CN1981809B - Danshen effective component, its preparation, making method and usage - Google Patents
Danshen effective component, its preparation, making method and usage Download PDFInfo
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- CN1981809B CN1981809B CN200510122346XA CN200510122346A CN1981809B CN 1981809 B CN1981809 B CN 1981809B CN 200510122346X A CN200510122346X A CN 200510122346XA CN 200510122346 A CN200510122346 A CN 200510122346A CN 1981809 B CN1981809 B CN 1981809B
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- 239000012588 trypsin Substances 0.000 description 1
- 210000003462 vein Anatomy 0.000 description 1
- 230000002861 ventricular Effects 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
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Abstract
An active component of red sage root is proportionally composed of alkannic acid, danshinolic acid B and danshinolic acid E. Its preparing process includes such steps as breaking red sage root, adding ethyl acetate and alcohol, thermal extracting, concentrating to obtain extract, mixing it with silicon gel, separating with positive-phase silicon gel column, washing with petroether and ethyl acetate, eluting with chloroform and methanol, concentrating the eluting liquid, drying, and separating by liquid-phase chromatography. Its application is also disclosed.
Description
Technical field
The present invention relates to a kind of Chinese medicine extract for the treatment of cardiovascular disease, relate in particular to the active component that from Radix Salviae Miltiorrhizae, extracts, preparation and preparation method thereof and purposes, this active component mainly contains alkannic acid (Lithospermic acid), salvianolic acid B (Salvianolic acid B) and three kinds of chemical compounds of salvianolic acid E (Salvianolic acid E).
Background technology
Coronary heart disease and angina pectoris are harm humans health " first killers ", in recent years, along with the variation of China's population senescence and people's work, life, dietary structure and environment etc., the incidence rate of cardiovascular and cerebrovascular diseases such as coronary heart disease also increases year by year, and the people's physical and mental health in serious threat.In the natural product many active substances have resist myocardial ischemia, anoxia functions, some of them have been developed to treatment coronary heart disease and anginal new drug, as treat the medicine DIAOXINXUE KANG of coronary heart disease, it is by 8 kinds of pure Chinese medicinal preparations that steroid saponin is made that extract from Chinese medicine; Flavonihippophae is to be that raw material extracts the pure natural medical that processes with the Fructus Hippophae, and its effective ingredient is a Fructus Hippophae total flavones.Thereby from natural product, seek have resist myocardial ischemia, the active substance of anoxia physiologically active, be find, one of effective way of developing new drug.China's medicinal organism resource is very abundant, and its biological active substances is research and finds new drug guide chemicals, the natural treasure-house of developing new drug.At present, China extracts active substance from natural product, be used to be developed to treatment coronary heart disease, safety is good, toxicity is low new drug also seldom, from natural product, extract active substance, be developed to have and resist myocardial ischemia, be used for the treatment of coronary heart disease and anginal new drug, have significant application value and wide development prospect.
Radix Salviae Miltiorrhizae, another name blood is taken root, red, Herba Wedeliae Wallichii, Arisaema balansae Engl..Radix Salviae Miltiorrhizae is from plant Salvia miltiorrhiza rhizome.Radix Salviae Miltiorrhizae is used as medicine and begins to be stated from the Shennong's Herbal of Han dynasty.The traditional Chinese medical science is thought, its bitter in the mouth cold nature has promoting blood circulation to restore menstrual flow, an effect of the silt pain relieving of dispelling, the relieving restlessness that clears away heart-fire, removing heat from blood eliminating carbuncle, is applicable to the treatment menoxenia, coronary heart disease, arteriosclerosis, hyperlipidemia, angina pectoris, dysmenorrhea, amenorrhea, metrorrhagia, leukorrhagia, traumatic injury blood stasis, insomnia, hepatitis B, neurasthenia, the schistosomicide hepatosplenomegaly, diseases such as arthralgia, particularly gynecological, internal medicine and external wounds card belongs to the blood stasis hot person that holds concurrently and uses always.Modern study shows, mainly contain Tanshinone I, IIA, IIB (Tanshinone I in the Radix Salviae Miltiorrhizae, IIA, IIB), iso tanshinone I, IIA (Isotanshinone I, IIA), cryptotanshinone (Cryptotanshinone), different cryptotanshinone (Isocryptotanshinone), methyltanshinone (Methyltanshinone), hydroxyl TANSHINONES etc.
Radix Salviae Miltiorrhizae is the common drug of treatment cardiovascular disease, and in recent decades, research institution has carried out a large amount of research to Radix Salviae Miltiorrhizae both at home and abroad, and the various patent medicine that contain red rooted salvia are developed diseases such as being used for the treatment of coronary heart disease, angina pectoris, cerebral infarction.Radix Salviae Miltiorrhizae has the influence of cardiovascular system: (1) increases coronary flow.But composition such as Radix Salviae Miltiorrhizae, element coronary blood flow increasing, promote collateral circulation, improve myocardium microcirculation, do not increase ventricular stroke work and myocardial oxygen consumption simultaneously.(2) microcirculation improvement.Radix Salviae Miltiorrhizae has the effective ingredient of microcirculation improvement.In addition, effective ingredient can influence multiple thrombin, improves hemorheological property, can reduce plasma viscosity, regulate cell electrophoresis rate and packed cell volume patients with coronary heart disease in the Radix Salviae Miltiorrhizae, microcirculation improvement has therapeutical effect preferably to blood stasis patient's blood " sticking, poly-, stagnate " tendency.Studies show that Radix Salviae Miltiorrhizae can anticoagulant and antithrombotic effect.Just because of Radix Salviae Miltiorrhizae has above pharmacological action to cardiovascular system, therefore, from Radix Salviae Miltiorrhizae, isolate active component, the modern Chinese medicine of developing the treatment cardiovascular disease has great importance.
Summary of the invention
The object of the present invention is to provide a kind of Danshen effective component.
Another object of the present invention is to provide the preparation method of this Danshen effective component.
The present invention also provides the preparation that contains this Danshen effective component and the purposes of this component.
The invention provides following technical scheme:
Danshen effective component of the present invention mainly contains alkannic acid and salvianolic acid.Wherein, by weight percentage, the content of alkannic acid is 2.8～4.2%, and the content of salvianolic acid B is 91.8～94.2%, and the content of salvianolic acid E is 0.8～2.2%.
Preferably, in the Danshen effective component of the present invention, the content of alkannic acid is 3～4%, and the content of salvianolic acid B is 92～94%, and the content of salvianolic acid E is 1～2%.
Best, in the Danshen effective component of the present invention, the content of alkannic acid is 3.2～3.8%, and the content of salvianolic acid B is 92.5～93.5%, and the content of salvianolic acid E is 1.2～1.8%.
The preparation method of Danshen effective component of the present invention, comprise the following steps: that red rooted salvia is pulverized the back adds ethyl acetate and ethanol, heating extraction gets extracting solution, and extracting solution is condensed into extractum, and itself and silica gel mixed sample, with normal phase silicagel column it is separated, at first use petroleum ether and ethyl acetate, get eluent I as mobile phase, abandon it, change chloroform and methanol then as mobile phase, get eluent II, will get sample behind the eluent II concentrate drying; Continue to separate the sample that obtains with preparative liquid chromatography; The separation condition of preparative hplc: chromatographic column is a semi-preparative column, and mobile phase is water and acetonitrile, gradient elution.
Preferably, the preparation method of Danshen effective component of the present invention, comprise the following steps: that red rooted salvia is pulverized the back adds ethyl acetate and ethanol (1: 0.8～1.2), reflux 0.8～1.2 hour, extract 1～3 time, merging filtrate gets extracting solution, extracting solution is condensed into extractum, and itself and silica gel are mixed sample, it is separated with normal phase silicagel column, at first use petroleum ether and ethyl acetate (47～53: 1) as mobile phase, get eluent I, abandon it, change chloroform and methanol (8～13: then 1) as mobile phase, get eluent II, will get sample behind the eluent II concentrate drying; Continue to separate the sample that obtains with preparative liquid chromatography; The separation condition of preparative hplc: chromatographic column is a semi-preparative column, and mobile phase is water and acetonitrile, and gradient elution, flow velocity are 2.8～3.2ml/min, and column temperature is a room temperature.
Best, Danshen effective component preparation method of the present invention comprises the following steps: to get red rooted salvia, it is pulverized the back add ethyl acetate and ethanol (1: 1), and reflux 1 hour is extracted 2 times, filtrate merge extracting solution I.Medicinal residues add 70% ethanol, reflux 1 hour is extracted 2 times, filtrate merge extracting solution II, II is condensed into extractum with extracting solution, with 50% dissolve with methanol extractum, adopt the ODS-C18 post that it is separated, at first use 5% methanol as mobile phase, get eluent I, change 60% methanol then as mobile phase, get eluent II, get sample behind the concentrate drying; Continue to separate the sample that obtains with preparative liquid chromatography: the separation condition of preparative hplc: chromatographic column is semi-preparative column (Lichrospher C18; 10.0mm * 250mm, 5 μ m), mobile phase is water (A) and acetonitrile (B), the gradient elution program is as follows: during 0min, mobile phase A is water+0.1% acetum of 85%, and Mobile phase B is 15% acetonitrile solution; During 10in, mobile phase A is water+0.1% acetum of 79%, and Mobile phase B is 21% acetonitrile solution; During 15min, mobile phase A is water+0.1% acetum of 69%, and Mobile phase B is 31% acetonitrile solution; During 30min, mobile phase A is water+0.1% acetum of 63%, and Mobile phase B is 37% acetonitrile solution; During 35min, mobile phase A is water+0.1% acetum of 15%, and Mobile phase B is 85% acetonitrile solution; During 45min, mobile phase A is water+0.1% acetum of 15%, and Mobile phase B is 85% acetonitrile solution; Flow velocity is 3ml/min, and column temperature is a room temperature; Sample 100% dissolve with ethanol separates through preparative liquid chromatography, collects solution at time period 17.5～23.6min, and solution obtains active component behind concentrate drying.
Danshen effective component of the present invention can be used as active component, adds the adjuvant of accepting on the pharmaceutics, makes preparation according to the preparation method of the preparation of putting down in writing on the pharmaceutics.
Danshen effective component of the present invention can also add the adjuvant of accepting on the pharmaceutics as one of active component, makes preparation according to the preparation method of the preparation of putting down in writing on the pharmaceutics.
Described preparation comprises injection, drip liquid, injectable powder, granule, tablet, electuary, powder, oral liquid, sugar coated tablet, film coated tablet, enteric coated tablet, capsule, hard capsule, soft capsule, sucks agent, granule, pill, unguentum, sublimed preparation, spray, drop pill, disintegrating agent, oral cavity disintegration tablet, micropill etc.
Danshen effective component of the present invention and preparation thereof can be used in preparation treatment, angiocardiopathy preventing medicine.
Beneficial effect of the present invention is:
1. use normal phase silicagel column in the extraction and separation process of the present invention, can remove impurity such as desaccharide, protein, aminoacid effectively, improved content of effective, adopted preparative hplc simultaneously, can obtain effective ingredient fast and accurately.
2. Danshen effective component chemical constituent provided by the invention is simply clear and definite, is easier to illustrate its mechanism of action on pharmacological research, is easier to the quality control of medicine aborning.
Description of drawings
Fig. 1 is the HPLC analysis chart of Danshen effective component of the present invention.
The specific embodiment
Further describe flesh and blood of the present invention and beneficial effect below in conjunction with embodiment, this embodiment only is used to the present invention is described but not limitation of the present invention.
The preparation of embodiment one Danshen effective component
The 200g red rooted salvia is pulverized the back add ethyl acetate and ethanol, heating extraction gets extracting solution I, abandons it.Medicinal residues add ethanol, heating extraction gets extracting solution II, extracting solution II is condensed into extractum 102.4g, use dissolve with methanol extractum, adopt the ODS-C18 post that it is separated, at first use low concentration methanol as mobile phase, get eluent I, change high concentration methanol then as mobile phase, get eluent II, get sample 10.24g behind the concentrate drying; Continue to separate the sample that obtains with preparative liquid chromatography; The separation condition of preparative hplc: chromatographic column is a semi-preparative column, and mobile phase is water and acetonitrile, gradient elution, sample 100% dissolve with ethanol, separate through preparative liquid chromatography, collect solution, obtain active component 4.22g behind the concentrate drying at time period 17.5～23.6min.
The preparation of embodiment two Danshen effective components
The 500g red rooted salvia is pulverized the back add ethyl acetate and ethanol (1: 0.8～1.2), reflux 0.8～1.2 hour is extracted 1～3 time, and merging filtrate gets extracting solution I, abandons it.In medicinal residues, add 60%～30% ethanol, reflux 0.8～1.2 hour is extracted 1～3 time, filtrate merge extracting solution II, extracting solution II is condensed into extractum 255.86g, with 40%～60% dissolve with methanol extractum, adopt the ODS-C18 post that it is separated, at first use 3%～8% methanol as mobile phase, get eluent I, change 50%～70% methanol then as mobile phase, get eluent II, get sample 25.31g behind the concentrate drying; Continue to separate the sample that obtains with preparative liquid chromatography; The separation condition of preparative hplc: chromatographic column is a semi-preparative column, and mobile phase is water and acetonitrile, gradient elution, flow velocity is 2.8～3.2ml/min, column temperature is a room temperature, collects solution at time period 17.5～23.6min, and solution obtains active component 10.54g behind concentrate drying.
The preparation of embodiment three Danshen effective components
Get red rooted salvia 250g, it is pulverized the back add ethyl acetate and ethanol (1: 1), reflux 1 hour is extracted 2 times, filtrate merge extracting solution I.Medicinal residues add 70% ethanol, reflux 1 hour is extracted 2 times, filtrate merge extracting solution II, extracting solution II is condensed into extractum gets 138.3g, with 50% dissolve with methanol extractum, adopt the ODS-C18 post that it is separated, at first use 5% methanol as mobile phase, get eluent I, change 60% methanol then as mobile phase, get eluent II, get sample 13.83g behind the concentrate drying; Continue to separate the sample that obtains with preparative liquid chromatography: the separation condition of preparative hplc: chromatographic column is semi-preparative column (Lichrospher C18; 10.0mm * 250mm, 5 μ m), mobile phase is water (A) and acetonitrile (B), the gradient elution program is as follows: during 0min, mobile phase A is water+0.1% acetum of 85%, and Mobile phase B is 15% acetonitrile solution; During 10in, mobile phase A is water+0.1% acetum of 79%, and Mobile phase B is 21% acetonitrile solution; During 15min, mobile phase A is water+0.1% acetum of 69%, and Mobile phase B is 31% acetonitrile solution; During 30min, mobile phase A is water+0.1% acetum of 63%, and Mobile phase B is 37% acetonitrile solution; During 35min, mobile phase A is water+0.1% acetum of 15%, and Mobile phase B is 85% acetonitrile solution; During 45min, mobile phase A is water+0.1% acetum of 15%, and Mobile phase B is 85% acetonitrile solution; Flow velocity is 3ml/min, and column temperature is a room temperature; Sample 100% dissolve with ethanol separates through preparative liquid chromatography, collects solution at time period 17.5～23.6min, and solution obtains active component 5.70g behind concentrate drying.
The analysis of embodiment four Danshen effective components
(1) alkannic acid in the Radix Salviae Miltiorrhizae extract, salvianolic acid B, the assay of salvianolic acid E (high performance liquid chromatography)
Chromatographic conditionChromatographic column Agilent Zorbax SB-C
18Post (4.6mm * 150mm, 5 μ m); Adopt gradient elution, mobile phase A is 0.2% glacial acetic acid aqueous solution mutually, and Mobile phase B is mutually for containing the acetonitrile solution of 0.2% glacial acetic acid; The gradient elution program is as follows: in the time of 0 minute, mobile phase A is that 80% 0.2% glacial acetic acid aqueous solution, Mobile phase B are the acetonitrile solution of 20% 0.2% glacial acetic acid; In the time of 15 minutes, mobile phase A is that 50% 0.2% glacial acetic acid aqueous solution, Mobile phase B are the acetonitrile solution of 50% 0.2% glacial acetic acid; In the time of 20 minutes, mobile phase A is that 50% 0.2% glacial acetic acid aqueous solution, Mobile phase B are the acetonitrile solution of 50% 0.2% glacial acetic acid; Flow velocity 0.5mLmin
-1It is long to detect the wavelength all-wave; 30 ℃ of column temperatures; DAD detects in the interscan of 190-400nm scope; Sample size 5 μ L.
The preparation of need testing solutionTake by weighing this product, in volumetric flask, be diluted to scale, shake up, promptly with dissolve with methanol solution.
Assay methodThe accurate need testing solution of drawing injects chromatograph of liquid, measures, promptly.
(2) above-mentioned Radix Salviae Miltiorrhizae extract HPLC-ELSD finger printing
Assay methodReferring to the alkannic acid in (1) above-mentioned Radix Salviae Miltiorrhizae extract, salvianolic acid B, salvianolic acid E assay (high performance liquid chromatography).The record chromatograph time is 25 minutes.
Analysis resultThe HPLC-ELSD analysis of spectra of Danshen effective component is seen Fig. 1.1,2, No. 3 peak among Fig. 1 is respectively an alkannic acid, salvianolic acid B, and salvianolic acid E, its content is respectively 3.56%, 93.02%, and 1.51%.
The preparation of embodiment five dropping pill formulations
Get Danshen effective component 0.5g and 10.5g Polyethylene Glycol-6000 mix homogeneously of embodiment three, heating and melting moves in the drop pill drip irrigation behind the change material, and in ℃ liquid paraffin of medicine liquid droplet to 6～8, oil removing makes 300 of drop pill.
The preparation of embodiment six lyophilized injectable powders
Get Danshen effective component 0.5g, glucose 4.5g, sodium thiosulfate 0.9g and the distilled water 1ml of embodiment three, behind the said components mix homogeneously, lyophilization, 400 of packing, promptly.
The preparation of embodiment seven lyophilized injectable powders
Get Lignum Dalbergiae Odoriferae oil 1.5g, join in the saturated hydroxypropyl of 13ml, stirring and dissolving filters, and the filtrate cold drying gets the clathrate powder of Lignum Dalbergiae Odoriferae oil and hydroxypropyl.Except that above-mentioned Lignum Dalbergiae Odoriferae oil closes the clathrate powder of hydroxypropyl, get Radix Salviae Miltiorrhizae extract 0.5g, mannitol 5.5g, calcium disodium edetate 0.9g and the distilled water 2ml of embodiment three again, behind the said components mixing, lyophilization, 350 of packing, promptly.
The pharmacological evaluation of embodiment eight Danshen effective components
1 pharmacological model: neonatal rat myocardial cell hypoxic-ischemic model
Former generation myocardial cell cultivationAfter 1～3 day SD neonatal rat that is born was put to death, it was dirty to core, and subtracts into 1mm after PBS liquid cleans
3About fragment.With 0.125% trypsin Sigma, USA)+0.05% (Gibco is USA) in 37 degree digestion 3～4 times for collagenase II.The differential adherent method was separated into fibrocyte after 1.5 hours, and cell suspension is transferred to (every hole about 4 * 10 in 48 orifice plates
5Cell).(Gibco, (Falcon, USA) cell of cultivating 3-6 days supplies medicine efficacy screening USA) to add 10% hyclone through DMEM.Screen and replaced with serum-free medium in preceding 1 day.Cultivate 3～6 days myocardial cell, PBS washing back adds the pastille culture fluid.Place 95%N
2And 5%CO
2Anoxia is 6 hours in the incubator.Then at CO
2Reoxygenation is 3 hours in the incubator.Get cell conditioned medium liquid and measure LDH content.
Dosage regimenThe Danshen effective component that extraction obtains is made into 10 with sugar-free Hanks liquid
-4G/mL test liquid, liposoluble constituent can add a small amount of DMSO hydrotropy (the DMSO final concentration is controlled at below 0.2%).The final concentration of each component in culture fluid is controlled at 10
-5G/mL.
The lactic acid dehydrogenase assayLactic acid dehydrogenase content can be represented the cell injury degree in the cell culture fluid.The lactic acid dehydrogenase content that bleeds in the supernatant is high more, shows that cell injury is also remarkable.The determination of lactate dehydrogenase test kit builds up bio-engineering research institute available from Nanjing.Assay method is: get 30 microlitre cell conditioned medium liquid, add 50 microlitre substrate buffer and 10 microlitre lactic dehydrogenase enzyme cofactors, 37 degree vibrations 15 minutes add 50 microlitre 2,4 dinitrophenyl hydrazines again, 37 degree vibrations 15 minutes.Parallel blank.Extract reaction solution 50 microlitres and add 200 microlitre 0.4mol/L sodium hydroxide solutions, leave standstill 3～4 minutes after, measure shading value with microplate reader in 450nm.
Drug effect result
All data are represented with the mean value variance.Adopt monolateral variance analysis and T check carrying out statistical analysis.Compare with model group, P＜0.05 is thought significantly effectively.Drug effect the results are shown in Table 1.According to myocardial cell lactic acid dehydrogenase (LDH) testing result, Danshen effective component has the highly significant effect to reducing LDH release.
Table 1
2 pharmacological models: huve cell anoxia reoxygenation model
Former generation huve cell cultivationGet the healthy newborn umbilical cord, clean the interior residual bloodstain of vein blood vessel with 30～50mlHanks liquid.Look the 0.1% collagenase I that umbilical cord length adds respective amount, change incubator digestion 15～20 minutes over to.Subsequently Digestive system in the umbilical cord is carefully collected in the beaker, and with Hanks liquid with beaker rinse twice, collect in the lump and be sub-packed in centrifuge tube in the beaker, centrifugal 10 minutes of 900rpm.Supernatant is removed in suction, with the abundant dissolution precipitation of M199 culture fluid (contain 10% hyclone, 100U/ml is two anti-, 0.15mg/ml Heparin, 10uM VC).Gained cell suspension is sub-packed in 5cm
2Culture bottle places in the incubator and cultivates.Changed in second day and adopt the M199 culture fluid that contains 30ug/ml ECGS, changed a culture fluid and be paved with the bottom surface in per afterwards three days until cell.Used cell is former generation endotheliocyte.The culture bottle inner cell is seeded to 96 orifice plates before the modeling and cultivated one day, used cell concentration is 3.5～40,000/hole.During modeling, inhale and abandon old M199 culture fluid, adding pastille does not have phenol red M199 culture fluid, and every hole total amount is 100ul.Place 95%N
2And 5%CO
2Anoxia is 12 hours in the incubator, then at CO
2Reoxygenation is 2 hours in the incubator.Get cell conditioned medium liquid and measure LDH content and NO content.
Dosage regimenThe Danshen effective component that extraction obtains is made into 10 with sugar-free Hanks liquid
-4G/mL test liquid, liposoluble constituent can add a small amount of DMSO hydrotropy (the DMSO final concentration is controlled at below 0.2%).The final concentration of each component in no phenol red M199 culture fluid is controlled at 10
-5G/mL.
The NO assayAdopt Greiss reagent method to measure, get 90ul cell conditioned medium liquid, add equivalent Greiss reagent, left standstill under the room temperature 10 minutes, measure absorbance in 550nm with microplate reader.
Drug effect resultAll data are represented with the mean value variance.Adopt monolateral variance analysis and T check carrying out statistical analysis.Compare with model group, P＜0.05 is thought significantly effectively.Drug effect the results are shown in Table 2.According to endotheliocyte NO content detection result, the Fructus Aurantii active component has the highly significant effect to NO overexpression after suppressing endotheliocyte anoxia-reoxygenation.
Table 2
Claims (5)
1. a Danshen effective component is characterized in that, by weight percentage, wherein the content of alkannic acid is 2.8～4.2%, and the content of salvianolic acid B is 91.8～94.2%, and the content of salvianolic acid E is 0.8～2.2%;
Described Danshen effective component adopts following method preparation:
Get red rooted salvia, it is pulverized the back add 1: 1 ethyl acetate of volume ratio and ethanol, reflux 1 hour, extract 2 times, filtrate merge extracting solution I, medicinal residues add 70% ethanol, reflux 1 hour, extract 2 times, filtrate merge extracting solution II, II is condensed into extractum with extracting solution, with 50% dissolve with methanol extractum, adopt the ODS-C18 post that it is separated, at first use 5% methanol as mobile phase, get eluent I, change 60% methanol then as mobile phase, get eluent II, get sample behind the concentrate drying; Continue to separate the sample that obtains with preparative liquid chromatography: the separation condition of preparative hplc: chromatographic column is semi-preparative column LichrospherC18; 10.0mm * 250mm, 5 μ m, mobile phase A is that water and B are acetonitrile, the gradient elution program is as follows: during 0min, mobile phase A is water+0.1% acetum of 85%, and Mobile phase B is 15% acetonitrile solution; During 10min, mobile phase A is water+0.1% acetum of 79%, and Mobile phase B is 21% acetonitrile solution; During 15min, mobile phase A is water+0.1% acetum of 69%, and Mobile phase B is 31% acetonitrile solution; During 30min, mobile phase A is water+0.1% acetum of 63%, and Mobile phase B is 37% acetonitrile solution; During 35min, mobile phase A is water+0.1% acetum of 15%, and Mobile phase B is 85% acetonitrile solution; During 45min, mobile phase A is water+0.1% acetum of 15%, and Mobile phase B is 85% acetonitrile solution; Flow velocity is 3ml/min, and column temperature is a room temperature; Sample 100% dissolve with ethanol separates through preparative liquid chromatography, collects solution at time period 17.5～23.6min, and solution obtains active component behind concentrate drying.
2. Danshen effective component as claimed in claim 1 is characterized in that, by weight percentage, the content of alkannic acid is 3～4%, and the content of salvianolic acid B is 92～94%, and the content of salvianolic acid E is 1～2%.
3. Danshen effective component as claimed in claim 1 is characterized in that, by weight percentage, the content of alkannic acid is 3.2～3.8%, and the content of salvianolic acid B is 92.5～93.5%, and the content of salvianolic acid E is 1.2～1.8%.
4. contain the pharmaceutical formulation of the described Danshen effective component of the arbitrary claim of claim 1-3, it is characterized in that, make arbitrary pharmaceutical formulation that pharmaceutics is accepted according to the preparation method that pharmaceutics allows.
5. the application of the described Danshen effective component of the arbitrary claim of claim 1-3 in preparation treatment and angiocardiopathy preventing medicine.
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| 王美凤 等.丹参药材中丹酚酸B的提取及其HPLC法测定.现代中药研究与实践.2004,1862-64. |
| 王美凤等.丹参药材中丹酚酸B的提取及其HPLC法测定.现代中药研究与实践.2004,1862-64. * |
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