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Process for preparing rapamycin by using microbial fermentation method

A microbial fermentation method, a technology of rapamycin, applied in the directions of microorganism-based methods, microorganisms, biochemical equipment and methods, etc., can solve problems such as unseen production, reduce consumption costs, save time, and be suitable for industrial production. Effect

Active Publication Date: 2013-10-23
中科医药行业生产力促进中心有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In 1993, Kojima et al. reported that actinomycete N902-109 [Actino-planessp.N902-109 (FERM BP-3832)] can also synthesize rapamycin, and its synthesis ability is more than 10 times stronger than that of Streptomyces hygroscopicus ATCC 29253 , but there is no report that this strain is used for production

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Inoculate Streptomyces hygroscopicus, CGMCC No.5145, on an oat-potato slant at 28°C for 15 days, and black spots appear on the surface of the slant lawn. At this time, transfer it to a seed bottle; take a ring from the slant with an inoculation shovel Inoculate into an Erlenmeyer flask containing a seed medium, and cultivate at 28° C. and 200 rpm for 60 hr. The composition of the seed medium, mass (g) / volume (L) ratio: soluble starch 10.0, peptone 6.0, yeast extract 6.0, acid hydrolyzate 1.5, and the rest is water; the above cultured strains are transferred according to the inoculum size of 2%. Put it into a 100L fermenter (with 60L of sterilized fermentation medium inside), adjust the fermentation temperature to 28°C, the ventilation rate to 0.5v / min, the stirring speed to 250rmp, and ferment for 3 days. Among them, the composition of fermentation medium, mass (g) / volume (L) ratio: glucose 40, glycerol 10, soybean cake 10, yeast extract 6.0, peptone, L-Lysine 0.15, K ...

Embodiment 2

[0039] Inoculate Streptomyces hygroscopicus, CGMCC No.5145, on an oat-potato slant at 28°C for 15 days, and black spots appear on the surface of the slant lawn. At this time, transfer it to a seed bottle; take a ring from the slant with an inoculation shovel Inoculate into an Erlenmeyer flask containing a seed medium, and cultivate at 28° C. and 200 rpm for 60 hr. Among them, the composition of the seed medium, mass (g) / volume (L) ratio: soluble starch 10.0, peptone 6.0, yeast extract 6.0, acid hydrolyzate 1.5 and the rest is water; transfer the above cultured strains into In a 100L fermenter (with 60L sterilized fermentation medium inside), adjust the fermentation temperature to 28°C, the ventilation rate to 0.5v / min, the stirring speed to 250rmp, and ferment for 4 days. Among them, the composition of fermentation medium, mass (g) / volume (L) ratio: glucose 40, glycerol 10, soybean cake 10, yeast extract 6.0, peptone, L-Lysine 0.15, K 2 HPO 4 5.0, KH 2 PO 4 5.0, NaCl5.0, M...

Embodiment 3

[0042] Inoculate Streptomyces hygroscopicus, CGMCC No.5145, on an oat-potato slant at 28°C for 15 days, and black spots appear on the surface of the slant lawn. At this time, transfer it to a seed bottle; take a ring from the slant with an inoculation shovel Inoculate into an Erlenmeyer flask containing a seed medium, and cultivate at 28° C. and 200 rpm for 60 hr. Wherein the seed medium composition (g / L) comprises soluble starch 10.0, peptone 6.0, yeast extract 6.0, acid hydrolyzed casein 1.5; The above cultured bacterial strain is transferred in the fermentation tank of 100L according to the inoculation amount of 2% (built-in 60L through sterilized fermentation medium), the fermentation temperature was adjusted to 28°C, the ventilation rate was 0.5v / min, the stirring speed was 250rmp, and the fermentation was carried out for 5 days. The fermentation medium composition (g / L) includes: glucose 40, glycerin 10, soybean cake 10, yeast extract 6.0, peptone, L-Lysine 0.15, K 2 HP...

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Abstract

The invention provides a process for preparing rapamycin by using a microbial fermentation method. The method comprises the following steps of: culturing streptomyces hygroscopicus of which the collection number is CGMCC No.5145 in a slant culture medium, and inoculating into a fresh seed culture medium for culturing; and inoculating 2 percent of strain cultured by using the seed culture medium into a fresh fermentation culture medium for fermenting and culturing to obtain a fermentation liquor containing rapamycin, and separating and purifying the fermentation liquor containing the rapamycin according to the solubility difference of the rapamycin in water and an organic solvent. In a method provided by the invention, a production technology for producing high-purity rapamycin at low cost is provided. By reducing the cost of a raw material, an immunosuppressive medicament which is economically acceptable, safe and effective can be provided for an organ transplantation patient. The product purity of the rapamycin prepared with the process is over 95 percent, and the recrystallization yield is over 84 percent.

Description

technical field [0001] The invention relates to the technical field of microbial fermentation, in particular to the technical field of preparing rapamycin by microbial fermentation. Background technique [0002] Rapamycin (RAPA), also known as Sirolimus, is a new macrolide immunosuppressant. The molecular formula of RAPA is C 51 h 79 NO 13 , molecular weight 991KD, white solid crystal, melting point 183-185°C, lipophilic, soluble in methanol, ethanol, acetone, chloroform and other organic solvents, very slightly soluble in water, almost insoluble in ether. RAPA blocks signal transduction through different cytokine receptors, and blocks the progress of T lymphocytes and other cells from G1 phase to S phase, thereby exerting an immunosuppressive effect. [0003] Rapamycin was originally a metabolite produced by Streptomyces AYB-944 isolated from soil samples of Ester Island (Rapa Nui) in the mid-1970s by Canada's Wyeth-Ayerst Research Laboratory. The hyphae of the fungus ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P17/18C07D498/18C12R1/55
Inventor 王福清徐莉张桐
Owner 中科医药行业生产力促进中心有限公司
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