JPH07501689A - Method for producing 14α-hydroxy-4-androstene-3,17-dione - Google Patents
Method for producing 14α-hydroxy-4-androstene-3,17-dioneInfo
- Publication number
- JPH07501689A JPH07501689A JP5504864A JP50486493A JPH07501689A JP H07501689 A JPH07501689 A JP H07501689A JP 5504864 A JP5504864 A JP 5504864A JP 50486493 A JP50486493 A JP 50486493A JP H07501689 A JPH07501689 A JP H07501689A
- Authority
- JP
- Japan
- Prior art keywords
- androstene
- dione
- hydroxy
- producing
- fermentation
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P33/00—Preparation of steroids
- C12P33/06—Hydroxylating
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07J—STEROIDS
- C07J1/00—Normal steroids containing carbon, hydrogen, halogen or oxygen, not substituted in position 17 beta by a carbon atom, e.g. estrane, androstane
- C07J1/0003—Androstane derivatives
- C07J1/0011—Androstane derivatives substituted in position 17 by a keto group
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Wood Science & Technology (AREA)
- Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Zoology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Microbiology (AREA)
- General Chemical & Material Sciences (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Genetics & Genomics (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Steroid Compounds (AREA)
Abstract
(57)【要約】本公報は電子出願前の出願データであるため要約のデータは記録されません。 (57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.
Description
【発明の詳細な説明】 14α−ヒドロキシ−4−アンドロステン−3,17−ジオンの製造方法 本発明は、カルプラリアルナータ((:urvularia 1unata ) の培養菌を用いて4−アンドロステン−3,17−ジオンを発酵させることによ り14α−ヒドロキシ−4−アンドロステン−3,17−ジオンを製造する方法 において、4−アンドロステン−3,17−ジオンを水中に懸濁させ、得られた @/R液を発酵させることを特徴とする、14α−ヒドロキシ−4−アンドロス テン−3,17−ジオンの製造方法に関する。[Detailed description of the invention] Method for producing 14α-hydroxy-4-androstene-3,17-dione The present invention relates to carpularia lunata ((:urvularia 1unata) by fermenting 4-androstene-3,17-dione using cultured bacteria. Method for producing 14α-hydroxy-4-androstene-3,17-dione , 4-androstene-3,17-dione was suspended in water, and the obtained 14α-hydroxy-4-andros, characterized by fermenting @/R liquid The present invention relates to a method for producing ten-3,17-dione.
ソ連特許第801517号明細書(C,A、110゜133697u参照)には 、カルブラリアルナータの培養菌を用いて4−アンドロステン−3,17−ジオ ンを発酵させることによる、14α−ヒドロキシ−4−アンドロステン−3,1 7−ジオンの製造方法が記載されている。しかしながら該特許明細書には、該微 生物がどの公開菌株寄託機関からのものであるか、及びどの番号下にあるのかと いう指示が含まれていないので、記載された方法を追試することができない。該 明細書の技術的手段によれば4−アンドロステン−3゜17−ジオンをメタノー ル又はジメチルフォルムアミド中に溶解させ、基質0.5〜1.5g/lを40 〜66時間発酵させて、明らかに著しく強度に不純化された14α−ヒドロキシ −4−アンドロステン−3,17−ジオン粗製生成物35〜70%が得られる。In Soviet Patent No. 801517 (see C, A, 110゜133697u) 4-androstene-3,17-dio 14α-hydroxy-4-androstene-3,1 by fermenting A method for producing 7-dione is described. However, the patent specification does not include the Which public strain depository does the organism come from and under what number? Since there are no instructions included, it is not possible to retry the method described. Applicable According to the technical means in the specification, 4-androstene-3゜17-dione is converted into methanol. 0.5-1.5 g/l of substrate dissolved in dimethylformamide or dimethylformamide at 40% After ~66 hours of fermentation, the 14α-hydroxy was apparently severely contaminated. 35-70% of -4-androstene-3,17-dione crude product is obtained.
(そのことは、後処理方法から明白である、すなわち発酵バッチをクロロホルム で抽出し、クロロホルム相を濃縮し、残留物をヘキサンとともに煮沸する。(This is evident from the work-up method, i.e. the fermentation batch is chloroformed. The chloroform phase is concentrated and the residue is boiled with hexane.
従って、このようにして得られた生成物はすべて抽出可能であり、ヘキサン中に 不溶な物質を含有しなければならない。該生成物は、発酵パッチ中に含有される すべてのステロイドだけでなく、相応するカルブラリア含有物質及び媒体成分で もある)。Therefore, all the products obtained in this way are extractable and in hexane Must contain insoluble substances. The product is contained in a fermentation patch With all steroids as well as corresponding calbularia-containing substances and vehicle components ).
これとは異なり、本発明の方法により1リットル当り5gより多い4−アンドロ ステン−3,17−ジオンを約12時間以内で再現可能で14α−ヒドロキシ− 4−アンドロステン−3,17−ジオンに転化することができる。該処理生成物 の収率は、純粋生成物約60%である。In contrast, the method according to the invention provides more than 5 g of 4-andro per liter. Stene-3,17-dione can be reproduced within about 12 hours and 14α-hydroxy- It can be converted to 4-androstene-3,17-dione. the processed product The yield is about 60% pure product.
本発明による方法は、カルブラリアルナータ培養菌を用いた公知の微生物学的な 基質の転化にも使用される発酵条件下で行われる。The method according to the present invention is based on the known microbiological method using cultured microorganisms of Calburial Inata. It is carried out under fermentation conditions that are also used for substrate conversion.
該微生物のために通常使用される培養条件下で、適した培地中で、通気下で液体 培養菌を増殖させる。次いで、培養菌に4−アンドロステン−3,17−ジオン の水性!15液を加え、基質転化が最大に達するまで発酵させる。4−アンドロ ステン−3,17−ジオンの水性懸濁液は容易な方法で製造することができる、 例えば該基質を、2〜5倍の量の水とともに非イオン性界面活性剤を添加しなが らボールミル内で、該基質が約1〜5μmの粒度を有するまで粉砕する。適当な 界面活性剤は、例えばポリグリコールの酸化エチレン付加物又は脂肪酸エステル である。適当な界面活性剤としては、例えば市販の湿潤剤、例えばTegin( R)、Tween(R)又は5pan (R)が挙げられる。The liquid under aeration in a suitable medium under culture conditions normally used for the microorganism. Grow cultured bacteria. Next, the cultured bacteria were treated with 4-androstene-3,17-dione. Water-based! Add liquid 15 and ferment until maximum substrate conversion is reached. 4-Andro Aqueous suspensions of sten-3,17-dione can be prepared in a simple manner. For example, the substrate is mixed with 2 to 5 times the amount of water while adding a nonionic surfactant. The substrate is ground in a ball mill until the substrate has a particle size of approximately 1-5 μm. Appropriate Surfactants include, for example, ethylene oxide adducts of polyglycols or fatty acid esters. It is. Suitable surfactants include, for example, commercially available wetting agents such as Tegin ( R), Tween (R) or 5pan (R).
発酵を実施した後、培養菌を常法で後処理する1例えば難水溶性アルコール、ケ トン又はエステルで抽出し、該抽出液を濃縮し、次いで得られた残留物をクロマ トグラフィー及び/又は晶出により精製する。After fermentation, the cultured bacteria is post-treated using a conventional method. The extract was concentrated and the resulting residue was chromatographed. Purification by tography and/or crystallization.
得られた14α−ヒドロキシ−4−アンドロステン−3,17〜ジオンの薬理学 的に作用するステロイドへの次の処理は公知であるか、又は自体公知の方法で行 う(J、 Amer、 Chew、 Sac、、 91,1969.1228以 降:ヨーロッパ特許出願公開第0300062号明細書、その他)。Pharmacology of the obtained 14α-hydroxy-4-androstene-3,17-dione The subsequent treatments for steroids that act as (J, Amer, Chew, Sac, 91, 1969.1228 onwards) (Reference: European Patent Application Publication No. 0300062, etc.).
次に、本発明を以下の実施例につき詳細に説明する例。The invention will now be explained in detail with reference to the following examples.
a)コーンステイーブリーカー 1% 大豆パウダー 1,25% pH値6.2に調整 を含有する無菌培地1リツトルを有する2リツトルの工−レンマイヤーに、カル ブラリアルナータNRRL2380培養薗の!!!濁液濁液5奢l種し、30℃ で60時間、1分間に180回転で振盪させた。a) Corn staple breaker 1% Soybean powder 1.25% Adjust to pH value 6.2 In a 2 liter factory containing 1 liter of sterile medium containing Burarialnata NRRL2380 Culture Sono! ! ! Seed 5 liters of suspension, 30℃ The mixture was shaken at 180 revolutions per minute for 60 hours.
b)a)に記載と同様の無菌培養液40リツトルを有する50リツトルの発酵槽 に、カルブラリアルナータ増殖培11リットルを接種し、30’Cで18時間、 1時間に2m3の通気及び1分間に220回転の撹拌下で温室した。b) 50 liter fermentor with 40 liters of sterile culture medium as described in a) was inoculated with 11 liters of Calburial Innata growth medium and incubated at 30'C for 18 hours. The greenhouse was kept under aeration of 2 m3 per hour and stirring at 220 revolutions per minute.
C)コーンステイーブリーカー 1% 大豆パウダー 1.25% β−シクロデクストリン 0.3% pH値6,2に調整 を含有する無菌培養液40リツトルを有する50リツトルの発酵槽に、カルブラ リアルナータ前培養菌4リットルを接種し、1時間に2.4m’の通気及び1分 間に150回転の撹拌下で30℃で7時間湿地した。C) Corn staple breaker 1% Soybean powder 1.25% β-cyclodextrin 0.3% Adjust to pH value 6.2 In a 50 liter fermenter containing 40 liters of sterile culture medium containing Inoculate 4 liters of Rialnata preculture, aerate 2.4 m' per hour and aerate for 1 minute. The mixture was wetted at 30° C. for 7 hours with stirring at 150 rpm.
次いで、該培養菌4−アンドロステン−3,17−ジオン250gを粉砕物とし てTween(R)8025 gを有する水10100O中に加え、更に12時 間、1分間に220回転の撹拌及び1時間に2.4m’の通気下で発酵させた。Next, 250 g of the cultured bacteria 4-androstene-3,17-dione was ground. and added to 10,100 O of water containing 8,025 g of Tween(R), and further added at 12 o'clock. Fermentation was carried out under stirring at 220 revolutions per minute and aeration of 2.4 m' per hour.
d)発酵を実施した後に、該培養菌の液体をそれぞれメチルイソブチルケトン2 0リツトルで3回抽出し、該抽出液を回転蒸発器中、最大50℃で真空下で濃縮 した。引続き、ヘキサン−アセトン傾斜を用いたシリカゲル上でのクロマトグラ フィーによる精製を行った融点257〜259℃の14α−ヒドロキシ−アンド ロスタ−4−エン−3,17−ジオン150gが得られた。d) After carrying out the fermentation, the liquid of the culture is treated with methyl isobutyl ketone 2 Extract 3 times with 0 liters and concentrate the extract under vacuum in a rotary evaporator at a maximum of 50 °C. did. Followed by chromatography on silica gel using a hexane-acetone gradient. 14α-hydroxy-and, melting point 257-259°C, purified by 150 g of rost-4-ene-3,17-dione were obtained.
国際調査報告 。、71.。。5/n17゜7PCT/EP 92101797International search report. , 71. . . 5/n17゜7PCT/EP 92101797
Claims (2)
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
DE19914129005 DE4129005A1 (en) | 1991-08-28 | 1991-08-28 | METHOD FOR PRODUCING 14 (ALPHA) -HYDROXY-4-ANDROSTEN-3,17-DION |
DE4129005.4 | 1991-08-28 | ||
PCT/EP1992/001797 WO1993005170A1 (en) | 1991-08-28 | 1992-08-07 | METHOD OF PREPARING 14α-HYDROXY-4-ANDROSTENE-3,17-DIONE |
Publications (1)
Publication Number | Publication Date |
---|---|
JPH07501689A true JPH07501689A (en) | 1995-02-23 |
Family
ID=6439592
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
JP5504864A Pending JPH07501689A (en) | 1991-08-28 | 1992-08-07 | Method for producing 14α-hydroxy-4-androstene-3,17-dione |
Country Status (4)
Country | Link |
---|---|
EP (1) | EP0600946A1 (en) |
JP (1) | JPH07501689A (en) |
DE (1) | DE4129005A1 (en) |
WO (1) | WO1993005170A1 (en) |
Family Cites Families (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
FR1267758A (en) * | 1956-12-19 | 1961-07-28 | Merck & Co Inc | Preparation process |
ATE78825T1 (en) * | 1987-02-06 | 1992-08-15 | Snow Brand Milk Products Co Ltd | NEW ANDROST-4-EN-3,17-DIONE DERIVATIVES AND METHODS OF PRODUCTION. |
-
1991
- 1991-08-28 DE DE19914129005 patent/DE4129005A1/en not_active Withdrawn
-
1992
- 1992-08-07 JP JP5504864A patent/JPH07501689A/en active Pending
- 1992-08-07 EP EP92917129A patent/EP0600946A1/en not_active Withdrawn
- 1992-08-07 WO PCT/EP1992/001797 patent/WO1993005170A1/en not_active Application Discontinuation
Also Published As
Publication number | Publication date |
---|---|
DE4129005A1 (en) | 1993-03-04 |
WO1993005170A1 (en) | 1993-03-18 |
EP0600946A1 (en) | 1994-06-15 |
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