CN103613630A - Preparation method of mussaendoside Q - Google Patents

Preparation method of mussaendoside Q Download PDF

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Publication number
CN103613630A
CN103613630A CN201310631864.9A CN201310631864A CN103613630A CN 103613630 A CN103613630 A CN 103613630A CN 201310631864 A CN201310631864 A CN 201310631864A CN 103613630 A CN103613630 A CN 103613630A
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China
Prior art keywords
ethanol
mussaendoside
water
preparation
macroporous resin
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CN201310631864.9A
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Chinese (zh)
Inventor
姜平川
李嘉
杨海船
张颖
张赟赟
陈锋
何春欢
李红
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Guangxi Institute Of Chinese Medicine & Pharmaceutical Science
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Guangxi Institute Of Chinese Medicine & Pharmaceutical Science
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Priority to CN201310631864.9A priority Critical patent/CN103613630A/en
Publication of CN103613630A publication Critical patent/CN103613630A/en
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Abstract

The invention discloses a preparation method of mussaendoside Q, which comprises the following steps: (1) adding Mussaenda pubescens crude powder into an ethanol solution, heating under reflux for extraction, and recycling the ethanol to obtain a crude extract; (2) adding distilled water into the crude extract, mixing and dissolving, adding equivalent water saturated n-butanol to extract many times, and concentrating to obtain an extractum; and (3) dissolving the extractum in distilled water, filtering, carrying out primary purification on the filtrate by macroporous resin column chromatography, and carrying out HPLC (high performance liquid chromatography) preparative chromatography separation to obtain the high-purity mussaendoside Q. The method has the advantages of simple and reasonable technique, high yield and high purity, is convenient to operate and can be used for preparing a standard substance.

Description

The preparation method of a kind of mussaendoside Q
Technical field
The invention belongs to pharmaceutical chemistry field, especially relate to the preparation method of a kind of mussaendoside Q.
Background technology
Mussaendoside Q is tetracyclic triterpene saponin(e, molecular formula C 60h 95nO 23, molecular weight 1197, molecular structure is:
Mussaendoside Q is the main component of madder wort Radix Mussaendae.Radix Mussaendae has clearing away summer heat, the effect of removing pattogenic heat from the blood and toxic material from the body.Radix Mussaendae is rich in terpene and triterpenoid saponins.At present, Radix Mussaendae is widely used aspect Chinese patent medicine, and China's treatment first flows first Chinese patent medicine-golden flower and in the side of sense, contains Radix Mussaendae clearly; In YUYE JIEDU KELI, yuye qinghuo sheet, Radix Mussaendae is all used as monarch drug in a prescription simultaneously.
The method of existing separated mussaendoside Q generally adopts reversed-phase silica gel column chromatography, dextrane gel column chromatography, and product purity is lower than 90%, and yield is lower.
Summary of the invention
Technical problem to be solved by this invention is to provide the preparation method of a kind of mussaendoside Q, adopts HPLC preparative chromatography, and simple to operate, products obtained therefrom purity is higher, and the productive rate of the general chromatographic column separation purification method of productivity ratio increases.
The present invention addresses the above problem with following technical scheme:
The preparation method of a kind of mussaendoside Q of the present invention, comprises following operation steps:
(1) Radix Mussaendae meal is joined in ethanolic soln, heating and refluxing extraction at least 3 times, obtains crude extract by extracting solution vacuum concentration, and reclaims solvent, and the weight ratio of Radix Mussaendae meal and ethanolic soln is 1:6~10, and the concentration of ethanolic soln is 60~95%;
(2) gained crude extract is added water miscible after, then add equivalent water-saturated n-butanol extraction at least 3 times, collected organic layer vacuum concentration, must soak paste substance;
(3) the above-mentioned paste substance that soaks is dissolved in water, filters, filtrate is crossed macroporous resin column, adopt successively distilled water, 20% ethanol, 40% ethanol, 60% ethanol and 95% ethanolic soln wash-out, collect 60%-95% ethanol eluate, reclaim ethanol, evaporate to dryness, obtains total saponins;
(4) total saponins is dissolved in to methyl alcohol, adopts HPLC preparative chromatography separated, UV-detector on-line monitoring, collects mussaendoside Q, and concentrated evaporate to dryness, gets product.
In described step (3), macroporous resin column is selected DA201 macroporous resin column or D101 macroporous resin column.
In described step (4), the condition of HPLC preparative chromatography is: elutriant is that volume ratio is the acetonitrile of 25:75~40:60: water or acetonitrile: the mixed solution of 0.01% phosphoric acid solution; Volumetric flow rate: 10~20ml/min; Detect wavelength: 265nm; Column temperature: 25 ℃.
In described step (4), the time period of collecting mussaendoside Q is 11~30.5min.
The advantage of the inventive method is: technique advantages of simple, easy to operate, products obtained therefrom purity is higher.Embodiment
Below in conjunction with specific embodiment, further illustrate the inventive method, but the scope of protection of present invention is not limited to following embodiment.
Embodiment 1
By raw material powder, be that 5kg Radix Mussaendae meal is that 60% ethanol mixes with the volumetric concentration of 8 times of raw material powder weight, heating and refluxing extraction 3 times, each 2h, reclaims solvent by extracting solution vacuum concentration and obtains crude extract medicinal extract.Gained medicinal extract is added to water and add again equivalent water-saturated n-butanol extraction 3 times after miscible, organic layer is merged to final vacuum concentrated, concentrated the paste substance that soaks, the filtration that is dissolved in water, D101 macroporous resin chromatography column on filtrate, adopts distilled water, 20% ethanol, 40% ethanol, 60% ethanol and 95% ethanolic soln wash-out successively, collect 60%~95% ethanol eluate, reclaim ethanol, evaporate to dryness to water ratio is 14%, i.e. total saponins; Total saponins is dissolved in methyl alcohol, and (condition of HPLC preparative chromatography is: moving phase is acetonitrile: water (35:65) to adopt the separation of HPLC preparative chromatography; Volumetric flow rate: 10ml/min; Detect wavelength: 265nm; Column temperature: 25 ℃), UV-detector on-line monitoring, collects mussaendoside Q in 26.104min, and concentrated evaporate to dryness obtains powder 115mg, and purity is 94%, and productive rate is 0.023mg/g.
Embodiment 2
By raw material powder, be that 5kg Radix Mussaendae meal is that 95% ethanol mixes with the volumetric concentration of 10 times of raw material powder weight, heating and refluxing extraction 3 times, each 2h, reclaims solvent by extracting solution vacuum concentration and obtains crude extract medicinal extract.Gained medicinal extract is added to water and add again equivalent water-saturated n-butanol extraction 3 times after miscible, organic layer is merged to final vacuum concentrated, concentrated the paste substance that soaks, the filtration that is dissolved in water, D101 macroporous resin chromatography column on filtrate, adopts distilled water, 20% ethanol, 40% ethanol, 60% ethanol, 95% ethanolic soln wash-out successively, collect 60%-95% ethanol eluate, reclaim ethanol, evaporate to dryness to water ratio is 13%, i.e. total saponins; Total saponins is dissolved in methyl alcohol, and (condition of HPLC preparative chromatography is: moving phase is acetonitrile: water (35:65) to adopt the separation of HPLC preparative chromatography; Volumetric flow rate: 20ml/min; Detect wavelength: 265nm; Column temperature: 25 ℃), UV-detector on-line monitoring, collects mussaendoside Q in 26.588min, and concentrated evaporate to dryness obtains powder 90mg, and purity is 93%, and productive rate is 0.018mg/g.
Embodiment 3
By raw material powder, be that 5kg Radix Mussaendae meal is that 95% ethanol mixes with the volumetric concentration of 8 times of raw material powder weight, heating and refluxing extraction 3 times, each 2h, reclaims solvent by extracting solution vacuum concentration and obtains crude extract medicinal extract.Gained medicinal extract is added to water and add again equivalent water-saturated n-butanol extraction 3 times after miscible, organic layer is merged to final vacuum concentrated, concentrated the paste substance that soaks, the filtration that is dissolved in water, DA201 macroporous resin chromatography column on filtrate, adopts distilled water, 20% ethanol, 40% ethanol, 60% ethanol, 95% ethanolic soln wash-out successively, collect 60%~95% ethanol eluate, reclaim ethanol, evaporate to dryness to water ratio is 15%, i.e. total saponins; Total saponins is dissolved in methyl alcohol, and (condition of HPLC preparative chromatography is: moving phase is acetonitrile: 0.01% phosphoric acid solution (40:60) to adopt the separation of HPLC preparative chromatography; Volumetric flow rate: 15ml/min; Detect wavelength: 265nm; Column temperature: 25 ℃), UV-detector on-line monitoring, collects mussaendoside Q in 11.107min, and concentrated evaporate to dryness obtains powder 75mg, gets product, and purity is 94%, and productive rate is 0.015mg/g.
Embodiment 4
By raw material powder, be that 5kg Radix Mussaendae meal is that 85% ethanol mixes with the volumetric concentration of 6 times of raw material powder weight, heating and refluxing extraction 3 times, each 2h, reclaims solvent by extracting solution vacuum concentration and obtains crude extract medicinal extract.Gained medicinal extract is added to water and add again equivalent water-saturated n-butanol extraction 3 times after miscible, organic layer is merged to final vacuum concentrated, so concentrated that to soak paste substance, the filtration that is dissolved in water, DA201 macroporous resin chromatography column on filtrate, adopt successively distilled water, 20% ethanol, 40% ethanol, 60% ethanol, 95% ethanolic soln wash-out, collect 60%~95% ethanol eluate, reclaim ethanol, evaporate to dryness to water ratio is 15%,, obtain total saponins; Total saponins is dissolved in methyl alcohol, and (condition of HPLC preparative chromatography is: moving phase is acetonitrile: 0.01% phosphoric acid solution (32:68) to adopt the separation of HPLC preparative chromatography; Volumetric flow rate: 15ml/min; Detect wavelength: 265nm; Column temperature: 25 ℃), UV-detector on-line monitoring, collects mussaendoside Q in 28.359min, and concentrated evaporate to dryness obtains powder 100mg, and purity is 94%, and productive rate is 0.020mg/g.
Embodiment 5
By raw material powder, be that 5kg Radix Mussaendae meal is that 75% ethanol mixes with the volumetric concentration of 8 times of raw material powder weight, heating and refluxing extraction 3 times, each 2h, reclaims solvent by extracting solution vacuum concentration and obtains crude extract medicinal extract.Gained medicinal extract is added to water and add again equivalent water-saturated n-butanol extraction 3 times after miscible, organic layer is merged to final vacuum concentrated, so concentrated that to soak paste substance, the filtration that is dissolved in water, DA201 macroporous resin chromatography column on filtrate, adopt successively distilled water, 20% ethanol, 40% ethanol, 60% ethanol, 95% ethanolic soln wash-out, collect 60%~95% ethanol eluate, reclaim ethanol, evaporate to dryness to water ratio is 14%,, i.e. total saponins; Total saponins is dissolved in methyl alcohol, and (condition of HPLC preparative chromatography is: moving phase is acetonitrile: 0.01% phosphoric acid solution (38:62) to adopt the separation of HPLC preparative chromatography; Volumetric flow rate: 10ml/min; Detect wavelength: 265nm; Column temperature: 25 ℃), UV-detector on-line monitoring, collects mussaendoside Q in 13.657min, and concentrated evaporate to dryness obtains powder 75mg, and purity is 95%, and productive rate is 0.015mg/g.
Embodiment 6
By the volumetric concentration of 8 times of raw material powder 5kg Radix Mussaendae meal and raw material powder weight, be 95% ethanol, heating and refluxing extraction 3 times, each 2h, reclaims solvent by extracting solution vacuum concentration and obtains crude extract medicinal extract.Gained medicinal extract is added to water and add again equivalent water-saturated n-butanol extraction 3 times after miscible, organic layer is merged to final vacuum concentrated, concentrated the paste substance that soaks, the filtration that is dissolved in water, D101 macroporous resin chromatography column on filtrate, adopts distilled water, 20% ethanol, 40% ethanol, 60% ethanol, 95% ethanolic soln wash-out successively, collect 60%~95% ethanol eluate, reclaim ethanol, evaporate to dryness to water ratio is 15%, i.e. total saponins; Total saponins is dissolved in methyl alcohol, and (condition of HPLC preparative chromatography is: moving phase is acetonitrile: water (30:70) to adopt the separation of HPLC preparative chromatography; Volumetric flow rate: 20ml/min; Detect wavelength: 265nm; Column temperature: 25 ℃), UV-detector on-line monitoring, collects mussaendoside Q in 30.288min, and concentrated evaporate to dryness obtains powder 110mg, and purity is 94%, and productive rate is 0.022mg/g.

Claims (4)

1. a preparation method of mussaendoside Q, is characterized in that, comprises following operation steps:
(1) Radix Mussaendae meal is joined in ethanolic soln, heating and refluxing extraction at least 3 times, obtains crude extract by extracting solution vacuum concentration, and reclaims solvent, and the weight ratio of Radix Mussaendae meal and ethanolic soln is 1:6~10, and the concentration of ethanolic soln is 60~95%;
(2) gained crude extract is added water miscible after, then add equivalent water-saturated n-butanol extraction at least 3 times, collected organic layer vacuum concentration, must soak paste substance;
(3) the above-mentioned paste substance that soaks is dissolved in water, filters, filtrate is crossed macroporous resin column, adopt successively distilled water, 20% ethanol, 40% ethanol, 60% ethanol and 95% ethanolic soln wash-out, collect 60%~95% ethanol eluate, reclaim ethanol, evaporate to dryness, obtains total saponins;
(4) total saponins is dissolved in to methyl alcohol, adopts HPLC preparative chromatography separated, UV-detector on-line monitoring, collects mussaendoside Q, and concentrated evaporate to dryness, gets product.
2. the preparation method of mussaendoside Q according to claim 1, is characterized in that, in described step (3), macroporous resin column is selected DA201 macroporous resin column or D101 macroporous resin column.
3. the preparation method of mussaendoside Q according to claim 1, it is characterized in that, in described step (4), the condition of HPLC preparative chromatography is: elutriant is that volume ratio is the acetonitrile of 25:75~40:60: water or acetonitrile: the mixed solution of 0.01% phosphoric acid solution; Volumetric flow rate: 10~20ml/min; Detect wavelength: 265nm; Column temperature: 25 ℃.
4. the preparation method of mussaendoside Q according to claim 1, is characterized in that, in described step (4), the time period of collecting mussaendoside Q is 11~30.5min.
CN201310631864.9A 2013-11-29 2013-11-29 Preparation method of mussaendoside Q Pending CN103613630A (en)

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Country Status (1)

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Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
JUN-PING XU, ET AL.: "MUSSAENDOSIDES M AND N, NEW SAPONINS FROM MUSSAENDA PUBESCENS", 《JOURNAL OF NATURAL PRODUCTS》 *
WEIMIN ZHAO, ET AL.: "NEW TRITERPENOID SAPONINS FROM MUSSAENDA PUBESCENS", 《JOURNAL OF NATURAL PRODUCTS》 *
张颖等: "玉叶金花化学成分研究", 《中药新药与临床药理》 *

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Application publication date: 20140305