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Fermentation method for producing epothilone B by sorangium cellulosum and fermentation medium

A technology of C. celluliformis and a fermentation medium, which is applied to the field of fermentation and fermentation medium for the production of Epothilone B by C. cellulosum, and can solve the problems of difficulty in culturing, low yield, and mutagenesis screening of Myxobacter celluliformis Difficulty and other problems, to achieve the effect of stable strain performance, increased yield and significant effect

Inactive Publication Date: 2010-10-06
SHANGHAI INST OF PHARMA IND
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although Epothilone B can be produced by fermenting myxobacterium cellulotrophs using M26 medium, the yield is low; if mutagenesis screening is used, it is very difficult to perform mutagenesis screening because myxobacterium cellulosomes are difficult to cultivate

Method used

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  • Fermentation method for producing epothilone B by sorangium cellulosum and fermentation medium
  • Fermentation method for producing epothilone B by sorangium cellulosum and fermentation medium

Examples

Experimental program
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Effect test

Embodiment 1-4

[0031] Table 1 has provided the formula of fermentation medium embodiment 1~4 of the present invention, according to listed component and its content in Table 1 and its content (percentage is mass percentage), simply mix uniformly, make up content to mass percentage with deionized water 100%, sterilized and sterilized, that is, each fermentation medium is prepared.

[0032] Among them, trace element solution preparation (1L system): MnCl 2 4H 2 O 100mg, CoCl 2 20mg, CuSO 4 10 mg, Na 2 MoO 4 2H 2 O 10mg, ZnCl 2 20mg, LiCl 5mg, SnCl 2 2H 2 O5mg, H 3 BO 3 10mg, KBr 20mg, KI 20mg, EDTA Na-Fe 3+ 8g.

[0033]

[0034]

Embodiment 5

[0036]Myxobacterium cellulosus seeds: purchased from the American Type Culture Collection ATCC, the storage number is ATCC15384, and the seeds can be stored in glycerol tubes or freeze-dried tubes.

[0037] Trace element solution preparation (1L system): MnCl 2 4H 2 O 100mg, CoCl 2 20mg, CuSO 4 10 mg, Na 2 MoO 4 2H 2 O 10mg, ZnCl 2 20mg, LiCl 5mg, SnCl 2 2H 2 O 5 mg, H 3 BO 3 10mg, KBr 20mg, KI 20mg, EDTANa-Fe 3+ 8g.

[0038] 1. Spore culture: connect the myxobacterium Somatella cellulosus to the VY / 2 solid slant medium and cultivate it for 5-7 days. After forming a diffuse colony, use it as a seed. The composition of VY / 2 medium is: 0.5wt% fresh yeast, 0.1wt% CaCl 2 2H 2 O, 0.5mg / L VB 12 , 1.5wt% agar, adjust the pH to 7.2 with KOH, and sterilize at 121°C for 20min.

[0039] 2. Seed cultivation:

[0040] i. Use an inoculation needle to scrape a few colonies on the slant of Myxobacterium cellulosus VY / 2 and connect them to the M7 liquid seed medium. The bott...

Embodiment 6

[0046] Except for the fermentation medium used, the operating steps and conditions of spore culture, seed culture, and result analysis are the same as in Example 5.

[0047] Wherein, the composition of fermentation medium is: 0.8wt% potato starch, 0.2wt% soybean peptone, 0.2wt% glucose, 0.2wt% yeast extract, 0.1wt% MgSO 4 ·7H 2 O, 0.1wt% CaCl 2 2H 2 O, 1ml / L trace element solution, 2% (V:V) XAD-16 resin and 2-10mg / L threonine; the pH value is 7.0.

[0048] Result analysis The output of epothilone B is shown in the following table:

[0049] Threonine (mg / L)

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Abstract

The invention discloses a sorangium cellulosum fermentation medium, which comprises a carbon source, a nitrogen source, inorganic salts, a macroporous absorption resin, and water. The fermentation medium also comprises amino acid. When the fermentation medium is used for preparing the epothilone B by fermentation culture of sorangium cellulosum, the performance of strains is stable, the growth is rapid, and the yield of the product epothilone B is obviously improved. The invention also provides a fermentation method for producing the epothilone B by the fermentation culture of the sorangium cellulosum by using the sorangium cellulosum fermentation medium.

Description

technical field [0001] The invention relates to a fermentation method and a fermentation medium for producing epothilone B by S. cellulosus. Background technique [0002] Epothilone B, as a secondary metabolite of the myxobacterium S. cellulosus, is a new type of anti-tumor active substance with ultra-stable microtubules. The mechanism of action of epothilone B is the same as that of paclitaxel, but it is superior to paclitaxel in terms of anti-tumor spectrum, anti-tumor activity, safety, water solubility and synthesis method. It is a new type of anti-tumor drug with broad prospects. [0003] For the large-scale production of epothilone, biological fermentation is an ideal way. Epothilone B is generally produced by fermentation of the myxobacterium Sorangium cellulosum 90. It is reported in patent WO93 / 10121 that a low-yield epothilone B can be produced by cultivating Sonocystis cellulosus strains in a medium containing carbon sources, nitrogen sources and inorganic salts,...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P17/18C12R1/01
Inventor 陈少欣田刚
Owner SHANGHAI INST OF PHARMA IND
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