KR970061883A - Method for mass-extraction and purification of taxol from taxa plants - Google Patents

Method for mass-extraction and purification of taxol from taxa plants Download PDF

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KR970061883A
KR970061883A KR1019960002621A KR19960002621A KR970061883A KR 970061883 A KR970061883 A KR 970061883A KR 1019960002621 A KR1019960002621 A KR 1019960002621A KR 19960002621 A KR19960002621 A KR 19960002621A KR 970061883 A KR970061883 A KR 970061883A
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taxus
taxol
methanol
extraction
plant
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KR100259396B1 (en
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송봉규
김진현
이현수
홍승서
임창배
김광욱
강인선
박흥복
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김경환
주식회사 삼양제넥스
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Priority to KR1019960002621A priority Critical patent/KR100259396B1/en
Priority to JP8533189A priority patent/JP2965168B2/en
Priority to CNB961904208A priority patent/CN1166780C/en
Priority to PCT/KR1996/000059 priority patent/WO1996034973A1/en
Priority to AU55165/96A priority patent/AU5516596A/en
Priority to EP96912315A priority patent/EP0774010B1/en
Priority to US08/652,493 priority patent/US5900367A/en
Priority to DE69615499T priority patent/DE69615499T2/en
Publication of KR970061883A publication Critical patent/KR970061883A/en
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D305/00—Heterocyclic compounds containing four-membered rings having one oxygen atom as the only ring hetero atoms
    • C07D305/14—Heterocyclic compounds containing four-membered rings having one oxygen atom as the only ring hetero atoms condensed with carbocyclic rings or ring systems
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D11/00—Solvent extraction
    • B01D11/02—Solvent extraction of solids
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    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
    • C12P17/02—Oxygen as only ring hetero atoms

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Abstract

본 발명은 택서스속(Taxusgenus) 식물체로부터 택솔을 대량으로 추출 정제하는 방법에 관한 것이다. 본 발명에 따른 택솔의 대량 추출 정제방법은 택서스속(Taxusgenus) 식물체의 시료를 메탄올로 추출하는 제1용매 추출공정; 전기에서 수득한 메탄올 추출액에 디클로로메탄을 가하여 디클로로메탄 추출액을 수득하는 제2용매 추출공정; 전기에서 수득한 디클로로메탄 추출액을 합성 흡착제로 처리하고 여과시켜 여액을 수득하는 공정; 전기에서 수득한 여액을 핵산에 첨가하여 조 정제 택솔을 침전시키는 공정; 전기에서 수득한 조 정제 택솔을 메탄올과 정제수 중에서 분별침전시켜 택솔 분말을 수득하는 공정; 및, 전기에서 수득한 택솔 분말을 고속 액체 크로마토그래피로 정제하여 정제된 택솔을 얻는 공정을 포함하여 이루어진다. 본 발명에 따른 택솔 대량 추출 정제방법에 의하면, 택서스속 식물체 시료의 수분함량에 관계없이 추출용매로서 메탄올, 디클로로메탄 및 핵산 3종류의 추출용매만을 사용하고 적은 추출용매량으로 단시간내에 약 99% 이상의 고순도를 지닌 결정성 택솔을 고수율로 추출 정제할 수 있으며, 분별 침전시키기 전에 합성 흡착제를 사용하여 타르 등의 불순물을 제거함으로써 분별침전 공정을 대량으로 수행할 수 있을 뿐만 아니라, 고속액체 크로마토그래피의 효율을 증대시킬 수 있었다.The present invention relates to a method of mass-extracting and purifying Taxol from a Taxus genus plant. The method for massive extraction and purification of taxol according to the present invention comprises a first solvent extraction step of extracting a sample of Taxus genus plant with methanol; A second solvent extraction step of adding dichloromethane to the methanol extract obtained in the previous step to obtain a dichloromethane extract; Treating the dichloromethane extract obtained in the previous step with a synthetic adsorbent and filtering to obtain a filtrate; A step of adding the filtrate obtained in the previous step to the nucleic acid to precipitate the crude drug; A step of fractionally precipitating the crude tablets obtained in the above process in methanol and purified water to obtain Taxol powder; And a step of purifying the taxol powder obtained in the foregoing by high performance liquid chromatography to obtain a purified taxol. According to the method for mass extraction and purification of taxol in accordance with the present invention, only methanol, dichloromethane and nucleic acid extraction solvents are used as the extraction solvent regardless of the water content of the taxus plant sample, and 99% The purified taxol having high purity can be extracted and refined at a high yield. In addition, it is possible to carry out the fractional precipitation process in a large amount by removing impurities such as tar by using a synthetic adsorbent before fractional precipitation, It is possible to increase the efficiency of the system.

Description

택서스속 식물체로부터 택솔의 대량 추출 정제방법Method for mass-extraction and purification of taxol from taxa plants

본 내용은 요부공개 건이므로 전문내용을 수록하지 않았음Since this is a trivial issue, I did not include the contents of the text.

Claims (15)

(i) 택서스속(Taxusgenus) 식물체의 시료를 메탄올로 추출하는 제1용매 추출공정; (ii)전기에서 수득한 메탄올 추출액에 디클로로메탄을 가하여 디클로로메탄 추출액을 수득하는 제2용매 추출공정; (iii)전기에서 수득한 디클로로메탄 추출액을 합성 흡착제로 처리하고 여과시켜 여액을 수득하는 공정; (iv) 전기에서 수득한 여액을 핵산에 첨가하여 조 정제 택솔을 침전시키는 공정; (v) 전기에서 수득한 조 정제 택속을 메탄올과 정제수 중에서 분별침전시켜 택솔 분말을 수득하는 공정; 및, (vi)전기에서 수득한 택솔 분말을 고속액체 크로마토그래피로 정제하여 정제된 택솔을 얻는 공정을 포함하는 택서스속 식물체로부터 택솔의 대량 추출 정제방법.(i) a first solvent extraction step of extracting a sample of Taxus genus plant with methanol; (ii) a second solvent extraction step of adding dichloromethane to the methanol extract obtained in the foregoing to obtain a dichloromethane extract; (iii) treating the dichloromethane extract obtained in the previous step with a synthetic adsorbent and filtering to obtain a filtrate; (iv) a step of adding the filtrate obtained in the above process to the nucleic acid to precipitate the crude drug; (v) fractionally separating the crude extract obtained in the previous step in methanol and purified water to obtain Taxol powder; And (vi) purifying taxol powder obtained in the past by high performance liquid chromatography to obtain a purified taxol. 제1항에 있어서, 전기 택서스속 식물체의 시료는 주목의 잎 또는 껍질을 분쇄하고 분말화하여 얻은 시료, 또는 주목의 조직세포를 배양하여 얻은 시료를 사용하는 것을 특징으로 하는 택서스속 식물체로부터 택솔의 대량 추출 정제방법.The method according to claim 1, wherein the sample of the plant in the electrical taxus is a sample obtained by pulverizing and pulverizing a leaf or a shell of interest, or a sample obtained by culturing a main tissue cell. Mass extraction and purification of taxol. 제1항 또는 제2항에 있어서, 전기 택서스속 식물체는 택서스 브레비폴리아(Taxus brevifolia), 택서스 카나덴시스(Taxus canadensis), 택서스 쿠스피다타(Taxus cuspidata), 택서스 바카타(Taxus baccata), 택서스 글로보사(Taxus globosa), 택서스 플로리다나(Taxus floridana), 택서스 월리치아나(Taxus wallichiana), 택서스 메디아(Taxus media) 및 택서스 치넨시스(Taxus chinensis)로 구성된 그룹으로부터 선택되는 것을 특징으로 하는 택서스속 식물체로부터 택솔의 대량의 추출 정제방법.The plant according to claim 1 or 2, wherein the plant belongs to the genus Taxus brevifolia , Taxus canadensis , Taxus cuspidata , Taxus cuspidata , Taxus baccata , Taxus globosa , Taxus floridana , Taxus wallichiana , Taxus media , and Taxus chinensis . Lt; RTI ID = 0.0 > of Taxas. ≪ / RTI > 제1항에 있어서, 전기 제1용매 추출공정은 택서스속 식물체의 시료를 메탄올에 20 내지 200%(w/v)의 양으로 가하고, 상온에서 교반한 후 여과하여 메탄올 추출액을 수득함으로써 수행되는 것을 특징으로 하는 택서스속 식물체로부터 택솔의 대량 추출 정제방법.The method according to claim 1, wherein the first solvent extraction step is carried out by adding a sample of Taxus sp. Plant to methanol in an amount of 20 to 200% (w / v), stirring at room temperature and then filtering to obtain a methanol extract Wherein the method comprises the steps of: 제1항에 있어서, 전기 제2용매 추출공정은 디클로로메탄을 제1용매 추출공정으로부터 수득한 메탄올 추출액에 부피비로 10 내지 50%의 양으로 첨가하고, 상온에서 교반하여 방치한 다음, 메탄올층을 제거하여 디클로로메탄 추출액을 수득함으로써 수행되는 것을 특징으로 하는 택서스속 식물체로부터 택솔의 대량 추출 정제방법.The method according to claim 1, wherein the second solvent extraction step is carried out by adding dichloromethane to the methanol extract obtained from the first solvent extraction step in an amount of 10 to 50% by volume, stirring and stirring at room temperature, To obtain a dichloromethane extract. The method of mass-extraction and purification of taxol from Taxus sp. 제1항에 있어서, 전기 합성 흡착제 처리공정은 전기 제2용매 추출공정에서 수득한 디클로로메탄 추출액에 합성 흡착제를 전기 디클로로 메탄 추출액의 건고 중량의 10 내지 200%의 양으로 첨가하고, 30 내지 40℃의 온도에서 교반시키고 여과하여 여액을 수득함으로서 수행되는 것을 특징으로 하는 택서스속 식물체로부터 택솔을 대량 추출 정제방법.The process for treating an electrochemically adsorbent according to claim 1, wherein the synthetic adsorbent is added to the dichloromethane extract obtained in the second solvent extraction step in an amount of 10 to 200% of the dry weight of the electrodichloromethane extract, Followed by filtration to obtain a filtrate. The method of mass-extracting taxol from Taxus sp. 제1항 또는 제6항에 있어서, 전기 합성 흡착제로서 백토 또는 활성탄을 사용하는 것을 특징으로 하는 택서스속 식물체로부터 택솔을 대량 추출 정제방법.The method according to any one of claims 1 to 6, wherein clay or activated carbon is used as the electro-synthetic adsorbent. 제1항에 있어서, 전기 조 정제 택솔의 침전공정은 전기 합성 흡착제 처리공정에서 수득한 여액을 부피비로 500 내지 1,500%량의 핵산에 적가하여 침전시키고 침전을 여과함으로써 수행되는 것을 특징으로 하는 택서스속 식물체로부터 택솔을 대량 추출 정제방법.The process according to claim 1, wherein the precipitation step of the electrotooligosaccharide is carried out by dropwise adding the filtrate obtained in the electrochemical adsorbent treatment step to a nucleic acid amount of 500 to 1,500% by volume and precipitating and filtering the precipitate. A method for massive extraction and purification of Taxol from a plant. 제1항에 있어서, 전기 분별침전공정은 전기 조 정에 택솔을 메탄올에 1 내지 10%(w/v)의 농도로 용해시켜 수득한 조 정제 택솔의 메탄올 용액에 부피비로 30 내지 80%의 정제수를 소량씩 나누어 첨가하고, -20 내지 10℃의 저온에서 1 내지 3이간 방치함으로써 수행되는 것을 특징으로 하는 택서스속 식물체로부터 택솔의 대량 추출 정제방법.The electrolytic precipitation process according to claim 1, wherein the electrolytic precipitation step comprises adding 30 to 80% of purified water to the methanol solution of the crude tabasol obtained by dissolving taxol in methanol at a concentration of 1 to 10% (w / v) Is added in small portions and left at a low temperature of -20 to 10 占 폚 for 1 to 3 times. The method for massive extraction and purification of taxol from Taxus sp. 제1항에 있어서, 전기 고속액체 크로마토그래피 정제 공정은 1차로 소수성 수지 컬럼 고속액체 크로마토그래피를 사용하여 비극성 불순물을 제거하고, 이어서 실리카 컬럼을 사용하여 극성 불순물을 제거하는 2단계 공정으로 수행되는 것을 특징으로 하는 택서스속 식물체로부터 택솔의 대량 추출 정제방법.The process according to claim 1, wherein the electrophoretic high performance liquid chromatography purification step is carried out in a two-step process in which non-polar impurities are first removed using a hydrophobic resin column high-performance liquid chromatography, and then a polar impurity is removed using a silica column A method for massive extraction and purification of taxol from a plant belonging to the genus Taxus. 제10항에 있어서, 전기 소수성 수지 컬럼 고속액체 크로마토그래피 단계는 전기 분별침전공정에서 수득한 택솔 분말을 유기용매에 0.5 내지 20%(w/v)의 농도로 용해시켜 ODS(octadecylsilylated, C18)컬럼에 주입시켜 용출시킴으로써 수행되는 것을 특징으로 하는 택서스속 식물체로부터 택솔의 대량 추출 정제방법.11. The method of claim 10, wherein the electrohydrophobic resin column high-performance liquid chromatography step comprises dissolving the taxol powder obtained in the electrochemical precipitation step in an organic solvent at a concentration of 0.5 to 20% (w / v) to form octadecylsilylated (C 18 ) Wherein the step of extracting and eluting the taxol from the plant is carried out by injecting the extract into a column. 제11항에 있어서, 전기 유기용매로서 디메틸설폭사이드 또는 메탄올을 사용하는 것을 특징으로 하는 택서스속 식물체로부터 택솔의 대량 추출 정제방법.12. The method according to claim 11, wherein dimethyl sulfoxide or methanol is used as an electric organic solvent. 제11항에 있어서, 전기 ODS컬럼 고속 액체 크로마토그래피의 용리액으로 메탄올과 물의 1:0.3 내지 1:0.8(v/v) 혼합용액을 사용하는 것을 특징으로 하는 택서스속 식물체로부터 택솔을 대량 추출 정제방법.12. The method according to claim 11, wherein a mixture solution of methanol and water in an amount of 1: 0.3 to 1: 0.8 (v / v) is used as an eluent in an electric ODS column high performance liquid chromatography. Way. 제10항에 있어서, 전기 실리카 컬럼고속액체 크로마토그래피의 용리액으로 디클로로메탄과 메탄올의 100:0.5 내지 100:2(v/v) 혼합용액을 사용하는 것을 특징으로 하는 택서스속 식물체로부터 택솔의 대량 추출 정제방법.11. The method according to claim 10, wherein a large amount of taxol is obtained from a plant belonging to the Taxus family selected from the group consisting of a 100: 0.5 to 100: 2 (v / v) mixed solution of dichloromethane and methanol as an eluent of an electric silica column high performance liquid chromatography Extraction purification method. 제1항의 방법에 의해 택서스속(Taxusgenus) 식물체로부터 정제된 택솔. Taxus purified from a Taxus genus plant by the method of claim 1. ※ 참고사항 : 최초출원 내용에 의하여 공개하는 것임.※ Note: It is disclosed by the contents of the first application.
KR1019960002621A 1995-04-29 1996-02-03 Method for mass production of high purity taxol from taxus genus plant KR100259396B1 (en)

Priority Applications (8)

Application Number Priority Date Filing Date Title
KR1019960002621A KR100259396B1 (en) 1996-02-03 1996-02-03 Method for mass production of high purity taxol from taxus genus plant
JP8533189A JP2965168B2 (en) 1995-04-29 1996-04-27 Method for mass production of taxol from Taxus plants
CNB961904208A CN1166780C (en) 1995-04-29 1996-04-27 Method for mass production of taxol from taxus genus plant
PCT/KR1996/000059 WO1996034973A1 (en) 1995-04-29 1996-04-27 A method for mass production of taxol from taxus genus plant
AU55165/96A AU5516596A (en) 1995-04-29 1996-04-27 A method for mass production of taxol from taxus genus plant
EP96912315A EP0774010B1 (en) 1995-04-29 1996-04-27 A method for mass production of taxol from taxus genus plant
US08/652,493 US5900367A (en) 1995-04-29 1996-04-27 Method for purifying taxol from taxus biomass
DE69615499T DE69615499T2 (en) 1995-04-29 1996-04-27 MASS PRODUCTION METHOD OF TAXOL FROM PLANTS OF THE GENERAL TAXUS

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KR1019960002621A KR100259396B1 (en) 1996-02-03 1996-02-03 Method for mass production of high purity taxol from taxus genus plant

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KR970061883A true KR970061883A (en) 1997-09-12
KR100259396B1 KR100259396B1 (en) 2000-07-01

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