CN104435067B - A kind of Cortex Eucommiae components composition and preparation thereof with myocardium protecting action - Google Patents
A kind of Cortex Eucommiae components composition and preparation thereof with myocardium protecting action Download PDFInfo
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Abstract
The invention belongs to the field of Chinese medicines, be specifically related to a kind of Cortex Eucommiae components composition of Cardiovarscular.The composition of said composition is all from Traditional Chinese medicine eucommia bark, and it is specifically the eucommiae total flavones of 10 ~ 80% by percentage by weight, the Cortex Eucommiae iridoid of 10 ~ 80% and the Cortex Eucommiae lignanoid composition of 10 ~ 80%.The percentage by weight of described compositions is preferably the eucommiae total flavones of 20 ~ 40%, the Cortex Eucommiae iridoid of 40 ~ 60% and the Cortex Eucommiae lignanoid composition of 20 ~ 50%.The present invention, by combining the single effective site of the Cortex Eucommiae, by adjustment associated weight proportioning, obtains a kind of clear curative effect, has the Cortex Eucommiae components composition of clinical development prospect.
Description
Technical field
The invention belongs to the field of Chinese medicines, be specifically related to a kind of Cortex Eucommiae components composition of Cardiovarscular.
Background technology
The Cortex Eucommiae is the dry bark of Eucommiaceae plant Cortex Eucommiae EucommiaulmoidesOliv., another name thinks celestial being, kapok etc., begin to be loaded in Shennong's Herbal, be classified as Chinese medicine top grade, there is invigorating the liver and kidney, bone and muscle strengthening, the effect such as antiabortive, be applied to the habitual abortion, hypertension etc. for the treatment of deficiency of the liver and kindey, soreness of waist and knee joint, gestation clinically, the normal and compatibility such as Fructus Psoraleae, Radix Dipsaci.
Cardiovascular disease, is also called blood circulation diseases, and blood circulation refers to hemophoric Organ and tissue in human body, mainly comprises heart and blood vessel, can be subdivided into acute and chronic, generally all relevant with arteriosclerosis.Myocardial ischemia refers to that the hemoperfusion of heart reduces, and cause the oxygen-supplying amount of heart to reduce, energy metabolism of myocardial is abnormal, can not support a kind of pathological state of normal heart action.The coronary stenosis that coronary atherosclerosis causes causes main, the modal cause of disease of myocardial ischemia, and then cause myocardial ischemia-anoxemia, and therefore coronary heart disease is the main cause of myocardial ischemia.Myocardial ischemia is mainly divided into latent coronary heart disease, angina pectoris, myocardial infarction type, ischemic cardiomyopathy and sudden death 5 class.
The common reason of myocardial ischemia is coronary atherosclerosis, secondly also has inflammation (rheumatic, syphilis, mucocutaneous lymphnode syndrome and thromboangitis obliterans etc.), spasm, thromboembolism, connective tissue disease, wound and congenital malformation etc. multiple.Epidemiological study finds, the significant risk factor relevant to atherosclerosis is hyperlipemia, hypertension, diabetes, smoking, obesity, homocysteine increases, physical exertion is few, advanced age and male etc.The damage mechanisms of ischemic myocardium comprises energy deficiency, calcium overload, free radical generate increase, the aspect such as cardiac damage, permeability of cell membrane increase that acidosis causes.
Research shows that aqueous extract from Eucommia ulmoides significantly can reduce the synthesis of fatty acid, reduce 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase activity, improve the level of high density lipoprotein (HDL) cholesterol, reduce plasma cholesterol and triglyceride concentration.In addition, aqueous extract from Eucommia ulmoides can also suppress Cu
2+the oxidative modification of the low density lipoprotein, LDL (LDL) of mediation.As everyone knows, the activation of HMG-CoA reductase, plasma cholesterol and triglyceride levels rising can promote the formation of atheromatous plaque; HDL or HDL cholesterol has very strong antiatherogenic effect, the oxidative modification of another kind of lipoprotein LDL then advances the pathological changes of atherosclerosis (AS) to be formed, so aqueous extract from Eucommia ulmoides is by reducing the oxidative modification level of plasma cholesterol and LDL, improve HDL cholesterol levels, suppress developing of AS.Aqueous extract from Eucommia ulmoides can also produce hypotensive effect by vasodilator, and has endothelium-dependent relaxation.In addition, separately there are some researches show, the activated monomer in the Cortex Eucommiae also shows at pharmacologically actives such as blood pressure lowering, blood fat reducing, antioxidation, atherosclerosiss.Such as, iridoids monomer genipin and geniposide in the Cortex Eucommiae, phenylpropionic acid compound ferulic acid and caffeic acid, flavone compound Quercetin, rutin and baicalin, triterpenoid compound betulic acid and ursolic acid.
Although current research shows that the Cortex Eucommiae has certain therapeutical effect to cardiovascular disease, before Eucommiales, the product of main listing mainly concentrates on blood pressure lowering field, and the application of other field is less.CN200710035744 discloses the application of a kind of Cortex Eucommiae lignans extract in anti-cardiovascular reconstruction, its mechanism remains relevant to the hypotensive effect of the Cortex Eucommiae (by regulating renin angiotensin RAS system), and adopt the single effective site of the Cortex Eucommiae (lignanoid), active component is comparatively single, and the therapeutic effect for cardiovascular disease is also extremely limited.
Summary of the invention
The present invention finds the single effective site of the Cortex Eucommiae by research, and such as, iridoid, lignanoid or flavones ingredient, in the curative effect of myocardial ischemia, have effect, but curative effect is not obvious, cannot be used for the treatment of cardiovascular patient as medicine listing.The present invention, by combining the single effective site of the Cortex Eucommiae, by adjustment associated weight proportioning, obtains a kind of clear curative effect, has the Cortex Eucommiae components composition of clinical development prospect.
An object of the present invention is to provide a kind of Cortex Eucommiae components composition with myocardium protecting action; the composition of said composition is all from Traditional Chinese medicine eucommia bark; it is specifically the eucommiae total flavones of 10 ~ 80% by percentage by weight, the Cortex Eucommiae iridoid of 10 ~ 80% and the Cortex Eucommiae lignanoid composition of 10 ~ 80%.
In one embodiment of the invention, the percentage by weight of described compositions is preferably the eucommiae total flavones of 20 ~ 40%, the Cortex Eucommiae iridoid of 40 ~ 60% and the Cortex Eucommiae lignanoid composition of 20 ~ 50%.In the present invention further embodiment, the percentage by weight of described compositions can be the eucommiae total flavones of 30%, the Cortex Eucommiae iridoid of 50% and the Cortex Eucommiae lignanoid composition of 20%; The percentage by weight of described compositions also can be the eucommiae total flavones of 25%, the Cortex Eucommiae iridoid of 45% and the Cortex Eucommiae lignanoid composition of 30%.
Effective ingredient in Chinese extract of the present invention with prior art self-control or can be obtained by commercial channel, also can the method listed of the present invention be prepared.
The preparation of eucommiae total flavones: take Folium Eucommiae coarse powder 5.0kg, the ethanol 40L of preparation 45%, pH is adjusted to be 4 with concentrated hydrochloric acid, at 60 ~ 70 DEG C, carrying out stirring extraction twice, (first time 25L alcoholic solution extracts 2 hours, second time 15L alcoholic solution extracts 1 hour), filter, merge extractive liquid, be concentrated into leave standstill after 4L 1.0 hours centrifugal.Get supernatant to add in the macroporous resin column handled well and carry out saturated adsorption, eluting is carried out successively with water, 45% ethanol, 65% ethanol, the flow velocity of adsorbing and extracting liquid be 0.5 retention volume/hour, flow velocity during desorbing be 2 retention volumes/hour, discard water lotion, collect 65% alcoholic solution eluent, be condensed into thick paste, drying obtains the dry powder 0.31kg that Flavonoids from Eucommia ulmoides Leaves content is 80%, yield is 4.96%.
The preparation of Cortex Eucommiae iridoid: Cortex Eucommiae is dried, pulverizes, sieved is good with Cortex Eucommiae Cortex Dictamni.Take the Cortex Eucommiae after process 20 kilograms, adopt conventional lixiviate, add 70% with the liquid-solid ratio of 15 times.Ethanol-water solution is extractant, extracts 2 hours, repeat 3 times under 60 DEG C of conditions.By extracting liquid filtering, dense institute, be adjusted to pH2.5 with hydrochloric acid; Adopt n-butanol extraction, collect extract, concentrated solution is cooled to-10 ~-20 DEG C, be placed in freeze dryer again and be cooled to-45 ~-65 DEG C, vacuum <20Pa, time 24 ~ 48 low-temperature vacuum drying, obtains iridoid active site 675.3 grams, and total iridoid compounds content is 78.5%.
The preparation of Cortex Eucommiae lignanoid: get eucommia bark (removing crust) 2Kg, with 8L65% alcohol reflux twice, each lh, filters, and merges extracted twice liquid and is concentrated into 800ml.Mix sample loading (macroporous resin column volume 3L) with D101 macroporous resin, wash 3 times of column volumes, 1% ammonia continues to wash 3 times of column volumes, then is washed to neutrality.With 20% ethanol elution, 2 times of column volumes, wash 2.5 times of column volumes with 45% ethanol.45% ethanol elution is concentrated into 600ml.Powder is dried to after eluent is concentrated.With pinoresinol diglucoside in contrast, extract is 85% through ultraviolet detection lignans content, and detecting pinoresinol diglucoside content through HPLC is 12.5%.
The present invention also provides Cortex Eucommiae components composition through the pharmaceutical preparation of the applicable any unit dosage form taken of processing preparation, these pharmaceutical preparatioies are selected from following dosage form: tablet, sugar coated tablet, film coated tablet, enteric coated tablet, capsule, hard capsule, soft capsule, oral liquid, granule, electuary, pill, powder, powder, solution, suppository, ointment, plaster, cream, spray, drop, patch, can make slow releasing preparation, enteric coated preparation when needing.
Cortex Eucommiae components composition of the present invention, medicine acceptable carrier can be added when being prepared into pharmaceutical preparation, described medicine acceptable carrier is selected from: antioxidant, chelating agen, surfactant, filler, disintegrating agent, wetting agent, dispersant, lubricant, rectify and hide agent, pigment etc., conventional carrier is as mannitol, dextran, lactose, glucose, sorbitol, mannitol, xylitol, oxidation is received, silicon derivative, cellulose and cellulose derivative, sodium sulphite, pyrosulfurous acid is received, alginate, gelatin, glycerol, Tween 80, agar, calcium carbonate, calcium bicarbonate, Polyethylene Glycol, cyclodextrin, cyclodextrin, phospholipid material, Kaolin, Pulvis Talci, calcium stearate, stearic acid steamed bun etc.
Detailed description of the invention
Below will further by for example bright the present invention.It is pointed out that following explanation is only illustrating the technical scheme that application claims is protected, any restriction not to these technical schemes.The content that protection scope of the present invention is recorded with appended claims is as the criterion.
The preparation of each effective site of embodiment 1 Cortex Eucommiae
The preparation of eucommiae total flavones: take Folium Eucommiae coarse powder 5.0kg, the ethanol 40L of preparation 45%, pH is adjusted to be 4 with concentrated hydrochloric acid, at 60 ~ 70 DEG C, carrying out stirring extraction twice, (first time 25L alcoholic solution extracts 2 hours, second time 15L alcoholic solution extracts 1 hour), filter, merge extractive liquid, be concentrated into leave standstill after 4L 1.0 hours centrifugal.Get supernatant to add in the macroporous resin column handled well and carry out saturated adsorption, eluting is carried out successively with water, 45% ethanol, 65% ethanol, the flow velocity of adsorbing and extracting liquid be 0.5 retention volume/hour, flow velocity during desorbing be 2 retention volumes/hour, discard water lotion, collect 65% alcoholic solution eluent, be condensed into thick paste, drying obtains the dry powder 0.34kg that Flavonoids from Eucommia ulmoides Leaves content is 87%.
The preparation of Cortex Eucommiae iridoid: Cortex Eucommiae is dried, pulverizes, sieved is good with Cortex Eucommiae Cortex Dictamni.Take the Cortex Eucommiae after process 20 kilograms, adopt conventional lixiviate, add 70% with the liquid-solid ratio of 15 times.Ethanol-water solution is extractant, extracts 2 hours, repeat 3 times under 60 DEG C of conditions.By extracting liquid filtering, dense institute, be adjusted to pH2.5 with hydrochloric acid; Adopt n-butanol extraction, collect extract, concentrated solution is cooled to-10 ~-20 DEG C, be placed in freeze dryer again and be cooled to-45 ~-65 DEG C, vacuum <20Pa, time 24 ~ 48 low-temperature vacuum drying, obtains iridoid active site 675.3 grams, and total iridoid compounds content is 78.5%.
The preparation of Cortex Eucommiae lignanoid: get eucommia bark (removing crust) 2Kg, with 8L65% alcohol reflux twice, each lh, filters, and merges extracted twice liquid and is concentrated into 800ml.Mix sample loading (macroporous resin column volume 3L) with D101 macroporous resin, wash 3 times of column volumes, 1% ammonia continues to wash 3 times of column volumes, then is washed to neutrality.With 20% ethanol elution, 2 times of column volumes, wash 2.5 times of column volumes with 45% ethanol.45% ethanol elution is concentrated into 600ml.Powder is dried to after eluent is concentrated.With pinoresinol diglucoside in contrast, extract is 87.3% through ultraviolet detection lignans content.
The experimentation of embodiment 2 Cortex Eucommiae components composition
Copying and medication of animal cardiac muscle ischemia model
Male SD rat, body weight 220 ± 50g, often organizes 12.Sham operated rats: the aqueous solution gavaging equivalent; Model group: the aqueous solution gavaging equivalent; Each treated animal all gives test medicine (5ml/kg) by body weight gavage.Every day 1 time, altogether administration 7 times.After last administration 30 minutes, 1% pentobarbital sodium (50mg/kg) intraperitoneal injection of anesthesia, continued to monitor ECG Change.ECG electrode pin is embedded in left lower extremity and right upper extremity is subcutaneous, the great cardiac vein of accompanying row with left coronary artery is found between left auricle and pulmonary conus, 2mm place ophthalmology pin 6-0 silk thread threading below left auricle, depth of needle is about 1-1.5mm, wide about 2-3mm, together with great cardiac vein following coronary artery occlusion left anterior descending branch (a sham operated rats threading not ligation).Electrocardiogram display ST section significantly raise or abnormality Q wave time; Namely acute myocardial ischemia modeling success is shown.
The mensuration of myocardial ischemia infarct size percentage ratio: postoperative 24h, postanesthetic rat is opened thoracic cavity, take off rapidly heart and left ventricle is cut into the 4-5 sheet that thickness is 2mm below ligature, put into 1%TTC solution, dye 15min under 37 DEG C of water-baths, lucifuge condition, infarcted region not painted (white), redness is dyed in non-infarcted region, according to formula 1 Image-J (NIH) computed in software Infarct area percentage ratio.Myocardial infarction area percentage ratio (S) %=Infarct area/left ventricle gross area × 100%.
Left room hemodynamic index measures: postoperative 24h, again rats by intraperitoneal injection 1% pentobarbital sodium (50mg/kg) is anaesthetized, then intravenous injection heparin whole body anticoagulant, operation is applied physiograph and is detected cardiac hemodynamic parameter after being separated right common carotid artery.Intubate, to left ventricular recording pressure curve, records left room maximum collapse (+) and diastole (-) speed (± dp/dtmax).
The mensuration of serum myocardial enzyme level: postoperative 24h, opens abdominal cavity by postanesthetic rat, from abdominal aortic blood 5 ~ 8ml, places 1 ~ 2h, centrifugal 10min (3000rpm), measures the level of Serum LDH and CK-MB.
The dosage of each administration group is 100mg/kg, and each administration group is composed as follows:
The eucommiae total flavones of 30%, the Cortex Eucommiae iridoid of 50% and the Cortex Eucommiae lignanoid composition of 20%; The percentage by weight of described compositions also can be the eucommiae total flavones of 25%, the Cortex Eucommiae iridoid of 45% and the Cortex Eucommiae lignanoid composition of 30%.
Concrete outcome is as follows:
Myocardial ischemia infarct size:
| Grouping | Infarct size (%) |
| Sham operated rats | 0 |
| Model group | 39.4±6.2 |
| Administration group 1 | 29.7±5.8 |
| Administration group 2 | 31.5±5.8 * |
| Administration group 3 | 26.8±6.1 * |
| Administration group 4 | 29.7±5.3 * |
| Administration group 5 | 22.5±5.2 ** |
| Administration group 6 | 23.1±4.8 ** |
| Administration group 7 | 11.3±3.3 **▲ |
| Administration group 8 | 10.6±3.8 **▲ |
| Administration group 9 | 33.5±6.4 |
* or * * be medicine group compared with model group, T checks P<0.05 or 0.01,
▲compared with administration group 1-6, T checks P<0.05 or 0.01.
Left room hemodynamic index:
| Grouping | +dp/dt | －dp/dt |
| Sham operated rats | 4551±389 | 4635±447 |
| Model group | 1953±321 ## | 2054±307 ## |
| Administration group 1 | 2355±337 | 2279±217 |
| Administration group 2 | 2628±287 | 2544±233 |
| Administration group 3 | 2856±209 * | 2942±221 * |
| Administration group 4 | 3235±220 ** | 3289±256 ** |
| Administration group 5 | 3125±233 ** | 3218±211 ** |
| Administration group 6 | 3382±285 ** | 3425±301 ** |
| Administration group 7 | 4087±428 **▲ | 4117±456 **▲ |
| Administration group 8 | 4256±468 **▲ | 4277±481 **▲ |
| Administration group 9 | 2162±324 | 2154±267 |
* or * * be medicine group compared with model group, T checks P<0.05 or 0.01,
▲compared with administration group 1-6, T checks P<0.05 or 0.01.
Serum myocardial enzyme level:
| Grouping | LDH(U/L) | CK-MB(U/L) |
| Sham operated rats | 456±48 | 261±34 |
| Model group | 1375±117 ## | 725±69 ## |
| Administration group 1 | 508±51 | 291±28 |
| Administration group 2 | 518±58 | 304±33 |
| Administration group 3 | 567±52 * | 343±29 * |
| Administration group 4 | 624±56 ** | 395±37 ** |
| Administration group 5 | 647±54 ** | 417±33 ** |
| Administration group 6 | 668±48 ** | 456±41 ** |
| Administration group 7 | 1078±115 **▲ | 647±59 **▲ |
| Administration group 8 | 982±103 **▲ | 656±61 **▲ |
| Administration group 9 | 485±45 | 288±31 |
* or * * be medicine group compared with model group, T checks P<0.05 or 0.01,
▲compared with administration group 1-6, T checks P<0.05 or 0.01.
Embodiment 3
Rat myocardial cell primary culture method: take out the SD neonatal rat 25 in raw 24h, after heart all takes out, Digestive system (0.125% pancreatin+0.05% II Collagenase Type) digests, collecting cell, the 200 order bore diameter stainless steel net filtration removings of the cell suspension of collection are not digested tissue, be inoculated in culture bottle, put cell culture incubator (37 DEG C, 95%O
2+ 5%CO
2) middle cultivation, purification myocardial cell.After 2h, the not yet adherent cell suspension of sucking-off blows even counting, adjustment density to 1 × 10
6implant (every hole 500 μ l) in 24 orifice plates, cell is uniformly distributed, adds Brdu to 0.1mmol/L, 24h changes liquid once, and every 3-4d changes liquid once later.The regularly situation of observation of cell growth under inverted microscope, after cultivating 48h, random packet is tested.
37 DEG C, 5%CO
2cultivate under saturated humidity condition.After primary myocardial cell culture 72h, Normal group and model group are normally cultivated with 500 μ lDMEM culture fluid, and each administration group adds the DMEM culture fluid containing medicinal liquid, after hatching 6h, rinses 2 times with ice-cold PBS, and model group and the every hole of administration group add 500 μ l through 95%N
2-5%O
2saturated Tyrode liquid, then puts into a hermetic container by culture plate, continues to pass to mist (95%N
2-5%O
2) hermetic container after 30min, put into incubator anoxia 8h, simulate at body ischemia model.Normal group adds the Tyrode liquid 500 μ l containing 10g/L glucose after cleaning old culture medium, CO
212h is cultivated in incubator.
It is that 0.5mg/mlMTT serum-free DMEM culture fluid 500 μ l cultivates 4h that every hole adds final concentration, then incline supernatant, adds the DMSO of 500 μ l, jolting 10min, the absorbance (OD570nm) at 570nm place is measured, for quantify cellular survival rate by full-automatic microplate reader.
200 μ l culture supernatant are drawn in the every hole of culture plate after process, illustrates according to LDH detection kit, detect LDH activity.
(2) concrete outcome is as follows:
Cell survival rate (MTT) and LDH activity:
| Grouping | LDH(U/L) | Cytoactive (%) |
| Sham operated rats | 33.5±4.1 | 100 |
| Model group | 105.4±6.7 ## | 55.6±2.9 ## |
| Administration group 1 | 45.4±3.5 | 65.4±3.1 |
| Administration group 2 | 48.6±4.1 | 68.3±2.8 |
| Administration group 3 | 51.3±4.3 | 69.4±3.5 |
| Administration group 4 | 65.4±4.6 | 75.3±3.7 |
| Administration group 5 | 63.2±3.9 | 73.6±3.3 |
| Administration group 6 | 66.4±5.2 | 76.1±3.2 |
| Administration group 7 | 85.4±5.6 | 91.2±3.5 |
| Administration group 8 | 87.2±5.4 | 90.6±3.7 |
| Administration group 9 | 41.5±3.7 | 61.8±3.6 |
Prepared by embodiment 4 tablet
Take 300g Cortex Eucommiae components composition (eucommiae total flavones 30%, Cortex Eucommiae iridoid 50%, Cortex Eucommiae lignanoid 20%), the mixing of 150g lactose, 300g microcrystalline Cellulose and 50g cross-linking sodium carboxymethyl cellulose; The material mixed is put in wet granulator, stirs 3 ~ 5min, has stirred in backward material and added 10% ethanol water wet granular, add after slurry completes, open shears, stir shearing, use pelletizing machine wet method granulate; Dry: wet granular is put into drying equipment, inlet temperature, not higher than 80 DEG C, controls moisture and is less than 2%; Granulate: use pelletizing machine dry method granulate; Total mixed: in the granule that granulate is complete, to add 5g magnesium stearate, mixing; Namely tabletting obtains (the heavy 300mg of sheet).
Content of the present invention merely illustrates some claimed specific embodiments; one of them or more described technical characteristic can be combined with arbitrary one or more technical scheme in technical scheme; these technical schemes obtained through combination also in the application's protection domain, just as these technical schemes obtained through combination in the disclosure of invention concrete record.
Claims (4)
1. one kind has the Cortex Eucommiae components composition of myocardium protecting action; it is characterized in that; the composition of said composition is all from Traditional Chinese medicine eucommia bark, and it is specifically the eucommiae total flavones of 10 ~ 80% by percentage by weight, the Cortex Eucommiae iridoid of 10 ~ 80% and the Cortex Eucommiae lignanoid composition of 10 ~ 80%.
2. compositions according to claim 1, is characterized in that, the percentage by weight of described compositions is preferably the eucommiae total flavones of 20 ~ 40%, the Cortex Eucommiae iridoid of 40 ~ 60% and the Cortex Eucommiae lignanoid composition of 20 ~ 50%.
3. compositions according to claim 2, is characterized in that, the percentage by weight of described compositions is the eucommiae total flavones of 30%, the Cortex Eucommiae iridoid of 50% and the Cortex Eucommiae lignanoid composition of 20%.
4. compositions according to claim 2, is characterized in that, the percentage by weight of described compositions is the eucommiae total flavones of 25%, the Cortex Eucommiae iridoid of 45% and the Cortex Eucommiae lignanoid composition of 30%.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101152234A (en) * | 2007-09-14 | 2008-04-02 | 欧阳冬生 | Application of cortex eucommiae lignans and its extract in against cardiovascular reconstruction |
| CN103494955A (en) * | 2013-09-25 | 2014-01-08 | 广元亿明生物科技有限公司 | Chinese medicine composition for treating cardiovascular and cerebrovascular diseases |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101152234A (en) * | 2007-09-14 | 2008-04-02 | 欧阳冬生 | Application of cortex eucommiae lignans and its extract in against cardiovascular reconstruction |
| CN103494955A (en) * | 2013-09-25 | 2014-01-08 | 广元亿明生物科技有限公司 | Chinese medicine composition for treating cardiovascular and cerebrovascular diseases |
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