Method for preparing 9alpha-hydroxide-androstenedione by utilizing microbial conversion

A technology for microbial transformation and androstenedione, which is applied in the directions of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., can solve the problems of low feeding concentration, long conversion time and high production cost, and achieves high feeding concentration and high production cost. Simple process and low cost effect

Active Publication Date: 2013-10-23
SHANGHAI ADVANCED RES INST CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Through this method, the disadvantages of low feed concentration, long conversion time and high production cost in the prior art can be overcome.

Method used

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  • Method for preparing 9alpha-hydroxide-androstenedione by utilizing microbial conversion

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] (1) Primary seed cultivation

[0038] Primary seed medium: peptone 0.5g / 100mL, yeast extract 1.3g / 100mL~1.4g / 100mL, glucose 1.2g / 100mL, water, adjust pH to 7.0-7.2, and sterilize.

[0039] Take an inoculation loop of Rhodococcus erythropolis (Rhodococcus erythropolis) ATCC14887 and put it in 20mL primary seed medium, and culture it at 30°C for 18h on a rotary shaker at 200rpm to obtain seeds suitable for inoculation.

[0040] (2) Secondary fermentation culture

[0041] Transformation medium: glucose 0.5g / 100mL, peptone 0.2g / 100mL, corn steep liquor 2.5g / 100mL, dipotassium hydrogen phosphate 0.02g / 100mL, magnesium sulfate 0.01g / 100mL, water, initial pH 6.8, and sterilized.

[0042] Inoculate 5 mL of the seeds obtained in step (1) into 95 mL of transformation medium [the inoculum size is 5% (V / V)], and shake at 200 rpm for 18 hours at 28°C to obtain a fermentation broth.

[0043] (3) Conversion of the substrate androstenedione

[0044] Dissolve 2 g of androstenedione wit...

Embodiment 2

[0050] (1) Primary seed cultivation

[0051] The steps were the same as in Example 1, but the culture conditions were changed to: culture at 35° C. for 12 hours on a rotary shaker at 150 rpm.

[0052] (2) Secondary fermentation culture

[0053] Secondary transformation medium: glucose 1g / 100mL, peptone 0.6g / 100mL, corn steep liquor 3.5g / 100mL, dipotassium hydrogen phosphate 0.06g / 100mL, magnesium sulfate 0.03g / 100mL, water, initial pH 7.0, and sterilized .

[0054] Inoculate 5 mL of the seeds obtained in step (1) into 95 mL of transformation medium [the inoculum size is 5% (V / V)], and culture at 30°C for 20 h with shaking at 250 rpm to obtain a fermentation broth.

[0055] (3) Conversion of the substrate androstenedione

[0056] At 90°C, dissolve 3.5g of androstenedione with 3mL of methanol, put it into about 100mL of the fermentation broth obtained in step (2) under sterile conditions, and continue to shake at 250rpm at 30°C for 56h to obtain the transformation liquid . ...

Embodiment 3

[0062] (1) Primary seed cultivation

[0063] The steps are the same as in Example 1, but the culture conditions are changed: culture at 26° C. for 15 h on a rotary shaker at 180 rpm.

[0064] (2) Secondary fermentation culture

[0065] Secondary transformation medium: glucose 1.5g / 100mL, peptone 1g / 100mL, corn steep liquor 5.0g / 100mL, dipotassium hydrogen phosphate 0.08g / 100mL, magnesium sulfate 0.05g / 100mL, water, initial pH 7.5, and sterilized .

[0066] Inoculate 5 mL of the seeds obtained in step (1) into 95 mL of transformation medium [the inoculum size is 5% (V / V)], and shake at 280 rpm for 16 hours at 32°C to obtain a fermentation broth.

[0067] (3) Conversion of the substrate androstenedione

[0068] At 85°C, dissolve 4.5g of androstenedione with 5mL of methanol, put it into about 100mL of the fermentation broth obtained in step (2) under sterile conditions, and continue to shake at 280rpm at 32°C for 72h to obtain the transformation liquid .

[0069] Among them,...

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Abstract

The invention discloses a method for preparing 9alpha-hydroxide-androstenedione by utilizing microbial conversion. The method comprises the steps of: 1) by taking rhodococcus erythropolis as a seed lot, inoculating rhodococcus erythropolis in a seed culture medium, and culturing to obtain seeds; 2) inoculating the seeds into a conversion culture medium, and culturing to obtain fermentation liquor; 3) adding an androstenedione solution into the fermentation liquor, continuing culturing to obtain conversion liquor, and detecting the content of 9alpha-hydroxide-androstenedione in the conversion liquor; and 4) separating and extracting the 9alpha-hydroxide-androstenedione in the conversion liquor. The method has the advantages of being high-efficiency and specific, environment-friendly, high in added materail concentration, high in conversion rate, product purity and yield, low in cost and the like, is simple in overall preparation process, and economic and practical, has wide application prospects, and has very important significance on the application for extending the steroid raw materail drug androstenedione.

Description

technical field [0001] The invention relates to a method for preparing 9α-hydroxyl-androstenedione, in particular to a method for preparing 9α-hydroxyl-androstenedione through microbial conversion. Background technique [0002] 9α-Hydroxy-androstenedione (9α-OH-AD) is a key intermediate product in the industrial production of antiestrous steroids and plays an important role in the industrial production of steroids. Using 9α-hydroxy-androstenedione as a precursor can synthesize hydrocortisone, 17α-OH progesterone, eplerenone, βmethasone, dexamethasone, cortisone, dexamethasone, fluprogesterone, Fludroxone, Temeifu, Triamcinolone and many other important steroid raw materials have important commercial value. The key to preparing 9α-hydroxy-androstenedione is to add a hydroxyl group to the 9th position of androstenedione. At present, chemical synthesis methods are mostly used at home and abroad, but chemical synthesis methods not only have many synthesis steps and low yields,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P33/06C12R1/01
Inventor 张保国魏长龙曾俊华王珊史吉平姜标
Owner SHANGHAI ADVANCED RES INST CHINESE ACADEMY OF SCI
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