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High purity lipiarmycin A4 preparation method

A leap year mycin, high-purity technology, applied in the field of biochemistry, to achieve the effect of simple operation, strong controllability, and elimination of interference

Active Publication Date: 2015-04-08
NCPC NEW DRUG RES & DEV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, the research on leap year mycin A4 is mainly concentrated in the field of its antibacterial activity research, but few people have conducted in-depth discussion and research on the preparation method of leap year mycin A4

Method used

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  • High purity lipiarmycin A4 preparation method
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  • High purity lipiarmycin A4 preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Preparation of leap year mycin A4 fermentation broth:

[0028] Inoculate the seed medium with Actinomycetes mobilis strain N12W0304, and shake the flask for 48 hours under the conditions of 30° C. and 220 rpm to obtain seed liquid. The seed solution was inoculated into a 50L fermenter with a fermentation medium at an inoculation amount of 4% by volume, and fermented for 190 hours at a tank temperature of 30°C, a tank pressure of 0.05±0.01Mpa, a ventilation rate of 25L / min, and a rotation speed of 400rpm. Obtain the leap year mycin A4 fermentation broth.

[0029] Wherein the preparation method of the seed medium is: beef extract 4.0g, peptone 4.0g, NaCl 2.5g, yeast powder 1.0g, soybean powder 10.0g, glucose 50.0g, CaCO 3 5.0g, add tap water to dissolve, dilute to 1000ml, add alkali to adjust the pH value to 7.0-7.2, and sterilize at 121°C for 30min.

[0030] The preparation method of the fermentation medium is: 250g of starch, 500g of glucose, 200g of wheat germ powde...

Embodiment 2

[0042] The leap year mycin A4 fermentation broth cultivation of the present embodiment is identical with embodiment 1, and the difference between the present embodiment and embodiment 1 mainly lies in the change of purification conditions, the purification method of the leap year mycin A4 of the present embodiment is as follows:

[0043]a. Take 20 L of Leapnianmycin A4 fermentation broth, and the fermentation unit is 506 μg / ml. Leapnianmycin A4 fermentation broth was filtered to obtain 4.0kg mycelia. Add 12.0 L of ethanol with a volume ratio concentration of 95% to the above-mentioned mycelia, stir and extract for 3 hours, and then filter to obtain Leapnianmycin A4 extract.

[0044] b. Concentrate the extract under reduced pressure at 40°C and a vacuum greater than 0.08Mpa until the ethanol concentration is less than 10%, to obtain 1360ml of concentrated solution, add 2700ml of ethyl acetate to extract for 3 hours, concentrate and dry to obtain 12.8g of Leapnianmycin A4 crude ...

Embodiment 3

[0048] The leap year mycin A4 fermentation broth cultivation of the present embodiment is identical with embodiment 1, and the difference between the present embodiment and embodiment 1 mainly lies in the change of purification conditions, the purification method of the leap year mycin A4 of the present embodiment is as follows:

[0049] a. Take 15 L of Leapnianmycin A4 fermentation broth, and the fermentation unit is 482 μg / ml. Leapnianmycin A4 fermentation broth was filtered to obtain 3.2kg mycelium. Add 6.4 L of acetone aqueous solution with a volume ratio concentration of 90% to the above-mentioned mycelia, stir and extract for 4 hours, and then filter to obtain the leach solution of Leapnianmycin A4.

[0050] b. Concentrate the extract under reduced pressure at 50°C and a vacuum greater than 0.08Mpa until the concentration of acetone is less than 10%, to obtain 1120ml of concentrated solution, add 1120ml of ethyl acetate to extract for 4 hours, concentrate and dry to obta...

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Abstract

The invention discloses a high purity lipiarmycin A4 preparation method which comprises the following steps: A, filtering lipiarmycin A4 fermentation broth to obtain mycelium, soaking the mycelium with a polar solvent for solid-liquid separation to obtain lipiarmycin A4 extracting solution; B, concentrating the lipiarmycin A4 extracting solution, extracting the lipiarmycin A4 extracting solution with ethyl acetate, concentrating and drying the extracting solution to obtain lipiarmycin A4 coarse powder; C, dissolving the lipiarmycin A4 coarse powder in an organic solvent, filtering with a 0.45 mum filter membrane to obtain a sample loading solution; and D, adding the sample loading solution into a medium pressure chromatography column filled with C18, adding an elution solvent, collecting eluant with the content greater than or equal to 98.5% by high performance liquid chromatography, concentrating, filtering and drying in vacuum to obtain a high purity lipiarmycin A4 product. The high purity lipiarmycin A4 preparation method has the advantages of simple operation, high sample recovery rate, and suitability for high purity antagonizing drug-resistance antibiotic production.

Description

technical field [0001] The invention belongs to the technical field of biochemistry, and relates to a preparation method of a macrolide compound, more precisely, relates to a preparation method of leapyearmycin A4. Background technique [0002] Leapyearmycin A4 is a new type of macrolide antibiotic with 18-membered ring structure produced by the fermentation of cystic bacteria. It is a narrow-spectrum antibacterial compound and has excellent antibacterial activity against Gram-positive aerobic and anaerobic bacteria effect. Its molecular formula is C 51 h 72 C l2 o 18 , the molecular weight is 1044.04, and the structural formula is as shown in formula I: [0003] [0004] Leapyearmycin A4 has the same antibacterial spectrum as Fidaxomycin, although its activity is slightly lower than that of Fidaxomicin (CN201210032395.4), but compared with the disclosed S-Tiacumicin B and C-19 ketones, Leapyearmycin A4 The activity against C. difficile strains in vitro is the best,...

Claims

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Application Information

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IPC IPC(8): C07H17/08C07H1/08A61P31/04
CPCC07H1/08C07H17/08
Inventor 张雪霞李晓露任风芝王海燕高月麒林旸张丽段宝玲
Owner NCPC NEW DRUG RES & DEV
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