CN1733753A - Epigallocatechin gallate monomer purification method - Google Patents
Epigallocatechin gallate monomer purification method Download PDFInfo
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- CN1733753A CN1733753A CN 200510028723 CN200510028723A CN1733753A CN 1733753 A CN1733753 A CN 1733753A CN 200510028723 CN200510028723 CN 200510028723 CN 200510028723 A CN200510028723 A CN 200510028723A CN 1733753 A CN1733753 A CN 1733753A
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- acid ester
- gallic acid
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Abstract
The invention discloses a new purification preparation method for epigallocatechin gallicin. Wherein, filtering the extract by microporous membrane to pass polar or feeble-polar macropore polystyrene resin; using dilute lower alcohol to resolve; collecting the dissolved part of EGCg; condensing in vacuum, crystallizing, filtering, freeze drying, and obtaining the product with purity more than 98%. This method is simple and nontoxic, needs short production cycle and low cost, and fit to industrial production in scale.
Description
Technical field
The present invention relates to a kind of method for preparing purified of epi-nutgall catechin gallic acid ester monomer.
Background technology
Tea-polyphenol is a kind of natural antioxidant, its main ingredient is a catechin, it is made up of the monomer more than six kinds, be D, L-catechin (D, L-C), L-l-Epicatechol (L-EC), L-epigallocatechin (L-EGC), D, L-l-Epigallocatechol (D, L-GC), L-L-Epicatechin gallate (L-ECG), L-NVP-XAA 723 (L-EGCG) etc.Studies show that L-EGCG is the main high reactivity composition of green tea, have suppress and opposing virus and bacterium, inhibition and preventing cardiovascular disease, prevention and anticancer, prevention and treat radiation injury, effect such as delay senility, can be applicable to fields such as medicine, food, protective foods, makeup.Therefore, research and development epi-nutgall catechin gallic acid ester monomer purifying novel method has important practical significance.
What the extraction separation purification process of epi-nutgall catechin gallic acid ester monomer had been delivered mainly is solvent-extraction process, and ion precipitation method, supercritical extraction, high-speed counter-current layer and membrane separation technique attached gel post separate preparation method.Solvent-extraction process is to leach from tealeaves with polar solvent, and used organic solvent is as acetone, ether, methyl alcohol, ethanol, hexane and trichloromethane etc.This method is used multiple organic solvent, and some deleterious organic solvent makes product safe not to the utmost with operation, and easily causes environmental pollution.The ion precipitation method is to utilize metal can precipitate tea-polyphenol, and it is separated with trimethyl-xanthine, as mantoquita, lead salt or aluminum chloride.This method has been used has the metal of harm to make precipitation agent to human body, and its product is that food and medicine industry can not be accepted.The supercritical extraction investment is big, is unfavorable for suitability for industrialized production.High-speed counter-current layer and membrane separation technique attached gel column chromatography, technological process is complicated, and column chromatography generally uses gel chromatography, and reagent consumption costs an arm and a leg greatly, and preparation amount is little and the cycle is long, obviously the improper large-scale industrial production that is used for.
Summary of the invention
The objective of the invention is to overcome the deficiency of prior art purifying epi-nutgall catechin gallic acid ester monomer method and technology and cost aspect, a kind of product purity height, with short production cycle is provided, simple, purifying process that can suitability for industrialized production.
With green tea extract, wherein catechin content is 60～80%, EGCG content is 40～60%, (for example nitrogen, helium, argon gas or hydrogen) is dissolved in the deionized water of 20～50 times of amounts in the presence of non-oxidizing gas, solution is filtered through 1-0.45 micron membranes strainer, remove undissolved impurity.Adopt the nonpolar or low-pole resin of macroporous type polystyrene, they are the D-101 that buys on the market, ADS-5, AB-8 or D-130 adorn column chromatography as stopping composition, under 20～60 ℃, 5%～40% low-alcohol solution is resolved, according to the ultraviolet detection result, collect the flow point of NVP-XAA 723, with the flow point collected at 35～70 ℃ of vacuum concentration to small volume, place, separate out crystallization, recrystallization, recrystallization solvent are methyl alcohol, ethanol, n-propyl alcohol, Virahol, ethylene glycol, propylene glycol or glycerol.Filtration, lyophilize get finished product, and this finished product detects through HPLC, and the EGCG product purity is greater than 98%, no agricultural chemicals and dissolvent residual.
What present method adopted is, with the non-oxidizing gas secluding air, under 20～60 ℃ of temperature condition, the catechin compounds such as nonpolar or low-pole resin absorption EGCg with the macroporous type polystyrene, utilize the nonpolar or low-pole resin of macroporous type polystyrene can to go out by wash-out under the lower alcohol condition of suitable concn, lower alcohol is C
1-C
3The principle of primary alconol or secondary alcohol EGCg.Effectively separate components such as EGCg, caffeine, the purity that can obtain EGCg greater than 98% theoretical yield 50 ~ 80%.
Prior art how with the method for gel filtration chromatography and high speed adverse current chromatogram separate, purifying, adopt chloroform, ethyl acetate equal solvent more, cause product dissolvent residual height, chromatography adopts imported product with filler more, the cost height is unfavorable for suitability for industrialized production.The present invention adopts domestic resin as column packing, and alcohol is desorbed solution, and product does not have the poisonous and harmful dissolvent residual, and cost is lower, is easy to suitability for industrialized production.
Description of drawings
Fig. 1 is the purity with the HPLC testing product.
Embodiment(per-cent that the present invention relates to all by weight percentage)
Embodiment one
The tea-polyphenol extract that takes by weighing 20g content and be 57%EGCg places a bottle with two necks, a mouth of bottle with two necks feed make when another mouthful of nitrogen derived gas in the 20g solid dissolving 120ml deionized water standby.Get D101 resin 2000ml and adorn post,, after concentrated, use the gas chromatography detected result until eluent with pore-creating agent and the polymerization single polymerization monomer remnants in the ethanol elution resin with ethanol, relatively do not have pore-creating agent and polymerization single polymerization monomer remnants with initial sample till.It is will be through nitrogen saturated and replace in the presence of nitrogen to replace ethanol in post water in this process with deionized water again.Ethanol in the post by wholly replace after, in the presence of nitrogen, standby solution annotated on the cylinder with 0.4 times of volume/hour flow velocity on sample, after treating that sample enters cylinder, the deionized water of at first using 2 times of volumes is with 0.2 times of volume/hour flow velocity wash-out, after washing, use 25% ethanol elution instead, elution volume is 3 times of volumes, through UV-detector and HPLC indication, collect EGCG stream part.Stream part merges, and to small volume, drips ethanol (AR) at 45 ℃ of following vacuum concentration in concentrating bottle, and heat makes to transfer to after the dissolving and is stored in the triangular flask in the refrigerator a little, to be crystallized finish to take out filter drying.Weighing gets 6.12g, and productive rate is 53.68%.Detecting purity through HPLC is 98.7%.
Embodiment two
The tea-polyphenol extract that takes by weighing 40g content and be 44%EGCg places a bottle with two necks, a mouth of bottle with two necks feed make when another mouthful of helium derived gas in the 40g solid dissolving 240ml deionized water standby.Get AB-8 resin 3000ml and adorn post,, after concentrated, use the gas chromatography detected result until eluent with pore-creating agent and the polymerization single polymerization monomer remnants in the ethanol elution resin with ethanol, relatively do not have pore-creating agent and polymerization single polymerization monomer remnants with initial sample till.It is will be through helium saturated and replace in the presence of helium to replace ethanol in post water in this process with deionized water again.Ethanol in the post by wholly replace after, in the presence of helium, standby solution annotated on the cylinder with 0.4 times of volume/hour flow velocity on sample, after treating that sample enters cylinder, the deionized water of at first using 2 times of volume volumes after washing, is used 2 times of volumes of 5% Virahol wash-out with 0.2 times of volume/hour flow velocity wash-out instead, then with 20% Virahol wash-out, elution volume is 3 times of volumes, detects indication through UV-detector in conjunction with HPLC, collects EGCG stream part.Stream part merges, and to small volume, drips ethanol (AR) at 45 ℃ of following vacuum concentration in concentrating bottle, and heat makes to transfer to after the dissolving and is stored in the triangular flask in the refrigerator a little, to be crystallized finish to take out filter drying.Weighing gets 14.25g, and productive rate is 80.97%.Detecting purity through HPLC is 98.2%.
Embodiment three
The tea-polyphenol extract that takes by weighing 25g content and be 65%EGCg places a bottle with two necks, a mouth of bottle with two necks feed make when another mouthful of argon gas derived gas in the 25g solid dissolving 150ml deionized water standby.Get D-130 resin 2000ml and adorn post,, after concentrated, use the gas chromatography detected result until eluent with pore-creating agent and the polymerization single polymerization monomer remnants in the ethanol elution resin with ethanol, relatively do not have pore-creating agent and polymerization single polymerization monomer remnants with initial sample till.It is will be through argon gas saturated and replace in the presence of argon gas to replace ethanol in post water in this process with deionized water again.Ethanol in the post by wholly replace after, in the presence of argon gas, standby solution annotated on the cylinder with 0.4 times of volume/hour flow velocity on sample, after treating that sample enters cylinder, the deionized water of at first using 2 times of volumes after washing, is used 2 times of volumes of 7% methanol-eluted fractions with 0.2 times of volume/hour flow velocity wash-out instead, then use 24% methanol-eluted fractions, elution volume is 3 times of volumes, detects indication through UV-detector in conjunction with HPLC, collects EGCG stream part.Stream part merges, and to small volume, drips ethanol (AR) at 45 ℃ of following vacuum concentration in concentrating bottle, and heat makes to transfer to after the dissolving and is stored in the triangular flask in the refrigerator a little, to be crystallized finish to take out filter drying.Weighing gets 10.24g, and productive rate is 63.15%.Detecting purity through HPLC is 98.0%.
Claims (6)
1. the method for preparing purified of an epi-nutgall catechin gallic acid ester monomer comprises the steps:
(1) with green tea extract in the presence of non-oxidizing gas, be dissolved in the saturated deionized water of non-oxidizing gas, solution filters through micron membranes, filtrate for later use;
(2) adopt macroporous type polarity or low-pole polystyrene resin dress post, resolve with 5%～40% alcoholic solution;
(3) adopt ultraviolet detection to collect the NVP-XAA 723 flow point, collection liquid, is placed and is separated out crystallization to small volume through 35～70 ℃ of vacuum concentration, the lower alcohol recrystallization, and HPLC detects.
2. the method for preparing purified of a kind of epi-nutgall catechin gallic acid ester monomer according to claim 1 is characterized in that non-oxidizing gas can be helium, argon gas, nitrogen and hydrogen.
3. the method for preparing purified of a kind of epi-nutgall catechin gallic acid ester monomer according to claim 1, it is characterized in that adopting the nonpolar or low-pole resin of macroporosity polystyrene is D-101, ADS-5, AB-8 and D-130.
4. the method for preparing purified of a kind of epi-nutgall catechin gallic acid ester monomer according to claim 1 is characterized in that resolving with the 5%-40%v/v low-alcohol solution.
5. the change preparation method of a kind of epi-nutgall catechin gallic acid ester monomer according to claim 1 is characterized in that described lower alcohol is C
1-C
3Primary alconol or secondary alcohol.
6. the method for preparing purified of a kind of epi-nutgall catechin gallic acid ester monomer according to claim 1 is characterized in that the lower alcohol recrystallization solvent is methyl alcohol, ethanol, n-propyl alcohol, Virahol, ethylene glycol, propylene glycol or glycerol.
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102311419A (en) * | 2011-09-09 | 2012-01-11 | 四川天予植物药业有限公司 | Refining and purification method of high content EGCG |
| CN101703130B (en) * | 2009-12-03 | 2012-05-02 | 应维强 | Technology for producing green tea extract |
| CN101492440B (en) * | 2008-01-24 | 2012-10-03 | 上海新康制药厂 | Separation purification process for main catechin component in tea polyphenol and glycosidase activity |
| CN102702163A (en) * | 2012-06-01 | 2012-10-03 | 广西济康生物科技有限公司 | Method for preparing high-purity monomer epigallocatechin gallate from processed leftovers of tea leaves |
| CN106967036A (en) * | 2017-04-19 | 2017-07-21 | 盛林 | A kind of EGCG preparation method |
| CN107556285A (en) * | 2017-10-31 | 2018-01-09 | 桂林纽泰生物科技有限公司 | The method that Epigallo-catechin gallate (EGCG) is extracted from litchi rind |
| CN107805235A (en) * | 2017-10-31 | 2018-03-16 | 桂林纽泰生物科技有限公司 | The extraction process of litchi rind Epigallo-catechin gallate (EGCG) |
| WO2019105216A1 (en) * | 2017-11-30 | 2019-06-06 | 江苏天晟药业股份有限公司 | Epigallocatechin gallate crystal form i and method for preparing same |
-
2005
- 2005-08-12 CN CNB2005100287233A patent/CN100482655C/en active Active
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101492440B (en) * | 2008-01-24 | 2012-10-03 | 上海新康制药厂 | Separation purification process for main catechin component in tea polyphenol and glycosidase activity |
| CN101703130B (en) * | 2009-12-03 | 2012-05-02 | 应维强 | Technology for producing green tea extract |
| CN102311419A (en) * | 2011-09-09 | 2012-01-11 | 四川天予植物药业有限公司 | Refining and purification method of high content EGCG |
| CN102702163A (en) * | 2012-06-01 | 2012-10-03 | 广西济康生物科技有限公司 | Method for preparing high-purity monomer epigallocatechin gallate from processed leftovers of tea leaves |
| CN106967036A (en) * | 2017-04-19 | 2017-07-21 | 盛林 | A kind of EGCG preparation method |
| CN106967036B (en) * | 2017-04-19 | 2022-03-22 | 盛林 | Preparation method of EGCG |
| CN107556285A (en) * | 2017-10-31 | 2018-01-09 | 桂林纽泰生物科技有限公司 | The method that Epigallo-catechin gallate (EGCG) is extracted from litchi rind |
| CN107805235A (en) * | 2017-10-31 | 2018-03-16 | 桂林纽泰生物科技有限公司 | The extraction process of litchi rind Epigallo-catechin gallate (EGCG) |
| WO2019105216A1 (en) * | 2017-11-30 | 2019-06-06 | 江苏天晟药业股份有限公司 | Epigallocatechin gallate crystal form i and method for preparing same |
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|---|---|
| CN100482655C (en) | 2009-04-29 |
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