CN109942515A - A method of extracting 10- deacetylate taxol - Google Patents
A method of extracting 10- deacetylate taxol Download PDFInfo
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- CN109942515A CN109942515A CN201910348505.XA CN201910348505A CN109942515A CN 109942515 A CN109942515 A CN 109942515A CN 201910348505 A CN201910348505 A CN 201910348505A CN 109942515 A CN109942515 A CN 109942515A
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- taxol
- deacetylate
- deacetylate taxol
- acetone
- concentrate
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- 229930012538 Paclitaxel Natural products 0.000 title claims abstract description 101
- 229960001592 paclitaxel Drugs 0.000 title claims abstract description 101
- RCINICONZNJXQF-MZXODVADSA-N taxol Chemical compound O([C@@H]1[C@@]2(C[C@@H](C(C)=C(C2(C)C)[C@H](C([C@]2(C)[C@@H](O)C[C@H]3OC[C@]3([C@H]21)OC(C)=O)=O)OC(=O)C)OC(=O)[C@H](O)[C@@H](NC(=O)C=1C=CC=CC=1)C=1C=CC=CC=1)O)C(=O)C1=CC=CC=C1 RCINICONZNJXQF-MZXODVADSA-N 0.000 title claims abstract description 99
- 238000000034 method Methods 0.000 title claims abstract description 32
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims abstract description 45
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims abstract description 36
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims abstract description 30
- 239000012141 concentrate Substances 0.000 claims abstract description 27
- 239000000706 filtrate Substances 0.000 claims abstract description 22
- 239000012043 crude product Substances 0.000 claims abstract description 21
- 239000000047 product Substances 0.000 claims abstract description 21
- LRHPLDYGYMQRHN-UHFFFAOYSA-N N-Butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 claims abstract description 14
- 238000007710 freezing Methods 0.000 claims abstract description 12
- 230000008014 freezing Effects 0.000 claims abstract description 12
- PHTQWCKDNZKARW-UHFFFAOYSA-N isoamylol Chemical compound CC(C)CCO PHTQWCKDNZKARW-UHFFFAOYSA-N 0.000 claims abstract description 12
- 239000012074 organic phase Substances 0.000 claims abstract description 11
- 230000002378 acidificating effect Effects 0.000 claims abstract description 10
- PNEYBMLMFCGWSK-UHFFFAOYSA-N aluminium oxide Inorganic materials [O-2].[O-2].[O-2].[Al+3].[Al+3] PNEYBMLMFCGWSK-UHFFFAOYSA-N 0.000 claims abstract description 10
- 238000001556 precipitation Methods 0.000 claims abstract description 10
- 239000002994 raw material Substances 0.000 claims abstract description 9
- 238000001953 recrystallisation Methods 0.000 claims abstract description 9
- 238000004440 column chromatography Methods 0.000 claims abstract description 8
- 238000003809 water extraction Methods 0.000 claims abstract description 7
- 230000001376 precipitating effect Effects 0.000 claims abstract description 6
- 239000006228 supernatant Substances 0.000 claims abstract description 6
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 51
- 238000001914 filtration Methods 0.000 claims description 26
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims description 24
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 21
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 19
- 239000000741 silica gel Substances 0.000 claims description 19
- 229910002027 silica gel Inorganic materials 0.000 claims description 19
- 229960001866 silicon dioxide Drugs 0.000 claims description 19
- IMNFDUFMRHMDMM-UHFFFAOYSA-N N-Heptane Chemical compound CCCCCCC IMNFDUFMRHMDMM-UHFFFAOYSA-N 0.000 claims description 13
- 239000013078 crystal Substances 0.000 claims description 13
- 239000011265 semifinished product Substances 0.000 claims description 12
- 239000003480 eluent Substances 0.000 claims description 10
- 239000003292 glue Substances 0.000 claims description 10
- 239000000203 mixture Substances 0.000 claims description 10
- 238000003756 stirring Methods 0.000 claims description 10
- 238000001035 drying Methods 0.000 claims description 9
- FDSGHYHRLSWSLQ-UHFFFAOYSA-N dichloromethane;propan-2-one Chemical compound ClCCl.CC(C)=O FDSGHYHRLSWSLQ-UHFFFAOYSA-N 0.000 claims description 6
- JYGYEBCBALMPDC-UHFFFAOYSA-N heptane;propan-2-one Chemical compound CC(C)=O.CCCCCCC JYGYEBCBALMPDC-UHFFFAOYSA-N 0.000 claims description 6
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 claims description 6
- 238000004458 analytical method Methods 0.000 claims description 5
- 238000005352 clarification Methods 0.000 claims description 5
- 238000011068 loading method Methods 0.000 claims description 5
- 239000006210 lotion Substances 0.000 claims description 5
- 239000000463 material Substances 0.000 claims description 5
- 238000005057 refrigeration Methods 0.000 claims description 5
- 235000013599 spices Nutrition 0.000 claims description 5
- WBKFWQBXFREOFH-UHFFFAOYSA-N dichloromethane;ethyl acetate Chemical compound ClCCl.CCOC(C)=O WBKFWQBXFREOFH-UHFFFAOYSA-N 0.000 claims description 4
- DEQYTNZJHKPYEZ-UHFFFAOYSA-N ethyl acetate;heptane Chemical compound CCOC(C)=O.CCCCCCC DEQYTNZJHKPYEZ-UHFFFAOYSA-N 0.000 claims description 4
- 238000005086 pumping Methods 0.000 claims description 2
- ZMQBBPRAZLACCW-UHFFFAOYSA-N acetic acid;dichloromethane Chemical compound ClCCl.CC(O)=O ZMQBBPRAZLACCW-UHFFFAOYSA-N 0.000 claims 1
- MVFGXYPEQHIKIX-UHFFFAOYSA-N acetic acid;heptane Chemical compound CC(O)=O.CCCCCCC MVFGXYPEQHIKIX-UHFFFAOYSA-N 0.000 claims 1
- 150000002148 esters Chemical class 0.000 claims 1
- 125000004494 ethyl ester group Chemical group 0.000 claims 1
- 238000010438 heat treatment Methods 0.000 claims 1
- 239000007788 liquid Substances 0.000 claims 1
- 238000004519 manufacturing process Methods 0.000 abstract description 10
- 239000000284 extract Substances 0.000 abstract description 6
- 238000004090 dissolution Methods 0.000 abstract description 3
- 239000000356 contaminant Substances 0.000 abstract description 2
- WVDDGKGOMKODPV-UHFFFAOYSA-N Benzyl alcohol Chemical compound OCC1=CC=CC=C1 WVDDGKGOMKODPV-UHFFFAOYSA-N 0.000 abstract 1
- 229960004217 benzyl alcohol Drugs 0.000 abstract 1
- 238000002386 leaching Methods 0.000 abstract 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 14
- 241001116500 Taxus Species 0.000 description 10
- 235000016408 Podocarpus macrophyllus Nutrition 0.000 description 8
- 244000162450 Taxus cuspidata Species 0.000 description 8
- 235000009065 Taxus cuspidata Nutrition 0.000 description 8
- 241001149649 Taxus wallichiana var. chinensis Species 0.000 description 8
- 238000000605 extraction Methods 0.000 description 8
- 239000012535 impurity Substances 0.000 description 6
- 230000000694 effects Effects 0.000 description 5
- 238000002425 crystallisation Methods 0.000 description 4
- 230000008025 crystallization Effects 0.000 description 4
- 206010054949 Metaplasia Diseases 0.000 description 3
- 206010028980 Neoplasm Diseases 0.000 description 3
- 229940123237 Taxane Drugs 0.000 description 3
- 201000011510 cancer Diseases 0.000 description 3
- 230000015689 metaplastic ossification Effects 0.000 description 3
- 239000002904 solvent Substances 0.000 description 3
- 241000196324 Embryophyta Species 0.000 description 2
- -1 acetyl paclitaxel Chemical compound 0.000 description 2
- 239000002246 antineoplastic agent Substances 0.000 description 2
- 229940041181 antineoplastic drug Drugs 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 2
- 239000006227 byproduct Substances 0.000 description 2
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- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 208000003174 Brain Neoplasms Diseases 0.000 description 1
- 206010009944 Colon cancer Diseases 0.000 description 1
- 208000001333 Colorectal Neoplasms Diseases 0.000 description 1
- 244000050510 Cunninghamia lanceolata Species 0.000 description 1
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- 240000001417 Vigna umbellata Species 0.000 description 1
- 125000000218 acetic acid group Chemical group C(C)(=O)* 0.000 description 1
- 150000001335 aliphatic alkanes Chemical class 0.000 description 1
- 230000001093 anti-cancer Effects 0.000 description 1
- 238000009835 boiling Methods 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
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- 238000006731 degradation reaction Methods 0.000 description 1
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- 201000003911 head and neck carcinoma Diseases 0.000 description 1
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- 239000012046 mixed solvent Substances 0.000 description 1
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- 238000002360 preparation method Methods 0.000 description 1
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- 238000011069 regeneration method Methods 0.000 description 1
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- Medicines Containing Plant Substances (AREA)
- Epoxy Compounds (AREA)
Abstract
The invention discloses a kind of methods for extracting natural 10- deacetylate taxol; this method is using branches and leaves of yew as raw material; it is extracted with industrial grade benzenemethanol; concentration leaching liquor obtains medicinal extract; the acetone freezing precipitation of addition isodose after medicinal extract is dissolved with methanol; supernatant after precipitating is concentrated drain after with acetonitrile dissolution filter; concentration is drained filtrate and is obtained containing 10- deacetylate taxol concentrate; concentrate adds n-butanol or isoamyl alcohol to dissolve; add water extraction primary; separate organic phase, be concentrated under reduced pressure drain content >=1% 10- deacetylate taxol first product；Its taxol first product chromatographs to obtain 10- deacetylate taxol crude product by an acidic alumina column; its content is greater than 25%; 25% crude product obtains content and is greater than 98% using primary column chromatography and primary recrystallization, 10- deacetylate taxol finished product of the single contaminant less than 0.2%.The method of the present invention is simple to operation, extracted in production process sufficiently, yield it is higher, it is at low cost, be suitable for industrial applications and marketing.
Description
Technical field
The present invention relates to the extraction field of natural organic-compound, specially one kind is extracted 10- from branches and leaves of yew and is gone
The method of acetyl paclitaxel (10-DAT).
Background technique
10- deacetylate taxol (10-DAT) is a kind of natural organic-compound extracted from Chinese yew, it is right
Cancer has certain effect, still, because of various reasons its structure and taxanes seemingly, commonly used in taxol biosynthesis and more
Western taxol etc..
Taxol is aobvious to treatment oophoroma, lung cancer, colorectal cancer, melanoma, head-neck carcinoma, lymthoma, brain tumor and other effects
It writes.Early in Food and Drug Adminstration of the US (FDA) just approval listing in 1992.For a long time, taxol is the tree from Chinese yew
It is extracted in skin, due to the bark category non-renewable resources of Chinese yew, causes the acquisition raw material of taxol extremely limited, thus made
It is standby at high cost, it is expensive.In addition, Chinese yew is universally acknowledged endangered natural rare plants for anticancer, in natural item
The speed of growth is slow under part, and power of regeneration is poor, only using bark of Ramulus et folium taxi cuspidatae as Japanese yew alcohol extracting raw material, it will seriously destroy red bean
China fir plant resources.
Currently, global cancer prevention and control work has obtained remarkable progress, treatment of cancer is greatly taken on a new look, and anticancer drug constantly gushes
It is existing.And taxol is usually extracted from bark of Ramulus et folium taxi cuspidatae as the most effective natural antitumor drug having now been found that, seriously
Destroy Chinese yew resource；The technique that taxol is extracted in part from branches and leaves of yew is not perfect, that there is yields is low,
The disadvantages of production cycle is long, so taxol price is relatively high；With Chinese yew while extracting isolation of taxol, extract pure
Change 10- deacetylate taxol (10-DAT)；Make full use of chemical means synthesis purple 10- deacetylate taxol (10-DAT)
China fir alkanes drug sufficiently, rationally utilizes existing taxus resource；Reduce taxanes bulk pharmaceutical chemicals costs, effective protection plant
Resource.
There are the following problems for 10- deacetylate purple alcohol (10-DAT) extraction in the prior art or conventional method:
(1) most of to extract taxol currently, about 10- deacetylate purple alcohol (10-DAT) report is extracted and with less
During do not utilize preferably, or as waste disposal；Due to the bark category non-renewable resources of Chinese yew, lead to Japanese yew
The acquisition raw material of alcohol is extremely limited, thus preparation cost is high, expensive；And taxol by-product 10- mentioned by the present invention
Deacetylate purple alcohol (10-DAT) does not utilize preferably and exploitation；
(2) some literature reports slave branches and leaves of yew in extract 10- deacetylate purple alcohol (10-DAT) method, there are yields
Problem low, the production cycle is long；
(3) prior art generallys use multiple column chromatography and selects different fillers, and during the extraction process, 10- deacetylate is purple
Alcohol (10-DAT) is by degradation and destroys, and there is a situation where that yield is low, impurity is more, influences quality, and increased costs；
(4) traditional technique is enriched to 10 contents from 3/10000ths or so content, needs through at least 3 column layers
Analysis, processing cost are higher.
Summary of the invention
In view of the deficiencies of the prior art, the present invention provides one kind extracts natural 10- deacetylate from branches and leaves of yew
The method of purple alcohol (10-DAT), this method is simple and easy, with short production cycle, extraction conditions require simple, high income；This method is used
Methanol impregnates branches and leaves of yew, acetone precipitation, acetonitrile crystallization treatment, and filtrate drains object n-butanol or isoamyl alcohol dissolution
Water extraction drains object after acidic alumina column column and chromatographs to obtain crude product, and crude product carries out a silica gel column chromatography again, and principal piece object is concentrated
Matter is simultaneously drained, and obtains semi-finished product with acetone and normal heptane crystallization are primary；Semi-finished product are carried out with steaming chloroform single-phase solvent again primary cold again
Freeze recrystallization to handle up to 10- deacetylate taxol finished product, content is greater than 98%, maximum single contaminant less than 0.2%.
The purpose of the present invention is achieved through the following technical solutions:
(1) using drying, crushing branches and leaves of yew as raw material, in mass volume ratio g:mL be 1:3~5 ratio add in the feed
Enter methanol, impregnate 6~8h, filtering at room temperature, filter residue continues after adding methanol to impregnate 1~2 time repeatedly, merging filtrate；It will merge
Filtrate afterwards is concentrated at 65~75 DEG C drains to paste, and the isometric acetone of lotion is then added and stirs and evenly mixs, and -5 DEG C~0 DEG C
Lower freezing precipitation 12~for 24 hours, the supernatant after taking precipitating is concentrated and dried at 60~65 DEG C；
(2) step (1) concentrate is dissolved with acetonitrile, refrigeration stands 10~14h, filtering, and pumping is concentrated in filtrate at 50~60 DEG C
It is dry, obtain 10- deacetylate taxol concentrate；
The acetonitrile solution temperature is 35~45 DEG C；
(3) 10- deacetylate taxol concentrate n-butanol or isoamyl alcohol are dissolved, add water extraction primary, collects organic phase,
Organic phase is concentrated under reduced pressure under 50~60 DEG C, vacuum condition, obtains the 10- deacetylate taxol first product of content >=1%；
(4) 10- deacetylate taxol first product is dissolved with methylene chloride, is filtered after dissolved clarification, filtrate is added to acidic alumina
Column chromatography is carried out in column, material glue specific mass ratio (10- deacetylate taxol first product: acidic alumina) is 1:3~4；Use dichloro
Methane-acetone mixture or dichloromethane-ethyl acetate mixed liquor are eluted, and TLC monitors eluate, and collection is gone containing 10-
The efflux of acetyl paclitaxel is concentrated at 45~50 DEG C, obtains the 10- deacetylate taxol crude product that content is greater than 25%；
The volume ratio of methylene chloride and acetone is 4~5:1 in the dichloromethane-acetone mixed liquor；Dichloromethane-ethyl acetate
The volume ratio of methylene chloride and ethyl acetate is 3~4:1 in mixed liquor；
(5) after 10- deacetylate taxol crude product being dissolved with methylene chloride, after spice glue is dry, loading to silicagel column middle layer
Analysis is eluted with normal heptane-acetone mixture or normal heptane-ethyl acetate mixtures, monitors eluate with TLC, collection contains 10-
The eluent of deacetylate taxol merges eluent concentration, obtains the concentrate that 10- deacetylate content of taxol is greater than 60%；
The silicagel column uses the technical grade silica gel of 200-300 mesh；
The volume ratio of normal heptane and acetone is 1:4~5 in the normal heptane-acetone mixture；Normal heptane-ethyl acetate mixtures
The volume ratio of middle normal heptane and ethyl acetate is 1:2~3；
10- deacetylate taxol crude product: the mass ratio of silica gel is 1:18~20；
(6) by step (5) concentrate acetone solution, normal heptane or n-hexane are added under stirring, until there is crystal analysis
Until out, 8~12h, filtering are stood, filter residue and drying obtains 10- deacetylate taxol semi-finished product；
(7) after 10- deacetylate taxol semi-finished product heat (45~50 DEG C) dissolutions with chloroform, filtering is placed in -5~0 DEG C
Under, freezing and crystallizing 20~for 24 hours, filtering, for crystal with chloroform dissolving-recrystallization 1 time, filtration drying is greater than 98% 10- up to content
Deacetylate taxol finished product.
The present invention compared with the existing technology the advantages of and technical effect:
1, present invention selection carries out deep processing to by-product, obtains a kind of synthesizing taxone during just mentioning taxol
Intermediate 10- deacetylate taxol, make full use of and save limited resources, for Chinese yew process save the cost,
2, the present invention has selected acetonitrile to medicinal extract processing, other taxanes can quickly be divided with 10- deacetylate taxol
From, save period and cost for later period process, method is simple, it is with short production cycle, save cost, be very suitable for industrial metaplasia
It produces；
3, the present invention is easy the characteristics of dissolving with 10- deacetylate taxol, adds water using n-butanol or isoamyl alcohol and water slightly soluble
Extraction, can be effectively retained 10- deacetylate taxol, remove a large amount of water-solubility impurities, and method is simple, with short production cycle, section
Cost-saving is very suitable for industrialized production；
4, the present invention has selected acidic alumina as first time column layer, can efficiently separate object, and prevents 10- deacetylate
Taxol is degraded in other fillers and difference is to change, using acidic alumina except the obvious method of impurity effect is simple, high income, section
Cost-saving is very suitable for industrialized production；
5, the present invention selects normal heptane and acetone or ethyl acetate as tomographic system, can preferably separate main matter, removal of impurities
Effect is obvious；2 kinds of mixed solvent boiling point differences are larger, can be very separated in concentration process；From quality and cost consideration, it is suitble to work
Industry metaplasia produces；
6, the present invention (is steamed) again using chloroform, carries out single solvent crystallization, and recrystallisation solvent dosage is small, and using the side of freezing and crystallizing
Formula, crystallization effect is good, and removal of impurities is obvious, high income, can effectively increase taxol to greatest extent by Control of Impurities within 0.2%
Product quality is very suitable for industrialized production；
Whole process of the present invention only needs to carry out 2 column chromatographies and 2 recrystallizations, the 10- of content 3/10000ths can be removed acetyl
For base Purification of Taxol to 98% or more, method is simple, technique is controllable, with short production cycle, saving cost, is very suitable for industry
Metaplasia produces and marketing.
Specific embodiment
Below by embodiment, invention is further described in detail, but the scope of the present invention is not limited in described
Hold.
Embodiment 1: the method for this extraction natural Japanese yew alcohol is as follows:
(1) dry using 500kg, crushing branches and leaves of yew is raw material, in the feed in the ratio that mass volume ratio g:mL is 1:3
Methanol is added, impregnates 6h at room temperature, filters, filter residue continues after adding methanol to impregnate 1 time repeatedly, merging filtrate；After merging
Filtrate is concentrated at 65 DEG C drains to paste, and the isometric acetone of lotion is then added and stirs and evenly mixs, freezing precipitation at -5 DEG C
12h, the supernatant after taking precipitating are concentrated and dried at 60 DEG C；
(2) step (1) concentrate acetonitrile to be dissolved at 45 DEG C, refrigeration stands 10h, filtering, and filtrate is concentrated at 50 DEG C and drains,
Obtain 10- deacetylate taxol concentrate；
(3) by 10- deacetylate taxol concentrate n-Butanol soluble, add water extraction primary, collect organic phase, organic phase exists
50 DEG C, be concentrated under reduced pressure under vacuum condition, obtain the 10- deacetylate taxol first product of content 1.5%；
(4) 10- deacetylate taxol first product is dissolved with methylene chloride, is filtered after dissolved clarification, filtrate is added to acidic alumina
Column chromatography is carried out in column, material glue specific mass ratio is 1:3；With the dichloromethane-acetone mixed liquor (volume ratio of methylene chloride and acetone
It being eluted for 4:1), TLC monitors eluate, and the efflux containing 10- deacetylate taxol is collected, is concentrated at 45 DEG C,
Obtain the 10- deacetylate taxol crude product of content 27%；
It (5) is 1:1.3's by the mass ratio of crude product and silica gel after dissolving 10- deacetylate taxol crude product with methylene chloride
Ratio spice glue after 45 DEG C of dry 4h, chromatographs, 10- deacetylate Japanese yew in loading to silica gel (the technical grade silica gel of 200 mesh) column
Alcohol crude product: the mass ratio of silica gel is 1:18, is eluted with normal heptane-acetone mixture (volume ratio of normal heptane and acetone is 1:4),
Eluate is monitored with TLC, collects the eluent containing 10- deacetylate taxol, merges and is concentrated in vacuo at 50 DEG C of eluent, obtained
The 10- deacetylate taxol concentrate of content 65%；
(6) step (5) concentrate is dissolved at 40 DEG C of acetone, normal heptane is added under stirring, until there is crystal precipitation
Until, 8h, filtering are stood, filter residue and drying obtains 10- deacetylate taxol semi-finished product；
(7) after 10- deacetylate taxol semi-finished product chloroform dissolves at 45 DEG C, filtering is placed at 0 DEG C, freezing and crystallizing
For 24 hours, it filters, crystal is with chloroform dissolving-recrystallization 1 time, and filtering crystals are up to 10- deacetylate taxol finished product, content 98.4%.
Embodiment 2: the method for this extraction natural Japanese yew alcohol is as follows:
(1) dry using 500kg, crushing branches and leaves of yew is raw material, in the feed in the ratio that mass volume ratio g:mL is 1:4
Methanol is added, impregnates 7h at room temperature, filters, filter residue continues after adding methanol to impregnate 2 times repeatedly, merging filtrate；After merging
Filtrate is concentrated at 70 DEG C drains to paste, and the isometric acetone of lotion is then added and stirs and evenly mixs, freezing precipitation at -2 DEG C
15h, the supernatant after taking precipitating are concentrated and dried at 65 DEG C；
(2) step (1) concentrate acetonitrile to be dissolved at 35 DEG C, refrigeration stands 14h, filtering, and filtrate is concentrated at 55 DEG C and drains,
Obtain 10- deacetylate taxol concentrate；
(3) by 10- deacetylate taxol concentrate n-Butanol soluble, add water extraction primary, collect organic phase, organic phase exists
60 DEG C, be concentrated under reduced pressure under vacuum condition, obtain the 10- deacetylate taxol first product of content 1.6%；
(4) 10- deacetylate taxol first product is dissolved with methylene chloride, is filtered after dissolved clarification, filtrate is added to acidic alumina
Column chromatography is carried out in column, material glue specific mass ratio is 1:4；With the dichloromethane-acetone mixed liquor (volume ratio of methylene chloride and acetone
It being eluted for 5:1), TLC monitors eluate, and the efflux containing 10- deacetylate taxol is collected, is concentrated at 50 DEG C,
Obtain the 10- deacetylate taxol crude product of content 26.5%；
It (5) is 1:1.4's by the mass ratio of crude product and silica gel after dissolving 10- deacetylate taxol crude product with methylene chloride
Ratio spice glue after 45 DEG C of dry 5h, chromatographs, 10- deacetylate Japanese yew in loading to silica gel (the technical grade silica gel of 300 mesh) column
Alcohol crude product: the mass ratio of silica gel is 1:20, is eluted with normal heptane-acetone mixture (volume ratio of normal heptane and acetone is 1:5),
Eluate is monitored with TLC, collects the eluent containing 10- deacetylate taxol, merges and is concentrated in vacuo at 60 DEG C of eluent, obtained
The 10- deacetylate taxol concentrate of content 63%；
(6) step (5) concentrate is dissolved at 45 DEG C of acetone, normal heptane is added under stirring, until there is crystal precipitation
Until, 10h, filtering are stood, filter residue and drying obtains 10- deacetylate taxol semi-finished product；
(7) after 10- deacetylate taxol semi-finished product chloroform dissolves at 50 DEG C, filtering is placed at -2 DEG C, freezing and crystallizing
20h, filtering, crystal is with chloroform dissolving-recrystallization 1 time, and filtering crystals are up to 10- deacetylate taxol finished product, content 98.5%.
Embodiment 3: the method for this extraction natural Japanese yew alcohol is as follows:
(1) dry using 500kg, crushing branches and leaves of yew is raw material, in the feed in the ratio that mass volume ratio g:mL is 1:5
Methanol is added, impregnates 8h at room temperature, filters, filter residue continues after adding methanol to impregnate 1 time repeatedly, merging filtrate；After merging
Filtrate is concentrated at 75 DEG C drains to paste, and the isometric acetone of lotion is then added and stirs and evenly mixs, at 0 DEG C freezing precipitation for 24 hours,
Supernatant after taking precipitating is concentrated and dried at 62 DEG C；
(2) step (1) concentrate acetonitrile to be dissolved at 40 DEG C, refrigeration stands 14h, filtering, and filtrate is concentrated at 60 DEG C and drains,
Obtain 10- deacetylate taxol concentrate；
(3) 10- deacetylate taxol concentrate is dissolved with isoamyl alcohol, adds water extraction primary, collect organic phase, organic phase exists
60 DEG C, be concentrated under reduced pressure under vacuum condition, obtain the 10- deacetylate taxol first product of content 1.4%；
(4) 10- deacetylate taxol first product is dissolved with methylene chloride, is filtered after dissolved clarification, filtrate is added to acidic alumina
Column chromatography is carried out in column, material glue specific mass ratio is 1:3；With dichloromethane-ethyl acetate mixed liquor (methylene chloride and ethyl acetate
Volume ratio be 3:1) eluted, TLC monitor eluate, collect the efflux containing 10- deacetylate taxol, 42
It is concentrated at DEG C, obtains the 10- deacetylate taxol crude product of content 25.5%；
It (5) is 1:1.5's by the mass ratio of crude product and silica gel after dissolving 10- deacetylate taxol crude product with methylene chloride
Ratio spice glue after 45 DEG C of dry 6h, chromatographs, 10- deacetylate Japanese yew in loading to silica gel (the technical grade silica gel of 200 mesh) column
Alcohol crude product: the mass ratio of silica gel is 1:19, and with normal heptane-ethyl acetate mixtures, (volume ratio of normal heptane and ethyl acetate is
It 1:2) elutes, monitors eluate with TLC, collect the eluent containing 10- deacetylate taxol, merge true at 50 DEG C of eluent
Sky concentration, obtains the 10- deacetylate taxol concentrate of content 63.5%；
(6) step (5) concentrate is dissolved at 40 DEG C of acetone, n-hexane is added under stirring, until there is crystal precipitation
Until, 12h, filtering are stood, filter residue and drying obtains 10- deacetylate taxol semi-finished product；
(7) after 10- deacetylate taxol semi-finished product chloroform dissolves at 45 DEG C, filtering is placed at -2 DEG C, freezing and crystallizing
22h, filtering, crystal is with chloroform dissolving-recrystallization 1 time, and filtering crystals are up to 10- deacetylate taxol finished product, content 98.7%.
Claims (7)
1. a kind of method for extracting 10- deacetylate taxol, which is characterized in that steps are as follows:
(1) using drying, crushing branches and leaves of yew as raw material, in mass volume ratio g:mL be 1:3~5 ratio add in the feed
Enter methanol, impregnate 6~8h, filtering at room temperature, filter residue continues after adding methanol to impregnate 1~2 time repeatedly, merging filtrate；It will merge
Filtrate afterwards is concentrated at 65~75 DEG C drains to paste, and the isometric acetone of lotion is then added and stirs and evenly mixs, and -5 DEG C~0 DEG C
Lower freezing precipitation 12~24 hours, the supernatant after taking precipitating are concentrated and dried at 60~65 DEG C；
(2) step (1) concentrate is dissolved with acetonitrile, refrigeration stands 10~14h, filtering, and pumping is concentrated in filtrate at 50~60 DEG C
It is dry, obtain 10- deacetylate taxol concentrate；
(3) 10- deacetylate taxol concentrate n-butanol or isoamyl alcohol are dissolved, add water extraction primary, collects organic phase,
Organic phase is concentrated under reduced pressure under 50~60 DEG C, vacuum condition, obtains 10- deacetylate taxol first product；
(4) 10- deacetylate taxol first product is dissolved with methylene chloride, is filtered after dissolved clarification, filtrate is added to acidic alumina
Column chromatography is carried out in column, material glue specific mass ratio is 1:3~4；With dichloromethane-acetone mixed liquor or dichloromethane-ethyl acetate
Mixed liquor is eluted, and TLC monitors eluate, the efflux containing 10- deacetylate taxol is collected, at 45~50 DEG C
Concentration, obtains 10- deacetylate taxol crude product；
(5) after 10- deacetylate taxol crude product being dissolved with methylene chloride, after spice glue is dry, loading to silicagel column middle layer
Analysis is eluted with normal heptane-acetone mixture or normal heptane-ethyl acetate mixtures, monitors eluate with TLC, collection contains 10-
The eluent of deacetylate taxol merges eluent concentration, obtains the concentrate that 10- deacetylate content of taxol is greater than 60%；
(6) by step (5) concentrate acetone solution, normal heptane or n-hexane are added under stirring, until there is crystal analysis
Until out, 8~12h, filtering are stood, filter residue and drying obtains 10- deacetylate taxol semi-finished product；
(7) after 10- deacetylate taxol semi-finished product are dissolved by heating with chloroform, filtering is placed at -5~0 DEG C, freezing and crystallizing
20~for 24 hours, filtering, crystal is with chloroform dissolving-recrystallization 1 time, and filtration drying is up to 10- deacetylate taxol finished product.
2. the method according to claim 1 for extracting 10- deacetylate taxol, it is characterised in that: step (2) acetonitrile is molten
Solving temperature is 35~45 DEG C.
3. the method according to claim 1 for extracting 10- deacetylate taxol, it is characterised in that: dichloromethane-acetone
The volume ratio of methylene chloride and acetone is 4~5:1 in mixed liquor.
4. the method according to claim 1 for extracting 10- deacetylate taxol, it is characterised in that: methylene chloride-acetic acid
The volume ratio of methylene chloride and ethyl acetate is 3~4:1 in ethyl ester mixed liquor.
5. the method according to claim 1 for extracting 10- deacetylate taxol, it is characterised in that: what silicagel column used
It is the technical grade silica gel of 200~300 mesh.
6. the method according to claim 1 for extracting 10- deacetylate taxol, it is characterised in that: normal heptane-acetone is mixed
Closing the volume ratio of normal heptane and acetone in liquid is 1:4~5.
7. the method according to claim 1 for extracting 10- deacetylate taxol, it is characterised in that: normal heptane-acetic acid second
The volume ratio of normal heptane and ethyl acetate is 1:2~3 in ester mixed liquor.
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