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Separation and purification method of all component monomers of Dalbavancin key intermediate A40926

An A40926, separation and purification technology, used in the field of medicine, can solve the problems of high cost, difficult to industrialize amplification, small sample loading volume, etc., and achieve the effects of improved efficiency, large sample loading, and safe operation.

Pending Publication Date: 2022-07-01
上海健启生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Aiming at the deficiencies of the prior art, the present invention provides a method for the separation and purification of each component monomer of the key intermediate A40926 of dalbavancin, which has the advantages of large sample loading in a single batch, high efficiency, low pollution, low cost, and easy operation. The advantages of safety and suitability for industrial production solve the problem of gel filtration, macroporous adsorption resin combined with reversed-phase chromatography with small sample loading volume, high requirements for column packing, low sample loading, difficult to achieve industrial scale-up, and high cost. There are problems of environmental pollution and potential safety hazards

Method used

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  • Separation and purification method of all component monomers of Dalbavancin key intermediate A40926

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Effect test

Embodiment 1

[0029] The 35L fermentation broth was added with 2mol / L sodium hydroxide solution to adjust the pH value to 12.0, and stirred for 30min. Take 1ml of the sample and centrifuge to take the supernatant for HPLC analysis and determination. The A40926 B0 fraction is 42.8g. Pour the fermentation broth into the ceramic membrane filtration equipment (ceramic membrane pore size is 0.1um), and control the temperature of the fermentation broth to be lower than 30°C for filtration. When the volume of the fermentation broth is lower than 20L, slowly add 180L of alkaline water with a pH value of 12.0 to maintain fermentation. The liquid volume is about 20L. The filtrate after the ceramic membrane filtration was adjusted to pH 8.0 with 1 mol / L hydrochloric acid, and the sample was loaded on a polyamide column (15CM*70CM) at a speed of 4BV / hr. After the sample was loaded, 3BV pure water, 5BV 1% ammonia water and 2% ammonia water of 5BV were eluted, and the eluates containing A40926 were comb...

Embodiment 2

[0031] The 50L fermentation broth was added with ammonia water to adjust the pH value to 11.0, and stirred for 30min. Take 1ml of the sample and centrifuge to take the supernatant for HPLC analysis and determination. The A40926 B0 fraction is 57.5g. Pour the fermentation broth into the ceramic membrane filtration equipment (ceramic membrane pore size 0.2um), control the temperature of the fermentation broth to be lower than 30°C for filtration, when the volume of the fermentation broth is lower than 20L, slowly add 180L of alkaline water with a pH value of 11.0 to maintain fermentation The liquid volume is about 20L. The filtrate after the ceramic membrane filtration was adjusted to pH 7.0 with 1 mol / L sulfuric acid, and the sample was loaded on a polyamide column (15CM*70CM) at a speed of 5BV / hr. After the sample was loaded, 3BV pure water, 10BV 10 BV of NaOH aqueous solution at pH 10, 10BV of pH 12 aqueous NaOH solution. The eluates containing A40926 were pooled. The elua...

Embodiment 3

[0033]45 L of fermentation broth was added with 1 mol / L potassium hydroxide solution to adjust the pH value to 13.0, and stirred for 30 min. Take 1ml of the sample and centrifuge to take the supernatant for HPLC analysis and determination. The A40926 B0 fraction is 52.9g. Pour the fermentation broth into the ceramic membrane filtration equipment (ceramic membrane pore size 0.05um), control the temperature of the fermentation broth to be lower than 25°C for filtration, when the volume of the fermentation broth is lower than 20L, slowly add 200L of alkaline water with a pH value of 13.0 to maintain fermentation The liquid volume is about 20L. The filtrate after the ceramic membrane filtration was adjusted to pH 7.0 with 1 mol / L phosphoric acid, and the sample was loaded on a polyamide column (15CM*70CM) with a sample loading speed of 3BV / hr. 5% urea in water and 8BV in 20% urea in water. The eluates containing A40926 were pooled. The eluate was adjusted to pH 3.2 with phospho...

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Abstract

The invention relates to the technical field of medicines, and discloses a separation and purification method of monomer components of a key intermediate A40926 of dalbavancin, which comprises the following steps: adjusting the pH value of fermentation liquor, and performing membrane filtration to obtain an extracting solution of A40926; the A40926 extracting solution is subjected to polyamide chromatographic purification and isoelectric precipitation, and a pasty A40926 semi-finished product is obtained; after the semi-pure product is dissolved, components such as high-purity A40926B0 and the like can be obtained through ion exchange chromatography purification, isoelectric precipitation, ethanol dehydration and vacuum drying. The separation and purification method of each component monomer of the Dalbavancin key intermediate A40926 comprises the following steps: filtering a fermentation broth by using a ceramic membrane, purifying isoelectric precipitation by using polyamide chromatography to obtain a pasty semi-pure product, purifying by using ion exchange chromatography, carrying out isoelectric precipitation, dehydrating by using ethanol, and carrying out vacuum drying to obtain high-purity A40926B0 and other components. The adopted equipment can be used for treating samples on a large scale, industrial amplification is conveniently realized and is mainly completed in a water phase, and compared with a traditional process, the method is safer and more environment-friendly.

Description

technical field [0001] The invention relates to the technical field of medicine, in particular to a method for separating and purifying each component monomer of dalbavancin key intermediate A40926. Background technique [0002] Dalbavancin is a novel second-generation semi-synthetic glycopeptide antibiotic with similar structure to vancomycin and teicoplanin. It is used for severe infections caused by Gram-positive pathogens. It is synthesized from A40926 as a precursor. of. A40926 is a glycopeptide substance discovered by scientists in 1984 when culturing and isolating Madura strains from soil. A40926 has 5 main components: PA, PB, A, B0 and B1, B0 and B1 are collectively referred to as B component. Dalbavancin is obtained by the modification of the secondary metabolite A40926 B produced by bacteria after chemical synthesis. Compared with vancomycin and teicoplanin, it has a wider antibacterial spectrum and is effective against staphylococcus, streptococcus, intestinal ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K9/00C07K1/18C07K1/22C07K1/30C07K1/34C07K1/36C07K1/14
CPCC07K9/008
Inventor 谭鑫张长清王小曼茅缪伟张立俊
Owner 上海健启生物科技有限公司
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