Culture medium for fermenting polymyxin B sulfate and method for fermenting and producing polymyxin B sulfate

A technology of polymyxin sulfate and culture medium, applied in the field of microbial fermentation, can solve problems such as difficult removal of impurities

Active Publication Date: 2017-06-20
SHANDONG LUKANG PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

And at present, industrially adopt microbial fermentation method to produce polymyxin B sulfate, there are also many related substances (impurities) produced in the fermentation process, and many impurities are structural analogues of main components, and these impurities are extracted in the follow-up operation process hard to remove

Method used

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  • Culture medium for fermenting polymyxin B sulfate and method for fermenting and producing polymyxin B sulfate

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Seed solution culture method: dig out about 1cm of polymyxin B sulfate slant culture with an inoculation shovel 2 The size is inserted into the seed medium of the shaker flask, wherein the shaker flask adopts a 500ml Erlenmeyer flask containing 70-80ml of the seed medium, and the shaker rotates at 220rpm and cultures at 29°C for 14-18 hours. The composition of the seed medium is: flour 8g / 100ml, yeast extract powder 0.3g / 100ml, dipotassium hydrogen phosphate 0.015g / 100ml, calcium carbonate 0.3g / 100ml, ammonium sulfate 0.45g / 100ml.

[0052] Put the seed culture of Paenibacillus polymyxa, a polymyxin B sulfate-producing bacterium, into the fermentation medium containing the following ingredients: flour 10g / 100ml, high-temperature amylase 4g / 100ml, cottonseed cake powder 1g / 100ml, corn steep liquor 0.5ml / 100ml, dipotassium hydrogen phosphate 0.015g / 100ml, ammonium sulfate 0.4g / 100ml, calcium carbonate 0.3g / 100ml, defoamer 0.05ml / 100ml, inoculum size 7%, temperature 29°C, ...

Embodiment 2

[0062] Put the seed culture of Paenibacillus polymyxa, a polymyxin B sulfate-producing bacterium, into the fermentation medium containing the following ingredients: flour 8g / 100ml, high-temperature amylase 5g / 100ml, cottonseed cake powder 0.8g / 100ml, corn steep liquor 0.8ml / 100ml, dipotassium hydrogen phosphate 0.012g / 100ml, ammonium sulfate 0.5g / 100ml, calcium carbonate 0.25g / 100ml, defoamer 0.06ml / 100ml, inoculum size 10%, temperature 30°C, culture for 46 hours and then end fermentation.

[0063] The fermentation broth and the extracted finished product were tested by high-pressure liquid phase with the external standard method, and the peak area ratios of the components and impurities are shown in Table 2 below.

[0064] Table 2 The peak area ratio of each component and impurity of the fermentation product in Example 2

[0065] Detection situation B 2P Proportion%

[0066] As can be seen from the results in Table 2, the main impurity B in the finished product ...

Embodiment 3

[0068]Put the seed culture of Paenibacillus polymyxa, a polymyxin B sulfate-producing bacterium, into the fermentation medium containing the following ingredients: flour 11g / 100ml, high-temperature amylase 3.5g / 100ml, cottonseed meal 1.5g / 100ml, corn steep liquor 0.4 ml / 100ml, dipotassium hydrogen phosphate 0.03g / 100ml, ammonium sulfate 0.3g / 100ml, calcium carbonate 0.5g / 100ml, defoaming agent 0.04ml / 100ml, inoculum size 15%, temperature 28°C, culture for 42 hours and end fermentation .

[0069] The fermentation broth and the extracted finished product were tested by high-pressure liquid phase with the external standard method, and the peak area ratios of the components and impurities are shown in Table 3 below.

[0070] Table 3 The peak area ratio of each component of the fermentation product and impurities in Example 3

[0071] Detection situation B 2P Proportion%

[0072] As can be seen from the results in Table 3, the main impurity B in the finished product ...

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Abstract

The invention provides a culture medium for fermenting polymyxin B sulfate. Each 100ml of the culture medium is prepared from the following components: 6g to 12g of flour, 0.03g to 0.06g of high-temperature amylase, 0.4g to 2g of cottonseed meal, 0.3g to 1g of corn pulp, 0.01g to 0.05g of dipotassium phosphate, 0.2g to 0.6g of ammonium sulfate, 0.2g to 0.6g of calcium carbonate, 0.02ml to 0.08ml of a de-foaming agent and the balance of water. A method for fermenting and producing the polymyxin B sulfate comprises the following the steps: inoculating the culture medium with strain seed liquid for producing the polymyxin B sulfate; fermenting and culturing fermentation liquid at 26 DEG C to 32 DEG C, wherein the pH (Potential of Hydrogen) value is 6.7 to 7.4 at the beginning of fermenting; controlling the pH value of the fermentation liquid to be natural during a fermentation process, wherein the dissolved oxygen volume concentration of the fermentation liquid is 10 percent to 30 percent. All the components of the culture medium are cooperatively matched and the content of impurities B2P is controlled to be lower than 1.5 percent.

Description

technical field [0001] The invention belongs to the technical field of microbial fermentation, and in particular relates to a culture medium for fermenting polymyxin B sulfate and a method for fermenting and producing polymyxin B sulfate. Background technique [0002] Polymyxin Bsulfate (PolymyxinBsulfate), also known as colistin B sulfate, is a basic cyclic polypeptide antibiotic composed of amino acids and fats produced by Paenibacillus polymyxa. Lambert-negative bacteria have strong inhibitory or bactericidal effects, such as Escherichia coli, Pseudomonas aeruginosa, paraescherichia coli, Klebsiella pneumoniae, acidophilus bacillus, pertussis bacilli and Shigella, etc., especially for Pseudomonas aeruginosa effect. Many countries in the world have approved the drug as a feed additive. Polymyxin B sulfate is also used as a drug, and it is mainly used clinically for infections caused by sensitive bacteria and urinary system infections, meningitis, sepsis, burn infections,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/04C12N1/20C12R1/01
CPCC07K7/62C12N1/20
Inventor 白芳静郑长春王博闫婷婷晁红英张玲
Owner SHANDONG LUKANG PHARMA
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