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Preparation method of cyclic lipopeptide compound

A compound and amino acid technology, applied in the field of biosynthetic cycloaliphatic peptide compounds, can solve the problems of unfavorable dissolved oxygen control, subsequent filtration operation, and high medium viscosity, and achieve easy dissolved oxygen control, reduce production costs, and reduce environmental damage. Effect

Active Publication Date: 2012-08-01
SHANGHAI TECHWELL BIOPHARMACEUTICALS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] However, the yield of the compound of formula I needs to be further improved, and the viscosity of the above-mentioned medium is still very high, and the bacteria grow in a spherical shape during the fermentation process, which is not conducive to the dissolved oxygen control and subsequent filtration operation during the fermentation process.

Method used

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  • Preparation method of cyclic lipopeptide compound
  • Preparation method of cyclic lipopeptide compound
  • Preparation method of cyclic lipopeptide compound

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] Fermentation Strain Preparation Coleophoma empetri F-11899 (FERM BP2635)

[0059] The seed liquid of Coleophoma empetri F-11899 (FERMBP2635) was prepared with reference to the cultivation method of seeds in the document Scale-up fermentation of echinocandin type antibiotic FR901379 (Journal of Bioscience and Bioengineering, VOL 109 No. 2, 138-144, 2010), for later The compound of formula I is used for fermentation.

[0060] Potato dextrose agar medium (PDA) was used as the slant medium, and its composition was: 30% potato, 2% glucose, and 1.5% agar.

[0061] Seed medium composition: sucrose 1%, cottonseed meal 2%, dry yeast 1%, peptone 1%, KH 2 PO 4 0.2%, CaCO 3 0.2%, defoamer 0.05%.

[0062] Strain Coleophoma empetri F-11899 (FERM BP2635) matures after being cultivated on a slant at 25°C for 6-10 days, picks mature mycelia or spores and inserts them into the seed medium, and then cultivates them on a shaker at 25°C with a rotation speed of 280RPM for 2 -4 days. ...

Embodiment 2

[0064] Fermentation strain preparation (mutagenic strain CGMCC 4129)

[0065] With reference to the cultivation method of seeds in the document Scale-up fermentation of echinocandin type antibiotic FR901379 (Journal of Bioscience and Bioengineering, VOL 109 No.2, 138-144, 2010), the seed liquid of the mutagenized bacterial strain CGMCC 4129 was prepared to prepare the following formula I compound For fermentation.

[0066] Potato dextrose agar medium (PDA) was used as the slant medium, and its composition was: 30% potato, 2% glucose, and 1.5% agar.

[0067] Seed medium composition: sucrose 1%, cottonseed meal 2%, dry yeast 1%, peptone 1%, KH 2 PO 4 0.2%, CaCO 3 0.2%, defoamer 0.05%.

[0068] Strain CGMCC 4129 matures after being cultivated on a slant at 25°C for 6-10 days, picks mature mycelia or spores and inserts them into the seed medium, and then cultivates them on a shaker at 280RPM for 2-4 days at 25°C.

Embodiment 3

[0070] Preparation of compounds of formula I

[0071] In a 250ml shake flask, add 50ml containing mannitol concentration 5%, yeast extract concentration 0.5%, L-proline concentration 1%, cottonseed cake powder concentration 1%, ammonium sulfate concentration 0.1%, magnesium sulfate concentration 0.06%, Trace element solution concentration 0.1%, the culture medium of MES concentration 2%, adjust pH to be 5.5 ± 0.5, 121 ℃ of sterilization 30min, the seed 1ml (2%) that obtains in embodiment 1 is inoculated in this nutrient solution, 25 ℃ After cultivating at 280r / m for 240 hours, the cultivation was completed, and sampling analysis showed that the viscosity of the final culture solution was 2200cp, and the content of the compound of formula I was 0.5g / L.

[0072] Trace elements: FeSO 4 ·7H 2 O 10g / L, MnSO 4 ·H 2 O 10g / L, ZnSO 4 ·7H 2 O 2g / L, CaCl 2 0.7g / L,H 3 BO 3 0.56g / L, CuCl 22H 2 O 0.25g / L, (NH 4 ) 6 Mo 7 o 24 ·7H 2 O0.19g / L, concentrated hydrochloric acid 5...

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Abstract

The invention discloses a fermentation medium which comprises 0.5-5.0 wt% of amino acid or its derivative, 0.5-3.0 wt% of hard-soluble organic nitrogen source, and 1.0-10.0 wt% of carbon source. By using the medium of the invention to carry out biologic fermentation of the compound of formula I, the fermentation level can stably reach more than 0.5g / L, with the highest fermentation level being 1.0g / L. By using the strain mutagenized with Coleophoma empetri F-11899 (FERM BP2635), the fermentation level can even reach 1.5 g / L. Because the fermentation level is high and the amount of organic solvent used in the post-treatment is relatively reduced, the production cost is reduced, and the invention has high economic value and meets the need of industrial production.

Description

technical field [0001] The present invention relates to a method for biosynthesizing cyclolipopeptide compounds, in particular to a method for preparing compounds of formula I through biological fermentation. Background technique [0002] Fungal infection has become the main cause of high morbidity and mortality in immunocompromised patients. The incidence of fungal infections has increased significantly over the past 20 years. High-risk groups for fungal infections include critically ill patients, surgical patients, and those with HIV infection, leukemia, and other neoplastic diseases. Those who have undergone organ transplantation are also at high risk of fungal infection. [0003] Echinocandins, a new class of antifungal drugs, are effective in the treatment of infections caused by Candida or Aspergillus. This class of drugs is represented by caspofungin and micafungin. Echinocandin drugs inhibit fungi by inhibiting the formation of 1,3-β glycosidic bonds, thereby bet...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/02C12R1/645
CPCC07K7/56C07K7/54C12R1/645C12N1/14C12P7/64A61P31/10C12R2001/645C12N1/145Y02A50/30C12P21/02
Inventor 刘石东张兆利陈懿王修胜周亮亮季晓铭
Owner SHANGHAI TECHWELL BIOPHARMACEUTICALS CO LTD
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