CN108409752B - Method for extracting and separating picrinine - Google Patents
Method for extracting and separating picrinine Download PDFInfo
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- CN108409752B CN108409752B CN201810562176.4A CN201810562176A CN108409752B CN 108409752 B CN108409752 B CN 108409752B CN 201810562176 A CN201810562176 A CN 201810562176A CN 108409752 B CN108409752 B CN 108409752B
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- picrinine
- chloroform
- triethylamine
- alkaloid
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- BDXYPHKGNUGUFG-VETGLWQVSA-N picrinine Chemical compound C12=CC=CC=C2N[C@]2(O3)[C@@H]4C[C@@H]5[C@@H](C(=O)OC)[C@]12C[C@H]3N4C\C5=C\C BDXYPHKGNUGUFG-VETGLWQVSA-N 0.000 title claims abstract description 66
- UBWVEGSADBOAEC-BASWHVEKSA-N Picrinine Natural products COC(=O)C1C2CC3C4Nc5ccccc5C14CCN3C/C/2=C/C UBWVEGSADBOAEC-BASWHVEKSA-N 0.000 title claims abstract description 65
- BDXYPHKGNUGUFG-WAYFEFRKSA-N deacetyl-deformopicraline Natural products COC(=O)[C@@H]1[C@H]2C[C@@H]3N(CC2=CC)[C@@H]4C[C@]15c6ccccc6N[C@]35O4 BDXYPHKGNUGUFG-WAYFEFRKSA-N 0.000 title claims abstract description 65
- 238000000034 method Methods 0.000 title claims abstract description 14
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims abstract description 114
- 239000002253 acid Substances 0.000 claims abstract description 8
- 239000011347 resin Substances 0.000 claims abstract description 7
- 229920005989 resin Polymers 0.000 claims abstract description 7
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 claims description 63
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 45
- 229930013930 alkaloid Natural products 0.000 claims description 38
- 150000003797 alkaloid derivatives Chemical class 0.000 claims description 38
- 239000000243 solution Substances 0.000 claims description 38
- 239000007864 aqueous solution Substances 0.000 claims description 37
- 239000003480 eluent Substances 0.000 claims description 36
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 34
- 238000001914 filtration Methods 0.000 claims description 31
- 239000000706 filtrate Substances 0.000 claims description 28
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 24
- 239000012043 crude product Substances 0.000 claims description 22
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 21
- 238000001035 drying Methods 0.000 claims description 21
- 239000013078 crystal Substances 0.000 claims description 18
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 16
- SKCNIGRBPJIUBQ-UHFFFAOYSA-N chloroform;ethyl acetate Chemical compound ClC(Cl)Cl.CCOC(C)=O SKCNIGRBPJIUBQ-UHFFFAOYSA-N 0.000 claims description 16
- 239000012535 impurity Substances 0.000 claims description 11
- JAYUDPKFDQGKFQ-UHFFFAOYSA-N n,n-diethylethanamine;ethanol Chemical compound CCO.CCN(CC)CC JAYUDPKFDQGKFQ-UHFFFAOYSA-N 0.000 claims description 10
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 9
- 239000003208 petroleum Substances 0.000 claims description 8
- 229920006122 polyamide resin Polymers 0.000 claims description 8
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 7
- 239000002026 chloroform extract Substances 0.000 claims description 7
- 238000002425 crystallisation Methods 0.000 claims description 7
- 230000008025 crystallization Effects 0.000 claims description 7
- 239000000741 silica gel Substances 0.000 claims description 7
- 229910002027 silica gel Inorganic materials 0.000 claims description 7
- IDGUHHHQCWSQLU-UHFFFAOYSA-N ethanol;hydrate Chemical compound O.CCO IDGUHHHQCWSQLU-UHFFFAOYSA-N 0.000 claims description 6
- 239000003513 alkali Substances 0.000 claims description 5
- 238000010828 elution Methods 0.000 claims description 4
- 238000001291 vacuum drying Methods 0.000 claims description 4
- 239000000047 product Substances 0.000 claims description 3
- DKAGJZJALZXOOV-UHFFFAOYSA-N hydrate;hydrochloride Chemical compound O.Cl DKAGJZJALZXOOV-UHFFFAOYSA-N 0.000 claims description 2
- GBMDVOWEEQVZKZ-UHFFFAOYSA-N methanol;hydrate Chemical compound O.OC GBMDVOWEEQVZKZ-UHFFFAOYSA-N 0.000 claims description 2
- QMYTXCUSWCFXLY-UHFFFAOYSA-N methanol;triethylazanium;hydroxide Chemical compound [OH-].OC.CC[NH+](CC)CC QMYTXCUSWCFXLY-UHFFFAOYSA-N 0.000 claims description 2
- FWFGVMYFCODZRD-UHFFFAOYSA-N oxidanium;hydrogen sulfate Chemical compound O.OS(O)(=O)=O FWFGVMYFCODZRD-UHFFFAOYSA-N 0.000 claims description 2
- 238000000605 extraction Methods 0.000 abstract description 9
- 238000000926 separation method Methods 0.000 abstract description 4
- 238000004440 column chromatography Methods 0.000 abstract description 2
- 239000002994 raw material Substances 0.000 abstract description 2
- 238000010898 silica gel chromatography Methods 0.000 abstract description 2
- 238000001953 recrystallisation Methods 0.000 abstract 1
- 238000003809 water extraction Methods 0.000 abstract 1
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 14
- 238000009835 boiling Methods 0.000 description 12
- 238000003756 stirring Methods 0.000 description 10
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 9
- HIXDQWDOVZUNNA-UHFFFAOYSA-N 2-(3,4-dimethoxyphenyl)-5-hydroxy-7-methoxychromen-4-one Chemical compound C=1C(OC)=CC(O)=C(C(C=2)=O)C=1OC=2C1=CC=C(OC)C(OC)=C1 HIXDQWDOVZUNNA-UHFFFAOYSA-N 0.000 description 7
- 238000002156 mixing Methods 0.000 description 6
- 239000012141 concentrate Substances 0.000 description 5
- 239000000284 extract Substances 0.000 description 5
- 239000012530 fluid Substances 0.000 description 5
- 239000002699 waste material Substances 0.000 description 5
- FKSLYSSVKFYJKE-UHFFFAOYSA-N n,n-diethylethanamine;methanol Chemical compound OC.CCN(CC)CC FKSLYSSVKFYJKE-UHFFFAOYSA-N 0.000 description 4
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 239000003960 organic solvent Substances 0.000 description 3
- 238000002360 preparation method Methods 0.000 description 3
- 206010011224 Cough Diseases 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000007796 conventional method Methods 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- JEUXZUSUYIHGNL-UHFFFAOYSA-N n,n-diethylethanamine;hydrate Chemical compound O.CCN(CC)CC JEUXZUSUYIHGNL-UHFFFAOYSA-N 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 206010006458 Bronchitis chronic Diseases 0.000 description 1
- 206010062717 Increased upper airway secretion Diseases 0.000 description 1
- GAMYVSCDDLXAQW-AOIWZFSPSA-N Thermopsosid Natural products O(C)c1c(O)ccc(C=2Oc3c(c(O)cc(O[C@H]4[C@H](O)[C@@H](O)[C@H](O)[C@H](CO)O4)c3)C(=O)C=2)c1 GAMYVSCDDLXAQW-AOIWZFSPSA-N 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 206010006451 bronchitis Diseases 0.000 description 1
- 208000007451 chronic bronchitis Diseases 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- -1 echinocandine Chemical class 0.000 description 1
- 150000002148 esters Chemical class 0.000 description 1
- 229930003944 flavone Natural products 0.000 description 1
- 150000002212 flavone derivatives Chemical class 0.000 description 1
- 235000011949 flavones Nutrition 0.000 description 1
- 239000008187 granular material Substances 0.000 description 1
- 241000411851 herbal medicine Species 0.000 description 1
- 229930005303 indole alkaloid Natural products 0.000 description 1
- 150000002475 indoles Chemical class 0.000 description 1
- AFJPGVUCVDCFPM-BBFZNYENSA-N methyl (1S,9R,10S,12S,13E,15S,18R)-13-ethylidene-10-hydroxy-18-(hydroxymethyl)-15-methyl-8-aza-15-azoniapentacyclo[10.5.1.01,9.02,7.09,15]octadeca-2,4,6-triene-18-carboxylate Chemical compound COC(=O)[C@]1(CO)[C@H]2C[C@H](O)[C@@]34Nc5ccccc5[C@@]13CC[N@@+]4(C)C\C2=C\C AFJPGVUCVDCFPM-BBFZNYENSA-N 0.000 description 1
- 208000026435 phlegm Diseases 0.000 description 1
- 150000004032 porphyrins Chemical class 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- VHBFFQKBGNRLFZ-UHFFFAOYSA-N vitamin p Natural products O1C2=CC=CC=C2C(=O)C=C1C1=CC=CC=C1 VHBFFQKBGNRLFZ-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D498/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms
- C07D498/22—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms in which the condensed system contains four or more hetero rings
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Medicines Containing Plant Substances (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention discloses a method for extracting and separating picrinine, which takes alstonia-leaf as a raw material and adopts the technical processes of acid water extraction, chloroform extraction, resin column chromatography separation, silica gel column chromatography separation, recrystallization and the like to prepare a product with the content of more than 98 percent.
Description
Technical Field
The invention relates to a method for extracting and separating picrinine from alstonia-leaf, belonging to the field of compound separation.
Background
The product is a medicine for minority nationalities of Guangdong province and Yunnan province, and is prepared from the leaf of common Alstoniae, Scholaris, the Scholaris, the southern province and the like. It is recorded in the local medical records such as Luchuan Ben Cao and Yunnan Chinese herbal medicine selection, and recorded in the Yunnan province pharmaceutical standards (1974) and the Chinese pharmacopoeia (1977 edition I); and various preparations such as alstonia-leaf granules, alstonia-leaf and the like are developed later and are used for relieving cough, eliminating phlegm and treating chronic bronchitis cough.
Alkaloid and flavone are main components of folium Alstoniae Scholaris, and stem and leaf contain various indole alkaloids, such as picrinine, ditaine, echinocandine, porphyrin, etc. Alkaloid is an important active substance of alstonia-leaf, wherein picrinine (picrinine) has the highest content, is a marking component of alstonia-leaf and is a control item of a preparation.
Picrinine is colorless needle crystal, is easily soluble in methanol, chloroform, and acetone, is soluble in ethyl acetate and ethanol, and is insoluble in water and petroleum ether; is easy to dissolve in dilute hydrochloric acid and dilute sulfuric acid, and precipitates are re-separated after the solution becomes turbid after the solution is neutralized by adding alkali. The chemical name is 2 alpha, 5 alpha-Epoxy-1, 2-dihydroakuammlian-17-oleacidmethyl ester, and the chemical structural formula is shown as formula I;
the alstonia-leaf is a drug for minority nationalities, so that research data on the alstonia-leaf is few, and picrinine is an identification component of the alstonia-leaf and is a control project of a preparation, so that a method for separating picrinine is found, and more pharmacology and activity of the picrinine are researched necessarily.
Disclosure of Invention
The invention aims to provide a method for extracting and separating picrinine from alstonia-leaf, which takes alstonia-leaf as a raw material; the purpose of the invention is realized by the following technical scheme:
(1) adding acid water with the mass 5-7 times of that of the dried alstonia-leaf into the dried alstonia-leaf, decocting for 2-3 times, 2-3 hours each time, and filtering; combining the filtrates, concentrating the filtrate to a relative density of 1.05-1.10, extracting with chloroform for three times, removing a chloroform layer, adjusting the pH of the residual raffinate to 9-10 with alkali, extracting with chloroform for three times, collecting combined chloroform extract, concentrating and drying to obtain alkaloid;
(2) dissolving alkaloid by using an ethanol solution or a methanol solution with the volume concentration of 80-90%, filtering, adding a triethylamine aqueous solution with the mass concentration of 0.01-0.02% into the filtrate to enable the concentration of ethanol or methanol in the filtrate to be 25-30%, adsorbing the alkaloid by using a resin column, eluting impurities by using an eluent I with the column volume of 3-4 times, eluting by using an eluent II, collecting an eluent of the picrinine section, concentrating, and drying to obtain a crude picrinine product, wherein the eluent I is triethylamine aqueous solution or triethylamine ethanol aqueous solution with the mass concentration of 0.01-0.02%, which is prepared by adding triethylamine into an ethanol aqueous solution or a methanol aqueous solution with the mass concentration of 40-45%; adding triethylamine into an ethanol water solution or a methanol water solution with the mass concentration of 50-55% to prepare a triethylamine methanol water solution or a triethylamine ethanol water solution with the mass concentration of 0.01-0.02%;
(3) dissolving the picrinine crude product with chloroform with the mass of 4-5 times of that of the picrinine crude product, putting the crude product into a silica gel column, eluting with chloroform with the volume of 2-3 times of that of the column, performing gradient elution with chloroform-ethyl acetate, collecting the picrinine section eluent, and concentrating and drying to obtain picrinine;
(4) dissolving picrinine in the step (3) by using acetone, then dropwise adding petroleum ether until crystals appear, standing for crystallization, filtering, repeatedly operating for 2-3 times, and finally drying the crystals in vacuum to obtain the picrinine.
The acid water in the step (1) is sulfuric acid water solution or hydrochloric acid water solution with the mass concentration of 0.1-0.2%.
The alkali in the step (1) is a sodium hydroxide aqueous solution with the mass concentration of 40-50%.
And (3) the resin in the resin column in the step (2) is polyamide resin.
And (3) dissolving the alkaloid in the step (2) by using an ethanol solution with the mass of 3-4 times that of the alkaloid.
And (3) eluting impurities by using 3-4 times of column volume of chloroform-ethyl acetate with the volume ratio of 75-80: 20-25 during gradient elution in the step (3), and eluting the alkaloid by using chloroform-ethyl acetate with the volume ratio of 65-70: 30-35.
The method has the advantages and the technical effects that: the picrinine content in the alstonia-leaf is very low, only about one thousandth of the picrinine content, and the separation and purification are difficult; according to the invention, 0.1-0.2% acid water is adopted for extraction according to the property that picrinine is easily dissolved in acid water during extraction, so that the extraction rate of picrinine is ensured, and the adoption of expensive organic solvents is avoided; extracting impurities with chloroform according to the characteristic that picrinine salt is easily soluble in acid water and insoluble in organic solvent; and the raffinate after chloroform extraction is adjusted to be alkaline according to the characteristic that picrinine is easily dissolved in an organic solvent in an alkaline environment, and then the alkaloid is extracted by using chloroform; the extracted alkaloid is subjected to column chromatography by polyamide resin, then subjected to silica gel column chromatography, and finally recrystallized to obtain picrinine with the content of more than 98%.
Detailed Description
The present invention will be described in further detail with reference to the following examples, but the scope of the present invention is not limited thereto, and the methods of the present invention are those using conventional methods unless otherwise specified, and those using reagents prepared by conventional methods or commercially available reagents unless otherwise specified.
Example 1: taking 10kg of alstonia-leaf, adding 70kg of sulfuric acid aqueous solution with the mass concentration of 0.1%, boiling for 3 hours, filtering, adding 50kg of sulfuric acid aqueous solution with the mass concentration of 0.1% into filter residue again, boiling and extracting for 3 hours, filtering, combining filtrates obtained by secondary filtering, and concentrating the filtrate under reduced pressure until the relative density is 1.05 to obtain 20kg of clear paste; adding 10kg chloroform into the fluid extract under stirring, stirring for 5 min, standing for 2 hr, separating chloroform layer, repeatedly extracting water layer with equal amount of chloroform for 2 times, mixing chloroform extracts, recovering chloroform, and treating the concentrate as waste; adjusting pH of water layer to =9 with 40% sodium hydroxide, extracting with chloroform for 3 times (10 kg each time), mixing 3 chloroform extractive solutions, concentrating, and drying to obtain alkaloid; dissolving alkaloid with 90% ethanol solution with 3 times of the mass of the alkaloid, filtering, adding 0.01% triethylamine aqueous solution into the filtrate to enable the ethanol concentration in the filtrate to be 25%, then passing through a polyamide resin column, eluting impurities with 4 column volumes of eluent I (triethylamine is added into 40% ethanol aqueous solution to prepare 0.01% triethylamine ethanol aqueous solution), eluting the alkaloid with eluent II (triethylamine is added into 50% ethanol aqueous solution to prepare 0.01% triethylamine ethanol aqueous solution), collecting eluent containing picrinine segments, concentrating and drying to obtain crude picrinine; dissolving the picrinine crude product by using chloroform with the mass 5 times of that of the picrinine crude product, putting the mixture into a silica gel column, eluting by using chloroform with the volume 2 times of the column volume, and then eluting by using chloroform with the volume ratio of 3 times of the column volume of 80:20 of chloroform-ethyl acetate eluent, then eluting with a volume ratio of 70: eluting alkaloid with chloroform-ethyl acetate eluate of 30, collecting picrinine eluate, concentrating, drying, dissolving with acetone, adding petroleum ether dropwise until crystal appears, standing for crystallization, filtering, repeating for 2 times, and vacuum drying to obtain picrinine crystal 4.2g with content of 98.2%.
Example 2: taking 10kg of alstonia-leaf, adding 60kg of hydrochloric acid aqueous solution with the mass concentration of 0.1%, boiling for 2 hours, filtering, adding 60kg of hydrochloric acid aqueous solution with the mass concentration of 0.1% again into filter residue, boiling and extracting for 2 hours, filtering, boiling and extracting again, combining filtrate obtained by filtering for three times, concentrating the filtrate under reduced pressure until the relative density is 1.08, and obtaining 22kg of clear paste; adding 10kg chloroform into the fluid extract under stirring, stirring for 5 min, standing for 2 hr, separating chloroform layer, repeatedly extracting water layer with equal amount of chloroform for 2 times, mixing chloroform extracts, recovering chloroform, and treating the concentrate as waste; adjusting the pH of a water layer to be =10 by using 50% sodium hydroxide, extracting for 3 times by using chloroform, wherein the dosage of each time is 10kg, combining 3 times of chloroform extraction solutions, and concentrating and drying to obtain alkaloid; dissolving alkaloid with 80% methanol solution 4 times of the mass of alkaloid, filtering, adding 0.02% triethylamine aqueous solution into the filtrate to make the methanol concentration in the filtrate 30%, then passing through a polyamide resin column, eluting impurities with 3 column volumes of eluent I (triethylamine is added into 45% methanol aqueous solution to prepare 0.02% triethylamine methanol aqueous solution), eluting alkaloid with eluent II (triethylamine is added into 55% methanol aqueous solution to prepare 0.02% triethylamine methanol aqueous solution), collecting eluent containing picrinine segment, concentrating and drying to obtain picrinine crude product; dissolving the picrinine crude product by using chloroform with the mass of 4 times of the picrinine crude product, putting the mixture into a silica gel column, eluting by using chloroform with the volume of 3 times of the column volume, and then eluting by using chloroform with the volume ratio of 4 times of the column volume of 75: 25, and then eluting with a chloroform-ethyl acetate eluent at a volume ratio of 65: 35, eluting alkaloid with chloroform-ethyl acetate eluent, collecting picrinine section eluent, concentrating and drying, dissolving with acetone, dripping petroleum ether until crystal appears, standing for crystallization, filtering, repeating for 3 times, and vacuum drying to obtain 3.8g of picrinine crystal with content of 98.7%.
Example 3: taking 10kg of alstonia-leaf, adding 60kg of sulfuric acid aqueous solution with the mass concentration of 0.2%, boiling for 2.5 hours, filtering, adding 50kg of sulfuric acid aqueous solution with the mass concentration of 0.1% into filter residue again, boiling and extracting for 2 hours, filtering, combining filtrates obtained by secondary filtration, and concentrating the filtrate under reduced pressure until the relative density is 1.1 to obtain 17kg of clear paste; adding 10kg chloroform into the fluid extract under stirring, stirring for 5 min, standing for 2 hr, separating chloroform layer, repeatedly extracting water layer with equal amount of chloroform for 2 times, mixing chloroform extracts, recovering chloroform, and treating the concentrate as waste; adjusting the pH of a water layer to be =9.5 by using sodium hydroxide with the mass concentration of 45%, extracting for 3 times by using chloroform, wherein the dosage of each time is 10kg, and combining 3 times of chloroform extraction liquid to obtain alkaloid; dissolving alkaloid with 85% ethanol solution with 3.5 times volume concentration by mass, filtering, adding 0.015% triethylamine water solution into filtrate to make the ethanol concentration in the filtrate be 28%, then passing through polyamide resin column, eluting with 3.5 column volumes of eluent I (triethylamine is added into 42% ethanol water solution to prepare 0.015% triethylamine ethanol water solution), eluting alkaloid with eluent II (triethylamine is added into 53% ethanol water solution to prepare 0.014% triethylamine ethanol water solution), collecting eluent containing picrinine segment, concentrating and drying to obtain picrinine crude product; dissolving the picrinine crude product by using chloroform with the mass of 4.5 times of the picrinine crude product, putting the solution into a silica gel column, eluting by using chloroform with the volume of 2.5 times of the column volume, and then using a volume ratio of 3.5 times of the column volume of 78: 22, and then eluting with a chloroform-ethyl acetate eluent at a volume ratio of 67: eluting alkaloid with chloroform-ethyl acetate eluate of 33, collecting eluate of picrinine section, concentrating, drying, dissolving with acetone, adding petroleum ether dropwise until crystal appears, standing for crystallization, filtering, repeating for 2 times, and vacuum drying to obtain 4.5g of picrinine crystal with content of 98.5%.
Example 4: taking 50kg of alstonia-leaf, adding 250kg of sulfuric acid aqueous solution with the mass concentration of 0.15%, boiling for 3 hours, filtering, adding 250kg of sulfuric acid aqueous solution with the mass concentration of 0.2% into filter residue again, boiling and extracting for 3 hours, filtering, boiling and extracting again, merging filtrate obtained by filtering for 3 times, concentrating the filtrate under reduced pressure until the relative density is 1.08, and obtaining 110kg of clear paste; adding 50kg chloroform into the fluid extract under stirring, stirring for 5 min, standing for 2 hr, separating chloroform layer, repeatedly extracting water layer with equal amount of chloroform for 2 times, mixing chloroform extracts, recovering chloroform, and treating the concentrate as waste; adjusting the pH of a water layer to be =9 by using 42% sodium hydroxide, extracting for 3 times by using chloroform, wherein the dosage of each time is 50kg, and combining 3 times of chloroform extraction liquid to obtain alkaloid; dissolving alkaloid with a methanol solution with the mass of 3 times and volume concentration of 88%, filtering, adding a triethylamine aqueous solution with the mass concentration of 0.02% into the filtrate to enable the methanol concentration in the filtrate to be 26%, then passing through a polyamide resin column, eluting impurities with 4 column volumes of eluent I (triethylamine is added into the methanol aqueous solution with the mass concentration of 40% to prepare the triethylamine methanol aqueous solution with the mass concentration of 0.02%), then eluting the alkaloid with an eluent II (triethylamine is added into the methanol aqueous solution with the mass concentration of 50% to prepare the triethylamine methanol aqueous solution with the mass concentration of 0.02%), collecting eluent containing picrinine segments, concentrating and drying to obtain a picrinine crude product; dissolving the picrinine crude product with chloroform with the mass of 5 times of the crude product, putting the crude product into a silica gel column, eluting by using chloroform with the volume of 3 times of the column volume, eluting impurities by using chloroform-ethyl acetate eluent with the volume ratio of 3 times of the column volume at 76:24, eluting alkaloid by using chloroform-ethyl acetate eluent with the volume ratio of 68:22, collecting picrinine section eluent, concentrating and drying, dissolving by using acetone, dropwise adding petroleum ether until crystals appear, standing for crystallization, filtering, repeating the operation for 2 times, and finally drying the crystals in vacuum to obtain 23.2g of picrinine crystals with the content of 98.7%.
Example 5: taking 50kg of alstonia-leaf, adding 300kg of hydrochloric acid aqueous solution with the mass concentration of 0.2%, boiling for 3 hours, filtering, adding 250kg of hydrochloric acid aqueous solution with the mass concentration of 0.2% into filter residue again, boiling and extracting for 2 hours, filtering, combining filtrates obtained by filtering for 2 times, and concentrating the filtrate under reduced pressure until the relative density is 1.1 to obtain 98kg of clear paste; adding 50kg chloroform into the fluid extract under stirring, stirring for 5 min, standing for 2 hr, separating chloroform layer, repeatedly extracting water layer with equal amount of chloroform for 2 times, mixing chloroform extracts, recovering chloroform, and treating the concentrate as waste; adjusting the pH of a water layer to be =10 by using sodium hydroxide with the mass concentration of 45%, extracting for 3 times by using chloroform, wherein the dosage of each time is 50kg, and combining 3 times of chloroform extraction liquid to obtain alkaloid; dissolving alkaloid with ethanol solution with the mass 3 times and volume concentration of 85%, filtering, adding triethylamine water solution with the mass concentration of 0.01% into filtrate to enable the ethanol concentration in the filtrate to be 27%, then passing through a polyamide resin column, eluting impurities with 3 column volume eluent I (triethylamine is added into the ethanol water solution with the mass concentration of 45% to prepare triethylamine ethanol water solution with the mass concentration of 0.01%), then eluting alkaloid with eluent II (triethylamine is added into the ethanol water solution with the mass concentration of 54% to prepare triethylamine ethanol water solution with the mass concentration of 0.02%), collecting eluent containing picrinine segments, concentrating and drying to obtain picrinine crude product; dissolving the picrinine crude product with chloroform with the mass of 4 times of the crude product, putting the crude product into a silica gel column, eluting by using chloroform with the volume of 3 times of the column volume, eluting impurities by using chloroform-ethyl acetate eluent with the volume ratio of 4 times of the column volume of 80:20, eluting alkaloid by using chloroform-ethyl acetate eluent with the volume ratio of 66:34, collecting the picrinine section eluent, concentrating and drying, dissolving by using acetone, dropwise adding petroleum ether until crystals appear, standing for crystallization, filtering, repeating the operation for 3 times, and finally drying the crystals in vacuum to obtain 25.8g of the picrinine crystal crystals with the content of 98.2 percent.
Claims (3)
1. The method for extracting and separating picrinine is characterized by comprising the following steps:
(1) adding acid water with the mass 5-7 times of that of the dried alstonia-leaf into the dried alstonia-leaf, decocting for 2-3 times, 2-3 hours each time, and filtering; combining the filtrates, concentrating the filtrate to a relative density of 1.05-1.10, extracting with chloroform for three times, removing a chloroform layer, adjusting the pH of the residual raffinate to 9-10 with alkali, extracting with chloroform for three times, collecting combined chloroform extract, concentrating and drying to obtain alkaloid;
the acid water in the step (1) is sulfuric acid water solution or hydrochloric acid water solution with the mass concentration of 0.1-0.2%;
(2) dissolving alkaloid by using an ethanol solution or a methanol solution with the volume concentration of 80-90%, filtering, adding a triethylamine aqueous solution with the mass concentration of 0.01-0.02% into the filtrate to enable the concentration of ethanol or methanol in the filtrate to be 25-30%, adsorbing the alkaloid by using a resin column, eluting impurities by using an eluent I with the column volume of 3-4 times, eluting by using an eluent II, collecting an eluent of the picrinine section, concentrating, and drying to obtain a crude picrinine product, wherein the eluent I is triethylamine aqueous solution or triethylamine ethanol aqueous solution with the mass concentration of 0.01-0.02%, which is prepared by adding triethylamine into an ethanol aqueous solution or a methanol aqueous solution with the mass concentration of 40-45%; adding triethylamine into an ethanol water solution or a methanol water solution with the mass concentration of 50-55% to prepare a triethylamine methanol water solution or a triethylamine ethanol water solution with the mass concentration of 0.01-0.02%;
the resin in the resin column is polyamide resin;
(3) dissolving the picrinine crude product with chloroform with the mass being 4-5 times of that of the picrinine crude product, putting the crude product into a silica gel column, eluting with chloroform with the volume being 2-3 times of that of the column, performing gradient elution with chloroform-ethyl acetate, collecting the picrinine section eluent, concentrating and drying to obtain the picrinine;
(4) dissolving picrinine in step (3) with acetone, dripping petroleum ether until crystals appear, standing for crystallization, filtering, repeating for 2-3 times, and finally vacuum drying the crystals to obtain picrinine
And (3) eluting impurities by using 3-4 times of column volume of chloroform-ethyl acetate with the volume ratio of 75-80: 20-25 during gradient elution in the step (3), and eluting the alkaloid by using chloroform-ethyl acetate with the volume ratio of 65-70: 30-35.
2. The method for extracting and separating picrinine according to claim 1, wherein the method comprises the following steps: the alkali in the step (1) is a sodium hydroxide aqueous solution with the mass concentration of 40-50%.
3. The method for extracting and separating picrinine according to claim 1, wherein the method comprises the following steps: and (3) dissolving the alkaloid in the step (2) by using an ethanol solution with the mass of 3-4 times that of the alkaloid.
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| CN101084894A (en) * | 2006-06-07 | 2007-12-12 | 中国科学院昆明植物研究所 | Medicine for treating diseases concerned with respiratory and use thereof |
| CN101658540A (en) * | 2009-09-24 | 2010-03-03 | 刘富来 | Method for preparing medicine for treating respiratory tract diseases caused by pig blue ear diseases and the like |
| CN102040613A (en) * | 2010-12-03 | 2011-05-04 | 昆明振华制药厂有限公司 | Picrinine reference substance in common alstonia leaf and preparation method thereof |
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| CN101084894A (en) * | 2006-06-07 | 2007-12-12 | 中国科学院昆明植物研究所 | Medicine for treating diseases concerned with respiratory and use thereof |
| CN101658540A (en) * | 2009-09-24 | 2010-03-03 | 刘富来 | Method for preparing medicine for treating respiratory tract diseases caused by pig blue ear diseases and the like |
| CN102040613A (en) * | 2010-12-03 | 2011-05-04 | 昆明振华制药厂有限公司 | Picrinine reference substance in common alstonia leaf and preparation method thereof |
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