Method for preparing 9a-hydroxy androstendione

A technology for hydroxyandrostenedione and seeds, which is applied in the field of preparing 9a-hydroxyandrostenedione, can solve the problem that it is not suitable for industrial production of 9a-OHAD, the cost of the substrate androstenedione is high, and the conversion rate of androstenedione is poor. Advanced problems, to achieve the effect of shortening fermentation time, high yield and low by-products

Inactive Publication Date: 2013-10-09
SHANXI ZUYUAN IND & TRADE CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] It can be seen that the reaction steps of the first pathway are simple, but there are two defects in this pathway: 1) the conversion rate of androstenedione (AD) is not high, and the product concentration is low; 2) the substrate androstenedione (AD) high cost
[0009] M.V.Donova et al. (Appl Microbiol Biotechnol67:671–678, 2005) reported the use of Mycobacterium sp.2-4M to bioconvert sitosterol (sitosterol) to generate 9a-OH AD. After 120 hours of conversion, although the substrate conversion rate can be as high as 95- More than 97%, but the 9a-OH AD yield is only 48-50%, and the by-product AD yield reaches 21-22%, which is not conducive to subsequent separation and purification
[0010] No matter from the fermentation process, or from the yield and purity of the product, the currently disclosed methods are not suitable for the application of industrial production of 9a-OH AD

Method used

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  • Method for preparing 9a-hydroxy androstendione

Examples

Experimental program
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Embodiment 1

[0027] Activated strain: Inoculate Mycobacterium fortuitum ATCC35855 into sterile slant medium (yeast extract powder 1wt%; beef extract 0.5wt%; glucose 0.1wt%; sodium chloride 0.9wt%; agar 2wt%; pH6.8-7.0), culture at 30°C for 4 days until the whole slope is covered with bacteria.

[0028] Expanded cultivation: Scrape a well-growing single colony with an inoculation loop, inoculate it into a 250ml Erlenmeyer flask containing 30ml sterile seed medium, and cultivate it for 24 hours at a cultivation temperature of 30°C and a rotational speed of 200rpm. The seed medium includes: based on the total mass of the seed medium, phytosterols 0.05wt%, glucose 0.1wt%, ammonium sulfate 0.2wt%, magnesium sulfate heptahydrate 0.001wt%, dipotassium hydrogen phosphate 0.1wt%, yeast powder 0.5wt%, peptone 0.5wt%, sodium chloride 0.1wt%, calcium carbonate 0.1wt%, the balance is water; and the pH of the fermentation broth is 6.5-7.5.

[0029] Fermentation culture: Inoculate 5ml of bacterial liqui...

Embodiment 2

[0033] Expanded culture: insert the slant cells of Mycobacterium fortitum ATCC35855 in good growth state into a 250ml Erlenmeyer flask equipped with 30ml of sterile seed medium, and then vibrate at a culture temperature of 35°C and a rotation speed of 230rpm for 18 hours;

[0034] Fermentation culture: Then draw 5ml of seed solution according to the inoculation amount of 10% by volume and put it into a 500ml Erlenmeyer flask filled with 50ml of sterile fermentation broth, then culture at a temperature of 35°C and a rotation speed of 250rpm for 120 hours.

[0035] Then, the HPLC method described in Example 1 was used to measure the conversion rate of phytosterols, 9a-OH AD, and the concentration of by-product AD in the fermentation culture.

[0036] The seed culture medium and fermented liquid used in the present embodiment are respectively as follows:

[0037] The seed medium includes: phytosterol 0.075wt%, glucose 0.5wt%, ammonium sulfate 0.6wt%, magnesium sulfate heptahydrat...

Embodiment 3

[0041] Expanded culture: insert the slant cells of Mycobacterium fortitum ATCC35855 in a good growth state into a 250ml Erlenmeyer flask equipped with 30ml sterile seed medium, and then vibrate at a culture temperature of 32°C and a rotation speed of 230rpm for 24 hours;

[0042] Fermentation culture: Then draw 5ml of seed solution according to the inoculation amount of 10% by volume and put it into a 500ml Erlenmeyer flask filled with 50ml of sterile fermentation broth, then culture at a temperature of 32°C and a rotation speed of 250rpm for 110 hours.

[0043] Then, the HPLC method described in Example 1 was used to measure the conversion rate of phytosterols, 9a-OH AD, and the concentration of by-product AD in the fermentation culture.

[0044] The seed culture medium and fermented liquid used in the present embodiment are respectively as follows:

[0045] The seed medium includes: phytosterol 0.1wt%, glucose 0.25wt%, ammonium sulfate 0.3wt%, magnesium sulfate heptahydrate ...

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Abstract

The invention provides a method for preparing 9a-hydroxy androstendione, which comprises a step of performing fermentation culture on phytosterol by use of mycobacterium fortuitum ATCC 35855 so as to convert the phytosterol into 9a-hydroxy androstendione. The method is used for preparing 9a-hydroxy androstendione by use of mycobacterium fortuitum ATCC 35855 and corresponding fermentation liquid and fermentation technology; the cheap phytosterol can be adopted as a substrate; and moreover, the yield of the prepared 9a-hydroxy androstendione is high, a few byproducts are produced, the fermentation time is short, and a good way is provided for high-efficiency industrial production of 9a-hydroxy androstendione.

Description

technical field [0001] The invention relates to a method for preparing 9a-hydroxyandrostenedione, in particular to a method for preparing 9a-hydroxyandrostenedione by using Mycobacterium fortuitum ATCC35855. Background technique [0002] 9a-hydroxy androstenedione (9a-hydroxy androstenedione), referred to as 9a-OH AD, the molecular structure shown in the following formula 1, was first obtained by R.M.Dodson in 1958 from Nocardia (Nocardia) A20-10 fermentation broth A steroidal substance was isolated (J. Amer. Chem. Soc 80:6148, 1958). [0003] [Formula 1] [0004] [0005] 9a-OH AD is an important intermediate in the synthesis of steroid hormones, which can be used to synthesize corticoids (J.Org.Chem., 57:961-965, 1992), antiandrogenic , antiestrogenic and antifertility drugs. In addition, since the C9a-hydroxyl group is easily dehydrogenated with C11 to generate C9,11-dehydrosteroids, it is more conducive to the synthesis of 9a-halogenated-11β-hydroxyl steroids (US P...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P33/02C12R1/33
Inventor 产启银常继发曲卫国孙捷
Owner SHANXI ZUYUAN IND & TRADE CO LTD
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