Preparation method of high purity ramoplanin single components

A single-component, filter aid technology, which is applied in peptide preparation methods, chemical instruments and methods, organic chemistry, etc., can solve the problems of large damage to downstream purification media, inability to realize large-scale production, and decline in silica gel chromatography. , achieve the effects of shortening the process cycle, guaranteeing safety technology, and reducing solvent consumption

Active Publication Date: 2012-12-26
NCPC NEW DRUG RES & DEV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method has the following disadvantages: (1) the filtrate is directly obtained without decolorization and impurity removal, and the crude extract contains more impurities, which causes great damage to the downstream purification medium; (2) silica gel column chromatography uses different concentrations of acid water and Acetonitrile gradient elution, water will cause the decline of silica gel chromatography, and it can be reused after regeneration, which cannot realize large-scale production
The medium is expensive, with a price of more than 10,000 yuan per liter, and the preparation cost is very high

Method used

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  • Preparation method of high purity ramoplanin single components
  • Preparation method of high purity ramoplanin single components
  • Preparation method of high purity ramoplanin single components

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Take 10L of remoplanin fermentation broth, the fermentation unit is 1036μg / mL (calculated as A1+A2+A3), containing 1.55g of A1, 8.08g of A2, and 0.73g of A3. Add 100g of perlite to the fermentation broth at room temperature, stir for 50 minutes, and then filter the plate and frame. The filtrate passes through the macroporous resin D290 column at a flow rate of 1-2BV / h for decolorization, and the decolorization solution is introduced into the macroporous resin D312 column at a flow rate of 1BV / h Carry out adsorption enrichment, after adsorption, use 30% ethanol / water solution and 60% ethanol / water solution for discontinuous gradient desorption, the desorption flow rate is controlled at 0.5BV / h, when HPLC detects that remoplanin flows out, start to collect the desorption liquid, desorption is complete Finally, the stripping solution was concentrated in vacuum at 30°C to a remoplanin concentration of 100g / L-120g / L to obtain a crude extract (see figure 1) 19.2g (51.6% cont...

Embodiment 2

[0039] Take 100L of remoplanin fermentation broth, the fermentation unit is 1137μg / mL (calculated as A1+A2+A3), containing 15.92g of A1, 89.82g of A2, and 7.96g of A3. Add 3Kg of diatomaceous earth to the fermentation broth at room temperature, stir for 90 minutes, and then filter the plate and frame. The filtrate passes through the macroporous resin LX-700 column at a flow rate of 1.5BV / h for decolorization, and the decolorization solution is introduced at a flow rate of 1.5BV / h The macroporous resin HZ816 column is used for adsorption and enrichment. After adsorption, it is desorbed with a discontinuous gradient of 40% methanol / water solution and 60% methanol / water solution. The desorption flow rate is controlled at 0.5BV / h. When HPLC detects that remoplanin flows out, it starts to collect The desorption solution, after the desorption is completed, was concentrated in vacuum at 40°C to a remoplanin concentration of 110g / L-120g / L in the desorption solution to obtain 210.8g of ...

Embodiment 3

[0041] Take 1000L of remoplanin fermentation broth, the fermentation unit is 1058μg / mL (calculated as A1+A2+A3), containing 148.12g of A1, 825.24g of A2, and 84.64g of A3. Add 100Kg of perlite to the fermentation broth at room temperature, stir for 90 minutes, and then filter the plate and frame. The filtrate passes through the macroporous resin LX-700 column at a flow rate of 1.5BV / h for decolorization, and the decolorization solution is introduced into the macropore at a flow rate of 1.5BV / h Resin D312 column for adsorption and enrichment, after adsorption, use 30% ethanol / water solution and 60% ethanol / water solution for discontinuous gradient desorption, the desorption flow rate is controlled at 0.5BV / h-1BV / h, and when remoplanin flows out as detected by HPLC Start to collect the desorption liquid. After the desorption is completed, the desorption liquid is vacuum concentrated at 50°C to a remoplanin concentration of 110g / L-120g / L to obtain 1978.5g of crude extract (50.8% c...

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Abstract

The invention discloses a method of utilizing an Actinoplanes sp. fermentation culture material to prepare three ramoplanin single components. The method includes: conducting decoloration on a ramoplanin fermentation solution through macroporous decolorization resin, then carrying out macroporous resin adsorption, desorption as well as concentration so as to obtain a ramoplanin crude extract containing the three single components: A1, A2, and A3; and dissolving the crude extract with a solvent, then performing chromatographic separation by means of a polymer microsphere, thus obtaining the products of three high purity ramoplanin single components A1, A2 and A3. The invention has the advantage that when using the polymer microsphere to perform chromatographic separation on the product, the sample treatment amount is large, and the obtained single components have high purity. As the price of a polymer microsphere medium is 30% of that of the reverse phase silica gel C18, and the separation effect is equivalent, the preparation cost is greatly reduced.

Description

technical field [0001] The invention belongs to the technical field of industrial microorganisms, and relates to a preparation method of pharmaceutical raw materials, in particular to a preparation method of separating and purifying three single components of remoplanin from a fermentation culture. Background technique [0002] The causative organisms of many clinically serious infections are Gram-positive bacteria. Due to the extensive use of antibiotics, the problem of bacterial resistance has become increasingly prominent. Finding new glycopeptide antibiotics has gradually become an urgent need in the pharmaceutical industry. At present, several promising new antibiotics are under development, and remoplanin is one of them. [0003] The structural formulas of the single components A1, A2 and A3 of remoplanin are as follows: [0004] [0005] Component R molecular weight A1 -CO-CH=CH-CH=CH-CH 2 -CH 2 -CH 2 2540 A2 -CO-CH=CH-CH=CH-CH 2 -...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K9/00C07K1/16
Inventor 段宝玲张雪霞李宁李晓露任风芝陈书红成晓迅林毅王海燕李丽红张金娟林旸张丽张艳立高月麒张艳哲蒋沁董爱华
Owner NCPC NEW DRUG RES & DEV
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