Producing strain of oritavancin intermediate and application of producing strain

A technology of oritavancin and intermediates, which is applied in the direction of bacteria, microorganisms, and methods based on microorganisms, can solve the problems that cannot meet the needs of scientific research and actual production, the difficulty of separating target products, and the low probability of natural mutations. Improve the yield of the extraction step, reduce the cost of fermentation and extraction, and reduce the effect of extraction cost

Pending Publication Date: 2021-10-12
SHANGHAI JIANHE PHARM & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the fermentation process reported in this patent and subsequent documents has the following three defects: the one is that the target component A82846B fermentation titer is low (150mg/L); Expensive raw materials lead to high fermentation costs; the third is that the fermentation broth contains three homologues: A82846A, A82846B, and A82846C (respectively referred to as "A

Method used

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  • Producing strain of oritavancin intermediate and application of producing strain
  • Producing strain of oritavancin intermediate and application of producing strain
  • Producing strain of oritavancin intermediate and application of producing strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0069] Obtainment of strain JH100-32B16B (CGMCC No.17931).

[0070] JH100 strain was obtained by natural isolation from a strain of Nocardia orientalis producing A82846 series compounds (Hamill et al. A82846 ANTIBIOTICS [P]. USP 5312738, May 17, 1994). Taking JH100 as the starting strain, the subsequent mutagenesis breeding operation was started.

[0071]Cell preparation: First, dilute the JH100 liquid culture with sterile water, add glass beads into the dilution test tube, shake fully on the shaker to ensure that the hyphae are dispersed; use a sterile graduated pipette to draw about 0.1mL of appropriate dilution The bacterial suspension was added dropwise on a sterile ISP2 plate, and coated with an L-shaped glass spatula. Prepare several coating plates by following this procedure.

[0072] The first round of mutagenesis: the above-mentioned starting strain (JH100) coated plate was placed under ultraviolet light irradiation. The parameters of the ultraviolet lamp irradiati...

Embodiment 2

[0080] Identification of strain JH100-32B16B (CGMCC No.17931).

[0081] (1) Morphological characteristics of strain JH100-32B16B

[0082] After culturing the strain JH100-32B16B on solid media (plates) with different formulations at 28°C for 5-7 days, observe the colony morphology, see Table 1:

[0083] Table 1

[0084]

[0085] The aerial hyphae of strain JH100-32B16B were picked and observed under an optical microscope. The characteristics were: spore filaments were straight, flexible, hooked, loose and tight spiral; spores were cylindrical and had cystic structures.

[0086] (2) Culture and physiological and biochemical characteristics of strain JH100-32B16B

[0087] The physiological and biochemical characteristics of the strain JH100-32B16B were investigated during the culture process. Table 2 shows the carbon source utilization characteristics; Table 3 shows the nitrogen source utilization characteristics of the strain; Table 4 shows other physiological and biochemi...

Embodiment 3

[0102] Shake flask fermentation of control strain JH100. The JH100 strains preserved in glycerol tubes were transferred to the ISP-2 slant, and placed in a constant temperature and humidity incubator at 28°C for activation and culture for 5-7 days. Pick the activated bacterial lawn and transfer it to the seed bottle. The liquid volume of the seed bottle is: 30mL seed medium in a 250mL triangular flask. The formula of the seed medium is (g / L): glucose 15, maltodextrin 15, hydrolyzed casein 3, yeast extract 3, pH 7.2. The seed bottle was shaken and cultivated on a constant temperature shaker at 28° C. for 48 hours, and the shaker rotated at 200 rpm. The mature seeds were transferred to fermentation shake flasks for shake flask fermentation. The shake flask fermentation medium formula is (g / L): glucose 30, maltodextrin 30, molasses 25, hydrolyzed casein 10, yeast extract 10, cottonseed cake powder 10, pH 7.5. The amount of liquid in the fermentation bottle is 20mL medium in a...

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Abstract

The invention discloses a producing strain of an oritavancin intermediate (A82846B) and application of the producing strain. The producing strain kibdelosporangium aridum JH100-32B16B of the oritavancin intermediate is preserved in China General Microbiological Culture Collection Center (CGMCC), and the preservation number is CGMCC No.17931. The fermentation method is a method for producing the oritavancin intermediate by fermenting the kibdelosporangium aridum CGMCC No.17931 in a fermentation culture medium. By adopting the producing strain and a corresponding fermentation process thereof, the fermentation titer of A82846B can be 1450mg/L on a 30L fermentation tank by adopting a fermentation culture medium with a low-cost formula, and the fermentation cost is reduced by more than 90% compared with that of a contrast process; and the "impurity-product ratio" (A/B ratio) is as low as 1.5-2.0, and is reduced by more than half compared with a control level, thereby being beneficial to simplifying a product extraction process and reducing the extraction cost. The advantages in two aspects are combined, so that the comprehensive production cost of the A82846B is obviously lower than a control process level reported in the existing literature.

Description

technical field [0001] The present invention relates to a production bacteria selection and fermentation technology of a pharmaceutical intermediate, in particular to a microbial strain breeding for producing oritavancin intermediate A82846B component and a corresponding fermentation process. Background technique [0002] At present, in the field of antibacterial infection drugs, β-lactam broad-spectrum antibiotics represented by penicillins and cephalosporin antibiotics are widely used in clinical practice. Due to long-term and high-dose inappropriate use, a large number of drug-resistant pathogenic bacteria have emerged in recent years and have continued to evolve, making traditional first-line antibiotics encounter the dilemma of poor or even ineffective antibacterial effects. [0003] Glycopeptide antibiotics have a good antibacterial effect on common drug-resistant bacteria with β-lactamase due to their different mechanisms of action from β-lactam antibiotics. As a rep...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P21/02C12R1/01
CPCC12N1/20C07K9/008
Inventor 赵宏程陈义朗郭文清
Owner SHANGHAI JIANHE PHARM & TECH CO LTD
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