as in Example 1, except that the gelatin sponge was not polarized.

[0045] Each of the laminated structures obtained by the fiber production methods of Example 1, Example 2, and Comparative Example was cut in half at the center to prepare specimens for electron microscopy. The cross section of each specimen for electron microscopy was then observed and photographed using a scanning electron microscope (JEOL Ltd. JSM-6309LT model, hereinafter referred to as "SEM").

[0046] In the laminated structure of the comparative example, as shown in FIG. 4(A), the gelatin fibers laminated on the gelatin sponge had large fiber intervals and were deposited in a cotton-like state rather than in a sheet-like state.

[0047] 4(B), the laminated structure of Example 1 had a gelatin sponge and a sheet-like gelatin nonwoven fabric layer laminated together. The thickness of the sheet-like gelatin nonwoven fabric layer was 10 μm.

[0048] In the laminated structure of Example 2, as shown in Fig. 4(C), a sheet-like gelatin nonwoven fabric layer was laminated integrally with a gelatin sponge. In Example 2, the thickness of the sheet-like gelatin nonwoven fabric layer was greater than that in Example 1, reaching 120 µm.

[0049] For each of the laminated structures obtained by the fiber production methods of Example 1, Example 3, and Comparative Example, the morphology of the center of the surface of each electron microscope specimen was observed and photographed using the aforementioned scanning electron microscope. Figure 5 shows SEM images of the gelatin fiber production method of Comparative Example, the gelatin fiber produced by the gelatin fiber production method of Example 1, and the PLLA fiber produced by the production method of Example 3. The bar at the bottom right of each SEM image in Figure 5 is a scale bar. The fiber diameter was also measured by image analysis using image analysis software (Furinx Sigma Scan Pro, Inc.). The fiber diameter of Example 3 was 170 ± 29 nm. The fiber diameter of both the Comparative Example and Example 1 was 465 ± 75 nm. As shown in Figure 5(A), the gelatin fiber of the Comparative Example had many gaps and a low density, while the gelatin fiber and PLLA fiber of the Examples had fewer gaps and a high density, as shown in Figures 5(B) and 5(C).

[0050] <<Changing the ionizer irradiation time interval>> As mentioned above, the flow rate of the gelatin solution extruded from the syringe pump was 20 μL / min. The ionizer irradiation time was fixed at 10 seconds, but experiments were conducted in which the interval between ionizer irradiation times was changed.

[0051] That is, the interval of irradiation time of the ionizer was changed to every 1 minute of spinning, every 2 minutes of spinning, and every 3 minutes of spinning.

[0052] In Figure 6, the horizontal axis is time (minutes) and the vertical axis is Temptation The voltage (kV) of the fiber deposited on the surface of the dielectric substrate. As shown in Figure 6, when the ionizer irradiation time is fixed, the shorter the interval between ionizer irradiation times, the better. Temptation This is suitable for neutralizing the amount of positive charges in the sheet-like fiber layer laminated on the surface of the electric body substrate, and as a result, the thickness of the sheet-like fiber layer can be further increased.

[0053] In addition, we fixed the ionizer irradiation time at 60 seconds and conducted an experiment in which the interval between ionizer irradiation times was changed, but it was found that a shorter interval between ionizer irradiation times was better. Temptation This is suitable for neutralizing the amount of positive charges in the sheet-like fiber layer laminated on the surface of the electric body substrate, and as a result, the thickness of the sheet-like fiber layer can be further increased. [Industrial Applicability]

[0054] It can be used in regenerative medicine.

Claims

1. A laminated structure in which a dielectric substrate containing a non-metallic material or a polymeric material and a sheet-like fiber layer are integrally laminated.

2. 2. The laminated structure according to claim 1, wherein the polymeric material of the fibers forming the sheet-like fiber layer is collagen, gelatin, polylactic acid, polyglycolic acid, lactic-glycolic acid copolymer, polycaprolactone, polyglycerol sebacic acid, polyhydroxyalkanoic acid, polybutylene succinate, polystyrene, polycarbonate, polyacrylic acid, polymethyl acrylate, polyvinyl chloride, polyethylene terephthalate, cellulose, polyamide, polyurethane, or polyvinyl alcohol.

3. 3. The laminated structure according to claim 1, wherein the dielectric substrate is a porous body.

4. A laminated structure in which a gelatin sponge and a sheet-like gelatin nonwoven fabric layer are laminated together.

5. 5. The laminated structure according to claim 4, which is used as a scaffold for tissue regeneration, for wound healing, for scar treatment, or for preventing scar and keloid formation.

6. A method for polarizing a dielectric substrate comprising a non-metallic material or a polymeric material by applying a voltage to a rear surface of the dielectric substrate, thereby charging a surface of the dielectric substrate with a first charge; While a voltage is applied to the fiber-forming material solution, the fiber-forming material solution is ejected toward the surface of the dielectric substrate by electrospinning to form fibers charged with a second electric charge; A method for manufacturing a laminated structure, in which a laminated structure is manufactured in which the dielectric substrate and the sheet-like fiber layer are integrally laminated by electrical attraction.

7. 7. The method for manufacturing a laminated structure according to claim 6, wherein the first charge is a negative charge and the second charge is a positive charge.

8. The method for manufacturing a laminated structure according to claim 6 or 7, characterized in that when the second charge amount of the sheet-like fiber layer laminated on the surface of the dielectric substrate becomes high, the charged second charge amount is neutralized.

9. A method for polarizing a gelatin sponge by applying a voltage to the back surface of the gelatin sponge as a dielectric substrate containing a non-metallic material or a polymeric material, thereby negatively charging the surface of the gelatin sponge; A voltage is applied to the gelatin fiber solution to positively charge the gelatin fibers, and the gelatin fiber solution is ejected toward the surface of the gelatin sponge by electrospinning; A method for producing a laminated structure in which the gelatin sponge and a sheet-like gelatin nonwoven fabric layer are integrally laminated by electrical attraction.

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