CN101037467B - Method for separating purifying ursolic acid from loquat leaf - Google Patents
Method for separating purifying ursolic acid from loquat leaf Download PDFInfo
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- CN101037467B CN101037467B CN200710008809A CN200710008809A CN101037467B CN 101037467 B CN101037467 B CN 101037467B CN 200710008809 A CN200710008809 A CN 200710008809A CN 200710008809 A CN200710008809 A CN 200710008809A CN 101037467 B CN101037467 B CN 101037467B
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- ursolic acid
- acid
- folium eriobotryae
- organic solvent
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- WCGUUGGRBIKTOS-RRHGHHQTSA-N Ursolic acid Natural products O=C(O)[C@@]12[C@@H]([C@@H](C)[C@@H](C)CC1)C=1[C@](C)([C@@]3(C)[C@@H]([C@]4(C)[C@H](C(C)(C)[C@@H](O)CC4)CC3)CC=1)CC2 WCGUUGGRBIKTOS-RRHGHHQTSA-N 0.000 title claims abstract description 413
- 229940096998 ursolic acid Drugs 0.000 title claims abstract description 206
- 241001092070 Eriobotrya Species 0.000 title claims description 75
- 235000009008 Eriobotrya japonica Nutrition 0.000 title claims description 74
- 239000002253 acid Substances 0.000 claims abstract description 90
- 239000000284 extract Substances 0.000 claims abstract description 57
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- 239000002904 solvent Substances 0.000 claims abstract description 30
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 21
- 238000000605 extraction Methods 0.000 claims abstract description 14
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- 238000001953 recrystallisation Methods 0.000 claims abstract description 12
- OKTJSMMVPCPJKN-UHFFFAOYSA-N carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims abstract description 9
- 150000007513 acids Chemical class 0.000 claims description 79
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- 239000000706 filtrate Substances 0.000 claims description 55
- 239000012043 crude product Substances 0.000 claims description 52
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 43
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- 238000005057 refrigeration Methods 0.000 claims description 31
- HEDRZPFGACZZDS-UHFFFAOYSA-N chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims description 26
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- 238000000034 method Methods 0.000 abstract description 2
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- MIJYXULNPSFWEK-GTOFXWBISA-N 3β-hydroxy-Olean-12-en-28-oic acid Chemical compound C1C[C@H](O)C(C)(C)[C@@H]2CC[C@@]3(C)[C@]4(C)CC[C@@]5(C(O)=O)CCC(C)(C)C[C@H]5C4=CC[C@@H]3[C@]21C MIJYXULNPSFWEK-GTOFXWBISA-N 0.000 description 11
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- MIJYXULNPSFWEK-LXLHWSNGSA-N Oleanolic acid Natural products C1C[C@H](O)C(C)(C)[C@H]2CC[C@@]3(C)[C@]4(C)CC[C@@]5(C(O)=O)CCC(C)(C)C[C@@H]5C4=CC[C@H]3[C@]21C MIJYXULNPSFWEK-LXLHWSNGSA-N 0.000 description 10
- 239000012153 distilled water Substances 0.000 description 9
- 241000196324 Embryophyta Species 0.000 description 7
- 238000002360 preparation method Methods 0.000 description 7
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 6
- RTZKZFJDLAIYFH-UHFFFAOYSA-N diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 6
- 238000005516 engineering process Methods 0.000 description 6
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- ZWEHNKRNPOVVGH-UHFFFAOYSA-N 2-butanone Chemical compound CCC(C)=O ZWEHNKRNPOVVGH-UHFFFAOYSA-N 0.000 description 3
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- XTLNYNMNUCLWEZ-UHFFFAOYSA-N ethanol;propan-2-one Chemical compound CCO.CC(C)=O XTLNYNMNUCLWEZ-UHFFFAOYSA-N 0.000 description 2
- 150000008131 glucosides Chemical group 0.000 description 2
- 150000007522 mineralic acids Chemical class 0.000 description 2
- 235000005985 organic acids Nutrition 0.000 description 2
- 239000002002 slurry Substances 0.000 description 2
- 239000000341 volatile oil Substances 0.000 description 2
- QARAXUHORATRQJ-UHFFFAOYSA-N 1,4-dioxane;pyridine Chemical compound C1COCCO1.C1=CC=NC=C1 QARAXUHORATRQJ-UHFFFAOYSA-N 0.000 description 1
- XUCIJNAGGSZNQT-SWRVSKMJSA-N Amygdalin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC[C@@H]1[C@@H](O)[C@H](O)[C@@H](O)[C@H](OC(C#N)C=2C=CC=CC=2)O1 XUCIJNAGGSZNQT-SWRVSKMJSA-N 0.000 description 1
- 235000012871 Arctostaphylos uva ursi Nutrition 0.000 description 1
- 235000016855 Biancospino Nutrition 0.000 description 1
- 239000004215 Carbon black (E152) Substances 0.000 description 1
- 229920001429 Chelating resin Polymers 0.000 description 1
- 235000017181 Crataegus chrysocarpa Nutrition 0.000 description 1
- 235000004423 Crataegus monogyna Nutrition 0.000 description 1
- 240000000171 Crataegus monogyna Species 0.000 description 1
- 240000005967 Eriobotrya japonica Species 0.000 description 1
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- NBIIXXVUZAFLBC-UHFFFAOYSA-K [O-]P([O-])([O-])=O Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
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- CURLTUGMZLYLDI-UHFFFAOYSA-N carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 1
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- 238000004156 green chemistry Methods 0.000 description 1
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Abstract
The invention provides a method for separating the ursolic acid from the folium eriobotryae, which is characterized in that: having the folium eriobotryae as material, extracting in the water solutionby the hydrophiled organic solvent, decompressed recovering the solvent, concentrating the folium eriobotryae total triterpenic acid extract; removing the impurities by solvent, active carbon decoloring, precipitation separation of the triterpenic acid extract to produce the coarse folium eriobotryae total triterpenic acid extract with the ursolic acid concentration >=25%; enrichment and purification, precipitation separation to the beige coarse ursolic acid with the ursolic acid concentration >=80%; extraction by hydrophilic organic solvent, crystalline purification and recrystallization ofthe coarse ursolic acid, and finally getting the purified ursolic acid crystal. The purity of the white ursolic acid crystal is >=99%. The invention has a simple producing process, a high separation efficiency, a low cost and a good extension.
Description
Technical field
The present invention relates to a kind of Green Chemistry production method of from plant, extracting active pharmaceutical ingredients, more specifically relate to the method for separating and purifying ursolic acid from Loquat Leaf.
Background technology
China is topmost in the world loquat producing country, and loquat cultivated area and output account for more than 2/3 of the whole world, and its main producing region is the Putian, Fujian, Zhe Jiangtang is dwelt and mountain, Dongting Lake, Jiangsu.160,000 mu of the existing planting loquat areas in Putian, Fujian, account for the whole world 1/6,1/4 and the Fujian Province in the whole nation 1/2.
The loquat whole body is all precious, Loquat Leaf, flower, really, position such as nuclear is rich in triterpenic acid (can up to 1.5%), volatile oil, amygdaloside, saponin(e, flavones, water-soluble polysaccharide and organic acid isoreactivity composition.At present, loquat is based on fresh fruit supply, does not carry out the deep development and the comprehensive utilization at other position as yet.
The leaf of loquat leaf system rosaceous plant loquat [Eriobotrya japonica (Thunb.) Lindl.], be rich in ursolic acid (Ursolic acid, UA) and Oleanolic Acid (Oleanolic acid, triterpenic acid such as OA), cold nature, bitter suffering, relieving cough and asthma, clearing lung-heat and stomach, lowering the adverse-rising QI to resolve phlegm records in going through an edition Chinese Pharmacopoeia.The tradition tcm clinical practice often becomes compound medicines to be used as medicine with sweet honey-fried FOLIUM ERIABOTRYAE or Loquat Leaf with other drug matching, has obtained certain preventing phlegm from forming and stopping coughing, anti-inflammatory clinical efficacy such as relieving asthma.
Ursolic acid and Oleanolic Acid are two isomerss, are pentacyclic triterpenoid, and molecular formula is C
30H
48O
3, character is very similar, and usually coexistence brings certain difficulty to separation determination in plant.Infrared spectra is according to regional A (1355～1392cm
-1) and area B (1245～1330cm
-1) hydrocarbon absorption distinguish framework types, Oleanolic Acid has two peaks in the A district, there are three peaks in the B district, ursolic acid respectively has three peaks in A district and B district, can distinguish both with this.
Because the complex structure of ursolic acid and Oleanolic Acid, the synthetic difficulty is all extracted in herbal medicine or plant both at home and abroad.Ursolic acid and Oleanolic Acid are white crystalline powder, 310 ℃ of fusing points, and odorless, tasteless, be soluble in the dioxane pyridine, dissolve in methyl alcohol, ethanol isopolarity organic solvent, be slightly soluble in fat-soluble solvents such as chloroform, ether, be insoluble to sherwood oil and water.
Ursolic acid is distributed widely in about 27 section's 62 kind of plant with free form or glucosides form, in plants such as hawthorn, black bearberry, dark plum, Sambucus chinensis, Spreading Hedyotis Herb, Loquat Leaf, psyllium, Glossy Privet Fruit; Oleanolic Acid extensively is present in about 60 section's 190 kind of plant with free form or glucosides form, in plants such as east area of the Liao River, coextensive with eastern and southern Liaoning Province Hu wood, Mile Swertia Herb, Glossy Privet Fruit, Loquat Leaf, hymsleya amabilis.Modern pharmacy studies show that, ursolic acid and Oleanolic Acid all have anti-inflammatory, enhancing immunity, inhibition platelet aggregation, hypoglycemic, clinical pharmacology effect such as anticancer, be the ideal medicament of treatment acute icterohepatitis type hepatitis and chronic viral hepatitis, and the low side effect of toxicity is few.
In the loquat producing region, Loquat Leaf, flower, examine these parts that have than high added value and be wasted basically, do not obtain due application.Usually do not influencing under the prerequisite that the Fractus Eriobotryae produces can gather in year 50000 tons of Loquat Leaf dry products of the whole nation.Therefore, Loquat Leaf is carried out the deep development utilization, help promoting the sustainable sound development of loquat industry,, improve the economic worth of loquat, develop new product, have crucial social benefit and economic worth comprehensive utilization loquat resource.
Summary of the invention
The object of the present invention is to provide the method for separating and purifying ursolic acid from Loquat Leaf.This preparation method adopts hydrophilic organic solvent extraction, removal of impurities, activated carbon decolorizing, precipitate and separate, macroporous adsorbent resin enrichment, purifying, a series of separation and purification integrated technologies such as hydrophilic organic solvent extraction, crystallization and recrystallization processing, preparation high-purity ursolic acid crystal from Loquat Leaf, not only preparation technology is simple, separation and purification efficient height, suitability for industrialized production easily, and ursolic acid purity height, the production cost of preparation are low.
Of the present invention from Loquat Leaf the method steps of separating and purifying ursolic acid be:
(1) solvent extraction of Folium Eriobotryae total triterpenic acids: with the Loquat Leaf is raw material, adopt the water content of 5～20 times of Loquat Leaf dry weights (W/W) to be no more than the hydrophilic organic solvent aqueous solution of 30% (W/W), under 50～80 ℃ of conditions, extract 1～3 time, each 1～3 hour, united extraction liquid, filter, filtrate-0.05～-0.09MPa, 50～70 ℃ of conditions under, become the Folium Eriobotryae total triterpenic acids extract through decompression and solvent recovery, vacuum concentration;
(2) removal of impurities of Folium Eriobotryae total triterpenic acids is handled: under 50～80 ℃ of conditions, the Folium Eriobotryae total triterpenic acids extract is no more than the hydrophilic organic solvent of 20% (W/W) with the water content that is not less than 3 times of amounts (W/W), the aqueous solution fully extracts Folium Eriobotryae total triterpenic acids, insolubles is removed in still aging, filtration, and filtrate is carried out activated carbon decolorizing while hot;
(3) precipitate and separate of Folium Eriobotryae total triterpenic acids: the filtrate after the decolouring adds triterpenic acid solubleness conditioning agent while hot, the hydrophilic organic solvent final concentration is 20～40% and pH value 1.5～5.5 in the hierarchy of control, slowly cool to and begin to separate out Folium Eriobotryae total triterpenic acids precipitation, 4～10 ℃ of refrigeration, place ageing, filtration, must the Folium Eriobotryae total triterpenic acids crude product;
(4) resin concentration of ursolic acid: the Folium Eriobotryae total triterpenic acids crude product is abundant with the hydrophilic organic solvent aqueous solution dissolving of 20～40% (W/W) that are not less than 3 times of amounts (W/W), filter and remove insolubles, the ursolic acid final concentration reaches 0.5～5g/L in the adjustment filtrate system; Filtrate is extremely saturated through the absorption with macroporous adsorbent resin ursolic acid, use the washing that is no less than 3 times of amount of resin (W/W) to remove debris and the more weak water-soluble impurity of adsorptive power that is not adsorbed earlier, with 20～40% the hydrophilic organic solvent aqueous solution flush away middle polarity impurity that is no less than 3 times of amount of resin (W/W), be no more than the hydrophilic organic solvent aqueous solution wash-out ursolic acid of 20% (W/W) at last with the water content that is not less than 3 times of amounts (W/W) again;
(5) precipitate and separate of ursolic acid: the ursolic acid elutriant-0.05～-0.09MPa, 50～70 ℃ of conditions under, behind the reclaim under reduced pressure partial solvent, add ursolic acid solubleness conditioning agent, the hydrophilic organic solvent final concentration is 20～40% and pH value 1.5～5.5 in the control elutriant system, slowly cool to and begin to separate out ursolic acid crystallization, 4～10 ℃ of refrigeration, leave standstill growing the grain, filtration, the ursolic acid crude product;
(6) crystallization purifying of ursolic acid: under 50～80 ℃ of conditions, the ursolic acid crude product fully extracts ursolic acid with the hydrophilic organic solvent that is not less than 3 times of amounts (W/W), filter, filtrate slowly cooling to begins to separate out ursolic acid, 4～10 ℃ of refrigeration, leaves standstill growing the grain, filtration, room temperature condition washing down, the ursolic acid crystallization.
Remarkable advantage of the present invention is:
One: the present invention adopts one or more mixed solvents in methyl alcohol, ethanol, acetone, propyl alcohol or the Virahol to extract ursolic acid as hydrophilic organic solvent from Loquat Leaf, advantage 1: the solubility property of hydrophilic organic solvents such as methyl alcohol, ethanol is good, penetrativity to the Loquat Leaf cell is strong, easier being penetrated in the Loquat Leaf internal organizational structure improved the dissolution rate and the extraction efficiency of ursolic acid in the Loquat Leaf greatly; Advantage 2: hydrophilic organic solvents such as the methyl alcohol of heat, ethanol are very big to the solubleness of ursolic acid in the Loquat Leaf, and less to the solubleness of impurity such as protein, polysaccharide, essential oil, inorganic salt, are more conducive to follow-up further separation and purification and obtain high-purity ursolic acid.
Two: the present invention preferentially selects for use methyl alcohol or acetone as the advantage of hydrophilic solvent to be: though the separating effect of hydrophilic organic solvents such as methyl alcohol, ethanol, acetone, propyl alcohol or Virahol is suitable, but for other hydrophilic organic solvent, the boiling point of methyl alcohol or acetone is lower, energy consumption is littler during solvent recuperation, the rate of recovery is higher, can significantly reduce the preparation cost of product.
Three: the present invention makes full use of macroporous adsorbent resin to different with to the adsorptive power of impurity such as protein, polysaccharide, inorganic salt of the adsorptive power of ursolic acid, and in the hydrophilic organic solvent aqueous solution, be the dissolubility difference of impurity such as the Folium Eriobotryae total triterpenic acids of main component and protein, inorganic salt with the ursolic acid, really reach the high efficiency separation of ursolic acid and impurity.
Four: compare with existing document, the present invention makes full use of ursolic acid and dissolves in methyl alcohol and ethanol isopolarity organic solvent, is slightly soluble in fat-soluble solvents such as chloroform and ether, is insoluble to the characteristics of sherwood oil and water, do not adopt the silica gel column chromatography technology, and directly adopt modes such as hydrophilic organic solvent extraction, crystallization and recrystallization processing to remove impurity, obtain high-purity ursolic acid, have bigger generalization.
Five: the present invention makes up and adopts hydrophilic organic solvent extraction, removal of impurities, activated carbon decolorizing, precipitate and separate, macroporous adsorbent resin enrichment, purifying, a series of separation and purification integrated technologies such as hydrophilic organic solvent extraction, crystallization and recrystallization processing, preparation high-purity ursolic acid crystal from Loquat Leaf has that theoretical novelty, reasonable in technology, operational safety, technology are easy, an economically feasible, advantages of environment protection.
Embodiment
Implement separating and purifying ursolic acid from Loquat Leaf according to the described method steps of content of the present invention.
Wherein, the ursolic acid crystallization in the step (6) can also be used the hydrophilic organic solvent recrystallization further at ambient temperature, washes brilliant removal of impurities with chloroform, water and sherwood oil respectively, gets the high-purity ursolic acid crystal.
Used hydrophilic organic solvent can be one or more mixed solvents in methyl alcohol, ethanol, acetone, propyl alcohol or the Virahol, preferentially selects methyl alcohol or acetone for use.
The chemical constitution of the skeleton of used macroporous adsorbent resin can be one or both in polystyrene and the polyacrylic acid.
Used triterpenic acid solubleness conditioning agent is one or more in inorganic acid aqueous solution, aqueous solutions of organic acids or the acid brine solution, preferentially selects aqueous hydrochloric acid or aqueous sulfuric acid for use; Ursolic acid solubleness conditioning agent is one or more in inorganic acid aqueous solution, aqueous solutions of organic acids or the acid brine solution, preferentially selects aqueous hydrochloric acid or aqueous sulfuric acid for use.
Above-mentioned Folium Eriobotryae total triterpenic acids is one or more compounds in ursolic acid compounds or the Oleanolic Acid compounds.
After measured, purity 〉=25% of ursolic acid in the Folium Eriobotryae total triterpenic acids crude product; The purity of ursolic acid crude product 〉=80%; Ursolic acid crystalline purity 〉=95%; High-purity ursolic acid crystalline purity 〉=99% that final separation and purification obtains.
Physical and chemical parameter measuring method of the present invention is as follows:
The content of Folium Eriobotryae total triterpenic acids, ursolic acid and Oleanolic Acid adopts high performance liquid chromatograph to measure.Condition determination: Agilent1100 type high performance liquid chromatograph (DAD diode-array detector), Waters Nova-Pak C
18Chromatographic column (Φ 3.9 * 150mm, 5 μ m), moving phase be methyl alcohol-0.2% phosphate aqueous solution (92: 8) (V/V), flow velocity 1.0mL/min, 34 ℃ of column temperatures, sample size 20 μ L detect wavelength and are respectively 220nm.Ursolic acid and Oleanolic Acid standard substance are purchased in Nat'l Pharmaceutical ﹠ Biological Products Control Institute.
After measured, test that the Loquat Leaf ursolic acid content is 7.8～11.3g/kg in the Loquat Leaf (water content 4.2%) that used Fujian Province Putian City produces.
Preparation method's of the present invention embodiment is presented below:
Embodiment 1
Producing Loquat Leaf with the Putian, Fujian is raw material, with the washing of bright Loquat Leaf, dry (air-dry, all can dry or dry), pulverize, cross 20～30 mesh sieves and get Loquat Leaf, the 2.6kg Loquat Leaf is put into extractor, methyl alcohol-acetone (1: 1) aqueous solution of 85% with 12 times of Loquat Leaf dry weights (W/W), refluxing extraction is 2 hours under 72 ℃ of conditions, filter, filtrate-0.05～-0.09MPa, 50～70 ℃ of conditions under, become the Folium Eriobotryae total triterpenic acids extract through decompression and solvent recovery, vacuum concentration; Under 72 ℃ of conditions, the Folium Eriobotryae total triterpenic acids extract is that 90% methyl alcohol-acetone (1: 1) aqueous solution (W/W) of 10.9 fully extracts Folium Eriobotryae total triterpenic acids with the pH value of 3 times of amounts (W/W), insolubles is removed in still aging, filtration, and filtrate is added 1.3% (W/V) gac while hot and decoloured; Filtrate after the decolouring adds 35% aqueous hydrochloric acid while hot, methyl alcohol-acetone in the hierarchy of control (1: 1) aqueous solution final concentration reaches 40% (W/W) and the pH value is 1.5, slowly cool to and begin to separate out Folium Eriobotryae total triterpenic acids precipitation, 4～10 ℃ of refrigeration, place ageing, filtration, must the Folium Eriobotryae total triterpenic acids crude product.After measured, the purity 25.8% of ursolic acid in the Folium Eriobotryae total triterpenic acids crude product.
The Folium Eriobotryae total triterpenic acids crude product with the pH value of 3 times of amounts (W/W) be 12.0 20% methanol aqueous solution (W/W) dissolving fully, filter and remove insolubles, adjust that the ursolic acid final concentration reaches 0.5g/L in the filtrate system; Filtrate is adopted the HZ816 macroporous adsorbent resin, with 0.96 times of amount of resin/hour last column flow rate (V/V), the dynamic adsorption ursolic acid is to saturated, debris and the more weak water-soluble impurity of adsorptive power that elder generation is not adsorbed with distilled water (V/V) flush away that is no less than 3 times of amount of resin, use 30% aqueous ethanolic solution (V/V) of 8 times of amount of resin to be divided into again 2 times, flush away middle polarity impurity under the mixing speed condition of 76r/min, use the anhydrous methanol of 3 times of amount of resin at last, with 0.56 times of amount of resin/hour flow velocity (V/V) wash-out ursolic acid, elutriant-0.05～-0.09MPa, under 50～70 ℃ of conditions, behind the reclaim under reduced pressure partial solvent, add 30% aqueous sulfuric acid, the methyl alcohol final concentration reaches 20% (W/W) and the pH value is 1.5 in the control elutriant system, slowly cool to and begin to separate out the ursolic acid crystallization, 4～10 ℃ of refrigerations, leave standstill growing the grain, filter, get beige ursolic acid crude product.After measured, the purity 80.2% of ursolic acid crude product.
Under 50 ℃ of conditions, beige ursolic acid crude product fully extracts ursolic acid with methyl alcohol-acetone (1: 1) solvent of 6 times of amounts (W/W), filter, filtrate slowly cooling to begins to separate out ursolic acid, 4～10 ℃ of refrigeration, leaves standstill under growing the grain, filtration, 12 ℃ of conditions and wash, must white ursolic acid crystallization.After measured, ursolic acid crystalline purity 95.6%.The ursolic acid crystallization can also be used recrystallizing methanol further at ambient temperature, washes brilliant removal of impurities with chloroform, water and sherwood oil respectively, gets the high-purity ursolic acid crystal.After measured, high-purity ursolic acid crystal purity 99.2%, the ursolic acid rate of recovery 50.2% in the Loquat Leaf.
Embodiment 2
The Loquat Leaf 2.8kg of 20～30 orders process petroleum ether degreasing is put into extractor, it with the pH value of 20 times of Loquat Leaf dry weights (W/W) 3.2 70% methanol aqueous solution, under 50 ℃ of conditions, extract 2 times, each 3 hours, united extraction liquid, filter, filtrate-0.05～-0.09MPa, 50～70 ℃ of conditions under, become the Folium Eriobotryae total triterpenic acids extract through decompression and solvent recovery, vacuum concentration; Under 50 ℃ of conditions, the Folium Eriobotryae total triterpenic acids extract is that 8.0 92% methanol aqueous solution (W/W) fully extracts Folium Eriobotryae total triterpenic acids with 15 times of amounts (W/W) pH value, insolubles is removed in still aging, filtration, and filtrate is added 0.5% (W/V) gac while hot and decoloured; Filtrate after the decolouring adds 50% aqueous sulfuric acid while hot, the methyl alcohol final concentration reaches 20% (W/W) and the pH value is 4.5 in the hierarchy of control, slowly cool to and begin to separate out Folium Eriobotryae total triterpenic acids precipitation, 4～10 ℃ of refrigeration, place ageing, filtration, must the Folium Eriobotryae total triterpenic acids crude product.After measured, the purity 27.9% of ursolic acid in the Folium Eriobotryae total triterpenic acids crude product.
The Folium Eriobotryae total triterpenic acids crude product with the pH value of 8 times of amounts (W/W) be 8.0 40% methanol aqueous solution (W/W) dissolving fully, filter and remove insolubles, adjust that the ursolic acid final concentration reaches 5g/L in the filtrate system; Filtrate is under the mixing speed condition of 68r/min, directly adopt the HZ801 macroporous adsorbent resin that medicinal extract is carried out enrichment absorption ursolic acid to saturated, debris and the more weak water-soluble impurity of adsorptive power that elder generation is not adsorbed with distilled water (V/V) flush away that is no less than 3 times of amount of resin, 35% the methanol aqueous solution (V/V) of using 6 times of amount of resin again with 1.0 times of amount of resin/hour flow velocity (V/V) flush away middle polarity impurity, use 80% methanol aqueous solution (V/V) of 6 times of amount of resin to be divided at last 2 times, under the mixing speed condition of 100r/min, the wash-out ursolic acid; Elutriant-0.05～-0.09MPa, 50～70 ℃ of conditions under, behind the reclaim under reduced pressure partial solvent, add 40% aqueous hydrochloric acid, the methyl alcohol final concentration reaches 40% (W/W) and the pH value is 4.5 in the control elutriant system, slowly cool to and begin to separate out ursolic acid crystallization, 4～10 ℃ of refrigeration, leave standstill growing the grain, filtration, beige ursolic acid crude product.After measured, the purity 82.5% of ursolic acid crude product.
Under 80 ℃ of conditions, beige ursolic acid crude product fully extracts ursolic acid with the methyl alcohol of 3 times of amounts (W/W), filters, and filtrate slowly cooling to begins to separate out ursolic acid, 4～10 ℃ of refrigeration, leaves standstill under growing the grain, filtration, 31 ℃ of conditions and wash, must white ursolic acid crystallization; After measured, ursolic acid crystalline purity 95.8%.The ursolic acid crystallization can also be used recrystallizing methanol further at ambient temperature, washes brilliant removal of impurities with chloroform, water and sherwood oil respectively, gets the high-purity ursolic acid crystal.After measured, high-purity ursolic acid crystal purity 99.1%, the ursolic acid rate of recovery 52.3% in the Loquat Leaf.
Embodiment 3
The Loquat Leaf 2.9kg that 20～30 orders are handled through the carbon dioxide upercritical fluid extraction de-oiling puts into extractor, anhydrous methanol with 5 times of Loquat Leaf dry weights (W/W), refluxing extraction is 2 hours under 80 ℃ of conditions, filter, filtrate-0.05～-0.09MPa, 50～70 ℃ of conditions under, become the Folium Eriobotryae total triterpenic acids extract through decompression and solvent recovery, vacuum concentration; Under 80 ℃ of conditions, the Folium Eriobotryae total triterpenic acids extract is that 8.9 80% methanol aqueous solution (W/W) fully extracts Folium Eriobotryae total triterpenic acids with the pH value of 10 times of amounts (W/W), insolubles is removed in still aging, filtration, and filtrate is added 2.5% (W/V) gac while hot and decoloured; Filtrate after the decolouring adds saturated aqueous citric acid solution while hot, the methyl alcohol final concentration reaches 25% (W/W) and the pH value is 5.5 in the hierarchy of control, slowly cool to and begin to separate out Folium Eriobotryae total triterpenic acids precipitation, 4～10 ℃ of refrigeration, place ageing, filtration, must the Folium Eriobotryae total triterpenic acids crude product.After measured, the purity 29.6% of ursolic acid in the Folium Eriobotryae total triterpenic acids crude product.
The Folium Eriobotryae total triterpenic acids crude product with the pH value of 10 times of amounts (W/W) be 20% methyl alcohol-acetone (1: 1) aqueous solution (W/W) dissolving of 9.3 fully, filter and remove insolubles, adjust that the ursolic acid final concentration reaches 0.5g/L in the filtrate system; Filtrate is adopted the HZ802 macroporous adsorbent resin, with 0.59 times of amount of resin/hour last column flow rate (V/V), the dynamic adsorption ursolic acid is to saturated, debris and the more weak water-soluble impurity of adsorptive power that elder generation is not adsorbed with distilled water (V/V) flush away that is no less than 3 times of amount of resin, methyl alcohol-acetone (1: 1) aqueous solution (V/V) of 40% of using 3 times of amount of resin again with 0.5 times of amount of resin/hour flow velocity (V/V) flush away middle polarity impurity, 90% methyl alcohol-acetone (1: 1) aqueous solution (V/V) of using 9 times of amount of resin at last with 1.05 times of amount of resin/hour flow velocity (V/V) wash-out ursolic acid; Elutriant-0.05～-0.09MPa, 50～70 ℃ of conditions under, behind the reclaim under reduced pressure partial solvent, add saturated aqueous citric acid solution, methyl alcohol-acetone (1: 1) final concentration reaches 20% (W/W) and the pH value is 5.5 in the control elutriant system, slowly cool to and begin to separate out ursolic acid crystallization, 4～10 ℃ of refrigeration, leave standstill growing the grain, filtration, beige ursolic acid crude product.After measured, the purity 84.2% of ursolic acid crude product.
Under 72 ℃ of conditions, beige ursolic acid crude product fully extracts ursolic acid with the methyl alcohol of 8 times of amounts (W/W), filters, and filtrate slowly cooling to begins to separate out ursolic acid, 4～10 ℃ of refrigeration, leaves standstill under growing the grain, filtration, 25 ℃ of conditions and wash, must white ursolic acid crystallization; After measured, ursolic acid crystalline purity 96.3%.The ursolic acid crystallization can also be used recrystallizing methanol further at ambient temperature, washes brilliant removal of impurities with chloroform, water and sherwood oil respectively, gets the high-purity ursolic acid crystal.After measured, high-purity ursolic acid crystal purity 99.0%, the ursolic acid rate of recovery 52.6% in the Loquat Leaf.
Embodiment 4
The Loquat Leaf 2.9kg that solvent degreasing was handled through 6# puts into extractor with 20～30 orders, it with the pH value of 5 times of Loquat Leaf dry weights (W/W) 6.1 95% aqueous ethanolic solution, under 80 ℃ of conditions, extract 3 times, each 1 hour, united extraction liquid, filter, filtrate-0.05～-0.09MPa, 50～70 ℃ of conditions under, become the Folium Eriobotryae total triterpenic acids extract through decompression and solvent recovery, vacuum concentration; Under 80 ℃ of conditions, the Folium Eriobotryae total triterpenic acids extract is that 10.5 95% aqueous ethanolic solution (W/W) fully extracts Folium Eriobotryae total triterpenic acids with the pH value of 7 times of amounts (W/W), insolubles is removed in still aging, filtration, and filtrate is added 2.5% (W/V) gac while hot and decoloured; Filtrate after the decolouring adds 20% acid phosphatase salt buffer solution while hot, the ethanol final concentration reaches 20% (W/W) and the pH value is 3.8 in the hierarchy of control, slowly cool to and begin to separate out Folium Eriobotryae total triterpenic acids precipitation, 4～10 ℃ of refrigeration, place ageing, filtration, must the Folium Eriobotryae total triterpenic acids crude product.
The Folium Eriobotryae total triterpenic acids crude product with the pH value of 15 times of amounts (W/W) be 10.0 20% aqueous ethanolic solution (W/W) dissolving fully, filter and remove insolubles, adjust that the ursolic acid final concentration reaches 4.3g/L in the filtrate system; Filtrate is adopted the D101 macroporous adsorbent resin, under the mixing speed condition of 75r/min, the absorption ursolic acid is to saturated, earlier with distilled water (V/V) flush away that is no less than 3 times of amount of resin debris and the more weak water-soluble impurity of adsorptive power not to be adsorbed, 40% the aqueous ethanolic solution (V/V) of using 7 times of amount of resin again with 1.03 times of amount of resin/hour flow velocity (V/V) flush away middle polarity impurity, use 80% aqueous ethanolic solution (V/V) of 9 times of amount of resin to be divided at last 3 times, under the mixing speed condition of 89r/min, the wash-out ursolic acid, elutriant-0.05～-0.09MPa, under 50～70 ℃ of conditions, behind the reclaim under reduced pressure partial solvent, add 20% acid phosphatase salt buffer solution, the ethanol final concentration reaches 20% (W/W) and the pH value is 3.8 in the control elutriant system, slowly cool to and begin to separate out the ursolic acid crystallization, 4～10 ℃ of refrigerations, leave standstill growing the grain, filter, get beige ursolic acid crude product.
Under 65 ℃ of conditions, beige ursolic acid crude product fully extracts ursolic acid with the ethanol of 4 times of amounts (W/W), filters, and filtrate slowly cooling to begins to separate out ursolic acid, 4～10 ℃ of refrigeration, leaves standstill under growing the grain, filtration, 20 ℃ of conditions and wash, must white ursolic acid crystallization; The ursolic acid crystallization can also be used ethyl alcohol recrystallization further at ambient temperature, washes brilliant removal of impurities with chloroform, water and sherwood oil respectively, gets the high-purity ursolic acid crystal.After measured, high-purity ursolic acid crystal purity 99.3%, the ursolic acid rate of recovery 50.1% in the Loquat Leaf.
Embodiment 5
20～30 order Loquat Leaf 2.7kg are put into extractor, 78% ethanol-Virahol (2: 1) aqueous solution with 18 times of Loquat Leaf dry weights (W/W), under 76 ℃ of conditions, extract 2 times, each 2 hours, united extraction liquid, filter, filtrate-0.05～-0.09MPa, 50～70 ℃ of conditions under, become the Folium Eriobotryae total triterpenic acids extract through decompression and solvent recovery, vacuum concentration; Under 76 ℃ of conditions, the Folium Eriobotryae total triterpenic acids extract is that 80% ethanol-Virahol (2: 1) aqueous solution (W/W) of 11.8 fully extracts Folium Eriobotryae total triterpenic acids with the pH value of 9 times of amounts (W/W), insolubles is removed in still aging, filtration, and filtrate is added 0.9% (W/V) gac while hot and decoloured; Filtrate after the decolouring adds 28% aqueous hydrochloric acid while hot, ethanol-Virahol in the hierarchy of control (2: 1) final concentration reaches 32% (W/W) and the pH value is 3.3, slowly cool to and begin to separate out Folium Eriobotryae total triterpenic acids precipitation, 4～10 ℃ of refrigeration, place ageing, filtration, must the Folium Eriobotryae total triterpenic acids crude product.
The Folium Eriobotryae total triterpenic acids crude product is that 28% ethanol-Virahol (2: 1) aqueous solution molten (W/W) of 10.8 is separated fully with the pH value of 6 times of amounts (W/W), filters and removes insolubles, adjusts that the ursolic acid final concentration reaches 1.8g/L in the filtrate system; Filtrate is adopted the DK110 macroporous adsorbent resin, with 1.5 times of amount of resin/hour flow velocity (V/V), the dynamic adsorption ursolic acid is to saturated, debris and the more weak water-soluble impurity of adsorptive power that elder generation is not adsorbed with distilled water (V/V) flush away that is no less than 3 times of amount of resin, use 20% aqueous ethanolic solution (V/V) of 12 times of amount of resin to be divided into again 4 times, flush away middle polarity impurity under the mixing speed condition of 68r/min, use 84% ethanol-Virahol (1: 1) aqueous solution (V/V) of 15 times of amount of resin to be divided at last 3 times, under the mixing speed condition of 90r/min, the wash-out ursolic acid; Elutriant-0.05～-0.09MPa, 50～70 ℃ of conditions under, behind the reclaim under reduced pressure partial solvent, add 28% aqueous hydrochloric acid, ethanol-Virahol (1: 1) final concentration reaches 28% (W/W) and the pH value is 3.3 in the control elutriant system, slowly cool to and begin to separate out ursolic acid crystallization, 4～10 ℃ of refrigeration, leave standstill growing the grain, filtration, beige ursolic acid crude product.
Under 78 ℃ of conditions, beige ursolic acid crude product fully extracts ursolic acid with the ethanol-acetone (1: 2) of 9 times of amounts (W/W), filter, filtrate slowly cooling to begins to separate out ursolic acid, 4～10 ℃ of refrigeration, leaves standstill under growing the grain, filtration, 28 ℃ of conditions and wash, must white ursolic acid crystallization; The ursolic acid crystallization can also with ethanol-acetone (1: 1) recrystallization, be washed brilliant removal of impurities with chloroform, water and sherwood oil respectively further at ambient temperature, gets the high-purity ursolic acid crystal.After measured, high-purity ursolic acid crystal purity 99.4%, the ursolic acid rate of recovery 52.0% in the Loquat Leaf.
Embodiment 6
20～30 order Loquat Leaf 2.7kg are put into extractor, it with the pH value of 20 times of Loquat Leaf dry weights (W/W) 11.2 70% aqueous ethanolic solution, under 50 ℃ of conditions, extract 3 times, each 1 hour, united extraction liquid, filter, filtrate-0.05～-0.09MPa, 50～70 ℃ of conditions under, become the Folium Eriobotryae total triterpenic acids extract through decompression and solvent recovery, vacuum concentration; Under 50 ℃ of conditions, the Folium Eriobotryae total triterpenic acids extract fully extracts Folium Eriobotryae total triterpenic acids with the dehydrated alcohol of 3 times of amounts (W/W), and insolubles is removed in still aging, filtration, and filtrate is added 0.5% (W/V) gac while hot and decoloured; Filtrate after the decolouring adds 32% aqueous hydrochloric acid while hot, the ethanol final concentration reaches 40% (W/W) and the pH value is 2.6 in the hierarchy of control, slowly cool to and begin to separate out Folium Eriobotryae total triterpenic acids precipitation, 4～10 ℃ of refrigeration, place ageing, filtration, must the Folium Eriobotryae total triterpenic acids crude product.
The Folium Eriobotryae total triterpenic acids crude product with the pH value of 3 times of amounts (W/W) be 11.5 40% aqueous ethanolic solution (W/W) dissolving fully, filter and remove insolubles, adjust that the ursolic acid final concentration reaches 5.0g/L in the filtrate system; Filtrate is under the mixing speed condition of 50r/min, directly adopt macroporous adsorbent resin HZ801: HZ802 (2: 1) that medicinal extract is carried out enrichment absorption ursolic acid to saturated, the absorption ursolic acid is to saturated, debris and the more weak water-soluble impurity of adsorptive power that elder generation is not adsorbed with distilled water (V/V) flush away that is no less than 3 times of amount of resin, 28% the aqueous ethanolic solution (V/V) of using 5 times of amount of resin again with 0.65 times of amount of resin/hour flow velocity (V/V) flush away middle polarity impurity, the dehydrated alcohol of using 3 times of amount of resin at last with 1.0 times of amount of resin/hour flow velocity (V/V) wash-out ursolic acid; Elutriant-0.05～-0.09MPa, 50～70 ℃ of conditions under, behind the reclaim under reduced pressure partial solvent, add 32% aqueous hydrochloric acid, the ethanol final concentration reaches 40% (W/W) and the pH value is 2.6 in the control elutriant system, slowly cool to and begin to separate out ursolic acid crystallization, 4～10 ℃ of refrigeration, leave standstill growing the grain, filtration, beige ursolic acid crude product.
Under 50 ℃ of conditions, beige ursolic acid crude product fully extracts ursolic acid with the ethanol of 15 times of amounts (W/W), filters, and filtrate slowly cooling to begins to separate out ursolic acid, 4～10 ℃ of refrigeration, leaves standstill under growing the grain, filtration, 14 ℃ of conditions and wash, must white ursolic acid crystallization; The ursolic acid crystallization can also be used ethyl alcohol recrystallization further at ambient temperature, washes brilliant removal of impurities with chloroform, water and sherwood oil respectively, gets the high-purity ursolic acid crystal.After measured, high-purity ursolic acid crystal purity 99.3%, the ursolic acid rate of recovery 51.8% in the Loquat Leaf.
Embodiment 7
Producing Loquat Leaf with the Putian, Fujian is raw material, 10～16 mesh sieves are washed, pulverize, crossed to bright Loquat Leaf get the Loquat Leaf slurry, 2.6kg (with dry weight basis) Loquat Leaf slurry is put into extractor, with the pH value of 20 times of Loquat Leaf dry weights (W/W) is 6.5 70% aqueous acetone solution, extracts 3 times each 1 hour under 50 ℃ of conditions, united extraction liquid, filter, filtrate-0.05～-0.09MPa, 50～70 ℃ of conditions under, become the Folium Eriobotryae total triterpenic acids extract through decompression and solvent recovery, vacuum concentration; Under 50 ℃ of conditions, the Folium Eriobotryae total triterpenic acids extract is that 9.3 85% aqueous acetone solution (W/W) fully extracts Folium Eriobotryae total triterpenic acids with the pH value of 15 times of amounts (W/W), insolubles is removed in still aging, filtration, and filtrate is added 2.5% (W/V) gac while hot and decoloured; Filtrate after the decolouring adds 40% aqueous sulfuric acid while hot, the acetone final concentration reaches 25% (W/W) and the pH value is 4.2 in the hierarchy of control, slowly cool to and begin to separate out Folium Eriobotryae total triterpenic acids precipitation, 4～10 ℃ of refrigeration, place ageing, filtration, must the Folium Eriobotryae total triterpenic acids crude product.
The Folium Eriobotryae total triterpenic acids crude product with the pH value of 12 times of amounts (W/W) be 9.0 35% aqueous acetone solution (W/W) dissolving fully, filter and remove insolubles, adjust that the ursolic acid final concentration reaches 3.2g/L in the filtrate system; Filtrate is adopted Amberlite XAD-4 macroporous adsorbent resin, with 0.98 times of amount of resin/hour flow velocity (V/V), the dynamic adsorption ursolic acid is to saturated, debris and the more weak water-soluble impurity of adsorptive power that elder generation is not adsorbed with distilled water (V/V) flush away that is no less than 3 times of amount of resin, use 20% aqueous acetone solution (V/V) of 12 times of amount of resin to be divided into again 3 times, flush away middle polarity impurity under the mixing speed condition of 82r/min, use 85% aqueous acetone solution (V/V) of 6 times of amount of resin to be divided at last 3 times, under the mixing speed condition of 85r/min, the wash-out ursolic acid; Elutriant-0.05～-0.09MPa, 50～70 ℃ of conditions under, behind the reclaim under reduced pressure partial solvent, add 40% aqueous sulfuric acid, the acetone final concentration reaches 35% (W/W) and the pH value is 4.2 in the control elutriant system, slowly cool to and begin to separate out ursolic acid crystallization, 4～10 ℃ of refrigeration, leave standstill growing the grain, filtration, beige ursolic acid crude product.
Under 70 ℃ of conditions, beige ursolic acid crude product fully extracts ursolic acid with the acetone of 5 times of amounts (W/W), filters, and filtrate slowly cooling to begins to separate out ursolic acid, 4～10 ℃ of refrigeration, leaves standstill under growing the grain, filtration, 20 ℃ of conditions and wash, must white ursolic acid crystallization; The ursolic acid crystallization can also be used acetone recrystallization further at ambient temperature, washes brilliant removal of impurities with chloroform, water and sherwood oil respectively, gets the high-purity ursolic acid crystal.After measured, ursolic acid white crystal purity 99.5%, the ursolic acid rate of recovery 53.6% in the Loquat Leaf.
Embodiment 8
20～30 order Loquat Leaf 2.8kg are put into extractor, anhydrous propanone with 5 times of Loquat Leaf dried bean noodles heavy (W/W), refluxing extraction is 2 hours under 80 ℃ of conditions, filter, filtrate-0.05～-0.09MPa, 50～70 ℃ of conditions under, become the Folium Eriobotryae total triterpenic acids extract through decompression and solvent recovery, vacuum concentration; Under 80 ℃ of conditions, the Folium Eriobotryae total triterpenic acids extract fully extracts Folium Eriobotryae total triterpenic acids with the anhydrous propanone of 6 times of amounts (W/W), and insolubles is removed in still aging, filtration, and filtrate is added 1.1% (W/V) gac while hot and decoloured; Filtrate after the decolouring adds 50% aqueous sulfuric acid while hot, close acetone final concentration reaches 38% (W/W) and the pH value is 2.5 in the hierarchy of control, slowly cool to and begin to separate out Folium Eriobotryae total triterpenic acids precipitation, 4～10 ℃ of refrigeration, place ageing, filtration, must the Folium Eriobotryae total triterpenic acids crude product.
The Folium Eriobotryae total triterpenic acids crude product with the pH value of 7 times of amounts (W/W) be 9.8 40% aqueous acetone solution (W/W) dissolving fully, filter and remove insolubles, adjust that the ursolic acid final concentration reaches 4.9g/L in the filtrate system; Filtrate is under the mixing speed condition of 100r/min, directly adopt the HZ806 macroporous adsorbent resin that medicinal extract is carried out enrichment absorption ursolic acid to saturated, debris and the more weak water-soluble impurity of adsorptive power that elder generation is not adsorbed with distilled water (V/V) flush away that is no less than 3 times of amount of resin, use 40% aqueous acetone solution (V/V) of 6 times of amount of resin to be divided into again 2 times, flush away middle polarity impurity under the mixing speed condition of 75r/min, use 80% aqueous acetone solution (V/V) of 12 times of amount of resin to be divided at last 3 times, under the mixing speed condition of 90r/min, the wash-out ursolic acid; Elutriant-0.05～-0.09MPa, 50～70 ℃ of conditions under, behind the reclaim under reduced pressure partial solvent, add 50% aqueous sulfuric acid, the acetone final concentration reaches 40% (W/W) and the pH value is 2.5 in the control elutriant system, slowly cool to and begin to separate out ursolic acid crystallization, 4～10 ℃ of refrigeration, leave standstill growing the grain, filtration, beige ursolic acid crude product.
Under 80 ℃ of conditions, beige ursolic acid crude product fully extracts ursolic acid with the acetone of 3 times of amounts (W/W), filters, and filtrate slowly cooling to begins to separate out ursolic acid, 4～10 ℃ of refrigeration, leaves standstill under growing the grain, filtration, 26 ℃ of conditions and wash, must white ursolic acid crystallization; The ursolic acid crystallization can also be used acetone recrystallization further at ambient temperature, washes brilliant removal of impurities with chloroform, water and sherwood oil respectively, gets the high-purity ursolic acid crystal.After measured, high-purity ursolic acid crystal purity 99.4%, the ursolic acid rate of recovery 52.9% in the Loquat Leaf.
Embodiment 9
The Loquat Leaf 2.6kg of 20～30 orders skimming treatment is put into extractor, 84% methyl alcohol-ethanol (5: 3) aqueous solution with 15 times of Loquat Leaf dry weights (W/W), refluxing extraction is 2 hours under 80 ℃ of conditions, filter, filtrate-0.05～-0.09MPa, 50～70 ℃ of conditions under, become the Folium Eriobotryae total triterpenic acids extract through decompression and solvent recovery, vacuum concentration; Under 80 ℃ of conditions, the Folium Eriobotryae total triterpenic acids extract is that 82% methyl alcohol-ethanol (5: 3) aqueous solution (W/W) of 10.5 fully extracts Folium Eriobotryae total triterpenic acids with the pH value of 8 times of amounts (W/W), insolubles is removed in still aging, filtration, and filtrate is added 1.8% (W/V) gac while hot and decoloured; Filtrate after the decolouring adds 50% aqueous sulfuric acid while hot, methyl alcohol-ethanol in the hierarchy of control (5: 3) final concentration reaches 40% (W/W) and the pH value is 3.1, slowly cool to and begin to separate out Folium Eriobotryae total triterpenic acids precipitation, 4～10 ℃ of refrigeration, place ageing, filtration, must the Folium Eriobotryae total triterpenic acids crude product.
The Folium Eriobotryae total triterpenic acids crude product with the pH value of 9 times of amounts (W/W) be 32% methyl alcohol-ethanol (2: 5) aqueous solution (W/W) dissolving of 10.5 fully, filter and remove insolubles, adjust that the ursolic acid final concentration reaches 2.1g/L in the filtrate system; Filtrate is adopted the HZ803 macroporous adsorbent resin, with 1.0 times of amount of resin/hour flow velocity (V/V), the dynamic adsorption ursolic acid is to saturated, debris and the more weak water-soluble impurity of adsorptive power that elder generation is not adsorbed with distilled water (V/V) flush away that is no less than 3 times of amount of resin, methyl alcohol-ethanol (2: 5) aqueous solution (V/V) of 26% of using 4 times of amount of resin again with 0.75 times of amount of resin/hour flow velocity (V/V) flush away middle polarity impurity, 95% methyl alcohol-ethanol (2: 5) aqueous solution (V/V) of using 5 times of amount of resin at last with 0.75 times of amount of resin/hour flow velocity (V/V), the dynamic desorption ursolic acid; Elutriant-0.05～-0.09MPa, 50～70 ℃ of conditions under, behind the reclaim under reduced pressure partial solvent, add 50% aqueous sulfuric acid, methyl alcohol-ethanol (2: 5) final concentration reaches 32% (W/W) and the pH value is 3.1 in the control elutriant system, slowly cool to and begin to separate out ursolic acid crystallization, 4～10 ℃ of refrigeration, leave standstill growing the grain, filtration, beige ursolic acid crude product.
Under 50 ℃ of conditions, beige ursolic acid crude product fully extracts ursolic acid with the methyl alcohol-ethanol (5: 3) of 12 times of amounts (W/W), filter, filtrate slowly cooling to begins to separate out ursolic acid, 4～10 ℃ of refrigeration, leaves standstill under growing the grain, filtration, 15 ℃ of conditions and wash, must white ursolic acid crystallization; The ursolic acid crystallization can also with methyl alcohol-ethanol (1: 1) recrystallization, be washed brilliant removal of impurities with chloroform, water and sherwood oil respectively further at ambient temperature, gets the high-purity ursolic acid crystal.After measured, high-purity ursolic acid crystal purity 99.8%, the ursolic acid rate of recovery 53.8% in the Loquat Leaf.
Above embodiment is intended to further describe for example the present invention, rather than limits the present invention by any way.
Of the present invention a kind of from Loquat Leaf the method for separating and purifying high-purity ursolic acid, technology is simple, separation and purification is effective, product yield is high, production cost is low, to the reasonable utilization of China's loquat plant resources and the extension of traditional Chinese medicine industrial chain, solve the agriculture, rural areas and farmers problem, all have important social value and realistic meaning.
Claims (3)
1. the method for a separating and purifying ursolic acid from Loquat Leaf, it is characterized in that: the step of described method is:
(1) solvent extraction of Folium Eriobotryae total triterpenic acids: with the Loquat Leaf is raw material, adopt the water content of 5～20 times of Loquat Leaf dry weights (W/W) to be no more than the hydrophilic organic solvent aqueous solution of 30% (W/W), under 50～80 ℃ of conditions, extract 1～3 time, each 1～3 hour, united extraction liquid, filter, filtrate-0.05～-0.09MPa, 50～70 ℃ of conditions under, become the Folium Eriobotryae total triterpenic acids extract through decompression and solvent recovery, vacuum concentration;
(2) removal of impurities of Folium Eriobotryae total triterpenic acids is handled: under 50～80 ℃ of conditions, the Folium Eriobotryae total triterpenic acids extract fully extracts Folium Eriobotryae total triterpenic acids with the hydrophilic organic solvent aqueous solution that the water content that is not less than 3 times of amounts (W/W) is no more than 20% (W/W), insolubles is removed in still aging, filtration, and filtrate is carried out activated carbon decolorizing while hot;
(3) precipitate and separate of Folium Eriobotryae total triterpenic acids: the filtrate after the decolouring adds triterpenic acid solubleness conditioning agent while hot, the hydrophilic organic solvent final concentration is 20～40% and pH value 1.5～5.5 in the hierarchy of control, slowly cool to and begin to separate out Folium Eriobotryae total triterpenic acids precipitation, 4～10 ℃ of refrigeration, place ageing, filtration, must the Folium Eriobotryae total triterpenic acids crude product;
(4) resin concentration of ursolic acid: the Folium Eriobotryae total triterpenic acids crude product is abundant with the hydrophilic organic solvent aqueous solution dissolving of 20～40% (W/W) that are not less than 3 times of amounts (W/W), filter and remove insolubles, the ursolic acid final concentration reaches 0.5～5g/L in the adjustment filtrate system; Filtrate is extremely saturated through the absorption with macroporous adsorbent resin ursolic acid, use the washing that is no less than 3 times of amount of resin (W/W) to remove debris and the more weak water-soluble impurity of adsorptive power that is not adsorbed earlier, with 20～40% the hydrophilic organic solvent aqueous solution flush away middle polarity impurity that is no less than 3 times of amount of resin (W/W), be no more than the hydrophilic organic solvent aqueous solution wash-out ursolic acid of 20% (W/W) at last with the water content that is not less than 3 times of amounts (W/W) again;
(5) precipitate and separate of ursolic acid: the ursolic acid elutriant-0.05～-0.09MPa, 50～70 ℃ of conditions under, behind the reclaim under reduced pressure partial solvent, add ursolic acid solubleness conditioning agent, the hydrophilic organic solvent final concentration is 20～40% and pH value 1.5～5.5 in the control elutriant system, slowly cool to and begin to separate out ursolic acid crystallization, 4～10 ℃ of refrigeration, leave standstill growing the grain, filtration, the ursolic acid crude product;
(6) crystallization purifying of ursolic acid: under 50～80 ℃ of conditions, the ursolic acid crude product fully extracts ursolic acid with the hydrophilic organic solvent that is not less than 3 times of amounts (W/W), filter, filtrate slowly cooling to begins to separate out ursolic acid, 4～10 ℃ of refrigeration, leaves standstill growing the grain, filtration, room temperature condition washing down, the ursolic acid crystallization;
Described hydrophilic organic solvent is one or more mixed solvents in methyl alcohol, ethanol, propyl alcohol or the Virahol; Described triterpenic acid solubleness conditioning agent is aqueous hydrochloric acid or aqueous sulfuric acid; The chemical constitution of described macroporous adsorbent resin skeleton is one or both in polystyrene or the polyacrylic acid; Described ursolic acid solubleness conditioning agent is aqueous hydrochloric acid or aqueous sulfuric acid.
2. according to claim 1 from Loquat Leaf the method for separating and purifying ursolic acid, it is characterized in that: the ursolic acid crystallization in the described step (6) further at ambient temperature, use the hydrophilic organic solvent recrystallization, wash brilliant removal of impurities with chloroform, water and sherwood oil respectively, get the high-purity ursolic acid crystal; Described hydrophilic organic solvent is one or more mixed solvents in methyl alcohol, ethanol, propyl alcohol or the Virahol.
3. according to claim 2 from Loquat Leaf the method for separating and purifying ursolic acid, it is characterized in that: described high-purity ursolic acid crystalline purity 〉=99%.
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| CN101985459B (en) * | 2010-11-18 | 2012-11-28 | 桂林安和药业有限公司 | Process for extracting greater than or equal to 98% of ursolic acid from loquat leaf |
| CN102276682B (en) * | 2011-09-20 | 2012-11-07 | 江南大学 | Method for extracting ursolic acid from loquat leaves |
| CN102617693B (en) * | 2012-03-07 | 2015-04-22 | 江南大学 | Method utilizing subcritical water extraction technology to extract and prepare ursolic acid from loquat leaves |
| CN102649808A (en) * | 2012-04-24 | 2012-08-29 | 南昌大学 | Method for separating ursolic acid from plantain herb |
| CN104147171A (en) * | 2014-08-28 | 2014-11-19 | 蒋君军 | Extraction method for triterpene acid component from herb residue of loquat leaf |
| CN105131076B8 (en) * | 2015-08-30 | 2016-10-12 | 宁波市江东林清环保科技有限公司 | A kind of method of separating and purifying ursolic acid from Folium Eriobotryae |
| CN112704663B (en) * | 2020-12-18 | 2022-07-19 | 吉林农业科技学院 | Preparation method of loquat leaf ursolic acid cream |
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