Method of separating and purifying bacillomycin Lb from amyloliquefaciens bacilli

A technology of amylolytic spores and bacitracin, applied in peptide preparation methods, chemical instruments and methods, bacitracin, etc., can solve the problems of limited chromatographic columns, environmental protection, and high cost, and achieve the effect of rapid separation and purification

Active Publication Date: 2019-08-13
HUNAN NORMAL UNIVERSITY
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Problems solved by technology

However, these methods use a large amount of acetone, which is costly and not environmentally friendly; the acid precipitation method has limited suitable chromatographic columns and slow preparation speed

Method used

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  • Method of separating and purifying bacillomycin Lb from amyloliquefaciens bacilli
  • Method of separating and purifying bacillomycin Lb from amyloliquefaciens bacilli
  • Method of separating and purifying bacillomycin Lb from amyloliquefaciens bacilli

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Embodiment

[0028] This embodiment includes the following steps:

[0029] 1. Preparation of crude lipopeptide extract

[0030] (1) DM301 macroporous adsorption resin (Shanghai Maiteng Biotechnology Co., Ltd.) was activated with absolute ethanol, and then washed with double distilled water until no alcohol smell, to obtain activated DM301 macroporous adsorption resin;

[0031] (2) Transfer Bacillus amyloliquefaciens BaX030 (preservation number CCTCC No: M 2014159, see CN104630111A) to LB medium (1wt% NaCl, 0.5wt% yeast extract, 1wt% tryptone) at a ratio of 1wt% , shake culture at 30°C and 120rpm / min for 40-48h, centrifuge to collect the fermentation supernatant, add the fermentation supernatant to the activated DM301 macroporous adsorption resin obtained in step (1), the activated DM301 macroporous adsorption resin The weight of the resin is equivalent to 5 times the volume of the fermentation broth, and placed at 4 ° C for 48 hours to obtain the adsorption resin Ⅰ;

[0032] (3) After th...

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Abstract

Provided is a method of separating and purifying bacillomycin Lb from amyloliquefaciens bacilli. The method comprises the following steps of BaX030 fermentation supernate is adsorbed by macroporous adsorption resin and subjected to ethanol elution and decompress concentration, and a crude extract product is obtained; the crude extract product is subjected to chromatographic separation one time byadopting a RPC reversed phase column at -KTA avant 25, fractions are collected and merged, and decompress concentration is conducted to obtain a bacillomycin Lb crude product; the bacillomycin Lb crude product is purified for the second time by adopting the RPC reversed phase column and subjected to decompress concentration, and the target compound bacillomycin Lb is obtained. The method has the advantages that a macroporous resin adsorption method is adopted to separate and purify the bacillomycin Lb, the technology is simple and easy to implement, and the obtained lipopeptide compound bacillomycin Lb is easy to recycle and reuse, low in production cost and suitable for industrial production and has broad-spectrum anti-tumor activity.

Description

technical field [0001] The invention belongs to the technical field of industrial microorganisms, and in particular relates to a method for rapidly separating and purifying an antitumor active substance of bacillomycin Lb (bacillomycin Lb) from bacillus amyloliquefaciens X030. Background technique [0002] Bacillus amyloliquefaciens can produce a variety of lipopeptide antibiotics, such as surfactin, iturin (iturin A and bacimycin) and fengycins, whose structure contains a Cyclic polypeptide composed of 7-10 amino acids and 1 C 13 -C 18 The β fatty acid chain is an important source of clinically used pharmaceuticals or bioactive molecules. Bacitracin Lb belongs to the iturin family lipopeptide, a cyclic peptide composed of asparagine (Asn), tyrosine (Tyr), serine (Ser), glutamic acid (Glu) and threonine (Thr). Heptapeptide, and a β-sheet fatty acid chain is connected to the threonine (Thr) tail. Bacitracin Lb has been recognized as one of the lipopeptides that exert impo...

Claims

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Application Information

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IPC IPC(8): C07K7/58C07K1/22C07K1/16C07K1/36
CPCC07K7/58
Inventor 丁学知路娇扬夏立秋
Owner HUNAN NORMAL UNIVERSITY
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