WO2023065541A1 - 一种具有快速凝血作用的复合材料及其制备方法 - Google Patents

一种具有快速凝血作用的复合材料及其制备方法 Download PDF

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WO2023065541A1
WO2023065541A1 PCT/CN2022/070418 CN2022070418W WO2023065541A1 WO 2023065541 A1 WO2023065541 A1 WO 2023065541A1 CN 2022070418 W CN2022070418 W CN 2022070418W WO 2023065541 A1 WO2023065541 A1 WO 2023065541A1
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parts
composite material
chitosan
blood coagulation
rapid blood
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PCT/CN2022/070418
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English (en)
French (fr)
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胡章
张兆宇
卢思彤
黄德坚
余佳音
程瑜
廖铭能
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广东海洋大学
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Publication of WO2023065541A1 publication Critical patent/WO2023065541A1/zh
Priority to US18/341,748 priority Critical patent/US11992563B2/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
    • A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
    • A61L15/42—Use of materials characterised by their function or physical properties
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
    • A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
    • A61L15/18—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons containing inorganic materials
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
    • A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
    • A61L15/20—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons containing organic materials
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
    • A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
    • A61L15/22—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons containing macromolecular materials
    • A61L15/28—Polysaccharides or their derivatives
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
    • A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
    • A61L15/42—Use of materials characterised by their function or physical properties
    • A61L15/425—Porous materials, e.g. foams or sponges
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
    • A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
    • A61L15/42—Use of materials characterised by their function or physical properties
    • A61L15/44—Medicaments
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
    • A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
    • A61L15/42—Use of materials characterised by their function or physical properties
    • A61L15/46—Deodorants or malodour counteractants, e.g. to inhibit the formation of ammonia or bacteria
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L24/00—Surgical adhesives or cements; Adhesives for colostomy devices
    • A61L24/0047—Composite materials, i.e. containing one material dispersed in a matrix of the same or different material
    • A61L24/0073—Composite materials, i.e. containing one material dispersed in a matrix of the same or different material with a macromolecular matrix
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L24/00—Surgical adhesives or cements; Adhesives for colostomy devices
    • A61L24/0047—Composite materials, i.e. containing one material dispersed in a matrix of the same or different material
    • A61L24/0073—Composite materials, i.e. containing one material dispersed in a matrix of the same or different material with a macromolecular matrix
    • A61L24/0089—Composite materials, i.e. containing one material dispersed in a matrix of the same or different material with a macromolecular matrix containing inorganic fillers not covered by groups A61L24/0078 or A61L24/0084
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
    • C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
    • C08B37/0006—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid
    • C08B37/0024—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid beta-D-Glucans; (beta-1,3)-D-Glucans, e.g. paramylon, coriolan, sclerotan, pachyman, callose, scleroglucan, schizophyllan, laminaran, lentinan or curdlan; (beta-1,6)-D-Glucans, e.g. pustulan; (beta-1,4)-D-Glucans; (beta-1,3)(beta-1,4)-D-Glucans, e.g. lichenan; Derivatives thereof
    • C08B37/0027—2-Acetamido-2-deoxy-beta-glucans; Derivatives thereof
    • C08B37/003—Chitin, i.e. 2-acetamido-2-deoxy-(beta-1,4)-D-glucan or N-acetyl-beta-1,4-D-glucosamine; Chitosan, i.e. deacetylated product of chitin or (beta-1,4)-D-glucosamine; Derivatives thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/10—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing inorganic materials
    • A61L2300/102—Metals or metal compounds, e.g. salts such as bicarbonates, carbonates, oxides, zeolites, silicates
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/20—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials
    • A61L2300/21—Acids
    • A61L2300/214—Amino acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/20—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials
    • A61L2300/23—Carbohydrates
    • A61L2300/236—Glycosaminoglycans, e.g. heparin, hyaluronic acid, chondroitin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/40—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
    • A61L2300/418—Agents promoting blood coagulation, blood-clotting agents, embolising agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2400/00—Materials characterised by their function or physical properties
    • A61L2400/04—Materials for stopping bleeding

Definitions

  • the invention relates to the field of agricultural technology, in particular to a composite material with rapid coagulation effect and a preparation method thereof.
  • hemostatic materials such as glutaraldehyde-crosslinked albumin and fibrin-based bandages have good hemostatic effects, it is difficult for traditional hemostatic agents to control this type of hemostasis in patients with coagulopathy who lack coagulation factors or have dysfunctional hemostasis. Blood loss, which increases the risk of death from massive blood loss. Therefore, controlling hemostasis with coagulopathy bleeding remains a great challenge.
  • CN104114198A discloses a hemostatic wound dressing comprising a non-colloidal porous dressing material and a plurality of fibrinogen-binding peptides immobilized to the non-colloidal porous dressing material.
  • the dressing can accelerate hemostasis, but has no effect on hemostasis caused by lack of coagulation factors or abnormal function in patients with coagulopathy.
  • the object of the present invention is to provide a kind of composite material with fast blood coagulation and preparation method thereof, to solve the problems in the above-mentioned prior art, by using chitosan, salicylaldehyde, 2-aminoisonicotinic acid, ferrous sulfate , dopa and disodium hydrogen phosphate as raw materials to prepare a composite material with remarkable effect on blood coagulation, the material has high stability and biological safety, and the preparation process is simple and easy for industrialized production.
  • the present invention provides the following scheme:
  • a composite material with rapid blood coagulation comprising the following raw materials in parts by mass: 8-20 parts of chitosan, 7-14 parts of salicylaldehyde, 2-aminoisonicotinic acid 8 to 16 parts, 1.5 to 3.5 parts of ferrous sulfate, 3 to 5 parts of dopa and 10 to 15 parts of disodium hydrogen phosphate.
  • the following raw materials are included: 14 parts of chitosan, 10 parts of salicylaldehyde, 12 parts of 2-aminoisonicotinic acid, 2.5 parts of ferrous sulfate, 4 parts of dopa and 13 parts of disodium hydrogen phosphate share.
  • Chitosan is a product of deacetylation after extracting chitin from the shells of marine crustaceans such as shrimps and crabs. It is non-toxic, non-irritating, non-immunogenic, non-pyrogenic, non-hemolytic, and biodegradable And good biocompatibility, widely used in the manufacture of artificial skin, surgical sutures, drug carriers, etc., and more and more attention has been paid in the field of hemostatic dressings.
  • the coagulation effect of chitosan is affected by many factors, such as structural unit, molecular weight, degree of deacetylation, cationic properties, etc.
  • Dopa is an amino acid organic compound extracted from quinoa beans or artificially synthesized. It is an important part of the mucus secreted by marine mussels and other organisms. It has strong adhesion and can not only adhere to the surface of inorganic materials, but also Adheres to the surface of organic materials. The super-adhesive properties of the adhesive protein secreted by the marine mussels' foot silk in a humid water environment are particularly noticeable, and this excellent performance is unmatched by other adhesives at present. The strong wet adhesion properties of marine mussel secretions are closely related to catechol structural fragments.
  • Hemoglobin is the functional unit of red blood cells, and red blood cells are the carrier of hemoglobin. Hemoglobin is a tetramer composed of four subunits, each of which has a pigment prosthetic group, namely heme. Heme is an iron-containing porphyrin compound, in which ferrous ions can reversibly combine with oxygen and participate in the transfer of oxygen in organisms.
  • the chitosan has a molecular weight of 50-300KDa and a deacetylation degree of 80-95%.
  • the chitosan has a molecular weight of 100KDa and a deacetylation degree of 90%.
  • the chitosan molecular weight and deacetylation degree are too low, the coagulation effect of the final composite material will not be significant; if the chitosan molecular weight and deacetylation degree are too high, the preparation process will be difficult and the cost will be greatly increased, which is not conducive to industrialization.
  • the second technical solution of the present invention a method for preparing the above-mentioned composite material with rapid blood coagulation, comprising the following steps:
  • step (2) After dissolving the chitosan-aminoisonicotinic acid graft prepared in step (1), adjust the pH value to 5.0 to 7.0, then add salicylaldehyde, and after stirring for a certain period of time, add ferrous sulfate, After the barium and disodium hydrogen phosphate are mixed evenly, they are left standing in the dark to obtain the composite material with rapid blood coagulation.
  • Ferric sulfate is added first, which is conducive to the formation of coordination compounds and produces a significant coagulation effect; if ferric sulfate is added later, the dopa or disodium hydrogen phosphate added first will easily generate hydrogen bonds with chitosan-based ligand groups or The electrostatic effect hinders the coordination of ferrous ions, which leads to the reduction of the adsorption and aggregation ability of the composite material on red blood cells, and the weakening of the coagulation effect.
  • the solvent in the chitosan solution is acetic acid aqueous solution, and the mass fraction of chitosan is 2%-5%.
  • the mass fraction of chitosan in the chitosan solution is 3%.
  • the mass fraction of acetic acid in the aqueous acetic acid solution is 2%.
  • the molar ratio of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride to N-hydroxysuccinimide is 5:1.
  • the volume of the absolute ethanol is 10%-30% of the volume of the acetic acid aqueous solution.
  • the volume of the absolute ethanol is 20% of the volume of the acetic acid aqueous solution.
  • the molar ratio of 2-aminoisonicotinic acid and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride in the ethanol solution described in step (1) is 2:3 .
  • the stirring reaction time in step (1) is 6 to 8 hours; the pH value in step (2) is 5.5, and the reagent used to adjust the pH value is 0.1mol/L HCl solution, and the stirring time is 4 hours. ⁇ 6h.
  • the pH value of the solution is preferably 5.0 to 7.0, more preferably 5.5.
  • the composite material of the present invention has rapid blood coagulation effect, and its blood coagulation effect is not dependent on the normal blood coagulation pathway, and is especially suitable for hemostasis accompanied by hemorrhage caused by coagulation disorders.
  • the present invention firstly adopts EDC ⁇ HCl and NHS coupling agent to introduce 2-aminoisonicotinic acid on the chitosan molecular chain through amidation reaction, and then through salicylaldehyde and the compound on the 2-aminoisonicotinic acid Amino condensation forms a Schiff base, constructs a ligand for ferrous ions, and forms a chitosan-Fe 2+ complex after reacting with ferrous sulfate; on the one hand, this complex cross-ligates with red blood cell hemoglobin
  • the protonated amino group (positively charged) in chitosan is used to interact with the negatively charged red blood cell surface to exert a synergistic effect of adsorption and aggregation of red blood cells.
  • the coagulation test in vitro proves that the composite material constructed by the chitosan-Fe 2+ complex promotes the adsorption and aggregation of red blood cells in the blood, and produces a significant coagulation effect.
  • the present invention utilizes the internal salt characteristic of amino acid in the dopa structure, realizes the formation of charge transfer complex, is conducive to the formation of intermolecular hydrogen bond in composite material, strengthens the stability of composite material;
  • the strong reducibility of the phenolic fragment can effectively protect the Fe 2+ in the composite material from being oxidized, and has strong stability;
  • the catechol structure in the dopa structure exerts the characteristics similar to mussel adhesion protein, resulting in the composite material After absorbing liquid to form a hydrogel, it has strong adhesion and can effectively block the bleeding port, especially suitable for emergency treatment of deep and narrow massive bleeding.
  • the disodium hydrogen phosphate added in the present invention utilizes the hydrogen phosphate radical anion and the hydroxyl characteristic in its structure, can produce electrostatic and hydrogen bonding with the amino group (cation) of chitosan protonation, thereby make composite material have
  • the stable porous structure can quickly absorb liquid and swell to form a gel with a certain volume, which is not easy to enter the blood and cause the risk of thrombus.
  • the small molecular structure of disodium hydrogen phosphate while promoting the stability of the composite structure, does not hinder the coordination of Fe 2+ , which is beneficial to blood coagulation.
  • the composite material prepared by the present invention has low hemolysis rate and high biological safety; the preparation process is simple and easy for industrial production.
  • Fig. 1 is the scanning electron micrograph of the composite material with rapid coagulation effect prepared in Example 1 of the present invention
  • Fig. 2 is the comparison chart of whole blood coagulation index of the composite material with rapid coagulation effect prepared by the present invention
  • Fig. 3 is the comparison chart of red blood cell adsorption rate of the composite material with rapid blood coagulation prepared by the present invention
  • Fig. 4 is the scanning electron micrograph of the red blood cell adsorbed by the composite material with rapid coagulation effect prepared in Example 1 of the present invention
  • Fig. 5 is a comparison chart of the hemolysis rate of the composite material with rapid coagulation effect prepared in the present invention.
  • a preparation method of a composite material with rapid blood coagulation is a preparation method of a composite material with rapid blood coagulation:
  • the chitosan-aminoisonicotinic acid graft that step (3) is prepared is dissolved with 4526g acetic acid aqueous solution (the mass fraction of acetic acid in the acetic acid aqueous solution is 2%), adjusts pH with the HCl solution of 0.1mol/L value to 5.5, then add 100g salicylaldehyde under stirring conditions, add 25g ferrous sulfate, 40g dopa and 130g disodium hydrogen phosphate successively after stirring for 5h, stir evenly at room temperature, keep away from light for 12h, and freeze-dry to obtain Composite material with rapid blood coagulation, the scanning electron microscope picture is shown in Figure 1.
  • step (3) the chitosan-aminoisonicotinic acid graft that step (3) is prepared is dissolved with 3840g acetic acid aqueous solution (the mass fraction of acetic acid in the acetic acid aqueous solution is 2%), adjusts pH with the HCl solution of 0.1mol/L value to 6.0, then add 120g salicylaldehyde under stirring conditions, add 20g ferrous sulfate, 35g dopa and 110g disodium hydrogen phosphate successively after stirring for 6h, stir evenly at room temperature, keep away from light for 12h, and freeze-dry to obtain Composite material with rapid coagulation.
  • step (3) the chitosan-aminoisonicotinic acid graft prepared by step (3) is dissolved with 3920g aqueous acetic acid (the mass fraction of acetic acid in the aqueous acetic acid solution is 2%), and the pH is adjusted with the HCl solution of 0.1mol/L value to 7.0, then add 70g salicylaldehyde under stirring conditions, add 15g ferrous sulfate, 30g dopa and 100g disodium hydrogen phosphate successively after stirring for 4h, stir evenly at room temperature, keep away from light for 12h, and freeze-dry to obtain Composite material with rapid coagulation.
  • 3920g aqueous acetic acid the mass fraction of acetic acid in the aqueous acetic acid solution is 2%
  • the pH is adjusted with the HCl solution of 0.1mol/L value to 7.0, then add 70g salicylaldehyde under stirring conditions, add 15g ferrous sulfate, 30g dopa and
  • step (3) the chitosan-aminoisonicotinic acid graft prepared by step (3) is dissolved with 3920g aqueous acetic acid (the mass fraction of acetic acid in the aqueous acetic acid solution is 2%), and the pH is adjusted with the HCl solution of 0.1mol/L value to 5.0, then add 140g salicylaldehyde under stirring conditions, add 35g ferrous sulfate, 50g dopa and 150g disodium hydrogen phosphate successively after stirring for 5h, stir evenly at room temperature, keep away from light for 12h, and freeze-dry to obtain Composite material with rapid coagulation.
  • Example 2 Same as Example 1, the difference is that the disodium hydrogen phosphate in step (4) is replaced by ⁇ -sodium glycerophosphate.
  • chitosan 120g 2-aminoisonicotinic acid, 100g salicylaldehyde, 25g ferrous sulfate, 40g dopa and 130g disodium hydrogen phosphate raw material, join 4526g acetic acid aqueous solution (the massfraction of acetic acid in the acetic acid aqueous solution is 2 %), uniformly suspended, and freeze-dried to obtain a composite material.
  • BCI Bood Clotting Index
  • PBS phosphate solution
  • the sample was placed in 1% Triton-X100 solution, placed at 37°C for 1 hour to lyse the adhered red blood cells, and its absorbance was measured at 540nm.
  • the red blood cell solution without composite material was used as the control group.
  • a sample is the absorbance value of each sample group
  • a control is the absorbance value of the control group without adding samples.
  • Examples 1-4 can use the chitosan-Fe 2+ complex to produce cross-coordination with the hemoglobin of red blood cells to promote the adsorption and aggregation of red blood cells; in addition, the chitosan protonated amino groups carry The positive charge of the red blood cells attracts more negatively charged red blood cells, which produces a synergistic effect on the adsorption and aggregation of red blood cells, prompting a large number of red blood cells to coagulate quickly to form blood clots.
  • the hemolysis rate test was carried out on the prepared composite materials.
  • the evaluation refers to the national standard GB/T14233.2-2005 Inspection method for medical infusion, blood transfusion and injection equipment, the method is as follows:
  • the hemolysis rate test is to evaluate the safety of the material on blood by detecting the amount of hemoglobin released from red blood cells after the material interacts with fresh anticoagulated blood by absorbance.
  • the national standard stipulates that the hemolysis rate is less than 5% to be judged qualified.
  • the hemolysis rate of the composite materials in Examples 1 to 4 and Comparative Examples 1 to 3 is all less than 5%, which meets the qualification criteria for biological materials in the national standard GB/T 14233.2-2005, Satisfying the hemolytic requirements of biological materials means that the prepared composite material has good blood compatibility and high biological safety, and the composite material prepared by the present invention can quickly absorb liquid and swell to form a gel, has a certain volume, and is not easy to Entering the bloodstream causes a risk of blood clots.

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Abstract

本发明公开了一种具有快速凝血作用的复合材料及其制备方法，属于生物医药技术领域。具有快速凝血作用的复合材料以质量份数计，包括以下原料：壳聚糖8～20份、水杨醛7～14份、2-氨基异烟酸8～16份、硫酸亚铁1.5～3.5份、多巴3～5份和磷酸氢二钠10～15份。本发明制备得到的具有快速凝血作用的复合材料具有良好的产品稳定性，生物安全性高，对血液凝结具有显著的效果，工艺简单，易于工业化生产。

Description

一种具有快速凝血作用的复合材料及其制备方法 技术领域
本发明涉及农业技术领域，特别是涉及一种具有快速凝血作用的复合材料及其制备方法。
背景技术
据统计，不受控制的出血是死亡的主要原因，每年失血性休克死亡占创伤性伤害死亡人数的30％～45％。因此，高效率止血材料的开发成为快速止血，降低出血死亡率的关键。虽然戊二醛交联白蛋白、纤维蛋白基绷带等止血材料具有良好的止血效果，但对于凝血病患者缺乏凝血因子或功能异常导致止血效率降低的止血，传统止血剂通常难以控制这种类型的失血，这增加了因大量失血而死亡的风险。因此，控制伴有凝血障碍出血的止血仍然是一个巨大的挑战。CN104114198A公开了一种止血伤口敷料，包含非胶体多孔敷料材料和固定到所述非胶体多孔敷料材料的多个纤维蛋白原结合肽。所述敷料能够加速止血，但对于凝血病患者缺乏凝血因子或功能异常导致止血都无效果。
在市售的止血材料中，QuikClot沸石止血粉用于严重出血创伤的急救，但在使用中会释放大量热量，致使局部高温，造成创面组织再次伤害。粉末止血材料很容易在脉管内腔残留，阻塞末梢动脉流动形成血栓。如WoundStat止血剂，存在进入血液循环系统引起末梢血栓的风险等。而如何制备得到一种性质稳定，且生物安全性较高的凝血材料成为本领域技术人员亟待解决的技术难题。
发明内容
本发明的目的是提供一种具有快速凝血作用的复合材料及其制备方法，以解决上述现有技术存在的问题，通过以壳聚糖、水杨醛、2-氨基异烟酸、硫酸亚铁、多巴和磷酸氢二钠为原料制备得到一种对血液凝结具有显著的效果的复合材料，该材料稳定性及生物安全性均较高，且制备工艺简单，易于工业化生产。
为实现上述目的，本发明提供了如下方案：
本发明的技术方案之一：一种具有快速凝血作用的复合材料，以质量份数计，包括以下原料：壳聚糖8～20份、水杨醛7～14份、2-氨基异烟酸8～16份、硫酸亚铁1.5～3.5份、多巴3～5份和磷酸氢二钠10～15份。
进一步地，以质量份数计，包括以下原料：壳聚糖14份、水杨醛10份、2-氨基异烟酸12份、硫酸亚铁2.5份、多巴4份和磷酸氢二钠13份。
壳聚糖是主要从海洋甲壳类生物如虾、蟹等外壳提取出甲壳素后脱乙酰化的产物，具有无毒性，无刺激性，无免疫原性，无热源反应，不溶血，可生物降解及良好的生物相容性，广泛用于制造人工皮肤、手术缝合线、药物载体等，在止血敷料领域也越来越受到关注。壳聚糖的凝血作用受到诸多因素的影响，如结构单元、分子量、脱乙酰度、阳离子特性等。
多巴是以藜豆为原料提取或人工合成的一种氨基酸有机化合物，是海洋贻贝等生物分泌黏液的重要组成部分，它具有很强的黏附性，不仅可以黏附在无机材料表面，也可以黏附在有机材料表面。海洋贻贝类生物足丝分泌的黏性蛋白在潮湿的水环境下所展现出的超强黏附性尤为引人注意，这种优异性能是目前其它黏合剂所无法比拟的。海洋贻贝分泌物的强湿粘附性能与儿茶酚结构片段密切相关。
硫酸亚铁中的亚铁离子是血红蛋白的重要组成成分。血红蛋白是红细胞功能单位，红细胞是血红蛋白的载体。血红蛋白是由四个亚基构成的四聚体，每个亚基中都有一个色素辅基，即血红素。血红素是一种含铁的卟啉化合物，其中的亚铁离子能够与氧发生可逆结合，参与生物体中氧的传递作用。
进一步地，所述壳聚糖的分子量为50～300KDa，脱乙酰度为80～95％。
进一步地，所述壳聚糖的分子量为100KDa，脱乙酰度为90％。
若壳聚糖分子量和脱乙酰度太低，导致最后制备的复合材料凝血效果不显著；若壳聚糖分子量和脱乙酰度太高，导致制备工艺难度提升，成本极大增高，不利于工业化。
本发明的技术方案之二：一种上述的具有快速凝血作用的复合材料的制备方法，包括以下步骤：
(1)在壳聚糖溶液中加入2-氨基异烟酸和偶联剂混合的乙醇溶液，搅拌反应得到壳聚糖-氨基异烟酸接枝物；所述偶联剂为1-乙基-3-(3-二甲基氨基丙基)碳二亚胺盐酸盐和N-羟基琥珀酰亚胺；
(2)将步骤(1)制备得到的壳聚糖-氨基异烟酸接枝物溶解后，调pH值至5.0～7.0，然后加入水杨醛，搅拌一定时间后依次加入硫酸亚铁、多巴和磷酸氢二钠混合均匀后，避光静置，得到所述具有快速凝血作用的复合材料。
硫酸铁先加入，有利于形成配位化合物，产生显著的凝血效果；若硫酸铁后加入，则先加入的多巴或者磷酸氢二钠易与壳聚糖基配体基团产生氢键作用或者静电作用而阻碍亚铁离子的配位，导致复合材料对红细胞的 吸附、聚集能力降低，凝血效果减弱。
进一步地，所述壳聚糖溶液中的溶剂为醋酸水溶液，壳聚糖的质量分数为2％～5％。
更进一步地，所述壳聚糖溶液中壳聚糖的质量分数为3％。
若壳聚糖溶液的质量浓度太低，提升了后续工艺的难度，且制备的材料凝血效果不理想；若壳聚糖溶液的质量浓度太高，溶液体系太粘稠，导致反应不均匀，复合材料凝血效果差。
进一步地，所述醋酸水溶液中醋酸的质量分数为2％。
进一步地，所述1-乙基-3-(3-二甲基氨基丙基)碳二亚胺盐酸盐和N-羟基琥珀酰亚胺的摩尔比为5：1。
更进一步地，所述无水乙醇的体积为醋酸水溶液体积的10％～30％。
更进一步地，所述无水乙醇的体积为醋酸水溶液体积的20％。
若无水乙醇的体积太小，则溶质浓度太大，随后添加到壳聚糖溶液中导致反应不均匀，复合材料凝血效果差；若无水乙醇的体积太大，则随后添加到壳聚糖溶液中导致壳聚糖直接析出，致使最后制备的复合材料无显著凝血效果。
进一步地，步骤(1)所述乙醇溶液中2-氨基异烟酸和1-乙基-3-(3-二甲基氨基丙基)碳二亚胺盐酸盐的摩尔比为2：3。
进一步地，步骤(1)中所述搅拌反应的时间为6～8h；步骤(2)中所述pH值为5.5，调节pH值所用的试剂为0.1mol/L的HCl溶液，搅拌时间为4～6h。
若溶液的pH值过小(＜5.0)，则酸性过强，水杨醛与氨基缩合的席夫 碱难以形成；若溶液的pH值过大(＞7.0)，亚铁离子在体系中不稳定，易以沉淀析出。因此，pH值优选为5.0～7.0，更优选为5.5。
本发明公开了以下技术效果：
(1)本发明复合材料具有快速凝血作用，其凝血作用是不依赖于正常的凝血途径，特别适用于伴有凝血障碍出血的止血。
(2)本发明首先采用EDC·HCl和NHS偶联剂通过酰胺化反应将2-氨基异烟酸引入到壳聚糖分子链上，再通过水杨醛和与2-氨基异烟酸上的氨基缩合形成席夫碱，构建亚铁离子的配体，与硫酸亚铁作用后，形成壳聚糖基-Fe 2+配合物；该配合物一方面通过Fe 2+与红细胞的血红蛋白产生交叉配位，促进红细胞聚集；另一方面利用壳聚糖中质子化的氨基(带正电)与带负电荷的红细胞表面相互作用，发挥协同吸附、聚集红细胞效应。体外凝血试验证明，壳聚糖基-Fe 2+配合物所构筑的复合材料促进血液中红细胞的吸附、聚集，产生显著的凝血效果。
(2)本发明利用多巴结构中氨基酸的内盐特性，实现电荷转移复合物的形成，有利于复合材料中分子间氢键的形成，增强复合材料的稳定性；同时利用结构中邻苯二酚片段的强还原性，可有效保护复合材料中Fe 2+不被氧化，具有较强的稳定性；此外，多巴结构中邻苯二酚结构发挥类似贻贝粘附蛋白特性，致使复合材料吸收液体成水凝胶后具有强粘附性，有效封堵出血口，特别适用于深、窄等大量出血的紧急治疗。
(3)本发明中添加的磷酸氢二钠，利用其结构中的磷酸氢根阴离子和羟基特性，可与壳聚糖质子化的氨基(阳离子)产生静电和氢键作用，从而使复合材料具有稳定的多孔结构，可快速吸液溶胀，形成凝胶，具有一 定体积，不易进入血液引起血栓风险。此外，磷酸氢二钠小分子结构，在促进复合材料结构稳定的同时，不阻碍Fe 2+的配位，有利于凝血。
(4)本发明所制备复合材料溶血率低，生物安全性高；制备工艺简单，易于工业化生产。
附图说明
为了更清楚地说明本发明实施例或现有技术中的技术方案，下面将对实施例中所需要使用的附图作简单地介绍，显而易见地，下面描述中的附图仅仅是本发明的一些实施例，对于本领域普通技术人员来讲，在不付出创造性劳动的前提下，还可以根据这些附图获得其他的附图。
图1是本发明实施例1制备的具有快速凝血作用的复合材料的扫描电镜图；
图2为本发明制备的具有快速凝血作用的复合材料全血凝血指数对比图；
图3是本发明制备的具有快速凝血作用的复合材料的红细胞吸附率对比图；
图4是本发明实施例1制备的具有快速凝血作用的复合材料吸附红细胞的扫描电镜图；
图5是本发明制备的具有快速凝血作用的复合材料的溶血率对比图。
具体实施方式
现详细说明本发明的多种示例性实施方式，该详细说明不应认为是对本发明的限制，而应理解为是对本发明的某些方面、特性和实施方案的更详细的描述。
应理解本发明中所述的术语仅仅是为描述特别的实施方式，并非用于限制本发明。另外，对于本发明中的数值范围，应理解为还具体公开了该范围的上限和下限之间的每个中间值。在任何陈述值或陈述范围内的中间值以及任何其他陈述值或在所述范围内的中间值之间的每个较小的范围也包括在本发明内。这些较小范围的上限和下限可独立地包括或排除在范围内。
除非另有说明，否则本文使用的所有技术和科学术语具有本发明所述领域的常规技术人员通常理解的相同含义。虽然本发明仅描述了优选的方法和材料，但是在本发明的实施或测试中也可以使用与本文所述相似或等同的任何方法和材料。本说明书中提到的所有文献通过引用并入，用以公开和描述与所述文献相关的方法和/或材料。在与任何并入的文献冲突时，以本说明书的内容为准。
在不背离本发明的范围或精神的情况下，可对本发明说明书的具体实施方式做多种改进和变化，这对本领域技术人员而言是显而易见的。由本发明的说明书得到的其他实施方式对技术人员而言是显而易见的。本申请说明书和实施例仅是示例性的。
关于本文中所使用的“包含”、“包括”、“具有”、“含有”等等，均为开放性的用语，即意指包含但不限于。
实施例1
一种具有快速凝血作用的复合材料的制备方法：
(1)在140g的壳聚糖(壳聚糖为分子量100KDa，脱乙酰度90％)中加入4526g醋酸水溶液(醋酸水溶液中醋酸的质量分数为2％)溶解后 得到壳聚糖溶液。
(2)将120g(0.8688mol)2-氨基异烟酸用无水乙醇(无水乙醇的用量为步骤(1)中醋酸水溶液体积的20％)溶解后，加入1.3032mol的EDC·HCl和0.2606mol的NHS，室温(25℃左右)搅拌3h后得到2-氨基异烟酸与偶联剂混合的乙醇溶液。
(3)将步骤(2)制备得到的2-氨基异烟酸与偶联剂混合的乙醇溶液加入步骤(1)中制备得到的壳聚糖溶液中室温搅拌8h，蒸馏水透析，冷冻干燥，得到壳聚糖-氨基异烟酸接枝物。
(4)将步骤(3)制备得到的壳聚糖-氨基异烟酸接枝物用4526g醋酸水溶液(醋酸水溶液中醋酸的质量分数为2％)溶解，用0.1mol/L的HCl溶液调节pH值至5.5，然后在搅拌条件下加入100g水杨醛，搅拌5h后依次加入25g硫酸亚铁、40g多巴和130g磷酸氢二钠，室温下搅拌均匀，避光静置12h，冷冻干燥，得到具有快速凝血作用的复合材料，扫描电镜图见图1。
实施例2
(1)在160g的壳聚糖(壳聚糖为分子量100KDa，脱乙酰度90％)中加入3840g醋酸水溶液(醋酸水溶液中醋酸的质量分数为2％)溶解后得到壳聚糖溶液。
(2)将100g(0.724mol)2-氨基异烟酸用无水乙醇(无水乙醇的用量为步骤(1)中醋酸水溶液体积的20％)溶解后，加入1.086mol的EDC·HCl和0.2172mol的NHS，室温(25℃左右)搅拌3h后得到2-氨基异烟酸与偶联剂混合的乙醇溶液。
(3)将步骤(2)制备得到的2-氨基异烟酸与偶联剂混合的乙醇溶液加入步骤(1)中制备得到的壳聚糖溶液中室温搅拌7h，蒸馏水透析，冷冻干燥，得到壳聚糖-氨基异烟酸接枝物。
(4)将步骤(3)制备得到的壳聚糖-氨基异烟酸接枝物用3840g醋酸水溶液(醋酸水溶液中醋酸的质量分数为2％)溶解，用0.1mol/L的HCl溶液调节pH值至6.0，然后在搅拌条件下加入120g水杨醛，搅拌6h后依次加入20g硫酸亚铁、35g多巴和110g磷酸氢二钠，室温下搅拌均匀，避光静置12h，冷冻干燥，得到具有快速凝血作用的复合材料。
实施例3
(1)在80g的壳聚糖(壳聚糖为分子量100KDa，脱乙酰度90％)中加入3920g醋酸水溶液(醋酸水溶液中醋酸的质量分数为2％)溶解后得到壳聚糖溶液。
(2)将80g(0.5792mol)2-氨基异烟酸用无水乙醇(无水乙醇的用量为步骤(1)中醋酸水溶液体积的30％)溶解后，加入0.8688mol的EDC·HCl和0.1738mol的NHS，室温(25℃左右)搅拌3h后得到2-氨基异烟酸与偶联剂混合的乙醇溶液。
(3)将步骤(2)制备得到的2-氨基异烟酸与偶联剂混合的乙醇溶液加入步骤(1)中制备得到的壳聚糖溶液中室温搅拌6h，蒸馏水透析，冷冻干燥，得到壳聚糖-氨基异烟酸接枝物。
(4)将步骤(3)制备得到的壳聚糖-氨基异烟酸接枝物用3920g醋酸水溶液(醋酸水溶液中醋酸的质量分数为2％)溶解，用0.1mol/L的HCl溶液调节pH值至7.0，然后在搅拌条件下加入70g水杨醛，搅拌4h后依 次加入15g硫酸亚铁、30g多巴和100g磷酸氢二钠，室温下搅拌均匀，避光静置12h，冷冻干燥，得到具有快速凝血作用的复合材料。
实施例4
(1)在200g的壳聚糖(壳聚糖为分子量100KDa，脱乙酰度90％)中加入3800g醋酸水溶液(醋酸水溶液中醋酸的质量分数为2％)溶解后得到壳聚糖溶液。
(2)将160g(1.1584mol)2-氨基异烟酸用无水乙醇(无水乙醇的用量为步骤(1)中醋酸水溶液体积的10％)溶解后，加入1.7376mol的EDC·HCl和0.3475mol的NHS，室温(25℃左右)搅拌3h后得到2-氨基异烟酸与偶联剂混合的乙醇溶液。
(3)将步骤(2)制备得到的2-氨基异烟酸与偶联剂混合的乙醇溶液加入步骤(1)中制备得到的壳聚糖溶液中室温搅拌8h，蒸馏水透析，冷冻干燥，得到壳聚糖-氨基异烟酸接枝物。
(4)将步骤(3)制备得到的壳聚糖-氨基异烟酸接枝物用3920g醋酸水溶液(醋酸水溶液中醋酸的质量分数为2％)溶解，用0.1mol/L的HCl溶液调节pH值至5.0，然后在搅拌条件下加入140g水杨醛，搅拌5h后依次加入35g硫酸亚铁、50g多巴和150g磷酸氢二钠，室温下搅拌均匀，避光静置12h，冷冻干燥，得到具有快速凝血作用的复合材料。
对比例1
同实施例1，区别在于，将步骤(2)中的2-氨基异烟酸替换成3-氨基苯甲酸。
对比例2
同实施例1，区别在于，将步骤(4)中的磷酸氢二钠替换成β-甘油磷酸钠。
对比例3
同实施例1，区别在于，步骤(4)中未加入硫酸亚铁。
对比例4
将140g壳聚糖、120g 2-氨基异烟酸、100g水杨醛、25g硫酸亚铁、40g多巴和130g磷酸氢二钠原料，加入到4526g醋酸水溶液(醋酸水溶液中醋酸的质量分数为2％)中，混悬均匀，冷冻干燥，得到复合材料。
效果例1
体外凝血试验：
采用全血凝血指数BCI(Blood Clotting Index)对材料的体外凝血性能进行评价。自新西兰兔耳缘静脉取血于真空采血管中(含有枸橼酸钠抗凝剂，3.8％枸橼酸钠：血＝1：9)，备用；分别取将实施例1～4、对比例1～4制备得到的复合材料以及阳性对照(云南白药)0.1g平铺在50mL的离心管底部。将离心管放入37℃恒温摇床孵育5min后，将20μL的新鲜抗凝兔血滴加到样品上，随即加入20μL 0.2M的CaCl 2溶液，然后在37℃恒温摇床孵育15min后，缓缓地加入25mL的去离子水于离心管中，37℃恒温摇床振摇5min。然后取出部分溶液，用紫外分光光度计在波长545nm处测其吸光度(A 样品)。另取20μL的新鲜抗凝兔血于离心管中，然后加入25mL的去离子水稀释，在同样的波长下测其吸光度(A 空白)。具体的BCI计算方法如下公式：
BCI＝A 样品/A 空白×100％
每组平行实验5次，实验结果取平均值
Figure PCTCN2022070418-appb-000001
并且用SPSS19.0对实验所得结果进行单因素ANOVA方差分析。p<0.05和p<0.01分别表示所得结果具有显著性和极显著性统计学差异，结果见图2。
凝血指数BCI值越小，则表示材料的凝血效果越好。从图2中可以看出不同受试材料的凝血指数结果。市售的云南白药，又称“曲焕章百宝丹”，俗称“白药”，是我国的活血化瘀中成药，对跌打损伤、创伤出血有很好的疗效，作为阳性对照。实施例1～4组的BCI值都远低于阳性对照组，具有极显著性差异(p<0.01)，表现出强的促凝血效果，尤以实施例1的BCI值最低。虽然对比例1和对比例2的BCI值与阳性对照组无显著差异，但对比例1～4组的BCI值都远高于实施例1～4组，表明对比例1-4组的促凝血效果远不如实施例1～4组。
效果例2
红细胞吸附试验：
将新鲜的抗凝全血以2000rpm离心15min，收集下层红细胞，用磷酸盐溶液(PBS，pH＝7.4)稀释到10％浓度，备用。分别将实施例1～4、对比例1～4制备得到的复合材料用手术刀切成0.4×0.4cm的小块，加入500μL红细胞溶液，37℃下温育1h，取出样品，在PBS中浸洗2次，以去除未粘附的红细胞，不加复合材料的红细胞溶液为对照组。
(1)将样品置于1％Triton-X100溶液中，37℃下放置1h，以裂解粘附的红细胞，540nm处测其吸光度，不加复合材料的红细胞溶液为对照组。
(2)将吸附红细胞的复合材料(实施例1)放入2.5％戊二醛中固定30min，依次用40％、60％、80％、100％的乙醇脱水15min，自然晾干， 然后喷金制样，扫描电镜观察并拍照记录，结果见图4。
Figure PCTCN2022070418-appb-000002
式中，A 样品为各样品组的吸光度值，A 对照为不加样品对照组的吸光度值。
材料对红细胞的吸附率越高，则表示材料的凝血效果越好。从图3中可以看出为不同受试材料红细胞吸附率的结果。实施例1～4组的红细胞吸附率都远高于对比例1-4组，具有极显著性差异(p<0.01)，表现出强的凝血效果，尤以实施例1效果最佳。这些结果表明，实施例1-4的复合材料能够利用壳聚糖基-Fe 2+配合物与血红细胞的血红蛋白产生交叉配位，促进红细胞吸附、聚集；此外，壳聚糖质子化氨基所携带的正电荷吸引更多带负电荷的红细胞，对红细胞的吸附、聚集产生协同效应，促使大量的红细胞快速凝集形成血块。
从图4中可以看出，实施例1的复合材料显著性地促进红细胞吸附、聚集，从而加速血液凝固。
效果例3
溶血率试验：
对制备的复合材料进行了溶血率试验。评价参考国家标准GB/T14233.2-2005医用输液、输血、注射器具检验方法，方法如下：
取新鲜抗凝兔血，在4℃2000rpm离心15min，移去上清液，下层为红细胞，用磷酸盐溶液(PBS，pH＝7.4)将红细胞稀释至体积浓度为2％，得到红细胞悬液备用。将0.050g实施例1～4、对比例1～4制备得到的复合材料分别加入1.5mL离心管中，再加入500μL PBS溶液；37℃恒温水浴30min，再加入500μL红细胞悬液，轻轻摇匀，37℃恒温水浴培养1h，阳 性对照加入500μL蒸馏水，阴性对照组加入500μL PBS溶液，每组平行3次。将样品管在4℃2000rpm离心10min，取上清液于孔板中，540nm处酶标仪测定其吸光度，其中阴性对照组的吸光度值不应大于0.03，阳性对照组的吸光度范围应为0.80±0.20，否则重新实验。溶血率计算公式如下：
Figure PCTCN2022070418-appb-000003
式中：A为样品吸光度；A 0为阴性组吸光度；A 100为阳性组吸光度。
溶血率试验是通过吸光度测定法检测材料与新鲜抗凝血作用后红细胞释放的血红蛋白量来评价材料对血液的安全性。按本试验检验溶血率时，国标中规定溶血率小于5％判定合格。
从图5可以看出，除了对比例4外，实施例1～4和对比例1～3的复合材料溶血率都小于5％，符合国标GB/T 14233.2-2005中生物材料的合格判定指标，满足生物材料对溶血性的要求，即说明制备的复合材料具有良好的血液相容性，生物安全性高，且本发明制备的复合材料可快速吸液溶胀，形成凝胶，具有一定体积，不易进入血液引起血栓风险。
以上所述的实施例仅是对本发明的优选方式进行描述，并非对本发明的范围进行限定，在不脱离本发明设计精神的前提下，本领域普通技术人员对本发明的技术方案做出的各种变形和改进，均应落入本发明权利要求书确定的保护范围内。

Claims (10)

  1. 一种具有快速凝血作用的复合材料，其特征在于，以质量份数计，包括以下原料：壳聚糖8～20份、水杨醛7～14份、2-氨基异烟酸8～16份、硫酸亚铁1.5～3.5份、多巴3～5份和磷酸氢二钠10～15份。
  2. 根据权利要求1所述的具有快速凝血作用的复合材料，其特征在于，以质量份数计，包括以下原料：壳聚糖14份、水杨醛10份、2-氨基异烟酸12份、硫酸亚铁2.5份、多巴4份和磷酸氢二钠13份。
  3. 根据权利要求1所述的具有快速凝血作用的复合材料，其特征在于，所述壳聚糖的分子量为50～300KDa，脱乙酰度为80～95％。
  4. 根据权利要求1所述的具有快速凝血作用的复合材料，其特征在于，所述壳聚糖的分子量为100KDa，脱乙酰度为90％。
  5. 一种根据权利要求1～4任一项所述的具有快速凝血作用的复合材料的制备方法，其特征在于，包括以下步骤：
    (1)在壳聚糖溶液中加入2-氨基异烟酸和偶联剂混合的乙醇溶液，搅拌反应得到壳聚糖-氨基异烟酸接枝物；所述偶联剂为1-乙基-3-(3-二甲基氨基丙基)碳二亚胺盐酸盐和N-羟基琥珀酰亚胺；
    (2)将步骤(1)制备得到的壳聚糖-氨基异烟酸接枝物溶解后，调pH值至5.0～7.0，然后加入水杨醛，搅拌一定时间后依次加入硫酸亚铁、多巴和磷酸氢二钠混合均匀后，避光静置，得到所述具有快速凝血作用的复合材料。
  6. 根据权利要求5所述的具有快速凝血作用的复合材料的制备方法，其特征在于，所述壳聚糖溶液中的溶剂为醋酸水溶液，壳聚糖的质量分数为2％～5％。
  7. 根据权利要求6所述的具有快速凝血作用的复合材料的制备方法，其特征在于，所述醋酸水溶液中醋酸的质量分数为2％。
  8. 根据权利要求5所述的具有快速凝血作用的复合材料的制备方法，其特征在于，所述1-乙基-3-(3-二甲基氨基丙基)碳二亚胺盐酸盐和N-羟基琥珀酰亚胺的摩尔比为5：1。
  9. 根据权利要求5所述的具有快速凝血作用的复合材料的制备方法，其特征在于，步骤(1)所述乙醇溶液中2-氨基异烟酸和1-乙基-3-(3-二甲基氨基丙基)碳二亚胺盐酸盐的摩尔比为2：3。
  10. 根据权利要求5所述的具有快速凝血作用的复合材料的制备方法，其特征在于，步骤(1)中所述搅拌反应的时间为6～8h；步骤(2)中所述pH值为5.5，调节pH值所用的试剂为0.1mol/L的HCl溶液，搅拌时间为4～6h。
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116099035A (zh) * 2022-12-29 2023-05-12 广东海洋大学 一种复合抗菌水凝胶敷料及其制备方法与应用

Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN113769150B (zh) * 2021-10-22 2022-05-17 广东海洋大学 一种具有快速凝血作用的复合材料及其制备方法
CN115836983B (zh) * 2023-02-26 2023-07-14 广州善元堂健康科技股份有限公司 一种含有改性壳聚糖的润肤护甲组合物及其制备方法

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103145738A (zh) * 2005-08-04 2013-06-12 西特里斯药业公司 作为SIRTUIN调节化合物的咪唑并[2,1-b]噻唑衍生物
CN111870731A (zh) * 2020-08-19 2020-11-03 明光市铭垚凹凸棒产业科技有限公司 一种复合海绵及其制备方法和应用
CN113769150A (zh) * 2021-10-22 2021-12-10 广东海洋大学 一种具有快速凝血作用的复合材料及其制备方法

Family Cites Families (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR101103423B1 (ko) * 2009-09-04 2012-01-06 아주대학교산학협력단 생체 주입형 조직 접착성 하이드로젤 및 이의 생의학적 용도
CN101791425B (zh) * 2010-03-30 2013-04-10 赵雪林 用于制备医用创伤敷料的抗菌促愈凝胶材料及其制备方法
WO2012143788A1 (en) * 2011-04-20 2012-10-26 Diabetes Science International Foundation Wound dressing comprising of silver chitosan
CN111234267B (zh) * 2020-03-25 2021-12-07 西安交通大学第二附属医院 一种导电光热自愈合复合水凝胶敷料及制备方法和应用
CN111777773B (zh) * 2020-07-01 2021-04-30 广东海洋大学 一种儿茶酚功能化壳聚糖/牡蛎肽温敏水凝胶的制备方法、产品及应用
CN112791223B (zh) * 2020-12-30 2022-02-25 河南亚都实业有限公司 一种抗菌壳聚糖基止血贴
CN112972749B (zh) * 2021-03-05 2022-04-12 武汉纺织大学 基于壳聚糖纤维的高效止血材料及其制备方法

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103145738A (zh) * 2005-08-04 2013-06-12 西特里斯药业公司 作为SIRTUIN调节化合物的咪唑并[2,1-b]噻唑衍生物
CN111870731A (zh) * 2020-08-19 2020-11-03 明光市铭垚凹凸棒产业科技有限公司 一种复合海绵及其制备方法和应用
CN113769150A (zh) * 2021-10-22 2021-12-10 广东海洋大学 一种具有快速凝血作用的复合材料及其制备方法

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
GUAN JING, HE QIN, WANG XIAO HUI, HUANG SHU JIE, LI ZHI HONG, LIU CHANG JUN, LI FAN, SHU ZHAN: "Preparation and Blood Coagulation Property of Ortho Hydroxytoluene Chitosan", MATERIALS SCIENCE FORUM, TRANS TECH PUBLICATIONS LTD, vol. 893, 1 January 2017 (2017-01-01), pages 62 - 68, XP093057705, DOI: 10.4028/www.scientific.net/MSF.893.62 *
JIANG YUEXIA, ET AL.: "Basis Properties and safety Assessment of DOPA-Chitosan Hemostatic Sponge", JOURNAL OF QIQIHAR MEDICAL COLLEGE, XX, XX, vol. 34, no. 16, 31 December 2013 (2013-12-31), xx , pages 2341 - 2342, XP093057704, ISSN: 1002-1256 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116099035A (zh) * 2022-12-29 2023-05-12 广东海洋大学 一种复合抗菌水凝胶敷料及其制备方法与应用
CN116099035B (zh) * 2022-12-29 2023-09-08 广东海洋大学 一种复合抗菌水凝胶敷料及其制备方法与应用

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