WO2013036082A2 - Composé polymère auquel est liée de la tris(2-carboxyéthyl)phosphine - Google Patents

Composé polymère auquel est liée de la tris(2-carboxyéthyl)phosphine Download PDF

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WO2013036082A2
WO2013036082A2 PCT/KR2012/007242 KR2012007242W WO2013036082A2 WO 2013036082 A2 WO2013036082 A2 WO 2013036082A2 KR 2012007242 W KR2012007242 W KR 2012007242W WO 2013036082 A2 WO2013036082 A2 WO 2013036082A2
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polymer compound
tcep
hydrogel
chondroitin sulfate
group
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WO2013036082A3 (fr
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노인섭
조성연
김수미
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서울과학기술대학교 산학협력단
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    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08L—COMPOSITIONS OF MACROMOLECULAR COMPOUNDS
    • C08L101/00—Compositions of unspecified macromolecular compounds
    • C08L101/02—Compositions of unspecified macromolecular compounds characterised by the presence of specified groups, e.g. terminal or pendant functional groups
    • C08L101/06—Compositions of unspecified macromolecular compounds characterised by the presence of specified groups, e.g. terminal or pendant functional groups containing oxygen atoms
    • C08L101/08—Carboxyl groups
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J3/00—Processes of treating or compounding macromolecular substances
    • C08J3/24—Crosslinking, e.g. vulcanising, of macromolecules
    • C08J3/246—Intercrosslinking of at least two polymers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J3/00—Processes of treating or compounding macromolecular substances
    • C08J3/02—Making solutions, dispersions, lattices or gels by other methods than by solution, emulsion or suspension polymerisation techniques
    • C08J3/03—Making solutions, dispersions, lattices or gels by other methods than by solution, emulsion or suspension polymerisation techniques in aqueous media
    • C08J3/075—Macromolecular gels
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J3/00—Processes of treating or compounding macromolecular substances
    • C08J3/24—Crosslinking, e.g. vulcanising, of macromolecules
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J2300/00—Characterised by the use of unspecified polymers
    • C08J2300/14—Water soluble or water swellable polymers, e.g. aqueous gels

Definitions

  • the present invention relates to a polymer compound having tris (2-carboxyethyl) phosphine (hereinafter referred to as TCEP), and more particularly, to a polymer compound having a TCEP attached to a side chain or a terminal thereof and
  • TCEP tris (2-carboxyethyl) phosphine
  • the present invention relates to a crosslinked polymer compound crosslinked by being bonded to each other by a Michael-type addition reaction between polymer compounds having an acrylic functional group, and also to a use thereof.
  • TCEP is a colorless, odorless, water-soluble, nonvolatile, non-reactive, functional, and oxidation-resistant reducing agent, and has the property of reducing disulfide bonds present in proteins within 5 minutes. Compared with DTT (DiThioThreitol) reducing agent, it is more stable, effective, and can be used as reducing agent in biochemistry and molecular biology because of its ability to reduce disulfide bonds in acid.
  • DTT DiThioThreitol
  • Such TCEP has only been used as a reducing agent and has never been used to form a crosslinked polymer by introducing into a functional group of the polymer, and has never been used to prepare a polymer-TCEP compound into a hydrogel, a film, or the like.
  • crosslinked polymer compound having a TCEP-bonded polymer compound and a polymer compound having an acrylic functional group crosslinked with each other by a Michael type addition reaction.
  • These crosslinked polymer compounds are prepared in the form of hydrogels or scaffolds, and medical materials and drugs that deliver cells, growth factors, hormones, and bioactive materials, such as drugs with medicinal therapeutic functions, where necessary. It is intended to be used as a carrier, tissue engineering support, or cell therapy.
  • the present invention provides a polymer compound bonded with TCEP.
  • the polymer compound is a biocompatible polymer compound having a carboxyl group in the side chain
  • the TCEP forms a bond by a chemical reaction with an amine group of the polymer compound, or the carboxyl group of the TCEP It is to bond with the carboxyl group of the polymer compound through a linker to form a bond by a chemical reaction of.
  • the biocompatible polymer compound includes a polysaccharide group consisting of chondroitin sulfate, carboxymethyl cellulose, hyaluronic acid, heparin, dextran, dermatan sulfate, and alginate; Protein groups consisting of peptides, collagen, gelatin, proteins, hormones and growth factors; And it may be selected from the group consisting of polyethylene oxide, polyvinyl alcohol, polyvinylpyrrolidone, polylactide, polyglycolide.
  • the linker is a compound having at least one amine group, in particular, the linker is dicarboxylic acid dihydrazide, and specifically, the linker may be adipic dihydrazide, hexamethylenediamine. .
  • the polymer compound is a biocompatible polymer compound having an amine group in the side chain, wherein the TCEP is bonded by the chemical reaction of the amine group of the polymer compound.
  • the biocompatible polymer compound is preferably chitosan.
  • the TCEP to be bonded to the polymer compound is preferably bonded to 0.01 ⁇ 1 per repeat unit of the polymer compound.
  • the present invention provides a crosslinked polymer compound crosslinked by the above-mentioned polymer compound and a polymer compound having an acrylic functional group are bonded to each other by a Michael type addition reaction.
  • the crosslinked polymer compound may be formed in the form of a hydrogel, a film, particles, or a tube.
  • the acrylic functional group is preferably acrylate or methacrylate.
  • the crosslinked polymer compound may be used as a drug carrier, tissue engineering support, or cell transfer agent.
  • the present invention also provides a hydrogel comprising the above-mentioned crosslinked polymer compound and comprising a bioactive material.
  • the bioactive material may be selected from the group consisting of cells, drugs, growth factors, hormones, nucleic acids, proteins, RNA, extracellular matrix material and drugs having a medicinal therapeutic function.
  • the polymer compound and the crosslinked polymer compound according to the present invention can be prepared simply by introducing TCEP into the side chain of the polymer and by reacting it with the polymer compound having an acrylic functional group on the side chain.
  • the crosslinked polymer compound of the present invention is made of a hydrogel, a support, a film is a material for artificial organ regeneration for tissue engineering, wrinkle improvement agent, nerve, bone and cartilage regeneration agent and therapeutic agent, dressing agent for burn treatment or beauty,
  • a drug delivery agent for the treatment of arthritis, arthritis and cancer it is expected to promote efficient delivery of drugs and cells and tissue regeneration by biodegradation of hydrogels.
  • 1 is a method for synthesizing chondroitin sulfate-acrylate and chondroitin sulfate-TCEP according to an embodiment of the present invention.
  • FIG. 2 is a chondroitin sulfate hydrogel (A) produced by the reaction of chondroitin sulfate-acrylate and chondroitin sulfate-TCEP prepared according to the method of FIG. 1 and its injectable (B), scaffold type (D ), Digital (C), particulate (F) and tubular (E) supports.
  • A chondroitin sulfate hydrogel
  • Figure 3 is the result of FTIR analysis of chondroitin sulfate-acrylate and chondroitin sulfate-TCEP prepared according to an embodiment of the present invention.
  • Figure 4 is a result of performing a cell culture evaluation for 7 days on the surface of chondroitin sulfate hydrogel, gelatin-containing chondroitin sulfate hydrogel and hyaluronic acid hydrogel prepared according to an embodiment of the present invention (A, B: chondroitin Sulfate hydrogel, C: chondroitin sulfate hydrogel with gelatin, D: hyaluronic acid hydrogel).
  • FIG. 8 is an FTIR analysis result of atelo collagen-TCEP prepared according to an embodiment of the present invention.
  • the present invention primarily relates to a polymer compound to which TCEP is bound.
  • Such polymer compounds are used as raw materials for preparing crosslinked polymer compounds as described below, rather than having their own uses.
  • the polymer compound used in the present invention is a biocompatible polymer compound, and the form in which TCEP is bonded thereto is not particularly limited, but one particularly preferred embodiment is to use a biocompatible polymer compound having a carboxyl functional group or an amine functional group in its side chain.
  • biocompatible polymer compounds examples include polysaccharide compounds such as chondroitin sulfate, carboxymethylcellulose, hyaluronic acid, dextran, chitosan, heparin, dermatan sulfate and alginate, peptides, collagen, RNA, DNA, proteins, hormones and growth factors. Nucleic acids and proteins, and synthetic polymer compounds such as polyethylene oxide, polyvinyl alcohol, polyvinylpyrrolidone, polylactide, and polyglycolide.
  • Linkers are required to introduce TCEP into the side chain of such biocompatible polymeric compounds having a carboxyl group in the side chain. Such linkers must be capable of forming a bond by chemical reaction with a carboxyl group of the polymer compound and also by a chemical reaction with a carboxyl group of TCEP. Representative examples of such linkers are compounds having at least one amine group. Representative compounds belonging to such linkers include dicarboxylic acid dihydrazide, and examples thereof include adipic acid dihydrazide.
  • the polymer compound used in the present invention is a biocompatible polymer compound, and another embodiment is to use a biocompatible polymer compound having an amine group in the side chain.
  • the side chain of the high molecular compound has an amine group
  • the amine group can be directly bonded to the carboxyl group of TCEP, and thus no linker is required.
  • biocompatible polymer compounds include chitosan.
  • the polymer compound in which the TCEP formed as described above is bonded to the side chain is formed to form a hydrogel by crosslinking with the polymer compound having an acrylic functional group on the side chain to form a crosslinked polymer compound. Therefore, in the polymer compound in which the TCEP is bonded to the side chain, the number of TCEPs bonded to the side chain should be such that such crosslinking reaction can occur sufficiently to form a substantial crosslinked polymer compound. To this end, TCEP bonded to the side chain of the polymer compound is preferably bound to 0.01 to 1 range per repeating unit of the polymer compound. If the number of TCEP bound to the side chain is too small, crosslinking to the extent that hydrogels can be formed is not achieved.
  • the polymer compound used in the present invention preferably has a size of 1 to 8,000 kDa, more preferably 3 kDa to 700 kDa.
  • the present invention also provides a crosslinked polymer compound in which a polymer compound having TCEP bonded to a side chain and a polymer compound having an acrylic functional group at the side chain are bonded to each other by a Michael-type addition reaction.
  • the ratio of the acrylic functional group and the TCEP functional group can be variously adjusted according to the use.
  • the molar ratio of the acrylic functional group and the TCEP functional group is 10: 1 to 1:10, preferably 3: 1 to 1: 2, more preferably 1: 1.
  • the acrylic functional group is preferably acrylate or methacrylate.
  • the polymer compound having an acrylic functional group may be the above-mentioned biocompatible polymer.
  • a linker is required in order to connect with the acrylate.
  • the linker can use the one described above. 1 shows an example of using adipic dihydrazide as a linker.
  • adipic dihydrazide In order to induce a reaction between the carboxyl group of chondroitin sulfate and the primary amine group of adipic dihydrazide, and to induce a reaction between the remaining primary amine groups of adipic dihydrazide and acrylic acid, -Ethyl-3- (3-dimethylaminopropyl) carbodiimide] is used.
  • the crosslinked polymer compound of the present invention can be prepared by simply mixing a solution of a polymer compound in which TCEP is bonded to a side chain, such as a PBS solution and a solution of a polymer compound having an acrylic functional group, such as a PBS solution.
  • Such crosslinked polymer compounds are basically prepared in the form of a hydrogel.
  • Hydrogel refers to the three-dimensional structure of the hydrophilic polymer having a sufficient amount of water. Therefore, the polymer compound having such a three-dimensional crosslinked structure is in the form of a hydrogel expanded by absorbing the moisture in the presence of moisture.
  • Such hydrogels may comprise bioactive substances.
  • a bioactive material means a material used for the treatment, healing, prevention or diagnosis of a disease, and examples thereof include proteins or peptides such as cells, growth factors and hormones, nucleic acids, extracellular matrix substances, and medicinal therapeutic functions. Drugs, and the like.
  • the bioactive material may be prepared to be included in one solution, and mixed with the other solution to proceed to synthesize the hydrogel. In this case, the bioactive material becomes a form supported on the crosslinked polymer hydrogel to be formed.
  • the hydrogel of the present invention can be used as a bioactive material transporter, cell transporter or drug transporter. It can also be used as a tissue engineering support or cell therapy.
  • Examples of the drug as a bioactive substance supported on the hydrogel of the present invention include antibiotics, anticancer agents, anti-inflammatory drugs, antiviral agents, antibacterial agents and the like.
  • Antibiotics include derivatives and mixtures of tetracycline, minocycline, doxycycline, opfloxacin, levofloxacin, ciprofloxacin, clarithromycin, erythromycin, cefacller, cefotaxime, imipenem, penicillin, gentamicin, streptomycin, vancomycin and the like. The antibiotic selected can be illustrated.
  • anticancer agent examples include an anticancer agent selected from derivatives and mixtures of methotrexate, carboplatin, taxol, cis-platin, 5-fluorouracil, doxorubicin, etoposide, paclitaxel, camptothecin, cytosine arabinose and the like.
  • the anti-inflammatory agent can be exemplified by an anti-inflammatory agent selected from derivatives and mixtures of indomethacin, ibuprofen, ketoprofen, pyroxicam, flubiprofen, diclofenac and the like.
  • the antiviral agent can be exemplified by an antiviral agent selected from derivatives and mixtures such as acicolober, lovabin and the like.
  • the antimicrobial agent may be exemplified by an antimicrobial agent selected from derivatives and mixtures of ketoconazole, itraconazole, fluconazole, amphotericin-B, griseo fulvin and the like.
  • Proteins and peptides that can be delivered in vivo by being supported on the hydrogel of the present invention include hormones, cytokines, enzymes, antibodies, growth factors, transcriptional regulators, blood factors, vaccines, structural proteins, Various bioactive peptides such as ligand proteins, polysaccharides and receptors, cell surface antigens, receptor antagonists, derivatives and analogs thereof can be exemplified. Specifically, bone growth factor, liver growth hormone, growth hormone releasing hormone and peptide, interferon and interferon receptors (e.g.
  • interferon-alpha, -beta and -gamma, soluble type I interferon receptor, etc. granulocyte colony stimulating factor ( G-CSF), granulocyte-macrophage colony stimulating factor (GM-CSF), glucacon-like peptides (GLP-1, etc.), g-protein-coupled receptors, interleukins (e.g. interleukin) -1, -2, -3, -4, -5, -6, -7, -8, -9, etc., interleukin receptors (e.g.
  • IL-1 receptor IL-4 receptor, etc.
  • enzymes Examples: Glucocerebrosidase, iduronate-2-sulfatase, alpha-galactosidase-A, agalsidase alpha, -beta, Alpha-L-iduronidase, chitinase, butyrylcholinesterase, lipase, glutamate Glutamate decarboxylase, imiglucerase, uricase, platelet-activating factor acetylhydrolase, neutral endopeptidase, myeloperoxidase (myeloperoxidase)), interleukin and cytokine binding proteins (e.g.
  • IL-18bp TNF-binding protein, etc.
  • macrophage activators macrophage peptides, B cell factor, T cell factor, protein A, allergic inhibitors, tumors Tumor Necrosis Factor (TNF) Alpha Inhibitor, Cell Necrosis Glycoprotein, Immunotoxin, Lymphotoxin, Tumor Necrosis Factor, Tumor Suppressor, Metastasis Growth Factor, Alpha-1 Antitrypsin, Albumin, Alpha-Lactalbumin -lactalbumin), apolipoprotein-E, erythropoietin, high glycated erythropoietin, angiopoietin, hemoglobin, thrombin, thrombin receptor activation Peptide, thrombomodulin, blood factor, blood factor a, blood factor XIII, plasminogen activator, fibrin-binding peptide, urokinase, streptokinase, hirudin
  • nerve growth factor ciliary neurotrophic factor, axogenesis factor
  • Axogenesis factor-1 brain-natriuretic peptide, glial derived neurotrophic factor, netrin, neutrophil inhibitor factor, neurotrophic factor, nu Neuturin), parathyroid hormone, relaxin, secretin, somatomedin, insulin-like growth factor, corticosteroids, glucagon, cholecystokinin, pancreatic polypeptide, gastrin releasing peptide, corticotropin releasing factor, thyroid stimulating hormone , Autotaxin, lactoferrin, myostatin, receptors (e.g.
  • TNFR TNFR
  • P55 IL-1 receptor
  • VEGF receptor VEGF receptor
  • B cell activator receptor etc.
  • receptor Antagonists e.g. IL1-Ra
  • cell surface antigens e.g. CD 2, 3, 4, 5, 7, 11a, 11b, 18, 19, 20, 23, 25, 33, 38, 40, 45, 69
  • monoclonal antibodies e.g. CD 2, 3, 4, 5, 7, 11a, 11b, 18, 19, 20, 23, 25, 33, 38, 40, 45, 69
  • monoclonal antibodies e.g. CD 2, 3, 4, 5, 7, 11a, 11b, 18, 19, 20, 23, 25, 33, 38, 40, 45, 69
  • monoclonal antibodies e.g. CD 2, 3, 4, 5, 7, 11a, 11b, 18, 19, 20, 23, 25, 33, 38, 40, 45, 69
  • monoclonal antibodies e.g. CD 2, 3, 4, 5, 7, 11a, 11b, 18, 19, 20, 23, 25, 33, 38, 40, 45, 69
  • monoclonal antibodies e
  • Nucleic acids that can be physically supported or chemically coupled to the hydrogel of the present invention to be delivered in vivo can be exemplified by DNA, RNA, PNA, oligonucleotide, and the like.
  • extracellular matrix material examples include collagen, fibronectin, gelatin, laminin, and vitronectin.
  • Examples of cells that can be physically supported on the hydrogel of the present invention and delivered in vivo include stem cells, fibroblasts, vascular endothelial cells, smooth muscle cells, neurons, chondrocytes, bone cells, skin cells, Schwann cells, and the like. .
  • the crosslinked polymer compound of the present invention may be formed into a film, particle or tube form by drying, in particular lyophilizing it.
  • chondroitin sulfate 0.4 g was dissolved in 80 ml of distilled water to prepare an aqueous solution of chondroitin sulfate (0.5%; w / v), and 0.3 ml of crosslinker EDC and adipic dihydrazide (adipic dihydrazide) were added to the aqueous solution of chondroitin sulfate. 0.3 g of ADH) was added to induce hydrazide functional groups in the polymer side chain of chondroitin sulfate.
  • step 1 0.3 ml EDC and 0.3 g ADH were added to the carboxyl functional group of 0.4 g chondroitin sulfate to induce the hydrazide functional group in the polymer side chain of chondroitin sulfate, followed by 0.6 ml EDC and 1.0 g TCEP. The reaction was induced and lyophilized to produce chondroitin sulfate-TCEP powder (FIG. 1).
  • Chondroitin sulfate-acrylate prepared in step 1 and chondroitin sulfate-TCEP prepared in step 2 were dissolved in PBS solution, respectively, to prepare a 5% chondroitin sulfate-acrylate solution and a 5% chondroitin sulfate-TCEP solution.
  • Two solutions were mixed in a 1: 1 ratio, and gelation (gelation) proceeded to gel within 1 minute (chondroitin sulfate hydrogel) was confirmed by the tilting method (tilting method) (A of FIG. 2).
  • the chondroitin sulfate-acrylate solution and the chondroitin sulfate-TCEP solution obtained in the first and second steps of Example 1 were mixed in a syringe to obtain an injected gel sample, which was confirmed to be prepared as an injectable gel (FIG. 1B).
  • the mixed solution was put into a mold having a certain form such as a film, a tube, and the like to induce gelation to prepare a support, a film form and a support in the form of a tube (C, D, E of Figure 2).
  • 5% chondroitin sulfate prepared by dissolving the chondroitin sulfate-acrylate powder and the chondroitin sulfate-TCEP powder in the first and second steps of Example 1 in a buffer to contain fibroblasts at 10 5 cells / cm 3 , respectively.
  • -Chondroitin sulfate hydrogel containing fibroblasts was prepared by mixing an acrylate solution (100 uL) and 5% chondroitin sulfate-TCEP solution (100 uL).
  • the chondroitin sulfate hydrogel containing the fibroblasts of Example 4 was cultured in vitro for 7 days and stained by a live & dead method and observed by fluorescence microscopy. As a result, all the cells inside the gel were confirmed to be alive (blue).
  • a solution containing 10 uL of the model drug 0.1% rhodamine B in 10% of the 5% chondroitin sulfate-acrylate solution of Example 1 was prepared, and a 5% chondroitin sulfate-TCEP solution (95 uL) And mixed with to prepare a 200 uL chondroitin sulfate hydrogel containing rhodamine.
  • Rhodamine-supported chondroitin sulfate hydrogel prepared in Example 5 induced release of rhodamine in PBS buffer, and it was confirmed that rhodamine was released over time from the chondroitin sulfate hydrogel and released to about 86% over 10 hours. (FIG. 5).
  • Polystyrene culture flasks in which neurons (PC-12) were cultured were loaded with chondroitin sulfate hydrogel containing NGF prepared in Example 6 and observed for NGF release and cell behavior during 7 days in vitro cell culture. While the neurites of the cells were not observed on the culture flask containing no growth factor, the growth factor was released from the hydrogel containing the growth factor, and the induction of the neurites of the neurons was observed under the light microscope. .
  • Example 1 The sample of Example 1 was dissolved in a buffer solution to prepare a 6.67% chondroitin sulfate-acrylate solution (75 uL) and a 6.67% chondroitin sulfate-TCEP solution (75 uL), followed by 10 5 in 1% gelatin solution (50 ul).
  • a gelatin-cell solution was prepared by including fibroblasts of cells / cm 3 .
  • the solution was mixed to prepare a 5% chondroitin sulfate hydrogel containing gelatin-cells.
  • Step 1 hyaluronic acid-acrylate preparation
  • Hyaluronic acid aqueous solution was prepared by dissolving 0.4 g of hyaluronic acid in 120 ml distilled water instead of the chondroitin sulfate of Example 1, and adding a hyaluronic acid-hydrazide sample by adding 0.4 ml EDC and 0.3 g ADH, and then again 0.7 ml Hyaluronic acid-acrylate was prepared by adding EDC and 0.3 mL acrylic acid and then lyophilizing for one day.
  • step 2 0.4 ml EDC and 0.4 g ADH are bound to a carboxyl functional group of 0.4 g hyaluronic acid, followed by addition of 0.7 ml EDC and 1.2 g TCEP, followed by lyophilization to obtain hyaluronic acid-TCEP powder. Prepared.
  • Step 3 preparing hyaluronic acid hydrogel
  • the prepared 0.01 g hyaluronic acid-acrylate and 0.005 g hyaluronic acid-TCEP were dissolved in 100 uL buffer solution, respectively, to prepare a 5% hyaluronic acid-acrylate solution and a 5% hyaluronic acid-TCEP solution.
  • Hyaluronic acid hydrogel was prepared by mixing in a ratio of 1: 1.
  • the PC peak of Lonic Acid-TCEP was penetrated to confirm that hyaluronic acid-acrylate and hyaluronic acid-TCEP were synthesized.
  • a carboxymethylcellulose-hydrazide compound was prepared using carboxymethylcellulose instead of the chondroitin sulfate of Example 1, followed by addition of 0.4 ml EDC and 0.1 mL acrylic acid, and then lyophilized to give carboxymethylcellulose-acrylate. Prepared.
  • step 2 In the same manner as in step 1, 0.4 ml EDC and 0.6 g ADH were added to the carboxyl functional group of 0.6 g carboxymethylcellulose, and then 0.6 g TCEP was reacted and lyophilized to prepare carboxymethyl cellulose-TCEP powder.
  • Step 1 preparing gelatin-acrylate
  • a gelatin-acrylate was prepared by preparing a 0.5% gelatin solution using gelatin instead of the chondroitin sulfate of Example 1, followed by addition of 0.3 m EDC and 0.1 mL acrylic acid, followed by lyophilization.
  • step 2 0.2 g EDC and 0.3 g TCEP were reacted with a 0.4 g gelatin carboxyl functional group, and lyophilized to prepare a gelatin-TCEP powder.
  • 20% gelatin-acrylate solution and 20% gelatin-TCEP solution were prepared by dissolving 0.02 g gelatin-acrylate prepared in step 1 and 0.02 g gelatin-TCEP prepared in step 2 in 100 uL PBS, respectively.
  • the solution was mixed in a 1: 1 ratio to synthesize gelatin hydrogels.
  • chondroitin sulfate of Example 2 0.1 g of gelatin was dissolved in 80 ml of solvent to prepare a collagen solution, which was combined by adding 0.2 ml EDC and 0.2 g ADH, followed by 0.2 ml EDC and 0.3 g of TCEP. was reacted and lyophilized to prepare collagen-TCEP powder.
  • the collagen-TCEP sample observed 848 cm -1 PC peak compared to the collagen to confirm that collagen-TCEP was synthesized.
  • atelo collagen solution 0.05 g was dissolved in a 40 ml solvent to prepare an atelo collagen solution, and chitosan-hydrazide was synthesized by adding 0.02 ml EDC and 0.02 g ADH.
  • atelo-collagen-TCEP was prepared by additionally adding 0.02 ml EDC and 0.03 g of TCEP, followed by freeze drying.
  • FTIR analysis of the atelo collagen-TCEP sample of Example 13 showed that the atelo collagen-TCEP sample had a P-CH2 peak at a wavelength of 1380 cm -1 compared to the atelo collagen. It was confirmed that the synthesis (Fig. 8).

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Abstract

La présente invention concerne un composé polymère auquel est liée de la tris(2-carboxyéthyl)phosphine (TCEP), et un composé polymère réticulé dans lequel ledit composé polymère et un composé polymère possédant un groupe fonctionnel acryle sont liés l'un à l'autre par le biais d'une réaction d'addition de Michael et sont réticulés. Le composé polymère réticulé selon l'invention est préparé sous la forme d'un hydrogel et utilisé en tant que vecteur de médicament, d'échafaudage pour ingénierie tissulaire ou d'agent de thérapie cellulaire.
PCT/KR2012/007242 2011-09-09 2012-09-07 Composé polymère auquel est liée de la tris(2-carboxyéthyl)phosphine WO2013036082A2 (fr)

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CN110078945B (zh) * 2019-05-28 2021-07-30 陕西科技大学 一种明胶基高强度水凝胶的制备方法
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