CN1528757A - Technical method for extracting oridinin from rabdosia rubescens - Google Patents
Technical method for extracting oridinin from rabdosia rubescens Download PDFInfo
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- CN1528757A CN1528757A CNA031514774A CN03151477A CN1528757A CN 1528757 A CN1528757 A CN 1528757A CN A031514774 A CNA031514774 A CN A031514774A CN 03151477 A CN03151477 A CN 03151477A CN 1528757 A CN1528757 A CN 1528757A
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- rabdosia rubescens
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Abstract
The invention advances a new technique proper to separate and purify oridonin. It adopts alcohol solution extraction, then makes water float reflux treatment, where the water float liquor directly passes through the column of macroporous resin (like HZ-841 macroporous absorbing resin) polymerized of styrene and divinylbenzene as monomers to eliminate sugar, pigment, etc, adopts silica gel and magnesium chloride mixed column chromatography to separate and elute, and finally recrystallize to obtain higher purity oridonin. It is safe, low-cost, and applied to industrialized production.
Description
Technical field
The present invention relates to a kind of new processing method of from Rabdosia rubescens, extracting rubescensine A.
Background technology
Drug research shows that Rabdosia rubescens has good anti-inflammatory, antibiotic, analgesic activity, can effectively suppress first type, beta hemolysis type suis, golden yellow Portugal coccus etc.To the Hela cell of vitro culture, the human body esophageal carcinoma 109 cells and liver cancer BEL-7402 cell have tangible cytotoxicity, to multiple transplanting rerum natura animal tumor such as ECA, S
180, P
388, L
1210Liver cancer and AKS have obvious antitumor action, and rubescensine A has slight restraining effect to esophageal epithelial proliferation.Wherein diterpenes composition rubescensine A is an antineoplastic effective composition, thereby is described as " penicillin " and " natural antibiotics " in the Chinese medicine.And the control that Rabdosia rubescens is used for AIDS is also being studied by the U.S..The exploitation of Rabdosia rubescens will become this century " sunrise industry " and " gold industry ".
The Rabdosia rubescens extraction process is mainly by extracting---decolouring---technologies such as chromatography, extraction wherein important effective constituent rubescensine A and remove wherein plant pigments, carbohydrate and tannin class.The difference of the method for various bibliographical informations is embodied in:
(1) difference of extraction solvent as ether extraction, mainly contains the Zhao Qingzhi of China, the E.Fujita.T.Fujita of Wang Hanqing etc. and Japan, H, Katayama, M.Shibuya; Ethanol extraction method;
(2) extract the back to extracting solution treatment process difference, remove water-insoluble matter as methods such as useful filtrations, the most frequently used is sedimentation, then activated carbon decolorizing; Be directly to use activated carbon decolorizing in addition;
(3) the water extract after the processing generally adopts the technology of organic solvent extraction or chromatography or extraction+chromatography to obtain rubescensine A.
Except that above-mentioned difference was arranged on operational path, the concrete difference of different methods showed different on selected extraction agent and the column chromatography method.
These extraction processes normally adopt the solution extraction method, adopt activated carbon decolorizing then, directly carry out column chromatography for separation then after concentrating, this technology can not well be removed tannin and polysaccharide wherein, can not finely remove and only differ 2 rubescensine B with the rubescensine A molecular weight, be difficult to obtain the pure product of rubescensine A.
The structure of rubescensine A is:
Rubescensine A
The structure of rubescensine B is:
Rubescensine B
In order to obtain the higher rubescensine A of purity, remove wherein plant pigments, polysaccharide, resin, tannin class material well, therefore be necessary to improve above-mentioned traditional processing technology.
Summary of the invention
The purpose of this patent invention is the defective that overcomes prior art, develop a kind of simple, be suitable for suitability for industrialized production, and the novel process of effective constituent rubescensine A in the low extraction purifying Rabdosia rubescens of cost, and can be used for the development of new anti-cancer agent.
Technical conceive of the present invention is such:
Several extracting and purifying method of integrated use of the present invention carry out organic assembling with it, have developed the novel process of the suitable separation and purification rubescensine A of a cover.Adopt the alcoholic solution extraction method, carry out the floating reflow treatment of water then, it is that macroporous adsorbent resin (as the HZ-841 macroporous adsorbent resin) post of monomer polymerization is removed wherein sugar, plant pigments etc. that the water supernatant liquid is crossed with vinylbenzene and divinylbenzene, adopt silica gel to separate wash-out, carry out recrystallization at last and just can obtain the higher rubescensine A of purity with magnesium oxide mixing column chromatography.
The present invention is achieved through the following technical solutions:
1, with Rabdosia rubescens with 4-10 95% alcoholic solution doubly, the alcoholic solution of 6 times of amounts preferably, alcohol extracting 2-6 hour, best 4 hours; Again with 2-6 95% alcoholic solution doubly, the alcoholic solution of 4 times of amounts preferably, alcohol extracting 1-4 hour, best 2 hours, extracting temperature was 76 degrees centigrade;
2, merge extracted twice liquid, concentrating under reduced pressure reclaims ethanol, obtains green and brown look paste; It is floating that the water that adding 1-6 doubly measures carries out water, preferably 3 times; Temperature is 60-90 ℃, preferably 80 ℃; Return time 0.5-3h, preferably 1.5h;
3, filter, with the direct mistake of filtrate is macroporous adsorbent resin (as the HZ-841 macroporous adsorbent resin) post of monomer polymerization with vinylbenzene and divinylbenzene, the most of plant pigments of elder generation's water flush away, use lower concentration alcohol wash-out again, be generally 5%-30%, be preferably 10%, then use the pure wash-out of 50%-70%, 60% alcohol preferably, effect is better;
4, will cross macroporous adsorptive resins liquid concentrating under reduced pressure evaporate to dryness, silica gel separates wash-out with magnesium oxide mixing column chromatography, and eluent can adopt chloroform-methanol, chloroform-acetone, sherwood oil-acetone etc., preferably chloroform-acetone, the eluent ratio is 6: 1-9: 1, and preferably 85: 15; The ratio of raw material and sorbent material is 1: 5～1: 10, and the best is 1: 7, and sorbent material is silica gel and magnesian mixed adsorbent, is generally and is advisable at 15: 1;
5, carry out recrystallization with methyl alcohol-acetone, filter the back and is washed till white, obtain purity and be 99% rubescensine A with small amount of acetone.
With method of the present invention extracting effective components rubescensine A from Rabdosia rubescens, technical have following remarkable advantage and a progress:
(1) this technology is increased in after the ethanolic soln extraction, adopts the floating technique of backflow of water to remove a part of impurity, can tentatively improve the purity of product.
(2) adopting with vinylbenzene and divinylbenzene is macroporous adsorbent resin (as the HZ-841 macroporous adsorbent resin) column technology of monomer polymerization, and adopts silica gel+magnesian mixing column to separate rubescensine A, can largely improve product purity.
(3) handle with crossing macroporous resin column through water is floating, when crossing chromatography column because of on the sample purity of sample higher relatively, thereby the sorbent material volume containing the sample of same amount is just bigger, has so just saved the sorbent material consumption.
(4) only with a cover elution system, solvent load and elution time have been saved greatly.
(5) can obtain the higher rubescensine A of content (99.85%)
Description of drawings:
Fig. 1 is that thin-layer chromatogram, Fig. 2 rubescensine A reference substance and the comparison of sample thin-layer chromatogram, Fig. 3 of column chromatography wash-out different fractions is the HPLC test result, and Fig. 4 rubescensine A sample and reference substance HPLC figure comparison, Fig. 5 are H
ENMR spectrum, Fig. 6 are that IR spectrum, Fig. 7 are the MS spectrums.
Below in conjunction with embodiment and accompanying drawing the present invention is further specified.
Extract raw material 1kg and add 6 times of amount 95% extraction using alcohols 4 hours, pour out solution, added 4 times of amount 95% alcohol refluxs again 2 hours, concentrating under reduced pressure, reclaim ethanol, get green and brown look paste, adding 3 times of water gagings refluxed 1.5 hours, filter, it is macroporous adsorbent resin (as the HZ-841 macroporous adsorbent resin) post of monomer polymerization that this filtrate is crossed with vinylbenzene and divinylbenzene, and elder generation is washed till closely colourless with about 6 times of water gagings, measures 60% ethanol liquid wash-out with about 4 times then, with this elutriant concentrating under reduced pressure, obtain the about 12g of khaki color powder.
Column chromatography for separation will go up in the step gained khaki color powder and proper silica gel and mix thoroughly, will be equivalent to silica gel, magnesium oxide (15: 1) the dress post of 5 times of amounts of powder, and the upper strata covers a small amount of silica gel behind the application of sample, adds a small amount of absorbent cotton again.With 9: 1 chloroforms-acetone wash-out, carry out wash-out with the flow velocity of 5ml/min, collect by every part of 100ml, carry out the thin layer plate analysis, prove that the 6th part is pure first element (seeing accompanying drawing 1) later on.Concentrating under reduced pressure carries out recrystallization with methyl alcohol-acetone, behind the filtration under diminished pressure, is washed till white with small amount of acetone, weighs after the drying, altogether 1.5g.Thin layer chromatography analysis with rubescensine A reference substance and sample with dissolve with methanol after, point sample on same silica gel thin-layer plate, launch with 9: 1 chloroform-methanols, taking-up is dried, under ultraviolet lamp, shine, sample has identical Rf value (seeing accompanying drawing 2) with reference substance, and presents same bright orange green spot, inclusion-free spot in the sample.HPLC analyze with rubescensine A reference substance and sample with dissolve with methanol after, sample introduction under identical conditions obtains identical color atlas, retention time is identical, peak shape is also identical.Rubescensine A HPLC figure and contrast figure.See accompanying drawing 3,4.
The mensuration of NMR and IR shows that through infrared spectra and nuclear magnetic resonance spectrometry institute's product that get sample are identical with the reference substance structure, is same compound.See accompanying drawing 5,6.
It is rubescensine A by analysis that MS detects.See accompanying drawing 7.
HPLC, H
ENMR, IR, MS analyze:
IR, H
ENMR, MS check and analysis are rubescensine A, and are consistent with bibliographical information, product is carried out HPLC detect, and the results are shown in accompanying drawing 3.
Claims (12)
1, a kind of processing method of extracting rubescensine A from Rabdosia rubescens is characterized in that, comprises the steps:
(1) with 95% alcoholic solution of Rabdosia rubescens, extracted 2-6 hour with 4-10 times; With 2-6 95% alcoholic solution doubly, extracted 1-4 hour, extracting temperature is 76 degrees centigrade again;
(2) merge extracted twice liquid, concentrating under reduced pressure reclaims ethanol, obtains green and brown look paste; It is floating that the water that adding 1-6 doubly measures carries out water, and temperature 60-90 ℃, backflow 0.5-3h forms the plain suspension of the careless first of winter spirit;
(3) with after the plain suspension filtered of the careless first of winter spirit, is the direct mistake of filtrate the macroporous adsorptive resins of monomer polymerization with vinylbenzene and divinylbenzene, as the HZ-841 macroporous adsorptive resins, the most of plant pigments of elder generation's water flush away, use 5%-30% lower concentration alcohol wash-out again, then use the pure wash-out of 50%-70%;
(4) will cross with vinylbenzene and divinylbenzene is macroporous adsorbent resin (as the HZ-841 macroporous adsorbent resin) the post liquid concentrating under reduced pressure evaporate to dryness of monomer polymerization, separate wash-out with silica gel with magnesium oxide mixing column chromatography, eluent can adopt chloroform-methanol, chloroform-acetone, sherwood oil-acetone, the eluent ratio is 6: 1-9: 1, the ratio of raw material and sorbent material is 1: 5-1: 10, sorbent material is silica gel and magnesian mixed adsorbent, is generally 15: 1
(5) carry out recrystallization with methyl alcohol-acetone, filter the back and is washed till white, obtain purity and be 99% rubescensine A with small amount of acetone.
2, the processing method of extracting rubescensine A from Rabdosia rubescens as claimed in claim 1 is characterized in that, when extracting with 95% alcoholic solution, the first time is with the alcoholic solution of 6 times of amounts, and the second time is with the alcoholic solution of 4 times of amounts.
3, the processing method of extracting rubescensine A from Rabdosia rubescens as claimed in claim 2 is characterized in that, the alcohol extracting time is 4 hours for the first time, is extracted as 2 hours for the second time.
4, the processing method of extracting rubescensine A from Rabdosia rubescens as claimed in claim 3 is characterized in that, extracting temperature is 76 degrees centigrade.
5, the processing method of extracting rubescensine A from Rabdosia rubescens as claimed in claim 1 is characterized in that, carries out water and floats when refluxing, and the floating amount of water is 3 times.
6, the processing method of extracting rubescensine A from Rabdosia rubescens as claimed in claim 5 is characterized in that reflux temperature is 80 ℃.
7, the processing method of extracting rubescensine A from Rabdosia rubescens as claimed in claim 6 is characterized in that return time is 1.5h.
8, the processing method of from Rabdosia rubescens, extracting rubescensine A as claimed in claim 1, it is characterized in that crossing with vinylbenzene and divinylbenzene is the macroporous adsorptive resins of monomer polymerization, during as the HZ-841 macroporous adsorptive resins, earlier with 10% pure wash-out, again with 60% pure wash-out collection.
9, the processing method of extracting rubescensine A from Rabdosia rubescens as claimed in claim 8 is characterized in that, crosses silica gel and magnesium oxide mixing column, and elutriant is chloroform-acetone, and ratio is 85: 15.
10, the processing method of extracting rubescensine A from Rabdosia rubescens as claimed in claim 9 is characterized in that, crosses silica gel and magnesium oxide mixing column, and silica gel and magnesian ratio are 15: 1.
11, the processing method of extracting rubescensine A from Rabdosia rubescens as claimed in claim 10 is characterized in that, crosses silica gel and magnesium oxide mixing column, and the ratio of raw material and sorbent material is 1: 7.
12, the processing method of extracting rubescensine A from Rabdosia rubescens as claimed in claim 1 is characterized in that, adopts methyl alcohol and acetone mixed solution during recrystallization.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 03151477 CN1240697C (en) | 2003-09-29 | 2003-09-29 | Technical method for extracting oridinin from rabdosia rubescens |
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| CN 03151477 CN1240697C (en) | 2003-09-29 | 2003-09-29 | Technical method for extracting oridinin from rabdosia rubescens |
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| CN1528757A true CN1528757A (en) | 2004-09-15 |
| CN1240697C CN1240697C (en) | 2006-02-08 |
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| CN 03151477 Expired - Fee Related CN1240697C (en) | 2003-09-29 | 2003-09-29 | Technical method for extracting oridinin from rabdosia rubescens |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1304387C (en) * | 2005-04-13 | 2007-03-14 | 中国科学院武汉植物园 | Separation and preparation of blushed rabdosia extract-A as check sample |
| CN100402532C (en) * | 2006-02-10 | 2008-07-16 | 陈俊辉 | Preparation method for extracting oridonin from rabdosia |
| CN1868503B (en) * | 2005-05-24 | 2011-10-26 | 山东绿叶天然药物研究开发有限公司 | Preparation method of extractive of Herba Rabdosiae diterpene |
| CN101919903B (en) * | 2005-05-24 | 2011-11-23 | 山东绿叶天然药物研究开发有限公司 | Method for preparing rabdosia rubescens diterpene extract |
| CN104474011A (en) * | 2014-11-21 | 2015-04-01 | 郑州轻工业学院 | Method for decolorizing and purifying rabdosia rubescens extract |
| CN105685740A (en) * | 2016-01-20 | 2016-06-22 | 河南丰之源生物科技有限公司 | Method for preparing concentrated clear rabdosia rubescens juice |
| CN105732653A (en) * | 2016-02-03 | 2016-07-06 | 河南中医学院 | Method for preparing oridonin from isodon japonica |
-
2003
- 2003-09-29 CN CN 03151477 patent/CN1240697C/en not_active Expired - Fee Related
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1304387C (en) * | 2005-04-13 | 2007-03-14 | 中国科学院武汉植物园 | Separation and preparation of blushed rabdosia extract-A as check sample |
| CN1868503B (en) * | 2005-05-24 | 2011-10-26 | 山东绿叶天然药物研究开发有限公司 | Preparation method of extractive of Herba Rabdosiae diterpene |
| CN101919903B (en) * | 2005-05-24 | 2011-11-23 | 山东绿叶天然药物研究开发有限公司 | Method for preparing rabdosia rubescens diterpene extract |
| CN100402532C (en) * | 2006-02-10 | 2008-07-16 | 陈俊辉 | Preparation method for extracting oridonin from rabdosia |
| CN104474011A (en) * | 2014-11-21 | 2015-04-01 | 郑州轻工业学院 | Method for decolorizing and purifying rabdosia rubescens extract |
| CN104474011B (en) * | 2014-11-21 | 2018-03-16 | 郑州轻工业学院 | The method of Rabdosia rubescens extract decolouring removal of impurities |
| CN105685740A (en) * | 2016-01-20 | 2016-06-22 | 河南丰之源生物科技有限公司 | Method for preparing concentrated clear rabdosia rubescens juice |
| CN105685740B (en) * | 2016-01-20 | 2019-01-22 | 河南丰之源生物科技有限公司 | A method of preparing concentration Rabdosia rubescens juice |
| CN105732653A (en) * | 2016-02-03 | 2016-07-06 | 河南中医学院 | Method for preparing oridonin from isodon japonica |
| CN105732653B (en) * | 2016-02-03 | 2017-12-15 | 河南中医药大学 | A kind of method that Oridonin is prepared from Isodon Japonica Hara |
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|---|---|
| CN1240697C (en) | 2006-02-08 |
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