CN110508031A - Method for separating ursolic acid and oleanolic acid from plants - Google Patents
Method for separating ursolic acid and oleanolic acid from plants Download PDFInfo
- Publication number
- CN110508031A CN110508031A CN201910693402.7A CN201910693402A CN110508031A CN 110508031 A CN110508031 A CN 110508031A CN 201910693402 A CN201910693402 A CN 201910693402A CN 110508031 A CN110508031 A CN 110508031A
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- China
- Prior art keywords
- acid
- plant
- crude extract
- ursolic acid
- oleanolic acid
- Prior art date
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- WCGUUGGRBIKTOS-GPOJBZKASA-N (3beta)-3-hydroxyurs-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)CC[C@@H](C)[C@H](C)[C@H]5C4=CC[C@@H]3[C@]21C WCGUUGGRBIKTOS-GPOJBZKASA-N 0.000 title claims abstract description 43
- 229940096998 ursolic acid Drugs 0.000 title claims abstract description 43
- PLSAJKYPRJGMHO-UHFFFAOYSA-N ursolic acid Natural products CC1CCC2(CCC3(C)C(C=CC4C5(C)CCC(O)C(C)(C)C5CCC34C)C2C1C)C(=O)O PLSAJKYPRJGMHO-UHFFFAOYSA-N 0.000 title claims abstract description 43
- MIJYXULNPSFWEK-GTOFXWBISA-N 3beta-hydroxyolean-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 title claims abstract description 41
- JKLISIRFYWXLQG-UHFFFAOYSA-N Epioleonolsaeure Natural products C1CC(O)C(C)(C)C2CCC3(C)C4(C)CCC5(C(O)=O)CCC(C)(C)CC5C4CCC3C21C JKLISIRFYWXLQG-UHFFFAOYSA-N 0.000 title claims abstract description 41
- YBRJHZPWOMJYKQ-UHFFFAOYSA-N Oleanolic acid Natural products CC1(C)CC2C3=CCC4C5(C)CCC(O)C(C)(C)C5CCC4(C)C3(C)CCC2(C1)C(=O)O YBRJHZPWOMJYKQ-UHFFFAOYSA-N 0.000 title claims abstract description 41
- MIJYXULNPSFWEK-UHFFFAOYSA-N Oleanolinsaeure Natural products C1CC(O)C(C)(C)C2CCC3(C)C4(C)CCC5(C(O)=O)CCC(C)(C)CC5C4=CCC3C21C MIJYXULNPSFWEK-UHFFFAOYSA-N 0.000 title claims abstract description 41
- 229940100243 oleanolic acid Drugs 0.000 title claims abstract description 41
- HZLWUYJLOIAQFC-UHFFFAOYSA-N prosapogenin PS-A Natural products C12CC(C)(C)CCC2(C(O)=O)CCC(C2(CCC3C4(C)C)C)(C)C1=CCC2C3(C)CCC4OC1OCC(O)C(O)C1O HZLWUYJLOIAQFC-UHFFFAOYSA-N 0.000 title claims abstract description 41
- 238000000034 method Methods 0.000 title claims abstract description 27
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims abstract description 27
- 239000000287 crude extract Substances 0.000 claims abstract description 24
- 230000005526 G1 to G0 transition Effects 0.000 claims abstract description 22
- 239000003480 eluent Substances 0.000 claims abstract description 19
- 239000000523 sample Substances 0.000 claims abstract description 19
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 19
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 claims abstract description 16
- 239000012488 sample solution Substances 0.000 claims abstract description 16
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims abstract description 13
- 238000001035 drying Methods 0.000 claims abstract description 11
- 239000002904 solvent Substances 0.000 claims abstract description 9
- 230000002378 acidificating effect Effects 0.000 claims abstract description 7
- 239000012046 mixed solvent Substances 0.000 claims abstract description 7
- 239000011259 mixed solution Substances 0.000 claims abstract description 6
- 239000013076 target substance Substances 0.000 claims abstract description 5
- 238000001704 evaporation Methods 0.000 claims abstract description 3
- 241000196324 Embryophyta Species 0.000 claims description 26
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 19
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 19
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 18
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 claims description 18
- 239000002023 wood Substances 0.000 claims description 16
- 239000000243 solution Substances 0.000 claims description 14
- 239000000284 extract Substances 0.000 claims description 13
- 238000001914 filtration Methods 0.000 claims description 11
- 239000000843 powder Substances 0.000 claims description 11
- 238000010992 reflux Methods 0.000 claims description 10
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical group [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 claims description 9
- 241000722818 Aralia Species 0.000 claims description 9
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 9
- 239000000908 ammonium hydroxide Substances 0.000 claims description 9
- 239000000706 filtrate Substances 0.000 claims description 9
- 239000013049 sediment Substances 0.000 claims description 9
- 229960000583 acetic acid Drugs 0.000 claims description 8
- 239000007864 aqueous solution Substances 0.000 claims description 8
- 239000000463 material Substances 0.000 claims description 8
- 238000000605 extraction Methods 0.000 claims description 7
- 238000004128 high performance liquid chromatography Methods 0.000 claims description 7
- 230000014759 maintenance of location Effects 0.000 claims description 7
- 238000004809 thin layer chromatography Methods 0.000 claims description 7
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 6
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 claims description 6
- IDGUHHHQCWSQLU-UHFFFAOYSA-N ethanol;hydrate Chemical compound O.CCO IDGUHHHQCWSQLU-UHFFFAOYSA-N 0.000 claims description 6
- 230000007062 hydrolysis Effects 0.000 claims description 6
- 238000006460 hydrolysis reaction Methods 0.000 claims description 6
- USFZMSVCRYTOJT-UHFFFAOYSA-N Ammonium acetate Chemical compound N.CC(O)=O USFZMSVCRYTOJT-UHFFFAOYSA-N 0.000 claims description 5
- 239000005695 Ammonium acetate Substances 0.000 claims description 5
- 229940043376 ammonium acetate Drugs 0.000 claims description 5
- 235000019257 ammonium acetate Nutrition 0.000 claims description 5
- 239000012362 glacial acetic acid Substances 0.000 claims description 5
- 125000004836 hexamethylene group Chemical group [H]C([H])([*:2])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[*:1] 0.000 claims description 5
- 238000010829 isocratic elution Methods 0.000 claims description 5
- 239000000047 product Substances 0.000 claims description 5
- 238000013517 stratification Methods 0.000 claims description 5
- 238000012360 testing method Methods 0.000 claims description 5
- 239000000203 mixture Substances 0.000 claims description 4
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 3
- 229910052799 carbon Inorganic materials 0.000 claims description 3
- 239000012065 filter cake Substances 0.000 claims description 3
- 230000010355 oscillation Effects 0.000 claims description 3
- 239000003208 petroleum Substances 0.000 claims description 3
- 230000003113 alkalizing effect Effects 0.000 claims description 2
- 230000006837 decompression Effects 0.000 claims description 2
- 239000007788 liquid Substances 0.000 claims description 2
- 238000002360 preparation method Methods 0.000 claims description 2
- 238000005070 sampling Methods 0.000 claims description 2
- 238000010792 warming Methods 0.000 claims description 2
- 238000005406 washing Methods 0.000 claims 2
- 229920002472 Starch Polymers 0.000 claims 1
- 238000009835 boiling Methods 0.000 claims 1
- 238000010438 heat treatment Methods 0.000 claims 1
- 238000007689 inspection Methods 0.000 claims 1
- 238000000643 oven drying Methods 0.000 claims 1
- 235000019698 starch Nutrition 0.000 claims 1
- 239000008107 starch Substances 0.000 claims 1
- 238000011084 recovery Methods 0.000 abstract description 9
- 238000000926 separation method Methods 0.000 abstract description 9
- 238000007670 refining Methods 0.000 abstract description 5
- 239000000126 substance Substances 0.000 abstract description 5
- 230000008569 process Effects 0.000 abstract description 4
- 238000004185 countercurrent chromatography Methods 0.000 abstract description 2
- 238000002347 injection Methods 0.000 abstract 1
- 239000007924 injection Substances 0.000 abstract 1
- 239000012071 phase Substances 0.000 description 20
- 241000220225 Malus Species 0.000 description 18
- 235000011430 Malus pumila Nutrition 0.000 description 18
- 235000019441 ethanol Nutrition 0.000 description 10
- 241001092070 Eriobotrya Species 0.000 description 6
- 235000009008 Eriobotrya japonica Nutrition 0.000 description 6
- 230000000694 effects Effects 0.000 description 6
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 5
- 230000002411 adverse Effects 0.000 description 5
- 238000004064 recycling Methods 0.000 description 5
- FEWJPZIEWOKRBE-UHFFFAOYSA-N Tartaric acid Natural products [H+].[H+].[O-]C(=O)C(O)C(O)C([O-])=O FEWJPZIEWOKRBE-UHFFFAOYSA-N 0.000 description 4
- AUALQMFGWLZREY-UHFFFAOYSA-N acetonitrile;methanol Chemical compound OC.CC#N AUALQMFGWLZREY-UHFFFAOYSA-N 0.000 description 4
- 238000001514 detection method Methods 0.000 description 4
- 238000004519 manufacturing process Methods 0.000 description 4
- 235000002906 tartaric acid Nutrition 0.000 description 4
- 239000011975 tartaric acid Substances 0.000 description 4
- 229940122803 Vinca alkaloid Drugs 0.000 description 3
- 239000003795 chemical substances by application Substances 0.000 description 3
- 238000013461 design Methods 0.000 description 3
- 238000010262 high-speed countercurrent chromatography Methods 0.000 description 3
- 239000007791 liquid phase Substances 0.000 description 3
- 238000002156 mixing Methods 0.000 description 3
- 241001656831 Arctous alpina Species 0.000 description 2
- 206010011224 Cough Diseases 0.000 description 2
- 206010013786 Dry skin Diseases 0.000 description 2
- 206010030113 Oedema Diseases 0.000 description 2
- 240000007817 Olea europaea Species 0.000 description 2
- 208000004880 Polyuria Diseases 0.000 description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 238000004440 column chromatography Methods 0.000 description 2
- 239000012141 concentrate Substances 0.000 description 2
- 239000012043 crude product Substances 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 230000035619 diuresis Effects 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 210000004072 lung Anatomy 0.000 description 2
- 108090000765 processed proteins & peptides Proteins 0.000 description 2
- 238000012545 processing Methods 0.000 description 2
- 230000001737 promoting effect Effects 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 239000000741 silica gel Substances 0.000 description 2
- 229910002027 silica gel Inorganic materials 0.000 description 2
- 238000010898 silica gel chromatography Methods 0.000 description 2
- 239000007787 solid Substances 0.000 description 2
- -1 triterpene compound Chemical class 0.000 description 2
- 206010000087 Abdominal pain upper Diseases 0.000 description 1
- 102100036475 Alanine aminotransferase 1 Human genes 0.000 description 1
- 108010082126 Alanine transaminase Proteins 0.000 description 1
- 241000167549 Aralia chinensis Species 0.000 description 1
- 241000208340 Araliaceae Species 0.000 description 1
- 208000004429 Bacillary Dysentery Diseases 0.000 description 1
- 208000034656 Contusions Diseases 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 244000061508 Eriobotrya japonica Species 0.000 description 1
- 208000005577 Gastroenteritis Diseases 0.000 description 1
- 206010017915 Gastroenteritis shigella Diseases 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 206010018473 Glycosuria Diseases 0.000 description 1
- 208000000616 Hemoptysis Diseases 0.000 description 1
- 208000032843 Hemorrhage Diseases 0.000 description 1
- 208000008771 Lymphadenopathy Diseases 0.000 description 1
- 244000141359 Malus pumila Species 0.000 description 1
- 208000002193 Pain Diseases 0.000 description 1
- 206010035664 Pneumonia Diseases 0.000 description 1
- 206010037660 Pyrexia Diseases 0.000 description 1
- 108090000340 Transaminases Proteins 0.000 description 1
- 208000025865 Ulcer Diseases 0.000 description 1
- 206010047700 Vomiting Diseases 0.000 description 1
- 208000012873 acute gastroenteritis Diseases 0.000 description 1
- 206010001093 acute tonsillitis Diseases 0.000 description 1
- 239000003513 alkali Substances 0.000 description 1
- 229930013930 alkaloid Natural products 0.000 description 1
- 150000003797 alkaloid derivatives Chemical class 0.000 description 1
- 230000000202 analgesic effect Effects 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 230000002300 anti-fibrosis Effects 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 0.000 description 1
- 230000003064 anti-oxidating effect Effects 0.000 description 1
- 230000000767 anti-ulcer Effects 0.000 description 1
- 239000003963 antioxidant agent Substances 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 230000036528 appetite Effects 0.000 description 1
- 235000019789 appetite Nutrition 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 239000002585 base Chemical class 0.000 description 1
- 230000000975 bioactive effect Effects 0.000 description 1
- 230000003115 biocidal effect Effects 0.000 description 1
- 229960000074 biopharmaceutical Drugs 0.000 description 1
- 208000034158 bleeding Diseases 0.000 description 1
- 230000000740 bleeding effect Effects 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 206010006451 bronchitis Diseases 0.000 description 1
- 208000034526 bruise Diseases 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 206010012601 diabetes mellitus Diseases 0.000 description 1
- 238000010494 dissociation reaction Methods 0.000 description 1
- 230000005593 dissociations Effects 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 239000000975 dye Substances 0.000 description 1
- 208000022532 enlargement of lymph nodes Diseases 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 239000000469 ethanolic extract Substances 0.000 description 1
- 125000004494 ethyl ester group Chemical group 0.000 description 1
- 230000006870 function Effects 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 229940093915 gynecological organic acid Drugs 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 208000006454 hepatitis Diseases 0.000 description 1
- 231100000283 hepatitis Toxicity 0.000 description 1
- 230000003301 hydrolyzing effect Effects 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 238000011031 large-scale manufacturing process Methods 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 230000003908 liver function Effects 0.000 description 1
- 210000003141 lower extremity Anatomy 0.000 description 1
- 125000000955 oleanolic acid group Chemical group 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 235000005985 organic acids Nutrition 0.000 description 1
- 150000007530 organic bases Chemical class 0.000 description 1
- 206010033675 panniculitis Diseases 0.000 description 1
- 150000002966 pentacyclic triterpenoids Chemical class 0.000 description 1
- 201000001245 periodontitis Diseases 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- 230000001376 precipitating effect Effects 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 201000003068 rheumatic fever Diseases 0.000 description 1
- 210000000697 sensory organ Anatomy 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 201000005113 shigellosis Diseases 0.000 description 1
- 230000000192 social effect Effects 0.000 description 1
- 210000002784 stomach Anatomy 0.000 description 1
- 230000004206 stomach function Effects 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 238000007920 subcutaneous administration Methods 0.000 description 1
- 210000004304 subcutaneous tissue Anatomy 0.000 description 1
- 230000035922 thirst Effects 0.000 description 1
- 102000014898 transaminase activity proteins Human genes 0.000 description 1
- 231100000397 ulcer Toxicity 0.000 description 1
- 230000002485 urinary effect Effects 0.000 description 1
- 230000008673 vomiting Effects 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
Classifications
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/10—Selective adsorption, e.g. chromatography characterised by constructional or operational features
- B01D15/12—Selective adsorption, e.g. chromatography characterised by constructional or operational features relating to the preparation of the feed
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/10—Selective adsorption, e.g. chromatography characterised by constructional or operational features
- B01D15/18—Selective adsorption, e.g. chromatography characterised by constructional or operational features relating to flow patterns
- B01D15/1807—Selective adsorption, e.g. chromatography characterised by constructional or operational features relating to flow patterns using counter-currents, e.g. fluidised beds
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/26—Selective adsorption, e.g. chromatography characterised by the separation mechanism
- B01D15/38—Selective adsorption, e.g. chromatography characterised by the separation mechanism involving specific interaction not covered by one or more of groups B01D15/265 - B01D15/36
- B01D15/3861—Selective adsorption, e.g. chromatography characterised by the separation mechanism involving specific interaction not covered by one or more of groups B01D15/265 - B01D15/36 using an external stimulus
- B01D15/388—Selective adsorption, e.g. chromatography characterised by the separation mechanism involving specific interaction not covered by one or more of groups B01D15/265 - B01D15/36 using an external stimulus modifying the pH
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/90—Plate chromatography, e.g. thin layer or paper chromatography
Landscapes
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)
Abstract
A method for separating ursolic acid and oleanolic acid from plants, the method comprising: adding a mixed solvent system consisting of normal hexane, dichloromethane, methanol and water into a separating funnel, fully shaking, standing for layering, taking an upper phase, adding an acidic substance as a stationary phase, and adding an alkaline substance as a mobile phase into a lower phase; dissolving the crude extract of the plant by using a mixed solution of a stationary phase and a non-alkalized lower phase to obtain a sample solution; filling a separation column of a countercurrent chromatograph with the stationary phase, then injecting a sample solution, starting a speed controller after sample injection is finished, enabling the separation column to rotate forwards, continuously injecting a mobile phase, detecting by an ultraviolet detector, collecting eluent by an automatic part collector, merging the eluent containing a target substance, evaporating the solvent under reduced pressure, and drying to obtain a target product; the invention separates ursolic acid and oleanolic acid by pH zone refining countercurrent chromatography, and has simple process and high recovery rate.
Description
(1) technical field
The method of the present invention relates to a kind of from plant separating ursolic acid and oleanolic acid.
(2) background technique
Ursolic acid is to be present in one of natural plants triterpene compound, has calm, anti-inflammatory, antibacterial, anti-glycosuria
Various biologicals effect, the ursolic acid such as disease, antiulcer, reduction blood glucose also have apparent anti-oxidation function, thus by widely
As medicine and cosmetic material.Ursolic acid clinical manifestation has significant and reduces rapidly glutamic-pyruvic transaminase, serum transaminase, recession
Yellow subcutaneous ulcer improves a poor appetite, the effect of anti-fibrosis and recovery liver function, has the characteristics that quick, short treating period, effect stability.Together
Pier tartaric acid exists very extensively in plant, and general content 0.2%~2% is broad spectrum antibiotic, clinically drops enzyme for protect liver,
Treat bronchitis, pneumonia, acute tonsillitis, periodontitis, bacillary dysentery, acute gastroenteritis, urinary system infection contamination.In addition, clinical
On be also used to treat oxyhepatitis.
Aralia wood (Aralia chinensis), is Umbellales Araliaceae, has dispelling wind and eliminating dampness, inducing diuresis to remove edema, promoting blood circulation
The effects of analgesic, can be used for hepatitis, enlargement of lymph nodes, nephritic dropsy, diabetes, stomachache, rheumatic arthritis, pain in waist and lower extremities, bruise
The treatment of the diseases such as damage.Apple skin is the pericarp of rosaceae Malus apple (Malus domestica), in apple skin
Antioxidant content and bioactive substance rich in, it is beneficial to health to eat apple skin.Loguat leaf is that rosaceae loquat belongs to plant
The effect of dried leaf of object loquat (Eriobotrya japonica Thunb.) has a clearing lung and relieving cough, stomach function regulating diuresis, quenches the thirst and
There are treatment lung feverish cough, hemoptysis, bleeding from five sense organs or subcutaneous tissue, stomach hot vomiting effect.Main active in oleanolic acid Shi Aralia wood, ursolic acid with
Oleanolic acid is active constituent important in apple skin and loguat leaf.
The Aralia wood whole nation is widely distributed, early very high by the civil content using oleanolic acid in , Aralia wood in various regions as medicinal material,
In development of the West Regions, conceding the land to forestry, afforestation, this resource of active development is answered.China is the world's largest apple production
State and country of consumption can generate a large amount of apple skin in current consumption and processing apple.If apple skin is not handled well, will produce
Raw serious problem of environmental pollution.Ursolic acid is the most abundant pentacyclic triterpenoid of content in apple skin, is become in recent years
Research hotspot.China is that all there are plantation, therefore loguat leaf in most important loguat leaf producing country, each province on the south the Changjiang river in the world
Resource is very rich.However, most of loguat leaves are still used as garbage disposal, if dealt with improperly, such as burn, can also pollute ring
Border.The content of ursolic acid and oleanolic acid is higher in loguat leaf, is extracted by the deep processing of the effective active composition to loguat leaf
Purifying research makes its " turning waste into wealth ", then its market prospects is inevitable very optimistic.Therefore, Devoting Major Efforts To Developing rich in ursolic acid and
The plant of oleanolic acid, by with good economic efficiency and social effect.
Currently, the extraction of ursolic acid and oleanolic acid mainly passes through silica gel column chromatography, silica gel column chromatography is cumbersome, step
More, process is slowly increased the production cycle, and obtaining object ursolic acid and oleanolic acid, time-consuming, at high cost, it is difficult to obtain wide
General application.PH zone refining countercurrent (pH-zone-refining countercurrent chromatography) be by
A kind of special separation technology that common adverse current chromatogram develops has sample volume big (reaching gram-grade), separation purity height, separation
The advantages that time is short.The prominent feature of the technology is to joined reservation acid or alkali in stationary phase, joined and washes in mobile phase
Dealkalize or acid, according to the dissociation constant (pK of separate substancea) and hydrophobic difference and realize separation, target components are with rectangle peak
It is eluted out, and impurity is then enriched in rectangle peak edge, and different component is eluted out the variation along with pH value.
It is mainly used in the preparatives such as organic acids and base derivative, polypeptide, organic dyestuff, alkaloid, isomer, free peptide point
From.
(3) summary of the invention
The object of the present invention is to provide the methods of a kind of preparative separating ursolic acid and oleanolic acid, rapidly by ursolic acid
It is separated from plant with two kinds of substances of oleanolic acid, simple process, there is stronger purpose, quickly and efficiently.
Technical scheme is as follows:
A method of separating ursolic acid and oleanolic acid, the method from plant are as follows:
(1) mixed solvent system of n-hexane, methylene chloride, first alcohol and water composition is added in separatory funnel, sufficiently shakes
Shake rear stratification, take and be added to acidic materials as stationary phase, under be added to alkaline matter as mobile phase；
In the mixed solvent system, n-hexane, methylene chloride, methanol, water volume parts be respectively 6~8 parts, 3~5
Part, 2~4 parts, 6~8 parts, preferably respectively 7 parts, 3 parts, 2 parts, 8 parts；
The acidic materials are preferably trifluoroacetic acid or acetic acid, most preferably trifluoroacetic acid, and the acidic materials are in stationary phase
In concentration be 5~20mM, preferably 10mM；
The alkaline matter is preferably ammonium hydroxide (25~28wt%NH3) or triethylamine, most preferably ammonium hydroxide, the alkaline matter
Concentration in mobile phase is 5~20mM, preferably 10mM；
(2) mixed solution for the lower phase for taking the crude extract of plant stationary phase and not alkalizing dissolves, and obtains sample solution；
The plant is, for example: Aralia wood, apple skin, loguat leaf；
The volume ratio of the stationary phase and the lower phase not alkalized is 1:1；
The volumetric usage of the mixed solution of the stationary phase and the lower phase not alkalized is calculated as with the quality of the crude extract of plant
0.1mL/mg；
(3) it takes stationary phase to fill the splitter of counter-current chromatograph, is subsequently injected into sample solution, after the completion of sample introduction, open
Speed control rotates forward splitter, under 550~850r/min (preferably 600r/min) revolving speed, (preferably with 1~2mL/min
Flow velocity 1.5mL/min) persistently injects mobile phase, is detected with the UV detector of 200~254nm of wavelength, is received with automatic part
Storage collects eluent, merges the eluent containing target substance, evaporating solvent under reduced pressure and drying, obtains target product；
The sample solution can be injected by sample introduction circle or by being pumped into counter-current chromatograph；
The sampling volume of the sample solution is usually less than the 30% of splitter column volume, preferably 5%~10%；
The target substance can be monitored by thin-layer chromatography or high performance liquid chromatography, be merged and ursolic acid and olive
Sour standard items RfValue or the identical eluent containing ursolic acid and oleanolic acid of retention time；
The solvent of the thin-layer chromatography is that the mixing of hexamethylene, ethyl acetate and glacial acetic acid volume ratio 10:3:0.5 is molten
Agent；
The testing conditions of the high performance liquid chromatography are as follows: 5 HC-C of Agilent18(2) column (4.6 × 250mm, 5 μ
m)；25 DEG C of column temperature；Mobile phase is -0.5% ammonium acetate solution of acetonitrile-methanol (67:12:21, v/v/v), isocratic elution mode；Stream
Speed: 1mL/min；Detection wavelength: 210nm；Sample volume: 20 μ L.
In the present invention, the preparation method of the crude extract of the plant enumerates as follows:
The crude extract of Aralia wood is made as follows:
Qu Aralia wood root powder, with 95% ethanol water refluxing extraction of volume fraction 2 times, 1 hour every time, extracting solution was evaporated off molten
Agent adds 10wt%H at thick medicinal extract shape, thick paste object2SO4Aqueous solution is condensed back hydrolysis 5h, after the completion of hydrolysis, by water at 100 DEG C
Sediment filtering is solved, then sediment is washed till neutrality with water repeatedly, sediment adds 10wt%NaOH aqueous solution, pH to 8~9 is adjusted,
70 DEG C are heated to, is filtered while hot, filtrate adds 10wt%HCl aqueous solution tune pH to 1, stands 8~12h, filters, the sediment filtered out
Dry get Dao Aralia wood crude extract.
The crude extract of apple skin is made as follows:
Apple skin powder is added in 100 DEG C of water and decocts 10min, repeats to decoct 5~7 times, is clarified until the water of filtering becomes,
Apple skin powder after decoction is placed on (40 DEG C) dryings of air -oven, is then mentioned with the reflux of 95% ethanol water of volume fraction
It takes 2~3 times, 1 hour every time, after extracting solution concentration, 10wt%HCl aqueous solution is added, adjusts pH to 3, stands 8~12h, filters,
Filter cake petroleum ether, drying obtain the crude extract of apple skin.
The crude extract of loguat leaf is made as follows:
Dry loguat leaf is crushed, 20 meshes is crossed, weighs loguat leaf fine powder, 95% ethanol water of volume fraction is added to condense
Refluxing extraction 1h, filtering, filter residue repeat refluxing extraction 2~3 times, combined extract, and active carbon is added and is warming up to 70 DEG C, oscillation is de-
Color 20min, filtering, filtrate decompression are concentrated to dryness, and obtain the crude extract of loguat leaf.
The beneficial effects of the present invention are: pass through pH zone refining countercurrent separating ursolic acid and oleanolic acid, technique
Simply, the rate of recovery is high, provides new approaches for ursolic acid and isolating and purifying for oleanolic acid.
(4) Detailed description of the invention
Fig. 1: high speed adverse current chromatogram (HSCCC) figure of embodiment 1.
Fig. 2: high speed adverse current chromatogram (HSCCC) figure of embodiment 2.
Fig. 3: high speed adverse current chromatogram (HSCCC) figure of embodiment 3.
(5) specific embodiment
Technical solution of the present invention is described further with specific embodiment below, but protection scope of the present invention is not
It is limited to this.
Semi-preparative adverse current chromatogram is used in the embodiment of the present invention, (MP0106, Shanghai three are counter-current chromatograph by constant flow pump
Scientific instrument Co., Ltd), UV detector (UVD-680-3, Shanghai Jin Da biochemical instrument Co., Ltd), recorder (Jin Da
Work station, Shanghai Jin Da biochemical instrument Co., Ltd) etc. composition.
Embodiment 1:
Drying Aralia wood is crushed, 20 mesh , Qu Aralia wood powder 50g are crossed, 95% alcohol reflux measured with 10 times extracts 2 times
(every time 1 hour), filtering recycle ethyl alcohol into thick medicinal extract shape, and thick paste object adds the 10%H of 100mL2SO4It is condensed back at 100 DEG C
5h is hydrolyzed, after the completion of hydrolysis, hydrolytic precipitation object is filtered, then precipitating is washed till neutrality with water repeatedly, sediment adds 10%NaOH
Solution is adjusted pH to 8-9, is filtered while hot after being heated to 70 DEG C.Filtrate adds 10% hydrochloric acid tune pH to 1, and after standing overnight, drying is heavy
It forms sediment, obtains crude product 1.014g.
N-hexane, methylene chloride, first alcohol and water are configured in separatory funnel according to volume ratio 7:3:2:8, shake well
Stratification afterwards is above added to trifluoroacetic acid, makes the final concentration of 10mM of trifluoroacetic acid as stationary phase, under be added to ammonium hydroxide
(25%~28%NH3), make the final concentration of 10mM of ammonium hydroxide as mobile phase；Cheng Qu Aralia wood crude extract 100mg, is fixed with 5ml
The lower phased soln not alkalized with 5ml mutually, obtains sample solution.
Loquat Leave extract is separated using semi-preparative counter-current chromatograph, the column volume of splitter is 105ml.It is first
Stationary phase is first filled into splitter with the flow velocity of 10ml/min, above-mentioned sample solution is then injected into splitter by sample introduction circle,
After the completion of sample introduction, opening speed controller rotates forward splitter, and adjusting revolving speed is 600r/min, and the flow velocity of mobile phase is arranged
For 1.5ml/min, start to be pumped into mobile phase, receives eluent by the speed of 3 minutes/pipe with automatic fraction collector, use thin layer
(solvent is hexamethylene: ethyl acetate: glacial acetic acid=10:3:0.5) tracing detection is chromatographed into eluent without oleanolic acid group
Point, stop collecting.It collects obtained eluent to be detected by thin-layer chromatography and efficient liquid phase, merge and oleanolic acid standard
Product RfIt is worth (Rf=0.67) and the identical eluent of retention time (21.7min) with Rotary Evaporators recycling design obtains olive
Sour 38.56mg, the wherein purity 99.01% of oleanolic acid, the rate of recovery 90.23%.
The testing conditions of high performance liquid chromatography are as follows: 5 HC-C of Agilent18(2) column (4.6 × 250mm, 5 μm)；Column
25 DEG C of temperature；Mobile phase is -0.5% ammonium acetate solution of acetonitrile-methanol (67:12:21, v/v/v), isocratic elution mode；Detect wave
It is long: 210nm；Sample volume: 20 μ L.
Embodiment 2:
Dry apple skin is crushed, 20 meshes is crossed, weighs 50g apple skin powder, be added in 100 DEG C of water and decoct
10min is repeated several times (5-7 times), clarifies until remaining water becomes, places it in (40 DEG C) dryings of air -oven, is then added
10 times of 95% ethanol solutions of amount are condensed back 1h, filtering, and 95% ethanol solution that residue is measured with 10 times repeats above-mentioned reflux 2 times,
After concentrated extract to 50mL, 10% dilute hydrochloric acid is slowly added into concentrate, pH to 3 or so is adjusted, stood for merging filtrate
Night filters, and for filter cake with petroleum ether 2 times, drying obtains crude product 2.353g.
N-hexane, methylene chloride, first alcohol and water are configured in separatory funnel according to volume ratio 7:3:2:8, shake well
Stratification afterwards is above added to trifluoroacetic acid, makes the final concentration of 10mM of trifluoroacetic acid as stationary phase, under be added to ammonium hydroxide
(25%~28%NH3), make the final concentration of 10mM of ammonium hydroxide as mobile phase；Apple skin crude extract 100mg is weighed, it is solid with 5ml
The fixed lower phased soln not alkalized with 5ml mutually, obtains sample solution.
Loquat Leave extract is separated using semi-preparative counter-current chromatograph, the column volume of splitter is 105ml.It is first
Stationary phase is first filled into splitter with the flow velocity of 10ml/min, above-mentioned sample solution is then injected into splitter by sample introduction circle,
After the completion of sample introduction, opening speed controller rotates forward splitter, and adjusting revolving speed is 600r/min, and the flow velocity of mobile phase is arranged
For 1.5ml/min, start to be pumped into mobile phase, receives eluent by the speed of 3 minutes/pipe with automatic fraction collector, use thin layer
(solvent is hexamethylene: ethyl acetate: glacial acetic acid=10:3:0.5) tracing detection is chromatographed into eluent without ursolic acid and neat
Pier tartaric acid component stops collecting.It collects obtained eluent to be detected by thin-layer chromatography and efficient liquid phase, merging and black bearberry
Sour standard items RfIt is worth (Rf=0.60) and retention time (22.5min) is identical and oleanolic acid standard items RfIt is worth (Rf=0.67)
And the identical eluent of retention time (21.7min) obtains the mixing of ursolic acid and oleanolic acid with Rotary Evaporators recycling design
Object 65.6mg, the wherein purity 90.98% of ursolic acid, the rate of recovery 89.66%；The purity of oleanolic acid is 6.51%, recycling
Rate is 88.89%.
The testing conditions of high performance liquid chromatography are as follows: Agilent 5 HC-C18 (2) column (4.6 × 250mm, 5 μm)；Column
25 DEG C of temperature；Mobile phase is -0.5% ammonium acetate solution of acetonitrile-methanol (67:12:21, v/v/v), isocratic elution mode；Detect wave
It is long: 210nm；Sample volume: 20 μ L.
Embodiment 3:
Dry loguat leaf is crushed, 20 meshes is crossed, weighs the loguat leaf fine powder of 100g, add 10 times of 95% ethyl alcohol of amount molten
Liquid is condensed back 1h, filtering, and 95% ethanol solution that residue is measured with 10 times repeats above-mentioned reflux 2 times, and merging filtrate, filtrate adds
Enter 1.5% active carbon, 70 DEG C of decolorations are filtered after oscillation decoloration 20min, then concentrate filtrate to dry, obtain loguat leaf
Ethanol extract, obtain 1.535g crude extract.
N-hexane, methylene chloride, first alcohol and water are configured in separatory funnel according to volume ratio 7:3:2:8, shake well
Stratification afterwards is above added to trifluoroacetic acid, makes the final concentration of 10mM of trifluoroacetic acid as stationary phase, under be added to ammonium hydroxide
(25%~28%NH3), make the final concentration of 10mM of ammonium hydroxide as mobile phase；Loquat Leave extract 100mg is weighed, it is solid with 5ml
The fixed lower phased soln not alkalized with 5ml mutually, obtains sample solution.
Loquat Leave extract is separated using semi-preparative counter-current chromatograph, the column volume of splitter is 105ml.It is first
Stationary phase is first filled into splitter with the flow velocity of 10ml/min, above-mentioned sample solution is then injected into splitter by sample introduction circle,
After the completion of sample introduction, opening speed controller rotates forward splitter, and adjusting revolving speed is 600r/min, and the flow velocity of mobile phase is arranged
For 1.5ml/min, start to be pumped into mobile phase, receives eluent by the speed of 3 minutes/pipe with automatic fraction collector, use thin layer
(solvent is hexamethylene: ethyl acetate: glacial acetic acid=10:3:0.5) tracing detection is chromatographed into eluent without ursolic acid and neat
Pier tartaric acid component stops collecting.It collects obtained eluent to be detected by thin-layer chromatography and efficient liquid phase, merging and black bearberry
Sour standard items RfIt is worth (Rf=0.60) and retention time (22.5min) is identical and oleanolic acid standard items RfIt is worth (Rf=0.67)
And the identical eluent of retention time (21.7min) obtains the mixing of ursolic acid and oleanolic acid with Rotary Evaporators recycling design
Object 46.6mg, the wherein purity 74.35% of ursolic acid, the rate of recovery 86.59%；The purity of oleanolic acid is 23.61%, recycling
Rate is 90.99%.
The testing conditions of high performance liquid chromatography are as follows: 5 HC-C of Agilent18(2) column (4.6 × 250mm, 5 μm)；Column
25 DEG C of temperature；Mobile phase is -0.5% ammonium acetate solution of acetonitrile-methanol (67:12:21, v/v/v), isocratic elution mode；Detect wave
It is long: 210nm；Sample volume: 20 μ L.
Comparative example:
(Niu Huiying vinca alkaloids extract the separation purifying technique of ursolic acid and oleanolic acid in residue to Niu Huiying
Study [D] Northeast Forestry University, 2008.) ursolic acid and neat is extracted in residue using column chromatography purifying vinca alkaloids
Pier tartaric acid, specific process parameter are as follows: column chromatographic stuffing is 300-400 mesh silica gel, and applied sample amount is 0.2g/10g silica gel, is eluted molten
Agent is n-hexane: ethyl acetate=5:4, flow velocity 3ml/min, collects corresponding outflow component.4.84g vinca alkaloids acetic acid
Sample 2.28g is obtained after ethyl ester extract column chromatography, the purity of ursolic acid is 68.14% in column chromatographed product, and the rate of recovery is
86.05%；The purity of oleanolic acid is 17.79%, the rate of recovery 81.21%.
The production cost of the method is relatively high, the technique very complicated and production cycle is long, is not suitable for large-scale production.And
It is easy to operate using the ursolic acid and oleanolic acid in pH zone refining countercurrent separation loguat leaf in the present invention, when separation
Between it is short, the rate of recovery is high, and obtained ursolic acid and oleanolic acid purity is high has certain promotional value.
Claims (9)
1. a kind of method of the separating ursolic acid from plant and oleanolic acid, which is characterized in that the method are as follows:
(1) mixed solvent system of n-hexane, methylene chloride, first alcohol and water composition is added in separatory funnel, after shake well
Stratification takes and is added to acidic materials as stationary phase, under be added to alkaline matter as mobile phase；
In the mixed solvent system, n-hexane, methylene chloride, methanol, water volume parts be respectively 6~8 parts, 3~5 parts, 2
~4 parts, 6~8 parts；
The acidic materials are trifluoroacetic acid or acetic acid, and concentration of the acidic materials in stationary phase is 5~20mM；
The alkaline matter is ammonium hydroxide or triethylamine, and concentration of the alkaline matter in mobile phase is 5~20mM；
(2) mixed solution for the lower phase for taking the crude extract of plant stationary phase and not alkalizing dissolves, and obtains sample solution；
(3) it takes stationary phase to fill the splitter of counter-current chromatograph, is subsequently injected into sample solution, after the completion of sample introduction, opening speed
Controller rotates forward splitter, under 550~850r/min revolving speed, persistently injects mobile phase with the flow velocity of 1~2mL/min, with
The UV detector of 200~254nm of wavelength detects, and collects eluent with automatic fraction collector, merges washing containing target substance
De- liquid, evaporating solvent under reduced pressure and drying, obtain target product.
2. as described in claim 1 from plant separating ursolic acid and oleanolic acid method, which is characterized in that step (1)
In, in the mixed solvent system, n-hexane, methylene chloride, methanol, water volume parts be respectively 7 parts, 3 parts, 2 parts, 8 parts.
3. as described in claim 1 from plant separating ursolic acid and oleanolic acid method, which is characterized in that step (2)
In, the plant is: Aralia wood, apple skin, loguat leaf.
4. as described in claim 1 from plant separating ursolic acid and oleanolic acid method, which is characterized in that step (2)
In, the volume ratio of the stationary phase and the lower phase not alkalized is 1:1.
5. as described in claim 1 from plant separating ursolic acid and oleanolic acid method, which is characterized in that step (2)
In, the volumetric usage of the mixed solution of the stationary phase and the lower phase not alkalized is calculated as 0.1mL/ with the quality of the crude extract of plant
mg。
6. as described in claim 1 from plant separating ursolic acid and oleanolic acid method, which is characterized in that step (3)
In, the sample solution is injected by sample introduction circle or by being pumped into counter-current chromatograph.
7. as described in claim 1 from plant separating ursolic acid and oleanolic acid method, which is characterized in that step (3)
In, the sampling volume of the sample solution is less than the 30% of splitter column volume.
8. as described in claim 1 from plant separating ursolic acid and oleanolic acid method, which is characterized in that step (3)
In, the target substance is monitored by thin-layer chromatography or high performance liquid chromatography, is merged and ursolic acid and oleanolic acid standard
Product RfValue or the identical eluent containing ursolic acid and oleanolic acid of retention time；
The solvent of the thin-layer chromatography is the mixed solvent of hexamethylene, ethyl acetate and glacial acetic acid volume ratio 10:3:0.5；
The testing conditions of the high performance liquid chromatography are as follows: 5 HC-C of Agilent18(2)column；25 DEG C of column temperature；Mobile phase is second
The mixed solution of -0.5% ammonium acetate solution volume ratio 67:12:21 of nitrile-methanol, isocratic elution mode；Flow velocity: 1mL/min；Inspection
Survey wavelength: 210nm；Sample volume: 20 μ L.
9. as described in claim 1 from plant separating ursolic acid and oleanolic acid method, which is characterized in that the plant
Crude extract Wei Aralia wood, apple skin or loguat leaf crude extract, the crude extract of the plant the preparation method comprises the following steps:
The crude extract of Aralia wood is made as follows:
Qu Aralia wood root powder, with 95% ethanol water refluxing extraction of volume fraction 2 times, every time 1 hour, extracting solution be evaporated off solvent at
Thick medicinal extract shape, thick paste object add 10wt%H2SO4Aqueous solution is condensed back hydrolysis 5h at 100 DEG C, after the completion of hydrolysis, hydrolysis is heavy
Starch filtering, then sediment is washed till neutrality with water repeatedly, sediment adds 10wt%NaOH aqueous solution, adjusts pH to 8~9, heating
It to 70 DEG C, filters while hot, filtrate adds 10wt%HCl aqueous solution tune pH to 1, stands 8~12h, filters, the sediment drying filtered out
Crude extract of the get Dao Aralia wood；
The crude extract of apple skin is made as follows:
Apple skin powder is added in 100 DEG C of water and decocts 10min, repeats to decoct 5~7 times, clarifies, will decoct until the water of filtering becomes
Apple skin powder after boiling is placed on air -oven drying, then uses 95% ethanol water refluxing extraction of volume fraction 2~3 times,
1 hour every time, after extracting solution concentration, 10wt%HCl aqueous solution is added, adjusts pH to 3, stands 8~12h, filters, filter cake petroleum
Ether washing, drying obtain the crude extract of apple skin；
The crude extract of loguat leaf is made as follows:
Dry loguat leaf is crushed, 20 meshes is crossed, weighs loguat leaf fine powder, 95% ethanol water of volume fraction is added to be condensed back
1h, filtering are extracted, filter residue repeats refluxing extraction 2~3 times, combined extract, and active carbon is added and is warming up to 70 DEG C, oscillation decoloration
20min, filtering, filtrate decompression are concentrated to dryness, and obtain the crude extract of loguat leaf.
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Application publication date: 20191129 |