Biomatrix and manufacturing method therefor

Abstract

The present invention relates to a biomatrix and a manufacturing method therefor, the biomatrix comprising, as active ingredients, a fibrous acellular dermal matrix and a biocompatible material and having a layered cross-section and thus having enhanced physical properties.

Description

Biomatrix and method for manufacturing the same

[0001] The present invention relates to a biomatrix with enhanced physical properties and a method for manufacturing the same.

[0002] A wound is a condition in which the continuity of tissue is disrupted as skin or other tissues are severed or damaged by external pressure; typically, it refers to a phenomenon where the skin opens due to damage extending down to the dermis layer. Wound surgery fundamentally involves suturing and dressing the wound site to block exposure to the external environment, thereby preventing infection and suppressing inflammatory responses. Wound dressing materials can be broadly classified into allogeneic and xenogeneic dermis, and they can be manufactured using methods involving the extraction of specific polymers from the material or the utilization of the dermal matrix.

[0003] Acellular dermal matrix (ADM) refers to cadaveric skin tissue from which dermal cells have been removed to eliminate immune rejection, and there are various types depending on the origin. Human-derived tissues are widely used for soft tissue reconstruction and allografts for burn treatment; recently, in addition to the previously used bovine, porcine, and horse dermis, transplantation and wound treatment using fish dermis are also gaining attention. Dermal tissue is composed of 80–90% collagen, elastin, and glycosaminoglycans.

[0004] Meanwhile, in order to develop acellular dermal matrix for sheet-type therapeutic materials, which is in high demand among consumers, acellular dermal matrix is ​​being combined with gelatin or collagen. However, in the case of compositions in which acellular dermal matrix is ​​cross-linked with biocompatible polymers such as gelatin or collagen, although structural stability is high, the degree of cross-linking, viscoelasticity, hardening, and extrusion force are also high. While these are suitable for applications such as fillers and implants, they are still insufficient for application as sheet-type wound dressings. Furthermore, there were problems in that the cross-linking prevented easy dissolution or degradation, and the material failed to fix to the wound site, causing it to flow out and not be evenly distributed over the wound.

[0005] Therefore, there is an urgent need to develop a form of acellular dermal matrix that can be fixed to the wound site.

[0006] The objective of the present invention, derived to solve the problems described above, is to provide a biomatrix that is easy to apply to a wound site, has enhanced physical properties, and is soluble or degradable, as well as a method for manufacturing the same.

[0007] To achieve the above objective, a biomatrix according to one embodiment of the present invention comprises a fibrous acellular dermal matrix and a biocompatible material as active ingredients, wherein the content of the fibrous acellular dermal matrix satisfies 1 / 5 to 1 / 2 times the content of the biocompatible material and has a cross-section of a layered structure.

[0008] A method for manufacturing a biomatrix according to another embodiment of the present invention comprises: (a) a step of crushing an acellular dermal matrix to fibrose it; (b) a step of adding a biocompatible material to an acidic solvent, dispersing it, and then dividing it to prepare a biocompatible material mixture; (c) a step of mixing the acellular dermal matrix fibroseed in step (a) with the biocompatible material mixture prepared in step (b); (d) a step of drying and freezing the mixed mixture and then performing a first freeze-drying; (e) a step of immersing the first freeze-dried product in a cross-linking solution to cross-link it and then washing it; and (f) a step of freezing the cross-linked first freeze-dried product and then performing a second freeze-drying.

[0009] FIG. 1 is a process diagram for explaining a method for manufacturing a biomatrix in another embodiment of the present invention.

[0010] Figures 2 and 3 are SEM images showing the results of Experimental Example 1.

[0011] Figures 4 and 5 are graphs showing water absorption and tensile strength according to Experimental Example 2.

[0012] Figure 6 is a schematic diagram showing the method of Experimental Example 3.

[0013] Figure 7 is a photograph showing the results according to Experimental Example 3, and Figure 8 is a graph showing the results according to Experimental Example 3.

[0014] Figures 9 to 11 are photographs showing the results according to Experimental Example 4.

[0015] Hereinafter, the present invention will be described in detail with reference to the attached drawings and embodiments thereof. However, the following embodiments are presented as examples of the present invention, and if it is determined that a detailed description of a technology or configuration well known to those skilled in the art may unnecessarily obscure the essence of the present invention, such detailed description may be omitted, and the present invention is not limited thereby. The present invention is capable of various modifications and applications within the scope of the claims set forth below and the equivalent scope interpreted therefrom.

[0016] Furthermore, the terminology used in this specification is used to appropriately describe preferred embodiments of the present invention, and may vary depending on the intent of the user or operator, or the conventions of the field to which the present invention belongs. Accordingly, the definitions of these terms should be based on the content throughout this specification. Throughout the specification, when a part is described as “comprising” a certain component, unless specifically stated otherwise, this means that it does not exclude other components but may include additional components.

[0017] Throughout this specification, '%' used to indicate the concentration of a particular substance is %(w / w) for solid / solid, %(w / v) for solid / liquid, and %(v / v) for liquid / liquid, unless otherwise noted.

[0018] In this specification, the biomatrix may include a wound dressing composition, an implant composition, a skin substitute composition, and a drug delivery system composition, and may include, for example, a composition having a cross-section of a layered structure.

[0019] Hereinafter, a biomatrix according to one embodiment of the present invention will be described in detail.

[0020] The biomatrix of this embodiment may include a fibrotic acellular dermal matrix and a biocompatible material as active ingredients. For example, the fibrotic acellular dermal matrix has an open pore structure on its surface where fibers are interwoven in a net and a layered structure in its cross-section, thereby satisfying a water absorption capacity of 1500 to 3000% and a tensile strength of 0.5 to 1.2 MPa.

[0021] Fibrotic acellular dermal matrix (ADM) is a component widely used in plastic surgery and orthopedics for the reconstruction, regeneration, and reinforcement of skin, tendons, and ligaments due to its superior biocompatibility resulting from high cell adhesion ability and low immune response compared to existing animal-derived products. Fibrotic acellular dermal matrix may utilize materials derived from one or more sources selected from humans, fish by-products, fish skin, cattle, horses, and pigs; here, fish skin may refer to the skin of fish. For example, fibrotic acellular dermal matrix exhibits a coexistence of bundled forms resembling a ball of yarn and thin, long single fiber forms resembling threads; when used as a wound dressing, it demonstrates excellent in vivo preservation, maintains a moist environment well, and can exhibit a structure that is stably supported by biocompatible materials without the need for separate chemical bonding. The fibrotic acellular dermal matrix used in this embodiment may contain at least 90% of a long axis length of 100 to 3,000 μm, preferably 100 to 2,000 μm, and more preferably 50 to 90% of a length of 100 to 800 μm. This is because if a large portion is contained with a length of less than 100 μm, the shape may not be properly formed when attempting to process it into a sheet type, or it may be formed only into a film, making it difficult to flexibly adhere to the wound site. The content of this fibrotic acellular dermal matrix may satisfy 1 / 5 to 1 / 2 times the content of the biocompatible material.

[0022] For example, the fibrotic acellular dermal matrix may be included in an amount of 10 to 50 weight percent based on the total weight percent of the composition. If the fibrotic acellular dermal matrix is ​​less than 10 weight percent, the healing effect on the wound site is negligible, and the moisture absorption capacity is too high, making it difficult to manufacture into a sheet type. If it exceeds 50 weight percent, both the tensile strength and moisture absorption capacity may be below the values ​​required by the industry.

[0023] Biocompatible materials are major protein components of the skin and can be added to improve hemostatic properties, biocompatibility, cytotoxicity, low antigenicity, biodegradability, and cell adhesion. They can be extracted from various materials ranging from cattle, pigs, horses, and birds to marine tissues, and may be, for example, gelatin, hyaluronic acid, collagen, poloxamer, or mixtures thereof. These biocompatible materials may be included in an amount of 50 to 90 weight percent of the total weight of the composition. If the biocompatible material is less than 50 weight percent, the flowability increases, making it difficult to manufacture into a sheet type, and if it exceeds 90 weight percent, the viscosity increases, making it difficult to achieve desired physical properties and requiring a long time to dissolve.

[0024] In this embodiment, the term "fibrotic acellular dermal matrix" may refer to individual particles of granulated acellular dermal matrix that are not spherical or streamlined but are in the form of thin, long fibers like threads.

[0025] In this embodiment, stem cells, growth factors, or a mixture thereof may be further included.

[0026] In addition, the present embodiment may further include one or more selected from pharmaceutically used antimicrobial agents, excipients, and additives.

[0027] Antimicrobial agents include short-chain alcohols, benzoalkonium chloride (BAC), didecyl dimethyl ammonium chloride (DDAC), zeolite (CWT-A), isothiazolone, alkyl dimethyl ammonium chloride, triazine, 2-thiocyanomethylthiobenzothiazole, methylenebis thiocyanate, acrolein, dodecylguanidine hydrogen chloride, chlorophenol, quaternary ammonium salts, gluteraldehyde, dithiocarbamate, 2-mercaptobenzothiazole, para-chloro-meth-xylenol, silver, chlorhexidine, polyhexamethylene biguanide, n-halamine, triclosan, phospholipids, alpha-hydroxy acids, and 2,2-dibromo-3-nitrilopropionamide. One or more selected from 2-bromo-2-nitro-1,3-propanediol, farnesol, iodine, bromine, hydrogen peroxide, chlorine dioxide, vegetable oil, plant extract, benzalkonium chloride, chlorine, and sodium hypochlorite may be used.

[0028] One or more selected from stabilizers, antioxidants, osmotic pressure adjusters, buffers, and pH adjusters may be used as excipients, and specifically, one or more selected from starch, cellulose, glucose, lactose, sucrose, gelatin, corn, rice, wheat flour, chalk, silica gel, magnesium stearate, sodium stearate, glycerol monostearate, sodium chloride, glycerol, propylene glycol, water, and ethanol may be used.

[0029] As additives, one or more selected from physiologically biocompatible buffers (tromethamine hydrochloride), chelating agents (DTPA or DTPA-bisamide), and calcium chelating complexes (calcium DTPA, CaNaDTPA-bisamide) may be used, and optionally, calcium or sodium salts (calcium chloride, calcium ascorbate, calcium gluconate, or calcium lactate) may be used.

[0030] In this embodiment, the biomatrix may be in one form selected from sheet type, hydrogel type, and injectable type, and may be, for example, a sheet type.

[0031]

[0032] Hereinafter, a method for manufacturing a biomatrix according to another embodiment of the present invention will be described in detail with reference to the drawings. Since the composition of the biomatrix has been described in detail above, a description that is redundant therewith will be omitted.

[0033] FIG. 1 is a process diagram for explaining a method for manufacturing a biomatrix according to another embodiment of the present invention.

[0034] Referring to Fig. 1, first, the acellular dermal matrix is ​​crushed and fibrosed (S10).

[0035] A fibrotic acellular dermal matrix can be prepared by grinding the acellular dermal matrix to a size of 100 to 700 μm using one or more grinders selected from a cutting mill, food processor, agate grinder, freeze grinder, micronizer, vibratory micro mill, jaw crusher, mortar grinder, planetary mill, disk mill, ball mill, knife mill, and variable speed rotor mill. At this time, it is desirable to set the grinder to a rotational speed of 500 to 2000 rpm to facilitate grinding in a short time.

[0036] Meanwhile, in this embodiment, it is preferable to use a cell-free dermal matrix that has been decellularized and defat-treated. For example, the cell-free dermal matrix may be decellularized and defat-treated by the steps of preparing human-derived skin tissue, immersing the skin tissue in a hypotonic solution containing a surfactant, washing the immersed skin tissue first with an isotonic solution, and washing the first washed skin tissue second with a defat-removing solution.

[0037]

[0038] Next, a biocompatible material is added to an acidic solvent to disperse it, and then divided to prepare a biocompatible material mixture (S20).

[0039] A biocompatible material mixture in which the biocompatible material is dispersed can be prepared by adding 0.1 to 5 wt% of the biocompatible material to an acidic solvent, dispersing it for 1 to 35 minutes, and then sifting it using a sieve with a size of 0.5 to 2 mm. The acidic solvent may consist of purified water and hydrogen chloride (HCl). If the biocompatible material is less than 0.1 wt%, the flowability increases, making it difficult to manufacture in a sheet type, and if it exceeds 5 wt%, the viscosity increases, making it difficult to achieve the desired physical properties.

[0040] Then, a mixture of fibrotic acellular dermal matrix and biocompatible material is mixed (S30).

[0041] The fibrotic acellular dermal matrix from step S10 and the biocompatible material mixture prepared in step S20 can be mixed in a weight ratio of 0.1 to 50 : 50 to 99.9 and mixed for 1 to 5 minutes. If the weight ratio of the fibrotic acellular dermal matrix and the biocompatible material mixture is deviated from the above, not only is it difficult to manufacture it in the form of a film, but it also fails to satisfy the water absorption capacity and tensile strength values ​​required by the biomatrix industry, and it may be difficult to dissolve or decompose within 7 days, for example, within 4 days. In addition, it may not provide an environment suitable for cells to attach and grow, so the therapeutic effect may be minimal.

[0042] In this embodiment, 'solubility' or 'degradability' means that the biomatrix can dissolve or degrade when it comes into contact with moisture or the like.

[0043]

[0044] After that, the mixture is dried and frozen, then freeze-dried once (S40).

[0045] The mixture prepared in step S30 above can be poured into a mold and dried for 10 to 60 minutes, frozen at -40°C or lower for 1 to 2 hours, and then freeze-dried for 6 to 48 hours. If the drying time is 10 minutes or less, it may be difficult to maintain the shape to fit the mold, and if it exceeds 60 minutes, the effect due to the drying time being longer than necessary may not be very significant.

[0046]

[0047] Next, the primary freeze-dried product is immersed in a crosslinking solution to form crosslinks, and then washed (S50).

[0048] In the above S40 step, the primary freeze-dried material is completely immersed in a crosslinking solution and stirred at 50 to 200 rpm for 30 minutes to 4 hours to crosslink it, after which it can be washed with purified water at 50 to 200 rpm for 1 to 24 hours; for example, at the 10 to 15-hour washing point, it can be washed with phosphate-buffered saline (PBS). If the S50 step is not performed or if the crosslinking speed and time fall outside the aforementioned ranges, the final product may dissolve easily in water, making it difficult to act on the wound site. On the other hand, if the conditions for the aforementioned crosslinking speed and time are satisfied, the physical strength is excellent, so when the dressing is applied to the wound site, it can be fixed in place, maintain its shape, and remain there for a longer period, thereby improving the therapeutic effect.

[0049] In this embodiment, the crosslinking solution may consist of a solvent and a crosslinking agent, and the solvent may be one or more selected from purified water, ethanol, methanol, isopropanol, butanol, and ethyl acetate, for example, ethanol, and the crosslinking agent may be a substance soluble in the solvent, such as 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC), N-hydroxysuccinamide (NHS), or a mixture thereof, for example, a mixture of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide and N-hydroxysuccinamide.

[0050]

[0051] Finally, the cross-linked primary freeze-dried product is frozen and then secondary freeze-dried (S60).

[0052] The primary freeze-dried product cross-linked in the above S50 step can be frozen at minus 40°C or lower for 1 to 2 hours and then secondary freeze-dried for 6 to 48 hours.

[0053] Once the second freeze-drying is complete, the product can be cut to the required specifications, packaged in packaging materials, and then sterilized and stored at room temperature.

[0054]

[0055] The present invention will be described in more detail below using examples. These examples are solely for the purpose of explaining the present invention more specifically, and it is obvious to those skilled in the art that the scope of the present invention is not limited by them.

[0056]

[0057] Preparation Example 1. Preparation of Fibrotic Acellular Dermal Matrix

[0058] Skin tissue was purchased from EURO skin bank, Allosource, CTS, and tissues with a thickness of 0.5 mm or more were selected. Adipose tissue attached to the skin tissue was removed using forceps, and the tissue was washed three times with sterile water. Next, the tissue was immersed in a hypotonic solution (Tris-HCl, EDTA, NaOH, and SDS) and treated for 6 hours. Afterward, the skin tissue was washed with PBS at 4°C to remove residual fat, epidermis, cells, and hypotonic solution, and incubated overnight. Subsequently, it was washed with an isotonic solution (Tris-HCl, EDTA, NaCl, NaOH) for 6 hours to deepidermize, delipidize, and decellularize. Finally, the tissue was ground to a size of 100–700 µm using a cutting mill or grinding equipment to prepare a fibrotic acellular dermal matrix (hereinafter referred to as 'ADM').

[0059]

[0060] Preparation Example 2. Preparation of a biocompatible material mixture

[0061] An acidic solution was prepared by adding 2 ml of 34–37% HCl to 990 g of purified water in a beaker and mixing, and collagen was added to make the concentration 0.1–5%, then dispersed with a spatula for 1–5 minutes. Afterward, the mixture was dispersed using a stirring device for 1–10 minutes, then dispersed with a rotary mixer at 400–600 rpm and 400–600 rpm for 5–20 minutes, and filtered through a sieve with a size of 0.5–2 mm to prepare a biocompatible material mixture.

[0062]

[0063] Examples 1 to 5. Preparation of biomatrix

[0064] The fibrotic acellular dermal matrix prepared in Preparation Example 1 was dispersed in the biocompatible material mixture prepared in Preparation Example 2 at a ratio of 1 / 500 to 1 / 200 based on the weight of the biocompatible material mixture using a spatula for 1 to 5 minutes, then dispersed using a mixer or stirring device for 1 to 10 minutes, and then dispersed using a rotary mixer at 300 to 600 rpm for 5 to 30 minutes. Then, it was poured into a mold and dried for 10 to 60 minutes to maintain its shape, then frozen by storing in an ultra-low temperature freezer for 1 to 2 hours, and finally freeze-dried in a freeze-dryer for 6 to 48 hours. Next, the sample was immersed in a crosslinking solution containing 0.5–4 mg of EDC and 0.125–1 mg of NHS per 1 ml of ethanol, and crosslinked by stirring with a shaker at 50–200 rpm for 0.5–4 hours. Afterward, the sample was cut larger than the standard size and washed with purified water (using PBS as a washing solution in between) in a beaker at 50–200 rpm for 1–24 hours. Then, it was frozen by storing in an ultra-low temperature freezer for 1–2 hours, followed by secondary freeze-drying for 6–48 hours. The sample was cut to the standard size to produce a biomatrix satisfying the ADM content listed in Table 1 below.

[0065] Classification Example 1 Example 2 Example 3 Example 4 Example 5 ADM Content (%) 10 20 30 40 50

[0066]

[0067] Comparative Example 1. Biomatrix manufacturing

[0068] It was prepared in the same manner as Example 2, except that it did not contain fibrotic acellular dermal matrix.

[0069]

[0070] Comparative Example 2. Biomatrix Manufacturing

[0071] After preparing the fibrotic acellular dermal matrix, it was placed on a sieve, and the sieve was placed in a tray filled with water to spread the fibrotic acellular dermal matrix widely in the form of a sheet. This process was repeated 2 to 4 times, after which the sieve was removed and left at room temperature for 1 to 4 hours. Then, it was frozen in an ultra-low temperature freezer for 1 to 2 hours, followed by freeze-drying for 12 to 48 hours to produce a biomatrix.

[0072]

[0073] Comparative Example 3. Biomatrix Manufacturing

[0074] It was prepared in the same manner as Example 2, except that fibrotic acellular dermal matrix was not used and elastin was additionally used.

[0075]

[0076] Experimental Example 1. Confirmation of the structure and morphology of the biomatrix

[0077] SEM images of the biomatrix of Examples 1 to 5 and Comparative Examples 1 to 3 were taken to confirm the structure and morphology of the biomatrix according to the content of ADM, and the results are shown in Figures 2 and 3.

[0078] Referring to FIGS. 2 and 3, it was confirmed that compared to Comparative Example 1 and Comparative Examples 2 and 3, which did not contain fibrotic acellular dermal matrix, Examples 1 to 5 had a layered structure in cross-section and an open pore structure in which fibers were intertwined in a net on the upper surface. In particular, Example 2, which contained 20% ADM, showed the densest layered structure in cross-section.

[0079]

[0080] Experimental Example 2. Evaluation of Physical Properties

[0081] To confirm the physical properties of the biomatrix according to the content of ADM, water absorption capacity and tensile strength were measured, and the results are shown in Figures 4 and 5 and Table 2.

[0082] Referring to Figure 4 and Table 2, it can be seen that the water absorption capacity gradually decreases from Comparative Example 1 to Example 5. In the case where ADM is not included, the water absorption capacity was highest at over 3000%, and it was found that the water absorption capacity decreased as the content of ADM increased.

[0083] Referring to Figure 5 and Table 2, it was confirmed that the tensile strength of Comparative Example 1 was relatively lower compared to Examples 1 to 5, and that the tensile strength decreased as we moved from Example 2 to Example 1, Example 3, Example 4, and Example 5. In other words, it was observed that the tensile strength increased from Comparative Example 1 to Example 2, reaching its highest value of 0.10 MPa or higher in Example 2, and then decreased as we moved toward Example 5.

[0084] As such, it can be seen that the biomatrix manufactured according to the embodiment of the present invention has excellent tensile strength and can be applied in a desired shape even to curved wound areas.

[0085] Classification Comparison Example 1 Example 1 Example 2 Example 3 Example 4 Example 5 Moisture absorption capacity (%) 30 27 27 31 260 72 257 17 89 15 77 Tensile strength (MPa) 0.5 81.0 51.1 40.8 40.7 20.60

[0086]

[0087] Experimental Example 3. Evaluation of therapeutic effect on the wound site

[0088] To confirm the therapeutic effect on the wound site, four wound model rats with 8 mm circular skin wounds on their backs were prepared, and as shown in Fig. 6, biomatrixes according to Example 2, Comparative Examples 2 and 3 were injected into each wound site and fixed with gauze. Then, each was weighed and skin wound photographs were taken for 14 days, and the change in wound size was measured using the Image J program, and the results are shown in Figs. 7 and 8 and Table 3.

[0089] Referring to Figures 7 and 8 and Table 3, it was confirmed that Example 2 exhibited similar decomposition and recovery processes to Comparative Examples 2 and 3 during the wound healing process; in particular, after 14 days, the average wound size of Example 1 was 0.7–0.8 mm 2 Comparative Example 2 (1.1mm 2 ), Comparative Example 3 (1.3mm 2 Approximately 0.4~0.5mm compared to ) 2 I could see that it was small.

[0090] As such, it can be seen that the biomatrix manufactured according to the embodiment of the present invention completely decomposed within 4 days, and that the wound healed safely without problems such as inflammation occurring during the wound healing process.

[0091] Classification Day 0 Day 1 Day 4 Day 7 Day 10 Day 14 Example 2 #150.347.438.19.03.50.3 #250.342.228.511.62.01.4 #350.346.337.34.30.80.1 #450.348.239.515.32.71.2 Comparative Example 2 #150.346.531.114.67.91.2 #250.343.029.94.72.40.6 #350.343.434.68.66.21.5 Comparative Example 3 #450.345.131.08.92.91.3

[0092] Experimental Example 4. Evaluation of the therapeutic effect on wound tissue

[0093] To confirm the therapeutic effect on wound tissue, wound tissue from a 14-day-old rat of Experimental Example 3 was excised and fixed in a 10% neutral formalin solution for 24 hours. Afterward, the damaged tissue was taken, dehydrated, and embedded in paraffin. The tissue was sectioned to a thickness of 5 µm using a tissue sectioner, attached to a slide, and subjected to paraffin removal and hydration. Subsequently, hematoxyllin-eosin (H&E) staining and Masson's trichrome (MT) staining were performed.

[0094] For H&E staining, tissue sections were paraffin-removed with xylene, hydrated with 100%, 90%, 80%, and 70% ethanol and distilled water for 5 minutes each, and then washed with distilled water before use. The tissues were stained with Harris hematoxylin for 3 minutes and washed with distilled water for 5 minutes. After washing, the tissues were stained with eosin for 5 minutes, dehydrated with 70%, 80%, 90%, and 100% ethanol and xylene, and then mounted in Shandon Synthetic Mountant (Thermo scientific, USA). For MT staining, tissue sections were paraffin-removed with xylene, hydrated with 100%, 90%, 80%, and 70% ethanol and distilled water for 5 minutes each, and then washed with distilled water before use. The tissues were reacted in 60°C Bouin's (IMEB, USA) solution for 1 hour and then washed with distilled water. After the reaction and washing were completed, the tissues were treated again with Biebrich scarlet-acid fuchsin, phosphomolybdic-phosphotungstic acid, and aniline blue stain solution (IMEB, USA) for 5 minutes each, followed by washing with distilled water. After dehydration using 70, 80, 90, and 100% ethanol and xylene, the tissues were mounted in Shandon Synthetic Mountant (Thermo Scientific, USA). The results of H&E and MT staining are shown in Figures 8 to 10.

[0095] Referring to FIGS. 9 and 11, it was confirmed that in Example 2, Comparative Example 2, and Comparative Example 3, angiogenesis occurred at the wound site and cell influx occurred into the formed tissue; in particular, it was observed that Example 2 had a larger number of formed blood vessels compared to Comparative Example 2. Referring to FIGS. 10 and 11, it was confirmed that in Example 2, Comparative Example 2, and Comparative Example 3, cell influx occurred as biocompatible material fibers were formed at the wound site. However, in the case of Comparative Example 3, an area suspected to be inflamed was identified.

[0096] As such, it can be seen that the biomatrix manufactured according to the embodiment of the present invention heals the wound by forming blood vessels and biocompatible material fibers at the wound site without inflammation, thereby facilitating cell influx.

[0097]

[0098] Although exemplary embodiments of the present invention have been described in detail above, the scope of the present invention is not limited thereto, and various modifications and improvements by those skilled in the art using the basic concept of the present invention as defined in the following claims also fall within the scope of the present invention.

[0099] All technical terms used in this invention, unless otherwise defined, are used in the sense generally understood by those skilled in the art in the relevant field of this invention. The contents of all publications cited as references in this specification are incorporated into this invention.

Claims

1. Includes fibrotic acellular dermal matrix and biocompatible material, The weight of the fibrotic acellular dermal matrix satisfies 1 / 5 to 1 / 2 times the weight of the biocompatible material, and A biomatrix having a cross-section of a layered structure.

2. In Paragraph 1, The above biomatrix is ​​a biomatrix having a surface with an open pore structure in which fibers are interwoven in a net.

3. In Paragraph 1, 10 to 50 weight% of the above fibrotic acellular dermal matrix; and A biomatrix comprising 50 to 90 weight percent of the above-mentioned biocompatible material.

4. In Paragraph 1, The above-mentioned fibrotic acellular dermal matrix comprises a biomatrix comprising a material derived from one or more selected from the human body, fish byproducts, fish skin, cattle, and pigs.

5. In Paragraph 1, The above biomatrix is ​​a biomatrix having a water absorption capacity of 1500~3000%.

6. In Paragraph 1, The above biomatrix is ​​a biomatrix having a tensile strength of 0.5 to 1.2 MPa.

7. In Paragraph 1, The above biomatrix is ​​a sheet-type biomatrix. 8.(a) A step of pulverizing and fibrosing the acellular dermal matrix; (b) A step of preparing a biocompatible material mixture by adding and dispersing a biocompatible material in an acidic solvent and then dispersing it; (c) a step of mixing the fibrotic acellular dermal matrix from step (a) above with the biocompatible material mixture prepared in step (b) above; (d) A step of drying and freezing the above-mentioned mixed mixture, followed by primary freeze-drying; (e) a step of immersing the above primary freeze-dried product in a crosslinking solution to crosslink it, and then washing it; and (f) a step of freezing the cross-linked primary freeze-dried product and then performing secondary freeze-drying; comprising a method for manufacturing a biomatrix.

9. In Paragraph 8, A method for manufacturing a biomatrix in which, in step (a) above, the acellular dermal matrix is ​​decellularized and defat-treated, and the grinding is performed to a size of 100 to 700 µm.

10. In Paragraph 8, A method for manufacturing a biomatrix, wherein in step (d) above, drying is performed for 10 to 60 minutes, freezing is performed for 1 to 2 hours, and primary freeze-drying is performed for 6 to 48 hours.

11. In Paragraph 8, A method for manufacturing a biomatrix, wherein in step (e) above, the crosslinking solution comprises a solvent and a crosslinking agent, wherein the solvent is one or more selected from purified water, ethanol, methanol, isopropanol, butanol, and ethyl acetate, and the crosslinking agent is 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide, hydroxysuccinamide, or a mixture thereof.

12. In Paragraph 8, A method for manufacturing a biomatrix, wherein in step (f) above, freezing is performed for 1 to 2 hours and secondary freeze-drying is performed for 6 to 48 hours.

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