CN103910705B - The method of rapid extraction separation and purification NVP-XAA 723 from the tankage of green tea - Google Patents
The method of rapid extraction separation and purification NVP-XAA 723 from the tankage of green tea Download PDFInfo
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- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D311/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings
- C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
- C07D311/04—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring
- C07D311/58—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring other than with oxygen or sulphur atoms in position 2 or 4
- C07D311/60—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring other than with oxygen or sulphur atoms in position 2 or 4 with aryl radicals attached in position 2
- C07D311/62—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring other than with oxygen or sulphur atoms in position 2 or 4 with aryl radicals attached in position 2 with oxygen atoms directly attached in position 3, e.g. anthocyanidins
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Abstract
The present invention relates to a kind of method of rapid extraction separation and purification NVP-XAA 723 from the tankage of green tea, comprise following steps: the tankage ordinary method extraction using alcohol of (1) green tea; (2) extracting solution less than 80 DEG C decompression recycling ethanols are extremely without alcohol taste; (3) alcohol extract polyamide resin fractionation by adsorption; (4) NVP-XAA 723 (EGCG) flow point is collected, with activated carbon decolorizing, concentrate under reduced pressure at low temperature; (5) with ethyl alcohol recrystallization, highly purified NVP-XAA 723 (EGCG) is obtained; The present invention is simple and easy to do, and only adopt a kind of resin can reach absorption, separation and purification one step completes, the nontoxic solvent used, safety, reliably, product purity is high, cost is low, is suitable for industrialized production.
Description
Technical field
The present invention relates to medical applications field, particularly relate to a kind of method of rapid extraction separation and purification NVP-XAA 723 from the tankage of green tea.
Background technology
Polyphenols in tealeaves accounts for the 18%-36% of dry weight of tea leaves, comprises the flavanol compound based on catechin, flavones and flavonols, anthocyanidin and leucoanthocyanidin and phenolic acid and depside.Wherein catechin compounds is the bulk composition of tea-polyphenol, accounts for the 12%-24% of dry weight of tea leaves, accounts for the 70%-80% of tea-polyphenol total amount.Modern medicine study finds, tea-polyphenol is a kind of novel natural antioxidants, has the physiological actions such as antitumor, anti-ageing, anti-sudden change, Tumor suppression growth, Weight-lossing antihypertensive and preventing cardiovascular disease.And these physiological actions are mainly showed by catechin wherein, effect that the NVP-XAA 723 (EGCG) wherein in ester catechin shows in many aspects is obviously better than other catechin compositions.
The molecular formula of EGCG is C
22h
18o
11,containing multiple phenolic hydroxyl group in the molecule of molecular weight 458.4, EGCG, active centre, on two rings, therefore has the ability of very strong anti-oxidant and scavenging free radicals.EGCG sterling is white powder, bitter and puckery flavor, nontoxic, very easily water-soluble, and can be dissolved in methyl alcohol, ethanol, acetonitrile and ethyl acetate, be insoluble to chloroform, fusing point is 218 DEG C.Easily oxidized in atmosphere, become micro mist look, also easy to be oxidized along with the rising of temperature.
That is familiar with tea component nourishing function along with people gos deep into, and the extraction and isolation of tea-polyphenol obtains research extensively and profoundly, and research prepared by the separation of catechin monomers especially ester catechin EGCG also more and more receives the concern of people.At present, the process patent of extraction and isolation EGCG is more.Wherein preparation method's patent of high purity EGCG mainly contains:
CN1465572A epi-nutgall catechin gallic acid ester monomer purifying method
CN1470510A epi-nutgall catechin gallic acid ester monomer purifying process
CN102702163A prepares the method for high purity catechin from the tankage of tealeaves
The separation purification method of CN1603319A catechin monomers
The method of CN101386614A preparing epigallocatechin-3-gallate by resin adsorption method
CN1733753A epi-nutgall catechin gallic acid ester monomer purifying method
CN101381359A mono-kind extracts the method for high-purity epigallocatechin-3-gallate from green tea
CN101723927A method for batch production, separation and purification of catechin monomers EGCG
The preparation method of the EGCG of a CN101074224A high-content
Eight above-mentioned patents, sample is all adopt conventional method to extract, comprise solvent-extraction process, precipitation by metallic ion method, resin adsorption method, supercritical fluid extraction, ultrasonic-leaching method etc., due in tea-polyphenol except EGCG, other materials also containing certain content, the EGCG purity therefore extracted is not high.To the purge process of single component EGCG, mostly first adopt resin absorption technology to carry out removal of impurities, comprising macroporous adsorptive resins, anion-cation exchange resin post etc., the resin column of usual employing post or two different in kinds has been connected, main effect is adsorbed sample, take water as elutriant, carries out desorption again after removing water-soluble impurity with rare alcohol.Only according to above-mentioned means, the purity requirement of EGCG cannot be reached, and the crucial purification step of above-mentioned 8 patents is that last a few step all have employed chromatographic separation technology means, comprise means such as using supercritical fluid chromatography, sephadex chromatography, mesolow silica gel column chromatography chromatogram, C18 liquid chromatography technology of preparing, finally by the method for recrystallization, make the purity that EGCG reaches very high.
The weak point of 8 above-mentioned patents is that operating process is complicated, harsh to requirement for experiment condition, owing to adopting high-end laboratory apparatus and expensive chemical reagent, makes preparation cost very high and operates wayward.And repeatedly have employed post adsorption separation process, loss is comparatively large, and make product yield lower, and introduce a large amount of organic solvents during the course, operation steps is only limitted to laboratory, and product can not realize industrialization.
Summary of the invention
The technical problem that the present invention mainly solves is to provide a kind of method that tankage that can make full use of green tea carry out rapid extraction separation and purification NVP-XAA 723, the tankage of green tea are made to obtain recycling as waste, also solve EGCG in purge process simultaneously, step is complicated, length consuming time, solvent for use toxicity is large, and product yield is low, can not realize the problems such as industrialization.
The method of rapid extraction separation and purification NVP-XAA 723 from the tankage of green tea of the present invention, comprises following steps:
(1) tankage of green tea extract with 60% EtOH Sonicate concussion, and extraction time is 30 minutes, extracts twice, united extraction liquid;
(2) extracting solution reclaims ethanol to without alcohol taste, is concentrated into the clear cream that relative density is 1.20 to 1.22;
(3) the clear cream polyamide resin column fractionation by adsorption will obtained in (2);
(4) thin-layer chromatography tracing detection, collects NVP-XAA 723 flow point, after activated carbon decolorizing, filters, and filtrate is in 60 DEG C of-80 DEG C of low-temperature reduced-pressures, and lead to nitrogen protection, being concentrated into relative density is 1.15-1.18, lyophilize;
(5) with ethyl alcohol recrystallization, highly purified NVP-XAA 723 is obtained;
Elutriant in described step (3) fractionation by adsorption comprises 95% ethanol and 36% acetic acid, wherein 95% ethanol: the volume ratio of 36% acetic acid is 5:1-1:5.
Preferably, the tankage of described step (1) green tea, some the old tea comprising some thick old bars, pruning branch or the tea dusts produced in the grade course of processing in the process of picking tea-leaves except a large amount of fresh leaf or overstock in storage, tea grounds.
Preferably, the described tankage of step (1) green tea and the ratio of 60% ethanol are 1:8 g/ml-1:20 g/ml.
Preferably, described step (2) extracting solution reclaims ethanol to without alcohol taste, refers to reclaim under reduced pressure below 80 DEG C.
Preferably, the granularity of described step (3) polyamide resin is 60-100 order, and consumption is: resin: clear cream=10-20:1(g/g), the blade diameter length ratio of resin column is 1:20.
Preferably, the consumption of described step (4) gac is the 2%-3% of liquor capacity, and the amount of namely adding gac in every 100ml solution is 2g-3g.
The invention has the beneficial effects as follows: the present invention does not need the preliminary purification of macroporous resin, by controlling chromatographic separation condition, only adopt a kind of separation system, can directly carry out the separation of polymeric amide chromatography, therefore, by thin-layer chromatography tracing detection, EGCG sterling flow point can be collected, make separation like this, purifying one step completes, make easy to operation; And in whole process, only used the etoh solvent of food grade, acetic acid and gac, ensure that product safety, nontoxic.
Accompanying drawing explanation
Fig. 1 is the high-efficient liquid phase chromatogram of EGCG reference substance;
Fig. 2 is the sample high-efficient liquid phase chromatogram that the method for rapid extraction separation and purification NVP-XAA 723 from the tankage of green tea of the present invention obtains.
Embodiment
embodiment 1
Green tea tankage 1 kg, adds the concussion of 8L60% EtOH Sonicate and extracts, extract twice, each each 30min, united extraction liquid; Extracting solution less than 80 DEG C decompression recycling ethanols, to without alcohol taste, are concentrated into the clear cream that relative density is 1.20 to 1.22; Cooling; (resin demand is heavy 10 times of cream to the absorption of upper polyamide resin column; the blade diameter length ratio of post is 1:20); with 95% ethanol: 36% acetic acid (volume ratio is 5:1) wash-out, thin-layer chromatography tracing detection, collects NVP-XAA 723 (EGCG) flow point; after activated carbon decolorizing with 2% (g/100ml); filter, it is 1.15-1.18 that filtrate low temperature under 60 DEG C-80 DEG C logical nitrogen protections is evaporated to relative density, lyophilize.With ethyl alcohol recrystallization, obtain the NVP-XAA 723 (EGCG) that purity is 98.3%, yield is 10%.
embodiment 2
Green tea tankage 1 kg, adds the concussion of 12L60% EtOH Sonicate and extracts, extract twice, each each 30min, united extraction liquid.Extracting solution less than 80 DEG C decompression recycling ethanols, to without alcohol taste, are concentrated into the clear cream that relative density is 1.20 to 1.22.Cooling; (resin demand is heavy 15 times of cream to the absorption of upper polyamide resin column; the blade diameter length ratio of post is 1:20); with 95% ethanol: 36% acetic acid (volume ratio is 2:1) wash-out, thin-layer chromatography tracing detection, collects NVP-XAA 723 (EGCG) flow point; after charcoal absorption with 2.5% (g/100ml); filter, it is 1.15-1.18 that filtrate low temperature under 60 DEG C-80 DEG C logical nitrogen protections is evaporated to relative density, lyophilize.With ethyl alcohol recrystallization, obtain the NVP-XAA 723 (EGCG) that purity is 98.6%, yield is 12%.
embodiment 3
Green tea tankage 1 kg, adds the concussion of 16L60% EtOH Sonicate and extracts, extract twice, each each 30min, united extraction liquid.Extracting solution less than 80 DEG C decompression recycling ethanols, to without alcohol taste, are concentrated into the clear cream that relative density is 1.20 to 1.22.Cooling; (resin demand is heavy 20 times of cream to the absorption of upper polyamide resin column; the blade diameter length ratio of post is 1:20); with 95% ethanol: 36% acetic acid (volume ratio is 1:1) wash-out, thin-layer chromatography tracing detection, collects NVP-XAA 723 (EGCG) flow point; after charcoal absorption with 3% (g/100ml); filter, it is 1.15-1.18 that filtrate low temperature under 60 DEG C-80 DEG C logical nitrogen protections is evaporated to relative density, lyophilize.With ethyl alcohol recrystallization, obtain the NVP-XAA 723 (EGCG) that purity is 98.9%, yield is 16%.
embodiment 4
Green tea tankage 1 kg, adds the concussion of 20L60% EtOH Sonicate and extracts, extract twice, each each 30min, united extraction liquid.Extracting solution less than 80 DEG C decompression recycling ethanols, to without alcohol taste, are concentrated into the clear cream that relative density is 1.20 to 1.22.Cooling; (resin demand is heavy 20 times of cream to the absorption of upper polyamide resin column; the blade diameter length ratio of post is 1:20); with 95% ethanol: 36% acetic acid (volume ratio is 1:2) wash-out, thin-layer chromatography tracing detection, collects NVP-XAA 723 (EGCG) flow point; after charcoal absorption with 3% (g/100ml); filter, it is 1.15-1.18 that filtrate low temperature under 60 DEG C-80 DEG C logical nitrogen protections is evaporated to relative density, lyophilize.With ethyl alcohol recrystallization, obtain the NVP-XAA 723 (EGCG) that purity is 99.0%, yield is 15%.
The tankage of the green tea in above-described embodiment, comprise in the process of picking tea-leaves some the thick old bars removed outside a large amount of fresh leaf, prune some old tea overstocked in branch or the tea dusts produced in the grade course of processing and storage, tea grounds.
The beneficial effect of above-described embodiment is: the present invention does not need the preliminary purification of macroporous resin, because EGCG is polyphenolic compound, by controlling chromatographic separation condition, adopting a kind of separation system, can directly carry out the separation of polymeric amide chromatography, therefore, by thin-layer chromatography tracing detection, EGCG sterling flow point can be collected, instead of mixture, make separation like this, purifying one step completes, make easy to operation; And in whole process, only used the etoh solvent of food grade, acetic acid and gac, ensure that product safety, nontoxic.
Above-described embodiment is only for illustrating technical conceive of the present invention and feature; its object is to person skilled in the art can be understood content of the present invention and be implemented; can not limit the scope of the invention with this; all equivalences done according to spirit of the present invention change or modify, and all should be encompassed in protection scope of the present invention.
Claims (6)
1. the method for rapid extraction separation and purification NVP-XAA 723 from the tankage of green tea, comprises following steps:
(1) tankage of green tea extract with 60% EtOH Sonicate concussion, and extraction time is 30 minutes, extracts twice, united extraction liquid;
(2) extracting solution reclaims ethanol to without alcohol taste, is concentrated into the clear cream that relative density is 1.20 to 1.22;
(3) the clear cream polyamide resin column fractionation by adsorption will obtained in (2);
(4) thin-layer chromatography tracing detection, collects NVP-XAA 723 flow point, after activated carbon decolorizing, filters, and filtrate is in 60 DEG C of-80 DEG C of low-temperature reduced-pressures, and lead to nitrogen protection, being concentrated into relative density is 1.15-1.18, lyophilize;
(5) with ethyl alcohol recrystallization, highly purified NVP-XAA 723 is obtained;
Elutriant in described step (3) fractionation by adsorption comprises 95% ethanol and 36% acetic acid, wherein 95% ethanol: the volume ratio of 36% acetic acid is 5:1-1:5.
2. method according to claim 1, it is characterized in that: the tankage of described step (1) green tea, comprise in the process of picking tea-leaves some thick old bars, pruning branch or the tea dusts produced in the grade course of processing of removing outside a large amount of fresh leaf and some the old tea overstock in storage, tea grounds.
3. method according to claim 1, is characterized in that: the described tankage of step (1) green tea and the ratio of 60% ethanol are 1:8 g/ml-1:20 g/ml.
4. method according to claim 1, is characterized in that: described step (2) extracting solution reclaims ethanol to without alcohol taste, refers to reclaim under reduced pressure below 80 DEG C.
5. method according to claim 1, is characterized in that: the granularity of described step (3) polyamide resin is 60-100 order, and consumption is: resin: clear cream=10 g:1g-20 g:1g, the blade diameter length ratio of resin column is 1:20.
6. method according to claim 1, is characterized in that: the consumption of described step (4) gac is the 2%-3% of liquor capacity, and the amount of namely adding gac in every 100ml solution is 2g-3g.
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| CN104171157B (en) * | 2014-09-10 | 2016-06-15 | 河源市绿环生物科技有限公司 | A kind of green tea extract and preparation method thereof, application |
| CN104435369A (en) * | 2014-11-28 | 2015-03-25 | 桂林市和胤祥新型材料有限公司 | Preparation method of tea polyphenol |
| CN104926680B (en) * | 2015-05-08 | 2016-08-24 | 李会杰 | The method extracting bioactive substance from low-grade tea and tea making leftover bits and pieces |
| CN109851604A (en) * | 2017-11-30 | 2019-06-07 | 江苏天晟药业股份有限公司 | A kind of Epigallo-catechin gallate (EGCG) crystal form I and preparation method thereof |
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| CN1286252A (en) * | 1999-08-16 | 2001-03-07 | 弗·哈夫曼-拉罗切有限公司 | Process for preparing epigallocatechin gallate |
| CN101575326A (en) * | 2009-06-25 | 2009-11-11 | 合肥工业大学 | Method for extracting tea polyphenol from glede tea |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1286252A (en) * | 1999-08-16 | 2001-03-07 | 弗·哈夫曼-拉罗切有限公司 | Process for preparing epigallocatechin gallate |
| CN101575326A (en) * | 2009-06-25 | 2009-11-11 | 合肥工业大学 | Method for extracting tea polyphenol from glede tea |
Non-Patent Citations (3)
| Title |
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| 柱色谱分离制备无酯儿茶素;刘伟等;《食品工业科技》;20121231;第33卷(第18期);第58-62页 * |
| 聚酰胺色谱法分离制备高纯度表没食子儿茶素没食子酸酯;王传金等;《应用化学》;20070430;第24卷(第4期);第443-447页 * |
| 茶多酚制备高纯度EGCG的工艺研究;朱斌等;《食品研究与开发》;20090430;第30卷(第4期);第7-10页 * |
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