CN102626383A - Bilobalide injection and preparation method thereof - Google Patents
Bilobalide injection and preparation method thereof Download PDFInfo
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- CN102626383A CN102626383A CN2012101207876A CN201210120787A CN102626383A CN 102626383 A CN102626383 A CN 102626383A CN 2012101207876 A CN2012101207876 A CN 2012101207876A CN 201210120787 A CN201210120787 A CN 201210120787A CN 102626383 A CN102626383 A CN 102626383A
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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
- A61K9/0012—Galenical forms characterised by the site of application
- A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
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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
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- 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
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P37/00—Drugs for immunological or allergic disorders
- A61P37/08—Antiallergic agents
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- 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
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- 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
Abstract
The invention relates to a preparation method for a preparation of Chinese medicine, particularly relates to a bilobalide injection and a preparation method thereof and belongs to the field of medicine. The technical problem is solved by the invention by providing the bilobalide injection. The bilobalide injection has the advantages of clear components and fixed proportion; the bilobalide injection with high purity, clear component and fixed proportion is taken as an active ingredient; the content of the effective component is above 95%; and the bilobalide injection contains the following components.
Description
Technical field
The present invention relates to a kind of method for preparing of Chinese medicine preparation, particularly bilobalide injection and method for preparing, belong to drug world.
Background technology
From the sixties in last century, many countries adopt modern separation technology that the chemical constituent of Folium Ginkgo is studied, and through pharmacological evaluation and clinical verification, find that many-sided biological activity of Folium Ginkgo is relevant with its contained particular chemical composition.Germany Dr.Willar Schwabe has registered a kind of simple extract of Folium Ginkgo first; Applied for patent (W Schwabe DE 176708 and DE 2117429) in 1972; Name and be EGb761, be used to treat cardiovascular and cerebrovascular disease and nervous system disease, have significant curative effect; And have no side effect, ginkgolide compound (ginkgolids) has strong platelet activating factor (PAF) antagonism.There are Germany, France and Chinese in the country that ginkgo agent is classified as medicine, and other countries all use and are that health food or over-the-counter medications, the gingkgo health care food that the U.S. develops have obtained the FDA approval.
Bilobalide belongs to terpenoid, is called terpene lactone, is made up of sesquiterpene lactones and diterpenoid-lactone, is the important active component of Semen Ginkgo Ye Zhongyi class.Bilobalide (bilobalide) belongs to sesquiterpene lactones, is separated obtaining a present unique sesquiterpene lactones chemical compound of from Folium Ginkgo, finding in 1969 with K.Weinges in 1967 by R.T.Major.Ginkalide A, B, C, M, J (ginkgolidA, B, C, M, J) are diterpenic lactone; From Folium Ginkgo, separated first by S.Furukawa in 1932, just further separated and confirmed its chemical constitution in 1967 by people such as K.Nakanish, M.Maruyama and K.Okabe.See that from structure bilobalide quasi-molecule skeleton is made up of 15 carbon atoms, have 4 five-membered rings that condense mutually together, 1 five yuan of carbocyclic ring is wherein arranged, 3 five yuan of lactone carbocyclic rings are connected with the rare tert-butyl group in 1 natural product on the five-membered ring.Bilobalide has very strong biological activity; Has the effect that promotes nerve growth; Can prevent the changing function that brain cell mitochondrion oxidative stress causes, improve senile memory, prevent the generation of senile dementia; And preventing brain, spinal nerves demyelination, its neurotrophy, neuroprotective are stronger than bilobalide.Ginkalide B has effects such as antiinflammatory, shock, protection cardiovascular and cerebrovascular vessel, treatment acute pancreatitis.And the molecular skeleton of ginkgolide compound is made up of 20 carbon atoms; Have 6 five-membered rings, 2 five yuan of carbocyclic rings are wherein arranged, 3 lactone ring five membereds; 1 oxolane ring; Two five yuan of carbocyclic rings link together with the form of volution, and remaining ring connects with condensed mode, forms the special stereochemical structure of a rigidity verdant shape.All has the tert-butyl group rare in the natural product in the bilobalide molecule.Bilobalide comprises Diterpenes and sesquiterpenoids lactone, and diterpenes diterpenoids lactones mainly contains ginkalide A, B, C, J, M etc., and hemiterpene class lactone has bilobalide.
After early 1970s has been found PAF; Pharmacologist is studied bilobalide; Clear and definite Folium Ginkgo terpene lactones is strong platelet activating factor antagonist, and immune system, central nervous system, ischemic injuries have protective effect, and shock, antiallergic and antiinflammatory action are arranged.The position that the hydroxy number that ginkalide A, B, C, M, J structural difference are to contain is connected with hydroxyl is different.Bilobalide is strong platelet activating factor antagonist, is the active key component of special physiological in the Folium Ginkgo.
Ginkalide A structural formula ginkalide B structural formula
Molecular formula: C
20H
24O
9Molecular weight: 408.4 molecular formula: C
20H
24O
10Molecular weight: 424.4
Ginkalide C structural formula bilobalide structural formula
Molecular formula: C
20H
24O
11Molecular weight: 440.4 molecular formula: C
15H
18O
8Molecular weight: 326.3
Bilobalide has powerful specific inhibitory effect to the platelet activating factor paf receptor, and wherein the anti-PAF activity of bilobalide is the highest.PAF is a kind of endogenous phospholipid that platelet and multiple inflammation tissue secretion produce, and is the most effectively platelet aggregation derivant of finding so far, it with the generation of numerous disease, develop closely related.And bilobalide is considered to have most the natural paf receptor antagonists of potential applicability in clinical practice at present, and its antagonism is active closely related with chemical constitution.When R3 in the lactone structure is hydroxyl or hydroxy number when increasing, the antagonistic activity of PAF is weakened, and when R2 be hydroxyl and R3 when being H, then actively significantly strengthen, wherein the strongest to the antagonism that PAF produces with ginkalide B.
The extraction and the purification process of bilobalide are more, mainly contain: solvent extraction, post extraction method, solvent extraction-post extraction method, super critical extraction and chromatograph or column chromatography purification method etc.These methods all can not be isolated high-load bilobalide compositions effectively, and each proportion of composing of bilobalide is uncertain, therefore; Drug effect has nothing in common with each other in clinical use, because its content is not high, also has certain security risk; No complete pharmacological toxicology and clinical testing data, therefore, said method is all under test; Be not implemented on drug production process, though in China the bilobalide injection related patent U.S. Patent No. is arranged, its composition is all different with the present invention; Through the retrieval of how tame official website of ICH member state, still there is not other bilobalide injector launch so far.At present HPLC-UV method, HPLC-MS and the HPLC-ELSD methods of adopting of the mensuration of bilobalide constituents more; These methods only can be measured the content of each composition of bilobalide; Owing to lack strictness control to harmful substance in its product; The mass property that can not reflect medicine does not really form a perfect drug quality hierarchy of control.
The meglumine of employing or sodium hydroxide are arranged as cosolvent in other patent of invention,, inevitably bring huge potential safety hazard to clinical application because bilobalide at unique five-membered ring structure, can cause the lactone open loop and lose curative effect of medication under alkali condition.The present invention desires to provide a kind of bilobalide injection with good production technology and strict quality method.
Summary of the invention
Because bilobalide is a liposoluble substance; Almost insoluble in water, adopt high pharmaceutic adjuvant ethanol of safety and glycerol as cosolvent of the present invention, strict control cosolvent ratio is between 1: 1～5: 1; The structure of bilobalide does not change; Solved the solubility problem of medicine effectively, as the pH regulator agent, can make medicine more stable with citric acid or hydrochloric acid; Avoid medicine put procedure crystallize to cause clarity and particulate matter defective, solved the potential safety hazard of clinical use.
The technical problem that the present invention solved provides a kind of bilobalide injection, has the advantage of the clear and fixed ratio of component, and has removed macromole, protein effectively, has avoided the generation of untoward reaction.The active component of injection of the present invention is to adopt high-purity, the bilobalide of the clear and fixed ratio of component, and effective site content is more than 95%, and wherein 4 kinds of components contain bilobalide (C
15H
18O
8) be 25.0%～50.0%, ginkalide A (C
20H
24O
9) be 20.0%～45.0%, ginkalide B (C
20H
24O
10) be 10.0%～30.0%, ginkalide C (C
20H
24O
11) be 5.0%～15.0%.
Bilobalide injection of the present invention contains following compositions:
In above-mentioned prescription, there are two kinds of situations in essence:
1, when water for injection gets 0, adopt glycerol and ethanol as cosolvent.
Strict control cosolvent volume ratio is between 1: 1～5: 1, and promptly the volume ratio of ethanol, glycerol is 1～5 part of an ethanol, 1 part of glycerol.
Optimizing prescriptions is following:
Be that bilobalide injection of the present invention contains following compositions:
Bilobalide (in terpene lactone) 5mg/ml
Glycerol 0.4ml/ml
Ethanol 0.6ml/ml.
2, when water for injection is non-vanishing, adopt glycerol and ethanol as cosolvent.
Strict control cosolvent volume ratio is between 1: 1～5: 1, and promptly the volume ratio of ethanol, glycerol is 1～5 part of an ethanol, 1 part of glycerol.
Optimizing prescriptions is following:
Be that bilobalide injection of the present invention contains following compositions for every milliliter:
Ethanol and glycerol and water for injection have good intersolubility, mix back solution and have good stable property, can keep the original chemical constitution of medicine as solvent, help absorption of human body, the performance curative effect of medication.Bilobalide has fine solubility in ethanol; But it is fully bigger as solvent for injection to the zest of human body with ethanol; Therefore; With glycerol and a small amount of water for injection alcohol,diluted, can effectively alleviate pain and human body in the intravenous injection process to alcoholic acid toleration, can not change solution to the dissolubility of medicine and the stability of solution.
The method for preparing of bilobalide injection of the present invention is following:
A) preparation: mixed ethanol and glycerol (cosolvent proportion requirement 1: 1～5: 1), add bilobalide, stirring and dissolving is added ethanol or water for injection to full dose, regulates pH value to 2.5～6.5 with 5～10% citric acid solns or 1～10% hydrochloric acid solution; Preferred pH value is 3.2～3.8.
B) filtration sterilization;
C) embedding;
D) sterilization.
The method of quality control of injection of the present invention comprises:
Character: colourless clear liquid.
PH value: be 3.0～4.5.Preferred pH value is 3.2～3.8.
Solution colour: less than yellow No. 4 standard color solutions (Chinese Pharmacopoeia).
Tannin, resin, oxalates, potassium ion: must not detect.
Undue toxicity: process the solution that contains 0.2mg among every 1ml, meet the intravenous injection administration.
Pyrogen: dosage is by the every 1kg injection of rabbit body weight 5ml qualified (Chinese Pharmacopoeia).
Haemolysis and cohesion: get injection 6ml of the present invention and join among the 0.9% sodium chloride injection 100ml and dilute passed examination (Chinese Pharmacopoeia).
Hypersensitive test: get injection of the present invention, add 0.9% sodium chloride injection and be diluted to, in the intravenous injection need testing solution 30 minutes, anaphylaxis must not occur in terpene lactone 0.4mg/ml.
Irritation test: get injection of the present invention and use 0.9% sodium chloride injection to be diluted to concentration to be 0.5mg/ml solution; Get 3 of rabbit; Male and female are regardless of, weigh, every day the rabbit right edge intravenous drip need testing solution 10ml/kg that picks up the ears; The equal-volume of left side auricular vein instillation simultaneously 0.9% sodium chloride injection instils 3 times altogether.Before each administration with administration after after 1,6 hour and the last administration 48 hours, perusal injection site blood vessel has or not IR symptoms such as hyperemia, hemorrhage, necrosis.Injected back 48 hours carotid artery sacrificed by exsanguination rabbit, inserting needle place proximal part 1 centimeters that instils certainly, 2 centimetres at clip ear edge in last; FAA is liquid-solid fixed, conventional dehydration, specimens paraffin embedding slices; HE dyeing is carried out pathological examination to vascular tissue under optical microscope, judge whether to have zest.The vascular stimulation reaction must not appear in the intravenous drip test sample.
Total ginkgolic acids: chromatographic column C18, mobile phase: methanol-1% glacial acetic acid (90: 10), the detection wavelength is 310nm.Calculate total ginkgolic acids content with Semen Ginkgo eo-acid reference substance external standard method and be no more than 1ppm.
Finger printing: chromatographic column C18, mobile phase: methanol-oxolane-water (25: 10: 65), evaporative light scattering detector; Drift tube temperature: 105 ℃; Flow rate of carrier gas: 3.00L/min, column temperature: 40 ℃ is object of reference with bilobalide reference substance, ginkalide A reference substance, ginkalide B reference substance, ginkalide C reference substance; Press chromatographic fingerprints of Chinese materia medica similarity evaluation system, four total peaks of test sample finger printing and reference fingerprint similarity are greater than 0.95.
Aseptic: as according to membrane-filter procedure,, and to wash filter membrane 3 times, aseptic qualified (Chinese Pharmacopoeia) with 0.9% aseptic sodium chloride solution with 0.9% aseptic sodium chloride solution dilute filtration.
Content: chromatographic column C18, mobile phase: methanol-oxolane-water (25: 10: 65), use evaporative light scattering detector, drift tube temperature: 105 ℃; Flow rate of carrier gas: 3.00L/min; Column temperature: 40 ℃.With measuring behind the Extrelut-20 post adsorption treatment sample, every 1ml contains terpene lactone with bilobalide (C
15H
18O
8), ginkalide A (C
20H
24O
9), ginkalide B (C
20H
24O
10) and ginkalide C (C
20H
24O
11) total amount count 4.5～5.5mg.
Bilobalide injection has blood circulation promoting and blood stasis dispelling, the effect of dredge the meridian passage.Be used for apoplexy apoplex involving the channels and collaterals (light moderate cerebral infarction) convalescent period congestion resistance network disease, disease is seen hemiplegia, crooked mouth and tongue, and speech is stuttering puckery, numb limbs and tense tendons etc.
Injection of the present invention adopts high-purity, component is clear and the bilobalide of fixed ratio, and effective site content is more than 95%, and wherein 4 kinds of components contain bilobalide (C
15H
18O
8) be 25.0%～50.0%, ginkalide A (C
20H
24O
9) be 20.0%～45.0%, ginkalide B (C
20H
24O
10) be 10.0%～30.0%, ginkalide C (C
20H
24O
11) be 5.0%～15.0%, total ginkgolic acids meets European Union's requirement less than 5ppm.Whole production technology has been guaranteed the stable and clinical efficacy of terpene lactone component ratio; And gel chromatography and liquid-matter GC-MS be applied to detection range; Macromolecular substances in the conventional Chinese medicine injection agent producing process, polymer and proteinic residual have been solved; Reduced the generation of untoward reaction widely, 4 total peaks of finger printing similarity is greater than 0.95.To the water-fast character of terpene lactone; Filter out effective cosolvent---ethanol-glycerol mixed solvent; Promptly solved the technical barrier of common process; Avoid adopting other Acidity of Aikalinity adjuvant to cause the bilobalide open loop and destroy the main constituent structure, make effective ingredient more stable, also reached the aseptic requirement of injection simultaneously.Quality control index is from multi-angle, comprehensive control product quality; Set up perfect Control of Impurities method effectively; Adopted the strictest detection technique control product quality of Chinese medicine injection at present such as finger printing, undue toxicity, irritation test, haemolysis and cohesion, hypersensitive test; Improved the safety of clinical use greatly, Quality Control Technology considerably beyond generally acknowledging gingko leaf extract injection at present in the world, is reached advanced world standards.
The present invention is the effective site Chinese medicine; Macromole, protein have been removed effectively; Avoided the generation of untoward reaction, and have that energy consumption is low, determined curative effect, simple to operate, easy to control the quality, characteristics such as production cost is low, higher than other Semen Ginkgo class injection safety.
Description of drawings
Figure 101 criticizes bilobalide injection LC-MS figure (molecular weight 400～1000).
1 crowd of bilobalide injection LC-MS figure of Figure 20 (molecular weight 400～3000).
2 crowdes of bilobalide injection LC-MS figure of Figure 30 (molecular weight 400～1000).
2 crowdes of bilobalide injection LC-MS figure of Figure 40 (molecular weight 400～3000).
3 crowdes of bilobalide injection LC-MS figure of Figure 50 (molecular weight 400～1000).
3 crowdes of bilobalide injection LC-MS figure of Figure 60 (molecular weight 400～3000).
Fig. 7 Semen Ginkgo eo-acid linear relationship chart.
Fig. 8 bilobalide injection reference fingerprint;
Wherein total coneincone 2: ginkalide C, peak 3: bilobalide, peak 4: ginkalide A, peak 5: ginkalide B.
The specific embodiment
Raw material bilobalide of the present invention contains bilobalide (C
15H
18O
8) 25.0%～50.0%, ginkalide A (C
20H
24O
9) 20.0%～45.0%, ginkalide B (C
20H
24O
10) 10.0%～30.0%, ginkalide C (C
20H
24O
11) 5.0%～15.0%, and bilobalide, ginkalide A, ginkalide B, ginkalide C total amount are greater than 95%.
It below is the foundation of selecting cosolvent and pH regulator agent.
One, the selection of cosolvent: according to the character of supplementary material and preparation technology's requirement of injection, the usage ratio of screening solvent and cosolvent.
Test 1: get ethanol 1200ml and glycerol 800ml, mixing adds bilobalide 10g, stirring and dissolving, adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Test 2: get ethanol 400ml and glycerol 1600ml, mixing adds bilobalide 10g, stirring and dissolving, adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Test 3: get ethanol 300ml and glycerol 1700ml, mixing adds bilobalide 10g, stirring and dissolving, adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Test 4: get ethanol 1200ml and glycerol 400ml, mixing adds bilobalide 10g, and stirring and dissolving adds the injection water to 2000ml, adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Test 5: get ethanol 400ml and glycerol 1000ml, mixing adds bilobalide 10g, and stirring and dissolving adds the injection water to 2000ml, adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Above-mentioned sample is carried out low temperature freezing-thawing test circulation 3 times, check that respectively character, particulate matter and the pH value of solution of solution changes, and whether have crystal to separate out after the viewing test.Result of the test is seen table 1.
Table 1 cosolvent screening test result
When not adding the injection water, the volume ratio of ethanol and glycerol is 1: 1～5: 1 in the medicinal liquid, and optimal proportion is 3: 2; Optimal proportion is 4: 1 when adding the injection water.Proportion of ethanol is big more, and the bilobalide dissolution velocity is fast more.Medicinal liquid pH, particulate matter and appearance character after freezing-thawing test all meet the pharmacopeia regulation.
Two, the selection of pH regulator agent:, screen with 5～10% citric acid solns or 1～10% hydrochloric acid solution respectively because bilobalide is relatively stable under solutions of weak acidity.
Test 1: get ethanol 1200ml and glycerol 800ml, mixing adds bilobalide 10g, stirring and dissolving, with 5% hydrochloric acid adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Test 2: get ethanol 400ml and glycerol 1600ml, mixing adds bilobalide 10g, stirring and dissolving, with 10% hydrochloric acid adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Test 3: get ethanol 1200ml and glycerol 400ml, mixing adds bilobalide 10g, and stirring and dissolving adds the injection water to 2000ml, with 10% citric acid soln adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Test 4: get ethanol 1200ml and glycerol 800ml, mixing adds bilobalide 10g, stirring and dissolving, with 10% citric acid soln adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Character, particulate matter and the sterilization front and back pH value of solution of checking solution respectively change, and sample was placed 3 days in low temperature (0～6 ℃), and whether observe under the low temperature placement condition has crystal to separate out.Result of the test is seen table 2.
Table 2 pH regulator agent screening test result
Through pH regulator agent screening test, available 5～10% citric acid solns of pH regulator agent or 1～10% hydrochloric acid solution are regulated pH with hydrochloric acid solution in these article, slightly raise after the sterilization, but change not quite, and citric acid is a weak acid, and it is more stable to regulate pH.Hydrochloric acid and citric acid all can be used as the pH value of solution regulator.
The inventor studies the foregoing invention content and explains, is used to prove technique effect of the present invention.Following test is used to further specify and explain the present invention, but does not limit the present invention.
Test Example 1 bilobalide quality control---assay
Chromatographic condition and system suitability test: with the octadecylsilane chemically bonded silica is filler; With methanol-oxolane-water (25: 10: 65) is mobile phase; Use evaporative light scattering detector, drift tube temperature: 105 ℃; Flow rate of carrier gas: 3.00L/min; Column temperature: 40 ℃; Number of theoretical plate calculates by the bilobalide peak should be not less than 2500.The separating degree at bilobalide peak and ginkalide C peak should be greater than 1.5.
The preparation of reference substance solution: it is an amount of that precision takes by weighing bilobalide reference substance, ginkalide A reference substance, ginkalide B reference substance, ginkalide C reference substance respectively; Add the also quantitative dilution of dissolve with methanol and process the mixed solution that every 1ml contains 0.15mg, 0.12mg, 0.1mg, 0.1mg respectively; Shake up, promptly get.
The preparation of need testing solution: get bilobalide 6mg, the accurate title, decide, and puts to add methanol 1ml dissolving in the 10ml measuring bottle, adds mobile phase and be diluted to scale, shakes up, and promptly gets.
Algoscopy: accurate respectively reference substance solution 10 μ l, 20 μ l and need testing solution 10～20 μ l of drawing; Inject high performance liquid chromatograph; The record chromatogram calculates the content of bilobalide, ginkalide A, ginkalide B and ginkalide C respectively with external standard two-point method logarithmic equation, promptly gets.
Press dry product and calculate, contain bilobalide (C
15H
18O
8) be 25.0%～50.0%, bilobalide-containing A (C
20H
24O
9) be 20.0%～45.0%, bilobalide-containing B (C
20H
24O
10) be 10.0%～30.0%, bilobalide-containing C (C
20H
24O
11) be 5.0%～15.0%, and bilobalide, ginkalide A, ginkalide B, ginkalide C total amount are not less than 95.0%.
Test Example 2 bilobalide injection quality controls---related substance inspection
(1) protein: get injection 2ml of the present invention, add water and process 50ml, as need testing solution.Take by weighing the about 50mg of Coomassie brilliant blue G-250, be dissolved in the 25ml ethanol, add the phosphoric acid 50ml of 85% (w/v) again, thin up shakes up to 500ml, filters, and precision is measured filtrating 5ml and put in the test tube, adds 1ml test sample stock solution again, shakes up, and places 3min.Do blank with method, under the 595nm wavelength, measure absorbance, the need testing solution absorbance is less than 0.05.
(2) tannin: get injection 1ml of the present invention and add 1 of spirit of vinegar, add 5 of gelatin sodium chloride test solutions again, shake up, placed 10 minutes, do not occur muddy or deposition.
(3) resin: get injection 5ml of the present invention, add 1 of hydrochloric acid, placed 30 minutes, should not have resinoid and separate out.
(4) oxalates: get injection 2ml of the present invention, regulate pH value to 1～2 with dilute hydrochloric acid, filter, it is 5～6 that filtrating uses ammonia to regulate pH value, adds 3 of 3% calcium chloride solutions, places 10 minutes, does not occur muddy or deposition.
(5) potassium ion: get injection 2ml of the present invention; Put in the 10ml nessler colorimetric tube; Add alkaline formaldehyde solution 0.6ml, 2 of 3%EDTA solution, 3% sodium tetraphenylborate solution 0.5ml, thin up is to 10ml, and other gets standard Klorvess Liquid 0.8ml; With the method test, the turbidity of need testing solution is lower than contrast solution.
Test Example 3 bilobalide injection quality controls---undue toxicity's inspection
The preparation of need testing solution: get injection of the present invention and add the chlorination sodium injection and process the solution that contains 0.2mg among every 1ml.
Inspection technique: get 5 of body weight 17～20g mices, inject mouse tail vein need testing solution 0.5ml respectively, do not have dead in 48 hours.
Test Example 4 bilobalide injection quality control---pyrogen tests
The preparation of need testing solution: get injection 2ml of the present invention, join among the 0.9% sodium chloride injection 50ml, shake up.
Inspection technique: get 3 of rabbit, measure after its normal body temperature in 15 minutes, 5ml slowly injects need testing solution from ear vein by the every 1kg injection of rabbit body weight; Whenever measured body temperature 1 time at a distance from 30 minutes; Survey altogether 6 times, fervescence all should be lower than 0.6 ℃, and 3 rabbit body temperature rising summations are lower than 1.3 ℃.
Test Example 5 bilobalide injection quality controls---haemolysis and cohesion inspection
The preparation of need testing solution: get injection 6ml of the present invention, join among the 0.9% sodium chloride injection 100ml and shake up.
Inspection technique: get 5 of clean teat glasses, numbering, 1, No. 2 pipe is for the test sample pipe, manages negative control tube No. 3, manages positive control tube, No. 5 pipe test sample control tube No. 4.By adding 2% red cell suspension, 0.9% sodium chloride solution, distilled water shown in the table 3 successively, put immediately behind the mixing in 37 ℃ ± 0.5 ℃ the calorstat and carry out incubation.
Test test solution addition is seen table 3.
Table 3 haemolysis and agglutination test test solution addition
| The test tube numbering | 1 | 2 | 3 | 4 | 5 |
| 2% red cell suspension/ml | 2.5 | 2.5 | 2.5 | 2.5 | |
| 0.9% sodium chloride solution/ml | 2.2 | 2.2 | 2.5 | 4.7 | |
| Distilled water/ml | 2.5 | ||||
| Need testing solution/ml | 0.3 | 0.3 | 0.3 |
Be clear and bright redness like the solution in the test tube, it is residual or have a small amount of erythrocyte residual that the bottom is acellular, and showing has haemolysis to take place; All sink like erythrocyte, the supernatant achromatism and clarity, though or supernatant coloured clear and bright, 1,2, No. 5 pipe perusal no significant difference then shows no haemolysis generation.If in the solution brownish red or rufous flocculent deposit are arranged, reverse 3 times gently and still do not disperse, show that having red blood cell condensation takes place.
Observe no haemolysis and aggregation after 3 hours.
Test Example 6 bilobalide injection quality controls---macromolecular substances and polymer inspection
A. test apparatus and reagent
Dextran reference substance D2000 (blue dextran 2000), Nat'l Pharmaceutical & Biological Products Control Institute, lot number 140646-2000-01
Glucose reference substance (D0), content: 99.5%, lot number 086K0166, SIGMA.
Ultra-pure water makes with Millipore-Q ultra-pure water system.
All the other reagent are analytical pure.
Sample source: bilobalide injection, lot number 100501,100502,100503, specification is: 2ml.
B. the selection of mobile phase
Choose 0.71% (including 0.02% Hydrazoic acid,sodium salt) metabisulfite solution as mobile phase.
C. the selection of detector
Select the common detector differential refraction detector for use, this detector all has good response for the material that has refraction coefficient difference.
D. quasi-definite chromatographic condition
Chromatographic column: Phenomenex BioSep-SEC-S2000,300 * 7.8mm, 5um
Mobile phase: 0.71% (including 0.02% Hydrazoic acid,sodium salt) metabisulfite solution
Column temperature: 35 ℃, detector temperature: 35 ℃, flow velocity: 0.5ml/min
E. the molecular weight of bilobalide
| Bilobalide | Ginkalide A | Ginkalide B | Ginkalide C | Bilobalide | Bilobalide J |
| Molecular formula | C 20H 24O 9 | C 20H 24O 10 | C 20H 24O 11 | C 15H 18O 8 | C 20H 24O 10 |
| Molecular weight | 408.4 | 424.4 | 440.4 | 326.3 | 424.4 |
F. methodological study
1. dextran and glucose reference substance are added the solution that mobile phase is processed 10mg/ml respectively, accurate respectively each 20 μ l of reference substance solution, the injecting chromatograph drawn; The record chromatogram; Dextran is at retention time 9.816 ' go out peak as a result, and glucose shows the employing gel chromatography in retention time 18.712 ' go out the peak; The material that molecular weight is big goes out the peak earlier, goes out the peak behind the little material of molecular weight.
2. get bilobalide injection 2ml, add dextran reference substance solution (10mg/ml) 1ml, mixing; The accurate 10 μ l that draw, injecting chromatograph, record chromatogram; The result is in retention time 9.698 ' detect dextran, and bilobalide injection becomes swarming all to go out the peak later at 18min, show molecular weight at 180～450 appearance times about 18min; Molecular weight 5000～2000000 appearance times show that it is feasible adopting gel chromatography to detect macromolecular substances about 9min.
Do not contain macromole, polymer in these article in order to verify again, so carried out the test of LC-MS LC-MS again.
A. chromatographic condition
Mobile phase: methanol-water (90: 10), chromatographic column: Agilent RX-C
18(2.1 * 50mm)
Column temperature: 25 ℃ of flow velocity: 0.3ml/min
B. the preparation of need testing solution
Precision is measured bilobalide injection 2ml of the present invention and is put in the 10ml measuring bottle, adds mobile phase and is diluted to scale, shakes up the preparation of need testing solution.
C.LC-MS coupling test
According to the test method of confirming, get each 10 μ l of three batches of need testing solutions respectively, injecting chromatograph, at 400～1000 and 400～3000 molecular weight ranges build-in test molecular weight, result of the test is seen table 4.
Table 4 LC-MS coupling molecular weight determination result
| ?[M+Na] + | 419.1、431.5、447.4、463.3、475.7、532.2、588.8、701.8 |
| M | 396.1、408.5、424.4、440.3、452.7、509.2、678.8 |
From LC-MS coupling molecular weight determination result; Detect ginkalide A (molecular weight 408.5), ginkalide B (molecular weight 424.4), ginkalide C (molecular weight 440.4) respectively, in full accord with the active component molecular weight of bilobalide since the test molecule weight range respectively 400～1000 and 400-3000 between; Bilobalide molecular weight (326.3) is not tested; Do not detect the material greater than 700 above molecular weight in the injection of the present invention, other micro-low-molecular-weight material exists, and does not influence injection quality of the present invention; Through LC-MS to bilobalide in the molecular weight test of different components, explain not contain macromole or polymer in these article.
Three batches of bilobalide injection determining molecular weight scopes are respectively between 400～1000 and 400～3000, and the LC-MS collection of illustrative plates is seen Fig. 1～6.
Test Example 7 bilobalide injection quality controls---total ginkgolic acids inspection
Chromatographic condition and system suitability test: with the octadecylsilane chemically bonded silica is filler; With methanol-1% glacial acetic acid (90: 10) is mobile phase; Flow velocity 1.0ml/min; The detection wavelength is 310nm.Number of theoretical plate should be not less than 4000 by the Semen Ginkgo neo-acid peak.
The preparation of reference substance solution: it is an amount of to get Semen Ginkgo eo-acid reference substance, accurate claims surely, adds methanol and processes solution that every 1ml contains 5 μ g as reference substance solution; It is an amount of that other gets the total ginkgolic acids reference substance, and accurate the title decides, and adds methanol and processes the solution that every 1ml contains 100 μ g, uses contrast solution as the location.
The need testing solution preparation: get injection 1ml of the present invention, thin up shakes up to 10ml; With n-hexane extraction 3 times (20ml, 15ml, 15ml), merge normal hexane liquid, put evaporate to dryness in the water-bath; Residue adds dissolve with methanol and is diluted to 2ml, shakes up, as need testing solution.
Algoscopy: accurate need testing solution, reference substance solution and the location of drawing is with each 20 μ l of contrast solution; Inject chromatograph of liquid; Calculate in the need testing solution total peak area with the corresponding chromatographic peak of total ginkgolic acids reference substance, calculate total ginkgolic acids content with Semen Ginkgo eo-acid reference substance external standard method.Contain total ginkgolic acids and be lower than 1ppm.
Methodological study:
A. linear test
The preparation of reference substance solution: get Semen Ginkgo eo-acid reference substance, add methanol and process the solution that every 1ml contains Semen Ginkgo eo-acid 0.104mg, as reference substance solution (1); Precision is measured reference substance solution (1) 0.5ml and is put in the 100ml measuring bottle, is diluted to scale with methanol, shakes up, as reference substance solution (2).
Algoscopy: precision is measured reference substance solution (1) 2.5 μ l, 5 μ l, 10 μ l, 20 μ l and reference substance solution (2) 10 μ l respectively, injects chromatograph of liquid, record chromatogram under above-mentioned chromatographic condition.Result of the test is seen table 5.
Table 5 Semen Ginkgo eo-acid linear test result
| Sample size (ng) | 5.2 | 260 | 520 | 1040 | 2080 |
| Peak area | 2545 | 102162 | 213093 | 426512 | 843249 |
With the sample size is vertical coordinate, is abscissa with the peak area, carries out linear regression and gets regression equation: Y=0.0025X-0.6448r=0.9999.
Result of the test shows that Semen Ginkgo eo-acid sample size is between 5.2～2080ng, and sample size and peak area are good linear relationship.
Semen Ginkgo eo-acid linear relationship chart is seen Fig. 7.
B. quantitative limit detectability test
Algoscopy: precision is measured reference substance solution (2) 5 μ l and 2 μ l under the linear test item respectively, injects chromatograph of liquid, the record chromatogram, and the quantitative limit and the detectability of calculating Semen Ginkgo eo-acid, the Semen Ginkgo eo-acid quantitatively is limited to 2.24ng, detects to be limited to 0.69ng.
C. precision test
Algoscopy: precision is measured Semen Ginkgo eo-acid reference substance solution (2) 20 μ l, continuous sample introduction 5 times, and the record chromatogram calculates the RSD value.Result of the test is seen table 6.
Table 6 Precision test result
| |
1 | 2 | 3 | 4 | 5 |
| Peak area | 4850 | 4874 | 4845 | 4873 | 4837 |
Reference substance solution peak area RSD value is 0.35%, and result of the test shows that precision is good.
D. stability test
Algoscopy: precision is measured Semen Ginkgo eo-acid reference substance solution (2) 20 μ l, and respectively at 0,2,4,8,12 hour injection chromatograph of liquid, under above-mentioned chromatographic condition, the record chromatogram calculated the RSD value.Result of the test is seen table 7.
Table 7 reference substance solution stability test result
| Time (h) | 0 | 2 | 4 | 8 | 12 |
| Peak area | 4861 | 4853 | 4865 | 4843 | 4870 |
Result of the test shows that reference substance solution is stable in 12 hours.
E. recovery test
The preparation of reference substance solution: precision takes by weighing Semen Ginkgo eo-acid reference substance 5.03mg and puts in the 10ml measuring bottle, adds dissolve with methanol and is diluted to scale, shakes up, as reference substance solution.
The need testing solution preparation: get 9 parts of injection of the present invention, per 3 parts add reference substance solution 1.6ml, 2.0ml and 2.4ml respectively, shake up; With n-hexane extraction 3 times (20ml, 15ml, 15ml); Merge n-hexane extract, put evaporate to dryness in the water-bath, residue adds the chloroform dissolving and quantitatively is diluted to 2ml; Shake up, as need testing solution.
Algoscopy: precision is measured reference substance solution and each 20 μ l of need testing solution respectively, under above-mentioned chromatographic condition, and record chromatogram, calculate recovery rate.Result of the test is seen table 8.
Table 8 recovery test result
Average recovery rate is 98.04%, and RSD is 1.41%, and result of the test shows that method detects limiting the quantity of accurately and reliably of ginkgoic acid.
Test Example 8 bilobalide injection quality controls---finger printing inspection
Chromatographic condition and system suitability test: with the octadecylsilane chemically bonded silica is filler; With methanol-oxolane-water (25: 10: 65) is mobile phase; Use evaporative light scattering detector, drift tube temperature: 105 ℃; Flow rate of carrier gas: 3.00L/min; Column temperature: 40 ℃; Number of theoretical plate calculates by the bilobalide peak should be not less than 2500.The separating degree at bilobalide peak and ginkalide C peak should be greater than 1.5.
The preparation of object of reference solution: it is an amount of that precision takes by weighing bilobalide reference substance, ginkalide A reference substance, ginkalide B reference substance, ginkalide C reference substance respectively; Add methanol and process the mixed solution that every 1ml contains 0.15mg, 0.12mg, 0.1mg, 0.1mg respectively; Shake up, as object of reference solution.
The preparation of need testing solution: precision is measured injection 1ml of the present invention, adds phosphate buffer solution (pH6.5) 14ml, shakes up last Extrelut-20 post; Adsorbed 15 minutes, and, collected eluent with ethyl acetate 100ml eluting; Put evaporate to dryness in the water-bath, residue dissolves with mobile phase and is transferred in the 10ml measuring bottle, adds mobile phase and is diluted to scale; Shake up, filter with 0.45 μ m microporous filter membrane, as need testing solution.
Algoscopy: accurate respectively object of reference solution and each 20 μ l of need testing solution of drawing, inject chromatograph of liquid, write down 60 minutes chromatogram.
Press chromatographic fingerprints of Chinese materia medica similarity evaluation system, test sample finger printing and reference fingerprint calculate greater than 0.95 through similarity.
The inventor studies the foregoing invention content and explains, is used to prove technique effect of the present invention.Following test is used to further specify and explain the present invention, but does not limit the present invention.
In the bilobalide finger printing, wherein peak 2 is that ginkalide C, peak 3 are ginkalide B for bilobalide, peak 4 for ginkalide A, peak 5, and 4 characteristic peaks all can be corresponding one by one in finger printing in the injection of the present invention.The bilobalide injection reference fingerprint is seen Fig. 8.
At first adopting committee of pharmacopeia finger printing designated software in 2004---the chromatographic fingerprints of Chinese materia medica similarity evaluation A of system version generates reference fingerprint to 10 batches of bilobalide injections; Press the chromatographic fingerprints of Chinese materia medica similarity evaluation B of system version again, calculate 10 batches of bilobalide test sample finger printing and reference fingerprint similarity.Result of the test is seen table 9.
10 batches of bilobalide similarity result of table 9
| Lot number | Similarity |
| 100501 | 0.991 |
| 100502 | 0.987 |
| 100503 | 0.995 |
| 100504 | 0.996 |
| 110201 | 0.993 |
| 110202 | 0.989 |
| 110203 | 0.992 |
| 110701 | 0.989 |
| 110702 | 0.993 |
| 110703 | 0.991 |
The similarity of 4 total peaks of 10 batches of bilobalide injection finger printing of the present invention and reference fingerprint is all greater than 0.95.
Test Example 9 bilobalide injection quality control---assays
Chromatographic condition and system suitability test: with the octadecylsilane chemically bonded silica is filler; With methanol-oxolane-water (25: 10: 65) is mobile phase; Use evaporative light scattering detector, drift tube temperature: 105 ℃; Flow rate of carrier gas: 3.00L/min; Column temperature: 40 ℃; Number of theoretical plate calculates by the bilobalide peak should be not less than 2500.The separating degree at bilobalide peak and ginkalide C peak should be greater than 1.5.
The preparation of reference substance solution: it is an amount of that precision takes by weighing bilobalide reference substance, ginkalide A reference substance, ginkalide B reference substance, ginkalide C reference substance respectively; Add methanol and process the mixed solution that every 1ml contains 0.15mg, 0.12mg, 0.1mg, 0.1mg respectively; Shake up, as reference substance solution.
The preparation of need testing solution: precision is measured injection 1ml of the present invention, adds phosphate buffer solution (pH6.5) 14ml, shakes up last Extrelut-20 post; Adsorbed 15 minutes, and, collected eluent with ethyl acetate 100ml gradation eluting; Evaporate to dryness in water-bath, residue dissolves with mobile phase and is transferred in the 10ml measuring bottle, adds mobile phase and is diluted to scale; Shake up, filter with 0.45 μ m microporous filter membrane, as need testing solution.
Algoscopy: accurate respectively reference substance solution 10 μ l, 20 μ l and the need testing solution 15 μ l of drawing; Inject high performance liquid chromatograph; Write down chromatogram, calculate the content of bilobalide, ginkalide A, ginkalide B and ginkalide C with external standard two-point method logarithmic equation respectively.
The every 1ml of injection of the present invention contains the Semen Ginkgo terpene lactone with bilobalide (C
15H
18O
8), ginkalide A (C
20H
24O
9), ginkalide B (C
20H
24O
10) and ginkalide C (C
20H
24O
11) the total amount meter, content is 1-10mg/ml, preferred 4.25～5.75mg.
Following injection preparation example of the present invention.
Prescription:
Get the ethanol of glycerol and total amount 80% by prescription, mixing adds bilobalide, and stirring and dissolving adds ethanol to full dose, stir, with 1～10% hydrochloric acid adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Prescription:
Get the ethanol of glycerol and total amount 80% by prescription, mixing adds bilobalide, and stirring and dissolving adds ethanol to full dose, stir, with 5～10% citron acid for adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Prescription:
Get the ethanol of glycerol and total amount 80% by prescription, mixing adds bilobalide, and stirring and dissolving adds ethanol to full dose, stir, with 5～10% citron acid for adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Embodiment 4
Prescription:
Get ethanol and glycerol by prescription, mixing adds bilobalide, and stirring and dissolving adds to the full amount of water for injection, stir, with 5～10% citron acid for adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
Prescription:
Get ethanol and glycerol by prescription, mixing adds bilobalide, and stirring and dissolving adds to the full amount of water for injection, stir, with 1～10% hydrochloric acid adjusting pH to 3.2～3.8, intermediate after the assay was approved, be filtered to clear and bright, embedding, sterilization.
The active component content of the foregoing description 1～5, injection quality all meet bilobalide injector prescription of the present invention.
Claims (8)
1. bilobalide injection, it is characterized in that: said injection contains following compositions:
2. bilobalide injection according to claim 1 is characterized in that: said injection contains following compositions: bilobalide: in terpene lactone 5mg/ml
Glycerol 0.4ml/ml
Ethanol 0.6ml/ml.
3. bilobalide injection according to claim 1 is characterized in that: said injection contains following compositions:
4. bilobalide injection according to claim 1 is characterized in that: the volume ratio of ethanol and glycerol is 1～5 part of an ethanol, 1 part of glycerol.
5. according to each described bilobalide injection of claim 1-4; It is characterized in that: bilobalide is pressed dry product and is calculated; Containing bilobalide is 25.0%～50.0%; Bilobalide-containing A is 20.0%～45.0%, and bilobalide-containing B is 10.0%～30.0%, and bilobalide-containing C is 5.0%～15.0%; And bilobalide, ginkalide A, ginkalide B, ginkalide C total amount are not less than 95.0%.
6. the method for preparing of the described bilobalide injection of claim 1 is characterized in that: contain the following compositions preparation by injection:
Step is following:
A) preparation: mixed ethanol and glycerol, add bilobalide, dissolving, add ethanol or water for injection to full dose, regulate pH value to 2.5～6.5 with 5～10% citric acid solns or 1～10% hydrochloric acid solution;
B) filtration sterilization;
C) embedding;
D) sterilization.
7. the method for preparing of bilobalide injection according to claim 6 is characterized in that: step a) preparation adjust pH to 3.2～3.8.
8. according to the method for preparing of claim 6 or 7 described bilobalide injections, it is characterized in that: during the step a) preparation, the volume ratio of ethanol and glycerol is 1～5 part of an ethanol, 1 part of glycerol.
Priority Applications (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012101207876A CN102626383A (en) | 2012-04-23 | 2012-04-23 | Bilobalide injection and preparation method thereof |
| PCT/CN2012/075631 WO2013159411A1 (en) | 2012-04-23 | 2012-05-17 | Ginkgolide injection and preparation method thereof |
| SG11201405423PA SG11201405423PA (en) | 2012-04-23 | 2012-05-17 | Ginkgolide injection and preparation method thereof |
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|---|---|---|---|
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|---|---|
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|---|---|---|---|---|
| CN106580872A (en) * | 2015-10-16 | 2017-04-26 | 上海现代药物制剂工程研究中心有限公司 | Ginkgolides B injection and preparation method thereof |
| CN109925310A (en) * | 2013-12-10 | 2019-06-25 | 成都百裕制药股份有限公司 | Blood-pressure drug containing therapeutically effective amount ginkgolides |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1430968A (en) * | 2003-01-30 | 2003-07-23 | 复旦大学 | Injection liquid of extractive gingko leaves and its preparing method |
| CN1887283A (en) * | 2006-07-26 | 2007-01-03 | 孙毅 | Medicine composition containing bailobalide |
| CN1887282A (en) * | 2006-07-26 | 2007-01-03 | 孙毅 | Medicine composition containing bailobalide |
| CN101926835A (en) * | 2009-06-19 | 2010-12-29 | 山东新时代药业有限公司 | Method for extracting gingko total terpene lactones and injection containing same |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN100349568C (en) * | 2006-04-18 | 2007-11-21 | 张国清 | Intravenous administered injection of ginkgolide B, its preparation method and application |
| CN101301267B (en) * | 2008-05-21 | 2011-07-13 | 南京海陵中药制药工艺技术研究有限公司 | Bilobalide B injection and preparation thereof |
-
2012
- 2012-04-23 CN CN2012101207876A patent/CN102626383A/en active Pending
- 2012-05-17 WO PCT/CN2012/075631 patent/WO2013159411A1/en active Application Filing
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1430968A (en) * | 2003-01-30 | 2003-07-23 | 复旦大学 | Injection liquid of extractive gingko leaves and its preparing method |
| CN1887283A (en) * | 2006-07-26 | 2007-01-03 | 孙毅 | Medicine composition containing bailobalide |
| CN1887282A (en) * | 2006-07-26 | 2007-01-03 | 孙毅 | Medicine composition containing bailobalide |
| CN101926835A (en) * | 2009-06-19 | 2010-12-29 | 山东新时代药业有限公司 | Method for extracting gingko total terpene lactones and injection containing same |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109925310A (en) * | 2013-12-10 | 2019-06-25 | 成都百裕制药股份有限公司 | Blood-pressure drug containing therapeutically effective amount ginkgolides |
| CN106580872A (en) * | 2015-10-16 | 2017-04-26 | 上海现代药物制剂工程研究中心有限公司 | Ginkgolides B injection and preparation method thereof |
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