CN103288914B - Preparation method of traditional Chinese medicine manyflower tickclove herb extract and application in anti-senile dementia medicaments - Google Patents
Preparation method of traditional Chinese medicine manyflower tickclove herb extract and application in anti-senile dementia medicaments Download PDFInfo
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Abstract
The invention provides a preparation method of a manyflower tickclove herb extract, as well as an active component and an active compound thereof. A pentacyclic triterpenoid type active compound can be obtained by crushing traditional Chinese medicine manyflower tickclove herb, then leaching and separating to obtain an ester layer active component, and further performing HPLC (high performance liquid chromatography) purification. According to the preparation method provided by the invention, PC12 cells are adopted as an effective activity identification system, and the manyflower tickclove herb ethanol extract, as well as the active component and the active compound thereof are found to be capable of inducing neurite outgrowth of the PC12 cells, have the significant activity of imitating a nerve growth factor and be capable of being applied in preparation of medicaments and health care foods for preventing senile dementia and other neurodegenerative diseases. As the sources of the manyflower tickclove herb are wide, and the material is convenient to take, the preparation method provided by the invention can enable the development of the new pharmaceutical application of the manyflower tickclove herb to be possible.
Description
Technical field
The invention belongs to medical art, be specifically related to the preparation method of manyflower tickclover herbal extract and active ingredient and active compound, and its food in the nerve degenerative diseases such as Prevention and Curation senile dementia and the application in medicine.
Background technology
The quick growth of elderly population, make people more be concerned about the healthy state of this colony, it is healthy that senile dementia then has a strong impact on the elderly, one of its four Etiologicals having become grownup's death.Point out in international Alzheimer's disease association 2009 annual report: 2010, the whole world about has 3,600 ten thousand people to suffer from dementia, and will with every speed doubled for 20 years, 6,600 ten thousand people are reached in the year two thousand thirty, and the year two thousand fifty will reach 100,000,000 1,540 ten thousand people, middle and low income National Patient quantity will account for 71% of whole world sum.In 2010 annual reports, the overall society cost that the whole world causes due to senile dementia is approximately 6,040 hundred million dollars, exceedes 1.0% of global GDP.
China's old dementia patients is estimated to exceed 5,000,000, accounts for 1/4 of the total case load in the world; And along with the quickening of China's aging population process, this numeral will be more huge, bring great impact to social stability and development.According to statistics, the sickness rate over-65s of Aged in China dementia is 5%, within more than 70 years old, is 10%, within more than 80 years old, is 30%, by more than 85 years old then up to 40%.After 20 years, middle-aged people of today will step into the ranks of the elderly, and Dementia patients's quantity will sharply increase, and the health of the elderly also will be related to the stable of entire society and development.Therefore, the medicine of researching and developing the nerve degenerative diseases such as effective prophylactic treatment senile dementia has become whole world medical problem in the urgent need to address.
In senile dementia three macrotaxonomy, AD is that sickness rate is the highest, is also most important a kind of dementia form disease.AD is the one group of syndrome caused by many reasons such as nerve retrograde affection, cerebrovascular disease, infection, wound, tumour, Nutrition and Metabolism obstacles, it is the lasting comprehensive hypophrenia that patient occurs under the state of Consciousness, show as memory, computing power, judgement, attention, abstract thinking ability, linguistic function go down, affective disorder, activity of daily living and work capacity Progressive symmetric erythrokeratodermia go down until lose, and have various neuropsychic symptom and behavior disorder.The definite pathomechanism of AD it be unclear that, and main academic viewpoint has following several at present: 1. beta-amyloyd polypeptide (A β) toxicity and deposition; 2. cholinergic deficiency theory; 3. nerve retrograde affection (Neurodegeneration); 4. other many factors, as transgenation theory, oxidative stress theory.
The medicine being used for the treatment of AD is now a lot, mainly contain cholinergic agent, wherein acetylcholinesterase (Acetylcholinesterase, AChE) inhibitor, the medicine of main listing has tacrine (tacrine), rivastigmine (rivastigmine), selagine (huperzine A), E2020 (donepezil) etc.; β, gamma secretase Depressant; Brain metabolism regulators, as vincamine, nimodipine, cinnarizine; Affect the medicine of Radical Metabolism, if vitamins C is in conjunction with vitamin-E etc.But these medicines can only delay the progress of the AD course of disease, and reduce gradually with PD curative effect of medication, have side effect, sight is turned to the research and development of new anti senile dementia drug by old friends, find the medicine for the AD cause of disease and method, become focus and the difficult point of current research.
Research shows, the lysis of neurotrophic factor on neurodevelopment and Adult Nervous System has important impact.In neurodegeneration animal model, find that nerve growth factor (nerve growth factor, NGF) can stop or reduce neuronic regression.NGF is first neurotrophic factor that the mankind find, is also most important neurotrophic factor; It is the biologically active polypeptides that there is important regulating and controlling effect the aspects such as a kind of growth to neurocyte, growth, differentiation and function maintenance; Unusual effect is had to the treatment of the sacred diseases such as neuratorphy, neurodegeneration, wound reparation.Research finds, NGF to a certain degree can stop AD to be in progress, and it promotes that nerve growth and neuroprotective are long-term study hotspots.But it is a polypeptide be made up of more than 100 amino acid; Due to reasons such as molecular weight are large and polarity is strong, not by hemato encephalic barrier (Blood Brain Barrier), and be difficult to the factors such as extensive preparation, limited to its practical clinical, NGF does not also find better methods for the treatment of except operation in brain directly dispensing.Therefore, find and there is similar NGF activity (NGF mimics) maybe can strengthen its activity (NGF enhancer) and low molecular compound by hemato encephalic barrier just becomes study hotspot naturally.Due to PC12 cell (Pheochromocytoma cells, from Clonal Rat Pheochromocytoma, clone obtains), have the general feature of neurocyte and the feature that can go down to posterity, under the effect of NGF, cell can stop division, grow projection, change into neuron cell.Therefore, cellular and molecular level research NGF function PC12 cell be a good model.At present, NGF mimics has been had in the phase iii clinical trial stage.
Herbal medicine is the basic substance of Chinese materia medica, is the cornucopia of natural radioactivity organic compound.There is the traditional Chinese medical science civilization of several thousand in China, and vast territory and abundant resources, has a lot of valuable famous-region drug, because utilizing of Chinese material medicine resource is more convenient, so also just very important to the research of its new composition and new activity.Research shows, Chinese medicine has significant curative effect to senile dementia, wherein foremost is the alkaloid-selagine (huperzine A) extracted from Herba Lycopodii serrati, and it is the original new drug with independent intellectual property right of institute of materia medica of Chinese Academy of Sciences exploitation; It is not only a kind of acetylcholinesterase (AChE) Depressant of efficient highly selective, and can also reduce the nerve cell death that L-glutamic acid brings out; There is the response to oxidative stress that obvious neuroprotective cell antagonism beta-amyloyd polypeptide (A β) produces; Can to the nerve cell apoptosis effect of the induction such as resistive to hydrogen peroxide, inhibitors of protein kinase C; It is one of optimal drug of current global treatment senile dementia.
Herba desmodii multifloi is the complete stool of leguminous plants beggar-ticks plant Herba desmodii multifloi Desmodium tiflorum, is distributed in the ground such as China Guangxi, Guizhou, Yunnan, Fujian.Taste is sweet, bitter, cool in nature, has promoting blood circulation and stopping pain, detoxify and promote the subsdence of swelling function, for diseases such as stomachache, infantile malnutrition, women's emaciation due to blood disorder, lumbar sprain, wound, urethritis, parotitis and venomous snake bites.
Summary of the invention
The object of this invention is to provide the preparation method of manyflower tickclover herbal extract, active ingredient, active compound, realized by following steps:
(1) pulverizing and lixiviate:
After 1000g Chinese medicine Herba desmodii multifloi is pulverized, with lixiviate under 3L ethanol room temperature (shaking once in a while).After suction filtration concentrate drying, obtain ethanol extraction study 50g, used water and ethyl acetate to replace extracting and separating repeatedly, obtain ester layer active ingredient (A) 10g;
(2) abstraction and purification:
Active ingredient (A) is first separated (200-300 order through a silica gel opening column, following solvent systems all by volume, first time is separated into chloroform: ethyl acetate=100:0,90:10,80:20,70:30,50:50,0:10), combined chloroform: ethyl acetate is the eluted fraction of 90:10 to 50:50, obtains active ingredient (B) 1g; Then, active ingredient (B) is separated (solvent systems is methyl alcohol by volume: water=6:4,7:3,8:2,9:1,10:0) by ODS opening column further, merges methyl alcohol: water is the eluted fraction of 7:3 to 9:1, obtains active ingredient (C) 150mg; Active ingredient (C) purifies with HPLC that (chromatographic condition is: chromatographic column
flow velocity 3ml/min, determined wavelength 210nm, moving phase is 80%-85% aqueous methanol gradient wash-out 60 minutes), obtain three compounds, pentacyclic triterpenoid (1) 50mg is defined as, compound (2) 25mg and compound (3) 2.5mg through nuclear magnetic resonance spectrum, mass spectrum.
Active ingredient C prepared by the present invention, the wherein total content >50% of pentacyclic triterpenoid, determine three pentacyclic triterpenoids simultaneously, compound (1) is 3 β, 23, 28-trihydroxy--12-oleanene-3 β-caffeic acid ester (3 β, 23, 28-trihydroxy-12-oleanene-3 β-caffeate), compound (2) is 3 β, 23, 28-trihydroxy--12-oleanene-23-caffeic acid ester (3 β, 23, 28-trihydroxy-12-oleanene-23-caffeate), compound (3) is 3 β, 23, 28-trihydroxy-volatile oil-12-alkene (3 β, 23, 28-triol olean-12-ene).Concrete structure formula is as follows:
Compound (1):
Compound (2):
Compound (3):
Another object of the present invention is to provide the application in the medicine and protective foods of the nerve degenerative diseases such as preparation prevention senile dementia of manyflower tickclover herbal extract and active ingredient and active compound thereof.Described active ingredient is active ingredient (A)-(C), and described active compound is pentacyclic triterpenoid (1)-(3).
Manyflower tickclover herbal extract, active ingredient (A)-(C), active compound (1)-(3) have the significant activity intending nerve growth factor, therefore effective constituent can be it can be used as, add pharmacy or food acceptable carrier or vehicle, the protective foods of the nerve degenerative diseases such as preparation prevention, treatment senile dementia and medicine.
Described protective foods acceptable carrier refers to the carrier of field of food routine, the weighting agents such as such as starch, sucrose, Microcrystalline Cellulose, the tackiness agents such as starch slurry, hydroxypropylcellulose, gelatin, polyoxyethylene glycol, the wetting agents such as Magnesium Stearate, micropowder silica gel, polyethylene glycols, the absorption enhancers such as poly-sorb fat, Yelkin TTS, the tensio-active agents such as smooth, the poly-sorb fat of poloxamer, lipid acid sorb, can also add other assistant agent such as flavouring agent, sweeting agent in addition.
Described pharmaceutically acceptable carrier can refer to prior art.After described activeconstituents or compound are made medicine, can give use in a unit, route of administration can be enteron aisle or parenterai administration.
The formulation of described medicine can be solid preparation, liquid preparation or semisolid dosage form, and above-mentioned various formulation can adopt existing production method to be prepared.
The present invention also provides a kind of pharmaceutical composition preventing the nerve degenerative diseases such as senile dementia further, and this pharmaceutical composition contains the manyflower tickclover herbal extract of physiology significant quantity and active ingredient (A)-(C) thereof and active compound and pharmaceutically acceptable carrier or thinner.
Pharmaceutically acceptable carrier described here refers to the pharmaceutical carrier of pharmaceutical field routine, and in this way etc., weighting agent is as starch, sucrose, Microcrystalline Cellulose etc. for such as thinner, vehicle; Tackiness agent is as starch slurry, hydroxypropylcellulose, gelatin, polyoxyethylene glycol etc.; Wetting agent is as Magnesium Stearate, micropowder silica gel, polyethylene glycols etc.; Absorption enhancer gathers sorb fat, Yelkin TTS etc., and smooth, poly-sorb fat of tensio-active agent poloxamer, lipid acid sorb etc., can also add other assistant agent in addition in the composition as flavouring agent, sweeting agent etc.
Manyflower tickclover herbal extract of the present invention and active ingredient (A)-(C) thereof and active compound (1)-(3) can administrations in a unit, and route of administration can be enteron aisle and non-bowel, comprises oral, muscle, subcutaneous and nasal cavity.
Compound administration approach of the present invention can be intravenously administrable.Injection comprises intravenous injection, intramuscular injection, subcutaneous injection and acupoint injection therapy.
The various formulations of pharmaceutical composition of the present invention can be prepared according to the conventional production process of pharmaceutical field, such as, make activeconstituents and one or more carriers or mixed with excipients, be then made into required formulation.
Form of administration can be solid preparation, capsule or liquid preparation, comprises tablet, capsule, dispersible tablet, oral liquid, infusion solutions, little pin, freeze-dried powder, ointment, liniment or suppository.
The manyflower tickclover herbal extract prepared with the present invention and active ingredient (A)-(C) thereof and active compound (1)-(3), there is the significant activity intending nerve growth factor, medicine and the protective foods of the nerve degenerative diseases such as prevention senile dementia can be prepared.Chinese medicinal materials Herba desmodii multifloi in the present invention is cheap and easy to get, and has without it relevant report intending Nerve Growth Factor Activity so far.Therefore, the present invention is to the new pharmacology activity research of this Chinese medicine and develop significant.
The present invention adopts PC 12 cell as effective activity identification system, finds Herba desmodii multifloi ethanol extraction and active ingredient thereof and active compound, PC 12 cellular neural projection can be induced to extend, have and intend Nerve Growth Factor Activity significantly.In addition, Herba desmodii multifloi wide material sources, the advantages such as facility of drawing materials.So Herba desmodii multifloi is developed as the new drug of the nerve degenerative diseases such as prevention and therapy senile dementia and protective foods significant.
Accompanying drawing explanation
Fig. 1 adds manyflower tickclover herbal extract and active ingredient (A)-(C) thereof, the nervous process differentiation rate of PC 12 cell after 48 hours.
Fig. 2 adds pentacyclic triterpenoid, PC 12 cellular neural projection differentiation after 48 hours.
Fig. 3 adds manyflower tickclover herbal extract and active ingredient thereof and active compound, PC 12 microcytoscope picture after 48 hours.
Embodiment
The present invention is further described with specific embodiment by reference to the accompanying drawings.
The preparation of embodiment 1 manyflower tickclover herbal extract and active ingredient and active compound
1. pulverize and lixiviate:
After 1000g Chinese medicine Herba desmodii multifloi is pulverized, with lixiviate under 3L ethanol room temperature (shaking once in a while).After suction filtration concentrate drying, obtain ethanol extraction study 50g, used water and ethyl acetate to replace extracting and separating repeatedly, obtain ester layer active ingredient (A) 10g.
2. abstraction and purification:
Active ingredient (A) is first separated (200-300 order through a silica gel opening column, following solvent systems all by volume, first time is separated into chloroform: ethyl acetate=100:0,90:10,80:20,70:30,50:50,0:10), combined chloroform: ethyl acetate is the eluted fraction of 90:10 to 50:50, obtains active ingredient (B) 1g; Then, active ingredient is separated (solvent systems is methyl alcohol by volume: water=6:4,7:3,8:2,9:1,10:0) by ODS opening column further, merges methyl alcohol: water is the eluted fraction of 7:3 to 9:1, obtains active ingredient (C) 150mg; Active ingredient (C) purifies with HPLC that (chromatographic condition is: chromatographic column
flow velocity 3ml/min, determined wavelength 210nm, moving phase is 80%-85% aqueous methanol gradient wash-out 60 minutes), obtain pentacyclic triterpenoid (1) 50mg, pentacyclic triterpenoid (2) 25mg and pentacyclic triterpenoid (3) 2.5mg.
Embodiment 2
Qualitative Identification to embodiment 1 gained compound physicochemical characteristics and chemical structure:
Compound (1), the structure of (2) and (3) through MS,
1h NMR,
13c NMR tests, and determines after contrasting with data in literature.
Compound 1: white solid;
(c1.0, MeOH); C
39h
56o
6(HR ESI-TOF-MS m/z (M+Na)
+calcd.for C
39h
56o
6na:643.3969, Found:643.3981);
1h NMR (500MHz, CD
3oD) δ 7.51 (1H, d, J=15.8Hz), 7.03 (1H, d, J=2.0Hz), 6.94 (1H, dd, J=8.2,2.0Hz), 6.78 (1H, d, J=8.2Hz), 6.23 (1H, d, J=15.8Hz), 5.20 (1H, brt), 4.97 (1H, dd, J=9.1,7.6Hz), 3.52 (2H, d, J=10.0Hz), 3.35 (d, 10Hz, 1H), 3.16 (1H, d, J=12.7Hz), 3.11 (1H, d, J=11.8Hz), 1.23 (3H, s), 1.07 (3H, s), (1.02 3H, s), 0.89 (6H, s), 0.83 (3H, s).
13C NMR(125MHz,CD
3OD)δ169.2,149.5,146.7,146.6,145.8,127.7,123.4,122.9,116.5,115.6,115.1,75.8,69.8,64.7,49.1,47.9,47.8,43.8,43,43,41,39.2,38.1,37.7,35.3,33.7,33.1,32.3,31.8,26.6,26.6,24.6,24.1,24,22.9,18.8,17.3,16.5,13.9。
Compound 2: white solid;
(c1.0, MeOH); It is C that high resolution mass spectrum demonstrates its molecular formula
39h
56o
6(HR ESI-TOF-MS m/z (M+Na)
+calcd.for C
39h
56o
6na:643.3969, Found:643.3972);
1h NMR (500MHz, CD
3oD) δ 7.57 (1H, d, J=15.9Hz), 7.04 (1H, brs), 6.94 (1H, dd, J=8.2), 6.79 (1H, d, J=8.2Hz), 6.27 (1H, d, J=15.9Hz), 5.19 (1H, brt), 4.13 (1H, d, 11.3Hz), 4.03 (1H, d, 11.3Hz), 3.63 (1H, d, 11.1Hz, 6.3Hz), 3.50 (1H, d, J=12Hz), 3.09 (1H, d, J=11.0Hz), 1.13 (3H, s), 1.02 (3H, s), (1.00 3H, s), 0.88 (6H, s), 0.79 (3H, s).
13C NMR(125MHz,CD
3OD)δ169.13,149.74,146.99,146.92,145.82,127.69,123.48,123.07,116.58,115.15,115.15,72,69.7,66.5,49.39,49.05,47.7,43.86,43.31,42.85,41.12,39.8,38.1,37.94,35.28,33.78,33.52,32.32,31.89,27.47,26.6,26.54,24.6,24,22.8,19.2,17.3,16.4,12.82。
The physico-chemical property of compound 3: white solid;
+ 52 ° of (c1.0, CHCl
3); Molecular formula is C
30h
52o
3;
1hNMR (500MHz, CDCl
3) δ 5.18 (brt, 1H), 3.67 (m, 1H), 3.73 (d, J=10.3Hz, 1H), 3.44 (d, J=10.3Hz, 1H), 1.15 (s, 3H), 3.55 (d, 10.9Hz, 1H), 3.20 (d, 11.0Hz, 1H), 0.97 (s, 3H), 0.93 (s, 3H), 0.90 (s, 3H), 0.87 (s, 3H), 0.88 (s, 3H).
Embodiment 3
The biological activity of Chinese medicine manyflower tickclover herbal extract and active ingredient and active compound.
In nerve retrograde affection animal model, research finds that NGF can stop or reduce neuronic regression, and AD to a certain degree can be stoped to be in progress, and has and promotes nerve growth and neuroprotective.Because PC12 cell has the general feature of neurocyte, under the effect of NGF, PC12 cell can stop division, grows projection, changes into neuron cell.Therefore, adopt PC 12 cell as effective activity identification system, the activated effective constituent of screening tool, will the active drug for the treatment of senile dementia be become.
Experimental technique:
The cultivation of 1.PC 12 cell (adult rat adrenal tissue is addicted to chromium tumor cell strain): connect 20 × 10
4individual PC 12 cell, in the culture dish of 100mm, containing 10mL DMEM substratum (wherein containing 10% horse serum, 5% foetal calf serum), changes a subculture, after three days subcultures two days later.First use PBS(phosphate buffered saline buffer) cell is washed twice, then add 10mL PBS in culture dish, and at 37 DEG C, 5%CO
2incubator in cultivate 10 minutes, purge, transfers to the disposable centrifuge tube of 15mL, and centrifugal rear blood counting chamber counts.The 24 every holes of porocyte culture plate first add the DMEM substratum of 1mL containing serum, and after cell counting, every hole connects 2 × 10
4individual cell, CO
2application of sample after 24 hours cultivated by incubator.
2. active testing: with DMSO(dimethyl sulfoxide (DMSO)) be negative control, NGF 40ng is positive control, and by manyflower tickclover herbal extract, active ingredient (A)-(C) and compound (1)-(3) are configured to the DMSO solution of different concns.Contain the DMEM solution (not containing serum) of 1%DMSO and sample by after former for every hole of 24 porocyte plates substratum replacement with 1mL, put into 37 DEG C, 5%CO
2incubator in cultivate.Every 24 hours, continuous 6 days observation of cell metamorphosis under inverted microscope, the nervous process differentiation rate of the record cell ratio of cell number (under the nervous process cell number of being longer than cell space diameter one times and the visual field always), about 100 cells under each visual field, random selecting 3 place, and add up mapping analysis.
3. experimental result:
Under finite concentration, add manyflower tickclover herbal extract, ester layer active ingredient (A), active ingredient (B), active ingredient (C) and latter 48 hours of compound (1)-(3), manyflower tickclover herbal extract, ester layer active ingredient (A), active ingredient (B), active ingredient (C) and compound (1)-(3) all have the effect promoting that PC 12 cell process extends, and demonstrate and intend Nerve Growth Factor Activity significantly.See Fig. 1-3, the DMSO using 1% is as negative control, and the biological activity of sample activity component (B) and (C) and compound (1) and compound (2) is particularly outstanding.In Fig. 1: 1%DMSO is negative control; NGF:40ng/mL, positive control; Concentration unit: μ g/mL; * * P<0.001.
In Fig. 2, C:1%DMSO is negative control; NGF:40ng/mL, positive control; Concentration unit: μ Μ; * P<0.05, * * * P<0.001.Fig. 3 adds manyflower tickclover herbal extract and active ingredient thereof and active compound, PC 12 microcytoscope picture after 48 hours.The effect all significantly inducing PC 12 cell generation nervous process to extend in manyflower tickclover herbal extract, ester layer active ingredient (A), active ingredient (B), active ingredient (C) and compound (1)-(3) can be observed from Fig. 3.In Fig. 3: a, negative control is 1%DMSO; B, NGF 40ng/mL is positive control; C, manyflower tickclover herbal extract (100 μ g/mL); D, ester layer active ingredient (A) (10 μ g/mL); E, active ingredient (B) (10 μ g/mL); F, active ingredient (C) (1 μ g/mL); G, compound (1) (1 μM); F, compound (2) (1 μM).
Claims (2)
1. a Herba desmodii multifloi active ingredient (
c) preparing the application in prevention of neurodegenerative diseases medicine or protective foods, it is characterized in that, described nerve degenerative diseases is senile dementia; Described Herba desmodii multifloi active ingredient (
c) obtain by the following method:
(1) pulverize and lixiviate: 1000 g Chinese medicine Herba desmodii multiflois pulverize after, with lixiviate under 3 L ethanol room temperatures, after suction filtration concentrate drying, obtain ethanol extraction study 50 g, used water and ethyl acetate to replace extracting and separating repeatedly, obtain ester layer active ingredient (
a) 10 g;
(2) abstraction and purification: by active ingredient (
a) be first separated through a silica gel opening column, solvent systems is chloroform by volume: ethyl acetate=100:0,90:10,80:20,70:30,50:50,0:10, combined chloroform: the eluted fraction of ethyl acetate from 90:10 to 50:50, obtain active ingredient (
b), then, by active ingredient (
b) be separated by ODS opening column further, solvent systems is methyl alcohol by volume: water=6:4,7:3,8:2,9:1,10:0, merges methyl alcohol: the eluted fraction of water from 7:3 to 9:1, obtain active ingredient (
c).
2. application according to claim 1, is characterized in that, described medicine or protective foods by described active ingredient (
c) make with pharmacy or food acceptable carrier, the formulation of described medicine or protective foods is solid preparation, liquid preparation or semi-solid preparation.
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| Two Bioactive Pentacyclic Triterpene Esters from the Root Bark of Hibiscus syriacus;Bong-Sik Yun et al.;《Journal of Natural Products》;19990403;第62卷(第5期);第764页第1段至第766页倒数第1段 * |
| 李传宽等.饿蚂蝗化学成分研究.《中国中药杂志》.2010,第35卷(第18期),第2420页第1段至第2423页倒数第1段. * |
| 饿蚂蝗化学成分研究;李传宽等;《中国中药杂志》;20100930;第35卷(第18期);第2420页第1段至第2423页倒数第1段 * |
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