CN105687262A

CN105687262A - Folium ginkgo tincture and preparing method thereof

Info

Publication number: CN105687262A
Application number: CN201610053171.XA
Authority: CN
Inventors: 王海涛; 杜秀琴; 王秋平
Original assignee: BEIJING HUARUN GAOKE NATURAL MEDICINE Co Ltd
Current assignee: BEIJING HUARUN GAOKE NATURAL MEDICINE Co Ltd
Priority date: 2016-01-26
Filing date: 2016-01-26
Publication date: 2016-06-22

Abstract

The invention provides a folium ginkgo tincture which comprises traditional Chinese medicine composition formed by folium ginkgo extracts and ethanol water for dissolving the traditional Chinese medicine composition. The traditional Chinese medicine composition is prepared from the follwing raw materials in percentage by weight: 24-40 percent of gingko general flavone, 6-16 percent of bilobalide of which ginkgoic acid is smaller than 5ppm, 2.08-3.46 percent of kaempferol-3-O-p-coumaric acyl glucose rhamnoside and 2.11-3.53 percent of quercetin-3-O-2',6'-rhamanopyranosyl glucoside. According to the folium ginkgo tincture, the content of the gingko general flavone and the content of the bilobalide in the folium ginkgo extracts are clearly limited. Meanwhile, the kaempferol-3-O-p-coumaric acyl glucose rhamnoside and the quercetin-3-O-2', 6'-rhamanopyranosyl glucoside which have important effects on the folium ginkgo tincture are definite. The medical effects of the folium ginkgo extracts are improved by further defining the content of effective ingredients and further defining the effective ingredients. Thus, the medical effects of the tincture prepared from the folium ginkgo extracts are further improved.

Description

A kind of Folium Ginkgo tincture and preparation method thereof

Technical field

The present invention relates to a kind of Folium Ginkgo tincture and preparation method thereof, belong to Chinese medicine synthesis technical field。

Background technology

Folium Ginkgo is the dried leaves of Ginkgoaceae plant Ginkgo biloba, Folium Ginkgo extract is the enriched products of the active substance extracted from Folium Ginkgo through modern extraction process, can be used for the treatment of the diseases such as Alzheimer, depression, diabetes, sacred disease, sexual impotence, dysmnesia, peripheral blood vessel, intermittent claudication, vertigo and tinnitus。Its main active is flavonoid and terpenoid。Flavones ingredient includes single flavone, flavonol glycosides, acetyl-flavones alcohol glycosides, bisflavone, flavan-3-alcohol class and proanthocyanidin etc.。Terpenoid bilobalide has Ginkgolide A. B. C, J, M and bilobalide。

Current state food pharmaceuticals administration general bureau (CFDA) have approved tens of kinds of Folium Ginkgo extract dosage forms, including Folium Ginkgo, capsule, granule, soft capsule, dispersible tablet, ball, tincture, drop, oral liquid, gingko leaf extract injection etc., its patent medicine form is all Folium Ginkgo extract and other adjuvant be mixed to form, and ginkgo tree leave tincture is exactly a kind of dosage form being dissolved in the ethanol of certain depth to be formed Folium Ginkgo extract。There is suppression platelet aggregation, reduce Blood denseness, improve microcirculatory effect, and can pass through to strengthen cardiac muscular tissue's metabolism, suppress myocardial fibrosis, by vasodilator, improve microcirculation etc., keep the supply of nutritional labeling and oxygen, myocardial function sorrow is stoped to be moved back, oxygen-derived free radicals can be reduced simultaneously, carry the tolerance still to anoxia, regulate blood official's function, strengthen tissue energy metabolism, reduce serum total cholesterol and low-density lipoprotein cholesterol, high density lipoprotein increasing cholesterol, the risk factor etc. reducing cerebral infarction and other cardiovascular and cerebrovascular diseases acts on。It is mainly used in treating the obstruction of qi in the chest and cardialgia that causes for obstruction of collaterals by blood stasis, apoplexy, hemiplegia, stiff tongue language stuttering；Treating Stable Angina Pectoris of Coronary Artery Disease, cerebral infarction are waited indefinitely disease。It is known that, Folium Ginkgo extract is an extremely complex compound enriched products, except above-mentioned main active component, it contains from inorganic matter to Organic substance, from polarity to various compositions nonpolar, from little molecule to biomacromolecule, according to incompletely statistics containing more than 240 chemical composition in Folium Ginkgo。By the restriction of technical development, in current Folium Ginkgo extract, each chemical composition is not identified completely, and active ingredient, mechanism of action, pharmacokinetics and untoward reaction are not probed into clear completely yet。And the technical staff with general knowledge known in this field all knows, mechanism of action mentioned above, pharmacokinetics and untoward reaction all have inevitable contacting with the chemical composition in patent medicine。Therefore, the effective ingredient in further clear and definite Folium Ginkgo extract, and the content of further accurately effective ingredient, improve the drug effect of Folium Ginkgo extract, reduce untoward reaction, have great importance for improving patient health and drug safety thereof。

Summary of the invention

It is an object of the invention to provide a kind of effective ingredient and content Folium Ginkgo tincture definitely the preparation method that this extract is further provided。

For this, the technical scheme that the application takes is,

A kind of Folium Ginkgo tincture, including the Chinese medicine composition formed by Folium Ginkgo extract with for dissolving the ethanol water of described Chinese medicine composition, to account for the mass percent of described Chinese medicine composition, described Chinese medicine composition includes, the Ginkgo total flavones of 24-40%, the bilobalide of 6-16%, ginkgoic acid is less than 5ppm。

In above-mentioned Folium Ginkgo tincture, described Chinese medicine composition includes, the kaempferol-3-O-p-coumaroyl glucose rhamnoside of 2.08-3.46%, the Quercetin-3-O-2 of 2.11-3.53% ", 6 "-two rhamanopyranosyl glucosides。

In above-mentioned Folium Ginkgo tincture, described Chinese medicine composition includes, the ginkalide A of 1.4-3%, the ginkalide B of 0.9-1.8% and the ginkalide C of 1.2-1.3%。

In above-mentioned Folium Ginkgo tincture, described Chinese medicine composition also includes bilobalide and rutin。

In above-mentioned Folium Ginkgo tincture, the preparation method of described Chinese medicine composition includes,

The preparation method of S1 kaempferol-3-O-p-coumaroyl glucose rhamnoside,

(1) Folium Ginkgo extract is dissolved in the ethanol-water solution that volumetric concentration is 40-80%, prepares the extract solution that concentration is 10-1000mg/mL of Folium Ginkgo extract；

(2) extract solution that described step (1) is obtained carries out one-dimensional liquid chromatograph separation, obtains one-dimensional thick product, and the condition that in this step, one-dimensional liquid chromatograph separates is, the hydrophilic chromatograph packing material that it is substrate with silica gel that chromatographic column adopts；Organic facies in mobile phase is ethanol or methanol, and aqueous phase is water；Elution requirement with 0-15min organic facies 95% be down to 90% gradient carry out or etc. degree carry out；Collect component and remaining ingredient that retention time is 15～20min, the component that the retention time collected is 15～20min is removed solvent seasoning and obtains the one-dimensional thick product of described kaempferol-3-O-p-coumaroyl glucose rhamnoside；

(3) the one-dimensional thick product that step (2) obtains is dissolved in the methanol-water solution or ethanol-water solution that volumetric concentration is 40-80%, obtains the thick product solution that concentration is 20-200mg/mL of described one-dimensional thick product；

(4) the thick product solution that described step (3) is obtained carries out two-dimensional liquid chromatography and prepares; obtain kaempferol-3-O-p-coumaroyl glucose rhamnoside; wherein two-dimensional liquid chromatography condition is, chromatographic column adopts with silica gel for substrate-bound C18 reverse phase filler；Organic facies in mobile phase is ethanol or methanol, and aqueous phase is water；Elution requirement with 0-60min organic facies 15% increase to 80% gradient carry out or etc. degree carry out；Collecting retention time is the component of 50-55min, dries and obtains kaempferol-3-O-p-coumaroyl glucose rhamnoside；

Quercetin-3-O-2 described in S2 ", 6 " preparation method of-two rhamanopyranosyl glucosides,

A Folium Ginkgo extract is dissolved in the ethanol-water solution that volumetric concentration is 40-80% by (), prepare the extract solution that concentration is 10-1000mg/mL of Folium Ginkgo extract；

B the extract solution that described step (a) obtains is carried out one-dimensional liquid chromatograph separation by (), obtain one-dimensional thick product；The condition that in this step, one-dimensional liquid chromatograph separates is, the hydrophilic chromatograph packing material that it is substrate with silica gel that chromatographic column adopts；Organic facies in mobile phase is ethanol or acetonitrile, and aqueous phase is water；Elution requirement with 0-15min organic facies 95% be down to 90% gradient carry out or etc. degree carry out；Collecting retention time is component and the residual components of 28-32min, the component that retention time is 28-32min is removed solvent seasoning and obtains containing Quercetin-3-O-2 ", 6 " the one-dimensional thick product of-two rhamanopyranosyl glucosides；

C one-dimensional thick product that step (b) is obtained by () is dissolved in the methanol-water solution or ethanol-water solution that volumetric concentration is 40-80%, obtains the thick product solution that concentration is 20-200mg/mL of described one-dimensional thick product；

D thick product solution that described step (c) is obtained by () carries out two-dimensional liquid chromatography to be prepared, and obtains Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides；

The preparation method of the Chinese medicine composition that S3 is formed by Folium Ginkgo extract

By described residual components and and described remaining ingredient remove mixing after volume; and dry obtain supplement; by described supplement, described kaempferol-3-O-p-coumaroyl glucose rhamnoside and described Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides are mixed to get the described Chinese medicine composition formed by Folium Ginkgo extract。

In above-mentioned Folium Ginkgo tincture,

In described step (d), two-dimensional liquid chromatography condition is, chromatographic column adopts with silica gel for substrate-bound C18 reverse phase filler；Organic facies in mobile phase is ethanol or acetonitrile, and aqueous phase is water；Isocratic elution in 0-60min, during eluting, the volumetric concentration of organic facies is 15-25%；Collecting retention time is the component of 40-45min, dries and obtains Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides。

In above-mentioned Folium Ginkgo tincture, in described step (2), one-dimensional liquid chromatograph separates, and organic facies is possibly together with formic acid, and formic acid volumetric concentration is 0.1%；Possibly together with formic acid in aqueous phase, formic acid volumetric concentration is 0.1%；

Prepared by described step (4) two-dimensional liquid chromatography, possibly together with formic acid in mobile phase, formic acid volumetric concentration is 0.1%；Possibly together with formic acid in aqueous phase, formic acid volumetric concentration is 0.1%。

In above-mentioned Folium Ginkgo tincture, in described step (c), one-dimensional liquid chromatograph separates, and organic facies is possibly together with formic acid, and formic acid volumetric concentration is 0.1%；Possibly together with formic acid in aqueous phase, formic acid volumetric concentration is 0.1%；

Prepared by described step (d) two-dimensional liquid chromatography, possibly together with formic acid in mobile phase, formic acid volumetric concentration is 0.1%；Possibly together with formic acid in aqueous phase, formic acid volumetric concentration is 0.1%。

The preparation method that present invention also provides a kind of any of the above-described Folium Ginkgo tincture, including, the Chinese medicine composition taking the formation of described Folium Ginkgo extract is dissolved in the ethanol water that volumetric concentration is 60%, obtaining, the mass volume ratio of wherein said Chinese medicine composition and described ethanol water is 0.4g/10ml。

In the preparation method of above-mentioned Folium Ginkgo tincture, also include the step adding adjuvant。

Compared with prior art, the invention have the advantages that,

(1) in the ginkgo tree leave tincture of the present invention, the content of Ginkgo total flavones in Folium Ginkgo extract therein is clearly arrived 24-40%, the content of bilobalide clearly arrives 6-16%, specify that two kinds of kaempferol-3-O-p-coumaroyl glucose rhamnosides that its medicine is had material impact and Quercetin-3-O-2 " simultaneously, 6 "-two rhamanopyranosyl glucoside, and further its content is accurately defined to 2.08-3.46% and 2.11-3.53%, further, clearly define its ginkalide A 1.4-3%, ginkalide B 0.9-1.8% and ginkalide C 1.2-1.3%。By in Folium Ginkgo extract active constituent content further accurately, and the further identification of effective ingredient, improve the drug effect of Folium Ginkgo extract, and then improve the drug effect of the tincture prepared by it。

(2) in the ginkgo tree leave tincture of the present invention, by carrying out refining and purifying to Folium Ginkgo extract, obtain the compositions with definite content, while improving Folium Ginkgo extract drug effect, impurity is decreased, thus decreasing the untoward reaction of tincture by aforesaid operations process。

(3) the application adopts two-dimensional liquid chromatography method to isolate clear and definite medicinal compound kaempferol-3-O-p-coumaroyl glucose rhamnoside and Quercetin-3-O-2 from Folium Ginkgo extract "; 6 "-two rhamanopyranosyl glucosides; its preparation method is simple; lock out operation is convenient, is beneficial to the raising of Folium Ginkgo extract drug quality。

Accompanying drawing explanation

In order to make present disclosure be more likely to be clearly understood, below according to specific embodiments of the invention and in conjunction with accompanying drawing, the present invention is further detailed explanation, wherein

The mass spectrum of the kaempferol-3-O-p-coumaroyl glucose rhamnoside prepared in Fig. 1 embodiment of the present invention 1；

The H spectrum of the kaempferol-3-O-p-coumaroyl glucose rhamnoside prepared in Fig. 2 embodiment of the present invention 1；

The C spectrum of the kaempferol-3-O-p-coumaroyl glucose rhamnoside prepared in Fig. 3 embodiment of the present invention 1；

Quercetin-the 3-O-2 prepared in Fig. 4 embodiment of the present invention 1 ", 6 " mass spectrum of-two rhamanopyranosyl glucosides；

Quercetin-the 3-O-2 prepared in Fig. 5 embodiment of the present invention 1 ", 6 " H of-two rhamanopyranosyl glucosides spectrum；

Quercetin-the 3-O-2 prepared in Fig. 6 embodiment of the present invention 1 ", 6 " C of-two rhamanopyranosyl glucosides spectrum。

Detailed description of the invention

The percent % occurred in the embodiment of the present invention, what represent if no special instructions is percentage by volume, for instance, " ethanol-water solution of 40% " represents that the percentage by volume of the aqueous solution wherein ethanol of ethanol is 40%；

" methanol-water solution of 40% " represents that the percentage by volume of the aqueous solution wherein methanol of methanol is 40%；

" ethanol (containing 0.1% formic acid) " represents that the percentage by volume of the mixed solution wherein formic acid of ethanol and formic acid is 0.1%；" water (containing 0.1% formic acid) " represents that the percentage by volume of the mixed solution wherein formic acid of water and formic acid is 0.1%。

The Folium Ginkgo extract of the application is applicable for use with existing extraction process and extracts the extract obtained from Folium Ginkgo, and for ease of comparing, the Folium Ginkgo extract adopted in the embodiment of the present application is provided by Shuanghe High-Tech. Natural Medicine Co., Ltd., Beijing。

Embodiment 1

The preparation of kaempferol-3-O-p-coumaroyl glucose rhamnoside

Weighing Folium Ginkgo extract 10g, be dissolved in the ethanol-water solution of 50mL40%, prepare Folium Ginkgo extract solution, concentration is 200mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph separation。Hydrophilic chromatograph packing material 50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm (Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts the methanol containing 0.1% volumetric concentration formic acid to be organic facies, water containing 0.1% volumetric concentration formic acid is aqueous phase, gradient elution mode: 0-15min organic phase concentration is down to 90% stepwise gradient from 95% and is carried out。DAD UV-detector 360nm is adopted to select absorbing wavelength; preparation temperature is room temperature; sample size is 500 μ L/ pins; flow rate of mobile phase is 90mL/min; collect fraction and the remaining ingredient of 15～20 minutes; the fractions of collect 15～20 minutes are rotated evaporation and concentration to dry, prepare kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product for one-dimensional。Ethanol-water solution with 50% dissolves kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product; concentration is 80mg/mL; through filtering with microporous membrane; carry out two-dimensional liquid chromatography to prepare; hydrophilic chromatograph packing material (50 × 150mm that chromatographic column is is substrate with silica gel; 5 μm; Hua Puxinchuan Science and Technology Ltd.) mobile phase selects methanol containing 0.1% volumetric concentration formic acid to be organic facies; water containing 0.1% volumetric concentration formic acid is aqueous phase, adopts the 15-80% organic facies gradient elution of 0-60min。DAD UV-detector 360nm is adopted to select absorbing wavelength; preparation temperature is room temperature; sample size is 1000 μ L/ pins; flow rate of mobile phase is 90mL/min; collect the retention time fraction at 50-55min; rotary evaporated to dryness; obtain kaempferol-3-O-p-coumaroyl glucose rhamnoside compound; through liquid-phase chromatographic analysis; purity is 95.5%, and one-dimensional to prepare the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside in kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product be 30%。

Be respectively adopted mass spectrum, the end-product obtained is analyzed by 1H-NMR and 13C-NMR (MeOD), wherein mass spectrum, 1H-NMR spectrogram as illustrated in fig. 1 and 2,

13C-NMR (MeOD) resolves as follows,

Kaempferol female ring: 156.98 (C2), 135.42 (C3), 178.14 (C4), 161.75 (C5), 98.40 (C6), 164.18 (C7), 93.46 (C8), 156.88 (C9), 104.68 (C10), 121.25 (C1), 130.56 (C2,, C6), 115.00 (C3, C5,), 159.67 (C4)。

3-O-rhamnose: 101.12 (C1 "), 82.26 (C2 "), 70.38 (C3 "), 72.10 (C4 "), 70.55 (C5 "), 16.33 (C6 ")。

2 "-glucose: 105.66 (C1 " '), 73.99 (C2 " '), 76.29 (C3 " '), 70.78 (C4 " '), 73.73 (C5 " '), 63.12 (C6 " ')。

6 "-p-coumaroyl: 167.40 (C1 " "); 113.29 (C2 " "); 145.16 (C3 " "); 125.58 (C4 " "); 129.62 (C5 " ", C9 " "), 115.23 (C6 " "; C8 " "), 160.02 (C7 " ")。

Comprehensive identification is kaempferol-3-O-p-coumaroyl glucose rhamnoside, and the molecular formula of compound is C36H36O17, and molecular weight 740.1982, structural formula is as follows

Quercetin-3-O-2 ", 6 " preparation of-two rhamanopyranosyl glucosides

Weighing Folium Ginkgo extract 10g, be dissolved in the ethanol-water solution of 50mL40%, prepare Folium Ginkgo extract solution, concentration is 200mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph separation。Hydrophilic chromatograph packing material 50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm (Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts acetonitrile (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, gradient elution mode: 0-15min organic phase concentration is down to 90% stepwise gradient from 95% and is carried out。DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is room temperature, sample size is 500 μ L/ pins, flow rate of mobile phase is 90mL/min, collect component and the residual components of 28-30 minute, the component of 28-30 minute collected is rotated evaporation and concentration to dry, prepares Quercetin-3-O-2 for one-dimensional ", 6 "-two rhamanopyranosyl glucoside crude products。Ethanol-water solution with 50% dissolves Quercetin-3-O-2 "; 6 "-two rhamanopyranosyl glucoside crude products, concentration is 80mg/mL, through filtering with microporous membrane, carry out two-dimensional liquid chromatography to prepare, hydrophilic chromatograph packing material (50 × 150mm that chromatographic column is is substrate with silica gel, 5 μm, Hua Puxinchuan Science and Technology Ltd.) mobile phase adopt acetonitrile (containing 0.1% formic acid) be organic facies, water (containing 0.1% formic acid) is aqueous phase, adopts 0-60min20% organic facies isocratic elution。Adopting DAD UV-detector 360nm to select absorbing wavelength, preparation temperature is room temperature, and sample size is 1000 μ L/ pins, flow rate of mobile phase is 90mL/min, collects the retention time component at 40-45min, rotary evaporated to dryness, obtain Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucoside compounds。

Be respectively adopted mass spectrum,¹H-NMR and¹³The end-product obtained is analyzed by C-NMR (MeOD), wherein mass spectrum,¹H-NMR spectrum as shown in Figures 4 and 5,

¹³C-NMR (MeOD) resolves as follows,

Quercetin female ring: 157.04 (C2), 133.07 (C3), 177.88 (C4), 161.73 (C5), 98.38 (C6), 164.24 (C7), 93.32 (C8), 157.53 (C9), 104.51 (C10), 122.16 (C1 '), 116.04 (C2 '), 144.51 (C3 '), 148.14 (C4 '), 114.68 (C5 '), 122.08 (C6 ')

Glucose: 101.24 (C1 "), 72.48 (C2 "), 75.66 (C3 "), 70.47 (C4 "), 72.67 (C5 "), 66.89 (C6 ")。

2 "-rhamnose: 100.85 (C1 " '), 78.64 (C2 " '), 70.74 (C3 " '), 71.01 (C4 " '), 68.32 (C5 " '), 16.13 (C6 " ')。

6 "-rhamnose: 99.10 (C1 " "), 77.51 (C2 " "), 70.87 (C3 " "), 70.91 (C4 " "), 68.56 (C5 " "), 16.43 (C6 " ")。

Comprehensive identification is Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides, the molecular formula of compound is C₃₃H₄₀O₂₀, molecular weight 756.2103, structural formula is as follows

Mix after described residual components and described remaining ingredient rotary evaporation are removed volume; and dry obtain supplement; by described supplement, described kaempferol-3-O-p-coumaroyl glucose rhamnoside and described Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides are mixed to get the Chinese medicine composition formed by Folium Ginkgo extract of the present embodiment。

Adopt high performance liquid chromatography that the Chinese medicine composition of the application is detected; mass percent with described Chinese medicine composition; Ginkgo total flavones 24%, bilobalide 6%, ginkgoic acid is less than 5ppm; kaempferol-3-O-p-coumaroyl glucose rhamnoside 2.08%; Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides 2.11%, ginkalide A 1.4%; ginkalide B 1.8% and ginkalide C 1.3%, also detection has the chemical composition such as bilobalide and rutin。

The preparation of S4 ginkgo tree leave tincture

Take the above-mentioned Chinese medicine composition 0.4g formed by Folium Ginkgo extract, be dissolved in the ethanol water that 10ml volumetric concentration is 60% and form ginkgo tree leave tincture。

Embodiment 2

The preparation of kaempferol-3-O-p-coumaroyl glucose rhamnoside

Weighing Folium Ginkgo extract 1g, be dissolved in the ethanol-water solution of 100mL volumetric concentration 80%, prepare Folium Ginkgo extract solution, concentration is 10mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol to be organic facies, water is aqueous phase, organic phase concentration is 90% degree such as grade, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is 40 DEG C, sample size is 200 μ L/ pins, flow rate of mobile phase is 60mL/min, collect fraction and the remaining ingredient of 15～20 minutes, the fractions of collect 15～20 minutes are rotated evaporation and concentration to dry, kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product is prepared for one-dimensional。Ethanol-water solution with 40% dissolves kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product; concentration is 20mg/mL; through filtering with microporous membrane; carry out two-dimensional liquid chromatography to prepare; chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm; 10 μm; Hua Puxinchuan Science and Technology Ltd.); mobile phase selects the ethanol containing 0.1% volumetric concentration formic acid to be organic facies; water containing 0.1% volumetric concentration formic acid is aqueous phase; adopting isocratic elution mode, organic phase concentration is 20% totally 60 minutes。DAD UV-detector 360nm is adopted to select absorbing wavelength; preparation temperature is room temperature; sample size is 1000 μ L/ pins; flow rate of mobile phase is 100mL/min, collects the fraction of retention time 50-55min, rotary evaporated to dryness; obtain kaempferol-3-O-p-coumaroyl glucose rhamnoside compound; in liquid-phase chromatographic analysis, the kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product of two dimension preparation, the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside is 94.0%

Be respectively adopted mass spectrum with embodiment 1, the end-product obtained is analyzed by 1H-NMR and 13C-NMR (MeOD), and what final confirmation obtained is kaempferol-3-O-p-coumaroyl glucose rhamnoside。。

Quercetin-3-O-2 ", 6 " preparation of-two rhamanopyranosyl glucosides

Weighing Folium Ginkgo extract 1g, be dissolved in the ethanol-water solution of 100mL volumetric concentration 80%, prepare Folium Ginkgo extract solution, concentration is 10mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol to be organic facies, water is aqueous phase, organic phase concentration is 90% degree such as grade, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is 40 DEG C, sample size is 200 μ L/ pins, flow rate of mobile phase is 60mL/min, collect component and the residual components of 28～32 minutes, by the component of collect 28-32 minute, rotate evaporation and concentration to dry, Quercetin-3-O-2 is prepared " for one-dimensional, 6 "-two rhamanopyranosyl glucoside crude product。Ethanol-water solution with 40% dissolves Quercetin-3-O-2 "; 6 "-two rhamanopyranosyl glucoside crude products, concentration is 20mg/mL, through filtering with microporous membrane, carry out two-dimensional liquid chromatography to prepare, chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase selects ethanol (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, adopts isocratic elution mode, and organic phase concentration is 20% totally 60 minutes。Adopting DAD UV-detector 360nm to select absorbing wavelength, preparation temperature is room temperature, and sample size is 1000 μ L/ pins, flow rate of mobile phase is 100mL/min, collects the component of retention time 40-45min, rotary evaporated to dryness, obtain Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucoside compounds。

With embodiment 1 be respectively adopted mass spectrum,¹H-NMR and¹³The end-product obtained is analyzed by C-NMR (MeOD), and that final confirmation obtains is Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides。

Adopt high performance liquid chromatography that the Chinese medicine composition of the application is detected; mass percent with described Chinese medicine composition; Ginkgo total flavones 40%, bilobalide 16%, ginkgoic acid 11ppm; kaempferol-3-O-p-coumaroyl glucose rhamnoside 2.45%; Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides 3.53%, ginkalide A 3%; ginkalide B 1.8% and ginkalide C 1.3%, also detection has the chemical composition such as bilobalide and rutin。

The preparation of S4 ginkgo tree leave tincture

Embodiment 3

The preparation of kaempferol-3-O-p-coumaroyl glucose rhamnoside

Weighing Folium Ginkgo extract 100g, be dissolved in the ethanol-water solution of 200mL50%, prepare Folium Ginkgo extract solution, concentration is 500mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μ, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, adopts 95% organic equality mode eluting。DAD UV-detector 360nm is adopted to select absorbing wavelength; preparation temperature is 30 DEG C; sample size is 3000 μ L/ pins; flow rate of mobile phase is 120mL/min; collect component and the remaining ingredient of 15～20 minutes; the components of collect 15～20 minutes are rotated evaporation and concentration to dry, prepare kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product for one-dimensional。Ethanol-water solution with 60% dissolves kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product; concentration is 80mg/mL; through filtering with microporous membrane; carry out two-dimensional liquid chromatography to prepare; chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm; 10 μ; Hua Puxinchuan Science and Technology Ltd.); mobile phase selects ethanol (containing 0.1% formic acid) to be organic facies; water (containing 0.1% formic acid) is aqueous phase, adopts organic facies to rise to 80% gradient elution from 20%。DAD UV-detector 360nm is adopted to select absorbing wavelength; preparation temperature is 25 DEG C; sample size is 3000 μ L/ pins; flow rate of mobile phase is 120mL/min; collect the fraction of retention time 50-55min; rotary evaporated to dryness; obtain kaempferol-3-O-p-coumaroyl glucose rhamnoside compound; through liquid-phase chromatographic analysis; purity is 96.5%, and one-dimensional to prepare the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside in kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product be 29%。

Be respectively adopted mass spectrum with embodiment 1, the end-product obtained is analyzed by 1H-NMR and 13C-NMR (MeOD), and what final confirmation obtained is kaempferol-3-O-p-coumaroyl glucose rhamnoside。

Quercetin-3-O-2 ", 6 " preparation of-two rhamanopyranosyl glucosides

Weighing Folium Ginkgo extract 100g, be dissolved in the ethanol-water solution of 200mL50%, prepare Folium Ginkgo extract solution, concentration is 500mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μ, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, adopts 95% organic equality mode eluting。DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is 30 DEG C, sample size is 3000 μ L/ pins, flow rate of mobile phase is 120mL/min, collect component and the residual components of 28～32 minutes, the component of 28-32 minute collected is rotated evaporation and concentration to dry, prepares Quercetin-3-O-2 for one-dimensional ", 6 "-two rhamanopyranosyl glucoside crude products。Ethanol-water solution with 60% dissolves Quercetin-3-O-2 "; 6 "-two rhamanopyranosyl glucoside crude products, concentration is 80mg/mL, through filtering with microporous membrane, carries out two-dimensional liquid chromatography and prepares, chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm, 10 μ, Hua Puxinchuan Science and Technology Ltd.s), mobile phase selects ethanol (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, adopts organic facies 25% isocratic elution。Adopting DAD UV-detector 360nm to select absorbing wavelength, preparation temperature is 25 DEG C, and sample size is 3000 μ L/ pins, flow rate of mobile phase is 120mL/min, collects the component of retention time 40-45min, rotary evaporated to dryness, obtain Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucoside compounds。

Adopt high performance liquid chromatography that the Chinese medicine composition of the application is detected; mass percent with described Chinese medicine composition; Ginkgo total flavones 36%, bilobalide 12%, ginkgoic acid 0.1ppm; kaempferol-3-O-p-coumaroyl glucose rhamnoside 2.95%; Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides 2.31%, ginkalide A 2.4%; ginkalide B 1.6% and ginkalide C 1.25%, also detection has the chemical composition such as bilobalide and rutin。

The preparation of S4 ginkgo tree leave tincture

Embodiment 4

The preparation of kaempferol-3-O-p-coumaroyl glucose rhamnoside

Weighing Folium Ginkgo extract 100g, be dissolved in the ethanol-water solution of 100mL40%, prepare Folium Ginkgo extract solution, concentration is 1000mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μ, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, adopt gradient elution: organic phase concentration was reduced to 90% by 95% through 15 minutes, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is 25 DEG C, sample size is 2000 μ L/ pins, flow rate of mobile phase is 80mL/min, collect component and the remaining ingredient of 15～20 minutes, the components of collect 15～20 minutes are rotated evaporation and concentration to dry, kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product is prepared for one-dimensional。Ethanol-water solution with 80% dissolves kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product; concentration is 50mg/mL; through filtering with microporous membrane; carrying out two-dimensional liquid chromatography to prepare, chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm, 10 μ; Hua Puxinchuan Science and Technology Ltd.); mobile phase selects ethanol (containing 0.1% formic acid) to be organic facies, and water (containing 0.1% formic acid) is aqueous phase, adopts 15% organic phase concentration eluting。DAD UV-detector 360nm is adopted to select absorbing wavelength; preparation temperature is 40 DEG C; sample size is 200 μ L/ pins; flow rate of mobile phase is 60mL/min, collects retention time in 50-55min fraction, rotary evaporated to dryness; obtain kaempferol-3-O-p-coumaroyl glucose rhamnoside compound; through liquid-phase chromatographic analysis, purity is 94.2%, and one-dimensional to prepare the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside in kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product be 35%。

Quercetin-3-O-2 ", 6 " preparation of-two rhamanopyranosyl glucosides

Weighing Folium Ginkgo extract 100g, be dissolved in the ethanol-water solution of 100mL40%, prepare Folium Ginkgo extract solution, concentration is 1000mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μ, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, adopt gradient elution: organic phase concentration was reduced to 90% by 95% through 15 minutes, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is 25 DEG C, sample size is 2000 μ L/ pins, flow rate of mobile phase is 80mL/min, collect component and the residual components of 28-32 minute, the component of 28-32 minute collected is rotated evaporation and concentration to dry, Quercetin-3-O-2 is prepared " for one-dimensional, 6 "-two rhamanopyranosyl glucoside crude product。Ethanol-water solution with 80% dissolves Quercetin-3-O-2 "; 6 "-two rhamanopyranosyl glucoside crude products, concentration is 50mg/mL, through filtering with microporous membrane, carries out two-dimensional liquid chromatography and prepares, chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm, 10 μ, Hua Puxinchuan Science and Technology Ltd.s), mobile phase selects ethanol (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, adopts 15% organic phase concentration eluting。Adopting DAD UV-detector 360nm to select absorbing wavelength, preparation temperature is 40 DEG C, and sample size is 200 μ L/ pins, flow rate of mobile phase is 60mL/min, collects retention time in 30-35min component, rotary evaporated to dryness, obtain Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucoside compounds。

Adopt high performance liquid chromatography that the Chinese medicine composition of the application is detected; mass percent with described Chinese medicine composition; Ginkgo total flavones 30%, bilobalide 8%, ginkgoic acid 1ppm; kaempferol-3-O-p-coumaroyl glucose rhamnoside 3.46%; Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides 3.14%, ginkalide A 2.6%; ginkalide B 1.8% and ginkalide C 1.3%, also detection has the chemical composition such as bilobalide and rutin。

The preparation of S4 ginkgo tree leave tincture

Embodiment 5

The preparation of kaempferol-3-O-p-coumaroyl glucose rhamnoside

Folium Ginkgo extract being dissolved in the ethanol-water solution of volumetric concentration 55%, prepares Folium Ginkgo extract solution, concentration is 550mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, type of elution 0-15min organic phase concentration is down to 90% gradient from 95% and is carried out, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is 30 DEG C, sample size is 2000 μ L/ pins, flow rate of mobile phase is 100mL/min, collect component and the remaining ingredient of 15～20 minutes, the components of collect 15～20 minutes are rotated evaporation and concentration to dry, kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product is prepared for one-dimensional。Methanol-water solution with 55% dissolves kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product; concentration is 60mg/mL; through filtering with microporous membrane; carry out two-dimensional liquid chromatography to prepare; chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm; 10 μm; Hua Puxinchuan Science and Technology Ltd.); mobile phase selects methanol (containing 0.1% formic acid) to be organic facies; water (containing 0.1% formic acid) is aqueous phase; adopting gradient elution mode, within 0-60 minute, organic phase concentration increases to 80% from 20%。DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is room temperature, sample size is 2000 μ L/ pins, flow rate of mobile phase is 90mL/min, collect the fraction of retention time 50-55min, rotary evaporated to dryness, obtain kaempferol-3-O-p-coumaroyl glucose rhamnoside compound, through liquid-phase chromatographic analysis, in the kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product of two dimension preparation, the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside is 98.2.0%, it is one-dimensional that to prepare the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside in kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product be 36%。

Quercetin-3-O-2 ", 6 " preparation of-two rhamanopyranosyl glucosides

By Folium Ginkgo extract 80g, being dissolved in the ethanol-water solution of volumetric concentration 55%, prepare Folium Ginkgo extract solution, concentration is 550mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, type of elution 0-15min organic phase concentration is down to 90% gradient from 95% and is carried out, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is 30 DEG C, sample size is 2000 μ L/ pins, flow rate of mobile phase is 100mL/min, collect component and the residual components of 28-32 minute, the component of 28-32 minute collected is rotated evaporation and concentration to dry, Quercetin-3-O-2 is prepared " for one-dimensional, 6 "-two rhamanopyranosyl glucoside crude product。Methanol-water solution with 55% dissolves Quercetin-3-O-2 "; 6 "-two rhamanopyranosyl glucoside crude products, concentration is 60mg/mL, through filtering with microporous membrane, carry out two-dimensional liquid chromatography to prepare, chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase selects acetonitrile (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, 0-60 minute organic facies 15% isocratic elution。Adopting DAD UV-detector 360nm to select absorbing wavelength, preparation temperature is room temperature, and sample size is 2000 μ L/ pins, flow rate of mobile phase is 90mL/min, collects the component of retention time 40-45min, rotary evaporated to dryness, obtain Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucoside compounds。

Adopt high performance liquid chromatography that the Chinese medicine composition of the application is detected; mass percent with described Chinese medicine composition; Ginkgo total flavones 35%, bilobalide 14%, ginkgoic acid 1ppm; kaempferol-3-O-p-coumaroyl glucose rhamnoside 3.21%; Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides 3.21%, ginkalide A 1.37%; ginkalide B 1.6% and ginkalide C 1.3%, also detection has the chemical composition such as bilobalide and rutin。

The preparation of S4 ginkgo tree leave tincture

Embodiment 6

The preparation of kaempferol-3-O-p-coumaroyl glucose rhamnoside

Folium Ginkgo extract being dissolved in the ethanol-water solution of volumetric concentration 60%, prepares Folium Ginkgo extract solution, concentration is 250mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, type of elution 0-15min organic phase concentration is down to 90% gradient from 95% and is carried out, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is 30 DEG C, sample size is 2000 μ L/ pins, flow rate of mobile phase is 100mL/min, collect component and the remaining ingredient of 15～20 minutes, the components of collect 15～20 minutes are rotated evaporation and concentration to dry, kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product is prepared for one-dimensional。Ethanol-water solution with 55% dissolves kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product; concentration is 100mg/mL; through filtering with microporous membrane; carry out two-dimensional liquid chromatography to prepare; chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm; 10 μm; Hua Puxinchuan Science and Technology Ltd.); mobile phase selects ethanol (containing 0.1% formic acid) to be organic facies; water (containing 0.1% formic acid) is aqueous phase; adopting gradient elution mode, within 0-60 minute, organic phase concentration increases to 80% from 20%。DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is room temperature, sample size is 2500 μ L/ pins, flow rate of mobile phase is 90mL/min, collect the fraction of retention time 50-55min, rotary evaporated to dryness, obtain kaempferol-3-O-p-coumaroyl glucose rhamnoside compound, through liquid-phase chromatographic analysis, in the kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product of two dimension preparation, the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside is 97.8%, it is one-dimensional that to prepare the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside in kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product be 36%。

Quercetin-3-O-2 ", 6 " preparation of-two rhamanopyranosyl glucosides

By Folium Ginkgo extract 50g, being dissolved in the ethanol-water solution of volumetric concentration 60%, prepare Folium Ginkgo extract solution, concentration is 250mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, type of elution 0-15min organic phase concentration is down to 90% gradient from 95% and is carried out, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is 30 DEG C, sample size is 2000 μ L/ pins, flow rate of mobile phase is 100mL/min, collect component and the residual components of 28-30 minute, the component of 28-30 minute collected is rotated evaporation and concentration to dry, Quercetin-3-O-2 is prepared " for one-dimensional, 6 "-two rhamanopyranosyl glucoside crude product。Ethanol-water solution with 55% dissolves Quercetin-3-O-2 "; 6 "-two rhamanopyranosyl glucoside crude products, concentration is 100mg/mL, through filtering with microporous membrane, carry out two-dimensional liquid chromatography to prepare, chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase selects acetonitrile (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, 0-60 minute organic facies 20% isocratic elution。Adopting DAD UV-detector 360nm to select absorbing wavelength, preparation temperature is room temperature, and sample size is 2500 μ L/ pins, flow rate of mobile phase is 90mL/min, collects the component of retention time 40-45min, rotary evaporated to dryness, obtain Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucoside compounds。

Adopt high performance liquid chromatography that the Chinese medicine composition of the application is detected; mass percent with described Chinese medicine composition; Ginkgo total flavones 36%, bilobalide 11%, ginkgoic acid 1ppm; kaempferol-3-O-p-coumaroyl glucose rhamnoside 2.98%; Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides 2.96%, ginkalide A 1.8%; ginkalide B 1.8% and ginkalide C 1.3%, also detection has the chemical composition such as bilobalide and rutin。

The preparation of S4 ginkgo tree leave tincture

Embodiment 7

The preparation of kaempferol-3-O-p-coumaroyl glucose rhamnoside

Folium Ginkgo extract being dissolved in the ethanol-water solution of volumetric concentration 45%, prepares Folium Ginkgo extract solution, concentration is 400mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol to be organic facies, water is aqueous phase, organic phase concentration is 90% degree such as grade, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is 40 DEG C, sample size is 2500 μ L/ pins, flow rate of mobile phase is 70mL/min, collect component and the remaining ingredient of 15-20 minute, the components of collect 15～20 minutes are rotated evaporation and concentration to dry, kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product is prepared for one-dimensional。Methanol-water solution with 50% dissolves kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product; concentration is 70mg/mL; through filtering with microporous membrane; carry out two-dimensional liquid chromatography to prepare; chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm; 10 μm; Hua Puxinchuan Science and Technology Ltd.); mobile phase selects methanol (containing 0.1% formic acid) to be organic facies; water (containing 0.1% formic acid) is aqueous phase; adopting isocratic elution mode, organic phase concentration is 20% totally 60 minutes。DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is room temperature, sample size is 2000 μ L/ pins, flow rate of mobile phase is 80mL/min, collect the fraction of retention time 50-55min, rotary evaporated to dryness, obtain kaempferol-3-O-p-coumaroyl glucose rhamnoside compound, through liquid-phase chromatographic analysis, in the kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product of two dimension preparation, the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside is 94.6%, it is one-dimensional that to prepare the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside in kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product be 34%。

Quercetin-3-O-2 ", 6 " preparation of-two rhamanopyranosyl glucosides

By Folium Ginkgo extract 20g, being dissolved in the ethanol-water solution of volumetric concentration 45%, prepare Folium Ginkgo extract solution, concentration is 400mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol to be organic facies, water is aqueous phase, organic phase concentration is 90% degree such as grade, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is 40 DEG C, sample size is 2500 μ L/ pins, flow rate of mobile phase is 70mL/min, collect component and the residual components of 30-32 minute, the component of 30-32 minute collected is rotated evaporation and concentration to dry, Quercetin-3-O-2 is prepared " for one-dimensional, 6 "-two rhamanopyranosyl glucoside crude product。Methanol-water solution with 50% dissolves Quercetin-3-O-2 "; 6 "-two rhamanopyranosyl glucoside crude products, concentration is 70mg/mL, through filtering with microporous membrane, carry out two-dimensional liquid chromatography to prepare, chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase selects acetonitrile (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, adopts isocratic elution mode, and organic phase concentration is 25% totally 60 minutes。Adopting DAD UV-detector 360nm to select absorbing wavelength, preparation temperature is room temperature, and sample size is 2000 μ L/ pins, flow rate of mobile phase is 80mL/min, collects the component of retention time 40-45min, rotary evaporated to dryness, obtain Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucoside compounds。

Adopt high performance liquid chromatography that the Chinese medicine composition of the application is detected; mass percent with described Chinese medicine composition; Ginkgo total flavones 39%, bilobalide 15%, ginkgoic acid 1ppm; kaempferol-3-O-p-coumaroyl glucose rhamnoside 2.62%; Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides 3.43%, ginkalide A 2.95%; ginkalide B 1.6% and ginkalide C 1.3%, also detection has the chemical composition such as bilobalide and rutin。

The preparation of S4 ginkgo tree leave tincture

Embodiment 8

The preparation of kaempferol-3-O-p-coumaroyl glucose rhamnoside

Folium Ginkgo extract being dissolved in the ethanol-water solution of volumetric concentration 70%, prepares Folium Ginkgo extract solution, concentration is 150mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol to be organic facies, water is aqueous phase, organic phase concentration is 95% degree such as grade, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is room temperature, sample size is 800 μ L/ pins, flow rate of mobile phase is 110mL/min, collect component and the remaining ingredient of 15-20 minute, the components of collect 15～20 minutes are rotated evaporation and concentration to dry, kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product is prepared for one-dimensional。Ethanol-water solution with 60% dissolves kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product; concentration is 200mg/mL; through filtering with microporous membrane; carry out two-dimensional liquid chromatography to prepare; chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm; 10 μm; Hua Puxinchuan Science and Technology Ltd.); mobile phase selects ethanol (containing 0.1% formic acid) to be organic facies; water (containing 0.1% formic acid) is aqueous phase; adopting isocratic elution mode, organic phase concentration is 15% totally 60 minutes。DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is room temperature, sample size is 2500 μ L/ pins, flow rate of mobile phase is 100mL/min, collect the fraction of retention time 50-55min, rotary evaporated to dryness, obtain kaempferol-3-O-p-coumaroyl glucose rhamnoside compound, through liquid-phase chromatographic analysis, in the kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product of two dimension preparation, the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside is 95.7%, it is one-dimensional that to prepare the content of kaempferol-3-O-p-coumaroyl glucose rhamnoside in kaempferol-3-O-p-coumaroyl glucose rhamnoside crude product be 33%。

Quercetin-3-O-2 ", 6 " preparation of-two rhamanopyranosyl glucosides

By Folium Ginkgo extract 100g, being dissolved in the ethanol-water solution of volumetric concentration 70%, prepare Folium Ginkgo extract solution, concentration is 150mg/mL, crosses 0.45 μm of microporous filter membrane, carries out one-dimensional liquid chromatograph and prepare。Hydrophilic chromatograph packing material (50 × 250mm that it is substrate with silica gel that one-dimensional liquid chromatograph adopts, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase adopts ethanol to be organic facies, water is aqueous phase, organic phase concentration is 95% degree such as grade, DAD UV-detector 360nm is adopted to select absorbing wavelength, preparation temperature is room temperature, sample size is 800 μ L/ pins, flow rate of mobile phase is 110mL/min, collect component and the residual components of 30-32 minute, the component of 30-32 minute collected is rotated evaporation and concentration to dry, Quercetin-3-O-2 is prepared " for one-dimensional, 6 "-two rhamanopyranosyl glucoside crude product。Ethanol-water solution with 60% dissolves Quercetin-3-O-2 "; 6 "-two rhamanopyranosyl glucoside crude products, concentration is 200mg/mL, through filtering with microporous membrane, carry out two-dimensional liquid chromatography to prepare, chromatographic column is with silica gel for substrate-bound C18 reverse phase filler (50 × 150mm, 10 μm, Hua Puxinchuan Science and Technology Ltd.), mobile phase selects acetonitrile (containing 0.1% formic acid) to be organic facies, water (containing 0.1% formic acid) is aqueous phase, adopts isocratic elution mode, and organic phase concentration is 20% totally 60 minutes。Adopting DAD UV-detector 360nm to select absorbing wavelength, preparation temperature is room temperature, and sample size is 2500 μ L/ pins, flow rate of mobile phase is 100mL/min, collects the component of retention time 40-45min, rotary evaporated to dryness, obtain Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucoside compounds。

Adopt high performance liquid chromatography that the Chinese medicine composition of the application is detected; mass percent with described Chinese medicine composition; Ginkgo total flavones 37%, bilobalide 14%, ginkgoic acid 1ppm; kaempferol-3-O-p-coumaroyl glucose rhamnoside 3.16%; Quercetin-3-O-2 ", 6 "-two rhamanopyranosyl glucosides 3.45%, ginkalide A 2.47%; ginkalide B 1.4% and ginkalide C 1.3%, also detection has the chemical composition such as bilobalide and rutin。

The preparation of S4 ginkgo tree leave tincture

Clinical experiment

This test specifies requirement according to " Kind protection system for CM regulations "; developed programs by expert technological guidance group; total Test adopts random controls open clinical trial, 3 medical institutions of Xiangya Hospital, Central-South China Univ., The Third Xiangya Hospital of Central South University and Lengshuijiang City institute of traditional Chinese medicine complete。

(1) diagnostic criteria

1, angina pectoris:

(1) can there be the rising of increased heart rate, blood pressure, anxiety, perspiration during angina pectoris attacks, sometimes audible and fourth heart sound, third heart sound or gallop rhythm, or apex systolic murmur occurs, paradoxical splitting of second heart sound, the double; two base of lung sound of even news。

(2) Electrocardioscopy electrocardiographic abnormality includes branch or rear fascicular block, left ventricular hypertrophy or arrhythmia etc. before ST section and the change of T ripple, atrioventricular block, bundle branch block, left bundle branch, and during panic attacks, electrocardiogram can be the change that typical ischemic ST force down

(3) can relief of symptoms person through rest or sublingual administration nitroglycerin。

2, cerebral infarction

(1), there is rapidly focal neurologic impairment sings and symptoms, continues more than 24h in unexpected onset；

(2) sings and symptoms of visible focal neurologic impairment, including hemiplegia, hemidysesthesia, hemianopsia, speech disorder, dizziness, dysphagia, ataxia etc.；

(3) neuroimaging inspection: in morbidity 24h, the conventional many nothings of skull CT scanning substantially change, can have infarcted region low-density to change after 24～48h；Head B-sonography scanning energy early discovery infarct, T, in low signal, T, in high signal, (DW [) and Perfusion weighted imaging (PWI) contribute to cerebral ischemic penumbra and judge diffusion-weighted imaging；Magnetic Resonance Angiography (MRA), CT blood vessel imaging (cTA) etc. check, to the clear and definite cause of disease or determine that the position of obturation or narrow blood vessel is useful。

(2) case selection standard

1, case standard is included in

All meet angina pectoris, cerebral embolism diagnosis belong to syndrome of blood stasis pattern of syndrome person, for including object in。

2, Excluded cases standard

(1) the severe primary diseases such as the above hypertension of moderate, severe pulmonary insufficiency, severe arrhythmia, liver, kidney, hemopoietic system, psychotic are merged；

(2) gestation or women breast-feeding their children and allergic constitution person。

(3) do not take medicine by regulation, it is impossible to judge curative effect or data not congruent affect the treatment judgement or safety judgement person。

(3) administrated method and requirement

1, administrated method.

(1) treatment group:

Angina pectoris:

Give each 2ml of Folium Ginkgo tincture that embodiment 1 prepares, 3 times on the 1st, serve on 4 weeks, sorbide nitrate can be taken simultaneously, enteric coated aspirin, during angina pectoris attacks can buccal nitroglycerin temporarily, the experimenter that is in hospital can give Low molecular heparin, Nitro-Bid Ⅳ etc. if desired。In addition, do not take the medicative other drug of primary disease。

Acute cerebral infarction:

Give each 2ml of Folium Ginkgo tincture that embodiment 2 prepares, 3 times on the 1st, serve on 4 weeks。Observation period two groups all gives enteric coated aspirin, and the experimenter that is in hospital can use mannitol dehydration to alleviate cerebral edema according to the state of an illness if desired, it is prevented that soda acid Electrolyte imbalance, neuronal differentiation agent, intensive care, it is prevented that complication etc.。In addition, do not take the medicative other drug ischemic cerebrovascular of primary disease:

Treatment group gives each 2ml of Folium Ginkgo tincture that embodiment 3 prepares, 3 times on the 1st, serve on 4 weeks, simultaneously Aspirin sheet and neuroprotective, blood pressure lowering, tune fat, control cranium pressing thing, and anticoagulant therapy should use as one sees fit。

(2) matched group angina pectoris:

(Yangtze River, Jiangsu Pharmaceutical produces to give Folium Ginkgo, lot number: 07042901) each 2,3 times on the 1st, it serve on 4 weeks, sorbide nitrate can be taken simultaneously, enteric coated aspirin, during angina pectoris attacks can buccal nitroglycerin temporarily, the experimenter that is in hospital can give Low molecular heparin, Nitro-Bid Ⅳ etc. if desired。In addition, do not take the medicative other drug of primary disease。

Acute cerebral infarction:

Give each 2 of Folium Ginkgo (Yangtze River, Jiangsu Pharmaceutical produces, lot number: 07042901), 3 times on the 1st, serve on 4 weeks。Observation period two groups all gives enteric coated aspirin, and the experimenter that is in hospital can use mannitol dehydration to alleviate cerebral edema according to the state of an illness if desired, it is prevented that soda acid Electrolyte imbalance, neuronal differentiation agent, intensive care, it is prevented that complication etc.。In addition, do not take the medicative other drug of primary disease。

Ischemic cerebrovascular

Giving each 2 of Folium Ginkgo (Yangtze River, Jiangsu Pharmaceutical produces, lot number: 07042901), 3 times on the 1st, serve on 4 weeks, simultaneously Aspirin sheet and neuroprotective, blood pressure lowering, tune fat, control cranium pressing thing, anticoagulant therapy should use as one sees fit。

(4) observation index and detection project

1, observation of curative effect:

(1) clinical symptoms, sign, picture of the tongue change etc.。

(2) routine blood test, routine urinalysis, electrolyte, blood fat, hepatic and renal function, hemorheology (includes whole blood viscosity, Plasma Viscosity, erythrocyte aggregation, red cell deformability etc.)。

2, safety observation:

(1) general physical examination project。

(2) routine blood test, routine urinalysis, just routine examination (respectively looking into 1 time before and after treatment) before and after treatment。

(5) stopped treatment test indication

(1) midway is uncooperative, lost to follow-up or died。

(2), during treatment, add with the Chinese and western drugs having similar effect with this reagent。

(3) in medication process, because having bleeding tendency and serious bad reflection, anaphylaxis and drug withdrawal person。

(6) efficacy assessment standard

Angina pectoris: curative effect is according to " new Chinese medicine guideline of clinical investigations "

Effective: angina pectoris symptom disappears or substantially disappears；

Effective: angina pectoris attacks number of times reduces 50%, and attack degree and persistent period substantially alleviate；

Invalid: symptom is basic identical with treating current symptom

Acute cerebral infarction: the standard passed through with reference to the 4th national cerebrovascular meeting, observes before treatment and hemorheology index and neurological deficits score and disability degree after treatment 14d。Efficacy evaluation is divided into

Basic healing: functional impairment scoring minimizing 91%～100%, disability degree is 0 grade:

Marked improvement: functional impairment scoring minimizing 46%～90%, disability degree is 1～3 grade:

Effective: functional impairment scoring minimizing 18%～45%, disability degree is 4 grades:

Invalid: functional impairment scoring reduces≤17%, and disability degree is 5 grades:

(5) worsen: neurological deficits score reduces or increases more than 18%。

Ischemic cerebrovascular: with reference to " new Chinese medicine guideline of clinical investigations ": clinical efficacy evaluation criteria:

It is almost recovered: functional impairment scoring reduces 90%-100%, disability degree 0 grade；

Marked improvement: functional impairment scoring reduces 46%-89%, disability degree 1-3 level；

Progressive: functional impairment scoring reduces 18%-45%；

Unchanged: functional impairment scoring reduces or increases within 18%；

Worsen: functional impairment scoring increases by more than 18%；

(7) observation of untoward reaction。

Record faithfully various untoward reaction and time of occurrence thereof, the order of severity and processing method,

Effect。(8) statistical method:。

Adopt SSPS/PC+ software, relevant data are carried out paired data t inspection and X 2 test。

Three, result of the test

Off-test is data collection 564 example altogether, does not meet case 8 example of inclusion criteria, drops by the wayside 15 examples。Curative effect statistics case load is 303 examples, clinical control 238 example。

(1) physical data

Angina pectoris

This laboratory observation treatment group (n=138), male 76 example, women 62 example, at 51.6 ± 5.6 years old age, the course of disease 4.5 ± 2.3 years, including stable angina pectoris 90 example, unstable angina pectoris 48 example；Matched group (n=106), male 64 example, women 42 example, 55.1 ± 6.2 years old age, the course of disease 4.2 ± 1.9 years, including stable angina pectoris 72 example, the aspect such as unstable angina pectoris 34 example two groups of ages, sex, the state of an illness, courses of disease there was no significant difference (P > 0.05) through statistical procedures, has comparability。

Acute cerebral infarction

Treatment group: treatment group (n=58), male 34 example, women 24 example, at 58.5 ± 4.6 years old age, wherein, blocking part: 31 examples are blocked in basal ganglia region, frontal lobe blocks 14 examples, cerebellar infarction 12 example, infarction of occipital lobe 2 example；Matched group (n=47), male 28 example, women 19 example, 57.9 ± 3.8 years old age, wherein blocking part: 26 examples are blocked in basal ganglia region, frontal lobe blocks 11 examples, cerebellar infarction 9 example, infarction of occipital lobe 1 example；The aspects such as two groups of ages, sex, the state of an illness, courses of disease there was no significant difference (P > 0.05) through statistical procedures, has comparability。

Ischemic cerebrovascular

Treatment group: treatment group (n=107), male 59 example, women 48 example, 55.9 ± 7.2 years old age；Matched group (n=85), male 51 example, women 34 example, 55.1 ± 6.2 years old age, the course of disease 4.2 ± 1.9 years, the aspects such as two groups of ages, sex, the state of an illness, the course of disease, CT scoring, Clinical Nerve Function Deficiency scorings there was no significant difference (P > 0.05) through statistical procedures, has comparability。

(2) curative effect statistics

Angina pectoris

1. before and after treatment group and treatment of control group, curative effect to treat angina pectoris compares, in Table 1

1 liang of table group curative effect to treat angina pectoris compares

As seen from the results in Table 1: effective 118 examples of curative effect to treat angina pectoris treatment group (85.5%), effective 20 examples (14.5%), invalid 0 example, total effective rate 100%；Effective 90 examples of matched group (84.9%), effective 15 examples (14.1%), invalid 1 example (0.9%), total effective rate is 99.1%。Two groups of therapeutic equivalences, no significant difference。

Before and after 2 liang of group treatments, ECG Change compares, in Table 2

Table 2 ECG Change compares

As seen from the results in Table 2, effective 72 examples of ECG curative effect treatment group (52.6%), effective 50 examples (35.9%), invalid 16 examples (11.5%), total effective rate 88.5%；Effective 49 examples of matched group (46.7%), effective 37 examples (35%), invalid 20 examples (18.3%), total effective rate 81.7%。Two groups of Different therapeutical effects are statistically significant。

Hemorheology change before and after 3 liang of group treatments

Hemorheology change before and after the group treatment of 3 liang of table

By table 3 result: compare before and after treatment, whole blood viscosity, Plasma Viscosity, packed cell volume are the difference comparsion before and after P<0.05, treatment group and treatment of control group, whole blood viscosity, P>0.05；Plasma Viscosity, P > 0.05；Packed cell volume P > 0.05。Comparing before and after visible treatment group and treatment of control group, whole blood viscosity, Plasma Viscosity, packed cell volume all substantially reduce。Treatment group and matched group compare, and the degree that after two groups of treatments, whole blood viscosity, Plasma Viscosity, packed cell volume reduce is suitable, no significant difference。

Acute cerebral infarction

1, comparitive study before and after two groups for the treatment of acute cerebral infarctions, in Table 4

4 liang of table group curative effect to treat angina pectoris compares

Before and after two groups of treatments, hemorheology changes in Table 5

Hemorheology change before and after the group treatment of 5 liang of table

By table 5 result: comparing before and after treatment, whole blood viscosity, Plasma Viscosity, Fibrinogen, P < 0.05, it is seen that compare before and after treatment group and treatment of control group, whole blood viscosity, Plasma Viscosity, Fibrinogen all substantially reduce, and its difference has statistical significance。

Ischemic cerebrovascular

Tcm syndrome Clinical efficacy comparison before and after two groups of treatments, in Table 6

6 liang of table group clinical efficacy improves situation

As seen from the results in Table 6: curative effect to treat angina pectoris treatment group is almost recovered 25 examples, marked improvement 42 example, progressive 35 examples, unchanged 3 examples, worsen 2 examples, total effective rate 96%；Matched group is almost recovered 22 examples, marked improvement 33 example, progressive 25 examples, and unchanged 2 examples worsen 3 examples, total effective rate 94.1%。Two groups of therapeutic equivalences, no significant difference。

Before and after two groups of treatments, neurological functional deficit improvement situation compares, in Table 7

Table 7 neurological functional deficit is improved situation and is compared (* two groups self compare p < 0.01)

Tcm symptom scoring change before and after two groups of treatments, in Table 8

Table 8 cures symptom score and total mark change

By table 8 result: compare before and after treatment, numb limbs and tense tendons, hemiplegia, facial hemiparalysis, language blocking P < 0.05, it is seen that compare before and after treatment group and treatment of control group, numb limbs and tense tendons, hemiplegia, facial hemiparalysis, language blocking symptom is all obviously improved, and its difference has statistical significance。

Obviously, above-described embodiment is only for clearly demonstrating example, and is not the restriction to embodiment。For those of ordinary skill in the field, can also make other changes in different forms on the basis of the above description。Here without also cannot all of embodiment be given exhaustive。And the apparent change thus extended out or variation are still among the protection domain of the invention。

Claims

1. a Folium Ginkgo tincture, including the Chinese medicine composition formed by Folium Ginkgo extract with for dissolving the ethanol water of described Chinese medicine composition, it is characterized in that, to account for the mass percent of described Chinese medicine composition, described Chinese medicine composition includes, the Ginkgo total flavones of 24-40%, the bilobalide of 6-16%, ginkgoic acid is less than 5ppm。

2. Folium Ginkgo tincture according to claim 1; it is characterized in that, described Chinese medicine composition includes, the kaempferol-3-O-p-coumaroyl glucose rhamnoside of 2.08-3.46%; Quercetin-the 3-O-2 of 2.11-3.53% ", 6 "-two rhamanopyranosyl glucosides。

3. Folium Ginkgo tincture according to claim 1 and 2, it is characterised in that described Chinese medicine composition includes, the ginkalide A of 1.4-3%, the ginkalide B of 0.9-1.8% and the ginkalide C of 1.2-1.3%。

4. according to the arbitrary described Folium Ginkgo tincture of claim 1-3, it is characterised in that described Chinese medicine composition also includes bilobalide and rutin。

5. according to the arbitrary described Folium Ginkgo tincture of claim 1-4, it is characterised in that the preparation method of described Chinese medicine composition includes,

The preparation method of S1 kaempferol-3-O-p-coumaroyl glucose rhamnoside,

6. Folium Ginkgo tincture according to claim 6, it is characterised in that

7. according to claim 5 or 6 Folium Ginkgo tincture, it is characterised in that

In described step (2), one-dimensional liquid chromatograph separates, and organic facies is possibly together with formic acid, and formic acid volumetric concentration is 0.1%；Possibly together with formic acid in aqueous phase, formic acid volumetric concentration is 0.1%；

8. Folium Ginkgo tincture according to claim 7, it is characterised in that

In described step (c), one-dimensional liquid chromatograph separates, and organic facies is possibly together with formic acid, and formic acid volumetric concentration is 0.1%；Possibly together with formic acid in aqueous phase, formic acid volumetric concentration is 0.1%；

9. a preparation method for the arbitrary described Folium Ginkgo tincture of claim 1-8, including,

The Chinese medicine composition taking the formation of described Folium Ginkgo extract is dissolved in the ethanol water that volumetric concentration is 60%, to obtain final product, and the mass volume ratio of wherein said Chinese medicine composition and described ethanol water is 0.4g/10ml。

10. the preparation method of Folium Ginkgo tincture according to claim 9, it is characterised in that also include the step adding adjuvant。