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A kind of method for separating and extracting epothilone b from fermented broth of Cystobacter celluli

A technology of S. cellulosus and epothilone, which is applied in the extraction of epothilone B and the field of separation and extraction of epothilone B, can solve the problems of low operation efficiency, low product yield, and high process cost. Achieve the effect of increasing production, high product purity and reducing production cost

Active Publication Date: 2015-09-30
SHAANXI UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The method includes four steps of resin adsorption, silica gel column gradient separation, molecular sieve chromatography and high-performance liquid chromatography separation. It has certain defects, and the steps are too complicated, so that the operation efficiency is low and the process cost is high. The yield of the product lower

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] A method for isolating and extracting epothilone B from S. cellulosus fermentation broth, comprising the following steps:

[0031] 1. Fermentation and cultivation of S. cellulosus

[0032] Shake flask culture: first, inoculate the S. cellulosus strain ATCC25532 into a Erlenmeyer flask equipped with M26 liquid medium, and culture it on a shaker at 30°C and 200 rpm for 72 hours to obtain a shake flask seed solution;

[0033] Wherein, the composition of the M26 liquid medium is: potato starch 7.5-8.0g / L, soybean peptone 1.5-2.0g / L, glucose 1.5-2.0g / L, yeast powder 1.5-2.0g / L, MgSO 4 ·7H 2 O1.0~1.2g / L, CaCl 2 1.0~1.2g / L, EDTA-Fe 3+ 1mL / L, trace elements 1mL / L, pH 7.2;

[0034] Primary seed tank culture: Inoculate the obtained shake flask seed liquid into a 5L fermenter equipped with the above-mentioned M26 liquid medium, at a temperature of 30°C, a rotation speed of 150rpm, an air flow ratio of 1:0.5, and a tank pressure of 0.04~0.1kg / cm 2 , under the condition that t...

Embodiment 2

[0047] A method for isolating and extracting epothilone B from S. cellulosus fermentation broth, comprising the following steps:

[0048] 1. Fermentation and cultivation of S. cellulosus

[0049] Shake flask culture: first, inoculate the S. cellulosus strain ATCC25569 into an Erlenmeyer flask equipped with M26 liquid medium, and culture it on a shaker at 30°C and 180rpm for 84 hours to obtain a shake flask seed solution;

[0050] Wherein, the composition of described M26 liquid culture medium is identical with embodiment 1;

[0051] Primary seed tank cultivation: Inoculate the obtained shake flask seed liquid into a 5L fermenter equipped with the above-mentioned M26 liquid medium, at a temperature of 30°C, a rotation speed of 200rpm, an air flow ratio of 1:0.5, and a tank pressure of 0.05kg / cm 2 , under the condition that the pH value is 7.2, cultivate for 84 hours to obtain the first-grade strain liquid;

[0052] Secondary seed tank cultivation: inoculate vigorously growin...

Embodiment 3

[0064] A method for isolating and extracting epothilone B from S. cellulosus fermentation broth, comprising the following steps:

[0065] 1. Fermentation and cultivation of S. cellulosus

[0066] Shake flask culture: first, inoculate the S. cellulosus strain ATCC25532 into a Erlenmeyer flask equipped with M26 liquid medium, and culture it on a shaker at 30°C and 220rpm for 96 hours to obtain a shake flask seed solution;

[0067] Wherein, the composition of described M26 liquid culture medium is identical with embodiment 1;

[0068] Primary seed tank culture: Inoculate the obtained shake flask seed liquid into a 5L fermenter equipped with the above-mentioned M26 liquid medium, at a temperature of 30°C, a rotation speed of 160rpm, an air flow ratio of 1:0.5, and a tank pressure of 0.1kg / cm 2 , under the condition that the pH value is 7.2, cultivate for 96 hours to obtain the first-grade strain liquid;

[0069] Secondary seed tank cultivation: inoculate vigorously growing prim...

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Abstract

The invention provides a method for separation and extraction of epothilone B from a sorangium cellulosum fermentation liquid, and belongs to the microbial fermentation pharmacy field. The method comprises the steps: after shake-flask cultivation of sorangium cellulosum strains, amplifying step by step, fermenting sorangium cellulosum, followed by carrying out macroporous resin adsorption, ethyl acetate desorption and methanol redissolution, and thus obtaining a crude product; carrying out column chromatography and HPLC analysis of the crude product to obtain a preliminary component of epothilone B, then adding a molecularly imprinted polymer for specific adsorption of epothilone B, and finally carrying out methanol elution and crystallization to obtain epothilone B. Sorangium cellulosum is produced by step-by-step fermentation, and at the same time, the molecularly imprinted polymer with the specific targeted adsorption capacity is adopted in the separation and extraction process, not only has a strong adsorption capacity on epothilone B, but also can easily extract epothilone B, so as to reduce the extraction cost of separation and extraction of epothilone drugs, and have more wide industrialized application prospects and economic benefits.

Description

technical field [0001] The invention belongs to the field of microbial fermentation pharmaceuticals, and relates to a method for extracting epothilone B, in particular to a method for separating and extracting epothilone B from a fermentation liquid of S. cellulosus. Background technique [0002] Epothilone (epothilone) is a class of macrolide compounds with multiple biological activities and can be used as antibiotic drugs. It was developed by G. It was first reported by et al. in 1993. Epothilone can be isolated from the fermentation liquid of the Myxobacteria suborder S. cellulosus strain, and its main components are Epothilone A and B. [0003] Soon after, the Merck laboratory discovered epothilone again in the experiment of screening paclitaxel analogues from 7,000 natural extracts. And found that it has the same mechanism of action as paclitaxel, can promote GTP (guanosine triphosphate)-dependent tubulin polymerization to form microtubules, and has a stabilizing eff...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P17/18C07D493/04C12R1/01
Inventor 龚国利赵婷峰李慧
Owner SHAANXI UNIV OF SCI & TECH
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