CN101353294A - Separation and purification method of high-content resveratrol - Google Patents
Separation and purification method of high-content resveratrol Download PDFInfo
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- CN101353294A CN101353294A CNA2008101201337A CN200810120133A CN101353294A CN 101353294 A CN101353294 A CN 101353294A CN A2008101201337 A CNA2008101201337 A CN A2008101201337A CN 200810120133 A CN200810120133 A CN 200810120133A CN 101353294 A CN101353294 A CN 101353294A
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Abstract
The invention relates to a separation and purification method of a high-content resveratrol. A crude product containing resveratrol is taken as raw material; the raw material is separated chromatographically by an alumina column and purified to produce effluent fractions, the effluent fractions are collected, treated, decompressed and condensed, devitrified by adding water, filtered and dried in vacuum to obtain the resveratrol. The purity of the effluent fractions leaving the column is high and can be made into the resveratrol with the purity being 99%; the effluent fractions has high yield and the product thereof can be directly used as medicine and health product. By adopting alumina to absorb non-target products in the crude product, and utilizing the special advantages of the technology in preparing the high-content resveratrol, the whole process of chromatography and elution is carried out in an ethanol-water system; the solvent is safe in use, no waste liquor is produced in the whole chromatography process, and all effluent components are target products so that the method has few effects on production staff and environment, is convenient in mass production, has no need for repeating chromatography and crystallization, and has simple operation procedure and low production cost.
Description
Technical field
The present invention relates to a kind of method of separation and purification, particularly a kind ofly make raw material, utilize the alumina column chromatography separation and purification to prepare the method for high-content of resveratrol separation and purification with the crude product that contains trans-resveratrol.
Background technology
Because of trans-resveratrol has multiple pharmacological effect, can be antitumor, anti-ageing, reducing cholesterol, anticoagulant etc., it mainly is present in the plants such as grape, giant knotweed, black false hellebore, and is wherein the highest with Resveratrol content in the giant knotweed.Trans-resveratrol is mainly used in the treatment of aspects such as anticancer and cardiovascular disorder, and relevant for this reason research report and patent documentation are a lot, and the whole bag of tricks all has different characteristics.
Patent No. CN00121100.5 " trans-resveratrol and polidatin separation method and application thereof ", its method relates to just extracting carries out ethyl acetate extraction and then adopts chloroform-methanol or ethyl acetate-ethanol to carry out silica gel column chromatography as the chromatography elutriant, is that recrystallisation solvent carries out recrystallization and obtains polydatin and obtain trans-resveratrol by secondary chromatography and recrystallization with methyl alcohol-chloroform again.Because of adopting silica gel column chromatography as essential separation means, there is the shortcoming that the sample separation amount is little, output is little, amount of filler is big in silicagel column, and filler silica gel is difficult to regeneration, so industrial production cost height, because of using halogenated hydrocarbon solution, not only the solution use cost is higher, also environment there is serious harm, the finished product unavoidably can relate to harmful dissolvent residual problem, and gained finished product content is lower simultaneously, need repeatedly recrystallization processing, schedule of operation trouble.
Patent No. CN200310112538.3 " new preparation process of polygonin and trans-resveratrol ", separate preparation polygonin and trans-resveratrol with polymeric amide chromatography method, but in actually operating, find, the trans-resveratrol yield is very low behind polyamide column chromatography, trans-resveratrol purity is not high, also need carry out the secondary chromatography or could improve purity remedying its deficiency, and gac easily causes effective constituent extremely to adsorb, so program is many, troublesome poeration, loss of effective components are big with activated carbon decolorizing.
The number of patent application CN200610161949.5 method of separation and purification polydatin and trans-resveratrol " a kind of from the Chinese medicine giant knotweed ", adopt high speed adverse current chromatogram to come the separation and purification trans-resveratrol, this method disposable apparatus has high input, and production capacity is few, be difficult to satisfy the industrial production requirement, what adopt in addition is that its product purity of macroporous resin chromatography is low, also need handle with activated carbon decolorizing and periodic crystallisation again, and therefore operation also bothers.
Summary of the invention
Technical problem to be solved by this invention is: a kind of method of high-content of resveratrol separation and purification is provided, and it is easy to have technology, the equipment less investment; simple to operate; and product has high yield, high purity, prepares free of contamination characteristics, and is easy to realize large-scale industrial production, practical.
The present invention addresses the above problem the technical scheme that is adopted: the method for high-content of resveratrol separation and purification, step: 1, select materials: with commercially available trans-resveratrol crude product is raw material, its Resveratrol content reaches 15%～90% by dry product, and raw material is medicinal extract shape or powder, particulate state;
2, go up the sample formulations prepared from solutions: the weight ratio with crude product is 1, is that 60%～80% aqueous ethanol dissolves crude product with concentration, as last sample solution, and aqueous ethanol consumption (L): crude product (Kg)=15～25: 1;
3, dress post: adopt conventional technology, aluminum oxide is with wet method or dry column-packing;
4, go up the sample wash-out: will contain alumina column chromatography on the aqueous ethanol solution of trans-resveratrol, flow rate control 0.1%～1.0% * column volume/minute, collect stream part, use again behind the end of the sample once with the aqueous ethanol flushing alumina column of the suitable column volume capacity of concentration, collect stream part simultaneously, twice stream part merged;
5, stream part processing: the stream part that will collect is evaporated to 1/3～1/10 of original volume, add 10～20 ℃ tap water again, amount of water is 1～6 times of concentrated volume, again through 2～14 hours placement crystallization, refilter and be target product resveratrol recrystallized powder at 50～60 ℃ of crystal that obtained in 6～8 hours through vacuum-drying.
Compared with prior art, the invention has the advantages that: the separation and purification of adopting the alumina column chromatography technology and using trans-resveratrol, by the absorption of aluminum oxide, given full play to the peculiar advantage of this technology at the preparation high-content of resveratrol to non-target product in the crude product; Alumina chromatographic column is used in whole upper prop resolving all to be the alcohol-water system, this solvent is safe in utilization, does not have waste liquid to produce simultaneously in the chromatography process, flows out composition and is target product, few to producers and environmental influence, the mass industrialized production condition is controlled easily; And equipment used is simple, simultaneous oxidation aluminium column chromatography enabling objective product recovery rate height, and lower prop stream part purity height does not need repeatedly repeatedly chromatography, crystallization, and schedule of operation is easy, so production cost is low.
Embodiment
Below embodiments of the invention are further described.
Step:
1, select materials: with commercially available trans-resveratrol crude product is raw material, and its Resveratrol content reaches 15%～90% by dry product, is medicinal extract shape or powder, particulate state; Or select for use the giant knotweed medicinal material to use the extract of ethyl acetate extraction gained again by ethanol-extracted in the back by fermentation; Giant knotweed makes polygonin convert trans-resveratrol to by fermentation after ethanol-extracted, again the extract of handling with ethyl acetate extraction; Also comprise the above-mentioned acetic acid ethyl ester extract crude product after the buck removal of impurities again;
2, go up the sample formulations prepared from solutions: the weight ratio with crude product is 1, is that 60%～80% aqueous ethanol dissolves crude product with concentration, as last sample solution, and aqueous ethanol consumption (L): crude product (Kg)=18～22: 1;
3, select alumina chromatographic column for use: adopt conventional technology wet method or dry column-packing;
Alumina column chromatography can be with neutral alumina or alkali alumina, and it is 60～250 purpose white particles;
4, go up the sample wash-out: will contain alumina column chromatography on the aqueous ethanol solution of trans-resveratrol, flow rate control 0.1%～1.0% * column volume/minute, collect stream part, use again behind the end of the sample once with the aqueous ethanol flushing alumina column of the suitable column volume capacity of concentration, collect stream part simultaneously, twice stream part merged;
5, stream part processing: the stream part concentrating under reduced pressure that will collect (vacuum degree control exists :-0.05～-0.09Mpa) to 1/3～1/10 of original volume, add 10～20 ℃ tap water again, amount of water is 1～6 times of concentrated volume, and top covering nylon membrane sealing and standing is after 2～14 hours, filter or centrifugal, 50～60 ℃ of temperature again through vacuum-drying (vacuum degree control exists :-0.08～-0.1Mpa) obtained the target product white crystalline powder in 6～8 hours.
Example 1
The 40g crude product (the giant knotweed medicinal material by fermentation after by ethanol-extracted, use the dried powder of the extract of ethyl acetate extraction gained again, detect through HPLC and to contain trans-resveratrol 18.6%) with the 70% ethanol stirring and dissolving of 800ml, (column volume is about 350ml to last sample to standby alkali alumina post, the alkali alumina specification: 100～200 orders, adopt wet method dress post), collect stream part, end of the sample is used 70% ethanol elution of 350ml again, collect stream part, merge two portions stream part about 1140ml, be evaporated to 200ml, with tap water 200ml, 12 hours crystallizatioies are placed in sealing, filter (60 ℃ of crystal vacuum-dryings 7 hours,-0.09～-0.1Mpa), final product 7.1g.Containing trans-resveratrol through the HPLC detection is 99.3%.
Example 2
The 40g crude product (the giant knotweed medicinal material by fermentation after by ethanol-extracted, use the dried powder of the extract of ethyl acetate extraction gained again, detect through HPLC and to contain trans-resveratrol 18.6%) with the 70% ethanol stirring and dissolving of 1000ml, be splined on that (column volume is 350ml on the standby neutral alumina post, the neutral alumina specification: 100～200 orders, adopt wet method dress post), collect stream part, end of the sample is used 70% ethanol elution of 350ml again, collect stream part, merge two portions stream part about 1340ml, be evaporated to 300ml, with tap water 600ml, 4 hours crystallizatioies are placed in sealing, filter (60 ℃ of crystal vacuum-dryings 7 hours,-0.09～-0.1Mpa), final product 7.3g.Containing trans-resveratrol through the HPLC detection is 99.1%.
Example 3
(giant knotweed makes polygonin convert trans-resveratrol to by fermentation after ethanol-extracted to the 40g crude product, the dried powder of the extract of handling with ethyl acetate extraction again, detect through HPLC and to contain trans-resveratrol 20.3%) with the 75% ethanol stirring and dissolving of 800ml, (column volume is about 350ml to last sample to standby alkali alumina post, alkali alumina specification: 100～200 orders, adopt wet method dress post), collect stream part, end of the sample is used 75% ethanol elution of 350ml again, collect stream part, merge two portions stream part about 1140ml, be evaporated to 200ml, with tap water 200ml, 12 hours crystallizatioies are placed in sealing, filter (60 ℃ of crystal vacuum-dryings 7 hours,-0.09～-0.1Mpa), final product 7.7g.Containing trans-resveratrol through the HPLC detection is 99.5%.
Example 4
40g crude product (the dried powder that the extract of ethyl acetate extraction processing is handled through the buck removal of impurities again, detect through HPLC and to contain trans-resveratrol 51.6%) with the 75% ethanol stirring and dissolving of 800ml, (column volume is about 350ml to last sample to standby alkali alumina post, alkali alumina specification: 100～200 orders, adopt wet method dress post), collection stream part, end of the sample is used 75% ethanol elution of 350ml again, collects stream part, merge two portions stream part about 1140ml, be evaporated to 200ml, with tap water 200ml, 12 hours crystallizatioies are placed in sealing, filter, crystal vacuum-drying 7 hours (60 ℃ ,-0.09～-0.1Mpa), final product 19.6g.Containing trans-resveratrol through the HPLC detection is 99.7%.
Example 5
The 40g crude product is (commercially available, detect through HPLC and to contain trans-resveratrol 52.3%) with the 75% ethanol stirring and dissolving of 800ml, (column volume is about 350ml to last sample to standby alkali alumina post, alkali alumina specification: 100～200 orders, adopt wet method dress post), collection stream part, end of the sample is used 75% ethanol elution of 350ml again, collects stream part, merge two portions stream part about 1140ml, be evaporated to 210ml, with tap water 210ml, 12 hours crystallizatioies are placed in sealing, filter, crystal vacuum-drying 7 hours (60 ℃ ,-0.09～-0.1Mpa), final product 19.8g.Containing trans-resveratrol through the HPLC detection is 99.6%.
Example 6 trans-resveratrol large-scale industrialization preparation methods
The 51kg crude product (the giant knotweed medicinal material by fermentation after by ethanol-extracted, use the dried powder of the extract of ethyl acetate extraction gained again, detect through HPLC and to contain trans-resveratrol 18.6%) with the 75% ethanol stirring and dissolving of 1000L, be splined on that (column volume is 450L on the standby alkali alumina post, the alkali alumina specification: 100～200 orders, adopt wet method dress post), collect stream part, end of the sample is used 75% ethanol elution of 450L again, collect stream part, merge two portions stream part about 1450L, be evaporated to 250L, with tap water 250L, 12 hours crystallizatioies are placed in sealing, filter (60 ℃ of crystal vacuum-dryings 7 hours,-0.09～-0.1Mpa), final product 9.05kg.Containing trans-resveratrol through the HPLC detection is 99.2%.
Claims (5)
1, a kind of method of high-content of resveratrol separation and purification, 1), select materials step:: with commercially available trans-resveratrol crude product is raw material, and its Resveratrol content reaches 15%～90% by dry product, raw material is medicinal extract shape or powder, particulate state;
2), go up the sample formulations prepared from solutions: the weight ratio with crude product is 1, is that 60%～80% aqueous ethanol dissolves crude product with concentration, as last sample solution, aqueous ethanol consumption (L): crude product (Kg)=15～25: 1;
3), dress post: adopt conventional technology, aluminum oxide is with wet method or dry column-packing;
4), go up the sample wash-out: will contain alumina column chromatography on the aqueous ethanol solution of trans-resveratrol, flow rate control 0.1%～1.0% * column volume/minute, collect stream part, use again behind the end of the sample once with the aqueous ethanol flushing alumina column of the suitable column volume capacity of concentration, collect stream part simultaneously, twice stream part merged;
5), stream part processing: the stream part that will collect is evaporated to 1/3～1/10 of original volume, add 10～20 ℃ tap water again, amount of water is 1～6 times of concentrated volume, again through 2～14 hours placement crystallization, refilter and be target product resveratrol recrystallized powder at 50～60 ℃ of crystal that obtained in 6～8 hours through vacuum-drying.
2, the method for high-content of resveratrol separation and purification according to claim 1, it is characterized in that raw material also can select for use the giant knotweed medicinal material after fermenting by ethanol-extracted, use the extract of ethyl acetate extraction gained again; Giant knotweed makes polygonin convert trans-resveratrol to by fermentation after ethanol-extracted, again the extract of handling with ethyl acetate extraction; Also comprise the above-mentioned acetic acid ethyl ester extract crude product after the buck removal of impurities again.
3, the method for high-content of resveratrol separation and purification according to claim 1 is characterized in that alumina column chromatography can be with neutral alumina or alkali alumina, and it is 60～250 purpose white particles.
4, the method for high-content of resveratrol separation and purification according to claim 1 is characterized in that sample solution aqueous ethanol consumption (L): crude product (Kg)=18～22: 1.
5, the method for high-content of resveratrol separation and purification according to claim 1, it is characterized in that the stream part of collecting through the reduced vacuum degree be controlled at-0.05～-0.09Mpa, filter the back 60 ℃ through vacuum-drying and make vacuum degree control-0.08～-0.1Mpa obtains crystal.
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| CNA2008101201337A CN101353294A (en) | 2008-07-23 | 2008-07-23 | Separation and purification method of high-content resveratrol |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104087623A (en) * | 2014-06-27 | 2014-10-08 | 湖南鑫利生物科技有限公司 | Method for extracting resveratrol from giant knotweed by enzymolysis |
| CN105420293A (en) * | 2015-11-06 | 2016-03-23 | 贵州天豪民族药业有限公司 | Method for separating and purifying resveratrol from traditional Chinese medicine polygonum cuspidatum extraction solution |
| CN108911952A (en) * | 2018-06-27 | 2018-11-30 | 成都华高生物制品有限公司 | A kind of preparation method for extracting resveratrol from giant knotweed |
| CN109265321A (en) * | 2018-10-23 | 2019-01-25 | 湖南美可达生物资源股份有限公司 | The tomographic system and method for continuous separating refining resveratrol from Gentrin Knotweed P.E |
-
2008
- 2008-07-23 CN CNA2008101201337A patent/CN101353294A/en active Pending
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104087623A (en) * | 2014-06-27 | 2014-10-08 | 湖南鑫利生物科技有限公司 | Method for extracting resveratrol from giant knotweed by enzymolysis |
| CN105420293A (en) * | 2015-11-06 | 2016-03-23 | 贵州天豪民族药业有限公司 | Method for separating and purifying resveratrol from traditional Chinese medicine polygonum cuspidatum extraction solution |
| CN108911952A (en) * | 2018-06-27 | 2018-11-30 | 成都华高生物制品有限公司 | A kind of preparation method for extracting resveratrol from giant knotweed |
| CN109265321A (en) * | 2018-10-23 | 2019-01-25 | 湖南美可达生物资源股份有限公司 | The tomographic system and method for continuous separating refining resveratrol from Gentrin Knotweed P.E |
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Open date: 20090128 |