CN107739399B - Method for extracting madecassic acid - Google Patents
Method for extracting madecassic acid Download PDFInfo
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- CN107739399B CN107739399B CN201711227437.9A CN201711227437A CN107739399B CN 107739399 B CN107739399 B CN 107739399B CN 201711227437 A CN201711227437 A CN 201711227437A CN 107739399 B CN107739399 B CN 107739399B
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Abstract
The invention discloses a method for extracting madecassic acid, which comprises the following steps of 1) crushing centella, carrying out reflux extraction by using an ethanol solution, filtering an extracting solution, recovering a solvent from a filtrate, dissolving the filtrate in water at 40-50 ℃, filtering, and dissolving filter residues in a 50-60 v% ethanol solution to obtain a crude extracting solution; 2) loading the crude extract on XAD-9 macroporous resin column, eluting with acid solution, and mixing effluent and acid washing solution; 3) adjusting the pH value of the combined solution to be neutral, filtering, introducing the filtrate into an XAD-3 macroporous resin column again, eluting with ethanol with the volume concentration of 30-40%, tracking and detecting by thin layer chromatography, and collecting ethanol eluate containing asiatic acid; and eluting with 55-65% ethanol, tracking and detecting by thin-layer chromatography, collecting ethanol eluent containing madecassic acid, recovering ethanol from the madecassic acid eluent, and drying to obtain madecassic acid. The madecassic acid extracted by the method has high purity and high yield.
Description
Technical Field
The invention belongs to the technical field of biological extraction, and particularly relates to a method for extracting madecassic acid from madecassic acid.
Background
Centella asiatica is a dry whole plant of Centella asiatica (L.) Urban of Umbelliferae, is widely distributed in the south of Yangtze river basin, is used as a medicine, can be collected all the year round, is rich in resources, has been applied to the medical field of many countries for thousands of years, is listed as a middle-quality product in Shen nong Ben Cao Jing, is bitter in taste and pungent in flavor, has the effects of clearing heat and promoting diuresis, and detoxifying and reducing swelling, and is mainly used for damp-heat jaundice, sunstroke diarrhea, carbuncle and sore toxin, traumatic injury and the like. The chemical components of centella asiatica include triterpenoid saponin, triterpenic acid, polyacetylenic group, volatile oil, etc. Modern researches prove that the centella asiatica extract can effectively promote skin injury and local collagen anabolism, and has a traditional Chinese medicine effect on tissue repair after skin injury. In recent years, the research also finds that the centella asiatica has the effects of resisting oxidation, resisting depression, protecting the liver, inhibiting tumor cell proliferation and the like. The main pharmacological component of centella asiatica is triterpenoid, so the research on extracting and separating the active components mainly focuses on separating and purifying asiatic acid and madecassic acid more quickly and better. The conventional extraction methods of triterpenoids are various, generally mainly solvent extraction, including extraction method, reflux method, decoction method and the like, and recently, some new extraction methods, including microwave method, enzyme method and the like, have been reported and have also gained wide attention. In the aspect of purification, the conventional column chromatography method for purifying the triterpenoids still occupies the mainstream, and other methods such as chromatography methods are rarely utilized due to the fact that the materials are more consumed and the cost is high.
Disclosure of Invention
The technical problem solved by the invention is to provide a method for extracting madecassic acid, and the madecassic acid extracted by the method has high yield and high purity.
The technical scheme provided by the invention is to provide a method for extracting madecassic acid, which comprises the following steps:
1) crushing centella asiatica, performing reflux extraction by using a 60-70 v% ethanol solution, filtering an extracting solution, recovering a solvent from filtrate, dissolving the solvent in water at 40-50 ℃, filtering, and dissolving filter residues in a 50-60 v% ethanol solution to obtain a crude extracting solution;
2) putting the crude extract into an XAD-9 macroporous resin column, eluting with acid liquor with the pH value of 2-4, and combining effluent liquid and acid washing liquor;
3) adjusting the pH value of the combined solution to be neutral, filtering, introducing the filtrate into an XAD-3 macroporous resin column again, eluting with ethanol with the volume concentration of 30-40%, tracking and detecting by thin layer chromatography, and collecting ethanol eluate containing asiatic acid; and eluting with 55-65% ethanol, tracking and detecting by thin-layer chromatography, collecting ethanol eluent containing madecassic acid, recovering ethanol from the madecassic acid eluent, and drying to obtain madecassic acid.
In the step 1), the ethanol with the volume concentration of 60-70% can fully dissolve triterpenoids in the centella asiatica, including triterpenoid saponins and triterpenoid acid substances. Because the asiaticoside molecules are often combined with a plurality of sugar molecules, the asiaticoside molecules have a large number of hydroxyl groups, and can show certain hydrophilicity, but the asiaticoside molecules have weak hydrophilicity and are slightly soluble in water, while the triterpenic acids have small polarity and cannot be dissolved in water. Therefore, the asiatic centella total saponins can be dissolved by warm water at 40-50 ℃, the asiatic centella triterpenic acid substances are insoluble in hot water, the filtrate is filtered, and the filter residue is dissolved in ethanol, so that the aim of removing the asiatic centella total saponins can be fulfilled.
XAD-9 resin mainly adsorbs liposoluble components and substances with strong macromolecule polarity, which have stronger adsorbability to the resin, and macroporous resin preferentially adsorbs the substances to quickly reach saturation. XAD-9 is a polar macroporous resin, has a polarity which is different from that of triterpenic acid, can not effectively adsorb the triterpenic acid, has a larger average pore diameter which is far larger than that of the triterpenic acid, and can not play a role in physically trapping the triterpenic acid, so that the triterpenic acid flows out along with effluent liquid. Acid liquor with the pH value of 2-4 can further elute a small amount of triterpenic acid compounds adsorbed in the resin, so that adsorption loss of the triterpenic acid compounds is avoided. In the step 2), the column feeding speed of the crude extract is 3-5 BV/h. The elution speed of the acid solution is 1-2 BV/h.
In step 3), the XAD-3 macroporous resin is nonpolar resin, the average pore diameter is 0.44 μm, and the specific surface area is 256m2(ii) in terms of/g. The resin has very good adsorption performance on the triterpenic acid compounds, the column loading speed of the filtrate is 0.5-1 BV/h, and the triterpenic acid compounds can be completely adsorbed. Because asiatic acid and madecassic acid have polarity difference, the asiatic acid can be preferentially eluted by adopting ethanol with the volume concentration of 30-40%. Then, the madecassic acid is eluted by using ethanol with the volume concentration of 55-65%, and the rest madecassic acid can be eluted.
Compared with the prior art, the method carries out adsorption and desorption on the target component, the yield of the extracted madecassic acid is up to more than 90%, and the purity of the extracted madecassic acid is up to more than 95.0%.
Detailed Description
The following specific examples further illustrate the invention but are not intended to limit the invention thereto.
The following percentages are by volume.
Example 1
1) Pulverizing herba Centellae, reflux-extracting with 60 v% ethanol solution, wherein the amount of 60 v% ethanol is 5 times of the weight of herba Centellae, reflux-extracting at 40 deg.C for 1 time, each time for 0.5 hr, filtering the extractive solution, recovering solvent from the filtrate, dissolving in 40 deg.C water, filtering, dissolving the residue in 50 v% ethanol solution to obtain crude extractive solution;
2) loading the crude extractive solution onto XAD-9 macroporous resin column at a speed of 3BV/h, eluting with acid solution with pH of 2 at a speed of 1BV/h, and mixing effluent and acid solution;
3) adjusting pH of the combined solution to neutral, filtering, loading the filtrate on XAD-3 macroporous resin column at a speed of 0.5BV/h, eluting with 30% ethanol at a speed of 2BV/h, performing thin layer chromatography tracking detection, and collecting ethanol eluate containing asiatic acid component; eluting with 55% ethanol at an ethanol elution speed of 2BV/h, performing thin-layer chromatography tracking detection, collecting ethanol eluate containing madecassic acid, recovering ethanol from madecassic acid eluate, and drying to obtain madecassic acid.
HLPC analysis shows that the recovery rate of madecassic acid is above 90.08%, and the purity is as high as 95.27%.
Example 2
1) Pulverizing herba Centellae, reflux-extracting with 70 v% ethanol solution, wherein the amount of 70 v% ethanol is 10 times of the weight of herba Centellae, reflux-extracting at 50 deg.C for 3 times, each time for 1 hr, filtering the extractive solution, recovering solvent from the filtrate, dissolving in 50 deg.C water, filtering, and dissolving the residue in 60 v% ethanol solution to obtain crude extractive solution;
2) loading the crude extractive solution onto XAD-9 macroporous resin column at 5BV/h, eluting with acid solution of pH 4 at 2BV/h, and mixing effluent and acid solution;
3) adjusting pH of the combined solution to neutral, filtering, loading the filtrate on XAD-3 macroporous resin column at a speed of 1BV/h, eluting with 40% ethanol at a speed of 3BV/h, detecting by thin layer chromatography, and collecting ethanol eluate containing asiatic acid; eluting with 65% ethanol at a speed of 3BV/h, detecting by thin layer chromatography, collecting ethanol eluate containing madecassic acid, recovering ethanol from madecassic acid eluate, and drying to obtain madecassic acid.
HLPC analysis shows that the recovery rate of madecassic acid is more than 90.11%, and the purity is as high as 95.46%.
Example 3
1) Crushing centella asiatica, performing reflux extraction by using a 65 v% ethanol solution, performing reflux extraction for 2 times at 45 ℃ with the use amount of 65 v% ethanol being 8 times of the weight of the centella asiatica, each time for 0.8h, filtering an extracting solution, recovering a solvent from filtrate, dissolving the filtrate in water at 40-50 ℃, filtering, and dissolving filter residues in a 50-60 v% ethanol solution to obtain a crude extracting solution;
2) loading the crude extractive solution onto XAD-9 macroporous resin column at a speed of 4BV/h, eluting with acid solution with pH of 3 at a speed of 1.5BV/h, and mixing effluent and acid washing solution;
3) adjusting pH of the combined solution to neutral, filtering, loading the filtrate on XAD-3 macroporous resin column at a speed of 0.8BV/h, eluting with 35% ethanol at a speed of 2.5BV/h, performing thin layer chromatography tracking detection, and collecting ethanol eluate containing asiatic acid component; eluting with 60% ethanol at an ethanol elution speed of 2.5BV/h, performing thin layer chromatography tracking detection, collecting ethanol eluate containing madecassic acid, recovering ethanol from madecassic acid eluate, and drying to obtain madecassic acid.
HLPC analysis shows that the recovery rate of madecassic acid is above 90.71%, and the purity is as high as 95.69%.
Example 4
1) Pulverizing herba Centellae, reflux-extracting with 60 v% ethanol solution, wherein the amount of 60 v% ethanol is 10 times of the weight of herba Centellae, reflux-extracting at 40 deg.C for 3 times, each time for 0.5 hr, filtering the extractive solution, recovering solvent from the filtrate, dissolving in 50 deg.C water, filtering, and dissolving the residue in 50 v% ethanol solution to obtain crude extractive solution;
2) putting the crude extract on an XAD-9 macroporous resin column, wherein the column putting speed of the crude extract is 5BV/h, eluting with acid liquor with the pH value of 2-4, and the elution speed of the acid liquor is 1BV/h, and combining effluent liquid and acid liquor;
3) adjusting pH of the combined solution to neutral, filtering, loading the filtrate on XAD-3 macroporous resin column at a speed of 1BV/h, eluting with 30% ethanol at a speed of 3BV/h, detecting by thin layer chromatography, and collecting ethanol eluate containing asiatic acid; eluting with 55% ethanol at an ethanol elution speed of 3BV/h, tracking and detecting by thin layer chromatography, collecting ethanol eluate containing madecassic acid, recovering ethanol from madecassic acid eluate, and drying to obtain madecassic acid.
HLPC analysis shows that the recovery rate of madecassic acid is more than 90.17%, and the purity is as high as more than 95.83%.
Claims (6)
1. The extraction method of madecassic acid is characterized by comprising the following steps: the method comprises the following steps:
1) crushing centella asiatica, performing reflux extraction by using a 60-70 v% ethanol solution, filtering an extracting solution, recovering a solvent from filtrate, dissolving the solvent in water at 40-50 ℃, filtering, and dissolving filter residues in a 50-60 v% ethanol solution to obtain a crude extracting solution;
2) putting the crude extract into an XAD-9 macroporous resin column, eluting with acid liquor with the pH value of 2-4, and combining effluent liquid and acid washing liquor;
3) adjusting the pH value of the combined solution to be neutral, filtering, introducing the filtrate into an XAD-3 macroporous resin column again, eluting with ethanol with the volume concentration of 30-40%, tracking and detecting by thin layer chromatography, and collecting ethanol eluate containing asiatic acid; and eluting with 55-65% ethanol, tracking and detecting by thin-layer chromatography, collecting ethanol eluent containing madecassic acid, recovering ethanol from the madecassic acid eluent, and drying to obtain madecassic acid.
2. The method for extracting madecassic acid according to claim 1, characterized in that: in the step 1), the using amount of 60-70 v% ethanol is 5-10 times of the weight of centella asiatica, and the reflux extraction is carried out for 1-3 times at 40-50 ℃ for 0.5-1 h each time.
3. The method for extracting madecassic acid according to claim 1, characterized in that: in the step 2), the column feeding speed of the crude extract is 3-5 BV/h.
4. The method for extracting madecassic acid according to claim 1, characterized in that: in the step 2), the elution speed of the acid solution is 1-2 BV/h.
5. The method for extracting madecassic acid according to claim 1, characterized in that: in the step 3), the column feeding speed of the filtrate is 0.5-1 BV/h.
6. The method for extracting madecassic acid according to claim 1, characterized in that: in the step 3), the ethanol elution speed is 2-3 BV/h.
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Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101991624A (en) * | 2009-08-27 | 2011-03-30 | 上海新康制药厂 | Method for preparing total asiatic acid, asiatic acid and madecassic acid from asiatic pennywort herb and use of prepared product |
| CN102389437A (en) * | 2011-06-24 | 2012-03-28 | 中国人民解放军第二军医大学 | Application of Centella triterpenes in preparing products for improving cerebral apoplexy sequelae |
| CN106336448A (en) * | 2016-08-25 | 2017-01-18 | 桂林益天成生物科技有限公司 | Method for extracting madecassic acid from herba centellae |
| CN106349320A (en) * | 2016-08-25 | 2017-01-25 | 桂林益天成生物科技有限公司 | Method for extracting asiatic acid from centella |
| KR20170050062A (en) * | 2015-10-29 | 2017-05-11 | 최경석 | Composition for Beverage and Cosmetics Containing Asiaticoside Madecassoside Asiatic acid Madecasic acid Extracted from Centella Asiatica Manufacturing Method Thereof |
| CN106967145A (en) * | 2017-04-28 | 2017-07-21 | 南宁馨艺荣生物科技有限公司 | A kind of technique that brahmic acid is extracted from centella |
-
2017
- 2017-11-29 CN CN201711227437.9A patent/CN107739399B/en active Active
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101991624A (en) * | 2009-08-27 | 2011-03-30 | 上海新康制药厂 | Method for preparing total asiatic acid, asiatic acid and madecassic acid from asiatic pennywort herb and use of prepared product |
| CN102389437A (en) * | 2011-06-24 | 2012-03-28 | 中国人民解放军第二军医大学 | Application of Centella triterpenes in preparing products for improving cerebral apoplexy sequelae |
| KR20170050062A (en) * | 2015-10-29 | 2017-05-11 | 최경석 | Composition for Beverage and Cosmetics Containing Asiaticoside Madecassoside Asiatic acid Madecasic acid Extracted from Centella Asiatica Manufacturing Method Thereof |
| CN106336448A (en) * | 2016-08-25 | 2017-01-18 | 桂林益天成生物科技有限公司 | Method for extracting madecassic acid from herba centellae |
| CN106349320A (en) * | 2016-08-25 | 2017-01-25 | 桂林益天成生物科技有限公司 | Method for extracting asiatic acid from centella |
| CN106967145A (en) * | 2017-04-28 | 2017-07-21 | 南宁馨艺荣生物科技有限公司 | A kind of technique that brahmic acid is extracted from centella |
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Effective date of registration: 20211220 Address after: 214100 2-307 and 308, Tian'an smart city, No. 228, Linghu Avenue, Xinwu District, Wuxi City, Jiangsu Province Patentee after: Wuxi daikesi Biotechnology Co.,Ltd. Address before: 541004 pavement 1-4 and 2-4, 1 Tung ring new village, Putuo Road, East Ring Road, Qixing District, Guilin, the Guangxi Zhuang Autonomous Region Patentee before: GUILIN RONGTONG TECHNOLOGY Co.,Ltd. |