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WO1993009869A1 - Adsorbant pour le facteur viii de la coagulation sanguine - Google Patents

Adsorbant pour le facteur viii de la coagulation sanguine Download PDF

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Publication number
WO1993009869A1
WO1993009869A1 PCT/JP1992/001480 JP9201480W WO9309869A1 WO 1993009869 A1 WO1993009869 A1 WO 1993009869A1 JP 9201480 W JP9201480 W JP 9201480W WO 9309869 A1 WO9309869 A1 WO 9309869A1
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WO
WIPO (PCT)
Prior art keywords
adsorbent
factor viii
molecule
carrier
water
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/JP1992/001480
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English (en)
Japanese (ja)
Inventor
Naofumi Ito
Shigeru Igarashi
Tohru Suzuki
Tsugikazu Tomono
Sadayoshi Sekiguchi
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Asahi Kasei Corp
Asahi Chemical Industry Co Ltd
Original Assignee
Asahi Chemical Industry Co Ltd
Asahi Kasei Kogyo KK
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Filing date
Publication date
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Publication of WO1993009869A1 publication Critical patent/WO1993009869A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/745—Blood coagulation or fibrinolysis factors
    • C07K14/755—Factors VIII, e.g. factor VIII C (AHF), factor VIII Ag (VWF)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • A61P7/04—Antihaemorrhagics; Procoagulants; Haemostatic agents; Antifibrinolytic agents
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/22—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising organic material
    • B01J20/24—Naturally occurring macromolecular compounds, e.g. humic acids or their derivatives
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/22—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising organic material
    • B01J20/26—Synthetic macromolecular compounds
    • B01J20/261—Synthetic macromolecular compounds obtained by reactions only involving carbon to carbon unsaturated bonds
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/22—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising organic material
    • B01J20/26—Synthetic macromolecular compounds
    • B01J20/264—Synthetic macromolecular compounds derived from different types of monomers, e.g. linear or branched copolymers, block copolymers, graft copolymers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/28—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties
    • B01J20/28002—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties characterised by their physical properties
    • B01J20/28004—Sorbent size or size distribution, e.g. particle size
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/30—Processes for preparing, regenerating, or reactivating
    • B01J20/32—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating
    • B01J20/3202—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating characterised by the carrier, support or substrate used for impregnation or coating
    • B01J20/3206—Organic carriers, supports or substrates
    • B01J20/3208—Polymeric carriers, supports or substrates
    • B01J20/321—Polymeric carriers, supports or substrates consisting of a polymer obtained by reactions involving only carbon to carbon unsaturated bonds
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/30—Processes for preparing, regenerating, or reactivating
    • B01J20/32—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating
    • B01J20/3231—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating characterised by the coating or impregnating layer
    • B01J20/3242—Layers with a functional group, e.g. an affinity material, a ligand, a reactant or a complexing group
    • B01J20/3268—Macromolecular compounds
    • B01J20/327—Polymers obtained by reactions involving only carbon to carbon unsaturated bonds
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/30—Processes for preparing, regenerating, or reactivating
    • B01J20/32—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating
    • B01J20/3231—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating characterised by the coating or impregnating layer
    • B01J20/3242—Layers with a functional group, e.g. an affinity material, a ligand, a reactant or a complexing group
    • B01J20/3268—Macromolecular compounds
    • B01J20/3276—Copolymers

Definitions

  • the present invention relates to an adsorbent for blood coagulation factor VIII (hereinafter often simply referred to as "factor VIII"). More specifically, molecules having a polycation suitable for efficiently adsorbing and recovering factor VIII from a solution containing various plasma proteins including factor VIII are bound and immobilized on a water-insoluble carrier. And an adsorbent made of a composite.
  • Hemophilia A is an example of a congenital bleeding disorder. Hemophilia A is caused by a deficiency of a protein called factor VIII.In order for a person with hemophilia A to live as a healthy person, it is necessary to replace the lacking factor VIII. is there.
  • Factor V II concentrates are usually prepared by using a precipitate from plasma called cryoprecipitate as a raw material and removing contaminating proteins such as fibrinogen and fibronectin.
  • the recipe recipe can be obtained by centrifuging the precipitate generated by thawing fresh frozen plasma at a low temperature.
  • the factor V III contained in plasma is 100%
  • the recovery of factor V III in cryoprecipitate is as low as 40 to 50%.
  • cryoprecipitate contains a large amount of contaminating proteins, it is necessary to remove those contaminating proteins.
  • adsorption is performed by immobilizing a low-molecular-weight ligand such as a getylaminoethyl group on a water-insoluble carrier. Michalskieta 1.: Vox Sang, Vo 1.55, pp.
  • factor IX blood coagulation factor IX
  • factor VIII blood coagulation factor IX
  • Factor VIII is a protein with a molecular weight of about 300,000. It is known from the amino acid sequence analysis that there is a region rich in acidic amino acids [G.A.V ehareta 1.: Natur'e, Vol. 3.12, pp. 3 3 7-3 4 2 (1 998 4)]. Furthermore, it is known to exist in blood as a complex with a von Willebrand factor having a multimeric structure with a molecular weight of 1,000,000 to 2,000,000 (Z.M. R uggerieta 1.: Blood, Vol. 57, PP. 114-11 4 3 (1 98 1)).
  • the present inventors have found that in order to efficiently adsorb such a protein having a huge molecular weight, it has a large number of anion-exchange groups and a sufficient length.
  • an adsorbent having a substance capable of firmly adsorbing by interacting with the complex at a plurality of points is suitable.
  • the adsorbent consisting of a complex in which a molecule having a polycation is bonded to and immobilized on a water-insoluble carrier has an extremely specific adsorption to factor VIII. — It showed that it was able to adsorb and recover Factor VIII with high selectivity and high yield from solutions containing various plasma proteins including Factor VIII.
  • the present invention has been completed based on these findings.
  • one and primary object of the present invention is to provide a solution containing various plasma proteins, including factor VIII, from a solution containing factor VIII.
  • An adsorbent forest capable of directly and specifically adsorbing factor I, and capable of easily desorbing and recovering adsorbed factor VIII.
  • the molecular weight of a polycation-containing molecule is at least 15,500, unlike a complex formed by binding and immobilizing a molecule having a polycation to a water-insoluble carrier.
  • an adsorbent for blood coagulation factor VIII wherein the positive charge density of the complex of 1 milliliter is from leq to 200 eq.
  • the molecule having a polycation immobilized on the adsorbent of the present invention refers to a molecule having a molecular weight of at least 150000 and having a large number of anion exchange groups in the molecule.
  • the anion exchange group include a primary amino group, a secondary amino group, a tertiary amino group, a primary ammonium base and the like.
  • One of these anion exchange groups may be present alone in the molecule having a polycation, or two or more may be present.
  • Examples of such a molecule include a homopolymer of a basic vinyl monomer having an anion exchange group in a side chain, or a vinyl copolymer which can be copolymerized with the basic vinyl monomer. Copolymer with monomer Can be mentioned. Further, instead of the basic vinyl monomer, a vinyl monomer having a side chain to which an anion exchange group can be added after polymerization may be used. Further, there may be mentioned polypeptides containing basic amino acids such as lysine and arginine.
  • Examples of the basic vinyl monomer having an anion exchange group in the side chain include a vinyl monomer having a primary amino group, a secondary amino group, a tertiary amino group, and a quaternary ammonium base. Body included.
  • Examples of such a biel monomer include acrylic acid derivatives, methacrylic acid derivatives, acrylic acid amide derivatives, and metaacrylic acid amines each having an amino group.
  • Examples thereof include vinyl derivatives having a nitrogen-containing aromatic ring group such as a pyridyl group and an imidazolyl group in the side chain, and styrene derivatives substituted with an amino group.
  • dimethylaminoethyl acrylate dimethylaminoethyl acrylate, dimethylaminoethyl acrylate, dimethylaminopropyl acrylate, 3-dimethylaminopropyl 2—hydroxypropylamine Rate, 3—Jethylamino2—Hydroxypropylacrylate, N—Dimethylaminoethylacrylate, N—Nethylethylaminoethylacrylate, Dimethylaminoethylmethacrylate Rate, getylaminoethyl methacrylate, getylaminopropyl methacrylate, 3—dimethylamine 2—hydroxypropyl propylmethacrylate, 3—getylamine 1—2— histamine Dro kiss Mouth pill methacrylate, N-dimethylamino methacrylate amide, N-ethylaminoethyl methacrylate, and the following formula:
  • R is one CH 3 or one C 2 H 5 , X is (CH 2 ) n is an integer of 3)
  • CHz -CH Such a vinyl monomer having a side chain that can be aminated after polymerization can also be used.
  • Specific preferred examples of such a vinyl monomer include hydroxyxethyl acrylate and hydroxypropynoleacrylate. And hydroxypropyl methacrylate, and hydroxypropyl methacrylate.
  • the basic butyl monomer may be an acrylic acid derivative, a metaacrylic acid derivative, an acrylic acid amide derivative or a methacrylic acid derivative having an amino group.
  • vinyl acrylamide derivatives vinyl monomers that can be copolymerized with basic vinyl monomers include acrylic esters, metaacrylic esters, and N-alkylacrylic acids. Copolymers obtained using amides or N-alkyl methacrylates are preferred.
  • the molecular weight of the molecule having a polycation of the present invention is at least 15,000. When the molecular weight is smaller than this, sometimes factor VIII is an extremely large protein, so that it is difficult to expect a sufficient interaction between the molecule and factor VIII.
  • the positive charge density of the complex (1 mil / liter) composed of a molecule having a polycation and a water-insoluble carrier constituting the adsorbent is leq to 200 eq. .
  • the positive charge density of the composite constituting the adsorbent of the present invention is determined by the following method. Swollen with water Put 1 milliliter of adsorbed material into a plastic column, wash with 1N hydrochloric acid, water, and 1N aqueous sodium hydroxide in that order, and adjust the pH of the liquid falling from the cam to 6.5.
  • any of a hydrophilic carrier and a hydrophobic carrier can be used, but the hydrophobic carrier causes non-specific adsorption of proteins other than factor VIII to the carrier. Gives better results.
  • the water-insoluble carrier is not particularly limited as long as it has a functional group that can be used for immobilizing a molecule having a polycation by chemical bonding.
  • the chemical bond is preferably a covalent bond. Even when the water-insoluble carrier itself does not have such a functional group, one that can generate a functional group that can be used for immobilizing a molecule having a rebolication by appropriate chemical modification is used in the present invention. be able to.
  • any known shape such as a particle shape, a fibrous shape, a hollow fiber shape, and a membrane shape can be used, but handling as an adsorbent In view of the above, particles and fibrous materials are preferred.
  • examples of the particulate carrier include agarose ⁇ , dextran-based, cellulose-based and other natural polymer-based carriers, polyacrylamide-based, polyamide-based, and poly- amide-based carriers. Synthetic polymer-based carriers such as ester-based, polyurethane-based, and vinyl compound polymers can be mentioned.
  • a carrier having a porous structure the surface area of the carrier is increased, so that the molecule having immobilized polythione and the factor VIII can be easily brought into contact with each other. It gives good results with increased volume.
  • a polymer of a vinyl compound hydrophilized by one unit of a hydrophilic monomer can easily form a carrier having a porous structure, and can easily obtain a carrier having high physical strength.
  • a crosslinked copolymer having a bul alcohol unit often reduces the non-specific adsorption of a protein to a carrier, and gives a favorable result in many cases.
  • the fibrous carrier known fibers such as a cellulose-based fiber and a polyester-based fiber can be used.
  • the fiber diameter is preferably in the range of 0.02 denier or 10 denier.
  • the particulate support having a porous structure has an exclusion limit molecular weight in the range of 1,000 to 100,000,000, and an average particle size of 5 to 1,000 ⁇ m. ⁇ is preferred.
  • Binding and immobilization of polycation-containing molecules to water-insoluble carriers It is better to use a chemical bond, especially a covalent bond, for the conversion.
  • a known method for activating a water-insoluble carrier and a method for immobilizing a molecule having a polycation which are generally used in immobilized enzymes and affinity chromatography, can be used [Kenichi Kasai et al .: “Affinity mouth mouth one”, published by Tokyo Chemical Industry Co., Ltd. (1991):].
  • a molecule (spacer) having an arbitrary length can be introduced between the water-insoluble carrier and the molecule having a polycation.
  • the active group generated by activating the water-insoluble carrier is a group capable of reacting with a nucleophilic reactive group having an active hydrogen such as an amino group, a carboxyl group, a hydroxyl group, or a thiol group of a molecule having a polycation.
  • a nucleophilic reactive group having an active hydrogen such as an amino group, a carboxyl group, a hydroxyl group, or a thiol group of a molecule having a polycation.
  • epoxy group tresil group (tresy 1 gr)
  • the adsorbent of the present invention can be obtained by contacting a solution containing various plasma proteins containing Factor VII
  • Adsorb factor II Adsorb factor II.
  • the solution containing various plasma proteins including factor VIII at this time may be plasma or a product from plasma.
  • the products from plasma include the above-mentioned cryoprecipitate, The supernatant after separation and removal of the pithate, plasma from which some or all of the contaminant components have been removed by appropriate treatment with an ion exchange resin or the like can be mentioned.
  • the adsorbent of the present invention is capable of adsorbing factor VIII in plasma under various conditions, but usually adsorbs at normal temperature and normal pressure. It is preferred in terms of sex.
  • the temperature should be within the stable range of Factor V I I I, but is usually between 0 and 50 ° C.
  • the adsorption time is not particularly limited, it is usually in the range of 10 minutes to 2 hours, and an appropriate time can be selected depending on the temperature and the method of adsorption.
  • either a batch method or a column method can be used, but when a large amount of plasma is to be treated, the batch method is often preferred because it can be processed in a shorter time.
  • a crosslinked copolymer having a vinyl alcohol unit was synthesized as shown below.
  • 8 g and Yo Li becomes homogeneous mixture of polyvinyl alcohol 1 wt 0/0, V phosphate 2 hydrogen isocyanatomethyl Li um dihydrate 0.0 5 wt.
  • the water-insoluble carrier obtained in the above step was used as a water-insoluble carrier (swelled with acetate). 3 Milliliters were suspended in acetonitrile 2.1 milliliters. To the suspension, add Tresyl Mouth Ride 45 microliters and pyridine 135 microliters and shake at room temperature for 10 minutes to obtain the active group. An activated carrier having a tresyl group was obtained. (1) above, 2-copolymer of hydroxyethyl methyl acrylate and methylaminoethyl methacrylate 30 Omg 5
  • blood coagulation factor VIII was used.
  • a commercial kit (Aceracrom FVIIIR: Ag, manufactured by Boehringer Mannheim Yamanouchi Co., Ltd., Japan)
  • the amount of blood coagulation factor VIII antigen in the plasma before and after the above adsorption experiment was measured.
  • the amount of factor VIII antigen after the adsorption experiment was 10% before the adsorption experiment.
  • the adsorbent after the adsorption experiment was transferred to a plastic column, and washed three times with 2 milliliters of a 10 mM aqueous sodium citrate solution (pH 7.4). Then, an aqueous solution containing 2 OmM sodium phosphate, 1% lysine, 1 M sodium chloride, 20% ethylene glycol, and 1% triton X100 ( Factor VIII was eluted using two PH 6.0) 2 milliliters. When the factor VIII activity contained in the eluate was measured, 69% of the factor VIII in the plasma used in the adsorption experiment was recovered.
  • Example 2 (1) 10 milliliters of the water-insoluble carrier obtained in Example 1 was suspended in 12 milliliters of dimethyl sulfoxide, and 8 milliliters of the suspension was added to the suspension. Add a little epichlorohydrin and 1 milliliter of 50% aqueous sodium hydroxide solution and shake at 30 ° C for 5 hours to activate the active group. As a result, an activated carrier having an epoxy group was obtained.
  • Molecule having polycation synthesized in Example 1 135 mg was dissolved in 50% aqueous solution of dimethylformamide in 13.5 milliliters of aqueous solution, and 9 milliliters of the activated carrier obtained as described above was added thereto. Adjust the pH to 12 with, shake for 20 hours at 45 ° C, and fix the copolymer of 2-hydroxymethyl methacrylate and getylaminoethyl methacrylate. Thus, a modified adsorbent was obtained. The positive charge density per milliliter of the adsorbent determined by titration with an aqueous sodium hydroxide solution was 110 / eq.
  • Sepharose which is an agarose-based carrier, was used as the water-insoluble carrier.
  • an activating carrier having a tresyl group as an active group commercially available tresyl-activated Sepharose 4B (Pharmacia, Sweden) 3 milliliters
  • the pH was adjusted to 8 with an aqueous sodium hydroxide solution, and the mixture was shaken at room temperature for 24 hours to obtain an adsorbent.
  • the positive charge density of the adsorbent per milliliter determined by titration with an aqueous solution of sodium hydroxide was 12 ⁇ eq. I got it.
  • Example 1 when an adsorption experiment was performed using plasma from a healthy subject, the factor VII I activity in the plasma after the adsorption experiment was 10% of that before the adsorption experiment.
  • Example 1 As a water-insoluble carrier, Toray Pearl (Tosoichi Japan, Ltd.), a hydrophilic bullpolymer carrier, was used.
  • an activating carrier having a tresyl group as an active group commercially available AF-tresyl toyopearl 6.50 (manufactured by Tosoh Corporation B, Japan) 3 milliliters was used in Example 1. 30 mg of a copolymer of 2-hydroxyhydroxymethyl methacrylate and getylaminoethyl methacrylate, which was synthesized in step 2, was dissolved in 6 milliliter of 50% aqueous dimethylformamide solution.
  • Example 2 Using a crosslinked copolymer having a butyl alcohol unit synthesized in Example 1 as a water-insoluble carrier, in the same manner as in Example 1, An activated carrier having an epoxy group as an active group was obtained using epichlorohydrin.
  • poly-L-lysine molecular weight: 20,000, ICN Immunobiological, America
  • PH10 aqueous solution of sodium phosphate
  • Example 1 when an adsorption experiment was performed using plasma from a healthy individual, the factor VIII activity in the plasma after the adsorption experiment was 28% of that before the adsorption experiment.
  • the adsorption rates of factor VIII and factor IX were determined by measuring the respective activities remaining in plasma after the adsorption experiment. The measurement of the factor VIII activity was carried out in the same manner as in Example 1.
  • the activity of Factor IX is determined by the Hard i sty—step method [R.M.Had i s t y e a 1.: Thro m b os D a he s.H a e m o h., V o
  • the adsorption rates of factor VIII and factor IX were determined by the respective activities remaining in the plasma after the adsorption experiment, as in Example 6. It was determined by measuring.
  • the activated carrier 3 was added to a solution prepared by dissolving 150 mg of the copolymer synthesized in Example 1 in 6% of a 75% aqueous dimethyl sulfoxide solution. Milliliter was added, the pH was adjusted to 8 with an aqueous solution of sodium hydroxide, and the mixture was shaken at room temperature for 24 hours to obtain an adsorbent.
  • the positive charge density of 1 milliliter of the adsorbent determined by titration with an aqueous sodium hydroxide solution was 43 ⁇ eq.
  • Example 1 As in Example 1, an adsorption experiment was carried out using plasma from a healthy subject. As a result, the factor VIII activity in the plasma after the adsorption experiment was 26% of that before the adsorption experiment. Further, factor VIII was eluted using the same eluate as in Example 1, and the activity of factor V factory II contained in the eluate was measured. Factor VIII had been recovered.
  • Example 8 In order to examine the multi-molecular structure of the von Willebrand factor in the eluate in Example 1, SDS electrophoresis using a 2% agarose gel was performed. The existence of multimers was recognized up to the same high molecular weight region. Industrial applicability
  • the blood coagulation factor VIII adsorbent of the present invention has a molecule having a polycation, it can be compared with a conventional adsorbent using a low-molecular-weight compound as a ligand. Interaction becomes more specific, and plasma can be used as it is without using the conventional method via cryoprecipitate.
  • the VI I I factor can be adsorbed at a high rate.
  • the adsorbent for blood coagulation factor VIII of the present invention makes a remarkable contribution to the production of a blood coagulation factor VIII preparation.

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Abstract

L'invention concerne un adsorbant pour le facteur VIII de la coagulation sanguine, comprenant un composite obtenu par immobilisation d'une molécule polycationique sur un vecteur non soluble dans l'eau, ladite molécule ayant une masse moléculaire d'au moins 15000 et la densité de charge positive étant de 1 à 200 νeq par millilitre de composite. L'adsorbant selon l'invention présente une telle capacité d'adsorption spécifique par rapport au facteur VIII qu'il peut adsorber et extraire ledit facteur d'une solution contenant différentes protéines plasmatiques qui elles-mêmes contiennent ledit facteur, cela avec une haute sélectivité et un rendement élevé.
PCT/JP1992/001480 1991-11-12 1992-11-12 Adsorbant pour le facteur viii de la coagulation sanguine Ceased WO1993009869A1 (fr)

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JP32235391 1991-11-12
JP3/322353 1991-11-12

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WO1993009869A1 true WO1993009869A1 (fr) 1993-05-27

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101010334B (zh) * 2004-08-30 2013-02-06 东丽株式会社 分级装置

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP4509341B2 (ja) * 2000-09-21 2010-07-21 シスメックス株式会社 凝固因子欠乏血漿およびその製造方法

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS5627090B2 (fr) * 1976-08-04 1981-06-23
JPS6241212B2 (fr) * 1977-07-25 1987-09-02 Monsanto Co
JPH01157000A (ja) * 1987-08-14 1989-06-20 Waander Riethorst 凝固因子の単離方法およびそれに適した吸着剤
JPH0236199A (ja) * 1988-07-22 1990-02-06 Asahi Medical Co Ltd 血液凝固因子の精製方法および精製用吸着材

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS5627090B2 (fr) * 1976-08-04 1981-06-23
JPS6241212B2 (fr) * 1977-07-25 1987-09-02 Monsanto Co
JPH01157000A (ja) * 1987-08-14 1989-06-20 Waander Riethorst 凝固因子の単離方法およびそれに適した吸着剤
JPH0236199A (ja) * 1988-07-22 1990-02-06 Asahi Medical Co Ltd 血液凝固因子の精製方法および精製用吸着材

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101010334B (zh) * 2004-08-30 2013-02-06 东丽株式会社 分级装置

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JPH05262664A (ja) 1993-10-12
AU2932992A (en) 1993-06-15

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