WO2007118363A1 - Composition médicinale chinoise, procédé de préparation et utilisation de celle-ci - Google Patents

Composition médicinale chinoise, procédé de préparation et utilisation de celle-ci Download PDF

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Publication number
WO2007118363A1
WO2007118363A1 PCT/CN2006/000724 CN2006000724W WO2007118363A1 WO 2007118363 A1 WO2007118363 A1 WO 2007118363A1 CN 2006000724 W CN2006000724 W CN 2006000724W WO 2007118363 A1 WO2007118363 A1 WO 2007118363A1
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ethanol
water
extract
chinese medicine
group
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PCT/CN2006/000724
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English (en)
French (fr)
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Jianxun Liu
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Jianxun Liu
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Priority to PCT/CN2006/000724 priority Critical patent/WO2007118363A1/zh
Priority to EP06722373.5A priority patent/EP2033653B1/en
Priority to KR1020087028170A priority patent/KR101126117B1/ko
Priority to CA2652826A priority patent/CA2652826C/en
Priority to JP2009505701A priority patent/JP5097196B2/ja
Priority to US12/297,643 priority patent/US8158169B2/en
Priority to EP12167540A priority patent/EP2526956A1/en
Priority to AU2006342350A priority patent/AU2006342350B2/en
Priority to EP12167532A priority patent/EP2486919A1/en
Publication of WO2007118363A1 publication Critical patent/WO2007118363A1/zh

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    • 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
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A61K9/1605—Excipients; Inactive ingredients
    • A61K9/1629—Organic macromolecular compounds
    • A61K9/1652—Polysaccharides, e.g. alginate, cellulose derivatives; Cyclodextrin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/16—Ginkgophyta, e.g. Ginkgoaceae (Ginkgo family)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/25—Araliaceae (Ginseng family), e.g. ivy, aralia, schefflera or tetrapanax
    • A61K36/258—Panax (ginseng)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/48—Fabaceae or Leguminosae (Pea or Legume family); Caesalpiniaceae; Mimosaceae; Papilionaceae
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/88—Liliopsida (monocotyledons)
    • 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
    • A61K9/08—Solutions
    • 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
    • A61K9/20—Pills, tablets, discs, rods
    • A61K9/2004—Excipients; Inactive ingredients
    • A61K9/2022—Organic macromolecular compounds
    • A61K9/205—Polysaccharides, e.g. alginate, gums; Cyclodextrin
    • A61K9/2059—Starch, including chemically or physically modified derivatives; Amylose; Amylopectin; Dextrin
    • 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
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/4841—Filling excipients; Inactive ingredients
    • A61K9/4866—Organic macromolecular compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
    • A61K35/13—Tumour cells, irrespective of tissue of origin
    • 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
    • A61K9/20—Pills, tablets, discs, rods

Definitions

  • the present invention relates to a traditional Chinese medicine composition, in particular to a traditional Chinese medicine composition for treating ischemic cerebrovascular disease and senile dementia, the composition of which may be a Chinese medicinal material which is directly pulverized into a powder and used as a medicine. / or extracted Chinese herbal extracts. Background technique
  • Cerebrovascular disease is usually divided into two categories: ischemic cerebrovascular disease and hemorrhagic cerebrovascular disease.
  • Ischemic cerebrovascular disease is common, and cerebral infarction accounts for 59.2% to 85%.
  • Ischemic cerebrovascular disease includes: (1) transient ischemic attack (TIA, also known as small stroke or transient ischemic attack), the cause of which is related to cerebral arteriosclerosis, is transient and lack of brain tissue Dysfunction caused by bloody and focal damage; (2) cerebral thrombosis, blood clotting caused by atherosclerosis, various arteritis, trauma and other physical factors, blood diseases caused by local cerebral vascular lesions The block is blocked and the disease occurs; (3) Cerebral embolism can be induced by the emboli of various diseases entering the blood and blocking the blood vessels in the brain.
  • TIA transient ischemic attack
  • TIA transient ischemic attack
  • cerebral thrombosis blood clotting caused by atherosclerosis, various arteritis, trauma
  • Dementia is an acquired, persistent intellectual impairment syndrome caused by organic brain lesions. According to the 2005 Global Epidemiological Survey, approximately 24 million people currently suffer from dementia and are growing at a rate of 460 per cent per person (an increase of one person per 7 seconds), doubling every 20 years. In China, conservative tricks will increase at a rate of 300% per year from 2001 to 2040. It is estimated that the number of people suffering from dementia in the world will reach 81 million in 2040. The incidence of dementia increases with age, and the dementia that occurs in the elderly is mainly divided into: A. Primary degenerative dementia, namely Alzheimer's disease (AD); B. Vascular dementia (Vascular dementia, VD); C. Mixed dementia (AD combined with VD); D. Other dementia (Pick disease, Louis inclusion body dementia).
  • AD Alzheimer's disease
  • VD Vascular dementia
  • C Mixed dementia
  • D Other dementia
  • AD and VD are the two most important types of Alzheimer's disease, and the prevalence rate accounts for more than 90% of all dementias. Among them, AD is the most common cause of dementia in the elderly over 65 years old, and it is very likely to be the leading cause of fatal diseases.
  • cholinesterase inhibitors such as tacrine, donepezil, rivastigmine, and galantamine
  • senile dementia Currently, cholinesterase inhibitors (such as tacrine, donepezil, rivastigmine, and galantamine) are the first choice for the treatment of senile dementia.
  • the diagnosis of senile dementia in traditional Chinese medicine includes AD, VD of Western medicine or a mixed type of both, that is, the allergic diseases referred to by the senile dementia.
  • cholinesterase inhibitors can only partially improve the cognitive and emotional symptoms of patients in the early and middle stages, and have no significant effect on the basic pathological changes. They can only be delayed (about 1-2 years) and cannot stop the disease. The progress, that is, only the symptoms can not be cured, the long-term application may lead to increased cholinesterase synthesis, tacrine is more likely to cause severe gastrointestinal reactions and liver toxicity; Chinese herbal compound has unclear active ingredients, long-term inconvenience, lack of Stable quality control standards and other issues.
  • the inventor originated from the theory of traditional Chinese medicine, summed up the experience of many parties, and was formed through clinical practice experience.
  • the invention is a traditional Chinese medicine composition, which is a pure traditional Chinese medicine preparation obtained by extracting and purifying four kinds of natural plants, and the experiment proves that the curative effect is exact, safe and effective.
  • the present invention provides a traditional Chinese medicine composition comprising 1-10 parts of ginseng, 1-10 parts of ginkgo leaves, 0. 05-0.5 parts of saffron, 5-10 parts of soybeans, said ginseng
  • the source of Ginkgo biloba, saffron and soybean may be Chinese medicinal materials or Chinese herbal extracts equivalent to the above-mentioned Chinese medicinal materials.
  • composition of the present invention is a systemic therapeutic prescription for ischemic cerebrovascular disease and senile dementia, which is composed of a traditional Chinese medicine compound, focuses on the improvement of the autoimmune system, and has a wide range of drug sources, non-toxic side effects, and prescriptions. Reasonable and effective.
  • the above-mentioned traditional Chinese medicine composition wherein the parts by weight of the components are preferably: ginseng 2-6 parts, ginkgo leaves 3-6 parts, saffron 0. 06-0. 2 parts, soybean 7-8 parts; more preferably ginseng 4 0 ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ . ⁇ .
  • the present invention also provides a composition comprising 1-10 parts of ginseng extract, 1-10 parts of ginkgo biloba extract, saffron extract 0. 5-5 parts, soybean extract 0. 1-1 parts, the above-mentioned parts are parts by weight, and the ginseng, ginkgo leaf, saffron and soybean extract are all alcohol extracts.
  • the composition of the present invention is reasonable in prescription, and according to the traditional Chinese medicine, the ginseng in the composition is Fangzhongjun.
  • Ben The AD standard of the invention is mainly related to the dysfunction of viscera function and the abnormal operation of qi and blood.
  • the viscera deficiency and phlegm and phlegm and blood stasis are the main pathological basis of the disease.
  • ginseng active ingredient is ginseng total saponin
  • Ginkgo biloba leaves active ingredients mainly ginkgo flavonoids and ginkgo total lactone
  • saffron main active ingredient saffron total glycosides
  • the blood stasis and phlegm detoxification; the adjuvant soybean eliminates the "endogenous poison", the whole body benefits Qi and activating blood circulation, detoxification and collaterals, soothe the nerves, and the specimens.
  • all the raw powders may be selected as medicine, or may be extracted as an extract, and the composition can achieve the treatment of ischemic cerebrovascular disease as long as the components of the above weight ratio are included. And the efficacy of Alzheimer's disease, that is, within the scope of protection of the present invention.
  • ginseng, ginkgo biloba, saffron and soybean are all ethanol extracts of the drug substance.
  • the components and the ratio (by weight) of the above pharmaceutical composition are preferably ginseng extracts: Ginkgo biloba extract: Saffron extract: Soybean extract is 5: 5: 1: 0 .
  • the ginseng extract can be prepared by any known method, and the present invention preferably adopts the following extraction method - the human participates in at least 2 times (preferably 8 times) the low drug concentration (such as 50-70%, preferably 60%) of ethanol, and extracts at least 1 (preferably 3 times), each time at least 1 hour (preferably 3 hours), the extracts are combined, concentrated to a relative density of about 1.
  • the low drug concentration such as 50-70%, preferably 60%
  • the concentrate is added at least 1 time (preferably 2 times) volume of steam Hydrophobic, filtered, weakly polar polystyrene macroporous resin (preferably AB-8) on the filtrate, eluted with distilled water, continue to elute the drug with 10% ethanol, discard water and 10% ethanol
  • Hydrophobic, filtered, weakly polar polystyrene macroporous resin preferably AB-8
  • the eluate, the drug-loaded resin was further eluted with 70% ethanol, and the ginseng extract containing ginseng total saponin was obtained by collecting 70% ethanol eluate to about 2.5 column volumes.
  • the invention adopts the technical requirement that the drug-loaded macroporous adsorption resin is eluted with water, 10% ethanol, and finally eluted with 70% ethanol, and the two ethanol concentration and the elution program are screened to ensure the total content of the traditional Chinese medicine. , can also ensure that the active ingredient can have a higher transfer rate and finished product yield.
  • Ginkgo biloba extract can be obtained by the following extraction method:
  • the extracts are combined, and the extract is concentrated under reduced pressure to a relative density of about 1.05 (50 ° C).
  • the concentrate is added with water to cool the precipitate, filtered, and the polar hydrogen bonds on the filtrate.
  • Styrene type macroporous adsorption resin (preferably ADS-17 type), the purpose is to enrich the active ingredient, the drug loading resin is first eluted with water, continue to elute with 60% ethanol, discard the water eluent, collect the ethanol eluent Concentrate to an alcohol-free taste, add 2 times the amount of crude water to the boiling water, settling at room temperature for 24 hours, filter, and filter the weakly polar polystyrene macroporous adsorption resin (preferably DM-130 type) on the filtrate.
  • the weakly polar polystyrene macroporous adsorption resin preferably DM-130 type
  • Impurities first eluted with water, continue to elute with 15% ethanol and 60% ethanol, discard water and 15% ethanol eluent, collect 60% ethanol eluent; also continue to concentrate and dry to obtain finished ginkgo leaf extract .
  • the harmful impurities of ginkgo phenolic acids are extremely small in water solubility, they mainly appear in the eluent of ethanol at a concentration of more than 60%, and most of impurities such as polysaccharides and inorganic salts can be eluted by 15% ethanol, so In the refining step of the macroporous adsorption resin twice, most of the impurities are removed, and eluted by 60% ethanol, which is equivalent to collecting the eluent of 15-60% ethanol, and the phenolic acid content is less than 5 ppm.
  • the ratio of the main active ingredients in the extract of Ginkgo biloba leaves, SP, Ginkgo biloba total flavonol glycosides and ginkgo total lactone content ratio is 24: 25-10, preferably 24: 20-10, more preferably 24: 15, medicine Efficacy experiments have shown that the ratio of Ginkgo biloba extract in the treatment of ischemic cerebrovascular disease and senile dementia is significantly better than the international standard Ginkgo biloba extract EGb761 total flavonoids and total lactones. The ratio is 24:6.
  • the saffron extract can be prepared by any known method, and the present invention preferably adopts the following extraction method:
  • the saffron raw material is added with at least 5 times (preferably 20 times) of 60-80% ethanol, 60-90 ° C extraction (preferably 80 ° C) at least 1 time (preferably 3 times), at least 1 hour each time (preferably 2 hours), filtered, combined extracts, concentrated to an alcohol-free flavor and diluted with water equivalent to more than 1 times the weight of the crude drug, filtered, and added to the weak polar polystyrene macroporous adsorption resin column (for example) Type AB-8), first eluted with water, eluted with low to high ethanol below 30% (for example, eluted with 20% ethanol, continued to elute with 30% ethanol), and finally eluted with 70% ethanol.
  • the drug-loaded resin discarding water, 30% ethanol eluate, and collecting 70% ethanol eluate to obtain saffron extract containing saffron total glycosides.
  • the present invention preferably elutes sequentially with 20% and 30% ethanol. On the surface, the difference between the two is 10% ethanol concentration. The key is that the inventors are In the experiment, it was found that 20% elution and 30% ethanol elution had little loss of crocin, while directly eluting with 30% ethanol or eluting with other concentrations of ethanol would lose a large amount of crocin.
  • Soy alcohol extract can be obtained by the following extraction method:
  • the raw soybean is extracted with 85-95% ethanol, filtered, and the residue is extracted with 60 80% (preferably about 70%) of ethanol, filtered, and the ethanol extract is combined and concentrated to an alcohol-free taste, and the weight of the raw material is added 1 time (preferably 2 times) above water, fully stirred, filtered, weakly polar polystyrene-type macroporous adsorption resin on the filtrate, such as AB-8 type macroporous adsorption resin, preferably eluting the drug-loaded resin with water, discarding water
  • the eluate is eluted with 50-65% (preferably about 60%) of ethanol, and the eluate is collected to obtain component A.
  • the components in A are mainly isoflavones; and then eluted with 90-95% ethanol, collected and washed. Deliquoring, recovery to dryness, adding absolute ethanol for esterification, washing with water, layering the solution, removing the lower aqueous solution (acid water) and depressurizing to 0.
  • IMPa degassing preferably using a rotary concentrator to reduce pressure to 0
  • IMPa degassing adding sodium hydroxide for alcoholysis, washing with water, discarding the lower layer of washing liquid (alkaline water), and depressurizing the upper layer of organic liquid to 0.
  • IMPa degassing and thin film distillation for the purpose of removing fatty acid ethyl ester , 133Pa, steaming, the residual liquid is subjected to molecular distillation (preferably at a pressure of 0.133 Pa, steamed) Condensation plate to plate distance of less than 0. 5mm), to give the component B, B component is mixed in the raw After the phenol is added, B and A are then mixed to obtain the soybean extract of the present invention.
  • the above extraction method of the extract of ginseng, ginkgo leaf, saffron and soybean is the preparation method of the main component in the capsule of the composition of Example 1.
  • the soybean extract of the present invention is mainly composed of soybean total isoflavones and vitamin E in a weight ratio of 4:2 to 0.5, preferably about 4:1.
  • the invention also provides a medicament for treating ischemic cerebrovascular disease and senile dementia, comprising the above traditional Chinese medicine composition and a pharmaceutically acceptable auxiliary material, and the selected auxiliary materials are also different according to the pharmaceutical dosage form, in the medicament, Ginseng extract active ingredient (ginseng total saponin) based on ginsenoside Re (C, fi H 82 0 IS ), not less than 13. 75mg / 0.
  • Ginseng extract active ingredient ginseng total saponin
  • C, fi H 82 0 IS ginsenoside Re
  • Ginkgo biloba active ingredient including ginkgo total lactone
  • ginkgo Ester A C 2 .H 24 0 9
  • Ginkgolide B C 2 .H 2i A confusion
  • Ginkgolide C C 2 .H 24 0u
  • Ginkgolide C 15 H l8 0 8
  • the weight of the drug is not less than 2.75 mg / 0.15 g, and the above-mentioned 0.15 g refers to the weight of the finished drug.
  • the above pharmaceutical preparation may be in any dosage form, preferably an oral dosage form, which is all available in a pharmaceutical form, preferably a granule, a capsule, a tablet, an oral solution and a syrup; in the embodiment of the present invention More preferred are granules and capsules.
  • the pharmaceutical composition of the invention was further studied by the applicant on the basis of the present invention. It was found that the ratio of total flavonol glycosides and total lactones in Ginkgo biloba extract EGb761, which is commonly used internationally, is 24:6. However, in the applicant's pre-pharmacodynamic experiments, it was confirmed that the optimal ratio of total flavonol glycosides and total lactones in the treatment of ischemic cerebrovascular disease and senile dementia in Ginkgo biloba extract was 24:15.
  • the study further analyzed the orthogonal design of ginseng extract, ginkgo biloba extract, saffron extract and soybean extract with actual error, and induced D-galactose-induced brain aging mice and normal small
  • the mouse is an animal model, and the Morris water maze test is used as a tool to detect the memory level of the animal, and the behavioral test results of the animal are comprehensively investigated.
  • the comprehensive analysis showed that the compatibility of the four components was better than that of the single component.
  • the invention also provides a preparation method of various oral dosage forms of the above drugs, which comprises:
  • Granule type Add each prescription amount of mixed paste to dextrin or other binder, mix well, dry at 60-8CTC, pulverize; mix well with flavoring agent (such as stevioside), granulate, Thousands of dry, that is.
  • flavoring agent such as stevioside
  • B. Tablets Add the mixed paste to dextrin or other binder, mix well, dry and pulverize at 60-80 ° C; with appropriate amount of binder (such as starch) and disintegrant (such as sodium carboxymethyl starch) Mixing, granulating, drying, adding appropriate amount of lubricant (such as magnesium stearate), disintegrating agent (sodium carboxymethyl starch), mixing, tableting, film coating as needed, that is.
  • Capsule Add each prescription amount of paste to dextrin or other binder, mix well, dry, smash; mix with flavoring agent (such as stevia), proper amount of filler (such as starch), dry , into the capsule.
  • D. Pills Add each prescription amount of the mixed paste to dextrin or other binder, mix well, dry, and pulverize; add honey or water or beeswax or rice flour or rice paste, according to the preparation method of other conventional pills.
  • the composition of the present invention can also be made into a honey pill, a water pill, a water pill, a paste pill, a wax pill, and a concentrated pill.
  • the process steps are all routine operations, and the process conditions can be changed as appropriate depending on the condition of the medicinal material. It is well known to those skilled in the art.
  • the preparation steps of the medicament of the present invention may be different depending on the desired product, but they are all known common-sense preparation processes and will not be described one by one.
  • the technical conditions of extraction are determined by using a three-factor three-level table for orthogonal test using the yield and effective component content as indicators.
  • the Chinese medicinal materials used in the composition of the present invention are all medicinal materials contained in the 2005 edition of the Pharmacopoeia, and all the indicators are in compliance with the Pharmacopoeia; according to the Chinese Pharmacopoeia 2005 edition, Appendix IXE, IXF, The arsenic salts and heavy metals of the three batches of samples were examined and the results were within the specified ranges.
  • the medicament of the present invention is examined by hygiene and conforms to the pharmacopoeia hygiene standards.
  • the present invention also provides the use of the above composition for the preparation of a medicament for the treatment of ischemic cerebrovascular disease and senile dementia.
  • This study suggests that the drug may have different effects than the current clinical use of AChEI (acetylcholinesterase inhibitor) and EGb761 (formal name: Ginkgo biloba extract tablets, ie Ginkgo biloba extract), showing better efficacy Therefore, it has more market advantages.
  • the mechanism of analysis may be that the drug acts on multiple targets of ischemic cerebrovascular disease and Alzheimer's disease, including the abnormal expression of the ⁇ -amyloid precursor protein (APP) gene, which is an upstream link of AD lesions, which is effective for Chinese medicine.
  • APP ⁇ -amyloid precursor protein
  • the capsule drug of the embodiment 1 of the present invention (hereinafter referred to as the drug of the present invention);
  • Reference drug Haberin (English name: Huperzine A Tablets, official name: Huperzine A tablets, Henan Zhulin Zhongsheng Pharmaceutical Co., Ltd., main components: Huperzine A C l5 H ls N 2 0); Dana Kang (English name: tanakan, official name: Ginkgo biloba extract tablets, produced by Beaufort-Epson Pharmaceutical Industries, France); Wei Naikang (laboratory, containing ginseng, ginkgo biloba and saffron extract).
  • the experimental method was carried out according to the method of "Morris RG, Garrud P, Rawlins JNP et al. Place navigation impaired in rats with hippocampal lesions. Nature; 297: 681-3.”.
  • the learning and memory abilities of the rats in the three dose groups of the present invention were significantly improved, and the time required to travel the maze was significantly shorter than that of the model group (0.05-0.01); Wei Naikang, Haberin The group also had the same effect (0.05-0.01); 2 months after the administration, the learning and memory ability of the control drug Danakang group was significantly improved (0.05); compared with the Weikankang group, the drug of the present invention was administered 1 to 2 After the month, the learning and memory abilities of the rats in the large and middle dose groups were significantly improved, and the time required to travel the labyrinth was significantly shorter than that in the model group (P ⁇ 0.05-0.01).
  • the present invention also provides a pharmacodynamic test for chemical damage caused by scopolamine, chlorpromazine, reserpine or sodium nitrite.
  • the mouse jumping test was carried out with the escape latency and the number of errors within 5 (10) miri.
  • both indicators were improved to varying degrees, indicating that the drug of the present invention may have an effect of improving the acquired and consolidation memory impairment of the model mice.
  • the ACh content in the whole brain of the model group was significantly decreased after injection of ⁇ ⁇ ⁇ in the bilateral hippocampal CA1 area (0.01).
  • the ACh content in the whole brain of the rats in the group of the present invention was significantly increased (0.05-0.01); the ACh 7 level in the whole brain of the positive drug Haberin group was increased. But there is no significant difference.
  • Table 5 Effect of whole brain ACh content in rat model of A ⁇ injury (soil SD) Group dose (rag/kg) n ACh content (yg/l) Sham operation group 6 242.8 ⁇ 39.7
  • the medicament of the invention 11.5 6 211.2 ⁇ 39.3*
  • the medicament of the invention 23 6 227.8 ⁇ 54.1*
  • the ACh decreased in the brain 3 months after 2V0, which was significantly different from that in the sham operation group (0.01).
  • the ACh in the brain of the rats in each group was significantly increased (P ⁇ 0.05). - 0.01); Weikangkang, Haberin and Danacon have the same effect (P ⁇ 0.05-0.01); compared with the Weikangkang group, the ACh in the brain of the large and middle dose groups of the invention is significantly improved (P ⁇ 0.05-0.01).
  • Table 6 The results are shown in Table 6.
  • the drug group of the invention 11.5 10 183.3 ⁇ 22.7**
  • the drug group of the invention 23 10 201.2 ⁇ 31.2** ⁇
  • the drug group of the present invention 46 10 218.1 ⁇ 34.9** ⁇
  • the ACh content in the whole brain or hippocampus of the animal is significantly increased, suggesting that the drug of the present invention can regulate the release and degradation of the ACh in the AD, VD animal model, increase the central ACh level, and thereby improve the central bile.
  • the ACh level of the central nervous system of the 2V0 rat was significantly increased compared with the retinoic acid of the present invention, indicating that the effect of adding the soybean component to the composition of the present invention is better.
  • the low level of monoamine neurotransmitter metabolism in rat brain with A ⁇ toxicity injury is similar to that in aging and AD patients, indicating that memory impairment in rats with ⁇ toxic injury may be related to monoamine neurotransmitter metabolism in the brain.
  • the drug of the present invention may delay the degradation of DA and 5-HT, and the relative levels of central DA and 5-HT are relatively increased, thereby improving the activity of the monoaminergic system in the brain.
  • DA, D0PAC, 5-HT and 5-HIAA After 12 weeks of administration of the drug of the present invention, DA, D0PAC, 5-HT and 5-HIAA all showed an increasing trend, and the levels of DA and 5-HT in the hippocampus of the high-dose group of the present invention were significantly elevated, indicating that the drug of the present invention may be improved.
  • the effect of DA, 5-HT system activity may be achieved by inhibiting the uptake of DA and 5-HT.
  • the results also indicate that the mechanism of the drug of the present invention for increasing the levels of DA and 5-HT may be different from that of Haberin and Danacon.
  • the effect of Haberin may inhibit the metabolism of DA in neurons and inhibit the degradation of 5-HT.
  • the role of Danacon may inhibit the metabolism of DA in neurons and inhibit the involvement of 5-HT uptake. .
  • the drug of the present invention has the functions of regulating the levels of NE, DA, 5-HT and regulating the activities of NE, DA and 5-HT systems.
  • the role of improving learning and memory may be related to the intervention of monoamine transmitters and degradation.
  • the detection method was carried out according to the instructions of each kit.
  • the MDA content in the whole brain of the elderly control group was significantly increased (0.05).
  • the MDA content in the whole brain of the rats in the drug administration group of the present invention was significantly decreased (0.05); the content of MDA in the whole brain of the positive drug Haberin and Danakang rats were all The trend is reduced, but there is no significant difference (AO.05).
  • Table 9 The results are shown in Table 9.
  • the medicament of the invention 11.5 7 46.84 ⁇ 3.47*
  • AChE activity of AChE in the whole brain of rats with natural aging cognitive impairment was significantly reduced, consistent with changes in aging and AD.
  • the medicine of the invention The substance can significantly increase the decreased AChE activity and regulate the abnormal metabolic state of ACh, which may improve the central cholinergic nervous system function of aging or AD, and improve learning and memory dysfunction.
  • the drug of the present invention may have the effect of improving the antioxidant and free radical scavenging ability of the aged animal body, which may contribute to anti-aging and improvement. Memory capacity.
  • the drug of the present invention may have the function of protecting ATPase, regulating cell transport function, thereby improving the function of neuronal plasma membrane and improving cell function. effect.
  • the hippocampal pyramidal cells were arranged closely, the cells were prominent in the upper and lower lines, and the cells were abundant; the nuclear membrane was clear and the nucleolus was obvious.
  • the hippocampal pyramidal cells were loosely arranged, the cell structure was fuzzy, the cell body was swollen, and the cells were irregular in line; some of the cells were pyknotic and deep-stained.
  • the hippocampal pyramidal cells of the positive drug Haberin group and the rats of the present invention were arranged neatly, and the cell outline was clear; the degree of nuclear pyknosis and deep staining was alleviated.
  • Haberin and the high- and medium-dose groups of the present invention have obvious effects.
  • the hippocampal pyramidal cells were densely arranged, the cells were lined up neatly, and the cells were full; the nuclear membrane was clear and the nucleolus was clearly visible.
  • the hippocampal pyramidal cells were loosely arranged, and some of the cell structures were incomplete; a few nuclei were shrunk and deep-stained, and they were triangular; compared with the elderly control group, Danacon and the rats of the present invention were hippocampus The level of somatic cells is clearer, the cell morphology is improved to varying degrees; the degree of nuclear pyknosis and deep staining is reduced. Among them, Danacon and the high- and medium-dose groups of the present invention have obvious effects.
  • the method was carried out according to the first and fourth pages of Modern Medical Experimental Methods edited by Wang Qian.
  • the ultrastructure of hippocampal neurons was normal, the nucleus was round or elliptical, and the euchromatin was uniformly distributed. The nucleolus was visible and the nucleus was intact. Cytoplasmic mitochondria and rough endoplasmic reticulum structure Clear; ribosomes, synapses are more abundant.
  • the hippocampal neurons in the hippocampus were collected and condensed; the cell volume was reduced, and the cytoplasm was concentrated; a few cells showed mild mitochondrial swelling; the rough endoplasmic reticulum, ribosome and other organelles were basically normal; presynaptic and posterior membranes were not Clear, synaptic gap disappeared, and synaptic vesicles were significantly reduced compared with the sham operation group.
  • the morphology and structure of hippocampal neurons in different groups of Haberin and the present invention were improved, and the membranes and synaptic membranes were clearer; most of the nuclear chromatin distribution was uniform; mitochondria were clearer.
  • the hippocampal neurons maintained normal morphological features.
  • the nucleus was round or elliptical, and the chromatin was evenly distributed, showing obvious nucleoli.
  • the cytoplasmic mitochondria and rough endoplasmic reticulum were clear; ribosomes and neurites Rich in touch.
  • the membrane was dissolved, and vacuoles and myeloid structures were observed.
  • the shape of the nucleus was irregular, the chromatin was agglomerated into blocks, and the perinuclear space was thickened.
  • the mitochondrial swelling was vacuolized. Quality network expansion; more common lipofuscin, oil droplets.
  • the positive drug Haberin, Danacom and the drug of each group have improved cell morphology and structure.
  • the membrane is clearer; some of the nuclear chromatin are evenly distributed; the mitochondrial lamellar layer is clear; the ribosome is increased; the number of synapses is increased.
  • Haberin, Danacon and the high- and medium-dose groups of the present invention have obvious effects.
  • the ultrastructure of hippocampal neurons in Tongwosheng non-transgenic mice is normal, the nucleus is round or elliptical, and the euchromatin is evenly distributed, showing obvious nucleoli; the mitochondria and rough endoplasmic reticulum in the paddle are clear; Rich in body; more synapses.
  • the nuclei of the blank group were multi-contracted, marginal, and irregularly arranged; the mitochondria were few, the sputum was disordered, and the sputum gap was enlarged; the rough endoplasmic reticulum was disordered and even disintegrated; the amount of lysosomes increased, and the shape was irregular; Vesicles and myeloid structures can be seen in the cytoplasm, and irregularly shaped lipofuscin can be seen.
  • the morphology and structure of the high-dose group of the present invention were improved, and some of the membranes were clear; some of the nuclear chromatin were evenly distributed.
  • the integrity of the morphological structure of hippocampal neurons is a prerequisite for maintaining its normal function.
  • Pathological factors such as ischemic cerebrovascular disease, aging or senile dementia cause damage to the morphological structure of nerve cells, which inevitably leads to abnormal functions, such as nuclei and rough surfaces. Damage to the protein network leads to a decline in protein synthesis, damage to the mitochondria leads to energy metabolism disorders, and a large amount of lipofuscin accumulation destroys the spatial order of the organelles. Loss of synapses means that the nerve cells lose their target tissues, eventually leading to senescence and death of nerve cells.
  • D-galactose-induced brain aging rats and natural aging cognitive impairment rats were used for HE staining, and A ⁇ -toxic injury rats, natural aging cognitive impairment rats, and APP transgenic mice were used as models for transmission.
  • the effect of the drug of the present invention on hippocampal neurons of model animals was investigated by electron microscopy. The results showed that the morphology and structure of hippocampal neurons and synapses were improved to different degrees after administration of different model animals. The number of synapses increased, and the apoptosis and loss of nerve cells were delayed or decreased, suggesting that the drug of the present invention may have protective nerves. The role of cells.
  • the rats were intragastrically administered twice a day (24h) at a maximum concentration (57.5 mg/ml) and a maximum volume (20 ml/kg body weight).
  • the maximum dose in one day was 2300mg/kg body weight, equivalent to 670 times the clinical dose (240mg / day).
  • the rats were continuously administered for 6 months at a dose of 70, 35 and 17. 5 times of the clinical dose, and the general condition, body weight and food intake were administered 3 months and 6 months after the administration and 4 weeks after the drug withdrawal. No obvious pathological changes were found in blood routine, blood coagulation, blood biochemistry, electrocardiogram, major organ index and gross and microscopic examination. Toxicological studies have shown that the drug of the present invention is safe and low in toxicity.
  • the therapeutic effect of the composition of the present invention on ischemic cerebrovascular disease and senile dementia is superior to the currently used drugs, and the effect is improved after the treatment.
  • the long-term stability investigation also indicates that the composition of the present invention is stable and reliable. . detailed description
  • Example 1 Formulation of the composition (extract) of the present invention
  • the dried ginkgo biloba leaves were added with 8 times the amount of 70% ethanol, and the mixture was extracted twice at 60 ° C, and extracted twice, each time for at least 1 hour, and the extracts were combined, and the extract was concentrated under reduced pressure to a relative density of about 1.05 (5 CTC).
  • the liquid was cooled with water, filtered, and the filtrate was coated with ADS-17 large-cell adsorption resin.
  • the drug-loaded resin was eluted with water, and then eluted with 60% ethanol. The water eluate was discarded and 60% ethanol eluate was collected.
  • the saffron raw material is added with 20 times of 60% ethanol, and extracted twice at 70-80 ° C, and the combined extracts are filtered, concentrated to a non-alcoholic flavor and diluted with water more than 1 time, and the diluted solution is added to AB- On the column of type 8 macroporous adsorption resin, first elute with water, then elute with 20% ethanol, continue to elute with 30% ethanol, finally elute the drug-loaded resin with 70% ethanol, discard the water, wash with 30% ethanol. The solution was deliquored, and a 70% ethanol eluate was collected to obtain a saffron extract containing saffron total glycosides.
  • Soybean is extracted with about 95% ethanol, filtered, and the residue is extracted with 70% ethanol. It is filtered, and the ethanol extract is concentrated and concentrated to an alcohol-free taste. The water is added twice the weight of the raw material, stirred well, filtered, and filtered.
  • AB- 8 type macroporous adsorption resin first elute the drug-loading resin with water, discard the water eluent, elute with 60% ethanol, collect the eluent to obtain component A, and the components in A are mainly isoflavones.
  • the dried extract is mixed and pulverized to 20 mesh, and starch is added to 86.75 g, and the capsule No. 3 is charged, thereby obtaining the present invention.
  • the identification and content determination of ginkgolides refer to the Chinese Pharmacopoeia 2005 edition of a ginkgo leaf extract; the identification and content determination of crocin-I refer to the Chinese Pharmacopoeia 2005 edition of a saffron; The identification and content determination of ginseng is based on a Chinese Pharmacopoeia 2005 edition of a ginseng.
  • Determination method of total isoflavone and genistein in soybean Accurately weigh the soybean extract and mix it with 25ml water, add 1ml acetate buffer solution ( P H4,5) and 15 ⁇ 1 ⁇ -glucosidase, 37° Hydrolysis in C water bath, solvent recovery to 1000, and dissolved in methanol, filtered through a microporous membrane, and analyzed by liquid chromatography (Zorbax-( ⁇ column, mobile phase is methanol-water-acetic acid 45: 55: 1, Flow rate 0.8ml/min, monitoring wavelength 260nm);
  • Determination method of VE content Take appropriate amount of this product, accurately weigh it, put it in the mortar and add two drops of absolute ethanol, and transfer it to the powder funnel with 20ml of absolute ethanol. Add 10ml of water and extract with n-hexane. 3 times, 5ml each time, combined with n-hexane extract, the solvent was recovered to dryness under reduced pressure, and the residue was quantitatively transferred to a 10 ml volumetric flask by mobile phase. The mobile phase was adjusted to volume, shaken, and filtered through 0.45 ⁇ m. The membrane, as the test solution, accurately draws 10 l of the reference solution and the test solution, respectively, and injects into a liquid chromatograph to determine (97: 3 ⁇ / ⁇ methanol-water as mobile phase, detection wavelength is 207 nm).
  • Each capsule of the capsule of the composition of the present invention contains the following ingredients (in terms of 1.5 g per capsule):
  • the total flavonoids are in rutin (C 27 H 3 personally0, s ), not less than 11.00 mg.
  • the total flavonol glycosides are not less than 6.60 rn g in terms of quercetin, kaempferol and isorhamnetin.
  • the total glucosides containing saffron are not less than 2.75 mg based on anhydrous safflorin-1 (3 ⁇ 4mon0 ⁇ ).
  • Vitamin E containing vitamin E (C 31 H 52 0 3 ), not less than 0.5 mg.
  • the clinically recommended amount of the composition capsule of the present invention is 150 mg/time, 3 times/day.
  • composition Preparation Method The above-mentioned medicinal material is pulverized over 20 mesh to obtain the composition of the present invention.
  • Preparation of granules After mixing the above powders, dextrin and stevioside are added, thoroughly mixed, dried under vacuum at 70 to 75 Torr, pulverized, and granulated to obtain granules of the composition of the present invention, which are brown granules.
  • the content was determined by referring to Example 1, and not less than 9.15 mg/g based on ginsenoside Re.
  • Prescription 2 parts of ginseng extract, 10 parts of Ginkgo biloba extract, 0.5 parts of saffron extract, 1 part of soybean extract.
  • Preparation method of tablet After the extract of the composition of the present invention is mixed, a binder such as dextrin is added, and the mixture is thoroughly mixed, vacuum-dried at 60-80 ° C, pulverized, and a filler such as starch or magnesium stearate is added.
  • a lubricant such as a disintegrant such as sodium carboxymethylcellulose is mixed, granulated, and tableted to obtain a tablet of the present invention.
  • Prescription 10 parts of ginseng extract, 3 parts of apricot leaf extract, 4 parts of saffron extract, and 0.2 parts of soybean extract.
  • the preparation method of the composition is the same as in the first embodiment.
  • Preparation of tablets After the above-mentioned extracts of the composition of the present invention are mixed, a binder such as dextrin is added, and the mixture is thoroughly mixed, vacuum-dried at 60-80 ° C, pulverized, and a filler such as starch or magnesium stearate is added.
  • a lubricant such as a disintegrant such as sodium carboxymethylcellulose is mixed, granulated, and tableted to obtain a tablet of the present invention.
  • Preparation method The above medicinal material is pulverized through 20 mesh to obtain a composition of the present invention.
  • the content was determined by referring to Example 1 and not less than 9.15 mg/g based on ginsenoside Re.

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Description

一种中药组合物及其制备方法和用途 技术领域
本发明涉及一种中药组合物， 具体地说， 是一种用于治疗缺血性脑血管病和老年痴呆的 中药组合物，该组合物的组分可以是直接粉碎成粉末入药的中药材和 /或经提取得到的中药提 取物。 背景技术
脑血管病通常分为缺血性脑血管病和出血性脑血管病两大类， 以缺血性脑血管病为多 见， 其中脑梗塞占 59. 2%〜85%。 缺血性脑血管病包括： （1)短暂性脑缺血发作（简称 TIA, 又 叫小中风或一过性脑缺血发作）， 其病因与脑动脉硬化有关， 是脑组织短暂性、 缺血性、 局灶 性损害所致的功能障碍； （2)脑血栓的形成， 多由动脉粥样硬化、 各种动脉炎、 外伤及其他物 理因素、 血液病引起脑血管局部病变形成的血凝块堵塞而发病； （3)脑栓塞， 可由多种疾病所 产生的栓子进入血液， 阻塞脑部血管而诱发。
现用于治疗缺血性脑血管病的药物很多， 西药多为溶栓、 抗血小板、 抗凝血药； 中药主 要有以丹参、三七皂苷为代表的活血化瘀中药注射液； 以醒脑静、 清开灵为代表的清热解毒、 醒脑开窍中药注射液类； 以人参再造丸、 华佗再造丸为代表的益气活血通络的口服制剂。 但 在临床应用中， 西药主要用于急症， 有明显的副作用； 中药注射液则无法长期用药， 中药复 方又有疗效不确切、 有效成分不明确、 缺乏稳定的质控标准等诸多问题。
痴呆是由于大脑器质性病变引起的一种获得性、 持续性智力损害综合征。 2005年全球流 行病学调査结果显示， 目前约有 2400万人患有痴呆， 并以每年 460力-人的速度增长 （每 7秒增 加 1人） ， 平均每 20年翻一番。 而在中国， 保守怙计从 2001年到 2040年， 将以每年 300% 的速 度递增。 预计 2040年， 全球受痴呆困扰的人数将达 8100万。 痴呆的发病率随年龄增长而增长， 发生于老年人的痴呆主要分为： A.原发性退行性痴呆， 即阿尔茨海默病（Alzheimer' s disease, 简称 AD)； B.血管性痴呆 (Vascular dementia, VD)； C.混合性痴呆 （AD合并 VD); D.其他痴呆 （匹克病、 路易包涵体痴呆） 。 老年痴呆中 AD和 VD是两大最主要的类型， 患病率 占所有痴呆的 90%以上， 其中 AD是 65岁以上老年人痴呆最常见的原因， 极有可能居于致死性疾 病之首。
目前治疗老年痴呆的药物临床首选胆碱酯酶抑制剂 （例如 tacrine、 donepezil , rivastigmine和 galantamine等临床首选治疗药物）。 中医关于老年痴呆的诊断多包括了西医 的 AD、 VD或二者的混合型， 即所针对的老年痴呆为泛指的全部相关病症。 中药治疗老年痴呆 的常用复方有定志小丸 （人参、 茯苓、 菖蒲、 远志）、 调心方 (党参、 茯苓、 甘草、 石菖蒲、 远志等）、 补肾方 (天冬、麦冬、 生地、 熟地、 山茱萸等）、 当归芍药散、黄连解毒汤、 钩藤散、 抑肝散、 小柴胡汤、 柴胡加龙骨牡蛎汤等。 但在临床应用中， 胆碱酯酶抑制剂仅能部分改善 早、 中期患者的认识障碍和情绪症状， 对基本病理改变均无明显影响， 只能延缓 （约 1-2年） 而不能阻止疾病的进展， 即仅能治标而不能治本， 长期应用反而可能引起胆碱酯酶合成增加， tacrine更可能导致严重的胃肠道反应和肝脏毒性； 中药复方存在有效成分不明确、长期用药 不便、 缺乏稳定的质控标准等问题。
由此可见， 目前治疗缺血性脑血管病和老年痴呆的西药、 中药均存在明显局限与不足， 因而市场急需幵发一种疗效确切并且显著、 工艺精良、 药品质量稳定的治疗缺血性脑血管病 和老年痴呆的药物。 发明内容
本发明人源于中医理论， 总结多方经验， 经过临床实践经验化裁而成。 本发明为中药组 合物， 是由四种天然植物经过提取精制而得的纯中药制剂， 实验证明其疗效确切， 安全有效。
本发明的目的在于提供一种中药组合物及包括该组合物的药物， 该药物可以治疗缺血性 脑血管病和老年痴呆。
本发明的目的还在于提供上述药物的制备方法， 通过该方法， 可得到用于治疗缺血性脑 血管病和老年痴呆的疗效显著、 质量稳定的药物组合物。
本发明的目的还在于提供上述药物组合物用于治疗缺血性脑血管病和老年痴呆的用途。 为了达到上述目的， 本发明提供了一种中药组合物， 包括人参 1-10份、银杏叶 1-10份、 西红花 0. 05- 0. 5份、 大豆 5- 10份， 所述人参、 银杏叶、 西红花和大豆的来源可以是中药材 或相当于上述中药材生药量的中药提取物。
本发明的上述组合物是针对缺血性脑血管病和老年痴呆设立的系统治疗方药， 其为中药 复方组成， 注重于自身免疫体系的提高， 原料药的药源广泛， 无毒副作用， 组方合理， 疗效 良好。
上述的中药组合物， 其中组分的重量份优选为： 人参 2-6 份、 银杏叶 3-6 份、 西红花 0. 06-0. 2份、 大豆 7-8份； 更优选人参 4. 0份、 银杏叶 4. 5份、 西红花 0. 1份、 大豆 7. 5份。
由于上述组合物中各中药材提取收率各不相同， 所以， 本发明还提供一种组合物， 其中 包括人参提取物 1-10份、银杏叶提取物 1-10份、西红花提取物 0. 5-5份、大豆提取物 0. 1-1 份， 以上所述的份为重量份数， 所述人参、 银杏叶、 西红花和大豆提取物均为醇提取物。
本发明的组合物处方合理， 按照中国传统医学的方解， 组合物中的人参为方中君药。 本 发明组合物主治的 AD本虚标实， 主要与脏腑功能虚损、 气血运行失常有关， 而脏腑亏虚、 浊 毒痹阻脑络是其主要病理基础， 贯穿该病始终。 方中君药人参 (有效成分为人参总皂苷) 大 补元气， 安神增智； 臣药银杏叶 （有效成分主要为银杏总黄酮和银杏总内酯）、 西红花（主要 有效成分西红花总苷） 活血化瘀排毒； 佐药大豆 （主要有效成分为大豆总异黄酮和维生素 E) 祛除 "内生之毒"， 全方益气活血， 排毒通络， 安神增智， 标本兼顾。
本发明的组合物中， 可选择药味全部生粉入药， 也可提取后以提取物的形式入药， 只要 包括了上述重量配比的组分， 该组合物即可达到治疗缺血性脑血管病和老年痴呆的药效， 即 在本发明所保护的范畴之内。 在本发明的优选实施例中， 人参、 银杏叶、 西红花和大豆均为 该原料药的乙醇提取物。
在本发明的优选实施方案中， 上述药物组合物的组分和配比（重量）优选为人参提取物： 银杏叶提取物： 西红花提取物： 大豆提取物为 5 : 5： 1： 0. 5。
其中人参提取物可釆用任何公知方法制备， 本发明优选下述提取方法- 人参加至少 2倍 （优选 8倍） 生药量低浓度 （如 50-70% , 优选 60%) 乙醇， 提取至少 1 次（优选 3次），每次至少 1小时 (优选 3小时），合并提取液，浓缩至相对密度约 1. 05 (50°C)， 浓缩液加至少 1倍 （优选 2倍） 体积的蒸镏水， 滤过， 滤液上弱极性聚苯乙烯型大孔吸附树 脂（优选 AB- 8型）， 以蒸馏水洗脱，继续以 10%乙醇洗脱载药树脂， 弃去水和 10%乙醇洗脱液， 载药树脂继续以 70%乙醇洗脱， 大约至 2. 5倍柱体积， 收集 70%乙醇洗脱液， 即得含有人参总 皂苷的人参提取物。
本发明采用将载药大孔吸附树脂分别以水洗脱、 10%乙醇洗脱， 最后以 70%乙醇洗脱， 筛 选上述两个乙醇浓度和洗脱程序既能保证中药总成分含量的技术要求， 又可以保证有效成分 能有较高的转移率和成品收率。
银杏叶提取物可采用下述提取方法制得：
取千燥银杏叶加至少 2倍量 （优选 8倍量） 的低浓度乙醇 （60-80 %， 优选 70%) 乙醇， 50-70Ό提取 （优选 60°C ) 至少 1次 （优选 3次）， 每次至少 1小时 (优选 4小时）， 合并提取 液， 提取液减压浓缩至相对密度 1. 05左右（50°C)， 浓缩液加水冷却沉淀， 滤过， 滤液上极性 氢键聚苯乙烯型大孔吸附树脂 （优选 ADS- 17型）， 目的是富集有效成分， 载药树脂先以水洗 脱， 继续 60%乙醇洗脱， 弃去水洗脱液， 收集乙醇洗脱液， 浓縮至无醇味， 加入生药 2倍量 水加热至沸， 室温沉降 24小时， 滤过， 滤液上弱极性聚苯乙烯型大孔吸附树脂（优选 DM-130 型）， 目的是除杂质， 先以水洗脱， 继续 15%乙醇和 60%乙醇洗脱， 弃去水和 15%乙醇洗脱液, 收集 60%乙醇洗脱液； 也可继续浓缩干燥， 得银杏叶提取物成品。 本发明的银杏叶提取物的提取尤其是精制工艺， 突破了目前有机溶剂萃取技术除去银杏 叶提取物中杂质 （主要是银杏酚酸、 多糠、 单糖、 无机盐等） 的局限。 由于银杏酚酸类有害 杂质水溶性极小， 主要出现在 60%以上浓度乙醇的洗脱液中， 而多糖、 无机盐等杂质绝大部 分能被 15 %的乙醇洗脱， 所以通过本发明的两次上大孔吸附树脂的精制步骤， 大部分杂质被 除去， 而且经过 60%乙醇洗脱， 相当于收集 15- 60 %乙醇的洗脱液， 可以保证酚酸含量小于 5ppm, 本发明经此筛选得到的银杏叶提取物中主要有效成分的比例， SP， 银杏总黄酮醇苷和 银杏总内酯的含量比例为 24: 25-10， 优选 24: 20-10， 更优选 24: 15， 药效学实验证明， 该比例的银杏叶提取物在治疗缺血性脑血管病和老年痴呆方面的效果显著优于国际上现通用 的标准银杏叶提取物 EGb761中银杏总黄酮醇苷和总内酯的比例为 24: 6的药效。
西红花提取物可采用任何公知方法制备， 本发明优选以下述提取方法：
西红花原料加至少 5倍量 (优选 20倍量）的 60-80 %乙醇， 60- 90°C提取 （优选 80°C ) 至 少 1次 (优选 3次）， 每次至少 1小时 (优选 2小时)， 滤过,合并提取液， 浓缩至无醇味加相当 于生药重量 1倍以上的水稀释， 滤过， 将滤液加到弱极性聚苯乙烯型大孔吸附树脂柱上 （例 如 AB- 8型）， 先以水洗脱， 再以 30 %以下的乙醇由低至高依次洗脱 （例如以 20%乙醇洗脱， 继续以 30%乙醇洗脱）， 最后用 70 %乙醇洗脱载药树脂， 弃去水、 30%以下乙醇洗脱液， 收集 70%乙醇洗脱液即得含有西红花总苷的西红花提取物。
由于所用原料的价格昂贵， 所以优选制备工艺的前提是确保高收率， 本发明优选用 20% 和 30%乙醇依次洗脱，表面上看两者相差 10%乙醇浓度，关键就在于发明人在实验中发现， 20% 洗脱后再用 30%乙醇洗脱对西红花苷损失很小，而直接以 30%乙醇洗脱或者以其他浓度的乙醇 洗脱会损失大量的西红花苷。
大豆醇提物可釆用下述提取方法制得：
原料大豆以 85- 95%的乙醇提取， 滤过， 残渣以 60 80% (优选 70%左右） 的乙醇提取， 滤 过， 将乙醇提取液合并浓缩至无醇味， 加入原料重量 1倍 （优选 2倍） 以上的水， 充分搅拌， 滤过， 滤液上弱极性聚苯乙型大孔吸附树脂， 例如 AB- 8型大孔吸附树脂， 优选先以水洗脱载 药树脂， 弃去水洗脱液， 再以 50- 65 % (优选大约 60 % ) 的乙醇洗脱， 收集洗脱液得到组分 A， A中成分主要是异黄酮； 再用 90- 95%乙醇洗脱， 收集洗脱液， 回收至干， 加入无水乙醇进 行酯化， 再加水洗涤， 溶液分层， 除去下层水液（酸水） 后减压至 0. IMPa脱气 （优选用旋转 浓缩仪减压至 0. IMPa脱气），加入氢氧化钠进行醇解， 再加水洗涤， 弃去下层水洗液（碱水）， 上层有机液减压至 0. IMPa脱气后进行薄膜蒸馏， 目的是除去脂肪酸乙酯， 残留液进行分子蒸 馏（优选压力为 0. 133Pa， 蒸发板到冷凝板距离小于 0. 5mm)， 得到组分 B , B中成分为混合生 育酚， 之后将 B与 A混合， 即可得到本发明的大豆提取物。
上述人参、 银杏叶、 西红花和大豆的提取物的提取方法即为实施例 1的组合物胶囊剂中 主要组分的制备方法。
经检测， 本发明的大豆提取物主要含有大豆总异黄酮和维生素 E， 其重量含量配比为 4: 2-0. 5， 优选约 4: 1。
本发明还提供了一种治疗缺血性脑血管病和老年痴呆的药物， 其包括上述中药组合物和 药学可接受的辅料， 根据药物剂型的不同， 所选择的辅料也不同， 在药剂中， 人参提取物有 效成分 （人参总皂苷） 以人参皂苷 Re ( C,fiH820IS ) 计， 不得少于 13. 75mg/0. 15g， 银杏叶 有效成分（含银杏总内酯）以银杏内酯 A(C2。H2409)、银杏内酯 B (C2。H2iA„)、银杏内酯 C (C2。H240u ) 和白果内酯 （C15Hl808) 含量之和计， 不得少于 2. 75mg/0. 15g， 所述的 0. 15g是指成药重量。
上述的药物制剂可以是任何剂型， 优选为口服剂型， 其为药学意义上的所有可供口服的 剂型， 优选颗粒剂、 胶囊剂、 片剂、 口服液和糖浆剂； 在本发明的实施例中， 更优选颗粒剂 和胶囊剂。 实验证明， 本发明药物的疗效明显优于已有的治疗缺血性脑血管病和老年痴呆的 药物， 与之相比， 本发明组合物最大程度地降低了用药量， 而且提高了药效， 确实达到了高 效低毒。
为了得到活性成分更稳定、 有效成分含量更！ ¾的药物组合物， 申请人在本发明的基础上 做了进一步的研究， 发现国际上现通用的标准银杏叶提取物 EGb761中， 银杏总黄酮醇苷和总 内酯的比例为 24: 6， 但在申请人前期药效学实验证实， 银杏叶提取物中总黄酮醇苷和总内 酯治疗缺血性脑血管病和老年痴呆的最佳疗效比例以 24: 15为更佳。在此基础上， 本研究再 以实际误差对人参提取物、 银杏叶提取物、 西红花提取物和大豆提取物进行全面分析正交设 计， 以 D-半乳糖致脑老化小鼠和正常小鼠为动物模型， 以 Morris水迷宫实验为检测动物记 忆水平的工具， 综合考察动物的行为学实验成绩。 结果综合分析表明， 四种组分组方配伍的 疗效优于单一组分， 大豆提取物与三组分组方 （人参、 银杏叶、 西红花） 中的人参、 银杏叶 提取物均存在交互作用， 表明对于治疗老年痴呆， 大豆或其提取物在该组方配伍中是必需的。
本发明还提供了上述药物各种口服剂型的制备方法， 其中包括：
A.颗粒剂型： 将每个处方量混合膏加入糊精或其它粘合剂， 充分混匀， 置 60-8CTC干燥， 粉碎； 与矫味剂 （例如甜菊素）， 充分混匀， 制粒， 千燥， 即得。
B.片剂： 将混合膏加入糊精或其它粘合剂， 充分混匀， 置 60- 80°C干燥粉碎； 与适量粘 合剂 (例如淀粉)和崩解剂（例如羧甲基淀粉钠）混匀， 制粒、干燥， 再加入适量润滑剂（例 如硬脂酸镁） 、 崩解剂 （羧甲基淀粉钠） 混匀， 压片， 还可根据需要进行薄膜包衣， 即得。 c.胶囊剂： 将每个处方量混合膏加入糊精或其它粘合剂， 充分混匀， 干燥， 粉碎； 与矫 味剂 （如甜菊素）、 适量填充剂 (如淀粉) 混匀， 干燥， 装入胶囊。
D.丸剂： 将每个处方量混合膏加入糊精或其它粘合剂， 充分混匀， 干燥， 粉碎； 加入蜂 蜜或水或蜂蜡或米粉或米糊， 按照其他常规丸剂的制备方法制得。
根据本领域常规技术， 本发明的组合物也可制成蜜丸、 水蜜丸、 水丸、 糊丸、 蜡丸、 浓 缩丸， 其工艺步骤均为常规操作， 可视药材情况不同酌情改变工艺条件， 其为本领域技术人 员所公知。
针对所需要的产品的不同， 本发明药物的制备步骤也可不同， 但均为公知常识性制剂工 艺， 不再一一描述。
在上述优选实施方案中， 提取的技术条件 （提取的最佳技术参数） 是以出膏率和有效成 分含量为指标， 釆用三因素三水平表进行正交试验来确定的。
本发明的组合物中所釆用的中药材均为 2005年版药典所收载的药材，经鉴定，各项指标 均符合药典规定； 按中国药典 2005年版一部， 附录 IXE、 IXF项下规定， 对 3批样品的砷盐 和重金属进行检査， 结果在规定范围内。
本发明药物经卫生学检査， 符合药典卫生学标准。
本发明还提供了上述组合物在制备治疗缺血性脑血管病和老年痴呆的药物中的应用。 本研究表明，该药可能具有与目前临床使用的 AChEI (乙酰胆碱酯酶抑制剂）以及 EGb761 (正式品名： 银杏叶萃取物片， 即银杏叶标准萃取物） 不同的作用特点， 显示出更佳疗效， 因而更具市场优势。分析机理可能是该药作用于缺血性脑血管病和老年痴呆病变的多个靶位， 包括 β -淀粉样蛋白前体蛋白 （APP)基因异常表达这一 AD病变的上游环节， 为中药有效组分 配伍防治缺血性脑血管病和老年痴呆提供了实验依据。
药效学试验
实验用药： 本发明实施例 1的胶囊剂药物 （以下称为本发明药物）；
对照药： 哈伯因 （英文名： Huperzine A Tablets, 正式品名： 石杉碱甲片， 河南竹林众 生制药股份有限公司生产， 主要成分： 石杉碱甲 Cl5HlsN20); 达纳康 （英文名： tanakan, 正式 品名： 银杏叶萃取物片， 法国博福-益普生制药工业公司生产）； 维脑康 （实验室自制， 含有 人参、 银杏叶和西红花提取物）。
1 行为学实验
1. 1 跳台实验
1. 1. 1 对氢溴酸东莨菪碱致获得性记忆障碍小鼠模型的影响 实验方法按徐叔云等主编的 《药理实验方法学》第三版 "神经系统药物实验法第五节" 项下进行。 与空白组相比， 模型组小鼠 5 niin内错误次数明显增多 （ 0.05)。 给药 15天后, 与模型组比较， 阳性药哈伯因、 达纳康组小鼠错误次数明显减少（ 0.05), 前者潜伏期显著 延长 （ 0.001); 本发明药物高、 中剂量组小鼠错误次数明显减少 （ 0.05- 0.01), 前者潜 伏期明显延长 （ 0.05)。 结果见表 1。 表 1 对氢溴酸东莨菪碱致获得性记忆障碍小鼠模型的影响 （ ±SD) 组别 剂量 (mg/kg) η 潜伏期 （S) 错误次数 空白组 10 279.2±65.8 0.3±0.9 模型组 13 192.8±107.3 1.2±1.3# 哈伯因组 0.08 12 271.8±54.6*** 0.6±0.9* 达纳康组 30 12 233.4±71.5 0.7±0.7* 本发明药物 11.5 12 209.8±96.3 0.9±1.0
本发明药物 23 12 226.3±111.3 0.6±0.7** 本发明药物 46 12 248.5±88.8* 0.5 + 0.7** 注： 与空白组比较: UK0.05； 与模型组比较： *P<Q.05, ** 0.01, *** ο.001。
1.2 避暗实验 ，
实验方法按徐叔云等主编 《药理实验方法学》 第三版 "神经系统药物实验法第五节"项 下进行。 与空白组相比， 模型组小鼠 5 min内错误次数明显增多 （ 0.05), 给药 15天后， 与模型组比较， 达纳康组小鼠潜伏期明显延长， 错误次数减少； 本发明高、 中剂量小鼠错误 次数明显减少 05)。 结果见表 2。 表 2 对乙醇致再现性记忆障碍小鼠模型的影响 （ 土 SD) 组别 剂量 (tng/kg) n 潜伏期 （s) 错误次数 空白组 10 234.3±130.3 0.4±0.9 模型组 10 204.9±102.4 2.1±2.0# 哈伯因组 0.08 10 214.1±127.8 1.0±1.3 达纳康组 30 10 255.8±69.5* 0.4±0.5* 本发明药物 11.5 10 221.5±92.6 1·3±1.1 本发明药物 23 10 212.2±128.2 0.7±0.7* 本发明药物 46 10 253.7±65.2 0.6±0.7* 注： 空白组比较： # 0.05; 丄 j模型组比较： * 0.05o
1.3 Morris水迷宫实验
实验方法按文献 "Morris RG , Garrud P, Rawlins JNP et al. Place navigation impaired in rats with hippocampal lesions. Nature; 297: 681-3. " 中的方法进行。
1.3.1 对 β -淀粉样蛋白 （Αβ ) 毒性损伤大鼠模型空间学习记忆的影响 双侧海马 CA1区注射 Α β Μο4周后， 模型组大鼠在 Morris水迷宫中的游泳持续时间、 路 径长度延长， 与假手术组比较均有显著差异 （ 0.05)， 搜索策略多为边缘式或随机式。 给药 4周后， 与模型组比较， 本发明药物大剂量组大鼠游泳路径长度明显缩短 （ 0.05)， 搜索策 略多为趋向式； 中剂量组大鼠游泳持续时间、 路径长度均显著缩短（ 0.05)， 搜索策略多为 趋向式。 结果见表 3。 表 3 对 Αβ毒性损伤大鼠模型空间学习记忆能力的影响 （ ±SD) 组别 "Γ、 n 持续时间 路径 L<：度 假手术组 11 9.3±5.1 268.9±186.9 模型组 11 23.0±9.1U 696.3±227. 哈伯因组 0.08 11 16.3±10.6 436.7±284.4* 本发明药物 11.5 11 11.6±10.0* 335.7±413.6* 本发明药物 23 11 11.3±5.6* 301, 1±179.1* 本发明药物 46 11 10.1 + 7.0* 280.5±250.6* 注： 与假手术组比较 #^0.05, ΚΟ. ΟΙ; 与模型组比较 * 0.05。
1.3, 2对大鼠双侧颈总动脉永久性结扎 （2V0)所致 VD大鼠模型空间学习记忆的影响
大鼠双侧颈总动脉结扎后 1个月， 虽然出现学习记忆障碍的趋势， 伹与假手术组比较无 明显差异。 结扎 2、 3个月后， 模型组大鼠游迷宫所需时间显著长于假手术组 （ 0.01), 表 明随着缺血时间的延长大鼠学习记忆能力显著降低。 给药 1至 2个月后， 本发明药物三个剂 量组大鼠学习记忆能力均明显提髙， 游迷宫所需时间与模型组比较明显缩短（ 0.05-0.01); 维脑康、 哈伯因组也有相同作用 （ 0.05- 0.01); 给药后 2个月， 对照药达纳康组大鼠可见 学习记忆能力明显提高（ 0.05); 与维脑康组比较， 本发明药物给药 1至 2个月后， 大、 中 剂量组大鼠学习记忆能力均明显提高， 游迷宫所需时间与模型组比较明显缩短 (P<0.05-0.01) c 结果见表 4。
表 4 本发明药物对 2V0大鼠游迷宫所需时间的影响 （
躕 时间 （S)
删 N
(mg/kg) 造模 1个月 造模 2个月 造模 3个月 假手术组 10 36· 8±23· 1 10.1±5.3 8.2±4.6
模型组 10 60.3±26· 1 55.2±26. l"" 45.1±23.2#" 哈伯因组 0.06 10 61.3±25.2 29.1±18.5* 10.8±5.6** 达纳康组 20 10 59.7±21.8 50.2±29.1 22.2±10.2* 维脑康组 15 10 60.9±27.5 34.3 ±16.9* 23.4±12.6 本发明药物组 11.5 10 61.2±23.5 26.6±10.2** 15.6±7.9** 本发明药物组 23 10 60.5±26.7 19.5±12.6**Δ 13.2±6.6**Δ 本发明药物组 46 10 60.8±24.0 17.1±11.4**Δ 7.6±5.5**ΔΔ 注： 与假手术组比较： m .oi; 与模型组比较： *κο.05, οι; 与 隹脑康组比较： Δ ο.θ5,
ΔΔ ο.01。
1.4讨论与小结
(1)本发明同时也做了东莨菪碱、 氯丙嗪、 利血平或亚硝酸钠所致化学损伤的药效学实 验。 在上述化学损伤基础上， 以 5 (10) miri内逃避潜伏期、 错误次数为指标进行小鼠跳台实 验。灌胃给予本发明药物 15天后， 二指标均获不同程度改善， 表明本发明药物可能有改善模 型小鼠获得性、 巩固性记忆障碍作用。
(2) 在乙醇化学损伤的基础上， 以 5min内逃避潜伏期、 错误次数为指标进行小鼠避暗 实验。 灌胃给予本发明药物 15天后， 二指标均获不同程度改善， 表明本发明药物有改善模型 小鼠再现性记忆障碍的作用。
(3)本研究以 D-半乳糖致脑老化大鼠、 A β毒性损伤大鼠、 自然衰老认知障碍大鼠和 ΑΡΡ 转基因小鼠为 AD模型， 以 2V0大鼠为 VD模型， 以 3 min内大 （小） 鼠在水迷宫中的游泳持 续时间、 游泳路径长度为主要指标， 以搜索策略等为辅助指标， 观察各药对小鼠游泳成绩的 影响。 结果表明， 灌胃给予大 （小） 鼠本发明药物后， 大 （小） 鼠的游泳成绩均获不同程度 改善，提示本发明药物可能有改善上述 AD模型动物和 VD动物模型空间学习记忆能力的作用。 同时本发明药物与维脑康比较， 可明显改善 2V0大鼠水迷宫实验各项指标， 表明本发明组成 中增加大豆成分后疗效更佳。
2 神经递质检测
实验方法按文献 "Liu JX, Cong WH, Xu L， Wang JN. Effect of combination of extracts of ginseng and ginkgo biloba on acetylcholine in amyloid beta- peptide— treated rats determined by an improved HPLC. Acta Pharmacol Sin. 2004; 25: 1118—23.，， 中公幵的 方法进行。
2.1乙酰胆碱 （ACh)
2.1.1 对 Αβ毒性损伤大鼠模型全脑 ACh含量的影响
与假手术组比较， 双侧海马 CA1 区注射 ΑβΜο 4周后， 模型组大鼠全脑 ACh含量显著降低 ( 0.01)。 给药 4 周后与模型组大鼠比较， 本发明药物各组大鼠全脑 ACh 含量显著升高 ( 0.05- 0.01); 阳性药哈伯因组大鼠全脑 ACh 7 平有升高趋势， 但无显著性差异。 结果见表 5。 表 5 对 A β损伤大鼠模型全脑 ACh含量的影响 （ 土 SD) 组别 剂量 (rag/kg) n ACh含量 （yg/l) 假手术组 6 242.8±39.7
模賴 6 155.7±15.5 哈伯 Kl组 0.08 6 180.0 ±23.5
本发明药物 11.5 6 211.2±39.3*
本发明药物 23 6 227.8±54.1*
本发明药物 46 6 235.4 ±25.4林
注： 与假手术组比较 ## 0.01； 与模型组比较 * 0.05， 0.01。
2.1.2对双侧颈总动脉永久性结扎所致 VD大鼠全脑 ACh的影响
模型组大鼠 2V0后 3个月脑内 ACh降低， 与假手术组比较有显著性差异（ 0.01); 给药 2个月后， 本发明各组大鼠脑内 ACh明显升高 （P<0.05- 0.01); 维脑康、 哈伯因及达纳康有 相同作用 （P<0.05- 0.01); 与维脑康组比较， 本发明大、 中剂量组大鼠脑内 ACh 明显提高 (P〈0.05- 0.01)。 结果见表 6。
表 6 本发明药物对 2V0大鼠脑内 ACh含量的影响 （ ±s) 删 n AC1^ ( Pg/l) 假手术组 10 284.4±51.2
模型组 10 146.4±13.2s"
哈伯因组 0.06 10 221.4±41.9**
达纳康组 20 10 182.4±40.3*
维脑康组 15 10 173.2±23.6**
本发明药物组 11.5 10 183.3±22.7**
本发明药物组 23 10 201.2±31.2**厶
本发明药物组 46 10 218.1±34.9**ΔΔ
注：与假手术组比较：" 01'、与模型组比较： *Ρ 0.05, 01 与 维脑康组比较： Ρ<0.05, ΔΔΡ<0.01。
2.1.3 讨论与小结
灌胃给予本发明药物后， 动物全脑或海马内 ACh含量均显著升高， 提示本发明药物有调 节上述 AD、 VD动物模型 ACh的释放和降解、升高中枢 ACh水平、进而改善其中枢胆碱能系统 功能的作用。 同时本发明药物与维脑康比较 2V0大鼠中枢 ACh水平明显升高， 表明本发明药 物组成中增加大豆成分后疗效更佳。
2.2 单胺类神经递质及其代谢产物
2.2.1 对 A β毒性损伤大鼠模型全脑单胺类神经递质及其代谢产物含量的影响
与假手术组相比， 模型组大鼠全脑 5-羟色胺 (5-ΗΤ) 含量明显降低 (Κ0.05), 多巴胺 (DA)、 去甲肾上腺素 （NE) 表现出下降趋势。 与模型组比较， 本发明各给药组大鼠全脑 DA、 5-HT 水平无显著变化 (P>0.05), 髙香草酸 (隱）、 5-羟吲哚乙酸 (5-HIAA) 水平呈下降趋 势， 其中本发明药物小剂量组 HVA显著下降 （ 0.05)。 结果见表 7。 表 7对 A β毒性损伤大鼠模型全脑单胺类递质及其代谢产物含量的影响 （ 土 SD) 单位： μ g/1 组别 n NE D0PAC DA 5-HIAA HVA 5-HT
105.7土 17.2士 160.4 + 39.7士 16.3士 59.3 + 假手术组 9
16.5 3.6 36.5 11.4 3.3 9.0
13.7士 139.4+ 35.6土 13.5 + 42.6土 模型组 6
2.0 14.2 13.2 4.2 7. m
75.9± 13.4土 136.2士 27.3士 15.4 + 41.7土 哈伯因组 0.08 6
11.3 1.3 8.5 3.1 9.4 5.9
12.6 + 137.3士 29.3土 9.4 + 39.9 + 本发明药物 11.5 6 85.6±5.8
o c 11— 2.0 12.9 6.2 1.5* 8.4
14.1土 135.7士 27.4士 13.1士 42.7士 本发明药物 23 6
2.4 15.6 3.6 4.6 8.8
13.8士 137.3± 26.1士 12.4土 37.5 + 本发明药物 46 6 84.7±8.4
1.9 14.4 2.0 2.5 10.8 注： 与假手术组比较 # 0.05； 与模型组比较 * 0.05。
2.2.2讨论与小结
(1) A β毒性损伤大鼠脑内单胺类递质代谢的低水平状态与衰老和 AD患者的病变相似， 表明 Α β毒性损伤大鼠的记忆障碍可能与脑内单胺类递质代谢的变化有关， 本发明药物可能 通过延缓 DA、 5- HT降解， 使中枢 DA、 5- HT水平相对升高， 继而改善脑内单胺能系统活性。
(2)本发明对自然衰老认知障碍模型也进行了同样的药效学实验， 结果测得模型大鼠海 马内 DA、 HVA, 5- HT和 5- HIAA均显著下降， NE、 二羟苯乙酸 （D0PAC) 呈下降趋势， 表明单 胺类递质代谢处于低水平状态， 这与 AD患者、 老年人中枢内 NE、 DA、 5-HT系统活性低下的 表现是一致的。 给予本发明药物 12周后， DA、 D0PAC、 5- HT和 5- HIAA均呈升高趋势， 其中 本发明药物高剂量组海马 DA、 5-HT水平明显升髙， 表明本发明药物可能有改善 DA、 5- HT系 统活性的作用， 这种作用可能是通过抑制 DA和 5- HT的摄取实现的。 结果还表明， 本发明药 物升高 DA、 5- HT水平的机理可能不同于哈伯因， 也有别于达纳康， 哈伯因的作用可能有抑制 神经元内 DA代谢、 抑制 5 - HT降解参与， 达纳康的作用可能有抑制神经元内 DA代谢、 抑制 5 - HT摄取参与。 .
(3) 本发明对 APP转基因小鼠也进行了同样的药效学实验， 与上述两种 AD模型脑内单 胺类递质变化不同， APP转基因小鼠全脑 5- HT、 5- HIAA显著升高， NE、 DA呈升高趋势， 表明 APP转基因动物由于 APP基因异常表达可能以不同于其他动物模型的方式影响中枢单胺类递 质的代谢， 这与文献报道一致。 阳性药哈伯因与本发明药物大剂量组 5- HT、 5- HIAA均呈不同 程度降低， 表明二者可通过调节中枢 5-HT的异常代谢而调节该系统活性。
结果表明， 本发明药物具有调节中枢 NE、 DA、 5- HT水平， 调节 NE、 DA和 5- HT系统活性 的作用。 鉴于胆碱能和单胺能系统之间的相互影响 （依赖和增强） 参与认知过程， 本发明药 物改善学习记忆的作用可能与干预单胺类递质的摄取和降解等有一定的关联。
3 其它生化指标检测
检测方法按各试剂盒说明书迸行。
3.1 乙酰胆碱酯酶 (AChE)
3.1.1对自然衰老认知障碍大鼠模型全脑 AChE活力的影响
与青年对照组比较， 老年对照组大鼠全脑 AChE活力显著降低 （ 0.05)。 给药 12周后， 与老年对照组大鼠比较， 阳性药哈伯因、达纳康和本发明各给药组大鼠全脑 AChE活力均显著 升高 (7X0.05-0.01)。 结果见表 8。 对自然衰老大鼠模型全脑 AChE活力的影响 （ ±SD) 组别 剂量 （mg/kg) n AChE活力 （U/mgProt)
青年对照组 7 1.31±0.26 老年对照组 15 0.88±0· 28# 哈伯因组 0.08 7 1.18±0, 19* 达纳康组 30 7 1.19±0.22* 本发明药物 11.5 7 1.49±0.47* 本发明药物 23 7 1.51±0.16** 本发明药物 46 7 1.31±0.17** 注： 与青年对照组比较 0.05; 与老年对照组比较 * 0.05, WO.01ο
3.2 丙二醛 (MDA)
3.2.1对自然衰老认知障碍大鼠模型全脑 MDA含量的影响
与青年对照组比较， 老年对照组大鼠全脑 MDA含量明显升高 （ 0.05)。 给药 12周后， 与老年对照组比较， 本发明药物各给药组大鼠全脑 MDA含量均显著降低（ 0.05); 阳性药哈 伯因、达纳康组大鼠全脑 MDA含量均有降低的趋势， 但无显著性差异（AO.05)。结果见表 9。 对自然衰老认知障碍大鼠模型全脑 MDA含量的影响 （ ± SD ) 组别 剂量 (mg/kg) n MDA含量 （nraol/mgProt) 靑年对照组 7 2.97±0.20 老年对照组 7 4.46±1.41# 哈伯冈组 0.08 7 3.50±0.43 达纳康组 30 7 4.35±0.66 本发明药物 11.5 7 3.16±0.61* 本发明药物 23 7 3.08±0.73* 本发明药物 46 7 3.17±0.25* 注: 与青年对照组比较 #P<0.05； 与老年对照组比较 *P<0.05o 3.3 超氧化物歧化酶 （SOD)
3.3.1对自然衰老认知障碍大鼠模型全脑 SOD活力的影响
与青年对照组比较， 老年对照组大鼠全脑 SOD活力显著降低 （ 0.05)。 给药 12周后， 与老年对照组大鼠比较， 阳性药哈伯因、 达纳康、 本发明药物各给药组大鼠全脑 SOD活力显 著升高 （ 0.05)。 结果见表 10。 对自然衰老认知障碍大鼠模型全脑 S0D活力的影响 （ X土 SD) 组别 n SOD活力 （NU/mgProt) 青年对照组 7 44.34±7.66 老年对照组 7 32.22±12.77tt
o b
哈伯因组 0.08 7 43.68±2.55*
达纳康组 30 7 40.18±4.74*
本发明药物 11.5 7 46.84±3.47* 本发明药物 23 7 47.94±6.07* 本发明药物 46 7 45.16±5.83* 注： 与青年对照组比较 H 0.05; 与老年对照组比较 * 0.05。
3.4 对 A β毒性损伤大鼠模型全脑 Na+- Κ+- ΑΤΡ酶活力的影响
与假手术组比较， 双侧海马 CA1区注射 1→。 4周后， 大鼠全脑 Na (+- ATP.酶活力显著 降低 （ 0.05)。 给药 4周后， 与模型组大鼠比较， 阳性药哈伯因、 本发明各给药组大鼠全脑 Na-K-ATP酶活力均有显著升高 ( 0.05-0.01)。 结果见表 11。 表 11 对 Αβ毒性损伤大鼠全脑 Na+- K+ ATP活力的影响 d土 SD)
Na'-Κ'-ΛΤΡ酶活力
组别 剂量 (mg/kg)
( μ moLPi/mgProt/h)
假手术组 7 0.205 + 0.037 模型组 7 0.161±0.013# 哈伯因组 0, 08 7 0.207±0.025* 本发明药物 11.5 • 7 0.218±0.018** 本发明药物 23 7 0.195±0.012** 本发明药物 46 7 0.193±0.032* 注： 与假手术组比较 # <0.05; 与模型组比较 *尸〈0.05, ** 0.0L
3.5 讨论与小结
3.5.1 对 AChE活力的影响
(1) 自然衰老认知障碍大鼠全脑 AChE活力显著降低， 与衰老、 AD变化一致。 本发明药 物可明显升髙降低的 AChE活力， 调节 ACh的异常代谢状态， 从而有可能改善衰老或 AD的中 枢胆碱能神经系统功能， 改善学习记忆障碍。
(2) 本发明对 D-半乳糖致脑老化大鼠和 APP转基因小鼠也进行了同样的药效学实验， 两种模型动物全脑 AChE活力呈显著升髙， 过高的活性将加速 ACh降解。本发明药物明显抑制 两种模型动物全脑 AChE活力， 从而有可能减缓 ACh降解代谢， 相对提高中枢 ACh水平， 改善 学习记忆。
3.5.2 对 MDA含量和 SOD活力的影响
给药后， 自然衰老认知障碍大鼠全脑 SOD活力提髙、 MDA水平降低， 表明本发明药物可 能具有提高老年动物机体抗氧化和清除自由基能力的作用， 可能有助于抗衰老、 提高记忆能 力。
3.5.3 对 A β毒性损伤大鼠模型全脑 Na+- Κ+- ΑΤΡ酶活力的影响
给药后， 毒性损伤大鼠全脑 Na+- K+- ATP酶活力显著提高， 表明本发明药物可能具有保 护 ATP酶活力， 调节细胞转运功能， 从而改善神经细胞质膜功能状态、 改善细胞功能的作用。
4. APP基因表达水平检测
4.1 对自然衰老认知障碍大鼠皮层和海马 APP基因表达水平的影响
与青年对照组比较， 老年对照组大鼠皮层和海马内 APP 基因的表达水平明显升高 ( 0.05)。 给药 12周后， 与老年对照组相比， 阳性药达纳康和本发明药物各剂量组 ΑΡΡ基 因的表达水平明显降低 （ Ο.Οδ) 结果见表 12。 表 12 对自然衰老认知障碍大鼠模型皮层和海马 ΑΡΡ基因表达水平的影响 （ ±SD) 组别 剂量 （mg/kg) n APP/ β- act in (%)
青年对照组 3 87.8±10.8 老年对照组 3 143.4±16.9tt 达纳康组 30 3 104.4±6.6* 本发明药物 11.5 3 101.4±20.9* 本发明药物 23 3 108.4±15, 0* 本发明药物 46 3 97.2 ±22.5* 注： 与靑年对照组比较 #/¾).05; 与老年对照组比较 * 0.05。
4.2讨论和小结
结果表明，本发明药物可明显纠正自然衰老认知障碍大鼠海马和皮层 ΑΡΡ基因的过表达， 提示其改善学习记忆障碍的作用可能是通过纠正皮层和海马内该基因的异常表达， 减少 Αβ 生成和异常沉积， 减少 SP形成， 从病变的上游干预 A3可能诱发的病理损害， 进而影响中枢 学习记忆功能的有关环节而发挥的。 5 组织细胞形态学实验
5. 1 HE染色
按龚志锦等主编 《病理组织制片和染色技术》 第 7章第 2节项下进行实验。
5. 1. 1 对 D-半乳糖致脑老化大鼠模型组织形态学的影响
空白组大鼠海马锥体细胞排列较为紧密， 细胞上下线明显， 细胞丰富； 核膜清楚， 核仁 明显。 模型组大鼠海马锥体细胞排列较为疏松， 细胞结构较模糊， 胞体肿胀， 细胞上下线不 规则； 部分细胞核固缩、 深染。 与模型组相比， 阳性药哈伯因组和本发明药物各组大鼠海马 锥体细胞排列较整齐， 细胞轮廓较清晰； 核固缩、 深染程度减轻。 其中哈伯因和本发明药物 高、 中剂量组效果较明显。
5. 1. 2 对自然衰老认知障碍大鼠模型组织形态学的影响
青年对照组大鼠海马锥体细胞排列密集， 细胞上下线整齐明显， 细胞丰满； 核膜清楚， 核仁清晰可见。 老年对照组大鼠海马锥体细胞排列较疏松， 部分细胞结构不完整； 少数细胞 核皱缩、 深染， 呈三角形； 与老年对照组相比， 达纳康和本发明药物各组大鼠海马锥体细胞 层次较清楚， 细胞形态有不同程度改善； 核固縮、 深染程度减轻。 其中达纳康和本发明药物 高、 中剂量组效果较明显。
5. 2 对海马神经细胞超微结构的影响
方法按汪谦主编的 《现代医学实验方法》 第 1-4页项下进行。
5. 2. 1 对 A β毒性损伤大鼠模型海马神经细胞超微结构的影响
假手术组大鼠海马神经细胞超微结构正常， 核大呈圆形或椭圆形， 常染色质均勾分布， 可 见明显的核仁， 核模完整； 胞浆内线粒体、 粗面内质网结构清晰； 核糖体、 突触均较丰富。 模 型组大鼠海马神经细胞核染色体边集、 固缩； 细胞体积缩小， 胞浆浓缩； 少数细胞可见线粒体 轻度肿胀； 粗面内质网、 核糖体等细胞器基本正常； 突触前、 后膜不清晰， 突触间隙消失， 突 触小泡较假手术组明显减少。 与模型组比较， 哈伯因和本发明各组海马神经细胞形态、 结构有 所改善， 胞膜和突触前后膜较清晰； 多数细胞核染色质分布均匀； 线粒体嵴较清楚。
5. 2. 2 对自然衰老认知障碍大鼠模型海马神经细胞超微结构的影响
青年对照组海马神经细胞保持正常的形态学特点， 核大呈圆形或椭圆形， 染色质均匀分 布， 可见明显的核仁； 胞浆内线粒体、 粗面内质网结构清晰； 核糖体、 突触丰富。 老年对照 组大鼠海马部分神经细胞明显碎裂， 膜溶解， 可见空泡及髓样结构等； 核形状不规则， 染色 质凝聚成块， 核周间隙明显增厚； 线粒体肿胀空泡化； 内质网扩张； 多见脂褐素、 油滴。 与 老年对照比较， 阳性药哈伯因、 达纳康和本发明药物各组部分细胞形态、 结构有所改善， 胞 膜较清晰； 部分细胞核染色质分布均匀； 线粒体板层状嵴清楚； 核糖体增多； 突触数目增加。 其中， 哈伯因、 达纳康和本发明药物高、 中剂量组效果较明显。
5. 2. 3 对 APP转基因小鼠模型海马神经细胞超微结构的影响
同窝生非转基因小鼠海马神经细胞超微结构正常， 核大呈圆形或椭圆形， 常染色质均匀 分布， 可见明显的核仁； 胞桨内线粒体、 粗面内质网结构清晰； 核糖体丰富； 突触较多。 空 白组小鼠细胞核多固缩、 边集， 排列不规则； 线粒体少， 嵴排列紊乱， 嵴间隙增大； 粗面内 质网排列紊乱， 甚至崩解； 溶酶体量增多， 形状不规则； 胞浆内可见空泡及髓样结构， 可见 形状不规则的脂褐素。 与空白组比较， 本发明药物高剂量组细胞形态、 结构有所改善， 部分 胞膜较清晰； 部分细胞核染色质分布均匀。
5. 3小结
海马神经细胞形态结构的完整是维持其正常功能的前提， 缺血性脑血管病、 衰老或老年 痴呆等病理因素导致神经细胞形态结构的损伤必然导致其功能的异常， 如胞核和粗面内质网 损害导致蛋白合成机能下降， 线粒体受损导致能量代谢障碍， 大量脂褐素堆积破坏细胞器的 空间有序性， 突触丢失意味着神经细胞丧失了靶组织， 最终导致神经细胞衰老、 死亡。 本研 究以 D-半乳糖致脑老化大鼠和自然衰老认知障碍大鼠为模型进行 HE染色，以 A β毒性损伤大 鼠、 自然衰老认知障碍大鼠、 APP转基因小鼠为模型进行透射电镜观察， 探讨本发明药物对 模型动物海马神经细胞的影响。结果表明， 不同模型动物给药后海马神经细胞及突触的形态、 结构均有不同程度改善， 突触数目增加， 神经细胞凋亡、 丢失有所延缓或减少， 提示本发明 药物可能有保护神经细胞的作用。
毒理学试验
大鼠禁食 16小时后， 以最大浓度 （57. 5mg/ml )、 最大体积 （20ml/kg体重） 的剂量， 一 日（24h)内灌胃给药 2次，一日内最大给药量为 2300mg/kg体重，相当于拟临床人用量（240mg/ 日） 的 670倍。 连续观察 14天， 无一例死亡。 按拟临床人用量的 70、 35及 17. 5倍的剂量给 大鼠连续服用 6个月， 分别给药后 3个月、 6个月及停药后 4周进行一般状况、 体重、 摄食 量、 血常规、 凝血、 血液生化、 心电图、 主要脏器指数及病理大体及镜下检查， 均未发现明 显的病理改变。 毒理学研究表明， 本发明药物安全、 低毒。
综上所述， 本发明组合物对缺血性脑血管病和老年痴呆的治疗作用优于目前常用药， 治 疗后有明显效果改善， 长期稳定性考察也表明， 本发明组合物质量稳定、 可靠。 具体实施方式
为了更清晰说明发明目的和技术方案， 借助下述实施例进一歩详述。 实施例 1 : 本发明的组合物 （提取物） 的处方
处方- 人参提取物 27. 5g 银杏叶提取物 27. 5g
西红花提取物 5. 5g 大豆提取物 2. 75g o
制备方法：
人参加 8倍生药量的 60%乙醇， 回流提取两次， 合并提取液， 浓缩至相对密度约 1. 05 (50 V) , 浓缩液加 2倍体积的蒸熘水， 滤过， 滤液上 AB- 8型大孔吸附树脂， 以蒸馏水洗脱， 继 续以 10%乙醇洗脱载药树脂， 弃去水和 10%乙醇洗脱液， 载药树脂继续以 70%乙醇洗脱， 大约 至 2. 5倍柱体积， 收集 70%乙醇洗脱液， 即得含有人参总皂苷的人参提取物。
取干燥银杏叶加 8倍量的 70%乙醇， 60°C温浸， 提取两次， 每次至少 1小时， 合并提取 液， 提取液减压浓缩至相对密度 1. 05左右（5CTC) , 浓缩液加水冷沉， 滤过， 滤液上 ADS - 17 型大孔吸附树脂， 载药树脂先以水洗脱， 继续 60%乙醇洗脱， 弃去水洗脱液，收集 60%乙醇洗 脱液， 浓缩至无醇味， 加入生药 2倍量水加热至沸， 室温沉降 24小时， 滤过， 滤液上 DM-130 型大孔吸附树脂， 先以水洗脱， 继续 15%乙醇和 60%乙醇洗脱， 弃去水和 15%乙醇洗脱液， 收 集 60%乙醇洗脱液， 继续浓缩干燥， 得银杏叶提取物。 其中银杏总黄酮醇苷和银杏总内酯的 含量比例为 24: 15。
西红花原料加 20倍的 60%乙醇， 70- 80°C温浸提取两次， 过滤合并提取液， 浓缩至无醇 味加生药 1倍以上体积的水稀释， 将稀释液加到 AB- 8型大孔吸附树脂柱上， 先以水洗脱， 再 以 20%乙醇洗脱， 继续以 30%乙醇洗脱， 最后用 70 %乙醇洗脱载药树脂， 弃去水、 30%乙醇洗 脱液， 收集 70%乙醇洗脱液， 即得西红花提取物， 其含有西红花总苷。
大豆以约 95%的乙醇提取， 滤过， 残渣以 70%的乙醇提取， 滤过， 将乙醇提取液合并浓缩 至无醇味， 加入原料 2倍重的水， 充分搅拌， 滤过， 滤液上 AB- 8型大孔吸附树脂， 先以水洗 脱载药树脂， 弃去水洗脱液， 再以 60 %的乙醇洗脱， 收集洗脱液得到组分 A， A中成分主要 是异黄酮； 再用 90- 95%乙醇洗脱， 收集洗脱液， 回收至干， 加入无水乙醇进行酯化， 再加水 洗涤， 溶液分层， 除去下层水液（酸水） 后用旋转浓缩仪减压至 O. lMPa脱气， 加入氢氧化钠 进行醇解， 再加水洗涤， 弃去下层水洗液 （碱水）， 上层有机液减压至 0. IMPa脱气后进行薄 膜蒸馏， 残留液进行分子蒸馏 （压力为 0. 133Pa， 蒸发板到冷凝板距离小于 0. 5mm)， 得到组 分 B， B中成分为混合生育酚， 之后将 B与 A混合， 即得到本发明的大豆提取物， 主要含大豆 总异黄酮和维生素 E， 其重量含量配比为 4: 1。
将上述干燥后的提取物混合粉碎至 20 目， 加淀粉 86. 75g， 装入 3号胶囊， 即得本发明 胶囊剂。
结果显示： 本发明组合物胶囊剂符合胶囊剂项下的有关各项规定。
定性和定量： 银杏内酯的鉴别及含量测定参照中国药典 2005 年版一部银杏叶提取物项 下； 西红花苷 -I的鉴别及含量测定参照中国药典 2005年版一部西红花项下； 人参鉴别及含 量测定参照中国药典 2005年版一部人参项下。
大豆总异黄酮和金雀异黄素的含量测定方法： 精密称取大豆提取物适量用 25ml水混悬， 加入 1ml醋酸盐缓冲溶液 （PH4，5) 和 15μ1β-葡萄糖苷酶， 37°C水浴水解， 减压回收溶剂至 千， 再用甲醇溶解， 微孔滤膜滤过， 用液相色谱分析（Zorbax- (^色谱柱， 流动相为甲醇-水- 乙酸 45: 55： 1， 流速 0.8ml/min， 监测波长 260nm);
VE的含量测定方法： 取本品适量， 精密称定， 放在乳钵中加两滴无水乙醇研磨， 用 20ml 无水乙醇分次定量转移至粉液漏斗内， 加水 10ml,以正己烷萃取 3次， 每次 5ml， 合并正己烷 萃取液，减压回收溶剂至干，残渣以流动相定量转移至 10ml容量瓶中，加流动相定容至刻度， 摇匀， 过 0.45 μπι的微孔滤膜， 作为供试品溶液， 分别精密吸取对照品溶液与供试品溶液各 10 l， 注入液相色谱仪中测定 （97: 3ν/ν的甲醇-水为流动相， 检测波长为 207nm)。
本发明组合物的胶囊剂中每粒胶囊含有各成分如下 （以每粒胶囊 1.5g计）：
(1) 含总皂苷以人参皂苷 Re (C4SHS201S) 计， 不少于 13.75mg;
(2) 含总皂苷以人参皂苷 Rgl (C«H72014)、 Re (C,8HS20IS) , 人参皂苷 Rb, (CMH92023 ) 计分别不低于 1.375mg、 0.825mg和 0.825mg;
(3) 含总黄酮以芦丁计 （C27H3„0,s)， 不少于 11.00mg。
(4) 含总黄酮醇苷分别以槲皮素、 山奈素及异鼠李素计不少于 6.60rng。
(5) 含萜类内酯以银杏内酯 A (C2() 0e)、 银杏内酯 B (C2U ,A。）、 银杏内酯 C (C2„HM0„) 和白果内酯 (CI5Hls08) 的含量之和计， 不少于 2.75mg;
(6) 含西红花总苷以无水西红花苷- 1 ( ,¾„0Μ) 计， 不少于 2.75mg。
(7) 含西红花苷- 1 (CJW)M) 不低于 1.375 mg;
(8) 含大豆总异黄酮以金雀异黄素计 (C2TH3()0lfi), 不少于 1.35mg;
(9) 含金雀异黄素 （C27 。0,B)， 不少于 0.5mg;
(10) 含维生素 E以维生素 E (C31H5203) 计， 不少于 0.5mg。
本发明组合物胶囊剂临床推荐量 150mg/次， 3次 /日。
实施例 2: 本发明的组合物的处方
人参 40份 银杏叶 45份 西红花 1份 大豆 75份。 组合物制备方法： 将上述药材粉碎过 20目， 即得本发明组合物。
颗粒剂的制备： 上述药粉混合后加入糊精、甜叶菊苷， 充分混匀， 置 70〜75Ό真空干燥, 粉碎， 制颗， 即得本发明组合物的颗粒剂， 其呈棕褐色颗粒。
结果显示：本发明组合物颗粒剂符合颗粒剂项下的有关各项规定（中国药典 2005年版一 部附录 I C)。
含量测定参照实施例 1， 以人参皂苷 Re计， 不少于 9. 15mg/g。
实施例 3:
处方： 人参提取物 2份、 银杏叶提取物 10份、 西红花提取物 0. 5份、 大豆提取物 1份。 片剂的制备方法： 上述本发明组合物的提取物混合后， 加入糊精等粘合剂， 充分混匀， 置 60- 80°C真空干燥， 粉碎， 加入淀粉等填充剂、 硬脂酸镁等润滑剂以及羧甲基纤维素钠等 崩解剂混匀， 制粒， 压片， 即得本发明片剂。
结果显示： 本发明组合物片剂符合片剂项下的有关各项规定。
定性定量检测同实施例 1 ; 本品以人参皂苷 Re计， 不少于 9. 15mg/g。
实施例 4:
处方： 人参提取物 10份、 锒杏叶提取物 3份、 西红花提取物 4份、 大豆提取物 0. 2份。 组合物的制备方法： 同实施例 1。
片剂的制备：上述本发明组合物的提取物混合后，加入糊精等粘合剂，充分混匀，置 60 - 80 °C真空千燥， 粉碎， 加入淀粉等填充剂、 硬脂酸镁等润滑剂以及羧甲基纤维素钠等崩解剂混 匀， 制粒， 压片， 即得本发明片剂。
结果显示： 本发明组合物片剂符合片剂项下的有关各项规定。
定性定量检测同实施例 1， 以人参皂苷 Re计， 不少于 9. 15mg/g。
实施例 5:
处方： 人参 10份、 银杏叶 1份、 西红花 0. 1份、 大豆 8份。
制备方法： 将上述药材粉碎过 20目， 即得本发明组合物。
含量测定参照实施例 1 , 以人参皂苷 Re计， 不少于 9. 15mg/g。

Claims

权 利 要 求
1、 一种中药组合物， 其由以下重量配比的组分组成： 人参 1-10份、 银杏叶 1-10份、 西红花 0. 05- 0. 5份、 大豆 5- 10份， 所述人参、 银杏叶、 西红花和大豆的来源可以是中药材 或相当于上述中药材生药量的中药提取物。
2、 权利要求 1所述的中药组合物， 其中所述的组分组成为： 人参 2- 6份、 银杏叶 3- 6 份、 西红花 0. 06- 0. 2份、 大豆 7 8份。
3、 一种中药组合物， 其由以下重量配比的组分组成： 人参提取物 1-10份、 银杏叶提取 物 1-10份、 西红花提取物 0. 5- 5份、 大豆提取物 0. 1- 1份。
4、 权利要求 3所述的中药组合物， 其中， 所述银杏叶提取物中至少包括银杏总黄酮醇 苷和银杏总内酯， 其在银杏叶提取物中的重量含量配比为 24 : 25-10。
5、权利要求 1或 3所述的中药组合物， 其中， 所述的银杏叶提取物可用下述方法得到： 取银杏叶加至少 2倍量的 60- 80 %的乙醇， 50- 70°C浸提至少一次， 合并提取液， 提取液浓缩 至相对密度 1. 05左右， 浓缩液加水， 滤过， 滤液上极性氢键聚苯乙烯型大孔吸附树脂， 树脂 先以水洗脱， 继续以 60%乙醇洗脱， 弃去水洗脱液， 收集 60%乙醇洗脱液， 浓缩至无醇味， 加 水， 滤过， 滤液上弱极性聚苯乙烯型大孔吸附树脂， 先以水洗脱， 继续 15%乙醇和 60%乙醇洗 脱， 弃去水和 15%乙醇洗脱液， 收集 60%乙醇洗脱液。
6、 权利要求 1或 3所述的中药组合物， 其中， 所述的大豆提取物中至少含有总大豆异 黄酮和维生素 E, 其重量配比为 4: 2-0. 5。
7、 权利要求 6所述的中药组合物， 其中， 所述的大豆提取物可用下述方法得到： 原料 大豆以 85- 95%的乙醇提取， 滤过， 残渣以 60- 80%的乙醇提取， 滤过， 将乙醇提取液合并浓缩 至无醇味， 加入原料重量 1倍以上的水， 滤过， 滤液上大孔吸附树脂， 以 50- 65%的乙醇洗 脱， 收集洗脱液得到组分 A; 大孔吸附树脂再用 90- 95%乙醇洗脱， 收集洗脱液， 回收至干， 加入无水乙醇进行酯化， 再加水洗涤， 分层， 除去下层水液后减压至 0. IMPa脱气， 加入氢氧 化钠进行醇解， 再加水洗涤， 弃去下层水洗液， 上层有机液减压至 0. IMPa脱气后进行薄膜蒸 馏， 残留液进行分子蒸馏得到组分 B, 将 B与 A混合。
8、权利要求 1或 3所述的中药组合物的制备方法， 包括将人参提取物、 银杏叶提取物、 西红花提取物和大豆提取物混合；
其中， 所述的人参提取物可用下述方法得到- 人参加至少 2 倍量的 50-70 %的乙醇， 回流提取至少一次， 提取液浓缩至相对密度约 1. 05， 浓缩液加至少 1倍体积的水， 滤过， 滤液上弱极性聚苯乙烯型大孔吸附树脂， 分别以 水和 10%乙醇洗脱树脂，弃去水和 10%乙醇洗脱液，树脂继续以 60- 75 %乙醇洗脱，收集 60-75 %乙醇洗脱液， 即得人参提取物；
所述的银杏叶提取物可采用下述提取方法得到：
取干燥银杏叶加至少 2倍量的 60-80%乙醇， 50- 70°C提取至少一次， 提取液减压浓缩 至相对密度 1. 05左右加水冷沉， 滤过， 滤液上极性氢键聚苯乙烯型大孔吸附树脂， 树脂先以 水洗脱， 继续 60%乙醇冼脱， 弃去水洗脱液， 收集 60%乙醇洗脱液， 浓缩至无醇味， 加入水加 热至沸， 室温沉降， 滤过， 滤液上弱极性聚苯乙烯型大孔吸附树脂， 先以水洗脱， 继续 15% 乙醇和 60%乙醇洗脱， 弃去水和 15%乙醇洗脱液， 收集 60%乙醇洗脱液；
所述的西红花提取物可釆用下述提取方法得到：
西红花加至少 5倍的 60%- 80 %乙醇， 70- 80°C提取至少一次， 提取液浓缩至无醇味， 加 生药 1倍以上体积的水稀释， 将稀释液加到弱极性聚苯乙烯型大孔吸附树脂柱上， 先以水洗 脱， 再以 30 %以下乙醇由低至高依次洗脱， 最后用 70 %乙醇洗脱载药树脂， 弃去水、 30%以 下乙醇洗脱液， 收集 70%乙醇洗脱液， 即得西红花提取物 ,·
所述的大豆醇提物可釆用下述提取方法得到：
原料大豆以 85 95%的乙醇提取， 滤过， 残渣以 60- 80%的乙醇提取， 滤过， 将乙醇提取 液合并浓缩至无醇味， 加入原料重量 1倍以上的水， 滤过， 滤液上大孔吸附树脂， 先以水洗 脱树脂， 弃去水洗脱液， 再以 50- 65 %的乙醇洗脱， 收集洗脱液得到组分 A， 树脂再用 90- 95% 乙醇洗脱， 收集洗脱液， 回收至千， 加入无水乙醇进行酯化， 再加水洗涤， 溶液分层， 除去 下层水液， 减压至 0. IMPa脱气， 加入氢氧化钠进行醇解， 再加水洗涤， 弃去下层水洗液， 上 层有机液减压至 O. lMPa脱气后进行薄膜蒸馏， 残留液进行分子蒸镏， 得到组分 B， 将 B与 A 混合， 即得到大豆提取物。
9、 一种治疗缺血性脑血管病和老年痴呆的药物， 其中包括权利要求 1-6任一项所述的 中药组合物和药学可接受的辅料。
10、权利要求 9所述的药物， 其中所述药物为口服剂型， 其为蜜丸、浓缩水蜜丸、水丸、 颗粒剂、 胶囊剂、 片剂、 散剂、 膏剂、 口服液或糖浆剂。
11、 权利要求 1所述的组合物在制备治疗缺血性脑血管病和老年痴呆的药物中的应用。
12、 权利要求 3所述的组合物在制备治疗缺血性脑血管病和老年痴呆的药物中的应用。
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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2009129833A1 (de) * 2008-04-22 2009-10-29 Merz Pharma Gmbh & Co. Kgaa Kompartimentspezifische pflanzenextraktkombination aus ginkgo biloba- und ginseng-extrakt mit tandemwirkung
WO2015159922A1 (ja) * 2014-04-16 2015-10-22 花王株式会社 ニンジン抽出物の製造方法
RU2674264C1 (ru) * 2014-12-30 2018-12-06 Шайнвэй Фармасьютикал Груп Лтд. Китайская лекарственная композиция для предотвращения или лечения сердечно-сосудистых и цереброваскулярных заболеваний или деменции и способ ее получения и применение

Families Citing this family (18)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090098228A1 (en) * 2007-10-10 2009-04-16 Tsumura & Co. Agent and method for improvement of impairment of learning and memory
CN102614460B (zh) * 2012-03-31 2013-10-30 陈欣 治疗心肌梗塞的中药组合物及其制备方法
JP2012214500A (ja) * 2012-08-03 2012-11-08 Jianxun Liu 漢方薬組成物ならびにこれらの調製方法および使用
SG11201501020WA (en) * 2012-08-15 2015-04-29 Chi Yu Fen Pharmaceutical composition for increasing content and availability of cyclic adenosine monophosphate in a body and the preparation thereof
CN103285104B (zh) * 2013-05-14 2016-01-06 张景红 一种具有修复心肌损伤作用的中药注射液
CN103536698B (zh) * 2013-11-04 2015-08-12 华润三九医药股份有限公司 一种中药组合物在制备治疗血管性痴呆的药物中的应用
CN104068703B (zh) * 2014-01-16 2016-01-13 赵正旭 一种玉石保健枕
CN104352671A (zh) * 2014-10-24 2015-02-18 神威药业集团有限公司 一种中药组合物的新的医药用途
EP4356967A3 (en) * 2014-11-14 2024-07-24 United Arab Emirates University Compounds for use as imaging agents
US10543243B2 (en) 2016-06-06 2020-01-28 Charsire Biotechnology Corp. Soybeam seed extract, method for producing the same and uses thereof
CN108079030B (zh) * 2018-01-26 2021-02-26 贵阳中医学院 苗药铁筷子提取物在制备抗ad产品中的应用
CN110433199B (zh) * 2019-05-22 2021-09-28 浙江中医药大学 一种治疗脑缺血的益气活血解毒组分中药组合及其制备方法
KR20210096704A (ko) 2020-01-28 2021-08-06 주식회사 다나을 간기능 개선을 위한 전통 한방 약제 조성물을 함유하는 약제학적 제형 및 이의 제조방법
KR20210096705A (ko) 2020-01-28 2021-08-06 주식회사 다나을 불안장애 및 불면증 치료를 위한 전통 한방 약제 조성물을 함유하는 약제학적 제형 및 이의 제조방법
CN111419958A (zh) * 2020-04-23 2020-07-17 唐荣成 一种治疗脑血管意外后遗症证的中药组合物及其制备方法
CN112931854B (zh) * 2021-02-04 2023-09-12 江西建昌帮食疗科技有限公司 一种生发、助消化组合物
CN113588808B (zh) * 2021-06-25 2023-02-28 西安千禾药业股份有限公司 一种同时检测血平胶囊中多种有效成分的含量测定方法
CN113634011B (zh) * 2021-08-24 2022-03-08 伽能生物科技(上海)有限公司 松红梅复合精萃组合物的生产集成系统、松红梅复合精萃组合物及其制备方法

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1413656A (zh) * 2002-10-14 2003-04-30 刘建勋 治疗缺血性脑血管疾病和老年性痴呆的药物维脑康及其制造方法

Family Cites Families (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5733926A (en) * 1996-12-13 1998-03-31 Gorbach; Sherwood L. Isoflavonoids for treatment and prevention of alzheimer dementia and reduced cognitive functions
JP2002199858A (ja) * 2001-01-08 2002-07-16 Fusao Takimoto 乾燥オカラ粉を主成分にした健康食品
EP1414469A2 (de) 2001-08-09 2004-05-06 Degussa Food Ingredient GmbH Formulierung enthaltend (lyso-) phosphatidylserin zur vorbeugung und behandlung von stresszuständen bei warmblütern
JP2003261456A (ja) * 2002-03-08 2003-09-16 Howaizu:Kk 脳の老化予防剤
KR20040062274A (ko) * 2003-01-02 2004-07-07 서울향료(주) 용안육, 산조인, 원지, 백복신 및 은행잎의 추출물을함유한 기억력 증진 및 치매 예방 및 치료용 조성물
JP2005281278A (ja) * 2004-03-31 2005-10-13 Daicho Kikaku:Kk 栄養剤、消化器剤、抗うつ剤、抗更年期障害剤、抗老人性痴呆症剤、抗アルツハイマー剤、強筋肉剤、抗炎症剤

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1413656A (zh) * 2002-10-14 2003-04-30 刘建勋 治疗缺血性脑血管疾病和老年性痴呆的药物维脑康及其制造方法

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
"Chinese Pharmacopoeia", 2005
LIU JX ET AL.: "Effect of combination of extracts of radix ginseng and ginkgo biloba on acetylcholine in amyloid beta-peptide-treated rats determined by an improved HPLC", ACTA PHARMACOL SIN., vol. 25, 2004, pages 1118 - 23
MORRIS RGM; GARRUD P; RAWLINS JNP ET AL.: "Place navigation impaired in rats with hippocampus lesions", NATURE, vol. 297, pages 681 - 3
See also references of EP2033653A4
SU L.: "Soybean, perfect nutritional and health food", FOOD AND NUTRITION IN CHINA, no. 9, 2004, pages 42 *

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2009129833A1 (de) * 2008-04-22 2009-10-29 Merz Pharma Gmbh & Co. Kgaa Kompartimentspezifische pflanzenextraktkombination aus ginkgo biloba- und ginseng-extrakt mit tandemwirkung
US9592261B2 (en) 2008-04-22 2017-03-14 Dr. Willmar Schwabe Gmbh & Co. Kg Compartment-specific plant extract combination of ginkgo biloba extract and ginseng extract having a tandem effect
WO2015159922A1 (ja) * 2014-04-16 2015-10-22 花王株式会社 ニンジン抽出物の製造方法
JP2015212260A (ja) * 2014-04-16 2015-11-26 花王株式会社 ニンジン抽出物の製造方法
RU2674264C1 (ru) * 2014-12-30 2018-12-06 Шайнвэй Фармасьютикал Груп Лтд. Китайская лекарственная композиция для предотвращения или лечения сердечно-сосудистых и цереброваскулярных заболеваний или деменции и способ ее получения и применение
US10967023B2 (en) 2014-12-30 2021-04-06 Shineway Pharmaceutical Group Ltd. Preparation of a chinese medicinal composition including ginseng, ginkgo leaf and stigma croci

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