CN1215031C - Process for preparing oridonin - Google Patents
Process for preparing oridonin Download PDFInfo
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- CN1215031C CN1215031C CN 02139049 CN02139049A CN1215031C CN 1215031 C CN1215031 C CN 1215031C CN 02139049 CN02139049 CN 02139049 CN 02139049 A CN02139049 A CN 02139049A CN 1215031 C CN1215031 C CN 1215031C
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- rubescensine
- oridonin
- acetone
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Abstract
The present invention discloses a process for preparing oridonin. Rabdosia is pulverized and refluxed with a mixed solvent of dichloromethane, acetone or chloroform and ethanol, an extraction liquid is decompressed and concentrated to obtain extract in the form of a thick paste, petroleum ether is added for degreasing, and the petroleum ether is filtered off and evaporated to obtain extract; the extract is processed by acetone or alcohol and is mixed with silica gel for column chromatography, and then, the mixed silica gel is uniformly loaded on an upper layer; the upper layer is eluted with dichloromethane, a mixed solvent of dichloromethane and acetone is used for stripping, and stripping liquid fraction containing oridonin is collected, decompressed and concentrated; the concentrated solution is put in a crystallizer for deposition and crystallization, so that oridonin crystals are separated from a mother liquid, the mother liquid can be continuously decompressed and concentrated for fractional crystallization until oridonin is completely separated from the mother liquor, and the oridonin crystals are recrystallized and refined with a methanol solution. The present invention has the advantages of short chromatography column course, high separation speed, favorable effect, high product purity and low production cost, and the content of oridonin can reach more than 90%.
Description
Technical field
The present invention relates to the method that medicinal plants extracts purifying, more specifically relate to a kind of preparation method of rubescensine A.
Technical background
The extracting method of traditional rubescensine A mainly contains ether extraction, ethanol extraction method.
The step of ether extraction is: the Rabdosia rubescens raw material adds 3～5 times of amount ether extraction 2～3 hours, reclaim ether, get green and brown look dope dissolve with methanol, add activated carbon decolorizing, with the faint yellow clear and bright methanol solution concentrating under reduced pressure that obtains, the concentrated solution standing over night is filtered, the filtrate reconcentration gets thick paste, last Al
2O
3Post is used the benzene wash-out earlier, uses the ether wash-out again, and the ether elutriant obtains coarse crystallization after reclaiming solvent, gets rubescensine A through acetone treatment.
This method has the extract yield advantage of higher, but owing to use poisonous, inflammable and explosive solvents such as ether, benzene in a large number, is absolutely unsafe, and is not suitable for industrial production.
The step of ethanol extraction method is: the Rabdosia rubescens raw material adds 10 times of amount 95% ethanol, heating was extracted 2 hours in extractor, reclaim ethanol, get green and brown look thickness lotion, use anhydrous alcohol solution, added the powdered carbon reflux decolour 1 hour, suction filtration remove concentrate behind the gac medicinal extract, the medicinal extract acetic acid ethyl dissolution is used 5%Na again
2CO
3Solution extraction reclaims solvent and gets syrup, and last silicagel column with chloroform, acetone mixed solvent or sherwood oil, acetone mixed solvent wash-out, is collected respective streams part concentrating under reduced pressure, and crystallization gets rubescensine A then.
This method replaced ether and methyl alcohol with industrial spirit, relatively be suitable for industrial production, but that shortcoming is an extract yield is low.
Though above-mentioned two methods are all considered Rabdosia rubescens extract is carried out column chromatography for separation, a common defective is arranged: all do not illustrate how the rubescensine A coarse-grain that obtains further is refined into highly purified rubescensine A crystallization.
Yuan Ke etc. rolled up in " column chromatography separates the novel process of preparation rubescensine A " of delivering on the 8th phase " CHINA JOURNAL OF CHINESE MATERIA MEDICA " at 1997 the 22nd, compared to be concentrated into medicinal extract shape → dissolve with methanol after the ether extraction and to mix thoroughly and be loaded on capital and carry out chromatographic separation → respectively with sherwood oil with a certain amount of back of activated carbon decolorizing → continue to be concentrated into and proper silica gel, chloroform, acetone is wash-out successively, can obtain the former technology of rubescensine A crystalline, pointing out that solvent ether equal solvent used in this technology is poisonous easily fires, and highly volatile, price is higher, the used solvent systems of wash-out is more, bothers during operational cost.So this method only is fit to the laboratory and prepares rubescensine A on a small quantity, is unsuitable for suitability for industrialized production.Therefore, its separation preparation is improved.The ethanol extraction that is about to Rabdosia rubescens is with an amount of 95% dissolve with ethanol, add 2 times of amounts of about alcohol extract dry extract (W/W) column chromatography silica gel, add an amount of 95% ethanol again, silica gel is all stirred in the moistening rearmounted water-bath of ethanol, fry and take out after being dry powder, it is standby to room temperature to dry in the air.With the column chromatography silica gel of handling well pack into the below be lined with in the glass column (the high 150cm of post, diameter 8cm) of absorbent cotton and filter paper.Dry extract and separatory silica gel usage ratio are 1: 10 (W/W).With a certain proportion of sherwood oil-acetone is that elutriant carries out wash-out.After collecting rubescensine A stream part merging evaporate to dryness, add an amount of methyl alcohol thermosol and place, promptly have the rubescensine A crystallization to separate out.Though the more preceding method solvent for use of this method system is simplified to some extent, but still could mix with column chromatography silica gel with 95% dissolve with ethanol again after extracting solution concentrated, selected chromatography column blade diameter length ratio is 1: 18.75, the desired equipment of suitability for industrialized production is huge, eluent flow rate is 1/150/ minute～1/220/ minute (effluent volume/sorbent material volume/minute), too slow, the obvious optimised process of suitability for industrialized production greatly.
Summary of the invention
The object of the present invention is to provide a kind of preparation method of rubescensine A, utilize mixed solvent to extract, by the short distance silica gel column chromatography, with the mixed solvent system desorb, fractional crystallization and recrystallization make highly purified rubescensine A then.
In order to achieve the above object, the present invention is by the following technical solutions: with the Rabdosia rubescens is raw material, through methylene dichloride: acetone (4: 1～9: 1) or trichloromethane: ethanol (4: 1～9: 1) reflux extraction, extracting solution obtains heavy paste extract through concentrating under reduced pressure, add 3～5 times of petroleum ether degreasings, filtering and volatilize sherwood oil, to obtain rubescensine A content be 10～15% extract medicinal extract, adopt the mixed solvent desorption system, by silicagel column short distance chromatography, collect stripping liquid, concentrate, crystallization can obtain content greater than 90% rubescensine A.Rabdosia rubescens Rabdosiarubescens (Hemsl.) Hara is a Labiatae Rabdosia plant, China's treatment esophageal carcinoma, carcinoma of gastric cardia and primary hepatocarcinoma of being mainly used among the people.Rubescensine A (oridonin) is antineoplastic main component in the Rabdosia rubescens.Experimental results show that rubescensine A all has tangible cytotoxicity external to Hela cell and esophageal squamous cell carcinoma cell strain (CaEs-17); In vivo multiple animal transplanting tumor such as ehrlich ascites tumor (ascitic type and solid-type), S-180 sarcoma (ascitic type and solid-type), liver cancer (ascitic type and solid-type), reticulum cell sarcoma (ARS) etc. all there is restraining effect; Can prevent that the caused rat esophagus epithelial proliferation of nitrosamine (precancerous lesion) from changing cancer into.Clinical trial proves, to the esophageal carcinoma, and carcinoma of gastric cardia, liver cancer, and breast cancer etc. has tangible relief of symptoms effect, the stable and effect of dwindling the knurl body and prolonging patient's life.Share with chemotherapy, can alleviate the untoward reaction of chemotherapeutics, improve curative effect, epithelium of esophagus severe hyperplasia is also had certain curative effect.
The step of extracting rubescensine A is as follows:
1, Rabdosia rubescens is crushed to 20～100 orders, successively uses 6～8 times, 4～6 times, 3～5 times, 4: 1～9: 1 methylene dichloride, acetone or trichloromethane, ethanol mixed solvent to carry out reflux extraction, each 3～5 hours;
2, extracting solution obtains heavy paste extract through concentrating under reduced pressure, adds 3～5 times of petroleum ether degreasings, and filtering and volatilize sherwood oil, to obtain rubescensine A content be 10～15% extract medicinal extract; 3, add 1～3 times of acetone or alcohol and handle, mix in 1: 1～2: 1 ratios and 200～300 order column chromatography silica gels and be mixed evenly, spread out drying, make solvent evaporates clean in the shady and cool ventilation place; 4, get 4～6 times and mix the fresh silica gel dress post that mixes silica gel, to mix again and mix the silica gel dress thereon, the blade diameter length ratio that is used for the chromatography column of column chromatography for separation is 1: 1.5～1: 4, chromatography column is made of stainless steel or enamel or glass or other corrosion resistant material, first methylene dichloride wash-out with 5～8 times of volumes, use 4: 1～9: 1 methylene dichloride, acetone mixed solvent desorb again, eluent flow rate is controlled at 1/60～1/70/ minute (the per minute effluent volume is with " effluent volume/sorbent material volume/minute " expression); 5, when mixed solvent washes out 5～7 times of dress column volumes, begin to collect the stream part that contains rubescensine A, the volume of collecting stripping liquid stops desorb during for 1.5～2.5 times of dress column volume, and stripping liquid concentrates 10～15 times under temperature is 45～55 ℃, vacuum tightness 0.5～0.6Mpa condition; 6, concentrated solution is put and is left standstill crystallization in 8～12 hours in the crystallizer, carries out centrifugally when waiting to reach capacity crystalline state, and the rubescensine A crystal is separated with mother liquor, and mother liquor can continue the concentrating under reduced pressure fractional crystallization, separates from mother liquor fully until rubescensine A; 7, the rubescensine A crystallization of above-mentioned prepared can be carried out recrystallizing and refining with 10～15 times methanol solution.
The present invention compared with prior art has the following advantages and effect: what the solvent of (1) extraction Rabdosia rubescens raw material was used is methylene dichloride: acetone (4: 1～9: 1) or trichloromethane: ethanol (4: 1～9: 1) mixed solvent; (2) because the post journey is short, medicinal extract rubescensine A content height, so just can finish the column chromatography for separation of rubescensine A with less silica gel; (3) because the chromatography column blade diameter length ratio only is 1: 1.5～1: 4, can save facility investment, can reduce again that the workshop takes up room and area; (4) eluent flow rate is controlled at 1/60/ minute～1/70/ minute (effluent volume/sorbent material volume/minute), and higher production efficiency has shortened the production cycle greatly; (5) select for use methylene dichloride and acetone (4: 1～9: 1) mixed solvent system to carry out silica gel column chromatography, make rubescensine A stream part of desorb comparatively concentrated; (6) select methylene dichloride, acetone or trichloromethane, ethanol equal solvent system for use, safe in utilization relatively; (7) on the basis of aforementioned technology, used crystallization and recrystallization technology can be prepared steady quality, highly purified rubescensine A, and content can reach more than 90%.(8) the chromatography column short distance is easy to operate, and velocity of separation is fast, and is effective, and cost is low, and the little yellow of color and luster is to white, and the further refining content that can get is greater than 99% pure product.
Embodiment
Embodiment 1. gets 30 kilograms of Rabdosia rubescens, be crushed to 40～80 orders, successively add 220 kilograms, 160 kilograms, 130 kilograms trichloromethanes: ethanol (9: 1) solution refluxes three times, each 3.5 hours, extracting solution obtains 1 kilogram of the Rabdosia rubescens medicinal extract of rubescensine A content 11.3% through concentrating under reduced pressure, and was even with Ethanol Treatment, mix mixed for 1.5 kilograms with 200～300 order silica gel, do at the stand, gets 8 kilograms of fresh silica gel dress posts, will mix mixed silica gel and add thereon.Washing post with 50 liters of methylene dichloride earlier removes preceding assorted, use the mixed solvent desorb of methylene dichloride and acetone (4: 1) again, treat that 65 liters of mixed solvents washed after the cylinder, begin to collect rubescensine A stream part to 15 liters, 50 ℃ of temperature, be concentrated into 1.5 liters under the vacuum tightness 0.6Mpa condition, concentrated solution leaves standstill crystallization in 10 hours in enamel tray, through the 0.8Mpa vacuum filtration, filtrate is continued concentrating under reduced pressure fractional crystallization 3 times, and rubescensine A is separated from mother liquor fully, and the rubescensine A crystallization that obtains merges back with 1.6 liters of dissolve with methanol solution, carry out recrystallization, volatilize solvent, in 45 ℃ of following vacuum (0.9Mpa) dry 3 hours, obtain 0.05 kilogram of rubescensine A.Rubescensine A content is 92% after testing.
Embodiment 2. is with 500 kilograms of commercially available medicinal material Rabdosia rubescens, be crushed to 40～80 orders, successively with 3000 kilograms, 2000 kilograms, 1000 kilograms of methylene dichloride: acetone (4: 1) solution refluxes three times, each 3 hours, united extraction liquid is through concentrating under reduced pressure, obtain about 10 kilograms of the medicinal extract of rubescensine A content 12.8%, even with acetone treatment, mix and mix in 18 kilogram of 200～300 order column chromatography silica gel, airing volatilizes solvent, in 15 chromatography columns, respectively pre-install earlier 8 kilograms of fresh silica gel, then sample is mixed and mixed the top that silica gel is loaded on 15 chromatography columns equably, each chromatography column is respectively earlier with 60 liters of methylene dichloride wash-outs, use methylene dichloride and acetone (4: 1) mixed solvent desorb again, when mixed solvent washes out about 70 liters, begin to collect the stripping liquid that contains rubescensine A stream part, each chromatography column is collected 18 liters of stripping liquids, stripping liquid is merged the back 50 ℃ of temperature, be concentrated into 18 liters under the vacuum tightness 0.55Mpa condition, concentrated solution is put and is left standstill crystallization in 10 hours in the crystallizer, carry out centrifugal when waiting to reach capacity crystalline state, the rubescensine A crystallization is separated with mother liquor, mother liquor can continue the concentrating under reduced pressure fractional crystallization, from mother liquor, separate fully until rubescensine A, merge isolated rubescensine A crystallization, with 20 liters of dissolve with methanol solution, carry out recrystallization, centrifugation in 45 ℃ of following vacuum (0.9Mpa) dry 3 hours, obtains 0.78 kilogram of 92.5% rubescensine A.
Claims (4)
1, a kind of preparation method of rubescensine A comprises the following steps:
A, Rabdosia rubescens is crushed to 20～100 orders, successively uses 6～8 times, 4～6 times, 3～5 times, 4: 1～9: 1 methylene dichloride, acetone or trichloromethane, ethanol mixed solvent to reflux, each 3～5 hours;
B, extracting solution obtain heavy paste extract through concentrating under reduced pressure, add 3～5 times of petroleum ether degreasings, and filtering and volatilize sherwood oil, to obtain rubescensine A content be 10～15% extract medicinal extract;
C, add 1～3 times of acetone or alcohol and handle, mix in 1: 1～1: 2 ratio and 200～300 order column chromatography silica gels and be mixed evenly, spread out drying in the shady and cool ventilation place;
D, get 4～6 times and mix the fresh silica gel dress posts that mix silica gel, will mix again and mix the silica gel dress thereon,, use 4: 1～9: 1 methylene dichloride, acetone mixed solvent desorb again with 5～8 times of methylene dichloride wash-outs of adorning column volumes;
E, collect the stream part contain rubescensine A, stop desorb when collecting the stripping liquid volume for 1.5～2.5 times of dress column volume, stripping liquid concentrates 10～15 times under temperature is 45～55 ℃, vacuum tightness 0.5～0.6Mpa condition;
F, concentrated solution are put and left standstill crystallization in 8～12 hours in the crystallizer, and be centrifugal, and the rubescensine A crystal is separated with mother liquor, and mother liquor can continue the concentrating under reduced pressure fractional crystallization, separates from mother liquor fully until rubescensine A;
G, rubescensine A crystallization are carried out recrystallizing and refining with 10～15 times methanol solution.
2, the preparation method of a kind of rubescensine A according to claim 1, the chromatography column blade diameter length ratio that it is characterized in that being used for column chromatography for separation is 1: 1.5～1: 4.
3, the preparation method of a kind of rubescensine A according to claim 2 is characterized in that chromatography column makes of stainless steel.
4, the preparation method of a kind of rubescensine A according to claim 1 is characterized in that eluent flow rate was controlled at 1/60～1/70/ minute, effluent volume/sorbent material volume/minute.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 02139049 CN1215031C (en) | 2002-09-13 | 2002-09-13 | Process for preparing oridonin |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 02139049 CN1215031C (en) | 2002-09-13 | 2002-09-13 | Process for preparing oridonin |
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|---|---|
| CN1482115A CN1482115A (en) | 2004-03-17 |
| CN1215031C true CN1215031C (en) | 2005-08-17 |
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Families Citing this family (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1311814C (en) * | 2004-09-14 | 2007-04-25 | 河南康鑫药业有限公司 | Rabdosia dripping pill and its preparing method |
| 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 |
| CN101003528B (en) * | 2006-01-18 | 2010-05-26 | 郑州大学 | Diterpene compound and derivative in kaurene class of new dissymmetry, and its preparation method and uses |
| CN100402532C (en) * | 2006-02-10 | 2008-07-16 | 陈俊辉 | Preparation method for extracting oridonin from rabdosia |
| CN101317837B (en) * | 2008-07-04 | 2011-07-27 | 浙江大学 | Effective component of rabdosia, preparation method and application thereof |
| CN102680588B (en) * | 2011-03-09 | 2015-10-28 | 常州博闻迪医药科技有限公司 | A kind of column chromatography for separation detection technique and uses thereof |
| CN107670033A (en) * | 2016-08-02 | 2018-02-09 | 天津中医药大学 | Antineoplastic pharmaceutical compositions and its application |
| CN110092796B (en) * | 2018-01-27 | 2021-04-09 | 南京工业大学 | Method for extracting and separating oridonin |
| CN110038322A (en) * | 2019-04-24 | 2019-07-23 | 上海复旦复华药业有限公司 | A kind of tripterygium glycosides chromatographic silica gel dress column method |
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