WO2016066128A1 - 一种含有银杏内酯B和Xa因子抑制剂的药物组合物及其制备方法和用途 - Google Patents

一种含有银杏内酯B和Xa因子抑制剂的药物组合物及其制备方法和用途 Download PDF

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WO2016066128A1
WO2016066128A1 PCT/CN2015/093305 CN2015093305W WO2016066128A1 WO 2016066128 A1 WO2016066128 A1 WO 2016066128A1 CN 2015093305 W CN2015093305 W CN 2015093305W WO 2016066128 A1 WO2016066128 A1 WO 2016066128A1
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parts
composition
ginkgolides
weight ratio
ginkgolide
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PCT/CN2015/093305
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English (en)
French (fr)
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孙毅
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成都百裕科技制药有限公司
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Priority to US15/523,239 priority Critical patent/US10058533B2/en
Priority to KR1020177014595A priority patent/KR101968190B1/ko
Priority to EP15854130.0A priority patent/EP3213749B1/en
Priority to CA2966249A priority patent/CA2966249C/en
Priority to JP2017542250A priority patent/JP6389009B2/ja
Publication of WO2016066128A1 publication Critical patent/WO2016066128A1/zh

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/365—Lactones
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/42—Oxazoles
    • A61K31/423—Oxazoles condensed with carbocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/4353—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/437—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems the heterocyclic ring system containing a five-membered ring having nitrogen as a ring hetero atom, e.g. indolizine, beta-carboline
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/4418—Non condensed pyridines; Hydrogenated derivatives thereof having a carbocyclic group directly attached to the heterocyclic ring, e.g. cyproheptadine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/4425—Pyridinium derivatives, e.g. pralidoxime, pyridostigmine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/4427—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems
    • A61K31/444—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems containing a six-membered ring with nitrogen as a ring heteroatom, e.g. amrinone
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/445—Non condensed piperidines, e.g. piperocaine
    • A61K31/4523—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems
    • A61K31/4545—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems containing a six-membered ring with nitrogen as a ring hetero atom, e.g. pipamperone, anabasine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/535—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one oxygen as the ring hetero atoms, e.g. 1,2-oxazines
    • A61K31/5375—1,4-Oxazines, e.g. morpholine
    • A61K31/5377—1,4-Oxazines, e.g. morpholine not condensed and containing further heterocyclic rings, e.g. timolol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • 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
    • A61P7/02—Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors

Definitions

  • the present invention relates to a pharmaceutical composition
  • a pharmaceutical composition comprising a ginkgolides B and a factor Xa inhibitor.
  • Xa factor inhibitors increase the anticoagulant effect while reducing the risk of major bleeding, without the need for blood monitoring and long-term treatment.
  • Factor Xa does not activate platelets and there is no positive feedback to the clotting cascade.
  • Direct inhibitor of factor Xa inhibits both thrombin-binding and prothrombin-binding factors.
  • Rivaroxaban has been officially launched as the first factor Xa inhibitor.
  • a large number of clinical studies have shown that rivaroxaban is superior to the current standard treatment drug enoxaparin in the field of prevention and treatment of venous thromboembolism.
  • it can be used to prevent the formation of deep vein thrombosis (DVT) and pulmonary embolism (PE) in patients with hip and knee joint replacement. It can also be used to prevent stroke and non-central nervous system embolism in patients with non-valvular atrial fibrillation. Reduce the risk of recurrence of coronary syndromes, etc.
  • overdose may lead to bleeding complications, and some subgroups have a higher risk of bleeding, and there is no specific antidote against rivaroxaban.
  • Apixaban an oral selective activated factor Xa inhibitor
  • the safety and efficacy of apixaban may depend on basic antiplatelet therapy.
  • Edusarban is a small molecule oral anticoagulant developed by Japan's First Sankyo Co., Ltd., which is a factor X (FXa) blocker. It is mainly used for patients undergoing total knee arthroplasty (TKA), total hip arthroplasty (THA), and hip fracture surgery (HFS) with venous thromboembolism (VTE).
  • TKA total knee arthroplasty
  • THA total hip arthroplasty
  • HFS hip fracture surgery
  • VTE venous thromboembolism
  • activated factor X (FXa) activates prothrombin (FII) to thrombin (FIIa), which promotes fibrin formation, thereby forming a thrombus, and FXa has become the main target for the development of a new generation of anticoagulant drugs. point.
  • Edesaban is an oral anticoagulant that inhibits thrombus formation by selectively, reversibly and directly inhibiting FXa, which is 104 times more selective for FXa than FIIa.
  • Clinical trials in Japan and abroad have confirmed that this product can effectively inhibit VTE in patients undergoing lower limb plastic surgery, and is safe and reliable.
  • the drug also has a relatively high cost limitation and may become the treatment of most patients. Stumbling block.
  • Rezashaban has a relative molecular mass of 528. As an orally effective peptide analog, it has a high affinity with factor Xa.
  • a phase II clinical study has shown its prevention of venous thromboembolism (VTE) after orthopedic surgery. It is vaild. This randomized, double-blind study included 656 patients undergoing total knee arthroplasty. Of the 438 patients, the incidence of rezashaban had a significant dose-response relationship with VTE. It is worth noting that the incidence of VTE in the 25 mg group of Rezashaban was significantly lower than that of the enoxaparin group, and the incidence of major bleeding was similar to that of the enoxaparin group. However, the probability of major bleeding in the 50-100 mg group of Rezashaban was higher than that of the enoxaparin group.
  • Omizaban is a highly selective and reversible thrombin factor Xa inhibitor for injection, which inhibits thrombosis in a dose-dependent manner with rapid onset and failure, and pharmacokinetics follows a linear law. Not easily eliminated by the kidneys.
  • Previous studies have shown that activation of factor X inhibitors in the treatment of ACS has a risk of thrombosis, so PCI should be adjusted to adjust these anticoagulants and other antithrombotic doses, but the application of medium doses of omeproxaban does not seem to be used Worried about these factors.
  • omeprazole may be a single anticoagulant for NSTE-ACS patients from emergency to interventional therapy. And it is likely to have great advantages. It has also been reported that the dose of omeproxacin excreted by the kidney is less than 25%, so there is no need to adjust the dose for patients with impaired renal function. It has also been reported that omeproxacin does not reduce the risk of ischemic events and increases the risk of bleeding, and does not support the use of omeproxaban in patients with NSTE-ACS who have undergone PCI.
  • the present invention provides a pharmaceutical composition
  • ginkgolides B comprising a ginkgolides B and a factor Xa inhibitor.
  • Xa factor inhibitor is a Shaban class drug.
  • the Shaban drugs are rivaroxaban, apixaban, edoxaban, rezashaban and/or omeproxaban.
  • the rivaroxaban structure is:
  • the apixaban structure is:
  • edoxaban The structure of edoxaban is:
  • the structure of the Rezashaban is:
  • the structure of the Omishaban is:
  • the weight ratio of the ginkgolides B to rivaroxaban is: bilobalide B1-20 parts and rivaroxaban 5-40 parts.
  • the weight ratio of the two is: bilobalide B5-15 parts and rivaroxaban 10-20 parts. Further preferably, the weight ratio of the two is: 10 parts of ginkgolides B and 15 parts of rivaroxaban.
  • the weight ratio of the ginkgolide B to apixaban is: ginkgolides B1-20 parts and Apixaban 0.5-20 parts.
  • the weight ratio of the two is: bilobalide B5-15 parts and apixaban 1-10 parts. Further preferably, the weight ratio of the two is: 10 parts of ginkgolides B and 5 parts of apixaban.
  • the weight ratio of the ginkgolides B to edoxaban is: ginkgolides B1-20 parts and edoxaban 8-50 parts.
  • the weight ratio of the two is: bilobalide B5-15 parts and edoxaban 10-30 parts. Further preferably, the weight ratio of the two is: 10 parts of ginkgolides B and 15 parts by edoxaban.
  • the weight ratio of the ginkgolides B to the rezazaban is: ginkgolides B1-20 parts and Rezashaban 1-20 parts.
  • the weight ratio of the two is: bilobalide B5-15 parts and Rezashaban 5-15 parts. Further preferably, the weight ratio of the two is: 10 parts of ginkgolides B and 10 parts of Rezazaban.
  • the weight ratio of the ginkgolides B to the omeproxaban is: bilobalide B1-20 parts and olmesaban 15-150 parts.
  • the weight ratio of the two is: 5-15 parts of ginkgolides and 30-125 parts of omeproxaban. Still preferably, the weight ratio of the two is: ginkgolides B8-12 parts and omiproxan 50-90 parts. Further preferably, the weight ratio of the two is: 10 parts of ginkgolides B and 60 parts of omeproxaban.
  • the invention also provides a method of preparing the pharmaceutical composition comprising the steps of:
  • the pharmaceutically acceptable excipients include: starch, pregelatinized starch, lactose, sucrose, talc, dextrin, cyclodextrin, microcrystalline cellulose, croscarmellose sodium, carboxymethyl Sodium starch, low-substituted hydroxypropyl cellulose, crospovidone, glucose, meglumine, magnesium stearate, dextran, glycerol, ethanol, propylene glycol, polyethylene glycol, mannitol, sorbitol, xylose Alcohol, fiber vegetable oil, sodium benzoate, sodium salicylate, hydrochloric acid, citric acid, sodium citrate, sodium dihydrogen phosphate, disodium hydrogen phosphate, gelatin, lecithin, vitamin C or one of several.
  • the pharmaceutical preparation comprises: a tablet, a capsule, a soft capsule, an oral liquid, a granule, a pill, a pill, a powder, a plaster, an agent, an injection, a suppository, a patch, a drop, a spray, a cream.
  • the invention also provides the use of the combination of ginkgolide B and a factor Xa inhibitor for preparing an anti-platelet aggregation drug.
  • Xa factor inhibitor is a Shaban class drug.
  • the Shaban drugs are rivaroxaban, apixaban, edoxaban, rezashaban and/or omeproxaban.
  • the weight ratio of the ginkgolides B to rivaroxaban is: bilobalide B1-20 parts and rivaroxaban 5-40 parts.
  • the weight ratio of the two is: bilobalide B5-15 parts and rivaroxaban 10-20 parts. Further preferably, the weight ratio of the two is: 10 parts of ginkgolides B and 15 parts of rivaroxaban.
  • the weight ratio of the ginkgolides B to apixaban is: ginkgolides B1-20 parts and apixaban 0.5-20 parts.
  • the weight ratio of the two is: bilobalide B5-15 parts and apixaban 1-10 parts. Further preferably, the weight ratio of the two is: 10 parts of ginkgolides B and 5 parts of apixaban.
  • the weight ratio of the ginkgolides B to edoxaban is: ginkgolides B1-20 parts and edoxaban 8-50 parts.
  • the weight ratio of the two is: bilobalide B5-15 parts and edoxaban 10-30 parts. Further preferably, the weight ratio of the two is: 10 parts of ginkgolides B and 15 parts by edoxaban.
  • the weight ratio of the ginkgolides B to the rezazaban is: ginkgolides B1-20 parts and Rezashaban 1-20 parts.
  • the weight ratio of the two is: bilobalide B5-15 parts and Rezashaban 5-15 parts. Further preferably, the weight ratio of the two is: 10 parts of ginkgolides B and 10 parts of Rezazaban.
  • the weight ratio of the ginkgolides B to the omeproxaban is: bilobalide B1-20 parts and olmesaban 15-150 parts.
  • the weight ratio of the two is: 5-15 parts of ginkgolides and 30-125 parts of omeproxaban. Still preferably, the weight ratio of the two is: ginkgolides B8-12 parts and omiproxan 50-90 parts. Further preferably, the weight ratio of the two is: 10 parts of ginkgolides B and 60 parts of omeproxaban.
  • the pharmaceutical composition provided by the invention can prevent rivaroxaban, apixaban, edoxaban, reza, by inhibiting platelet aggregation by different action mechanisms because it contains ginkgolides B and saban active ingredients.
  • the anti-platelet aggregation function of Shaban and Omibanban and the anti-platelet aggregation function of Ginkgolide B have synergistic effects.
  • the clinical use can reduce the dose of Shaban drugs. Increased efficacy, reduced costs, reduced side effects, and a better choice for clinical research.
  • the pharmaceutical composition of the invention has novel formula, simple composition, clear action mechanism and remarkable curative effect, and can realize industrialized large-scale production.
  • the present invention combines ginkgolide B with a banban drug, and the two can exert synergistic effects and have an excellent effect of inhibiting platelet aggregation.
  • the invention combines ginkgolide B with a class of drugs, and the two can exert a synergistic effect, and the effect of inhibiting platelet aggregation is excellent, and the dosage of the class of drugs can be reduced in clinical application, thereby reducing the use thereof.
  • the side effects can also reduce costs and have excellent clinical application prospects.
  • the ginkgolides B monomer compound of the present invention can be obtained by purchasing a commercially available product or by separating and purifying ginkgolides on the existing method; rivaroxaban, apixaban, and degree Shaban, Rezashaban, Omibanban, etc. can also be obtained by purchasing commercially available products, or synthesized by existing methods. All monomer compounds were tested to be consistent with the corresponding control structure, and their purity was above 95% by HPLC.
  • a pharmaceutically acceptable conventional excipient is added to prepare a tablet according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a capsule or a soft capsule according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a pellet according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare an oral liquid according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a dropping pill according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a spray according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a water injection or a powder injection according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a sustained release preparation or a controlled release preparation according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a targeted preparation according to a conventional process.
  • granules or suspensions are prepared according to a conventional process by adding a pharmaceutically acceptable conventional excipient.
  • a pharmaceutically acceptable conventional excipient is added to prepare a tablet according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a capsule or a soft capsule according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a pellet according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare an oral liquid according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a dropping pill according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a spray according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a water injection or a powder injection according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a sustained release preparation or a controlled release preparation according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a targeted preparation according to a conventional process.
  • granules or suspensions are prepared according to a conventional process by adding a pharmaceutically acceptable conventional excipient.
  • a pharmaceutically acceptable conventional excipient is added to prepare an oral liquid according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a dropping pill according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a spray according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a water injection or a powder injection according to a conventional process.
  • a pharmaceutically acceptable conventional excipient is added to prepare a sustained release preparation or a controlled release preparation according to a conventional process.
  • Test Example 1 Anti-PAF-induced platelet aggregation and its effect on Xa activity in vitro by combination of ginkgolide B and saban
  • micro-reaction plate microbiome method was used to evaluate the anti-PAF-induced platelet aggregation and Xa activity in vitro by the combination of ginkgolides B and Shaban produced by Chengdu Baiyu Technology Pharmaceutical Co., Ltd.
  • German Sartorius BS124S electronic balance (division value: 0.0001g), instrument number: BKY-YB-001; ACS-15LED electronic platform scale, produced by Shanghai Eagle Brand Weighing Apparatus Co., Ltd., instrument number BKY-YB-002; Changsha Xiangzhi DL -5M small vertical low speed large capacity refrigerated centrifuge, instrument number: BKY-ZX-040, Changsha Xiangzhi Centrifuge Instrument Co., Ltd.; Haier DW-40L188 cryopreservation box, instrument number: BKY-ZX-012, preserve plasma, etc.
  • Aggregation inhibition rate (time required for maximum aggregation of the drug-administered group - time required for maximum aggregation of the blank control group) / time required for maximum aggregation of the blank control group ⁇ 100%
  • Plasma anti-Xa activity was determined on a microplate reader using the chromogenic substrate method according to the kit instructions and references.
  • test data is the measurement data, and the aggregation inhibition rate and anti-Xa activity are calculated according to the corresponding calculation formula, and the average is calculated by Excel software.
  • Group Platelet aggregation time (min) Platelet aggregation inhibition rate (%) Saline 20 - Ginkgolide B 30 50 Livaraban 25 25 Apixaban group 40 100 Yidushaban group 30 80 Rezashaban 35 75 Omisaban 40 100 Composition 1 60 200 Composition 2 30 50 Composition 3 35 75 Composition 4 33 65 Composition 5 55 175 Composition 6 50 150 Composition 7 56 180 Composition 8 59 195 Composition 9 88 340 Composition 10 44 120 Composition 11 48 140 Composition 12 45 125 Composition 13 60 200 Composition 14 65 225 Composition 15 68 240 Composition 16 75 275
  • Composition 17 80 300 Composition 18 50 150 Composition 19 60 200 Composition 20 65 225 Composition 21 group 66 230 Composition 22 76 280 Composition 23 79 295 Composition 24 73 265 Composition 25 60 200 Composition 26 36 80 Composition 27 40 100 Composition 28 55 175 Composition 29 58 190 Composition 30 90 350 Composition 31 50 150 Composition 32 55 175 Composition 33 group 52 160 Composition 34 65 225 Composition 35 62 210 Composition 36 60 200 Composition 37 58 190 Composition 38 79 295 Composition 39 85 325 Composition 40 80 300 Composition 41 83 315
  • the platelet aggregation rate of each composition group was significantly different from that of the physiological saline group (p ⁇ 0.01, p ⁇ 0.05), indicating the respective pharmaceutical compositions of the present invention. Both can effectively inhibit platelet aggregation; compared with ginkgolides B and saban drugs alone, the anti-platelet aggregation function is significantly improved when the ginkgolide B is combined with the banban drug, indicating that ginkgolides B and The combination of Shaban drugs has a synergistic effect.
  • Anticoagulant is effectively defined as plasma anti-Xa activity ⁇ 0.5IU/ML ("Calculation of low-molecular-weight heparin for cardiovascular disease" report: based on the pharmacokinetic characteristics of LMWH in vivo.
  • the anti-hemagglutination factor Xa activity of each composition group was significantly different from that of the physiological saline group, indicating that each of the pharmaceutical compositions of the present invention can effectively inhibit factor Xa activity and anti-platelet aggregation;
  • the anti-hemagglutination factor Xa activity of the ginkgolide B in the present invention is significantly improved when the ginkgolide B is combined with the banban drug, indicating that the ginkgolide B is combined with the banban drug.
  • the role of synergy is significantly improved when the ginkgolide B is combined with the banban drug.
  • compositions 1 to 8 As can be seen from Tables 2 and 3, in the composition group (compositions 1 to 8) of ginkgolides B + rivaroxaban, the platelet aggregation inhibition rate and the factor Xa activity of the compositions 1, 5 to 8 were relatively higher. High, composition group 1 has the highest platelet aggregation inhibition rate and factor Xa activity. Therefore, in the composition of ginkgolides B + rivaroxaban, the ratio of ginkgolides B to rivaroxaban is preferably: ginkgo Ester B 5-15 parts and rivaroxaban 10-20 parts, most preferably: 10 parts of ginkgolides B and 15 parts of rivaroxaban.
  • compositions 9 to 16 of ginkgolides B + apixaban the platelet aggregation inhibition rate and the factor Xa activity of the composition groups 9, 13 to 16 were relatively high, and the platelet aggregation inhibition of the composition group 9 was suppressed.
  • the ratio of factor Xa activity is the highest. Therefore, in the composition of ginkgolide B + apixaban, the ratio of ginkgolide B to apixaban is preferably: 5-15 parts of ginkgolide B and apixaban. From 1 to 10 parts, most preferably: 10 parts of ginkgolides B and 5 parts of apixaban.
  • the ratio of ginkgolides B to edoxaban is preferably: ginkgolides B 5-15 parts and edoxaban 10-30 parts, most preferably: 10 parts of ginkgolides B and 15 parts of edoxaban.
  • the ratio of ginkgolides B to rezazaban is preferably: 5-15 parts of ginkgolides B and Rezashaban 5-15 parts, most preferably: 10 parts of ginkgolides B and 10 parts of Rezazaban.
  • compositions 30 to 41 of ginkgolides B+omisoxaban the platelet aggregation inhibition rate and the factor Xa activity of the composition 30, 38-41 group were relatively high, and the platelet aggregation inhibition of the composition group 30 was suppressed.
  • the rate and activity of factor Xa are the highest. Therefore, in the composition of ginkgolides B+omixampan, the ratio of ginkgolides B to omeproxa is preferably: ginkgolides B 8-12 parts and omeproxaban 50-90 parts, most preferably: 10 parts of ginkgolides B and 10 parts of omeproxaban.
  • the experimental results show that the present invention combines ginkgolide B and Shaban active ingredients, such as rivaroxaban, apixaban, edoxaban, rezashaban, and omeproxaban.
  • the anti-platelet aggregation function is obviously enhanced, the Xa factor activity is obviously increased, and the two are better than the two, and the combination of the two plays a synergistic effect.
  • the composition of the active ingredient of ginkgolide B and Shaban is anti-platelet aggregation. Strong function, high activity of factor Xa, anti-platelet aggregation through different mechanisms of action.
  • the anticoagulant effect and anticoagulant activity of the combination of ginkgolides B and Shaban in rabbit models of traumatic limb deep vein thrombosis were compared.
  • Japan Sysmex CA-660 automatic blood coagulation analyzer instrument number: BKY-ZX-044, Sysmex Biotech (Wuxi) Co., Ltd.
  • BC-117F type refrigerator instrument number: BKY-ZX-015, produced by Qingdao Haier Co., Ltd.
  • Multiskan MK3 microplate reader instrument number: BKY-ZX-014, American Thermo electroncorporation.
  • German Sartorius BS124S electronic balance (division value: 0.0001g), instrument number: BKY-TJ-002, produced by Sartorius Co., Germany.
  • ZT-14V2 biological tissue automatic dehydrator instrument number: BKY-BL-016, produced by Xiaogan Yaguang Medical Electronic Technology Co., Ltd.
  • Leica RM2126 Rotary Slicer instrument number: BKY-BL-004, produced by Shanghai Leica Instrument Co., Ltd.
  • YR-21 biological tissue automatic dyeing machine instrument number: BKY-BL-015, produced by Xiaogan Yaguang Medical Electronic Technology Co., Ltd.
  • Carmellose (provided by the client).
  • Lactose specifications: 100g / bottle, batch number: 2015041701, expiration date: 2018.06, Chengdu Kelon Chemical Reagent Factory.
  • Mannitol specifications: 500g / bottle, batch number: 201402201, expiration date: 2017.06, Chengdu Kelon Chemical Reagent Factory.
  • Coagulation quality control specifications: 1ml ⁇ 10 bottles / box, batch number: 528167B, expiration date: 2017.4.27, SIEMENS.
  • PT reagent specification: 2ml ⁇ 10 bottles / box, batch number: R5003, R5005, expiration date: 2017.1.1, 2017.3.1, produced by Xisen Meikang Biotechnology Co., Ltd.
  • aPTT reagent specification: 2ml ⁇ 10 bottles / box, batch number: R5001, R5006, expiration date: 2016.9.1, 2016.12.1, produced by Xisen Meikang Biotechnology Co., Ltd.
  • Anti-hemagglutination factor (Xa) assay kit (chromogenic substrate method), specification: 10 ⁇ 71 nkat, batch number: N1243555, manufacturer: Italy Chromogenix.
  • Chromogenic substrate S size: 25mg, batch number: N1143454, manufacturer: Italy Chromogenix.
  • Polyethylene glycol (PEG) 6000 specification: 50g, batch number: LB50P76, manufacturer: Beijing Bailingwei Technology Co., Ltd.
  • Sodium pentobarbital for rabbit anesthesia was formulated into a 3% concentration with purified water at the time of use. Formulated as a 3% sodium pentobarbital solution (g/v). Used for rabbit anesthesia during modeling.
  • Tissue specimen fixative 95% ethanol (AR grade), batch number: 2015052301, valid period: 2020.4, specification: 20kg/barrel, produced by Chengdu Kelon Chemical Reagent Factory; glacial acetic acid (AR grade), batch number: 2015040201, valid until: 2020.03, specification: 500ml/bottle, produced by Chengdu Kelon Chemical Reagent Factory; formaldehyde (AR grade), batch number: 2014120801, valid period: 2016.11, specification: 500ml/bottle, Chengdu Kelon Chemical Reagent Factory produce.
  • Preparation method separately measure 4200ml of 95% ethanol, 500ml of formaldehyde, 300ml of glacial acetic acid and 900ml of purified water into plastic bucket, and mix well.
  • Dehydration reagent anhydrous ethanol (AR grade), batch number: 2015052501, valid period: 2020.04, specification: 20kg/barrel, produced by Chengdu Kelon Chemical Reagent Factory.
  • Preparation method anhydrous ethanol was diluted with purified water to prepare 70%, 80% and 90% ethanol solution.
  • Transparent reagent xylene (AR grade), batch number: 2014071001, valid period: 2019.06, specification: 500ml / bottle, produced by Chengdu Kelon Chemical Reagent Factory.
  • Hematoxylin dyeing solution hematoxylin (AR grade), batch number: 2014010701, valid period: 2017.04, specification: 5g / bottle, Chengdu Kelon Chemical Reagent Factory production; aluminum sulfate potassium (AR grade), batch number: 2014091601, valid until: 2017.1 ,Specification: 500g/bottle, produced by Chengdu Kelon Chemical Reagent Factory; sodium iodate (AR grade), batch number: 20140711, valid period: 2017.1, specification: 100g/bottle, produced by Chengdu Kelon Chemical Reagent Factory; glycerin Glycerol, AR grade), batch number: 2014122401, valid until: 2017.12, produced by Chengdu Kelon Chemical Reagent Factory; glacial acetic acid (AR grade), Batch number: 2015040201, valid until: 2020.3, specification: 500ml / bottle, produced by Chengdu Kelon Chemical Reagent Factory.
  • Preparation method Weigh 1.5002g of hematoxylin, weigh 250ml of purified water and stir it to completely dissolve, then add 312.5ml of purified water, weigh 0.3003g of sodium iodate into the beaker, stir for 5 minutes, then add potassium aluminum sulfate 37.5004g, wait It is completely dissolved and added with 187.5 ml of glycerin, mixed, and finally added 7.5 ml of glacial acetic acid, and stirred well to obtain.
  • Yihong Dyeing Liquid Yihong (water soluble), batch number: 2013110501, valid period: 2016.05, specification: 25g/bottle, Chengdu Kelon Chemical Reagent Factory; anhydrous ethanol (AR grade), batch number: 2015052501, valid period: 2020.4 , Specification: 20kg/barrel, produced by Chengdu Kelon Chemical Reagent Factory.
  • Preparation method Measure 600 ml of absolute ethanol and 150 ml of purified water into a beaker, mix well, add 3.7505 g of eosin, stir well to dissolve, that is, obtain.
  • Hydrochloric acid-ethanol differentiation solution hydrochloric acid (AR grade), batch number: 2014072301, valid period: 2019.7.22, specification: 500ml/bottle, Chengdu Kelon Chemical Reagent Factory; anhydrous ethanol (AR grade), batch number: 2015052501, valid period To: 2020.4, Specification: 20kg/barrel, produced by Chengdu Kelon Chemical Reagent Factory.
  • Preparation method Take 520 ml of absolute ethanol and 222.5 ml of purified water, pour into a beaker, mix well, then slowly add 7.5 ml of concentrated hydrochloric acid, stir and mix, and obtain.
  • Sealing reagent neutral gum, specifications: 100g / bottle, batch number: 20140106, valid until: 2018.12, China Shanghai Hao Yang Instrument Co., Ltd.
  • Cover glass cleaning solution potassium dichromate: (AR grade), batch number: 20120822, valid period: 2016.07, specification: 500g / bottle, produced by Chengdu Kelon Chemical Reagent Factory; concentrated sulfuric acid: (AR grade), batch number: 2014072301, 20130607, valid until: 2010.7.22, 2011.06.06, specifications: 500ml / bottle, Chengdu Kelon Chemical Reagent Factory production.
  • Preparation method take 500ml of purified water, add 50.0003g of potassium dichromate, stir, dissolve and mix, then slowly add 50ml of concentrated sulfuric acid to it, stir and mix.
  • Drug (mg): Carboxymethylcellulose (mg): Lactose (mg): Mannitol (mg) 1: 2: 10: 20 After grinding evenly, add water to 1 ml of purified water, stir well, that is. For example: 50mg drug + 100mg carboxypropylmethylcellulose + 500mg lactose + 1000mg mannitol, evenly ground, add 50ml of water, stir evenly, that is, get 1mg / ml of liquid.
  • Each group of animals was divided into groups after model establishment, and administered intragastrically according to the administration scheme of Table 4, and administered continuously for 7 days, once a day.
  • Table 4 dosage regimen
  • the animals were sacrificed by bloodletting, and the femoral artery and vein were harvested.
  • HE staining was performed.
  • the endothelium injury, thrombosis and vascular rupture were examined by light microscopy.
  • the degree of the above lesions was graded. The specific scoring criteria are shown in the table. 5.
  • Coagulation function, coagulation factor and PAF content data were used as measurement data.
  • Excel software was used to calculate the mean and standard deviation to judge whether the data was normally distributed.
  • the excel F test was used to test the homogeneity of variance, and then the t test was selected. Statistics are performed; if non-normal distribution, nonparametric tests are performed.
  • the pathological detection data was graded data, and statistical analysis was performed using "multiple independent samples" of software SPSS13.0 "nonparametric test”.
  • Composition 15 21.55 ⁇ 7.89 34.59 ⁇ 7.76** 35.44 ⁇ 6.79 31.39 ⁇ 6.69 32.88 ⁇ 8.97
  • Composition 16 20.64 ⁇ 5.77 35.89 ⁇ 8.55** 36.75 ⁇ 8.66 31.84 ⁇ 9.48 31.84 ⁇ 8.96
  • Composition 17 29.30 ⁇ 8.54 35.80 ⁇ 12.36** 25.45 ⁇ 7.69* 30.28 ⁇ 6.64 33.75 ⁇ 4.95
  • Composition 18 32.82 ⁇ 7.69 29.57 ⁇ 7.99* 36.56 ⁇ 15.78 29.94 ⁇ 7.56 28.84 ⁇ 5.59
  • Composition 19 27.91 ⁇ 9.26 31.76 ⁇ 9.94* 29.58 ⁇ 9.74 28.67 ⁇ 5.95 29.57 ⁇ 5.64
  • Composition 20 33.65 ⁇ 7.85 33.46 ⁇ 7.87* 36.48 ⁇ 15.95 29.75 ⁇ 7.48 28.75 ⁇ 5.83
  • Table 6 shows that: 1 0.5 h to 4 h after administration, the aPTT (activated partial thromboplastin time) was not significantly prolonged in the model group compared with the normal control group (p>0.05). Explain that the model was successful. 2 Compared with the model group, most of the aPTTs were not significantly prolonged (p>0.05) from 0.5h to 4h after administration. The results showed that there were no adverse reactions of bleeding in ginkgolide B, Shaban and each composition group. 3 Compared with the model group, the aPTT of ginkgolide B and rivaroxaban group was significantly prolonged (p ⁇ 0.05), and aPPT of apixaban group, edoxaban group, rezashaban group and omeproxaban group.
  • composition 28 7.01 ⁇ 0.18** 6.69 ⁇ 0.42 6.77 ⁇ 0.52 6.95 ⁇ 0.25 6.85 ⁇ 0.26 Composition 29 7.00 ⁇ 0.31** 6.78 ⁇ 0.35 6.96 ⁇ 0.24 6.97 ⁇ 0.28 6.96 ⁇ 0.27 Composition 30 7.15 ⁇ 0.27** 6.86 ⁇ 0.14 6.88 ⁇ 0.29 6.95 ⁇ 0.24 6.88 ⁇ 0.17 Composition 31 6.89 ⁇ 0.16* 6.88 ⁇ 0.20 6.74 ⁇ 0.23 6.99 ⁇ 0.18 6.79 ⁇ 0.35 Composition 32 6.85 ⁇ 0.24* 6.95 ⁇ 0.24 6.78 ⁇ 0.56 6.97 ⁇ 0.36 7.01 ⁇ 0.26 Composition 33 group 6.86 ⁇ 0.19* 7.01 ⁇ 0.23 6.96 ⁇ 0.42 6.76 ⁇ 0.46 6.96 ⁇ 0.18 Composition 34 6.88 ⁇ 0.34* 6.76 ⁇ 0.26 6.79 ⁇ 0.34 6.85 ⁇ 0.28 6.98 ⁇ 0.24 Composition 35 6.85 ⁇ 0.22* 6.65 ⁇ 0.64 6.48 ⁇ 0.96 7.03 ⁇ 0.16 6.85 ⁇ 0.42 Composition 36 6.86 ⁇ 0.60* 6.98 ⁇ 0.26 6.67 ⁇ 0.51 6.95 ⁇
  • Table 7 shows that: 1 0.5 h to 4 h after administration, there was no significant prolongation of PT (prothrombin time) between the model group and the normal control group (p>0.05). Explain that the model was successful. 2 Compared with the model group, there was no significant prolongation of PT between 0.5h and 4h after administration in each drug-administered group (p>0.05). The results showed that there were no adverse reactions of bleeding in ginkgolide B, Shaban group and each composition group. 3 Compared with the model group, the PT (prothrombin time) of each composition group was significantly prolonged (p ⁇ 0.05) and significantly prolonged (p ⁇ 0.01), indicating that the pharmaceutical composition of the present invention can effectively prolong PT. And ginkgolide B and Shaban have a synergistic effect on the prolongation of PT.
  • composition 6 0.35 ⁇ 0.25*** 0.34 ⁇ 0.05*** 1.34 ⁇ 0.13*** 1.34 ⁇ 0.42*** 1.20 ⁇ 0.30*** Composition 7 0.34 ⁇ 0.09*** 0.29 ⁇ 0.08*** 1.22 ⁇ 0.23*** 1.35 ⁇ 0.32*** 1.28 ⁇ 0.40*** Composition 8 0.40 ⁇ 0.08*** 0.32 ⁇ 0.12*** 1.41 ⁇ 0.04*** 1.28 ⁇ 0.29*** 1.34 ⁇ 0.36*** Composition 9 0.46 ⁇ 0.04*** 0.24 ⁇ 0.21*** 1.03 ⁇ 0.21*** 1.30 ⁇ 0.18*** 1.26 ⁇ 0.31*** Composition 10 0.32 ⁇ 0.08*** 0.25 ⁇ 0.12*** 0.74 ⁇ 0.17*** 1.16 ⁇ 0.17*** 0.70 ⁇ 0.16*** Composition 11 0.25 ⁇ 0.05*** 0.29 ⁇ 0.17*** 1.21 ⁇ 0.06*** 1.31 ⁇ 0.15*** 1.12 ⁇ 0.07*** Composition 12 0.26 ⁇ 0.04*** 0.32 ⁇ 0.21*** 0.95 ⁇ 0.23*** 0.96 ⁇ 0.20*** 1.05 ⁇ 0.15*** Composition 13 0.45 ⁇ 0.09*** 0.35 ⁇ 0.20*** 1.05 ⁇ 0.22*** 1.21 ⁇ 0.11
  • Anticoagulant is effectively defined as plasma anti-Xa activity ⁇ 0.5IU/ML ("Calculation of low-molecular-weight heparin for cardiovascular disease" report: according to the pharmacokinetic characteristics of LMWH in vivo)
  • Table 8 shows that: 1 0.5 h to 4 h after administration, there was no abnormality in anti-clotting factor Xa activity between the model group and the normal control group (p>0.05). Explain that the model was successful. 2 Compared with the model group, the activity of anticoagulant factor Xa in ginkgolide B group increased with time from 0.5h to 4h after administration (P ⁇ 0.001), and both rivaroxaban and apixaban group had extreme Significant differences (p ⁇ 0.001) indicate that the anti-coagulant Xa activity of the class of Shaban is stronger than that of Ginkgolide B.
  • the anti-Xa activity of each single drug group was significantly increased (P ⁇ 0.05), and the anti-Xa activity of each composition group was significantly increased (P ⁇ 0.001), indicating that the pharmaceutical composition of the present invention can be
  • the activity of factor Xa is enhanced, and ginkgolide B has a synergistic effect with the anti-Xa activity of saban.
  • Table 9 shows that: 1 0.5 h to 4 h after administration, there was no significant difference in PAF (platelet activating factor) content between the model group and the normal control group (p>0.05). Explain that the model was successful. 2 Compared with the model group, the PAF content of Ginkgolide B group decreased significantly (P ⁇ 0.01) 2 h after administration, and the PAF content of rivaroxaban and apixaban group decreased significantly 4 h after administration (P ⁇ 0.001). The PAF content of the Rezashaban and Omishaban groups was significantly lower (P ⁇ 0.01).
  • the body has synergistic effect; 1 hour after administration, the PAF content of each composition group is significantly decreased (P ⁇ 0.001) and significantly decreased (P ⁇ 0.01), indicating that ginkgolides B and apixaban antagonize PAF receptor Synergistic effect; 2 hours after administration, the PAF content of each composition group was significantly decreased (P ⁇ 0.001) and significantly decreased (P ⁇ 0.01), indicating that ginkgolides B and apixaban antagonize PAF receptors The role of synergy.
  • compositions 1 to 8) of ginkgolides B+ rivaroxaban the groups 1 to 5 to 8 had longer aPPT, longer PT, higher activity of factor Xa, lower content of PAF, and combination.
  • the aPPT of the group 1 is the longest, the PT is the longest, and the Xa is the cause.
  • the highest activity and the lowest PAF content the ratio of ginkgolides B to rivaroxaban is preferably: ginkgolides B5-15 parts and rivaroxaban. 10-20 parts, most preferably: 10 parts of ginkgolides B and 15 parts of rivaroxaban.
  • composition group of Ginkgolide B + apixaban compositions 9 to 16
  • the platelets of Groups 9 and 13 to 16 had longer aPPT, longer PT, higher activity of factor Xa, and lower content of PAF.
  • the composition of group 9 has the longest aPPT, the longest PT, the highest activity of factor Xa and the lowest content of PAF. Therefore, the ratio of ginkgolide B to apixaban is preferred in the composition of ginkgolide B+ apixaban. It is: bilobalide B5-15 parts and apixaban 1-10 parts, most preferably: ginkgolides B10 parts and apixaban 5 parts.
  • compositions 17-24 In the composition group of Ginkgolide B + edoxaban (compositions 17-24), the composition 17 and 22-24 had longer aPPT, longer PT, higher activity of factor Xa, lower content of PAF, and combination.
  • the 17th group had the longest aPPT, the longest PT, the highest activity of factor Xa and the lowest content of PAF. Therefore, the ratio of ginkgolides B to edoxaban was better in the composition of ginkgolides B+ydoxaban. : Ginkgolide B5-15 parts and idoxaban 10-30 parts, most preferably: ginkgolides B10 parts and edoxaban 15 parts.
  • compositions 25-29 In the composition group of Ginkgolide B + Rezashaban (compositions 25-29), the composition 25, 28-29 group has longer aPPT, longer PT, higher activity of factor Xa, lower content of PAF, combination
  • the 25th group had the longest aPPT, the longest PT, the highest activity of factor Xa and the lowest content of PAF. Therefore, the ratio of ginkgolides B to rezashaban was better in the composition of ginkgolides B + rezashaban. : Ginkgolide B5-15 parts and Rezazaban 5-15 parts, most preferably: 10 parts of ginkgolides B and 10 parts of Rezazaban.
  • composition 30-41 the composition 30, 38-41 group has longer aPPT, longer PT, higher activity of factor Xa, lower content of PAF, combination
  • the 30th group had the longest aPPT, the longest PT, the highest activity of factor Xa and the lowest content of PAF. Therefore, the ratio of ginkgolides B to omeproxaban was better in the composition of ginkgolides B+omixampan.
  • the experimental results show that the present invention combines ginkgolide B and Shaban active ingredients, such as rivaroxaban, apixaban, edoxaban, rezashaban, and omeproxaban.
  • aPPT significantly prolonged
  • PT significantly prolonged
  • Xa factor activity was significantly enhanced
  • PAF content was significantly reduced
  • the composition, aPPT length, PT length, high factor Xa activity, low PAF content, can prevent platelet aggregation through different mechanisms of action.
  • the present invention combines ginkgolide B with a banban drug, and the two can exert synergistic effects.
  • the pharmaceutical composition comprising the ginkgolides B and the banban can effectively inhibit platelet aggregation. The effect is excellent and the clinical application prospect is excellent.

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Abstract

一种含银杏内酯B和Xa因子抑制剂沙班类药物的药物组合物，以及该药物组合物的制备方法和用途。其中，所述的沙班类药物选自利伐沙班、阿哌沙班、依度沙班、雷扎沙班和奥米沙班。

Description

一种含有银杏内酯B和Xa因子抑制剂的药物组合物及其制备方法和用途 技术领域
本发明涉及一种含银杏内酯B和Xa因子抑制剂的药物组合物。
背景技术
近年来，新型抗凝药物Ⅹa因子抑制剂的问世，开创了抗凝治疗的新方向。Ⅹa因子抑制剂在增加抗凝效果的同时，降低大出血危险性，无需血液监测、便于长期治疗。研究证实，Ⅹa因子在控制凝血酶形成和活化凝血瀑布中占有重要地位，在凝血瀑布的上游抑制凝血因子将产生更强的抗凝作用。Ⅹa因子不能活化血小板，对凝血瀑布也不存在正反馈作用。Ⅹa因子直接抑制剂既能抑制与血栓结合的因子也能抑制与前凝血酶结合的因子。
利伐沙班作为首个Ⅹa因子抑制剂已正式上市。大量临床研究结果显示，利伐沙班在静脉血栓栓塞的预防和治疗等领域疗效优于目前的标准治疗药物依诺肝素钠。临床上可用于预防髋关节和膝关节置换术后患者深静脉血栓(DVT)和肺栓塞(PE)的形成，也可用于预防非瓣膜性心房纤颤患者脑卒中和非中枢神经系统性栓塞，降低冠状动脉综合症复发的风险等。但由于利伐沙班的药效学性质，用药过量可能导致出血并发症，且一些亚群的患者的出血风险较高，暂时尚无对抗利伐沙班药效的特异性解毒剂。
阿哌沙班作为一种口服的选择性活化Ⅹa因子抑制剂，是雷扎沙班的变异体，由辉瑞与百时美施贵宝联合开发，已在美国和欧盟上市，其能预防血栓，用于接受过髋部或膝部置换手术患者的血栓预防。但在临床试验研究中发现，在发生急性冠脉综合征患者中，研究者观察到抗血小板治疗基础上加用阿哌沙班可引起剂量依赖的出血事件增加以及缺血事件减少的趋势。阿哌沙班的安全性及有效性可能依赖于基础抗血小板治疗。
依度沙班是日本第一三共株式会社研制的小分子口服抗凝药，为凝血因子X(FXa)阻滞剂。主要用于接受全膝关节置换术(TKA)、全髋关节置换术(THA)、髋关节骨折手术(HFS)患者并发静脉血栓栓塞(VTE)。凝血过程中，活化的凝血因子X(FXa)将凝血酶原(FII)激活成为凝血酶(FIIa)，促使纤维蛋白形成，由此形成血栓，因而FXa已成为开发新一代抗凝药物的主要靶点。依度沙班通过选择性、可逆性且直接抑制FXa达到抑制血栓形成的口服抗凝药物，其对FXa的选择性比FIIa高104倍。日本国内外临床试验均证实本品可有效抑制接受下肢整形外科手术患者并发VTE，且安全可靠。但是该药也存在一个费用相对较高的局限性，有可能会成为大多数患者治疗 的绊脚石。
雷扎沙班的相对分子质量为528，作为一种口服有效肽类似物，它可以和因子Xa具有高度的亲和力，一项II期临床研究显示其对预防矫形外科手术后静脉血栓栓塞(VTE)是有效的。这项随机的双盲研究有656例进行全膝关节置换术的病人参加，有效的438例病人中，雷扎沙班与VTE发生率有明显的剂量效应关系。值得注意的是雷扎沙班25mg组的VTE发生率显著低于依诺肝素组，大出血发生率与依诺肝素组相似。然而在雷扎沙班50～100mg组中大出血几率比依诺肝素组高。
奥米沙班是一种注射用的高选择性并且可逆的凝血酶因子Xa抑制剂，它通过一种剂量依赖的方式抑制血栓形成，起效和失效比较迅速，药代动力学遵循线性规律，不易经肾脏消除。此前有研究表明，活化X因子抑制剂治疗ACS有血栓形成风险，因此PCI术中应注意调整这些抗凝药物以及其他加用抗血栓因子的剂量，但是应用中等剂量的奥米沙班似乎并不用担心这些因素。此外，考虑到奥米沙班起效失效迅速、静脉给药、可预测的抗凝反应(无需监测)等优势，奥米沙班可能是NSTE-ACS患者从急诊到介入治疗的单一抗凝药物，且很可能具有很大优势。另有报道称：奥米沙班经肾脏排泄的药量不足25％，因此用于肾功能损害患者时无需调整剂量。也有报道称，奥米沙班不能降低缺血性事件发生风险且可增加出血风险，并不支持对早起行PCI手术的NSTE-ACS患者使用奥米沙班。
因此，如何在保证治疗效果的情况下，减少沙班类药物的大量使用以降低副作用、降低成本亟待解决。
发明内容
本发明的目的在于克服现有Xa因子抑制剂中的高出血风险及其他的缺陷，使患者具有更多选择而研制出的一种全新的药物组合物。
本发明提供了一种含银杏内酯B的药物组合物，它含有银杏内酯B和Xa因子抑制剂。
其中，所述的Xa因子抑制剂为沙班类药物。
其中，所述的沙班类药物为利伐沙班、阿哌沙班、依度沙班、雷扎沙班和/或奥米沙班。
所述利伐沙班结构式为：
Figure PCTCN2015093305-appb-000001
所述阿哌沙班结构式为：
Figure PCTCN2015093305-appb-000002
所述依度沙班结构式为：
Figure PCTCN2015093305-appb-000003
所述雷扎沙班结构式为：
Figure PCTCN2015093305-appb-000004
所述奥米沙班结构式为：
Figure PCTCN2015093305-appb-000005
其中，所述银杏内酯B与利伐沙班的重量配比为：银杏内酯B1-20份和利伐沙班5-40份。
优选地，二者的重量配比为：银杏内酯B5-15份和利伐沙班10-20份。进一步优选地，二者的重量配比为：银杏内酯B10份和利伐沙班15份。
或者，所述银杏内酯B与阿哌沙班的重量配比为：银杏内酯B1-20份和 阿哌沙班0.5-20份。
优选地，二者的重量配比为：银杏内酯B5-15份和阿哌沙班1-10份。进一步优选地，二者的重量配比为：银杏内酯B10份和阿哌沙班5份。
或者，所述银杏内酯B与依度沙班的重量配比为：银杏内酯B1-20份和依度沙班8-50份。
优选地，二者的重量配比为：银杏内酯B5-15份和依度沙班10-30份。进一步优选地，二者的重量配比为：银杏内酯B10份和依度沙班15份。
或者，所述银杏内酯B与雷扎沙班的重量配比为：银杏内酯B1-20份和雷扎沙班1-20份。
优选地，二者的重量配比为：银杏内酯B5-15份和雷扎沙班5-15份。进一步优选地，二者的重量配比为：银杏内酯B10份和雷扎沙班10份。
或者，所述银杏内酯B与奥米沙班的重量配比为：银杏内酯B1-20份和奥米沙班15-150份。
优选地，二者的重量配比为：银杏内酯B5-15份和奥米沙班30-125份。再优选地，二者的重量配比为：银杏内酯B8-12份和奥米沙班50-90份。进一步优选地，二者的重量配比为：银杏内酯B10份和奥米沙班60份。
本发明还提供了一种制备所述的药物组合物的方法，它包括以下步骤：
S1：按组份及重量比称取原料银杏内酯B和沙班类药物；
S2：将原料混合后，加入药学上可接受的辅料制备成药学上常用的药物制剂。
其中，所述的药学上可接受的辅料包括：淀粉、预胶化淀粉、乳糖、蔗糖、滑石粉、糊精、环糊精、微晶纤维素、交联羧甲基纤维素钠、羧甲基淀粉钠、低取代羟丙基纤维素、交联聚维酮、葡萄糖、葡甲胺、硬脂酸镁、右旋糖酐、甘油、乙醇、丙二醇、聚乙二醇、甘露醇、山梨醇、木糖醇、纤维植物油、苯甲酸钠、水杨酸钠、盐酸、枸橼酸、枸橼酸纳、磷酸二氢钠、磷酸氢二钠、明胶、卵磷脂、维生素C中的一种或是几种。
其中，所述的药物制剂包括：片剂、胶囊剂、软胶囊、口服液、颗粒剂、丸剂、滴丸剂、散剂、膏剂、丹剂、注射剂、栓剂、贴剂、滴剂、喷雾剂、霜剂、混悬剂、酊剂、乳剂、水针剂、粉针剂、靶向制剂、缓释制剂、控释制剂。
本发明还提供了银杏内酯B与Xa因子抑制剂联合用药在制备抗血小板聚集药物中的应用。
其中，所述的Xa因子抑制剂为沙班类药物。
其中，所述的沙班类药物为利伐沙班、阿哌沙班、依度沙班、雷扎沙班和/或奥米沙班。
其中，所述银杏内酯B与利伐沙班的重量配比为：银杏内酯B1-20份和利伐沙班5-40份。
优选地，二者的重量配比为：银杏内酯B5-15份和利伐沙班10-20份。进一步优选地，二者的重量配比为：银杏内酯B10份和利伐沙班15份。
或者，其中所述银杏内酯B与阿哌沙班的重量配比为：银杏内酯B1-20份和阿哌沙班0.5-20份。
优选地，二者的重量配比为：银杏内酯B5-15份和阿哌沙班1-10份。进一步优选地，二者的重量配比为：银杏内酯B10份和阿哌沙班5份。
或者，所述银杏内酯B与依度沙班的重量配比为：银杏内酯B1-20份和依度沙班8-50份。
优选地，二者的重量配比为：银杏内酯B5-15份和依度沙班10-30份。进一步优选地，二者的重量配比为：银杏内酯B10份和依度沙班15份。
或者，所述银杏内酯B与雷扎沙班的重量配比为：银杏内酯B1-20份和雷扎沙班1-20份。
优选地，二者的重量配比为：银杏内酯B5-15份和雷扎沙班5-15份。进一步优选地，二者的重量配比为：银杏内酯B10份和雷扎沙班10份。
或者，所述银杏内酯B与奥米沙班的重量配比为：银杏内酯B1-20份和奥米沙班15-150份。
优选地，二者的重量配比为：银杏内酯B5-15份和奥米沙班30-125份。再优选地，二者的重量配比为：银杏内酯B8-12份和奥米沙班50-90份。进一步优选地，二者的重量配比为：银杏内酯B10份和奥米沙班60份。
本发明提供的药物组合物，因其中含有银杏内酯B和沙班类活性成分，可以通过不同的作用机制抗血小板聚集，能使利伐沙班、阿哌沙班、依度沙班、雷扎沙班、奥米沙班等沙班类药物的抗血小板聚集功能和银杏内酯B的抗血小板聚集功能增强，两者具有协同增效的作用，临床使用可以减少沙班类药物的药量、增加药效，降低成本，减少副作用，为临床研究提供了更好的选择。本发明药物组合物配方新颖、组分简单、作用机制明确、且疗效显著，可实现工业化大生产。
本发明将银杏内酯B与沙班类药物联合使用，二者可以发挥协同增效作用，抑制血小板聚集的效果优良。
本发明将银杏内酯B与沙班类药物联合使用，二者可以发挥协同增效作用，抑制血小板聚集的效果优良，临床应用时可以降低沙班类药物的用量，进而减少其大量使用带来的副作用，还可以降低成本，临床应用前景优良。
以下通过实施例形式的具体实施方式，对本发明的上述内容作进一步的详细说明。但不应将此理解为本发明上述主题的范围仅限于以下的实施例。 凡基于本发明上述内容所实现的技术均属于本发明的范围。
具体实施方式
本发明所述的银杏内酯B单体化合物，可以通过购买市售产品得到，或通过将银杏内酯在现有方法上进行分离纯化制得；利伐沙班、阿哌沙班、依度沙班、雷扎沙班、奥米沙班等亦可通过购买市售产品得到，亦或通过现有方法合成而得。经检验，所有单体化合物均与相应对照品结构相符，且经HPLC检测其纯度均在95％以上。
实施例1
银杏内酯B 10份
利伐沙班15份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成片剂。
实施例2
银杏内酯B 5份
利伐沙班20份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成胶囊剂或软胶囊剂。
实施例3
银杏内酯B 15份
利伐沙班10份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成丸剂。
实施例4
银杏内酯B 1份
利伐沙班5份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成口服液。
实施例5
银杏内酯B 20份
利伐沙班40份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成滴丸剂。
实施例6
银杏内酯B 10份
阿哌沙班5份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成喷雾剂。
实施例7
银杏内酯B 5份
阿哌沙班1份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成水针剂或粉针剂。
实施例8
银杏内酯B 15份
阿哌沙班10份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成缓释制剂或控释制剂。
实施例9
银杏内酯B 20份
阿哌沙班20份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成靶向制剂。
实施例10
银杏内酯B 10份
依度沙班15份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成颗粒剂或混悬剂。
实施例11
银杏内酯B 5份
依度沙班30份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成片剂。
实施例12
银杏内酯B 15份
依度沙班10份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成胶囊剂或软胶囊剂。
实施例13
银杏内酯B 1份
依度沙班8份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成丸剂。
实施例14
银杏内酯B 20份
依度沙班50份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成口服液。
实施例15
银杏内酯B 10份
雷扎沙班10份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成滴丸剂。
实施例16
银杏内酯B 5份
雷扎沙班15份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成喷雾剂。
实施例17
银杏内酯B 15份
雷扎沙班5份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成水针剂或粉针剂。
实施例18
银杏内酯B 1份
雷扎沙班20份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成缓释制剂或控释制剂。
实施例19
银杏内酯B 20份
雷扎沙班1份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成靶向制剂。
实施例20
银杏内酯B 10份
奥米沙班60份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成颗粒剂或混悬剂。
实施例21
银杏内酯B 8份
奥米沙班50份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成口服液。
实施例22
银杏内酯B 12份
奥米沙班90份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成滴丸剂。
实施例23
银杏内酯B 15份
奥米沙班125份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成喷雾剂。
实施例24
银杏内酯B 1份
奥米沙班15份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成水针剂或粉针剂。
实施例25
银杏内酯B 20份
奥米沙班125份
药学上可接受的常规辅料
将上述原料混合后，加入药学上可接受的常规辅料按照常规工艺制备成缓释制剂或控释制剂。
以下用试验例的方式来说明本发明的有益效果：
试验例1银杏内酯B和沙班类组合在体外抗PAF诱导血小板聚集作用及对Xa活性影响
一、实验目的：
采用微量反应板酶标仪比浊法，评价成都百裕科技制药有限公司生产的银杏内酯B和沙班类组合在体外抗PAF诱导血小板聚集作用及对Xa活性影响。
二、材料与方法
2.1实验动物
日本大耳白兔，SPF级，雌雄各半，体重2.2-2.5kg。
2.2实验药物
银杏内酯B(成都百裕科技制药有限公司)、利伐沙班(成都百裕科技制药有限公司)、阿哌沙班(成都百裕科技制药有限公司)、依度沙班(成都百裕科技制药有限公司)、雷扎沙班(成都百裕科技制药有限公司)、奥米沙班(成都百裕科技制药有限公司)、组合物1(银杏内酯B：利伐沙班＝10:15)、组合物2(银杏内酯B：利伐沙班＝1:5)、组合物3(银杏内酯B：利伐沙班＝1:40)、组合物4(银杏内酯B：利伐沙班＝20:5)、组合物5(银杏内酯B：利伐沙班＝20:40)、组合物6(银杏内酯B：利伐沙班＝5:20)、组合物7(银杏内 酯B：利伐沙班＝15:10)、组合物8(银杏内酯B：利伐沙班＝15:20)、组合物9(银杏内酯B：阿哌沙班＝10:5)、组合物10(银杏内酯B：阿哌沙班＝1:20)、组合物11(银杏内酯B：阿哌沙班＝20：0.5)、组合物12(银杏内酯B：阿哌沙班＝20:20)、组合物13(银杏内酯B:阿哌沙班＝5:1)、组合物14(银杏内酯B：阿哌沙班＝5:10)、组合物15(银杏内酯B:阿哌沙班＝15:1)、组合物16(银杏内酯B:阿哌沙班＝15:10)、组合物17(银杏内酯B：依度沙班＝10:15)、组合物18(银杏内酯B：依度沙班＝1:8)、组合物19(银杏内酯B：依度沙班＝1：50)、组合物20(银杏内酯B：依度沙班＝20:8)、组合物21(银杏内酯B：依度沙班＝20:50)、组合物22(银杏内酯B：依度沙班＝5:10)、组合物23(银杏内酯B：依度沙班＝15:10)、组合物24(银杏内酯B：依度沙班＝5:30)、组合物25(银杏内酯B：雷扎沙班＝10:10)、组合物26(银杏内酯B：雷扎沙班＝1:20)、组合物27(银杏内酯B：雷扎沙班＝20:1)、组合物28(银杏内酯B：雷扎沙班＝5:15)、组合物29(银杏内酯B：雷扎沙班＝15:5)、组合物30(银杏内酯B：奥米沙班＝10:60)、组合物31(银杏内酯B：奥米沙班＝1:15)、组合物32(银杏内酯B：奥米沙班＝1:150)、组合物33(银杏内酯B：奥米沙班＝20:15)、组合物34(银杏内酯B：奥米沙班＝20:150)、组合物35(银杏内酯B：奥米沙班＝15:30)、组合物36(银杏内酯B：奥米沙班＝5:125)、组合物37(银杏内酯B：奥米沙班＝15:125)、组合物38(银杏内酯B：奥米沙班＝8:50)、组合物39(银杏内酯B：奥米沙班＝8:90)、组合物40(银杏内酯B：奥米沙班＝12:50)、组合物41(银杏内酯B：奥米沙班＝12:90)。
2.3试剂和仪器
仪器：
德国Sartorius BS124S型电子天平(分度值：0.0001g)，仪器编号：BKY-YB-001；ACS-15LED电子台秤，上海鹰牌衡器有限公司出品，仪器编号BKY-YB-002；长沙湘智DL-5M小立式低速大容量冷冻离心机，仪器编号：BKY-ZX-040，长沙湘智离心机仪器有限公司；海尔DW-40L188低温保存箱，仪器编号：BKY-ZX-012，保存血浆等；LSC-316C型星星立式陈列柜，仪器编号：BKY-ZX-041，保存试剂；Multiskan MK3型酶标仪，仪器编号：BKY-ZX-014；Wellwash 4MK2型洗板机，仪器编号：BKY-ZX-022；德国Brand
Figure PCTCN2015093305-appb-000006
S,D-1000微量移液器，仪器编号：BKY-ZX-036；德国Brand
Figure PCTCN2015093305-appb-000007
S,D-100微量移液器，仪器编号：BKY-ZX-038；MB-1830型全自动血液分析仪，四川美生科技有限公司，仪器编号：BKY-ZX-023。
试剂：
3.8％的枸橼酸钠(500g/瓶，成都市科龙化工试剂厂，批号：20130601)；血小板活化因子(PAF)(1mg/支，Sigma，批号：P7568)；抗血凝因子(Xa)测定试剂(发色底物法)(10×71nkat，意大利Chromogenix，批号：N1243555)；聚乙二醇(PEG)6000(50g，北京百灵威科技有限公司，批号：LB50P76)；肝素钠(1g/瓶，Bomei,批号：9041-08-1)。
2.4剂量设计和药物配制方法
①各组以体内试验建议给药浓度作为基础，根据不同药物比例，进行血小板聚集试验，具体见表1：
表1药物剂量设计
组别 给药剂量 每日给药次数
生理盐水 2ml 1
银杏内酯B 5.0mg/kg 1
利伐沙班组 5.0mg/kg 1
阿哌沙班组 5.0mg/kg 1
依度沙班组 5.0mg/kg 1
雷扎沙班组 5.0mg/kg 1
奥米沙班组 5.0mg/kg 1
组合物1组 5.0mg/kg 1
组合物2组 5.0mg/kg 1
组合物3组 5.0mg/kg 1
组合物4组 5.0mg/kg 1
组合物5组 5.0mg/kg 1
组合物6组 5.0mg/kg 1
组合物7组 5.0mg/kg 1
组合物8组 5.0mg/kg 1
组合物9组 5.0mg/kg 1
组合物10组 5.0mg/kg 1
组合物11组 5.0mg/kg 1
组合物12组 5.0mg/kg 1
组合物13组 5.0mg/kg 1
组合物14组 5.0mg/kg 1
组合物15组 5.0mg/kg 1
组合物16组 5.0mg/kg 1
组合物17组 5.0mg/kg 1
组合物18组 5.0mg/kg 1
组合物19组 5.0mg/kg 1
组合物20组 5.0mg/kg 1
组合物21组 5.0mg/kg 1
组合物22组 5.0mg/kg 1
组合物23组 5.0mg/kg 1
组合物24组 5.0mg/kg 1
组合物25组 5.0mg/kg 1
组合物26组 5.0mg/kg 1
组合物27组 5.0mg/kg 1
组合物28组 5.0mg/kg 1
组合物29组 5.0mg/kg 1
组合物30组 5.0mg/kg 1
组合物31组 5.0mg/kg 1
组合物32组 5.0mg/kg 1
组合物33组 5.0mg/kg 1
组合物34组 5.0mg/kg 1
组合物35组 5.0mg/kg 1
组合物36组 5.0mg/kg 1
组合物37组 5.0mg/kg 1
组合物38组 5.0mg/kg 1
组合物39组 5.0mg/kg 1
组合物40组 5.0mg/kg 1
组合物41组 5.0mg/kg 1
2.5检测血小板聚集率
动物心脏穿刺取血，枸橼酸钠(3.8％)1:9抗凝，以4℃×1000r/min离心10min，收集富血小板血浆(PRP)，剩余部分以3000r/min离心15min，收集贫血小板血浆(PPP)，用PPP将PRP的调整为200±50×109/L。
在微量反应板小孔中加入250μl的PRP，分别加入10μl对应浓度药物，每个浓度平行做两孔，最后再加入PAF致聚剂(浓度为0.38μg/ml)10μl；空白对照孔加入250μl的PPP和10μl的生理盐水调零；溶媒对照孔加入250μl的PPP和10μl的二甲基亚砜调零。记录不同时间的吸光度，直到吸光度不再下降为止。此时的时间即为血小板最大聚集所需时间，然后利用以下公式计算血小板的抑制率：
聚集抑制率＝(给药组最大聚集所需时间-空白对照组最大聚集所需时间)/空白对照组最大聚集所需时间×100％
2.6测试抗血凝因子Xa的活性
采用发色底物法，按照试剂盒说明书和参考文献在酶标仪上测定血浆抗Xa活性。
2.7统计学处理
试验数据为计量资料，根据对应计算公式计算聚集抑制率与抗Xa活性，并采用Excel软件计算平均数。
三、实验结果
3.1血小板聚集实验结果
表2银杏内酯B和沙班类组合在体外抗PAF诱导血小板聚集作用(n＝2)
组别 血小板最大聚集时间(min) 血小板聚集抑制率(％)
生理盐水 20 -
银杏内酯B 30 50
利伐沙班组 25 25
阿哌沙班组 40 100
依度沙班组 30 80
雷扎沙班组 35 75
奥米沙班组 40 100
组合物1组 60 200
组合物2组 30 50
组合物3组 35 75
组合物4组 33 65
组合物5组 55 175
组合物6组 50 150
组合物7组 56 180
组合物8组 59 195
组合物9组 88 340
组合物10组 44 120
组合物11组 48 140
组合物12组 45 125
组合物13组 60 200
组合物14组 65 225
组合物15组 68 240
组合物16组 75 275
组合物17组 80 300
组合物18组 50 150
组合物19组 60 200
组合物20组 65 225
组合物21组 66 230
组合物22组 76 280
组合物23组 79 295
组合物24组 73 265
组合物25组 60 200
组合物26组 36 80
组合物27组 40 100
组合物28组 55 175
组合物29组 58 190
组合物30组 90 350
组合物31组 50 150
组合物32组 55 175
组合物33组 52 160
组合物34组 65 225
组合物35组 62 210
组合物36组 60 200
组合物37组 58 190
组合物38组 79 295
组合物39组 85 325
组合物40组 80 300
组合物41组 83 315
由表2可以看出，在PAF诱导下，各个组合物组的血小板聚集率与生理盐水组相比，均发生显著性性差异(p<0.01，p＜0.05)，说明本发明各个药物组合物均可以有效抗血小板聚集；与银杏内酯B与沙班类药物单用相比，本发明将银杏内酯B与沙班类药物联用时，抗血小板聚集功能显著提高，说明银杏内酯B与沙班类药物联用具有协同增效的作用。
3.2抗血凝因子Xa活性实验结果
表3银杏内酯B和沙班类组合在体外抗血凝因子Xa活性
组别 抗Xa活性(IU/ml) 组别 抗Xa活性(IU/ml)
生理盐水 0 组合物18组 1.01
银杏内酯B 0.57 组合物19组 0.98
利伐沙班组 0.54 组合物20组 1.10
阿哌沙班组 1.07 组合物21组 0.85
依度沙班组 0.79 组合物22组 1.39
雷扎沙班组 0.86 组合物23组 1.40
奥米沙班组 1.03 组合物24组 1.35
组合物1组 1.53 组合物25组 1.52
组合物2组 0.95 组合物26组 0.96
组合物3组 1.06 组合物27组 0.91
组合物4组 0.96 组合物28组 1.46
组合物5组 1.47 组合物29组 1.39
组合物6组 1.04 组合物30组 1.56
组合物7组 1.39 组合物31组 1.11
组合物8组 1.45 组合物32组 0.96
组合物9组 1.55 组合物33组 1.89
组合物10组 0.87 组合物34组 0.98
组合物11组 0.92 组合物35组 1.15
组合物12组 1.11 组合物36组 1.06
组合物13组 1.50 组合物37组 1.17
组合物14组 1.38 组合物38组 1.49
组合物15组 1.45 组合物39组 1.52
组合物16组 1.52 组合物40组 1.38
组合物17组 1.45 组合物41组 1.50
备注：抗凝有效定义为血浆抗Xa活性≧0.5IU/ML(《心血管疾病人应用低分子肝素要规范化》报道：根据LMWH体内的药代动力学特点制定的。
由表3可以看出，各个组合物组的抗血凝因子Xa活性与生理盐水组相比，均差异显著，说明本发明各个药物组合物均可以有效抑制Xa因子活性、抗血小板聚集；与银杏内 酯B与沙班类药物单用相比，本发明将银杏内酯B与沙班类药物联用时，体外抗血凝因子Xa活性显著提高，说明银杏内酯B与沙班类药物联用具有协同增效的作用。
由表2和表3可以看出，银杏内酯B+利伐沙班的组合物组(组合物1～8)中，组合物1、5～8组的血小板聚集抑制率和Xa因子活性相对较高，组合物1组的血小板聚集抑制率和Xa因子活性相对最高，因此，银杏内酯B+利伐沙班的组合物中，银杏内酯B与利伐沙班的配比优选为：银杏内酯B 5-15份和利伐沙班10-20份，最优选为：银杏内酯B 10份和利伐沙班15份。
银杏内酯B+阿哌沙班的组合物组(组合物9～16)中，组合物9、13～16组的血小板聚集抑制率和Xa因子活性相对较高，组合物9组的血小板聚集抑制率和Xa因子活性最高，因此，银杏内酯B+阿哌沙班的组合物中，银杏内酯B与阿哌沙班的配比优选为：银杏内酯B 5-15份和阿哌沙班1-10份，最优选为：银杏内酯B 10份和阿哌沙班5份。
银杏内酯B+依度沙班的组合物组(组合物17～24)中，组合物17、22～24组的血小板聚集抑制率和Xa因子活性相对较高，组合物17组的血小板聚集抑制率和Xa因子活性最高，因此，银杏内酯B+依度沙班的组合物中，银杏内酯B与依度沙班的配比优选为：银杏内酯B 5-15份和依度沙班10-30份，最优选为：银杏内酯B 10份和依度沙班15份。
银杏内酯B+雷扎沙班的组合物组(组合物25～29)中，组合物25、28～29组的血小板聚集抑制率和Xa因子活性相对较高，组合物25组的血小板聚集抑制率和Xa因子活性最高，因此，银杏内酯B+雷扎沙班的组合物中，银杏内酯B与雷扎沙班的配比优选为：银杏内酯B 5-15份和雷扎沙班5-15份，最优选为：银杏内酯B 10份和雷扎沙班10份。
银杏内酯B+奥米沙班的组合物组(组合物30～41)中，组合物30、38～41组的血小板聚集抑制率和Xa因子活性相对较高，组合物30组的血小板聚集抑制率和Xa因子活性最高，因此，银杏内酯B+奥米沙班的组合物中，银杏内酯B与奥米沙班的配比优选为：银杏内酯B 8-12份和奥米沙班50-90份，最优选为：银杏内酯B 10份和奥米沙班10份。
实验结果说明，本发明将银杏内酯B和沙班类活性成分，如，利伐沙班、阿哌沙班、依度沙班、雷扎沙班、奥米沙班等沙班类药物联合使用，抗血小板聚集功能明显增强，Xa因子活性明显增加，且优于二者单独使用，二者联合使用发挥了协同增效作用，银杏内酯B和沙班类活性成分组成的组合物，抗血小板聚集功能强，Xa因子活性高，可以通过不同的作用机制抗血小板聚集。
试验例2银杏内酯B和沙班类在兔创伤性肢体深静脉血栓形成模型的抗凝作用与抗凝血因子活性作用比较研究
一、实验目的：
采比较银杏内酯B和沙班类组合在兔创伤性肢体深静脉血栓形成模型的抗凝作用与抗凝血因子活性作用差异。
二、材料与方法
2.1实验动物
选用Ⅰ级日本大耳白家兔384只，每组8只，试验时体重2.0～3.0kg，全雄性。
2.2实验药物
银杏内酯B(成都百裕科技制药有限公司)、利伐沙班(成都百裕科技制药有限公司)、阿哌沙班(成都百裕科技制药有限公司)、依度沙班(成都百裕科技制药有限公司)、雷扎沙班(成都百裕科技制药有限公司)、奥米沙班(成都百裕科技制药有限公司)、组合物1(银杏内酯B：利伐沙班＝10:15)、组合物2(银杏内酯B：利伐沙班＝1:5)、组合物3(银杏内酯B：利伐沙班＝1:40)、组合物4(银杏内酯B：利伐沙班＝20:5)、组合物5(银杏内酯B：利伐沙班＝20:40)、组合物6(银杏内酯B：利伐沙班＝5:20)、组合物7(银杏内酯B：利伐沙班＝15:10)、组合物8(银杏内酯B：利伐沙班＝15:20)、组合物9(银杏内酯B：阿哌沙班＝10:5)、组合物10(银杏内酯B：阿哌沙班＝1:20)、组合物11(银杏内酯B：阿哌沙班＝20：0.5)、组合物12(银杏内酯B：阿哌沙班＝20:20)、组合物13(银杏内酯B:阿哌沙班＝5:1)、组合物14(银杏内酯B：阿哌沙班＝5:10)、组合物15(银杏内酯B:阿哌沙班＝15:1)、组合物16(银杏内酯B:阿哌沙班＝15:10)、组合物17(银杏内酯B：依度沙班＝10:15)、组合物18(银杏内酯B：依度沙班＝1:8)、组合物19(银杏内酯B：依度沙班＝1：50)、组合物20(银杏内酯B：依度沙班＝20:8)、组合物21(银杏内酯B：依度沙班＝20:50)、组合物22(银杏内酯B：依度沙班＝5:10)、组合物23(银杏内酯B：依度沙班＝15:10)、组合物24(银杏内酯B：依度沙班＝5:30)、组合物25(银杏内酯B：雷扎沙班＝10:10)、组合物26(银杏内酯B：雷扎沙班＝1:20)、组合物27(银杏内酯B：雷扎沙班＝20:1)、组合物28(银杏内酯B：雷扎沙班＝5:15)、组合物29(银杏内酯B：雷扎沙班＝15:5)、组合物30(银杏内酯B：奥米沙班＝10:60)、组合物31(银杏内酯B：奥米沙班＝1:15)、组合物32(银杏内酯B：奥米沙班＝1:150)、组合物33(银杏内酯B：奥米沙班＝20:15)、组合物34(银杏内酯B：奥米沙班＝20:150)、组合物35(银杏内酯B：奥米沙班＝15:30)、组合物36(银杏内酯B：奥米沙班＝5:125)、组合物37(银杏内酯B：奥米沙班＝15:125)、组合物38(银杏内酯B：奥米沙班＝8:50)、组合物39(银杏内酯B：奥米沙班＝8:90)、组合物40(银杏内酯B：奥米沙班＝12:50)、组合物41(银杏内酯B：奥米沙班＝12:90)。
2.3试剂和仪器
仪器：
(1)家兔称重检查仪器
ACS-15LED电子台秤，分辨值1g，最大称量6kg，仪器编号BKY-YB-002，上海鹰牌衡器有限公司出品，用于称家兔体重。
(2)凝血功能及凝血因子活性测定仪器
日本希森美康CA-660型全自动血凝分析仪，仪器编号：BKY-ZX-044，希森美康生物科技(无锡)有限公司。
DL-5M低速冷冻离心机，仪器编号：BKY-ZX-040，长沙湘智离心机仪器有限公司生产。
BC-117F型冷藏箱，仪器编号：BKY-ZX-015，青岛海尔股份有限公司生产。
(3)PAF含量测定仪器
Multiskan MK3型酶标仪，仪器编号:BKY-ZX-014，美国Thermo electroncorporation。
(4)病理检查仪器
德国Sartorius BS124S型电子天平(分度值：0.0001g)，仪器编号：BKY-TJ-002，德国Sartorius股份有限公司生产。
ZT-14V2生物组织自动脱水机，仪器编号：BKY-BL-016，孝感市亚光医用电子技术有限公司出品。
常州中威BMJ-Ⅲ型包埋机，仪器编号：BKY-BL-002，江苏省常州市中威电子仪器厂出品。
常州中威PHY-Ⅲ病理组织漂烘仪，仪器编号：BKY-BL-003，江苏省常州市中威电子仪器厂出品。
徕卡RM2126轮转式切片机，仪器编号：BKY-BL-004，上海徕卡仪器有限公司出品。
日本奥林巴斯BX41-32P01显微镜，仪器编号：BKY-BL-013，日本奥林巴斯株式会社出品。
YR-21生物组织自动染色机，仪器编号：BKY-BL-015，孝感市亚光医用电子技术有限公司出品。
北京永光明202-0型台式干燥箱，仪器编号：BKY-BL-005，北京市永光明医疗仪器厂出品。
试剂：
(1)供试品配液所需试剂
羧丙甲纤维素(由委托方提供)。
乳糖，规格：100g/瓶，批号：2015041701，失效期：2018.06，成都市科龙化工试剂厂。甘露醇，规格：500g/瓶，批号：201402201，失效期：2017.06，成都市科龙化工试剂厂。
(2)凝血功能及凝血因子活性测定试剂
凝血质控品，规格：1ml×10瓶/盒，批号：528167B，失效期：2017.4.27，SIEMENS。
PT试剂，规格：2ml×10瓶/盒，批号：R5003，R5005，失效期：2017.1.1,2017.3.1，希森美康生物技术有限公司生产。
aPTT试剂，规格:2ml×10瓶/盒，批号：R5001，R5006，失效期：2016.9.1，2016.12.1，希森美康生物技术有限公司生产。
抗血凝因子(Xa)测定试剂盒(发色底物法)，规格：10×71nkat，批号：N1243555，生产企业：意大利Chromogenix。
发色底物S，规格：25mg，批号：N1143454，生产企业：意大利Chromogenix。
聚乙二醇(PEG)6000，规格：50g，批号：LB50P76，生产企业：北京百灵威科技有限公司。
肝素钠，规格：1g/瓶，批号：9041-08-1，生产企业：Bomei。
(3)PAF含量测定试剂
科研兔血小板活化因子(PAF)试剂盒，规格:96T/盒×5盒，48T/盒×1盒，批号：201509,201508，失效期：2016.02，上海酶联生物科技有限公司生产。
(4)麻醉剂
戊巴比妥钠，用于家兔麻醉。使用时用纯化水将戊巴比妥钠配制成3％浓度。配制成3％戊巴比妥钠溶液(g/v)。用于造模时家兔麻醉。
(5)病理检查剂
组织标本固定液(FAA液)：95％乙醇(AR级)，批号：2015052301，有效期至：2020.4，规格：20kg/桶，成都市科龙化工试剂厂生产；冰乙酸(AR级)，批号：2015040201，有效期至：2020.03，规格：500ml/瓶，成都市科龙化工试剂厂生产；甲醛(AR级)，批号：2014120801，有效期至：2016.11，规格：500ml/瓶，成都市科龙化工试剂厂生产。配制方法：分别量取95％乙醇4200ml、甲醛500ml、冰乙酸300ml和纯化水900ml倒入塑料桶中，充分混匀即可。
脱水试剂：无水乙醇(AR级)，批号：2015052501，有效期至：2020.04，规格：20kg/桶，成都市科龙化工试剂厂生产。配制方法：取无水乙醇用纯化水稀释配成70％、80％和90％乙醇溶液。
透明试剂：二甲苯(AR级)，批号：2014071001，有效期至：2019.06，规格：500ml/瓶，成都市科龙化工试剂厂生产。
苏木素染液：苏木素(AR级)，批号：2014010701，有效期至：2017.04，规格：5g/瓶，成都市科龙化工试剂厂生产；硫酸铝钾(AR级)，批号:2014091601，有效期至：2017.1，规格：500g/瓶，成都市科龙化工试剂厂生产；碘酸钠(AR级)，批号：20140711，有效期至：2017.1，规格：100g/瓶，成都市科龙化工试剂厂生产；甘油(丙三醇，AR级)，批号：2014122401，有效期至：2017.12，成都市科龙化工试剂厂生产；冰乙酸(AR级)， 批号：2015040201，有效期至：2020.3，规格：500ml/瓶，成都市科龙化工试剂厂生产。配制方法：称取苏木素1.5002g，加入250ml纯化水搅拌待其完全溶解，再加入312.5ml纯化水，称取碘酸钠0.3003g加入烧杯中，搅拌5分钟，再加入硫酸铝钾37.5004g，待其完全溶解加入甘油187.5ml，混匀，最后加入冰乙酸7.5ml，充分搅拌混匀即得。
伊红染液：伊红(水溶)，批号：2013110501，有效期至：2016.05，规格：25g/瓶，成都市科龙化工试剂厂；无水乙醇(AR级)，批号：2015052501，有效期至：2020.4，规格：20kg/桶，成都市科龙化工试剂厂生产。配制方法：量取无水乙醇600ml和纯化水150ml倒入烧杯中，混匀，加入伊红3.7505g，搅匀使溶解，即得。
盐酸－乙醇分化液：盐酸(AR级)，批号：2014072301，有效期至：2019.7.22，规格：500ml/瓶，成都市科龙化工试剂厂；无水乙醇(AR级)，批号：2015052501，有效期至：2020.4，规格：20kg/桶，成都市科龙化工试剂厂生产。配制方法：取无水乙醇520ml和纯化水222.5ml倒入烧杯中，混匀，再缓慢加入浓盐酸7.5ml，搅拌混匀，即得。
封片试剂：中性树胶，规格：100g/瓶，批号：20140106，有效期至：2018.12，中国上海懿洋仪器有限公司。
盖玻片清洗液：重铬酸钾：(AR级)，批号：20120822，有效期至：2016.07，规格：500g/瓶，成都市科龙化工试剂厂生产；浓硫酸：(AR级)，批号：2014072301、20130607，有效期至：2019.7.22、2018.06.06，规格：500ml/瓶，成都市科龙化工试剂厂生产。配制方法：取纯化水500ml，加入重铬酸钾50.0003g，搅拌、溶解、混匀，再向其中缓慢加入浓硫酸50ml，搅拌混匀即可。
2.4造模
取检疫及适应性观察合格的日本大耳白家兔392只，体重2.070-2.560kg，全雄性，先取8只作为正常组，其余均采用击打装置，击打接触圆面直径为1.5cm，创伤能量按Ep＝mgh计算，铁棒每打击一次能量为7.5J。具体方法：兔耳缘静脉注射3％戊巴比妥钠1ml/kg麻醉，右侧俯卧位，左大腿根部置打击平台上，触清兔左股骨大转子，将击打头平面紧贴股骨大转子至大转子下1.5cm范围内，使用自制击打装置，击打兔大腿近端外侧，石膏固定左下肢于屈髋屈膝位，上石膏绷带时检查右髋活动自如后，俯卧位待石膏定型,造模后动物自由饮水，颗粒饲料饲养，不用抗凝剂及抗生素。
2.5分组与给药方式
2.5.1药物配制方法
药物(mg)：羧丙甲纤维素(mg)：乳糖(mg)：甘露醇(mg)＝1:2:10:20研磨均匀后，再加水入纯化水1ml，搅拌均匀，即得。举例：50mg药物+100mg羧丙甲纤维素+500mg乳糖+1000mg甘露醇，研磨均匀，加水50ml，搅拌均匀，即得1mg/ml的药液。
2.5.2给药方案
各组动物于造模后分组，按下表4给药方案灌胃给药，连续给药7天，每天给药1次。表4给药方案
Figure PCTCN2015093305-appb-000008
Figure PCTCN2015093305-appb-000009
2.6指标检测
2.6.1凝血功能及凝血因子活性测定
首次给药后0.5、1.0、2.0、3.0和4.0h，耳中央动脉取血，分离血浆，在全自动血凝仪上测定aPPT、PT及Xa活性。
2.6.2 PAF含量变化
首次给药后0.5、1.0、2.0、3.0和4.0h，耳中央动脉取血，分离血清，采用ELISA法测定PAF含量变化。
2.6.3深静脉血栓形成测定
末次给药30min后放血处死动物，切取股动、静脉，HE染色，光镜镜检检查血管内皮损伤、血栓形成和血管破裂等情况，并对以上病变程度进行分级评分，具体评分标准详见表5。
表5深静脉血栓形成病理组织学检查病变程度评分标准
Figure PCTCN2015093305-appb-000010
Figure PCTCN2015093305-appb-000011
2.7统计学处理
凝血功能及凝血因子和PAF含量数据为计量资料，采用Excel软件计算平均数、标准差，判断数据是否呈正态分布，正态分布则采用excel中F检验进行方差齐性检验，然后选择t检验进行统计；若呈非正态分布，则进行非参数检验。
病理检测数据为等级资料数据，采用软件SPSS13.0“非参数检验”的“多个独立样本”进行统计分析。
三、实验结果
3.1凝血功能及抗凝血因子活性测定
表6凝血功能aPPT检查结果(
Figure PCTCN2015093305-appb-000012
秒)
组别 给药后0.5h 给药后1h 给药后2h 给药后3h 给药后4h
正常组 29.53±8.77 26.31±9.53 30.27±7.29 24.06±8.92 24.82±9.29
模型组 24.70±5.90 22.91±5.57 32.69±7.27 26.79±10.06 29.79±7.51
银杏内酯B组 25.89±8.15 30.65±6.79* 25.83±5.96 34.30±6.56 31.92±7.77
利伐沙班组 27.89±7.97 32.01±9.31* 34.63±5.57 35.92±10.85 29.20±6.32
阿哌沙班组 19.05±6.17 24.26±7.89 23.94±5.49 31.71±8.53 29.21±11.35
依度沙班组 26.77±7.65 25.01±9.52 33.63±5.45 36.47±9.12 27.20±6.31
雷扎沙班组 20.13±6.74 26.23±8.43 22.33±4.46 35.75±8.68 29.65±9.72
奥米沙班组 21.67±9.42 25.71±7.68 25.61±6.75 34.62±7.59 30.16±9.37
组合物1组 29.21±9.08 36.44±7.66** 24.48±8.23* 30.53±6.77 32.92±4.64
组合物2组 28.64±6.54 32.66±8.59* 35.24±4.34 36.99±7.65 30.90±8.15
组合物3组 25.64±3.41 33.75±8.16** 31.29±4.96 30.89±6.40 33.90±8.02
组合物4组 25.77±6.85 30.69±8.36* 36.63±5.64 36.46±9.31 31.20±6.52
组合物5组 19.13±6.56 34.23±9.46** 35.33±4.45 30.74±7.68 30.65±7.65
组合物6组 20.36±9.14 32.71±7.66* 35.61±6.78 36.62±7.57 32.16±8.67
组合物7组 21.36±7.69 34.63±7.91** 35.45±6.75 31.52±6.84 32.61±8.27
组合物8组 20.94±5.84 35.64±8.75** 36.14±8.17 31.67±9.17 31.67±8.19
组合物9组 29.71±8.42 35.54±12.24** 25.93±5.93* 30.25±6.04 33.23±4.18
组合物10组 32.77±7.24 29.06±7.22* 36.65±15.54 29.52±7.39 28.76±5.68
组合物11组 27.70±9.07 31.37±9.86* 29.62±9.51 28.88±5.37 29.54±5.92
组合物12组 28.67±8.14 33.71±7.48* 35.47±6.75 34.62±7.54 33.14±8.98
组合物13组 25.16±6.75 35.25±9.95** 35.74±5.67 31.64±7.75 31.64±7.57
组合物14组 20.78±9.48 34.71±7.74** 35.49±6.95 36.76±7.84 32.96±8.74
组合物15组 21.55±7.89 34.59±7.76** 35.44±6.79 31.39±6.69 32.88±8.97
组合物16组 20.64±5.77 35.89±8.55** 36.75±8.66 31.84±9.48 31.84±8.96
组合物17组 29.30±8.54 35.80±12.36** 25.45±7.69* 30.28±6.64 33.75±4.95
组合物18组 32.82±7.69 29.57±7.99* 36.56±15.78 29.94±7.56 28.84±5.59
组合物19组 27.91±9.26 31.76±9.94* 29.58±9.74 28.67±5.95 29.57±5.64
组合物20组 33.65±7.85 33.46±7.87* 36.48±15.95 29.75±7.48 28.75±5.83
组合物21组 34.81±7.75 33.68±7.82* 36.94±15.59 29.65±7.44 28.76±5.89
组合物22组 27.55±7.16 36.47±7.56** 35.26±6.49 31.69±6.72 32.73±8.18
组合物23组 26.64±5.29 35.99±8.48** 36.64±8.72 31.43±9.55 31.95±8.64
组合物24组 23.46±7.85 36.75±7.82** 35.48±6.80 31.19±6.48 32.15±8.73
组合物25组 29.25±8.78 35.65±12.85** 25.56±5.64* 30.85±6.96 33.49±8.62
组合物26组 30.67±7.00 33.52±7.96* 36.56±15.26 29.35±7.84 28.62±5.85
组合物27组 29.81±7.17 33.64±7.45* 36.44±10.64 29.46±7.52 28.43±5.42
组合物28组 27.48±5.96 36.72±7.63** 35.58±6.64 31.28±6.65 32.45±8.55
组合物29组 26.77±5.55 35.58±8.23** 36.66±8.75 31.44±9.28 31.63±8.45
组合物30组 28.30±8.46 35.75±10.64** 24.26±6.37* 30.58±6.57 33.56±5.00
组合物31组 32.42±7.22 31.69±7.41* 36.23±9.47 29.56±7.47 28.95±5.36
组合物32组 27.68±9.48 31.28±9.36* 29.59±9.82 28.63±5.41 29.45±5.69
组合物33组 33.55±7.96 34.36±7.85** 36.96±15.23 29.71±7.23 28.69±5.64
组合物34组 34.96±7.12 33.23±7.55* 36.64±15.38 29.45±7.26 28.82±5.38
组合物35组 30.65±9.22 32.65±9.64* 30.69±9.47 28.96±5.57 30.64±8.39
组合物36组 30.68±8.65 32.68±8.25* 35.68±8.64 29.58±7.84 28.58±5.28
组合物37组 31.47±5.94 33.75±7.11* 36.14±15.16 29.29±7.96 28.85±5.55
组合物38组 27.85±7.96 36.44±7.77** 35.95±6.05 31.39±6.41 32.65±8.45
组合物39组 26.72±5.58 35.83±8.64** 36.74±8.49 31.57±9.68 31.82±8.63
组合物40组 23.82±7.34 36.91±7.50** 35.53±6.67 31.58±6.65 32.42±8.95
组合物41组 29.74±8.45 36.65±12.27** 30.58±8.27 30.52±5.86 34.16±8.51
注：①模型组与正常对照组比较p＞0.05，②模型组与各给药组比较*p＜0.05**p＜0.01
由表6显示：①给药后0.5h～4h，模型组与正常对照组比较aPTT(活化部分凝血活酶时间)均未见明显延长(p>0.05)。说明造模成功。②与模型组比较，各给药组给药后0.5h～4h aPTT大部分未见明显延长(p>0.05)。说明银杏内酯B、沙班类及各组合物组无出血不良反应。③给药后1h与模型组比较，银杏内酯B和利伐沙班组aPTT出现明显延长(p﹤0.05)，阿哌沙班组、依度沙班组、雷扎沙班组、奥米沙班组aPPT均未见明显延长(P>0.05)；银杏内酯B+沙班类组aPPT出现显著延长(p﹤0.01)和明显延长(p﹤0.05)，表明本发明药物组合物可以有效延长aPPT，同时该比例下银杏内酯B与沙班对aPPT的延长具有协同增效的作用。
表7凝血功能PT检查结果(
Figure PCTCN2015093305-appb-000013
秒)
组别 给药后0.5h 给药后1h 给药后2h 给药后3h 给药后4h
正常组 6.83±0.31 6.89±0.39 6.83±0.17 6.83±0.30 6.78±0.23
模型组 6.59±0.23 6.69±0.26 7.07±1.47 6.66±0.22 6.73±0.16
银杏内酯B组 6.76±0.32 6.79±0.27 6.75±0.36 6.72±0.24 6.68±0.23
利伐沙班组 6.76±0.30 7.20±0.96 6.90±0.34 6.81±0.22 6.70±0.33
阿哌沙班组 6.78±0.21 6.76±0.21 6.70±0.19 6.70±0.24 7.14±0.87
依度沙班组 6.73±0.15 6.14±0.26 6.28±0.42 6.45±0.65 6.42±0.34
雷扎沙班组 6.71±0.25 7.41±0.51 6.47±0.25 6.47±0.41 6.70±0.25
奥米沙班组 6.58±0.45 6.74±0.26 6.52±0.43 6.62±0.42 7.82±0.31
组合物1组 7.03±0.13** 6.56±0.34 6.47±0.34 6.58±0.18 6.57±0.16
组合物2组 6.86±0.21* 6.73±0.25 6.85±0.30 6.70±0.28 6.71±0.28
组合物3组 6.82±0.22* 6.83±0.24 6.76±0.20 6.84±0.34 6.94±0.24
组合物4组 6.85±0.34* 6.56±0.77 6.48±0.56 6.75±0.35 6.75±0.26
组合物5组 6.99±0.32** 6.46±0.26 6.58±0.25 6.74±0.58 6.69±0.28
组合物6组 6.86±0.24* 6.74±0.58 6.74±0.34 6.81±0.29 6.83±0.50
组合物7组 6.95±0.26** 6.79±0.25 6.78±0.38 6.79±0.34 6.85±0.19
组合物8组 6.91±0.41** 6.78±0.34 6.95±0.18 6.84±0.24 6.67±0.26
组合物9组 7.10±0.23** 6.86±0.26 6.83±0.28 6.97±0.56 6.85±0.35
组合物10组 6.84±0.29* 6.88±0.33 6.72±0.15 6.75±0.20 6.75±0.30
组合物11组 6.88±0.12* 6.97±0.24 7.07±1.18 7.06±0.88 7.07±0.54
组合物12组 6.81±0.19* 6.83±0.31 6.83±0.13 6.89±0.62 6.74±0.13
组合物13组 7.02±0.33** 6.80±0.30 6.87±0.15 6.68±0.25 6.79±0.28
组合物14组 6.92±0.24** 6.64±0.21 6.82±0.34 6.79±0.19 6.95±0.30
组合物15组 7.00±0.18** 6.92±0.26 6.79±0.17 6.83±0.48 6.91±0.24
组合物16组 6.95±0.23** 6.95±0.29 6.85±0.36 6.58±0.24 6.86±0.28
组合物17组 7.07±0.52** 6.58±0.35 6.96±0.41 6.85±0.36 6.95±0.26
组合物18组 6.88±0.22* 6.93±0.24 6.85±0.25 6.92±0.21 6.84±0.28
组合物19组 6.86±0.31* 6.68±0.35 6.99±0.95 7.00±0.65 7.14±0.35
组合物20组 6.81±0.25* 6.79±0.29 6.85±0.26 6.79±0.53 6.88±0.26
组合物21组 6.89±0.27* 6.95±0.19 6.86±0.28 6.93±0.36 7.00±0.31
组合物22组 7.05±0.25** 6.85±0.36 6.84±0.28 6.86±0.26 6.89±0.19
组合物23组 6.98±0.26** 6.87±0.51 6.95±0.15 6.79±0.19 6.95±0.29
组合物24组 7.03±0.29** 6.98±0.24 6.79±0.17 6.86±0.27 6.88±0.35
组合物25组 7.05±0.45** 6.74±0.15 6.87±0.35 6.92±0.42 6.86±0.46
组合物26组 6.89±0.19* 6.82±0.46 6.94±0.47 6.98±0.29 6.79±0.18
组合物27组 6.83±0.24* 6.75±0.36 6.85±0.74 6.85±0.63 7.01±0.15
组合物28组 7.01±0.18** 6.69±0.42 6.77±0.52 6.95±0.25 6.85±0.26
组合物29组 7.00±0.31** 6.78±0.35 6.96±0.24 6.97±0.28 6.96±0.27
组合物30组 7.15±0.27** 6.86±0.14 6.88±0.29 6.95±0.24 6.88±0.17
组合物31组 6.89±0.16* 6.88±0.20 6.74±0.23 6.99±0.18 6.79±0.35
组合物32组 6.85±0.24* 6.95±0.24 6.78±0.56 6.97±0.36 7.01±0.26
组合物33组 6.86±0.19* 7.01±0.23 6.96±0.42 6.76±0.46 6.96±0.18
组合物34组 6.88±0.34* 6.76±0.26 6.79±0.34 6.85±0.28 6.98±0.24
组合物35组 6.85±0.22* 6.65±0.64 6.48±0.96 7.03±0.16 6.85±0.42
组合物36组 6.86±0.60* 6.98±0.26 6.67±0.51 6.95±0.62 6.80±0.23
组合物37组 6.84±0.63* 6.85±0.46 6.89±0.41 6.95±0.28 6.74±0.52
组合物38组 7.12±0.26** 6.96±0.55 6.89±0.45 6.78±0.27 6.95±0.35
组合物39组 7.05±0.65** 6.99±0.65 6.47±0.74 6.86±0.51 6.85±0.37
组合物40组 7.06±0.29** 6.75±0.29 6.76±0.42 6.83±0.36 6.96±0.28
组合物41组 6.98±0.52** 6.86±0.25 6.68±0.36 6.86±0.85 6.96±0.26
注：①模型组与正常对照组比较p＞0.05，②模型组与各给药组比较*p＜0.05
由表7显示：①给药后0.5h～4h，模型组与正常对照组比较PT(凝血酶原时间)均未见明显延长(p>0.05)。说明造模成功。②与模型组比较，各给药组给药后0.5h～4h PT均未见明显延长(p>0.05)。说明银杏内酯B、沙班类组及各组合物组无出血不良反应。③给药后0.5h与模型组比较，各组合物组PT(凝血酶原时间)出现明显延长(p﹤0.05)和显著延长(p﹤0.01)，表明本发明药物组合物可以有效延长PT，并且银杏内酯B与沙班类对PT的延长具有协同增效的作用。
表8抗凝血因子Xa活性检查结果(
Figure PCTCN2015093305-appb-000014
IU/ml)
组别 给药后0.5h 给药后1h 给药后2h 给药后3h 给药后4h
正常组 0.10±0.05 0.10±0.05 0.10±0.06 0.09±0.08 0.09±0.06
模型组 0.09±0.07 0.08±0.05 0.13±0.06 0.09±0.09 0.13±0.07
银杏内酯B组 0.08±0.07 0.10±0.10 0.35±0.22* 0.74±0.15*** 0.65±0.12***
利伐沙班组 0.37±0.09*** 0.31±0.13*** 0.18±0.05* 1.32±0.16*** 1.38±0.18***
阿哌沙班组 0.16±0.10*** 0.26±0.04*** 0.25±0.10* 1.22±0.22*** 1.12±0.15***
依度沙班组 0.25±0.06*** 0.21±0.08*** 0.20±0.10* 1.26±0.09*** 1.26±0.07***
雷扎沙班组 0.20±0.14*** 0.18±0.05*** 0.23±0.08* 1.31±0.30*** 1.32±0.09***
奥米沙班组 0.19±0.04*** 0.25±0.10*** 0.26±0.14* 1.25±0.12*** 1.24±0.06***
组合物1组 0.41±0.18*** 0.17±0.03*** 0.94±0.13*** 1.37±0.32*** 1.15±0.16***
组合物2组 0.32±0.05*** 0.31±0.13*** 1.10±0.19*** 1.27±0.22*** 1.06±0.21***
组合物3组 0.30±0.07*** 0.25±0.07*** 1.20±0.10*** 1.28±0.10*** 1.30±0.09***
组合物4组 0.24±0.12*** 0.30±0.05*** 1.23±0.08*** 1.23±0.52*** 1.42±0.05***
组合物5组 0.20±0.06*** 0.36±0.13*** 1.26±0.14*** 1.30±0.12*** 1.26±0.08***
组合物6组 0.35±0.25*** 0.34±0.05*** 1.34±0.13*** 1.34±0.42*** 1.20±0.30***
组合物7组 0.34±0.09*** 0.29±0.08*** 1.22±0.23*** 1.35±0.32*** 1.28±0.40***
组合物8组 0.40±0.08*** 0.32±0.12*** 1.41±0.04*** 1.28±0.29*** 1.34±0.36***
组合物9组 0.46±0.04*** 0.24±0.21*** 1.03±0.21*** 1.30±0.18*** 1.26±0.31***
组合物10组 0.32±0.08*** 0.25±0.12*** 0.74±0.17*** 1.16±0.17*** 0.70±0.16***
组合物11组 0.25±0.05*** 0.29±0.17*** 1.21±0.06*** 1.31±0.15*** 1.12±0.07***
组合物12组 0.26±0.04*** 0.32±0.21*** 0.95±0.23*** 0.96±0.20*** 1.05±0.15***
组合物13组 0.45±0.09*** 0.35±0.20*** 1.05±0.22*** 1.21±0.11*** 1.09±0.15***
组合物14组 0.35±0.08*** 0.32±0.17*** 1.14±0.08*** 1.20±0.09*** 1.06±0.08***
组合物15组 0.43±0.03*** 0.30±0.14*** 1.35±0.25*** 1.34±0.19*** 1.12±0.24***
组合物16组 0.40±0.10*** 0.32±0.15*** 1.54±0.20*** 1.26±0.31*** 1.34±0.20***
组合物17组 0.50±0.04*** 0.32±0.06*** 1.35±0.09*** 1.29±0.09*** 1.26±0.08***
组合物18组 0.30±0.06*** 0.35±0.24*** 1.24±0.16*** 1.28±0.06*** 1.21±0.23***
组合物19组 0.33±0.08*** 0.31±0.20*** 1.15±0.36*** 1.41±0.25*** 1.19±0.35***
组合物20组 0.31±0.03*** 0.29±0.16*** 1.09±0.06*** 1.24±0.04*** 1.14±0.05***
组合物21组 0.36±0.07*** 0.36±0.12*** 1.29±0.08*** 1.29±0.05*** 1.06±0.06***
组合物22组 0.46±0.06*** 0.39±0.06*** 1.46±0.19*** 1.42±0.26*** 1.46±0.18***
组合物23组 0.37±0.09*** 0.35±0.32*** 1.31±0.09*** 1.32±0.07*** 1.41±0.09***
组合物24组 0.42±0.07*** 0.40±0.07*** 1.25±0.07*** 1.26±0.08*** 1.28±0.06***
组合物25组 0.45±0.08*** 0.30±0.08*** 1.12±0.15*** 1.35±0.06*** 1.25±0.09***
组合物26组 0.34±0.06*** 0.34±0.05*** 1.21±0.08*** 1.29±0.08*** 1.41±0.09***
组合物27组 0.35±0.09*** 0.40±0.08*** 1.35±0.09*** 1.32±0.06*** 1.32±0.27***
组合物28组 0.43±0.07*** 0.31±0.30*** 1.36±0.02*** 1.28±0.01*** 1.38±0.05***
组合物29组 0.42±0.07*** 0.40±0.07*** 1.25±0.07*** 1.26±0.08*** 1.28±0.06***
组合物30组 0.47±0.08*** 0.28±0.26*** 1.29±0.05*** 1.24±0.06*** 1.39±0.05***
组合物31组 0.36±0.04*** 0.38±0.06*** 1.35±0.06*** 1.39±0.02*** 1.35±0.07***
组合物32组 0.34±0.03*** 0.28±0.04*** 1.25±0.09*** 1.41±0.08*** 1.28±0.04***
组合物33组 0.31±0.05*** 0.36±0.09*** 1.29±0.06*** 1.30±0.06*** 1.31±0.07***
组合物34组 0.34±0.01*** 0.29±0.06*** 1.32±0.10*** 1.36±0.08*** 1.29±0.05***
组合物35组 0.36±0.09*** 0.27±0.26*** 1.42±0.01*** 1.32±0.05*** 1.28±0.06***
组合物36组 0.37±0.05*** 0.35±0.06*** 1.38±0.13*** 1.29±0.04*** 1.34±0.07***
组合物37组 0.30±0.06*** 0.36±0.08*** 1.35±0.07*** 1.32±0.07*** 1.32±0.09***
组合物38组 0.45±0.06*** 0.31±0.05*** 1.32±0.04*** 1.36±0.09*** 1.28±0.08***
组合物39组 0.44±0.08*** 0.34±0.06*** 1.37±0.09*** 1.39±0.06*** 1.35±0.08***
组合物40组 0.44±0.10*** 0.35±0.25*** 1.38±0.06*** 1.41±0.04*** 1.37±0.02***
组合物41组 0.40±0.07*** 0.29±0.08*** 1.41±0.07*** 1.38±0.09*** 1.28±0.08***
注：①模型组与正常对照组比较p＞0.05，②模型组与各给药组比较*p＜0.05**p ＜0.01***p＜0.001
备注：抗凝有效定义为血浆抗Xa活性≧0.5IU/ML(《心血管疾病人应用低分子肝素要规范化》报道：根据LMWH体内的药代动力学特点制定的)
由表8显示：①给药后0.5h～4h，模型组与正常对照组比较抗凝血因子Xa活性均未见异常(p>0.05)。说明造模成功。②与模型组比较，给药后0.5h～4h，银杏内酯B组抗凝血因子Xa活性随着时间的推移而增高(P﹤0.001)，利伐沙班与阿哌沙班组均具有极显著差异(p﹤0.001)，表明沙班类药物的抗凝血因子Xa活性强于银杏内酯B。③给药后2h与模型组比较，各单药组抗Xa活性具有明显增高(P﹤0.05)，各组合物组抗Xa活性具有极显著增高(P﹤0.001)，表明本发明药物组合物可以提高Xa因子活性，并且银杏内酯B与沙班抗Xa活性具有协同增效的作用。
3.2 PAF含量变化
表9 PAF含量检查结果(
Figure PCTCN2015093305-appb-000015
ng/L)
Figure PCTCN2015093305-appb-000016
Figure PCTCN2015093305-appb-000017
Figure PCTCN2015093305-appb-000018
注：①模型组与正常对照组比较p＞0.05，②模型组与各给药组比较*p＜0.05,**p＜0.01***p＜0.001
由表9显示：①给药后0.5h～4h，模型组与正常对照组比较PAF(血小板活化因子)含量均未见明显差异(p>0.05)。说明造模成功。②与模型组比较，给药后2h，银杏内酯B组PAF含量显著降低(P＜0.01),给药后4h，利伐沙班与阿哌沙班组PAF含量极显著降低(P＜0.001)，雷扎沙班与奥米沙班组PAF含量显著降低(P＜0.01)。③与模型组比较，给药后0.5h，各组合物组PAF含量显著降低(P＜0.01)，说明本发明药物组合物可以有效降低PAF含量，并且银杏内酯B与沙班类拮抗PAF受体具有协同增效的作用；给药后1h，各组合物组PAF含量极显著降低(P＜0.001)和显著降低(P＜0.01)，说明银杏内酯B与阿哌沙班拮抗PAF受体具有协同增效的作用；给药后2h，各组合物组PAF含量极显著降低(P＜0.001)和显著降低(P＜0.01)，说明银杏内酯B与阿哌沙班拮抗PAF受体具有协同增效的作用。
由表6～表9可以看出：
银杏内酯B+利伐沙班的组合物组(组合物1～8)中，组合物1、5～8组的aPPT较长、PT更长、Xa因子活性更高、PAF含量更低，组合物1组的aPPT最长、PT最长、Xa因 子活性最高、PAF含量最低，因此，银杏内酯B+利伐沙班的组合物中，银杏内酯B与利伐沙班的配比优选为：银杏内酯B5-15份和利伐沙班10-20份，最优选为：银杏内酯B10份和利伐沙班15份。
银杏内酯B+阿哌沙班的组合物组(组合物9～16)中，组合物9、13～16组的血小板aPPT较长、PT更长、Xa因子活性更高、PAF含量更低，组合物9组的aPPT最长、PT最长、Xa因子活性最高、PAF含量最低，因此，银杏内酯B+阿哌沙班的组合物中，银杏内酯B与阿哌沙班的配比优选为：银杏内酯B5-15份和阿哌沙班1-10份，最优选为：银杏内酯B10份和阿哌沙班5份。
银杏内酯B+依度沙班的组合物组(组合物17～24)中，组合物17、22～24组的aPPT较长、PT更长、Xa因子活性更高、PAF含量更低，组合物17组的aPPT最长、PT最长、Xa因子活性最高、PAF含量最低，因此，银杏内酯B+依度沙班的组合物中，银杏内酯B与依度沙班的配比优选为：银杏内酯B5-15份和依度沙班10-30份，最优选为：银杏内酯B10份和依度沙班15份。
银杏内酯B+雷扎沙班的组合物组(组合物25～29)中，组合物25、28～29组的aPPT较长、PT更长、Xa因子活性更高、PAF含量更低，组合物25组的aPPT最长、PT最长、Xa因子活性最高、PAF含量最低，因此，银杏内酯B+雷扎沙班的组合物中，银杏内酯B与雷扎沙班的配比优选为：银杏内酯B5-15份和雷扎沙班5-15份，最优选为：银杏内酯B10份和雷扎沙班10份。
银杏内酯B+奥米沙班的组合物组(组合物30～41)中，组合物30、38～41组的aPPT较长、PT更长、Xa因子活性更高、PAF含量更低，组合物30组的aPPT最长、PT最长、Xa因子活性最高、PAF含量最低，因此，银杏内酯B+奥米沙班的组合物中，银杏内酯B与奥米沙班的配比优选为：银杏内酯B8-12份和奥米沙班50-90份，最优选为：银杏内酯B10份和奥米沙班10份。
实验结果说明，本发明将银杏内酯B和沙班类活性成分，如，利伐沙班、阿哌沙班、依度沙班、雷扎沙班、奥米沙班等沙班类药物联合使用，aPPT明显延长、PT明显延长、Xa因子活性明显增强、PAF含量明显降低，，且优于二者单独使用，二者联合使用发挥了协同增效作用，银杏内酯B和沙班类活性成分组成的组合物，aPPT长、PT长、Xa因子活性高、PAF含量低，可以通过不同的作用机制抗血小板聚集。
综上，本发明将银杏内酯B与沙班类药物联合使用，二者可以发挥协同增效作用，本发明银杏内酯B与沙班类药物组成的药物组合物，可以有效抑制血小板聚集，效果优良，临床应用前景优良。

Claims (10)

  1. 一种含银杏内酯B的药物组合物，其特征在于：它含有银杏内酯B和Xa因子抑制剂沙班类药物。
  2. 根据权利要求1所述的药物组合物，其特征在于：所述的沙班类药物为利伐沙班、阿哌沙班、依度沙班、雷扎沙班和/或奥米沙班。
  3. 根据权利要求2所述的药物组合物，其特征在于：
    所述银杏内酯B与利伐沙班的重量配比为：银杏内酯B1-20份和利伐沙班5-40份；
    优选地，二者的重量配比为：银杏内酯B5-15份和利伐沙班10-20份；进一步优选地，二者的重量配比为：银杏内酯B10份和利伐沙班15份。
  4. 根据权利要求2所述的药物组合物，其特征在于：
    所述银杏内酯B与阿哌沙班的重量配比为：银杏内酯B1-20份和阿哌沙班0.5-20份；优选地，二者的重量配比为：银杏内酯B5-15份和阿哌沙班1-10份；进一步优选地，二者的重量配比为：银杏内酯B10份和阿哌沙班5份。
    或者，所述银杏内酯B与依度沙班的重量配比为：所述各原料的重量配比为：银杏内酯B1-20份和依度沙班8-50份；优选地，二者的重量配比为：银杏内酯B5-15份和依度沙班10-30份；进一步优选地，二者的重量配比为：银杏内酯B10份和依度沙班15份；
    或者，所述银杏内酯B与雷扎沙班的重量配比为：银杏内酯B1-20份和雷扎沙班1-20份；优选地，二者的重量配比为：银杏内酯B5-15份和雷扎沙班5-15份；进一步优选地，二者的重量配比为：银杏内酯B10份和雷扎沙班10份；
    或者，所述银杏内酯B与奥米沙班的重量配比为：银杏内酯B1-20份和奥米沙班15-150份；优选地，二者的重量配比为：银杏内酯B5-15份和奥米沙班30-125份；进一步优选地，二者的重量配比为：银杏内酯B8-12份和奥米沙班50-90份；再进一步优选地，二者的重量配比为：银杏内酯B10份和奥米沙班60份。
  5. 一种制备权利要求1～4中任意一项所述的药物组合物的方法，它包括以下步骤：
    S1：按组份及重量比称取原料银杏内酯B和Xa因子抑制剂沙班类药物；
    S2：将原料混合后，加入药学上可接受的辅料制备成药学上常用的药物制剂。
  6. 权利要求1～4任意一项所述的药物组合物在制备抗血小板聚集药物中的用途。
  7. 银杏内酯B与Xa因子抑制剂沙班类药物在在制备抗血小板聚集的联合用药中的用途。
  8. 根据权利要求7所述的用途，其特征在于：所述的沙班类药物为利伐沙班、阿哌沙班、依度沙班、雷扎沙班和/或奥米沙班。
  9. 根据权利要求8所述的用途，其特征在于：所述银杏内酯B与利伐沙班的重量配比为：银杏内酯B1-20份和利伐沙班5-40份；
    优选地，二者的重量配比为：银杏内酯B5-15份和利伐沙班10-20份；进一步优选地，二者的重量配比为：银杏内酯B10份和利伐沙班15份。
  10. 根据权利要求8所述的用途，其特征在于：
    所述银杏内酯B与阿哌沙班的重量配比为：银杏内酯B1-20份和阿哌沙班0.5-20份；优选地，二者的重量配比为：银杏内酯B5-15份和阿哌沙班1-10份；进一步优选地，二者的重量配比为：银杏内酯B10份和阿哌沙班5份。
    或者，所述银杏内酯B与依度沙班的重量配比为：所述各原料的重量配比为：银杏内酯B1-20份和依度沙班8-50份；优选地，二者的重量配比为：银杏内酯B5-15份和依度沙班10-30份；进一步优选地，二者的重量配比为：银杏内酯B10份和依度沙班15份；
    或者，所述银杏内酯B与雷扎沙班的重量配比为：银杏内酯B1-20份和雷扎沙班1-20份；优选地，二者的重量配比为：银杏内酯B5-15份和雷扎沙班5-15份；进一步优选地，二者的重量配比为：银杏内酯B10份和雷扎沙班10份；
    或者，所述银杏内酯B与奥米沙班的重量配比为：银杏内酯B1-20份和奥米沙班15-150份；优选地，二者的重量配比为：银杏内酯B5-15份和奥米沙班30-125份；进一步优选地，二者的重量配比为：银杏内酯B8-12份和奥米沙班50-90份；再进一步优选地，二者的重量配比为：银杏内酯B10份和奥米沙班60份。
PCT/CN2015/093305 2014-10-30 2015-10-30 一种含有银杏内酯B和Xa因子抑制剂的药物组合物及其制备方法和用途 WO2016066128A1 (zh)

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KR1020177014595A KR101968190B1 (ko) 2014-10-30 2015-10-30 징코라이드 B(Ginkgolide B)와 Ⅹa 인자 억제제를 포함한 약물 조성물 및 그 제조방법과 용도
EP15854130.0A EP3213749B1 (en) 2014-10-30 2015-10-30 Pharmaceutical composition containing ginkgolide b and xa factor inhibitor, preparation method thereof and use thereof
CA2966249A CA2966249C (en) 2014-10-30 2015-10-30 Pharmaceutical composition containing ginkgolide b and xa factor inhibitor, preparation method thereof and use thereof
JP2017542250A JP6389009B2 (ja) 2014-10-30 2015-10-30 ギンコライドＢと第Ｘａ因子阻害剤とを含有する薬物組成物及びその製造方法と用途

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