CN108373474B - A bilobalide compound extracted from folium Ginkgo and its preparation method - Google Patents
A bilobalide compound extracted from folium Ginkgo and its preparation method Download PDFInfo
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Abstract
The invention relates to a ginkgolide compound extracted from folium ginkgo and a preparation method thereof, the ginkgolide compound is prepared by taking folium ginkgo as a raw material, extracting an extracted folium ginkgo extract by using an organic solvent, and separating and purifying by RP-18 column chromatography. The ginkgolide compound is named as ginkgolide HK, and the structural formula of the ginkgolide compound is as follows:
Description
Technical Field
The invention relates to a traditional Chinese medicine, in particular to a chemical component monomer of the traditional Chinese medicine and an extraction, separation and purification method, and especially relates to a ginkgolide compound extracted from ginkgo leaves and a preparation method thereof.
Background
Ginkgo biloba extract has been widely used as an active ingredient in pharmaceuticals. The active ingredients currently isolated from ginkgo biloba are mainly diterpene lactones and sesquiterpene lactones, namely bilobalide A, B, C, J, M and bilobalide. Since the 80 s in the 20 th century, researches have found that ginkgolides are strong antagonists of Platelet Activating Factor (PAF), are considered to be natural PAF receptor antagonists with the most clinical application prospect, have protection effects on ischemic injury and the central nervous system, and have remarkable curative effects on cardiovascular and cerebrovascular diseases. The inventor further discovers that an unknown component is extracted and separated in the process of preparing the ginkgo total lactone, and the content of the unknown component is between 0.1 and 0.8 percent, so the invention carries out extraction, separation and purification on the unknown component, carries out structural identification, carries out comparative analysis on the identification result and a reference document (Weinges K, Rummler M, SchickH.Herstellung von 1, 10-Dihydroxy-und 1,7, 10-trihydroxygiloid aus 1,3,7, 10-tetrahydroxygiloid. Liebigs Ann chem.1993,1023-1027.), and finally determines that the compound is 1, 10-dihydroxygiloid (1,10-Dihydroxy ginkgolide). The existing documents only report that main lactone components such as ginkgolide A, B, C and bilobalide are extracted and identified from ginkgo leaves, but no document reports about the research of extracting and identifying the unknown components from the ginkgo leaves, so that the further research has positive significance.
Disclosure of Invention
The invention aims to solve the technical problem of extracting and identifying unknown components in the ginkgo leaf extract from ginkgo leaves.
The invention provides a ginkgolide compound extracted from folium ginkgo.
The Ginkgolide compound identifies the structure through spectrum, mass spectrum and the like, determines that the chemical structure of the Ginkgolide compound is 1,10-dihydroxy Ginkgolide (1, 10-dihydroyginggolid), is named as Ginkgolide HK (Ginkgolide HK, GHK for short), and has the structural formula:
mass Spectrometric (MS) detection of ginkgolide HK compounds: the instrument is an Agilent LCMSD liquid chromatograph-mass spectrometer; the test conditions were: ESI 0.4 ml/min; the SCAN is m/z 100-1000, and a Positive mode; the ginkgolide HK Mass Spectrum (MS) result showed that the molecular weight was 408.
Ginkgolide HK was confirmed to be identical to the compound 1, 10-dihydroxyginkgolide.
Optical chromatography (ORD)) assay of ginkgolide HK compounds: the instrument is a RUDOLPH Autopol IV polarimeter; and (3) testing conditions are as follows: acetonitrile solution with the sample concentration of 1mg/mL, 25 ℃; the test result was [ alpha ] D25: -68 ° (c,0.1, Acetonitril).
Ginkgolide HK was confirmed to be identical to the compound 1, 10-dihydroxyginkgolide.
1H-nuclear magnetic resonance and 13C-nuclear magnetic resonance detection of ginkgolide HK compound: the instrument is a Bruker 600MHz NMR instrument, the solvent is DMSO-d6, and the NMR data of the hydrocarbon are shown in the table I.
TABLE I comparison of Hydrocarbon NMR data for the ginkgolide compound of the invention (ginkgolide HK compound) and the compound 1, 10-dihydroxyginkgolide (1, 10-dihydroginkgolid)
(DMSO-d6in 600MHz)
The detection confirms that the ginkgolide compound (ginkgolide HK compound) is the compound 1,10-dihydroxy ginkgolide.
The bilobalide compound (bilobalide HK) does not contain bilobalide (shown in figure 1). The content of the ginkgolide compound is more than or equal to 99 percent, and preferably the content of the ginkgolide compound is more than or equal to 99.5 percent.
Another objective of the present invention is to provide a method for preparing a ginkgolide compound (ginkgolide HK) extracted from ginkgo leaves.
The method comprises the following steps:
(1) extracting folium ginkgo three times by refluxing with an ethanol water solution, wherein the dosage of the ethanol water solution is 5-8 times of the weight of the medicinal materials, concentrating the extracting solution under reduced pressure until no ethanol smell exists, adding water, standing for clarification, filtering, passing the filtrate through macroporous adsorption resin, eluting, and concentrating under reduced pressure to obtain a folium ginkgo extract;
(2) dissolving folium Ginkgo extract in water, adding ethyl acetate or methyl acetate or their mixture, extracting, mixing extractive solutions, and concentrating under reduced pressure to dry to obtain total bilobalide extract;
(3) heating and dissolving the total ginkgolide extract with a solvent, cooling, standing, crystallizing, filtering, washing a filter cake with the solvent, and drying to obtain a crude product of the total ginkgolide;
(4) heating and refluxing the crude product of ginkgolide with ethyl acetate, methyl acetate or their mixture to dissolve completely, concentrating under reduced pressure until crystallization occurs, standing for crystallization, vacuum filtering, and concentrating the filtrate to dryness to obtain crude product of ginkgolide compound (ginkgolide HK);
(5) and (2) purifying the crude product of the ginkgolide compound (ginkgolide HK) by using a reverse phase column chromatography for 1-2 times, wherein the mobile phase is a mixed solution of methanol and water, the proportion range of the methanol is 10% -90%, isocratic elution or gradient elution is carried out, the wavelength of ultraviolet detection is 220nm, the effluent of the ginkgolide compound (ginkgolide HK) is collected, reduced pressure concentration is carried out until no solvent exists, and drying is carried out, so as to obtain the ginkgolide compound (ginkgolide HK).
The concentration of the ethanol water solution in the step (1) is 50-80% (V/V).
The amount of the ginkgo biloba extract dissolved in the water in the step (2) is 3-5 times of the weight of the ginkgo biloba extract, the organic solvent used for extraction is ethyl acetate, methyl acetate or a mixed solution of ethyl acetate and methyl acetate, the extraction volume is 0.5-1.5 times of the volume of the liquid medicine (V/V), preferably 0.8-1.2 times of the volume of the liquid medicine (V/V), and the extraction times are 1-4 times, preferably 3 times.
The solvent used for dissolving the total ginkgolide extract in the step (3) is 30-60% of aqueous methanol or aqueous ethanol, the using amount of the solvent is 0.5-1.5 times of the weight of the total lactone extract, preferably 0.8-1.2 times of the weight of the total lactone extract, the mixture is heated to 60 ℃ for dissolving, the standing crystallization time is 8-16 h, the standing crystallization temperature is 0-10 ℃, preferably 3-5 ℃, the crystal washing solvent is 30-60% of aqueous methanol or ethanol, and the crystal washing frequency is 0-4 times, preferably 1-2 times; the drying temperature is 55-65 ℃.
The solvent used for dissolving the crude product of the total ginkgolides in the step (4) is ethyl acetate, methyl acetate or a mixed solution of the ethyl acetate and the methyl acetate, and the mixture is concentrated under reduced pressure until the weight ratio of the liquid medicine to the crude product of the total ginkgolides is 10: 1-15: 1, the repeated operation times are 1-2 times.
And (3) purifying the ginkgolide compound (ginkgolide HK) by using a column chromatography in the step (5), wherein the column chromatography is used as a filler RP-18, the mobile phase is a mixed solution of methanol and water, the proportion range of the methanol is 10% -90%, isocratic elution or gradient elution is carried out, and the reduced pressure vacuum drying time at 80 ℃ is 2-4 hours.
The pressure of the steps (1), (2), (4) and (5) of the method for concentrating under reduced pressure is-0.08 to-0.1 MPa.
The ginkgolide compound obtained by the method is ginkgolide HK.
The invention is to extract, separate and purify ginkgolide compound (ginkgolide HK (GHK)) from folium ginkgo for the first time, and the compound is confirmed to be 1,10-dihydroxy ginkgolide through Mass Spectrum (MS) detection, optical rotation chromatography (ORD) detection and hydrocarbon nuclear magnetic resonance detection. The preparation method provided by the invention is simple to operate, economical and practical, and suitable for industrial production.
Drawings
FIG. 1: inventive example 4 HPLC-ELSD detection spectrum of ginkgolide compound (ginkgolide HK)
FIG. 2: EXAMPLE 4 Hydrogen Spectroscopy of ginkgolide Compound (ginkgolide HK) in example 4 of the present invention
FIG. 3: example 4 carbon spectra of ginkgolide Compound (ginkgolide HK) of the present invention
Detailed Description
For a further understanding of the present invention, reference will now be made in detail to the following examples.
The invention will be more readily understood by reference to the following examples, which are given to illustrate the invention, but are not intended to limit the scope of the invention.
The ginkgo biloba leaves and the reagents used in the examples of the present invention are commercially available, unless otherwise specified.
Example 1
Taking 500kg of dry folium Ginkgo (sold in market), crushing, extracting with 80% ethanol under reflux for three times, sequentially extracting with 4000L, 2500L and 2500L of solvent for 2h each time, mixing extractive solutions, concentrating at 60 deg.C under reduced pressure (-0.09MPa) until no ethanol smell exists, adding 1000L of purified water, standing, filtering, passing filtrate through macroporous adsorbent resin column (D101), washing with 1500L of purified water to remove impurities, eluting with 70% ethanol (2000L) until the color of eluate becomes lighter, concentrating the eluate at 60 deg.C under reduced pressure (-0.09MPa), oven drying at 60 deg.C for 4 hr to obtain folium Ginkgo extract (10.5kg), and detecting with HPLC-ELSD (high performance liquid phase-evaporative light scattering detector) that the content of total ginkgolides is 8.1%.
Example 2
Taking 500kg of dry ginkgo leaves (sold in market), crushing, carrying out reflux extraction with 70% ethanol for three times, sequentially carrying out extraction with solvent dosage of 4000L, 2500L and 2500L for 2h each time, combining extracting solutions, concentrating at 60 ℃ under reduced pressure (-0.09MPa) until no ethanol smell exists, adding 1000L of purified water, standing, filtering, introducing filtrate into a macroporous adsorption resin column (D101), washing with purified water (1500L) to remove impurities, eluting with 65% ethanol (2200L) until the color of eluent becomes light, concentrating the eluent at 60 ℃ under reduced pressure, drying at 60 ℃ for 4 hours to obtain a ginkgo leaf extract (10.6kg), and detecting the content of total ginkgolides by HPLC-ELSD to be 7.5%.
Example 3
Taking 1kg of ginkgo biloba extract, adding 5L of water, stirring at room temperature until the ginkgo biloba extract is completely dissolved, adding ethyl acetate, extracting for 3 times, wherein each time the ethyl acetate uses 4L, combining ethyl acetate extract, concentrating at 60 ℃ under reduced pressure (-0.09MPa) until no solvent exists, weighing 168.5g, adding 170mL of 50% ethanol, heating and refluxing until the ethyl acetate is completely dissolved, standing and refrigerating at 3 ℃ for 10 hours, filtering, washing a filter cake with 50% ethanol 100mL for 1 time, drying at 60 ℃ and weighing 17.2 g, adding 1700mL of ethyl acetate, heating and refluxing until the ethyl acetate is completely dissolved, concentrating at 60 ℃ under reduced pressure (-0.09MPa) until the weight of the liquid medicine is 190g, standing and refrigerating at 3 ℃ for 8 hours, filtering, collecting a filtrate, concentrating at 60 ℃ under reduced pressure (-0.09MPa) until no solvent exists, weighing 7.5g, purifying by RP-18 column chromatography, eluting with a flow rate of 5-10mL/min and a mobile phase of 40% methanol water, collecting bilobalide HK eluate, concentrating at 60 deg.C under reduced pressure (-0.09MPa) to dryness, repeatedly purifying the obtained compound under the same chromatographic conditions, collecting bilobalide HK eluate, concentrating at 60 deg.C under reduced pressure (-0.09MPa) to dryness, vacuum drying at 80 deg.C under reduced pressure for 2 hr, weighing to obtain bilobalide HK 62.5mg, and detecting by HPLC-ELSD to obtain product with purity of 99.64%.
The product is proved to be consistent with the compound 1,10-dihydroxy ginkgolide by Mass Spectrum (MS), optical rotation chromatography (ORD), 1H-nuclear magnetic resonance and 13C-nuclear magnetic resonance detection.
Example 4
Taking 10kg of ginkgo biloba extract, adding 50L of water, stirring until the ginkgo biloba extract is completely dissolved, adding ethyl acetate for extraction for 4 times, wherein the volume of ethyl acetate used each time is 50L, combining ethyl acetate extract, concentrating under reduced pressure (-0.09MPa) at 60 ℃ until no solvent exists, weighing 1.803kg, adding 1.8L of 40% ethanol, heating and refluxing until the ethyl acetate is completely dissolved, standing and refrigerating at 4 ℃ for 15 hours, performing suction filtration, washing a filter cake with 40% ethanol for 2 times, drying at 60 ℃, weighing 176.5g, adding 18L of ethyl acetate, heating and refluxing until the ethyl acetate is completely dissolved, concentrating under reduced pressure (-0.09MPa) at 60 ℃ until the weight of the liquid medicine is 1905g, standing and refrigerating for 12 hours at 4 ℃, performing suction filtration, collecting a filtrate, concentrating under reduced pressure (-0.09MPa) at 60 ℃ until no solvent exists, weighing 69.2g, continuously adding 7000 mL of ethyl acetate, heating and refluxing until the ethyl acetate is completely dissolved, concentrating under reduced pressure (-0.09MPa), standing at 4 deg.C, refrigerating for 8 hr, vacuum filtering, collecting filtrate, concentrating at 60 deg.C under reduced pressure (-0.09MPa) until no solvent exists, weighing 30.7g, purifying with RP-18 column chromatography once at flow rate of 5-10mL/min, eluting at constant rate with methanol and water mixed solution as mobile phase, methanol ratio of 50%, collecting bilobalide compound (bilobalide HK) effluent, concentrating at 60 deg.C under reduced pressure (-0.09MPa) to dry, vacuum drying at 80 deg.C under reduced pressure (-0.09MPa) for 4 hr, weighing to obtain bilobalide HK650mg, and detecting by HPLC-ELSD with purity of 98.1%. The obtained compound is further purified repeatedly under the same chromatographic conditions once to obtain ginkgolide HK 590mg with purity of 99.87% by HPLC-ELSD detection. (see the attached drawing)
The product is proved to be consistent with the compound 1,10-dihydroxy ginkgolide by Mass Spectrum (MS), optical rotation chromatography (ORD), 1H-nuclear magnetic resonance and 13C-nuclear magnetic resonance detection. (see the attached drawings and the table I)
Example 5
Taking 2kg of folium Ginkgo extract, adding 6L of water, stirring to dissolve completely, adding ethyl acetate, extracting for 3 times, each time using 5L of ethyl acetate, combining ethyl acetate extract, concentrating at 60 deg.C under reduced pressure (-0.09MPa) until no solvent exists, weighing 335.2g, adding 335 mL of 50% ethanol, heating and refluxing until complete dissolution, standing at 5 deg.C, refrigerating for crystallization for 16 hr, filtering, washing filter cake with 50% ethanol for 1 time, drying at 60 deg.C, weighing 35.8g, adding 3500mL of methyl acetate, heating and refluxing until complete dissolution, concentrating at 60 deg.C under reduced pressure (-0.09MPa) until the weight of medicinal liquid is 400g, standing at 5 deg.C, refrigerating for crystallization for 8 hr, filtering, collecting filtrate, concentrating at 60 deg.C under reduced pressure (-0.09MPa) until no solvent exists, weighing 16.4g, repeatedly operating at RP-18 column chromatography for 2 times, flow rate of 5-10mL/min, mobile phase is methanol and water mixed solution, gradient eluting with 30-75% methanol solution for 0-30 min, eluting with 75% methanol water solution at 30-60min, collecting bilobalide HK eluate, concentrating at 60 deg.C under reduced pressure (-0.09MPa) to dry, vacuum drying at 80 deg.C under reduced pressure (-0.09MPa) for 3 hr, weighing to obtain 115.8mg of bilobalide compound (bilobalide HK), and detecting by HPLC-ELSD to obtain product with purity of 99.84%.
The product is proved to be consistent with the compound 1,10-dihydroxy ginkgolide by Mass Spectrum (MS), optical rotation chromatography (ORD), 1H-nuclear magnetic resonance and 13C-nuclear magnetic resonance detection.
Example 6
Taking 500g of ginkgo biloba extract, adding 2.5L of water, stirring until the ginkgo biloba extract is completely dissolved, adding ethyl acetate for extraction for 3 times, wherein the volume of ethyl acetate used each time is 3L, combining ethyl acetate extract, concentrating at 60 ℃ under reduced pressure (-0.09MPa) until no solvent exists, weighing 85.2g, adding 85mL of 40% ethanol, heating and refluxing until complete dissolution, standing at 3 ℃ for refrigeration and crystallization for 16 hours, performing suction filtration, washing a filter cake with 40% ethanol for 2 times, drying, weighing 9.1g, adding 900mL of methyl acetate, heating and refluxing until complete dissolution, concentrating at 60 ℃ under reduced pressure (-0.09MPa) until the weight of the liquid medicine is 120g, standing at 3 ℃ for refrigeration and crystallization for 10 hours, performing suction filtration, collecting filtrate, concentrating at 60 ℃ under reduced pressure (-0.09MPa) until no solvent exists, weighing 3.5g, repeatedly performing RP-18 column chromatography for 2 times, wherein the flow rate is 5-10mL/min, and the mobile phase is a mixed solution of, isocratic elution with methanol ratio of 60%, collecting bilobalide HK eluate, concentrating at 60 deg.C under reduced pressure (-0.09MPa) to dryness, vacuum drying at 80 deg.C under reduced pressure (-0.09MPa) for 2 hr, weighing to obtain 29.7mg of bilobalide compound (bilobalide HK), and detecting by HPLC-ELSD to obtain product with purity of 99.74%.
The product is proved to be consistent with the compound 1,10-dihydroxy ginkgolide by Mass Spectrum (MS), optical rotation chromatography (ORD), 1H-nuclear magnetic resonance and 13C-nuclear magnetic resonance detection.
Example 7
Taking 5kg of ginkgo biloba extract, adding 25L of water, stirring until the ginkgo biloba extract is completely dissolved, adding ethyl acetate for extraction for 4 times, wherein the volume of ethyl acetate used in each time is 25L, combining ethyl acetate extract, concentrating at 60 ℃ under reduced pressure (-0.09MPa) until no solvent exists, weighing 840.2g, adding 840mL of 50% ethanol, heating and refluxing until the ethyl acetate is completely dissolved, standing and refrigerating at 4 ℃ for crystallization for 16 hours, performing suction filtration, washing a filter cake with 50% ethanol for 1 time, drying at 60 ℃, weighing 88.4g, adding 8000mL of ethyl acetate, heating and refluxing until the ethyl acetate is completely dissolved, concentrating at 60 ℃ under reduced pressure (-0.09MPa) until the weight of the liquid medicine is 1000g, standing and refrigerating and crystallizing at 4 ℃ for 10 hours, performing suction filtration, collecting filtrate, concentrating at 60 ℃ under reduced pressure (-0.09MPa) until no solvent exists, weighing 37.5g, continuously adding 3700mL of ethyl acetate, heating and refluxing until the ethyl acetate is completely dissolved, concentrating at 60 ℃ under, standing at 4 deg.C, refrigerating for 10 hr, vacuum filtering, collecting filtrate, concentrating at 60 deg.C under reduced pressure (-0.09MPa) until no solvent exists, weighing 15.9g, repeatedly performing RP-18 column chromatography for 2 times at flow rate of 5-10mL/min, eluting with methanol and water mixed solution at constant rate with methanol ratio of 40%, collecting bilobalide HK eluate, concentrating at 60 deg.C under reduced pressure (-0.09MPa) until dried, vacuum drying at 80 deg.C under reduced pressure (-0.09MPa) for 3 hr, weighing to obtain 310.5mg of bilobalide compound (bilobalide HK), and testing by HPLC-ELSD to obtain product with purity of 99.80%.
The product is proved to be consistent with the compound 1,10-dihydroxy ginkgolide by Mass Spectrum (MS), optical rotation chromatography (ORD), 1H-nuclear magnetic resonance and 13C-nuclear magnetic resonance detection.
Claims (8)
1. A preparation method of a ginkgolide compound extracted from folium ginkgo is characterized in that the ginkgolide compound is extracted from folium ginkgo, the content of the ginkgolide compound is more than or equal to 99%, the ginkgolide compound is named as ginkgolide HK, and the structural formula of the ginkgolide HK is as follows:
the preparation method comprises the following steps:
(1) extracting folium ginkgo three times by refluxing with an ethanol water solution, wherein the dosage of the ethanol water solution is 5-8 times of the weight of the medicinal materials, concentrating the extracting solution under reduced pressure until no ethanol smell exists, adding water, standing for clarification, filtering, passing the filtrate through macroporous adsorption resin, eluting and concentrating to obtain a folium ginkgo extract;
(2) dissolving folium Ginkgo extract in water, adding ethyl acetate or methyl acetate or their mixture, extracting, mixing extractive solutions, and concentrating under reduced pressure to dry to obtain total bilobalide extract;
(3) heating and dissolving the total ginkgolide extract with a solvent, cooling, standing, crystallizing, filtering, washing a filter cake with the solvent, and drying to obtain a crude product of the total ginkgolide;
(4) heating and refluxing the crude product of ginkgolide with organic solvent ethyl acetate, methyl acetate or their mixture to dissolve completely, concentrating under reduced pressure until crystallization occurs, standing for crystallization, vacuum filtering, and concentrating the filtrate to dryness to obtain ginkgolide compound, i.e. crude product of ginkgolide compound;
(5) purifying the crude product of the ginkgolide compound by a reverse phase column chromatography for 1-2 times, wherein the mobile phase is a mixed solution of methanol and water, the proportion range of the methanol is 10% -90%, isocratic elution or gradient elution is carried out, the wavelength of ultraviolet detection is 220nm, the effluent of the ginkgolide compound is collected, reduced pressure concentration is carried out until no solvent exists, and drying is carried out, so as to obtain the ginkgolide compound; the pressure of the reduced pressure concentration in the steps (1), (2) and (4) is-0.08 to-0.1 MPa; the bilobalide compound is bilobalide HK.
2. The method for preparing a ginkgolide compound extracted from ginkgo leaves according to claim 1, wherein the concentration of the ethanol aqueous solution in the step (1) is 50-80%.
3. The method for preparing a bilobalide compound extracted from ginkgo leaves according to claim 1, wherein the amount of the water-soluble ginkgo leaf extract in the step (2) is 3-5 times of the weight of the ginkgo leaf extract, the organic solvent used for extraction is ethyl acetate, methyl acetate or a mixed solution of ethyl acetate and methyl acetate, the extraction volume is 0.5-1.5 times of the liquid medicine volume, and the extraction times are 1-4 times.
4. The method of claim 3, wherein the amount of the water-soluble ginkgo biloba extract in the step (2) is 3 to 5 times the weight of the ginkgo biloba extract, the organic solvent used for extraction is ethyl acetate, methyl acetate or a mixture thereof, the extraction volume is 0.8 to 1.2 times the volume of the liquid medicine, and the extraction is performed for 3 times.
5. The method for preparing a ginkgolide compound extracted from ginkgo leaves according to claim 1, wherein the solvent used for dissolving the total ginkgolide extract in step (3) is 30-60% aqueous methanol or aqueous ethanol, the amount of the solvent is 0.5-1.5 times of the weight of the total ginkgolide extract, the mixture is heated to 60 ℃ for dissolution, the cooling and standing crystallization time is 8-16 h, the standing crystallization temperature is 0-10 ℃, the crystal washing solvent is 30-60% aqueous methanol or ethanol, and the crystal washing times are 1-4 times; the drying temperature is 55-65 ℃.
6. The method for preparing a ginkgolide compound extracted from folium Ginkgo according to claim 5, wherein the solvent used for dissolving the total ginkgolide extract in step (3) is 30% -60% aqueous methanol or aqueous ethanol, the amount of the solvent is 0.8-1.2 times of the weight of the total ginkgolide extract, the mixture is heated to 60 ℃ for dissolution, the cooling and standing crystallization time is 8-16 h, the standing crystallization temperature is 3-5 ℃, the crystal washing solvent is 30% -60% aqueous methanol or ethanol, and the crystal washing times are 1-2 times; the drying temperature is 55-65 ℃.
7. The method according to claim 1, wherein the solvent used for dissolving the crude ginkgolides in step (4) is ethyl acetate, methyl acetate or a mixture thereof, and the mixture is concentrated under reduced pressure until the ratio of the weight of the solution to the weight of the crude ginkgolides is 10: 1-15: 1, the repeated operation times are 1-2 times.
8. The method according to claim 5, wherein the purification of the bilobalide compound is performed by using column chromatography in step (5), wherein the column chromatography is RP-18 as a packing material, the mobile phase is a mixed solution of methanol and water, the ratio of methanol is 10% -90%, and isocratic elution or gradient elution is performed.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| GB2211841A (en) * | 1987-11-04 | 1989-07-12 | Scras | Ginkgolide derivatives |
| CN1837213A (en) * | 2006-04-26 | 2006-09-27 | 秦引林 | Bilobalide A derivatives and pharmaceutical application thereof |
| CN103393671A (en) * | 2013-08-21 | 2013-11-20 | 上海信谊百路达药业有限公司 | Extracting and refining method of ginkgolide |
| CN103588785A (en) * | 2013-11-28 | 2014-02-19 | 上海信谊百路达药业有限公司 | Refining method for ginkgolide A and ginkgolide B |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| GB2211841A (en) * | 1987-11-04 | 1989-07-12 | Scras | Ginkgolide derivatives |
| CN1837213A (en) * | 2006-04-26 | 2006-09-27 | 秦引林 | Bilobalide A derivatives and pharmaceutical application thereof |
| CN103393671A (en) * | 2013-08-21 | 2013-11-20 | 上海信谊百路达药业有限公司 | Extracting and refining method of ginkgolide |
| CN103588785A (en) * | 2013-11-28 | 2014-02-19 | 上海信谊百路达药业有限公司 | Refining method for ginkgolide A and ginkgolide B |
Non-Patent Citations (1)
| Title |
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| "Structure-activity Studies with Ginkgo biloba Extract Constituents as Receptor-gated Chloride Channel Blockers and Modulators";S. S. Chatterjee等，;《Pharmacopsychiatry》;20031231;第36卷;第68-77页 * |
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