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WO2019107887A2 - Composition pour hémostase et récipient comprenant celle-ci - Google Patents

Composition pour hémostase et récipient comprenant celle-ci Download PDF

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Publication number
WO2019107887A2
WO2019107887A2 PCT/KR2018/014752 KR2018014752W WO2019107887A2 WO 2019107887 A2 WO2019107887 A2 WO 2019107887A2 KR 2018014752 W KR2018014752 W KR 2018014752W WO 2019107887 A2 WO2019107887 A2 WO 2019107887A2
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Prior art keywords
collagen
thrombin
hemostasis
container
composition
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PCT/KR2018/014752
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English (en)
Korean (ko)
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WO2019107887A3 (fr
Inventor
박시내
배상희
고재형
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Dalim Tissen Inc
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Dalim Tissen Inc
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Publication date
Priority claimed from KR1020180127183A external-priority patent/KR101989054B1/ko
Application filed by Dalim Tissen Inc filed Critical Dalim Tissen Inc
Priority to JP2020549529A priority Critical patent/JP7223447B2/ja
Priority to US16/767,912 priority patent/US11628236B2/en
Priority to EP18883961.7A priority patent/EP3718578B1/fr
Priority to CN201880076999.4A priority patent/CN111465417B/zh
Publication of WO2019107887A2 publication Critical patent/WO2019107887A2/fr
Publication of WO2019107887A3 publication Critical patent/WO2019107887A3/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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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
    • A61L24/00—Surgical adhesives or cements; Adhesives for colostomy devices
    • 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
    • A61L24/04—Surgical adhesives or cements; Adhesives for colostomy devices containing macromolecular 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
    • A61L24/00—Surgical adhesives or cements; Adhesives for colostomy devices
    • A61L24/04—Surgical adhesives or cements; Adhesives for colostomy devices containing macromolecular materials
    • A61L24/10—Polypeptides; Proteins

Definitions

  • the present invention relates to a composition for hemostasis and a container containing the same.
  • tissue adhesives including sealants and hemostats
  • sealants and hemostats are growing rapidly.
  • tissue adhesives have been widely used as replacements for techniques such as suture, clip, and tampa, which have been used in surgical or medical operations.
  • tissue adhesives have many advantages such as quick adhesion time, easy use, and no need to be removed after the procedure, but they have a limitation in that adhesiveness is significantly lowered in the presence of low adhesiveness and tensile strength and moisture . Research has been continuing to overcome the limitations of such tissue adhesives.
  • Biocompatibility is required because the tissue adhesive is in direct contact with the tissue.
  • medical adhesive since it is usually used in vivo, when the adhesive flows into the body fluids and blood, the living body is directly involved. Therefore, there is no need to be toxic and harmful under more stringent conditions, and more strict biocompatibility and biodegradable material Is required.
  • Tissue adhesives that are currently commercialized and / or put to practical use are largely cyanoacrylate instant adhesives, fibrin glue, gelatin glue, and polyurethane-based adhesives. Cyanoacrylate instant adhesives have recently attracted attention in the research of instant adhesives having high functionality and high performance. In particular, medical instant adhesives for biocompatibility, flexibility and low toxicity biological tissue sealing are being actively developed in advanced countries because they can replace hemostatic and antibacterial effects as well as sutures.
  • Such cyanoacrylate-based tissue adhesives are currently commercially available as products such as Dermabond (Johnson & Johnson) and Indermil (US Surgical).
  • a cyanoacrylate-based tissue adhesive is a single substance, which is cured by moisture without an initiator at room temperature for a short time, has a transparent outer appearance and a large adhesive strength, but has a disadvantage that it is weak to impact and low in heat resistance. Also, its toxicity is so severe that it is rarely used at present and it is partially used in clinics in countries other than the United States, but its use is restricted due to some organizational toxicity and vulnerability.
  • Fibrin glue was first approved for use in cardiac surgery by the US FDA in 1998. Since then, fibrin tissue adhesives have been actively researched and developed, and products such as Tisseel VH (Baxer) and Evicel TM (Johnson & Johnson) are commercially available. Fibrin-based tissue adhesives, along with cyanoacrylate series, account for most of the tissue adhesive market. Fibrin tissue adhesives have been applied for the substitution or supplementation of sutures by applying fibrinogen, thrombin, calcium chloride and an enzyme inhibitor as a tissue adhesive to the peripheral nerve suture and microvascular suture using the crosslinking reaction of fibrin .
  • fibrin glue adhesives have the advantages of fast adhesion without being affected by the moisture at the adhesion site, free of platelet and coagulation trouble, and excellent in biocompatibility. However, it has a weak adhesive strength, a high rate of biodegradation, and a risk of infection of the blood.
  • gelatin glue has been developed by crosslinking with gelatin-resorcinol-formalin (GRF) as a tissue adhesive derived from living body.
  • GRF gelatin-resorcinol-formalin
  • tissue adhesives such as gelatin-glutaraldehyde have been developed.
  • formalin and glutaraldehyde which are used as cross-linking agents, have a disadvantage in that they cause cross-linking reaction with proteins in vivo and cause tissue toxicity.
  • the polyurethane-based adhesive has been developed in the form of an elastic adhesive which maintains the flexibility of the joint after curing.
  • This adhesive absorbs water on the surface of living tissue to enhance adhesion with the tissue, reacts with water to cure within a few minutes, and has a flexibility.
  • the adhesive has the advantage of biodegrading the cured adhesive appropriately.
  • the aromatic diisocyanate which is a starting material for synthesis, is disadvantageous in that it is biocompatible.
  • collagen alone, such as 'Avitene' (Alcon) and 'Helitene' (Dahamed), is a non-blood drug or a very expensive, sole component that is used only as a hemostatic agent because it has no tissue adhesion effect.
  • Floseal which contains a gelatin matrix component
  • Boxter which contains a gelatin matrix component
  • It is a product that exhibits a hemostatic effect by adding thrombin mixed in a calcium chloride solution to a gelatin matrix. This can be used for bleeding that is not effectively controlled or can not be controlled by ligature or general procedures, especially during surgical procedures in various areas for areas other than the eye.
  • thrombin gelatin
  • calcium chloride solution It takes some time to mix the solutions in two steps rather than mixing them one at a time. Therefore, during surgical operation, there is no room for mixing according to the instructions of the complicated product in the urgent state of severe ligation and hemorrhage, and the process of progressing sequentially may be life-threatening There is a problem that is very troublesome to the user.
  • hemostasis products are applied to humans, they must provide the highest safety standards, storage stability, and simple methods of use for the quality of the final product and its components.
  • they should be made in a "ready-to-use" form and should be provided in an easily mixed state.
  • Patent Document 1 U.S. Patent No. 5464471
  • Patent Document 2 US Patent No. 5773033
  • Patent Document 3 US Patent 5605887
  • the present inventors have developed a composition for hemostasis comprising collagen, a stabilizer and thrombin and a container containing the hemostatic composition to solve the above-mentioned problems.
  • the present invention can be applied to patients suffering from visual disturbance by simple use,
  • the present inventors completed the present invention by confirming that the biodegradation rate is fast and the hemostatic effect is excellent without the problem of blood infection.
  • an object of the present invention is to provide a composition for hemostasis comprising collagen, a stabilizer and thrombin, and a container containing the same.
  • Another object of the present invention is to provide a kit comprising the first container filled with the hemostatic composition and the second container filled with the diluent.
  • Still another object of the present invention is to provide a hydrogel produced using the hemostasis kit.
  • a composition for hemostasis comprising collagen, a stabilizer, and thrombin, wherein a stabilizer is disposed between the collagen and thrombin to form a hemostatic composition for collagen and thrombin .
  • the collagen may be crosslinked collagen.
  • the molecular weight of the crosslinked collagen may be 100,000 to 1,000,000 Dalton.
  • the collagen may be contained in an amount of 40 to 97% by weight based on the total weight of the hemostatic composition.
  • the stabilizer may be at least one selected from the group consisting of human serum albumin, mannitol, sodium acetate (C 2 H 3 NaO 2 ), sucrose, trehalose, sorbitol and glycine.
  • the stabilizing agent may be mannitol.
  • the stabilizer may be included in an amount of 1 to 30% by weight based on the total weight of the hemostatic composition.
  • the thrombin may be contained in an amount of 2 to 50% by weight based on the total weight of the hemostatic composition.
  • the hemostatic composition may be in powder form.
  • a container comprising the hemostatic composition.
  • a hemostatic composition comprising: a first container filled with a composition for hemostasis comprising collagen, a stabilizer and thrombin; And a second container filled with a diluent, wherein the first container is provided with a stabilizer between collagen and thrombin to provide a hemostasis kit in which collagen and thrombin are separated from each other.
  • the diluent may be at least one selected from the group consisting of purified water, a calcium chloride (CaCl 2 ) solution, a sodium chloride (NaCl) solution, a human serum albumin, and a sodium acetate (C 2 H 3 NaO 2 ) solution.
  • the diluent is a calcium chloride solution
  • the calcium chloride may be contained in an amount of 0.001 to 30% by weight based on the total weight of the diluent.
  • the first container and the second container may be bonded to each other, and the hemostatic composition and the diluent may be mixed to form the hydrogel.
  • the average pore size of the hydrogel may be from 50 ⁇ to 200 ⁇ .
  • the pH of the hydrogel may be from 6 to 8.
  • the first container may be filled with thrombin, a stabilizer, and collagen in this order.
  • composition for hemostasis of the hemostasis kit is the same as that described above, the following description will be omitted.
  • composition for hemostasis comprising the collagen, stabilizer and thrombin of the present invention is excellent in hemostatic effect.
  • the container containing the collagen, the stabilizer and the thrombin-containing composition of the present invention can be quickly and easily used, has a high bio-degradation rate without toxicity, no problems with blood infections, And its swelling rate is high, so it has excellent hemostatic effect.
  • the hemostatic composition of the present invention and the container containing the hemostatic composition can be used as a hemostasis kit.
  • Example 1 is an SEM image of each hydrogel prepared by mixing Example 2, Example 4, and Comparative Example 1 with a calcium chloride solution.
  • FIG. 2 is a graph obtained by measuring the respective expansion rates of crosslinked collagen and crosslinked gelatin.
  • FIG. 3 is a graph obtained by measuring the success rate of hemostasis (left) and the time taken for hemostasis (right) by treating Example 2 and Comparative Example 1 with a rabbit mesenteric shearing model, respectively.
  • Fig. 4 is an image of a tissue obtained by treating H & E staining of Example 2 and Comparative Example 1 on a porcine comparative vein injury model, respectively.
  • the present invention provides a composition for hemostasis comprising collagen, a stabilizer and thrombin, wherein a stabilizer is disposed between the collagen and thrombin to provide a composition for hemostasis wherein collagen and thrombin are separated from each other.
  • the collagen is contained in a composition for hemostasis, absorbs the blood at the bleeding site, and can attract platelets and cause a hemostatic effect.
  • the collagen is not bound to platelets in the absence of anything but binds to von Willebrand's factor (vWF), which acts like an adhesive, and binds to GPIb / IX complex, a membrane glycoprotein present on the cell surface of platelets And the first hemostasis can be performed.
  • vWF von Willebrand's factor
  • the collagen may be a protein that is treated by acid or alkali treatment of various animal tissues, or by treating with enzymes such as pepsin.
  • the collagen may be contained in an amount of 40 to 97% by weight based on the total weight of the hemostatic composition. If the content of the stabilizing agent is less than 40% by weight, the hemostatic effect may be very poor. If the content of the thrombin is more than 97% by weight, the thrombin and the stabilizer may be contained in a relatively small amount, have.
  • the collagen may be crosslinked collagen, and the crosslinked collagen preferably has a molecular weight of 100,000 to 1,000,000 Dalton, but is not limited thereto.
  • the molecular weight of the crosslinked collagen is less than 100,000 Dalton, the blood absorption capacity is rapid but the blood coagulation is delayed and the hemostatic effect may be deteriorated. If the molecular weight exceeds 1,000,000 Dalton, the blood absorption capacity may be significantly decreased.
  • the cross-linked collagen may be prepared by a chemical cross-linking method, a physical cross-linking method, or a combination thereof, wherein the cross-linked collagen by chemical cross-linking is a self-crosslinking collagen using a cross-linking agent.
  • crosslinked collagen by physical crosslinking is cross-linked collagen due to dry heat treatment, ultraviolet irradiation, and gamma irradiation.
  • the crosslinking agent may be any one or more crosslinking agents selected from the group consisting of Formaldehyde, Glutaraldehyde, Carbodiimides (EDC), and Polyepoxy compounds. Preferably, it is not limited to Carbodiimides (EDC).
  • the crosslinked collagen has an excellent mechanical strength and can exhibit a physical compression hemostatic effect, thereby allowing hemostasis of a large amount of hemostasis.
  • the crosslinked collagen may be a collagen obtained by crosslinking the esterified collagen.
  • the crosslinked collagen comprises
  • Step S1) is a step for treating collagen to ethanol or methanol, dissolving collagen in ethanol or methanol to facilitate mixing of the cross-linking agent, and producing collagen solution.
  • the method may further include the step of treating collagen with an acidic solution of animal skin tissue to extract atelocollagen.
  • step S2) an acid is added to the collagen treated in step S1) to prepare a collagen solution having a pH of 2 to 4.
  • An acid is added to the collagen solution to make the collagen completely dissolved in ethanol or methanol .
  • the step S3) is a step of preparing the esterified collagen by making the collagen solution prepared in the step S2) into a neutral state and then centrifuging to obtain a non-helical structure composed of about 12 to 27 amino acids at both ends of the collagen Removing the telopeptide, and esterifying the telopeptide-removed portion.
  • telopeptide is known to be the main cause of immune reaction when collagen is injected in vivo. Therefore, when telopeptide is used as a raw material for medicines and the like, telopeptide is removed to avoid immune reaction, and then atelocollagen or esterified collagen is used .
  • step S3) the telopeptide is removed and the esterified collagen having an ester function is firstly prepared by treating the collagen solution, which is made into a neutral state, with pepsin enzyme, followed by separation of high purity esterified collagen using centrifugation .
  • the centrifugation is preferably performed at 1000 rpm to 30,000 rpm for 5 minutes to 3 hours so that only the esterified collagen can be cleaved, but is not limited thereto.
  • a sample containing the atelocollagen is introduced into the filtration module provided with the filtration membrane from the container by a pumping action by a pump, and a pressure of about 10 to 30 psi is applied to the filtration module .
  • the ultrafiltration process can be stopped.
  • the same amount of purified water as the filtered solution through the ultrafiltration process may be added to the residue returned to the vessel.
  • the dyestuff filtration process in the steps (f) and (g) is preferably performed at least five times.
  • esterified collagen can be obtained by esterification of atelocollagen through steps (a) to (h).
  • cross-linking agent in step S4) may be any one or more cross-linking agents selected from the group consisting of Formaldehyde, Glutaraldehyde, Carbodiimides (EDC) and Polyepoxy compounds as described above, and a detailed description thereof will be omitted.
  • the step S5) is a step for dispersing the crosslinked collagen prepared in the step S4) into purified water and freeze-drying the crosslinked collagen and removing the remaining crosslinking agent and telopeptide-free collagen.
  • the crosslinked collagen prepared in step S4) is dispersed in the purified water to dissolve the collagen in which the trace amount of the crosslinking agent and the telopeptide have not been removed in the purified water, and the solution is freeze- .
  • freeze-drying may be a conventional method used in the technical field.
  • the stabilizing agent is included in the composition for hemostasis so that the collagen and the thrombin are interposed between the collagen and the thrombin to prevent the collagen and the thrombin from mixing with each other to maintain the intrinsic properties of the proteins. Therefore, by incorporating the stabilizing agent into the hemostatic composition, it is possible not only to separate collagen and thrombin having a strong hemostatic effect, but also to ensure the chemical stability and biological activity of the hemostatic composition with an inactive material.
  • the stabilizer may be at least one selected from the group consisting of human serum albumin, mannitol, sodium acetate (C 2 H 3 NaO 2 ), sucrose, trehalose, sorbitol and glycine.
  • the stabilizer includes at least one selected from the group consisting of albumin, mannitol, and sodium acetate, so that the chemical stability and biological activity of thrombin can be maintained.
  • the stabilizer includes at least one selected from the group consisting of mannitol, sucrose, trehalose, sorbitol, and glycine, so that prolongation of the hemostasis time can be suppressed and blood coagulation activity can be maintained.
  • the stabilizer is preferably, but not limited to, mannitol.
  • the above-mentioned mannitol is white needle-shaped or columnar crystals widely present in plants such as mushroom and pomegranate roots, and is well dissolved in water and used as a laxative agent, and an alternative roll of glycerol is used.
  • the stabilizing agent may be contained in an amount of 1 to 30% by weight based on the total weight of the hemostatic composition. If it is contained in an amount less than 1% by weight, it may be difficult to separate the collagen and thrombin, so that the chemical stability may deteriorate. If the collagen and thrombin are contained in an amount exceeding 30% by weight, the collagen and thrombin are less contained, Can be lowered.
  • the thrombin is contained in a composition for hemostasis, which acts as a strong hemostasis factor and acts as a catalyst in the reaction of converting the soluble fibrinogen in the blood into the insoluble fibrin by participating in blood coagulation.
  • the thrombin can induce platelet activation in the process of forming fibrin by the activation of the blood coagulation factor, which is a secondary hemostatic process.
  • the thrombin may be contained in an amount of 2 to 50% by weight based on the total weight of the hemostatic composition. If it is contained in an amount of less than 3% by weight, the hemostatic effect may be very poor. If it is contained in an amount of more than 50% by weight, the collagen and the stabilizer may be contained in a relatively small amount, have.
  • the composition for hemostasis may be in the form of powder, and the composition for hemostasis, which is a dry powder form, is preferably in powder form so as not to easily deteriorate into a liquid state by the external environment and moisture.
  • the composition for hemostasis is in the form of a powder, which may have chemical stability depending on each component, and may be easily maintained in a non-mixed state. It also has the advantage of convenient handling and storage.
  • a container containing the hemostatic composition can be provided.
  • the container containing the hemostatic composition is filled with collagen, stabilizer, and thrombin having a hemostatic effect, it is possible to apply the hemostatic composition to a patient suffering from visual disturbance. Especially, it can be applied to the hemorrhage site by mixing with the dilute solution only when the severe ligation and hemorrhage occurs during the surgical operation, and it can be easily used even if it is not a professional technician It is very simple to use.
  • a hemostatic composition comprising: a first container filled with a composition for hemostasis comprising collagen, a stabilizer and thrombin; And a second container filled with a diluent, wherein the first container is provided with a stabilizer between collagen and thrombin to provide a hemostasis kit in which collagen and thrombin are separated from each other.
  • hemostasis kit uses the hemostatic composition as described above, collagen, stabilizer, and thrombin are also the same, and a detailed description thereof will be omitted.
  • the first container is filled with a composition for hemostasis, and the stabilizer is filled between the collagen and the thrombin so that the order in which they are placed by the external physical force is not changed.
  • the collagen, the stabilizer, and the thrombin filled in the first container may be mixed with the diluent at a later time in order to change the flowability, as well as the rate of expansion and the rate of expansion. Accordingly, it is preferable that the first container is filled in the order of collagen, stabilizer, thrombin or thrombin, stabilizer, and collagen. More preferably, the order of thrombin, stabilizing agent and collagen is not limited thereto.
  • the second container is filled with a diluent, wherein the diluent is used to dissolve or disperse the hemostatic composition of the first container and may be used as a solvent or a dispersion medium.
  • the diluent may be at least one selected from the group consisting of purified water, a calcium chloride (CaCl 2 ) solution, a sodium chloride (NaCl) solution, a human serum albumin, and a sodium acetate (C 2 H 3 NaO 2 ) solution.
  • a diluent is used to dissolve or disperse the hemostatic composition of the first container and may be used as a solvent or a dispersion medium.
  • it may be at least one selected from the group consisting of purified water, a calcium chloride (CaCl 2 ) solution, a sodium chloride (NaCl) solution, a human serum albumin, and a sodium acetate (C 2 H 3 NaO 2 ) solution.
  • CaCl 2 calcium chlor
  • the calcium chloride may be contained in an amount of 0.001 to 30% by weight based on the total weight of the diluent, and is not particularly limited as long as it is in the range of mixing well with the hemostatic composition.
  • the shape and material of the first container or the second container are not particularly limited and may be those conventionally used in the fields of medicine and biotechnology.
  • the shape of the first container or the second container is a syringe
  • the material of the first container or the second container is a material that does not chemically react with any one selected from collagen, stabilizer, and thrombin.
  • the first container and the second container may be bonded to each other, and the hemostatic composition and the diluent may be mixed to form a hydrogel.
  • the front end of the first container may include a fastening protrusion such that the first container and the second container can engage with each other and the second container may include a fastening groove to engage with the fastening protrusion at the front end of the first container .
  • the hydrogel is porous, so that it has a high blood absorption rate when used as a hemostatic agent, maintains the shape of the hydrogel even after absorption of blood, and does not deteriorate the hemostatic effect, thereby exhibiting an excellent hemostatic effect.
  • the average pore size of the hydrogel may be 50 ⁇ to 200 ⁇ . If the average pore size is less than 50 ⁇ , the blood absorption rate is reduced. If the average pore size is more than 200 ⁇ , the hemostatic effect may be deteriorated because the hydrogel can not be maintained after absorbing the blood.
  • the hydrogel-forming time may be as short as 5 seconds to 5 minutes, so as to prevent serious ligation and bleeding. Further, by forming the hydrogel in a short time, the user can easily use it.
  • the hydrogel is mixed with a composition for hemostasis and a diluted solution to have a neutral pH, so that biocompatibility and biosafety can be ensured when the hydrogel is applied to a living body.
  • a neutral pH so that biocompatibility and biosafety can be ensured when the hydrogel is applied to a living body.
  • the pH of the hydrogel may be 6 to 8.
  • Esterified collagen powder was prepared by the following procedure.
  • Crosslinked collagen powder was prepared by the following procedure.
  • EDC (1-ethyl-3- (3-dimethylaminopropyl) -carbodiimide) was added to the well-mixed esterified collagen solution.
  • the above powder was sieved to select particles of 200 ⁇ 355 ⁇ m size.
  • Gelatin powder (FloSeal Hemostatic Matirx, Baxter AG, USA), which is commercially available as a hemostatic agent instead of the esterified collagen powder, was used to obtain crosslinked gelatin.
  • the concentration of collagen, mannitol and thrombin in the composition for hemostasis was determined according to the order in which the calcium chloride solution (concentration) was diluted. At this time, mannitol and thrombin were powdered products commercially available, and the esterified collagen obtained from Preparation Example 1 was used, and the crosslinked collagen obtained from Preparation Example 2 was used. The cross-linked gelatin obtained from Preparation Example 3 was used for flowability comparative analysis.
  • the force for injecting the calcium chloride solution into the syringe filled with the hemostatic composition is opposite to the force for injecting the hemostatic composition into the syringe filled with the calcium chloride solution. It is relatively large. We measured the force (N1, N3, N5, N7) required to inject the calcium chloride solution into the syringe filled with the hemostatic composition, and confirmed the difference in flowability during mixing.
  • Example 1 Number of mixing times (times) Maximum load (N) Example 1
  • Example 2 Example 3
  • Example 4 Comparative Example 1 N1 4.36 3.94 22.57 21.84 19.43 N3 10.83 7.05 5.12 13.1 8.39 N5 10.18 6.52 5.83 21.98 61.07 N7 9.97 7.04 22.32 7.69 22.16 Average 8.84 6.14 13.96 16.15 27.76
  • Examples 1 to 4 were lower in the magnitude of the force required for mixing, that is, the magnitude of the pressure generated during mixing, as compared with Comparative Example 1.
  • the pressure generated during mixing was small. Therefore, it was confirmed that thrombin and mannitol were placed at the inlet side of the syringe to reduce the pressure required for raw material mixing from the beginning of the mixing and to help the raw materials to be well mixed.
  • Comparative Example 1 in which crosslinked gelatin was packed had no uniform pressure pattern that occurred in Examples 1 to 4.
  • the cross-linked gelatin exhibited a higher overall pressure, including the average, as compared with Example 1, which was filled with cross-linked collagen, and Examples 2 to 4, while the pressure at the time of mixing was about 3 times as high as about 10 times Times as much.
  • Example 2 and Comparative Example 1 were prepared according to the method described in Table 1 of Experimental Example 1, in order to confirm the rate of expansion of the composition for hemostasis according to the present invention after mixing with the diluted solution.
  • the particle areas of the prepared Example 2 and Comparative Example 1 were measured. Then, each sample was mixed with calcium chloride solution, and the particle area was measured at intervals of 1, 2, 3, 5, 10, 20, and 30 minutes, and the rate of area change was calculated to calculate the expansion rate. The results are shown in Fig.
  • Example 2 and Comparative Example 1 expanded swiftly within the first minute and continued to expand for 2, 3, 5, and 10 minutes, but the swelling rate became saturated after 10 minutes .
  • Example 2 containing crosslinked collagen the swelling rate was faster than that of Comparative Example 1 and the area change was large, so that the crosslinked collagen performed physical pressing action due to swelling of the particles within the same time as the crosslinked gelatin in the same time And it can be a great help for the hemostatic effect.
  • the biodegradability and the degree of degradation of the collagen contained in the hemostatic composition according to the present invention were measured and the degradation degree test was conducted in order to confirm whether the biodegradability or the degradability of the biodegradable collagen was low.
  • the esterified collagen obtained from Preparation Example 1 the crosslinked collagen obtained from Preparation Example 2 and the crosslinked gelatin obtained from Preparation Example 3 were prepared, and the weight of each sample was measured.
  • each sample was placed in a micro-centrifuge tube and filled with 1-2 mL of 1XPBS buffer containing 25 units / mL of collagenase.
  • the tube containing the sample was placed in a 37 ° C water bath and allowed to stand for 24 to 72 hours. After 24, 48, and 72 hours, samples were taken out, lyophilized and weighed.
  • the weight of the sample measured before and after the decomposition test was calculated using the following equation. The results are shown in Table 3.
  • Remaining weight ratio (%) degradation degree Dry weight / resolution of sample after test Dry weight of sample before test * 100
  • crosslinked collagen and the esterified collagen were confirmed to be completely decomposed by the collagenase in 72 hours.
  • the crosslinked gelatin had a residual weight ratio of 3.3 ⁇ 1.2% after 72 hours, and the sample was not completely decomposed by the collagenase. Therefore, it was confirmed that cross-linked collagen and esterified collagen had faster degradability than the crosslinked gelatin under the same conditions, and it was confirmed that the biodegradability was high and the risk of infection was very low.
  • Example 2 and Comparative Example 1 were prepared in the same manner as Experimental Example 1 to confirm the hemostatic effect of the composition for hemostasis according to the present invention. Comparisons of hemostatic performance were made using a rabbit mesenteric shear model. Hemorrhage was induced by cutting the mesentery and then applying Hemorrhage for 30 seconds after applying Example 2 or Comparative Example 1. Hemorrhage was checked for 30 seconds and hemorrhagic compression was repeated for 30 seconds for hemorrhage. The probability of successful hemostasis in 90 seconds and the time taken for hemostasis were measured and the results are shown in FIG.
  • Example 2 15 rabbits succeeded in hemostasis within 90 seconds, whereas in Comparative Example 1, only 10 of 15 rabbits completed hemostasis within 90 seconds. Also, in the time taken for hemostasis, Example 2 was 64 ⁇ 4.957 seconds, whereas Comparative Example 1 was 86 ⁇ 8.718 seconds, and t-test showed statistical significance (p ⁇ 0.05). Accordingly, it has been confirmed that when the composition for hemostasis according to the present invention is used, the hemostasis rate is high and the hemostasis time is shortened.
  • EXPERIMENTAL EXAMPLE 6 Comparison of Degradation Intensity and Inflammatory Response in a Porcine Comparative Venous Injury Model
  • Example 2 and Comparative Example 1 were prepared in the same manner as Experimental Example 1 to confirm the decomposition degree and inflammation reaction degree of the hemostatic composition according to the present invention. Hemorrhage was induced by partial incision of the superior vein of the pig, and hemostasis was achieved by applying Example 2 or Comparative Example 1. One week after the experiment, the pigs were sacrificed and the specimens were collected and the amount of hemostatic composition remaining and inflammatory response were compared through H & E staining. The results are shown in Table 4, and the image of the tissue after H & E staining is shown in FIG.
  • Example 2 Comparative Example 1 Inflammatory reaction + ++ Residue + ++
  • FIG. 4 is an image of the tissue after H & E staining of Example 2 and Comparative Example 1, which shows that the amount of the residue of Example 2 is relatively smaller than the amount of the residue of Comparative Example 1, The number of inflammatory cells is smaller than that of Comparative Example 1.
  • the composition having a high resolution is highly biodegradable and the infection risk such as inflammation reaction is low. Accordingly, when the composition for hemostasis according to the present invention is used, it is possible to improve the biocompatibility, reduce the side effect, and reduce the risk of newly occurring infection due to hemostatic agent.
  • Example 2 and Comparative Example 1 were prepared in the same manner as in Experimental Example 1 to confirm the biocompatibility of the composition for hemostasis according to the present invention.
  • a portion of the kidney of the white paper was excised to induce hemorrhage. Then, Example 2 or Comparative Example 1 was applied to the cut surface for hemostasis.
  • the samples were sacrificed at the sacrifice of the white paper, and the biocompatibility according to the difference of the hemostatic composition was confirmed by pathological examination. The results are shown in Table 5 below.
  • Example 2 Comparative Example 1 Object 1 Object 2 Object 1 Object 2 Renal cortex Tissue alteration, bleeding 0 0 2 One Epithelial regressive changes (edema, necrosis) 0 0 3 One Glomerular necrosis 0 0 0 0 Glomerular hyaline microthrombi 0 0 0 0 Inflammation 4 4 5 5 Extraglomerular hemorrhage 0 0 0 0 0 0 Intraglomerular hemorrhage 0 0 0 0 0 Fibroblast activation 2 2 3 3 Fibrosis 2 One 3 3 Fistula 0 0 0 0 0 Microvascular proliferation 2 2 2 One Hyaline in the tubules 2 One 2 One Interstitial hemorrhage One 0 2 3 Polymorphonuclears 3 3 2 2 Collagen scar 0 0 0 0 Extra fat tissue Granuloma / inflammation around kidney 2 4 4 5 Destruction score 18 17 28 25
  • tissue deformation and hemorrhage were observed in the rats subjected to the Comparative Example 1, but no tissue deformation or postoperative bleeding was observed in the rats treated with the Example 2.
  • Fibroblast activity or fibrosis caused by fibroblast proliferation was larger in the subject to which Comparative Example 1 was applied, and intercellular hemorrhage was also larger in Comparative Example 1.
  • granuloma and inflammatory reaction were significantly increased in the adipose tissue around the renal resection surface of Comparative Example 1.
  • Example 2 On the other hand, the destruction score, which is an indicator of pathological harm, was found to be 17.5 in Example 2 and 26.5 in Comparative Example 1. Thus, it can be seen that the biocompatibility of Example 2 is superior to that of Comparative Example 1.

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  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
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  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

La présente invention concerne une composition pour hémostase, comprenant du collagène, un stabilisant et de la thrombine, et un récipient comprenant celle-ci. La composition pour hémostase et le récipient la comprenant peuvent être appliqués à des patients souffrant d'hémorragie qui sont engagés dans une course contre la montre par un procédé d'utilisation simple, et peuvent être utilisés en tant qu'agent hémostatique car n'ayant pas de toxicité, n'ayant pas de problèmes vis-à-vis des infections sanguines, ayant un taux de biodégradation rapide, et ayant un excellent effet hémostatique.
PCT/KR2018/014752 2017-11-28 2018-11-28 Composition pour hémostase et récipient comprenant celle-ci Ceased WO2019107887A2 (fr)

Priority Applications (4)

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JP2020549529A JP7223447B2 (ja) 2017-11-28 2018-11-28 止血用組成物及びこれを含む容器
US16/767,912 US11628236B2 (en) 2017-11-28 2018-11-28 Composition for hemostasis and container comprising same
EP18883961.7A EP3718578B1 (fr) 2017-11-28 2018-11-28 Composition pour hémostase et récipient comprenant celle-ci
CN201880076999.4A CN111465417B (zh) 2017-11-28 2018-11-28 止血用组合物以及包含上述止血用组合物的容器

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KR1020180127183A KR101989054B1 (ko) 2017-11-28 2018-10-24 지혈용 조성물 및 이를 포함하는 용기

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EP3903808B1 (fr) * 2019-05-28 2025-07-30 Dalim Tissen Co., Ltd. Composition hémostatique et contenant associé

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US5773033A (en) 1995-01-23 1998-06-30 The Regents Of The University Of California Fibrinogen/chitosan hemostatic agents
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