PL219992B1 - Method for producing 3ß, 7ß-dihydroxy-5α-androst-17-one - Google Patents
Method for producing 3ß, 7ß-dihydroxy-5α-androst-17-oneInfo
- Publication number
- PL219992B1 PL219992B1 PL400204A PL40020412A PL219992B1 PL 219992 B1 PL219992 B1 PL 219992B1 PL 400204 A PL400204 A PL 400204A PL 40020412 A PL40020412 A PL 40020412A PL 219992 B1 PL219992 B1 PL 219992B1
- Authority
- PL
- Poland
- Prior art keywords
- androst
- dihydroxy
- hydroxy
- substrate
- formula
- Prior art date
Links
- 238000004519 manufacturing process Methods 0.000 title claims 2
- 238000000034 method Methods 0.000 claims description 13
- 239000000758 substrate Substances 0.000 claims description 5
- 230000009466 transformation Effects 0.000 claims description 5
- 230000000813 microbial effect Effects 0.000 claims description 4
- 241000122824 Aspergillus ochraceus Species 0.000 claims description 3
- 102000004190 Enzymes Human genes 0.000 claims description 3
- 108090000790 Enzymes Proteins 0.000 claims description 3
- 239000000376 reactant Substances 0.000 claims description 2
- 230000036983 biotransformation Effects 0.000 claims 2
- QGXBDMJGAMFCBF-UHFFFAOYSA-N Etiocholanolone Natural products C1C(O)CCC2(C)C3CCC(C)(C(CC4)=O)C4C3CCC21 QGXBDMJGAMFCBF-UHFFFAOYSA-N 0.000 description 5
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 4
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 4
- QGXBDMJGAMFCBF-LUJOEAJASA-N epiandrosterone Chemical compound C1[C@@H](O)CC[C@]2(C)[C@H]3CC[C@](C)(C(CC4)=O)[C@@H]4[C@@H]3CC[C@H]21 QGXBDMJGAMFCBF-LUJOEAJASA-N 0.000 description 4
- 150000003431 steroids Chemical class 0.000 description 4
- 230000033444 hydroxylation Effects 0.000 description 3
- 238000005805 hydroxylation reaction Methods 0.000 description 3
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 3
- 238000002360 preparation method Methods 0.000 description 3
- 241000233866 Fungi Species 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- RGVHVYXPVVRNAJ-UHFFFAOYSA-N 3-(3-methylindol-1-yl)propanenitrile Chemical compound C1=CC=C2C(C)=CN(CCC#N)C2=C1 RGVHVYXPVVRNAJ-UHFFFAOYSA-N 0.000 description 1
- 241000235556 Cunninghamella elegans Species 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 206010021143 Hypoxia Diseases 0.000 description 1
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 102000008109 Mixed Function Oxygenases Human genes 0.000 description 1
- 108010074633 Mixed Function Oxygenases Proteins 0.000 description 1
- 238000005481 NMR spectroscopy Methods 0.000 description 1
- 241000235546 Rhizopus stolonifer Species 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 0.000 description 1
- 239000003963 antioxidant agent Substances 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 239000011942 biocatalyst Substances 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000003197 catalytic effect Effects 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 239000012043 crude product Substances 0.000 description 1
- 239000003480 eluent Substances 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- OSVMTWJCGUFAOD-KZQROQTASA-N formestane Chemical compound O=C1CC[C@]2(C)[C@H]3CC[C@](C)(C(CC4)=O)[C@@H]4[C@@H]3CCC2=C1O OSVMTWJCGUFAOD-KZQROQTASA-N 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 230000007954 hypoxia Effects 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 230000003961 neuronal insult Effects 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- 239000007800 oxidant agent Substances 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 230000001590 oxidative effect Effects 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 230000003595 spectral effect Effects 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Description
Opis wynalazkuDescription of the invention
Przedmiotem wynalazku jest sposób wytwarzania 3β,7β-dihydroksy-5α-androst-17-onu, o wzorze 2 przedstawionym na rysunku.The present invention relates to a process for the preparation of 3β, 7β-dihydroxy-5α-androst-17-one of the formula 2 shown in the drawing.
Wynalazek może znaleźć zastosowanie w przemyśle farmaceutycznym.The invention may find application in the pharmaceutical industry.
Epimeryczne 7-hydroksypochodne 3β-hydroksy-5α-androst-17-onu (epiandrosteronu) są natywnymi metabolitami ssaków, które odznaczają się różnorodną aktywnością biologiczną (B. Dudas, M. Rose, E. Wulfert, Neurobiol. Dis. 2004, 15, ss. 262-268; A. Pringle, W. Schmidt, J. Deans, E. Wulfert, K. Reymann, L. Sundstrom, Eur. J. Neurosci. 2003,18, ss. 117-24).Epimeric 7-hydroxy derivatives of 3β-hydroxy-5α-androst-17-one (epiandrosterone) are native metabolites of mammals, which are characterized by various biological activity (B. Dudas, M. Rose, E. Wulfert, Neurobiol. Dis. 2004, 15, pp. 262-268; A. Pringle, W. Schmidt, J. Deans, E. Wulfert, K. Reymann, L. Sundstrom, Eur. J. Neurosci. 2003, 18, pp. 117-24).
Szczególnie wysoką aktywnością charakteryzuje się ekwatorialna C7-hydroksypochodna 3β,7β-dihydroksy-5α-androst-17-on ^-hydroksyepiandrosteron) (O. Hennebert, M-A Pelissier, S. Le Mee, R. Morfin, J. Steroid Biochem. Mol. Biol, 2008, 110, ss. 255-262).The equatorial C7-hydroxy derivative 3β, 7β-dihydroxy-5α-androst-17-one4-hydroxyepiandrosterone is particularly active (O. Hennebert, MA Pelissier, S. Le Mee, R. Morfin, J. Steroid Biochem. Mol. Biol, 2008, 110, pp. 255-262).
3β,7β-dihydroksy-5α-androst-17-on posiada właściwości przeciwutleniające i przeciwzapalne oraz zapobiega uszkodzeniom neuronów spowodowanym niedotlenieniem (S. Niro, E. Pereira, M.A. Pelissier, R. Morfin, O. Hennebert, Steroids, 2012, 77, ss. 542-551; L. El Kihel, Steroids, 2011 77, ss. 10-26). Dlatego aktualnie podejmowane są badania nad opracowaniem metod syntezy 7-hydroksypochodnych epiandrosteronu na drodze chemicznej i enzymatycznej (C. Ricco, G. Revial, C. Ferroud, O. Hennebert, R. Morfin, Steroids, 2011, 76, ss. 28-30; T. Kołek, N. Milecka, A. Świzdor, A. Panek, A. Białońska, Org. Biomol. Chem., 2011, 9, ss. 5414-5422).3β, 7β-dihydroxy-5α-androst-17-one has antioxidant and anti-inflammatory properties and prevents neuronal damage caused by hypoxia (S. Niro, E. Pereira, MA Pelissier, R. Morfin, O. Hennebert, and Steroids, 2012, 77, pp. 542-551; L. El Kihel, Steroids, 2011 77, pp. 10-26). Therefore, research is currently being undertaken on the development of methods for the synthesis of 7-hydroxy derivatives of epiandrosterone by chemical and enzymatic methods (C. Ricco, G. Revial, C. Ferroud, O. Hennebert, R. Morfin, Steroids, 2011, 76, pp. 28-30). ; T. Kołek, N. Milecka, A. Świzdor, A. Panek, A. Białońska, Org. Biomol. Chem., 2011, 9, pp. 5414-5422).
Enzymatyczna hydroksylacja, w której utleniaczem jest atmosferyczny tlen, jest przyjazną dla środowiska metodą utleniania wiązania C-H, która umożliwia m.in. wprowadzanie grupy hydroksylowej w nieaktywowaną pozycję substratu (P. Fernandes, A. Gruz, B. Angelova, H.M. Pinheiro, J.M.S. Carbal, Enzyme. Microbial. Technology, 2003, 32, ss. 688-705). Jako biokatalizatory najczęściej stosowane są grzyby strzępkowe, ze względu na różnorodność produkowanych przez te grzyby hydroksylaz, jak i ich wysoką aktywność katalityczną.Enzymatic hydroxylation, in which the oxidant is atmospheric oxygen, is an environmentally friendly method of oxidation of the C-H bond, which allows, among others, introduction of the hydroxyl group into the unactivated position of the substrate (P. Fernandes, A. Gruz, B. Angelova, H. M. Pinheiro, J.M.S. Carbal, Enzyme. Microbial. Technology, 2003, 32, pp. 688-705). Filamentous fungi are most often used as biocatalysts, due to the variety of hydroxylases produced by these fungi and their high catalytic activity.
Znane są sposoby mikrobiologicznej hydroksylacji epiandrosteronu przy C-7, np. szczep Rhizopus nigricans przekształcał 3β-hydroksy-5α-androst-17-onu do mieszaniny: 7a-hydroksy- (15%), 7β-hydroksy- (12%) i 6a-hydroksy- (12%) pochodnych (V. E. M. Chambers, W.A. Denny, J.M. Evans, E.R.H. Jones, A. Kasal, G.D. Meakins, J. Pragnell, J. Chem. Soc., Perkin Trans. 1, 1973, ss. 1500-1511).There are known methods of microbial hydroxylation of epiandrosterone at C-7, e.g. the Rhizopus nigricans strain transformed 3β-hydroxy-5α-androst-17-one into a mixture of: 7a-hydroxy- (15%), 7β-hydroxy- (12%) and 6a -hydroxy- (12%) derivatives (VEM Chambers, WA Denny, JM Evans, ERH Jones, A. Kasal, GD Meakins, J. Pragnell, J. Chem. Soc., Perkin Trans. 1, 1973, pp. 1500- 1511).
W transformacji przez szczep Cunninghamella elegans z octanu epiandrosteronu otrzymywano mieszaninę 7a-hydroksy- (2,5%) i 7β-hydroksy- (22%) epiandro-steron obok 3β-hydroksy-5α-androst-7,17-dionu (2,5%) (T.A. Crabb, J.A. Saul, R.O. Williams, J. Chem Soc. Perkin Trans. 1, 1981, ss. 1041-1045).In the transformation by the Cunninghamella elegans strain from epiandrosterone acetate, a mixture of 7a-hydroxy- (2.5%) and 7β-hydroxy- (22%) epiandro-sterone was obtained along with 3β-hydroxy-5α-androst-7,17-dione (2, 5%) (TA Crabb, JA Saul, RO Williams, J. Chem. Soc. Perkin Trans. 1, 1981, pp. 1041-1045).
Wynalazek dotyczy sposobu wytwarzania 3β,7β-dihydroksy-5α-androsta-17-onu ^β-hydroksyepiandrosteronu) o wzorze 2, na drodze mikrobiologicznej hydroksylacji 3β-hydroksy-5α-androst-17-onu (epiandrosteronu), o wzorze 1.The invention relates to a process for the preparation of 3β, 7β-dihydroxy-5α-androsta-17-one (β-hydroxyepiandrosterone) of formula 2 by microbial hydroxylation of 3β-hydroxy-5α-androst-17-one (epiandrosterone) of formula 1.
Istota wynalazku polega na tym, że 3β-hydroksy-5α-androst-17-on, przekształca się do 3β,7β-dihydroksy-5a-androsta-17-onu ^-hydroksyepiandrosteronu) za pomocą kultury szczepu Aspergillus ochraceus AM456.The essence of the invention is that 3β-hydroxy-5α-androst-17-one is transformed into 3β, 7β-dihydroxy-5a-androsta-17-one (β-hydroxyepiandrosterone) by the culture of the Aspergillus ochraceus AM456 strain.
Korzystne jest, gdy proces transformacji prowadzi się wodną kulturą szczepu, przy wstrząsaniu reagentów, w temperaturze 293-300 K.It is advantageous if the transformation process is carried out with an aqueous strain culture, while agitating the reactants, at a temperature of 293-300 K.
Zasadniczą zaletą wynalazku jest otrzymanie 3β,7β-dihydroksy-5α-androst-17-onu, z łączną wydajnością 60,1%, w temperaturze pokojowej i pH bliskim obojętnemu.The main advantage of the invention is the preparation of 3β, 7β-dihydroxy-5α-androst-17-one, with a total yield of 60.1%, at room temperature and near neutral pH.
Wynalazek jest bliżej objaśniony na przykładzie wykonania.The invention is explained in more detail using an exemplary embodiment.
P r z y k ł a d. Do kolby Erlenmayera o pojemności 300 cm3, w której znajduje się 100 cm3 ste3 rylnej pożywki zawierającej 3 g glukozy i 1 g aminobaku, wprowadza się 2 cm3 zawiesiny komórek Aspergillus ochraceus AM456 wzrastających przez trzy dni na tym samym podłożu przy stałym wstrząsaniu w temperaturze 293-300 K. Po trzech dniach wzrostu dodaje się 20 mg 3β-hydroksy-5α3 Example d. To the Erlenmeyer flask with a capacity of 300 cm 3, which is 100 cm 3 ste 3 rylnej medium containing 3 g of glucose and 1 g aminobaku introduced 2 cm 3 cell suspension of Aspergillus ochraceus AM456 growing for three days in the same medium with constant shaking at a temperature of 293-300 K. After three days of growth, 20 mg of 3β-hydroxy-5α 3 are added
-androst-17-onu, o wzorze 1, rozpuszczonego w 1 cm3 acetonu. Transformację prowadzi się przy ciągłym wstrząsaniu przez kolejne 24 godzin w warunkach, w których prowadzona jest hodowla mikroorganizmu. Następnie uzyskane roztwory transformacyjne ekstrahuje się trzykrotnie chloroformem, osusza bezwodnym siarczanem magnezu i odparowuje rozpuszczalnik. Otrzymuje się 26 mg surowego produktu, który oczyszcza się chromatograficznie używając jako eluentu mieszaniny: heksan: :aceton:chloroform (1:2:2). Na tej drodze otrzymuje się 12,0 mg 3β,7β-dihydroksy-5α-androst-17-onu (wydajność 60,1%).-androst-17-one, formula I, dissolved in 1 cm 3 of acetone. The transformation is carried out under continuous shaking for a further 24 hours under the conditions under which the microorganism is cultivated. The resulting transformation solutions were then extracted three times with chloroform, dried with anhydrous magnesium sulfate, and the solvent was evaporated. 26 mg of crude product are obtained which are purified by chromatography using as eluent: hexane: acetone: chloroform (1: 2: 2). In this way, 12.0 mg of 3β, 7β-dihydroxy-5α-androst-17-one are obtained (yield 60.1%).
PL 219 992 B1PL 219 992 B1
Uzyskany produkt charakteryzuje się następującymi danymi spektralnymi:The obtained product is characterized by the following spectral data:
1H-NMR (δ ppm) w CDCI3: 0,85 (3H, s, I9-CH3); 0,88 (3H, s, I8-CH3); 3,47 (1H, m, Wh=15,8 Hz, 7a-H); 3,59 (1H, m, Wh=18,2 Hz, 3a-H); 1 H-NMR (δ ppm) in CDCl 3: 0.85 (3H, s, 19-CH 3); 0.88 (3H, s, 18-CH3); 3.47 (1H, m, Wh = 15.8 Hz, 7a-H); 3.59 (1H, m, Wh = 18.2Hz, 3a-H);
13C NMR (δ ppm): 221,4 (C-17); 74,46 (C-7); 70,9 (C-3); 52,4 (C-14); 51 (C-9); 48,2 (C-13); 42,8 (C-8); 42 (C-5); 38,7 (C-12); 37,5 (C-1); 36,7 (C-6); 36 (C-4); 13 C NMR (δ ppm): 221.4 (C-17); 74.46 (C-7); 70.9 (C-3); 52.4 (C-14); 51 (C-9); 48.2 (C-13); 42.8 (C-8); 42 (C-5); 38.7 (C-12); 37.5 (C-1); 36.7 (C-6); 36 (C-4);
35,1 (C-10); 31,4 (C-16); 31,3 (C-2); 24,9 (C-15); 20,7 (C-11); 14 (C-18);35.1 (C-10); 31.4 (C-16); 31.3 (C-2); 24.9 (C-15); 20.7 (C-11); 14 (C-18);
12,4 (C-19);12.4 (C-19);
IR vmax (cm-1): 3457, 1739.IR vmax (cm -1 ): 3457, 1739.
Claims (3)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| PL400204A PL219992B1 (en) | 2012-08-01 | 2012-08-01 | Method for producing 3ß, 7ß-dihydroxy-5α-androst-17-one |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| PL400204A PL219992B1 (en) | 2012-08-01 | 2012-08-01 | Method for producing 3ß, 7ß-dihydroxy-5α-androst-17-one |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| PL400204A1 PL400204A1 (en) | 2013-05-13 |
| PL219992B1 true PL219992B1 (en) | 2015-08-31 |
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Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
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| PL400204A PL219992B1 (en) | 2012-08-01 | 2012-08-01 | Method for producing 3ß, 7ß-dihydroxy-5α-androst-17-one |
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| Country | Link |
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| PL (1) | PL219992B1 (en) |
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- 2012-08-01 PL PL400204A patent/PL219992B1/en unknown
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| Publication number | Publication date |
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| PL400204A1 (en) | 2013-05-13 |
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Effective date: 20141222 |