CN102863430A - Preparation method of sesquiterpenes saussurea lappa clarke alkaloid monomer - Google Patents
Preparation method of sesquiterpenes saussurea lappa clarke alkaloid monomer Download PDFInfo
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- CN102863430A CN102863430A CN 201210362835 CN201210362835A CN102863430A CN 102863430 A CN102863430 A CN 102863430A CN 201210362835 CN201210362835 CN 201210362835 CN 201210362835 A CN201210362835 A CN 201210362835A CN 102863430 A CN102863430 A CN 102863430A
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Abstract
The invention belongs to the filed of natural product separating, and provides a preparation method of sesquiterpenes saussurea lappa clarke alkaloid monomer. The preparation method comprises the steps of extracting with ethanol solution, extracting with organic solvent, carrying out middle-pressure magnesium oxide column chromatography, and purifying with preparative high performance liquid chromatography. The product prepared by the preparation method provided by the invention is high in purity, high in quality, simple in preparation method, large in preparation amount, and easy for industrialization.
Description
Technical field
The invention belongs to the Separation of Natural Products field, especially relate to a kind of method that adopts the high performance liquid preparative chromatography method from the Saussurea lappa Clarke root, to prepare simultaneously Saussurea lappa Clarke amine A and Saussurea lappa Clarke amine B.
Background technology
Saussurea lappa Clarke is the feverfew Saussurea lappa Clarke
Saussurea lappaThe root of Clarke, the another name banksia rose, Radix Aucklandiae (Radix Vladimiriae) have the effects such as promoting qi circulation, resolving food retention, soothing the liver, stomach invigorating, can treat the stagnation of the circulation of vital energy, stop eating gather, the illnesss such as fullness sensation in chest abdominal distension, vomiting dysentery.Contain multiple volatile oil and alkaloid compound in the Saussurea lappa Clarke.
Saussurea lappa Clarke amine A is to separate the sesquiterpenoids alkaloid compound that obtains from the Saussurea lappa Clarke root with Saussurea lappa Clarke amine B, these two components all have antiulcer action, Saussurea lappa Clarke amine A is when 100mg/kg, be 59.8% to the rat gastric ulcer inhibiting rate that is caused by HCl/ ethanol, Saussurea lappa Clarke amine B is 87.2% to the rat gastric ulcer inhibiting rate that is caused by HCl/ ethanol when 50mg/kg.
Relatively less to alkaloidal correlative study report in the Saussurea lappa Clarke in the existing document, there are no preparing simultaneously the alkaloidal method report of sesquiterpenoids in the Saussurea lappa Clarke.
Summary of the invention
The technical problem to be solved in the present invention provides a kind of preparation method of sesquiterpenoids Saussurea lappa Clarke alkaloid monomer, and the method is simple to operate, and products obtained therefrom content is high.
The present invention is achieved by the following technical solutions:
A kind of preparation method of sesquiterpenoids Saussurea lappa Clarke alkaloid monomer is characterized in that comprising following steps:
1) get the Saussurea lappa Clarke root and pulverize, add 70 ~ 90%(V/V) aqueous ethanolic solution microwave extraction 2 ~ 3 times, medicinal material and solvent quality ratio are 1:(6-15), united extraction liquid, concentrating under reduced pressure gets crude extract;
2) above-mentioned crude extract is outstanding soluble in water, use ethyl acetate extraction, the extraction liquid concentrating under reduced pressure gets medicinal extract;
3) above-mentioned medicinal extract is carried out middle pressure magnesium oxide column chromatography, with sherwood oil-acetone gradient elution, collect each wash-out part, then carry out thin-layer chromatography and collect the target elutriant, reclaim reagent to small volume, the gained crude product extracts 3 times with the aqueous hydrochloric acid of 0.2mol/L, merge hydrochloric acid extraction liquid, regulate pH to 9.5-10.5 with the 0.5mol/L ammonia soln again, filter, throw out deionized water wash 3-5 time gets the Saussurea lappa Clarke total alkaloids;
4) with above-mentioned Saussurea lappa Clarke total alkaloids dissolve with methanol, through the preparative liquid chromatography separation and purification, acetonitrile-water mixing solutions wash-out is collected the corresponding compound flow point of each chromatographic peak, dry Saussurea lappa Clarke amine A and the Saussurea lappa Clarke amine B of getting of concentrating under reduced pressure.
Described step 2) the ethyl acetate consumption in is that the 1-4 of extract volume doubly measures.
Magnesium oxide order number in the described step 3) is the 100-300 order, and the column chromatography elution requirement is: adopt sherwood oil-acetone to carry out gradient elution for 20:1,10:1,5:1,2:1,1:1 by volume.
Preparative liquid chromatography adopts common C18 post in the described step 4), and the acetonitrile take volume ratio as 75:25 and water mixed solvent are moving phase, the flow velocity of 1.5-4ml/min.
The invention has the beneficial effects as follows: the inventive method is by adopting the steps such as microwave extraction, solvent extraction, middle pressure magnesium oxide column chromatography, crystallization and preparative liquid chromatography separation to prepare Saussurea lappa Clarke amine A, Saussurea lappa Clarke amine B, three kinds of sesquiterpenoids alkaloid compounds of Saussurea lappa Clarke amine C, consuming time short, yield is high, simple to operate, be easy to industrialization.
Embodiment:
Further specify the present invention below in conjunction with embodiment, but the scope of protection of present invention is not limited to following embodiment.
Embodiment 1:
Getting Saussurea lappa Clarke root 50kg is ground in the meal input microwave extraction tank, 85% ethanolic soln microwave extraction of 10 times of raw material weights of adding 20 minutes, extracting solution leaches, so repeatedly extract 2 times, united extraction liquid, filtrate decompression reclaims and obtains crude extract, crude extract is added water outstanding water-soluble, with the ethyl acetate extraction of 2 times of volumes 4 times, concentrating under reduced pressure gets ethyl acetate extract, ethyl acetate extract is pressed the magnesium oxide post with dissolve with methanol wet method loading to filler 200-300 purpose, with the mixed solvent of sherwood oil-acetone, 20:1 by volume successively, 10:1,5:1,2:1,1:1 carries out gradient elution, each flow point of Fractional Collections will contain Saussurea lappa Clarke sesquiterpenoids alkaloid flow point and merge, and decompression and solvent recovery is to small volume, then the aqueous hydrochloric acid with 0.2mol/L extracts 3 times, merge hydrochloric acid extraction liquid, regulate pH to 10.2 with the 0.5mol/L ammonia soln again, filter, throw out deionized water wash 5 times get the Saussurea lappa Clarke total alkaloids; The Saussurea lappa Clarke total alkaloids is added dissolve with methanol, separate at the anti-phase preparative column of C18, acetonitrile-water (75:25, V/V) is moving phase, and the flow velocity of 2.5ml/min adopts on-line ultraviolet to detect, and detects wavelength: 276nm.Collect respectively compound flow point corresponding to each chromatographic peak, obtain Saussurea lappa Clarke amine A and two kinds of sesquiterpenoids Saussurea lappa Clarke of Saussurea lappa Clarke amine B alkaloid after the reduced vacuum drying.
Embodiment 2:
Getting Saussurea lappa Clarke root 50kg is ground in the meal input microwave extraction tank, 70% ethanolic soln microwave extraction of 15 times of raw material weights of adding 20 minutes, extracting solution leaches, so repeatedly extract 3 times, united extraction liquid, filtrate decompression reclaims and obtains crude extract, crude extract is added water outstanding water-soluble, with the ethyl acetate extraction of 1 times of volume 4 times, concentrating under reduced pressure gets ethyl acetate extract, ethyl acetate extract is pressed the magnesium oxide post with dissolve with methanol wet method loading to filler 200-300 purpose, with the mixed solvent of sherwood oil-acetone, 20:1 by volume successively, 10:1,5:1,2:1,1:1 carries out gradient elution, each flow point of Fractional Collections will contain Saussurea lappa Clarke sesquiterpenoids alkaloid flow point and merge, and decompression and solvent recovery is to small volume, then the aqueous hydrochloric acid with 0.2mol/L extracts 3 times, merge hydrochloric acid extraction liquid, regulate pH to 9.5 with the 0.5mol/L ammonia soln again, filter, throw out deionized water wash 4 times get the Saussurea lappa Clarke total alkaloids; The Saussurea lappa Clarke total alkaloids is added dissolve with methanol, separate at the anti-phase preparative column of C18, acetonitrile-water (75:25, V/V) is moving phase, and the flow velocity of 3.0ml/min adopts on-line ultraviolet to detect, and detects wavelength: 276nm.Collect respectively compound flow point corresponding to each chromatographic peak, obtain Saussurea lappa Clarke amine A and two kinds of sesquiterpenoids Saussurea lappa Clarke of Saussurea lappa Clarke amine B alkaloid after the reduced vacuum drying.
Embodiment 3:
Getting Saussurea lappa Clarke root 50kg is ground in the meal input microwave extraction tank, 80% ethanolic soln microwave extraction of 8 times of raw material weights of adding 15 minutes, extracting solution leaches, so repeatedly extract 3 times, united extraction liquid, filtrate decompression reclaims and obtains crude extract, crude extract is added water outstanding water-soluble, with the ethyl acetate extraction of 4 times of volumes 4 times, concentrating under reduced pressure gets ethyl acetate extract, ethyl acetate extract is pressed the magnesium oxide post with dissolve with methanol wet method loading to filler 100-200 purpose, with the mixed solvent of sherwood oil-acetone, 20:1 by volume successively, 10:1,5:1,2:1,1:1 carries out gradient elution, each flow point of Fractional Collections will contain Saussurea lappa Clarke sesquiterpenoids alkaloid flow point and merge, and decompression and solvent recovery is to small volume, then the aqueous hydrochloric acid with 0.2mol/L extracts 3 times, merge hydrochloric acid extraction liquid, regulate pH to 10.0 with the 0.5mol/L ammonia soln again, filter, throw out deionized water wash 3 times get the Saussurea lappa Clarke total alkaloids; The Saussurea lappa Clarke total alkaloids is added dissolve with methanol, separate at the anti-phase preparative column of C18, acetonitrile-water (75:25, V/V) is moving phase, and the flow velocity of 1.5ml/min adopts on-line ultraviolet to detect, and detects wavelength: 276nm.Collect respectively compound flow point corresponding to each chromatographic peak, obtain Saussurea lappa Clarke amine A and two kinds of sesquiterpenoids Saussurea lappa Clarke of Saussurea lappa Clarke amine B alkaloid after the reduced vacuum drying.
Embodiment 4:
Getting Saussurea lappa Clarke root 50kg is ground in the meal input microwave extraction tank, 90% ethanolic soln microwave extraction of 6 times of raw material weights of adding 10 minutes, extracting solution leaches, so repeatedly extract 2 times, united extraction liquid, filtrate decompression reclaims and obtains crude extract, crude extract is added water outstanding water-soluble, with the ethyl acetate extraction of 3 times of volumes 4 times, concentrating under reduced pressure gets ethyl acetate extract, ethyl acetate extract is pressed the magnesium oxide post with dissolve with methanol wet method loading to filler 200-300 purpose, with the mixed solvent of sherwood oil-acetone, 20:1 by volume successively, 10:1,5:1,2:1,1:1 carries out gradient elution, each flow point of Fractional Collections will contain Saussurea lappa Clarke sesquiterpenoids alkaloid flow point and merge, and decompression and solvent recovery is to small volume, then the aqueous hydrochloric acid with 0.2mol/L extracts 3 times, merge hydrochloric acid extraction liquid, regulate pH to 9.8 with the 0.5mol/L ammonia soln again, filter, throw out deionized water wash 4 times get the Saussurea lappa Clarke total alkaloids; The Saussurea lappa Clarke total alkaloids is added dissolve with methanol, separate at the anti-phase preparative column of C18, acetonitrile-water (75:25, V/V) is moving phase, and the flow velocity of 3.5ml/min adopts on-line ultraviolet to detect, and detects wavelength: 276nm.Collect respectively compound flow point corresponding to each chromatographic peak, obtain Saussurea lappa Clarke amine A and two kinds of sesquiterpenoids Saussurea lappa Clarke of Saussurea lappa Clarke amine B alkaloid after the reduced vacuum drying.
Embodiment 5:
Getting Saussurea lappa Clarke root 50kg is ground in the meal input microwave extraction tank, 70% ethanolic soln microwave extraction of 6 times of raw material weights of adding 15 minutes, extracting solution leaches, so repeatedly extract 2 times, united extraction liquid, filtrate decompression reclaims and obtains crude extract, crude extract is added water outstanding water-soluble, with the ethyl acetate extraction of 2 times of volumes 4 times, concentrating under reduced pressure gets ethyl acetate extract, ethyl acetate extract is pressed the magnesium oxide post with dissolve with methanol wet method loading to filler 100-200 purpose, with the mixed solvent of sherwood oil-acetone, 20:1 by volume successively, 10:1,5:1,2:1,1:1 carries out gradient elution, each flow point of Fractional Collections will contain Saussurea lappa Clarke sesquiterpenoids alkaloid flow point and merge, and decompression and solvent recovery is to small volume, then the aqueous hydrochloric acid with 0.2mol/L extracts 3 times, merge hydrochloric acid extraction liquid, regulate pH to 10.5 with the 0.5mol/L ammonia soln again, filter, throw out deionized water wash 5 times get the Saussurea lappa Clarke total alkaloids; The Saussurea lappa Clarke total alkaloids is added dissolve with methanol, separate at the anti-phase preparative column of C18, acetonitrile-water (75:25, V/V) is moving phase, and the flow velocity of 4.0ml/min adopts on-line ultraviolet to detect, and detects wavelength: 276nm.Collect respectively compound flow point corresponding to each chromatographic peak, obtain Saussurea lappa Clarke amine A and two kinds of sesquiterpenoids Saussurea lappa Clarke of Saussurea lappa Clarke amine B alkaloid after the reduced vacuum drying.
Embodiment 6:
Getting Saussurea lappa Clarke root 50kg is ground in the meal input microwave extraction tank, 90% ethanolic soln microwave extraction of 15 times of raw material weights of adding 10 minutes, extracting solution leaches, so repeatedly extract 3 times, united extraction liquid, filtrate decompression reclaims and obtains crude extract, crude extract is added water outstanding water-soluble, with the ethyl acetate extraction of 1.5 times of volumes 4 times, concentrating under reduced pressure gets ethyl acetate extract, ethyl acetate extract is pressed the magnesium oxide post with dissolve with methanol wet method loading to filler 100-200 purpose, with the mixed solvent of sherwood oil-acetone, 20:1 by volume successively, 10:1,5:1,2:1,1:1 carries out gradient elution, each flow point of Fractional Collections will contain Saussurea lappa Clarke sesquiterpenoids alkaloid flow point and merge, and decompression and solvent recovery is to small volume, then the aqueous hydrochloric acid with 0.2mol/L extracts 3 times, merge hydrochloric acid extraction liquid, regulate pH to 10.0 with the 0.5mol/L ammonia soln again, filter, throw out deionized water wash 3 times get the Saussurea lappa Clarke total alkaloids; The Saussurea lappa Clarke total alkaloids is added dissolve with methanol, separate at the anti-phase preparative column of C18, acetonitrile-water (75:25, V/V) is moving phase, and the flow velocity of 2.0ml/min adopts on-line ultraviolet to detect, and detects wavelength: 276nm.Collect respectively compound flow point corresponding to each chromatographic peak, obtain Saussurea lappa Clarke amine A and two kinds of sesquiterpenoids Saussurea lappa Clarke of Saussurea lappa Clarke amine B alkaloid after the reduced vacuum drying.
Claims (4)
1. the preparation method of a sesquiterpenoids Saussurea lappa Clarke alkaloid monomer is characterized in that comprising following steps:
1) get the Saussurea lappa Clarke root and pulverize, add 70 ~ 90%(V/V) aqueous ethanolic solution microwave extraction 2 ~ 3 times, medicinal material and solvent quality ratio are 1:(6-15), united extraction liquid, concentrating under reduced pressure gets crude extract;
2) above-mentioned crude extract is outstanding soluble in water, use ethyl acetate extraction, the extraction liquid concentrating under reduced pressure gets medicinal extract;
3) above-mentioned medicinal extract is carried out middle pressure magnesium oxide column chromatography, with sherwood oil-acetone gradient elution, collect each wash-out part, then carry out thin-layer chromatography and collect the target elutriant, reclaim reagent to small volume, the gained crude product extracts 3 times with the aqueous hydrochloric acid of 0.2mol/L, merge hydrochloric acid extraction liquid, regulate pH to 9.5-10.5 with the 0.5mol/L ammonia soln again, filter, throw out deionized water wash 3-5 time gets the Saussurea lappa Clarke total alkaloids;
4) with above-mentioned Saussurea lappa Clarke total alkaloids dissolve with methanol, through the preparative liquid chromatography separation and purification, acetonitrile-water mixing solutions wash-out is collected the corresponding compound flow point of each chromatographic peak, dry Saussurea lappa Clarke amine A and the Saussurea lappa Clarke amine B of getting of concentrating under reduced pressure.
2. the preparation method of a kind of sesquiterpenoids Saussurea lappa Clarke alkaloid monomer as claimed in claim 1 is characterized in that described step 2) in the ethyl acetate consumption be that the 1-4 of extract volume doubly measures.
3. the preparation method of a kind of sesquiterpenoids Saussurea lappa Clarke alkaloid monomer as claimed in claim 1, it is characterized in that the magnesium oxide order number in the described step 3) is the 100-300 order, the column chromatography elution requirement is: adopt sherwood oil-acetone to carry out gradient elution for 20:1,10:1,5:1,2:1,1:1 by volume.
4. the preparation method of a kind of sesquiterpenoids Saussurea lappa Clarke alkaloid monomer as claimed in claim 1, it is characterized in that preparative liquid chromatography adopts common C18 post in the described step 4), acetonitrile take volume ratio as 75:25 and water mixed solvent are moving phase, the flow velocity of 1.5-4ml/min.
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| CN111454267A (en) * | 2019-12-07 | 2020-07-28 | 中国科学院昆明植物研究所 | Coriander soda nail and preparation method and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN111454267A (en) * | 2019-12-07 | 2020-07-28 | 中国科学院昆明植物研究所 | Coriander soda nail and preparation method and application thereof |
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Application publication date: 20130109 |