CN102219764A - Method for separating and purifying paclitaxel industrially - Google Patents
Method for separating and purifying paclitaxel industrially Download PDFInfo
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- 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 100
- 229960001592 paclitaxel Drugs 0.000 title claims abstract description 98
- 229930012538 Paclitaxel Natural products 0.000 title claims abstract description 96
- 238000000034 method Methods 0.000 title claims abstract description 45
- 238000000926 separation method Methods 0.000 claims abstract description 23
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- NTIZESTWPVYFNL-UHFFFAOYSA-N Methyl isobutyl ketone Chemical compound CC(C)CC(C)=O NTIZESTWPVYFNL-UHFFFAOYSA-N 0.000 claims description 2
- OFBQJSOFQDEBGM-UHFFFAOYSA-N Pentane Chemical compound CCCCC OFBQJSOFQDEBGM-UHFFFAOYSA-N 0.000 claims description 2
- 229950005499 carbon tetrachloride Drugs 0.000 claims description 2
- HPNMFZURTQLUMO-UHFFFAOYSA-N diethylamine Chemical compound CCNCC HPNMFZURTQLUMO-UHFFFAOYSA-N 0.000 claims description 2
- VZGDMQKNWNREIO-UHFFFAOYSA-N tetrachloromethane Chemical compound ClC(Cl)(Cl)Cl VZGDMQKNWNREIO-UHFFFAOYSA-N 0.000 claims description 2
- DBXFAPJCZABTDR-KUEXGRMWSA-N Cephalomannine Natural products O=C(O[C@@H]1C(C)=C2[C@@H](OC(=O)C)C(=O)[C@]3(C)[C@@H](O)C[C@@H]4[C@](OC(=O)C)([C@H]3[C@H](OC(=O)c3ccccc3)[C@@](O)(C2(C)C)C1)CO4)[C@@H](O)[C@H](NC(=O)/C(=C\C)/C)c1ccccc1 DBXFAPJCZABTDR-KUEXGRMWSA-N 0.000 abstract description 19
- DBXFAPJCZABTDR-WBYYIXQISA-N cephalomannine Chemical compound O([C@@H]1[C@]2(O)C[C@@H](C(=C([C@@H](OC(C)=O)C(=O)[C@]3(C)[C@@H](O)C[C@H]4OC[C@]4([C@H]31)OC(C)=O)C2(C)C)C)OC(=O)[C@H](O)[C@@H](NC(=O)C(/C)=C/C)C=1C=CC=CC=1)C(=O)C1=CC=CC=C1 DBXFAPJCZABTDR-WBYYIXQISA-N 0.000 abstract description 19
- 239000002994 raw material Substances 0.000 abstract description 15
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 abstract description 13
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- OVMSOCFBDVBLFW-VHLOTGQHSA-N 5beta,20-epoxy-1,7beta,13alpha-trihydroxy-9-oxotax-11-ene-2alpha,4alpha,10beta-triyl 4,10-diacetate 2-benzoate Chemical compound O([C@@H]1[C@@]2(C[C@H](O)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)O)C(=O)C1=CC=CC=C1 OVMSOCFBDVBLFW-VHLOTGQHSA-N 0.000 description 4
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- BEHTXUBGUDGCNQ-IEAAAIHOSA-N taxol c Chemical compound O([C@@H]1[C@]2(O)C[C@@H](C(=C([C@@H](OC(C)=O)C(=O)[C@]3(C)[C@@H](O)C[C@H]4OC[C@]4([C@H]31)OC(C)=O)C2(C)C)C)OC(=O)[C@H](O)[C@@H](NC(=O)CCCCC)C=1C=CC=CC=1)C(=O)C1=CC=CC=C1 BEHTXUBGUDGCNQ-IEAAAIHOSA-N 0.000 description 4
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- CXURGFRDGROIKG-UHFFFAOYSA-N 3,3-bis(chloromethyl)oxetane Chemical compound ClCC1(CCl)COC1 CXURGFRDGROIKG-UHFFFAOYSA-N 0.000 description 2
- 229940123237 Taxane Drugs 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
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- ZDZOTLJHXYCWBA-VCVYQWHSSA-N N-debenzoyl-N-(tert-butoxycarbonyl)-10-deacetyltaxol Chemical compound O([C@H]1[C@H]2[C@@](C([C@H](O)C3=C(C)[C@@H](OC(=O)[C@H](O)[C@@H](NC(=O)OC(C)(C)C)C=4C=CC=CC=4)C[C@]1(O)C3(C)C)=O)(C)[C@@H](O)C[C@H]1OC[C@]12OC(=O)C)C(=O)C1=CC=CC=C1 ZDZOTLJHXYCWBA-VCVYQWHSSA-N 0.000 description 1
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- Epoxy Compounds (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention discloses a method for separating and purifying paclitaxel industrially and efficiently, and belongs to the field of the separation and purification of compounds. In the method, the paclitaxel is separated by column chromatography, namely the paclitaxel is separated and purified by taking a combination of any three or four of compounds of alkane having 1 to 10 carbon atoms, chlorohydrocarbons, alcohols having 1 to 4 carbon atoms, saturated alkane esters and alkane ketones, and amine compounds as a mobile phase and 200 to 300-mesh ordinary silica gel as a stationary phase by normal phase column chromatography. By selecting a multielement system mixed solvent as the mobile phase, the separation degree of substances which are difficult to separate is increased, and the problem that the paclitaxel cannot be separated from cephalomannine effectively in the conventional separation method is solved. After the paclitaxel is separated and purified by the column chromatography, the purity of paclitaxel crude products serving as raw materials can be improved to be over 98 percent, and according to the condition of the raw materials, the total yield is up to 90 percent only by eluting the paclitaxel crude products once to twice. In the method, the common solvent and silica gel are used, so the cost is low, the method is easy and convenient to operate, the sample volume is large, and industrial production is easy to realize.
Description
Technical field
The present invention relates to the separation purification method of taxol, be specifically related to the method for a kind of industrial high efficiency separating and purifying taxol and Cephalomannine, belong to compound separation purifying field.
Background technology
Taxol is a kind of new type anticancer medicine with Taxan diterpene skeleton of separation and Extraction from Chinese yew genus plants, also is unique a kind of microtubule polymerization and stable medicine of polymerization microtubule of promoting of being understood at present.It has powerful restraining effect to most of solid tumors, and to normal this nothing of cell based influence, especially less to determined curative effect, the side effect of advanced ovarian cancer, mammary cancer, nonsmall-cell lung cancer and Kaposi.Along with further going deep into of research, the antitumous effect of taxanes medicine more and more is more widely used, and has now become the antitumor drug of a line cancer therapy drug and wide spectrum.
Ramulus et folium taxi cuspidatae class plant as the taxol main source belongs to rare species, is listed in the first-grade state protection plant in China.Content of taxol is extremely low in the bark of Ramulus et folium taxi cuspidatae, accounts for 0.01%, can not satisfy the demand of market to taxol far away.Over nearly 20 years, the preparation method of taxol has multidisciplinary in succession, with multi-angle the exploration report.Taxol cell cultures, fungi fermentation, the semi-synthetic and total synthesis method of chemistry are arranged.No matter take where legal system is equipped with taxol, purifying comes out all to need to separate taxol also from paclitaxel analogs.Common paclitaxel analogs mainly contains Cephalomannine, baccatin III, 10-deacetylate taxol, 7-table-taxol, taxol C etc.Because structure is more close with character, thereby causes taxol very difficult with separating of its analogue.In addition, as a kind of biomacromolecule material, taxol is subjected to the influence of envrionment conditionss such as temperature, organic solvent, acid, alkali, degraded or isomery take place easily and generates other taxanes materials.For example taxol can be degraded to baccatin III or isomery takes place and generate 7-table-taxol under strongly-acid or weak basic condition; DeR also can take place in taxol when temperature was higher, generated corresponding small-molecule substance.As the medicine in the clinical use, taxol its purity requirement in actual applications is very high.Taxol resource preciousness, hold at high price, isolating purity and yield become the key factor of restriction pharmaceutical paclitaxel cost.Based on above-mentioned several aspects, make and develop more efficient, cheap industrial separation purification technique and novel production separation purifying technique, become the focal issue in the taxol research.
The separating and purifying taxol commonly used at present and the method for paclitaxel analogs mainly contain column chromatography, tlc, the precipitator method, miccellar electrokinetic chromatography method, membrane separation process, resin absorption partition method, chemical reaction method etc.But what be applicable to large-scale industrialization production only has column chromatography a kind of, and additive method all is confined to laboratory study, and column chromatography is divided into normal phase column method and two kinds of methods of reversed-phase column again.
The patent application of relevant taxol separation and purification has multinomial: as Chinese patent CN 1377882A " a kind of combined preparation high-purity taxol, Cephalomannine, the method for 10-deacetylation Bakating III "; Chinese patent CN 1611496A " a kind of method of utilizing high pressure liquid chromatography to prepare high-purity taxol "; Chinese patent CN 101391989A " a kind of method for preparing polyhydroxy taxone and taxol ".
The weak point of existing taxol separation purification method mainly is: the method for present various isolation of taxol is isolation of taxol and Cephalomannine effectively still; With regard to can the heavy industrialization separating and purifying taxol and the column chromatography of paclitaxel analogs with regard to, there are defective again separately in normal phase method and reversed phase method.The normal phase column chromatography often needs to introduce oxidizing reaction, by changing the chemical structure of Cephalomannine, to increase the resolution of Cephalomannine and taxol, this has not only changed the chemical structure of separate object, and to repeatedly cross post, repeatedly recrystallization carries out purifying.The loaded down with trivial details yield that not only reduces the target taxol of operation steps increases production cost, and separating resulting only can obtain a kind of product of taxol, and the Cephalomannine that changes after the structure can only be abandoned as waste material, has not only wasted resource but also contaminate environment.Though the reversed phase column chromatography method need not change the chemical structure of separate object, in the practical application, this method need be used special reversed-phase column silica gel, and its market value is high, reaches 500,000 yuan/kilogram.In the suitability for industrialized production, per minute can be used 50 kilograms reversed-phase column silica gel at least from 1 kilogram of taxol, and only the cost of one in reversed-phase column silica gel will 2,500 ten thousand yuan, has increased the production cost of isolation of taxol greatly, has limited its large-scale industrialization and has produced.
Summary of the invention
The object of the present invention is to provide a kind of separation efficiency height, with low cost, the taxol separation purification method that is easy to industrialization operation.
Be to realize the object of the invention, adopt the normal phase column chromatography, polynary by selecting for use (quaternary or five yuan) system mixed solvent carries out separation and purification as moving phase to the thick product of taxol.Moving phase is the combination of any three kinds or four kinds and aminated compounds in C1-10 alkanes, chlorinated hydrocarbons, C1-4 alcohols, saturated alkane ester class, the alkane ketone compounds.
Described amine solvent is selected from diethylamine or triethylamine; The C1-10 alkane solvents is selected from normal hexane, Skellysolve A or hexanaphthene; The chlorinated hydrocarbons solvent is selected from methylene dichloride, trichloromethane or tetrachloromethane; Described C1-4 alcoholic solvent is selected from methyl alcohol or ethanol; Described saturated alkane esters solvent is selected from ethyl acetate; Described alkane ketone kind solvent is selected from acetone or isopropyl acetone.The consumption of amine solvent accounts for 8%～16% of mobile phase volume.
Usually through one to the twice chromatographic post wash-out the taxol of purity 93%～95% account for 80%～90%; Through one to the twice chromatographic post wash-out the taxol of purity 30%～90% account for 5%～15%; Through one to the twice chromatographic post wash-out purity be lower than 30% taxol and account for 1%～3%; Sample carries out recrystallization one time, gets the target taxol.
In the practical application, the content of raw material taxol is high more, and the wash-out number of times of required chromatography column is few more.When adopting present method isolation of taxol, be higher than 70%, then only need a chromatography column wash-out just can obtain the ideal separating effect as the raw material content of taxol; As the raw material content of taxol is 50%～70%, and twice chromatographic post wash-out just can obtain the ideal separating effect.
The selection of system moving phase that innovative point of the present invention is is polynary (quaternary or five yuan), make and saved chemical reaction step such as bromination method in the separation and purification process, avoided the change of chemical structure in the separate object, reduced the post number of times excessively of material sample, only crossed a chromatography column and just the taxol more than 80% can be separated.Simultaneously, separating resulting obtains highly purified taxol and two kinds of materials of Cephalomannine.Cephalomannine also can be made into Docetaxel or taxol except the bulk drug that itself can be used as high added value directly utilizes (150,000 yuan/kilogram), obtain more economic benefit.
Innovative point of the present invention is that moving phase selects the combination of any three kinds or four kinds and aminated compounds in C1-10 alkanes, chlorinated hydrocarbons, C1-4 alcohols, saturated alkane ester class, the alkane ketone compounds for use; Stationary phase is common 200 orders～300 order normal phase column silica gel of 50 yuan/Kg.In the separation and purification process material therefor be simple and easy to, material with low cost, be easy to carry out suitability for industrialized production and reduce production costs.Compare with reversed-phase column isolation of taxol method at present commonly used, effectively isolation of taxol and Cephalomannine, mistake post number of times is few, the separation efficiency height, low production cost can be carried out suitability for industrialized production on a large scale.
Innovative point of the present invention is the application of amine reagent, regulated the pH value of moving phase, sepn process is carried out under alkaline condition, this moment, separated material all existed with molecular form, reduced the absorption on silica gel, and improved the resolution of difficult separate substance, solved in the various in the past separation methods the effectively difficult problem of isolation of taxol and Cephalomannine well taxol and Cephalomannine.
Innovative point of the present invention is in the elutriant of chromatography column, and the liquid that contains Cephalomannine flows out earlier, flows out after containing the liquid of taxol.This use with polynary (quaternary or five yuan) system moving phase and amine reagent is relevant, and alkaline isolating environment makes Cephalomannine flow out prior to taxol.
Adopt the inventive method effectively isolation of taxol and Cephalomannine, separation and purification is effective, can easily the thick product purity of taxol be increased to more than 98%, according to raw material condition, only once can be up to 90% to twice wash-out total recovery.With existing taxol separation method ratio, it is few, easy and simple to handle that present method is crossed post wash-out number of times, safety, and the separation efficiency height, with low cost, applied sample amount is big, is easy to industrialization and carries out scale operation.
Embodiment
For the present invention is illustrated better, embodiment is as follows:
Embodiment 1
The sample of getting content of taxol 70% is a raw material, and raw material and silica gel weight ratio are 2:100.Raw material is standby with methylene dichloride dissolving back.Quaternary system moving phase is normal hexane: methylene dichloride: methyl alcohol: triethylamine volume ratio=3:6:2:2,200 orders～300 order silica gel are as stationary phase.Adopt the wet method upper prop, adopt the wet method upper prop, behind the last sample, behind a column volume, separate with the moving phase for preparing again with dichloromethane solvent.After moving phase is dashed 1.4 column volumes, begin to have taxol C to flow out, and then flow out for Cephalomannine, behind 0.3 column volume, for single taxol flows out, taxol can be separated fully behind 2 column volumes again, and the taxol of purity more than 90% accounts for 85% behind the wash-out.Merge the taxol effluent liquid of purity 30%～90% and carry out the secondary wash-out.(sepn process can be monitored with TLC or HPLC) merges the taxol effluent liquid of twice elutriant moderate purity more than 90%, under 45 ℃, carry out vacuum concentration, enriched material at room temperature (25 ℃) is used acetone: normal hexane=1:6 volume ratio is carried out recrystallization, there are a large amount of white precipitates to separate out after stirring 10min, after stirring 20min, taxol can be separated out fully.After carrying out suction filtration and vacuum-drying, obtain purity and reach 98.6% pure product of paclitaxel, the taxol total recovery reaches 97.6%.
Embodiment 2
The sample of getting content of taxol 60% is a raw material, and raw material and silica gel weight ratio are 1.5:100.Raw material is standby with methylene dichloride dissolving back.Penton is that moving phase is normal hexane: methylene dichloride: acetone: triethylamine: ethyl acetate volume ratio=6:3:1:1:1.5,200 orders～300 order silica gel are as stationary phase.Adopt the wet method upper prop, behind the last sample, behind a column volume, separate with the moving phase for preparing again with dichloromethane solvent.After moving phase is dashed 1.7 column volumes, begin to have taxol C to flow out, and then flow out for Cephalomannine, behind 0.5 column volume, for single taxol flows out, taxol can be separated fully behind 2 column volumes again, and the taxol of purity more than 90% accounts for 75% behind the wash-out.Merge the taxol effluent liquid of purity 30%～90% and carry out the secondary wash-out.(sepn process can be monitored with TLC or HPLC) merges the taxol effluent liquid of twice elutriant moderate purity more than 90%, the effluent liquid that (sepn process can be monitored with TLC or HPLC) contains taxol carries out vacuum concentration under 45 ℃, enriched material at room temperature (25 ℃) is used acetone: normal hexane=1:6 volume ratio is carried out recrystallization, there are a large amount of white precipitates to separate out after stirring 10min, after stirring 20min, taxol can be separated out fully.After carrying out suction filtration and vacuum-drying, obtain purity and reach 98.5% pure product of paclitaxel, the taxol total recovery reaches 94.6%.
Embodiment 3
The sample of getting content of taxol 50% is a raw material, and raw material and silica gel weight ratio are 2:100.Raw material is standby with trichloromethane dissolving back.Penton is that moving phase is normal hexane: trichloromethane: acetone: diethylamine: ethyl acetate volume ratio=4:3:1:1:1.5,200 orders～300 order silica gel are as stationary phase.Adopt the wet method upper prop, behind the last sample, behind 2 column volumes, separate with the moving phase for preparing again with the trichloromethane solvent.After moving phase is dashed 2 column volumes, begin to have taxol C to flow out, and then flow out for Cephalomannine, behind 0.5 column volume, for single taxol flows out, taxol can be separated fully behind 2.5 column volumes again, and the taxol of purity more than 90% accounts for 66.7% behind the wash-out.Merge the taxol effluent liquid of purity 30%～90% and carry out the secondary wash-out.The effluent liquid that (sepn process can be monitored with TLC or HPLC) contains taxol carries out vacuum concentration under 45 ℃, enriched material at room temperature (25 ℃) is used acetone: normal hexane=1:6 volume ratio is carried out recrystallization, there are a large amount of white precipitates to separate out after stirring 10min, after stirring 20min, taxol can be separated out fully, after carrying out suction filtration and vacuum-drying, obtain purity and reach 98.3% pure product of paclitaxel, the taxol total recovery reaches 90.7%.
Claims (3)
1. the method for an industrial separation purification of paclitaxel, adopt normal phase column chromatography isolation of taxol, it is characterized in that moving phase is the combination of any three kinds or four kinds and aminated compounds in C1-10 alkanes, chlorinated hydrocarbons, C1-4 alcohols, saturated alkane ester class, the alkane ketone compounds.
2. in the separating and purifying taxol method as claimed in claim 1, it is characterized in that aminated compounds is selected from diethylamine or triethylamine; The C1-10 alkane derivative is selected from normal hexane, Skellysolve A or hexanaphthene; The chlorinated hydrocarbons compound is selected from methylene dichloride, trichloromethane or tetrachloromethane; The C1-4 alcohol compound is selected from methyl alcohol or ethanol; The saturated alkane ester compound is selected from ethyl acetate; The alkane ketone compounds is selected from acetone or isopropyl acetone.
3. in the separating and purifying taxol method as claimed in claim 1 or 2, it is characterized in that the consumption of aminated compounds accounts for 8%～16% of mobile phase volume.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 201110121650 CN102219764B (en) | 2011-05-12 | 2011-05-12 | Method for separating and purifying paclitaxel industrially |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 201110121650 CN102219764B (en) | 2011-05-12 | 2011-05-12 | Method for separating and purifying paclitaxel industrially |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102219764A true CN102219764A (en) | 2011-10-19 |
| CN102219764B CN102219764B (en) | 2013-06-26 |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102417492A (en) * | 2011-12-07 | 2012-04-18 | 福建紫杉园生物有限公司 | Method for separating and purifying paclitaxel |
| CN103232416A (en) * | 2013-04-22 | 2013-08-07 | 无锡尔云科技有限公司 | Method for separating and purifying 10-deacetylated paclitaxel (10-DAP) |
| CN105457337A (en) * | 2015-11-17 | 2016-04-06 | 重庆臻源红豆杉发展有限公司 | A silica gel loading and sample loading method of a chromatographic column |
| CN109384749A (en) * | 2018-12-26 | 2019-02-26 | 重庆市碚圣医药科技股份有限公司 | A kind of purification process of taxol |
| CN112645906A (en) * | 2020-12-29 | 2021-04-13 | 重庆臻源红豆杉发展有限公司 | Method for separating and purifying paclitaxel by high-efficiency chromatography |
| CN113444060A (en) * | 2021-08-09 | 2021-09-28 | 怀化市盛德生物科技有限责任公司 | Paclitaxel separation and purification process |
| CN113717131A (en) * | 2021-08-27 | 2021-11-30 | 常熟纳微生物科技有限公司 | Separation and purification method of paclitaxel |
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Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102417492A (en) * | 2011-12-07 | 2012-04-18 | 福建紫杉园生物有限公司 | Method for separating and purifying paclitaxel |
| CN103232416A (en) * | 2013-04-22 | 2013-08-07 | 无锡尔云科技有限公司 | Method for separating and purifying 10-deacetylated paclitaxel (10-DAP) |
| CN103232416B (en) * | 2013-04-22 | 2015-03-18 | 无锡尔云科技有限公司 | Method for separating and purifying 10-deacetylated paclitaxel (10-DAP) |
| CN105457337A (en) * | 2015-11-17 | 2016-04-06 | 重庆臻源红豆杉发展有限公司 | A silica gel loading and sample loading method of a chromatographic column |
| CN109384749A (en) * | 2018-12-26 | 2019-02-26 | 重庆市碚圣医药科技股份有限公司 | A kind of purification process of taxol |
| CN112645906A (en) * | 2020-12-29 | 2021-04-13 | 重庆臻源红豆杉发展有限公司 | Method for separating and purifying paclitaxel by high-efficiency chromatography |
| CN113444060A (en) * | 2021-08-09 | 2021-09-28 | 怀化市盛德生物科技有限责任公司 | Paclitaxel separation and purification process |
| CN113717131A (en) * | 2021-08-27 | 2021-11-30 | 常熟纳微生物科技有限公司 | Separation and purification method of paclitaxel |
| CN113717131B (en) * | 2021-08-27 | 2023-09-22 | 常熟纳微生物科技有限公司 | Separation and purification method of paclitaxel |
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| CN102219764B (en) | 2013-06-26 |
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