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WO2019233367A1 - Souche de streptomyces et son application dans la production de staurosporine - Google Patents

Souche de streptomyces et son application dans la production de staurosporine Download PDF

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WO2019233367A1
WO2019233367A1 PCT/CN2019/089782 CN2019089782W WO2019233367A1 WO 2019233367 A1 WO2019233367 A1 WO 2019233367A1 CN 2019089782 W CN2019089782 W CN 2019089782W WO 2019233367 A1 WO2019233367 A1 WO 2019233367A1
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streptomyces
powder
culture
seed
medium
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WO2019233367A8 (fr
Inventor
方佳双
张辉
项仁鑫
邓爱文
王微芬
应灵萍
姜南
王继栋
滕云
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Zhejiang Hisun Pharmaceutical Co Ltd
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C12N1/205—Bacterial isolates
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
    • C12R2001/46—Streptococcus ; Enterococcus; Lactococcus
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
    • C12P17/18—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms containing at least two hetero rings condensed among themselves or condensed with a common carbocyclic ring system, e.g. rifamycin
    • C12P17/188—Heterocyclic compound containing in the condensed system at least one hetero ring having nitrogen atoms and oxygen atoms as the only ring heteroatoms

Definitions

  • the invention belongs to the technical field of microbial engineering, and particularly relates to a strain of Streptomyces and its application in preparing staurosporine.
  • Staurosporine (the structural formula is shown in Formula 1), originally found in the fermentation product of Streptomyces staurosporeus (AM-2282) strain, has antibacterial, antifungal, blood pressure lowering, platelet aggregation, antitumor and neuroprotection, etc.
  • AML acute myeloid leukemia
  • Rydapt was approved for use with the companion diagnostic reagent LeukoStrat CDx FLT3 Mutation Assay to detect FLT3 mutations in patients with AML.
  • Rydapt is also approved for the treatment of adult patients with certain types of rare hematological disorders, including aggressive systemic mastocytosis (ASM), systemic mast cell proliferation with hematological tumors Disease (SM-AHN) and mast cell leukemia (MCL).
  • ASM systemic mastocytosis
  • SM-AHN systemic mast cell proliferation with hematological tumors Disease
  • MCL mast cell leukemia
  • One of the objectives of the present invention is to provide a new Streptomyces HS-HY-153, which has the ability to produce staurosporin, and its deposit number is CGMCC No. 14806.
  • the microbial strain of the streptomyces HS-HY-153 of the present invention was deposited on October 13, 2017 at the Common Microbial Center (CGMCC) of the China Microbial Strain Collection Management Committee (Address: No. 3, No. 1, Beichen West Road, Chaoyang District, Beijing, China) ), Its deposit number is CGMCC No. 14806, it is classified as Streptomyces sp., And it is registered to prove its survival.
  • CGMCC Common Microbial Center
  • the object of the present invention is also to provide the application of a new streptomyces HS-HY-153 (CGMCC No. 14806) in preparing staurosporine or a pharmaceutical composition containing staurosporine.
  • the invention also provides a method for preparing staurosporin.
  • the method includes the process of aerobic fermentation by using Streptomyces HS-HY-153 (CGMCC No. 14806) in a nutrient medium containing an assimitable carbon source and / or nitrogen source.
  • the assimitable carbon source is selected from the group consisting of corn starch, corn soluble starch, maltodextrin, sucrose, glucose, sorbitol, mannitol, glycerol, maltose, lactose, galactose, xylan, One of industrial molasses, soybean oil, methyl oleate, or any combination thereof.
  • the assimilated nitrogen source is selected from beef extract powder, beef extract, yeast extract, yeast extract, malt extract powder, yeast powder, peptone, gluten powder, wheat bran , Wheat germ flakes, cottonseed meal powder, cottonseed meal powder, peanut meal powder, soybean meal, wheat germ meal, soybean meal, fish meal, dried corn slurry meal, corn meal, urea, ammonium salt, nitrate, or one of the above substances random combination.
  • the nutrient medium further includes an inorganic salt selected from the group consisting of zinc sulfate, magnesium sulfate, ferrous sulfate, copper sulfate, manganese sulfate, diammonium hydrogen phosphate, potassium chloride, and chloride.
  • an inorganic salt selected from the group consisting of zinc sulfate, magnesium sulfate, ferrous sulfate, copper sulfate, manganese sulfate, diammonium hydrogen phosphate, potassium chloride, and chloride.
  • the nutrient medium contains glucose 0-30.0g / L, glycerol 5.0-90.0g / L, peanut cake powder 5.0-60.0g / L, yeast powder 3.0-20.0g / L, and soybean cake Flour 0-40.0g / L, wheat germ flakes 0-30.0g / L, soybean flour 0-40.0g / L, magnesium sulfate 0-5.0g / L, calcium carbonate 1.0-5.0g / L, ferrous sulfate 0- 5.0 ⁇ 10 -2 g / L, zinc sulfate 0-2.0 ⁇ 10 -3 g / L, manganese sulfate 0-2.0 ⁇ 10 -4 g / L, cobalt chloride 0-5.0 ⁇ 10 -5 g / L, molybdenum Sodium 0-2.0 ⁇ 10 -4 g / L; preferably, the nutrient medium contains glucose 0-10.0g / L, glycerol 70.0.0-9
  • the temperature of the aerobic fermentation is 20 ° C-30 ° C, preferably 24 ° C-28 ° C;
  • the pH of the culture medium is 6.0-8.0, preferably 7.0-7.8;
  • the culture time is 24-300 hours, preferably 120-168 hours.
  • the streptomyces HS-HY-153 is inoculated into the nutrient medium through a seed liquid to perform the fermentation culture;
  • the seed solution is obtained by seed culture of Streptomyces HS-HY-153 in a seed medium; the conditions for the seed culture are: the temperature of the seed culture is 20 ° C-30 ° C, preferably 24 ° C-28 ° C
  • the pH of the medium is 6.0-8.0, preferably 6.5-7.8; the culture time is 17-80 hours, preferably 19-72 hours.
  • the seed medium contains corn starch 2.0-40.0g / L, glucose 0-20.0g / L, maltodextrin 0-10.0g / L, glycerin 5.0-50.0g / L, peanut Cake powder 5.0-40.0g / L, yeast powder 5.0-20.0g / L, soybean cake powder 0-10.0g / L, calcium carbonate 1.0-4.0g / L.
  • the staurosporin of the present invention can be detected by HPLC under the following conditions:
  • Chromatographic column C18 column (4.6mm ⁇ 250mm, 5 ⁇ m);
  • Injection volume 20 ⁇ L
  • the main biological characteristics of the Streptomyces HS-HY-153 (CGMCC No. 14806) of the present invention are: round colony shape, convex surface, colony diameter of about 5-7mm, smooth colony surface, grooves, and a slight depression in the middle
  • the matrix mycelium is well developed, tightly combined with the culture medium, not easy to provoke, the color is colorless, beige or light yellow, the aerial mycelium is white, the spores are rich, the cyan is in the early stage, and the teal is turned into dark cyan, cyan grey, and agate gray in the later stage No soluble pigments.
  • the strain HS-HY-153 of the present invention has high production capacity; and its ability to produce staurosporin has been greatly improved compared with other strains in the prior art, which is beneficial to the realization of industrial production.
  • Fig. 1 is a micrograph (400 ⁇ ) of strain HS-HY-153 (CGMCC No. 14806) on ISP2 medium.
  • Figure 2 is a colony characteristic diagram of strain HS-HY-153 (CGMCC No. 14806) on ISP2 medium.
  • Figure 3 is a colony characteristic diagram of strain HS-HY-153 (CGMCC No. 14806) on ISP1, ISP3, ISP4, ISP5, calcium malate, Gao No. 1, nutrient agar medium.
  • the actinomycete DNA extraction kit used was a product of Beijing Sanbo Yuanzhi Biotechnology Co., Ltd.
  • the SanPrep column PCR product purification kit used for the purification and recovery of PCR products is a product of Biotech Bioengineering (Shanghai) Co., Ltd.
  • Yeast extraction powder (YEAST EXTRACT) is a product of OXOID Company, article number LP0021.
  • Malt Extract is a product of BD Co., Ltd., product number 218630.
  • Corn starch is a product of Weifang Shengtai Pharmaceutical Co., Ltd.
  • Peanut cake powder is a product of Jinan Huilong Biotechnology Co., Ltd.
  • Wheat germ flakes are products of Haining Dawei Food Co., Ltd.
  • the staurosporine standard was made by Zhejiang Hisun Pharmaceutical Co., Ltd., and its chromatographic purity was 99%.
  • HPLC detection conditions are as follows:
  • Chromatographic column C18 column (4.6mm ⁇ 250mm, 5 ⁇ m);
  • Injection volume 20 ⁇ L
  • the Streptomyces HS-HY-153 (CGMCC No. 14806) of the present invention is obtained by isolating the original strain from a soil sample in Tiantai Mountain, Zhejiang province, and then breeding it through mutagenesis.
  • Solid medium formula ISP2. After sterilizing at 121 ° C for 20 minutes, cool to 50-60 ° C and pour the plate. The mutagenized bacteria suspension was appropriately diluted, and 0.2 mL of the bacterial solution was pipetted onto the ISP2 plate. The non-mutagenic bacteria suspension was also appropriately diluted and coated on the ISP2 plate as a control. Box culture, colony culture for 7 days, mycelium is mature.
  • Seed medium formula yeast powder 7.5g / L, glucose 8.0g / L, corn starch 10.0g / L, glycerin 20.0g / L, peanut cake powder 20.0g / L, calcium carbonate 2.0g / L, the balance is tap water , Adjust the pH to 7.2-7.4 with 6mol / L NaOH solution; the volume of the triangle shake bottle is 25mL / 250mL, and sterilize at 121 °C for 20min. Each seed shake flask was inoculated with about 0.5 colonies, and cultured at 28 ⁇ 1 ° C and 250 rpm for 24 hours to obtain seed liquid.
  • Formulation of fermentation medium glycerin 50.0g / L, glucose 5.0g / L, peanut cake powder 20.0g / L, yeast powder 5.0g / L, calcium carbonate 3.0g / L, the balance is tap water, using 6mol / L NaOH solution Adjust the pH to 7.4-7.6; the volume of the triangle shake bottle is 20mL / 250mL, and sterilize at 121 ° C for 20min.
  • the seed liquid is inserted into the fermentation medium with an inoculation amount of 5-10% (volume percentage), the cultivation temperature is 28 ⁇ 1 ° C., the rotation speed is 250 rpm, and the cultivation is performed for 120 hours.
  • Example 2 The morphology, culture characteristics and physiological and biochemical characteristics of Streptomyces HS-HY-153 (CGMCC No. 14806).
  • strain HS-HY-153 (CGMCC No. 14806) was cultured on ISP2 medium at 28 ° C for 7-10 days, the colonies were round in shape and the surface was convex, and the diameter of the colonies was about 5-7mm. The surface of the colony is smooth, with grooves, with a slight depression in the middle, and the matrix hyphae is developed. It is tightly combined with the culture medium and difficult to provoke. The color is pale yellow, the aerial hyphae is white, the spore production is rich, the early stage is blue, and the late stage is dark Cyan, cyan gray, no soluble pigment, the results are shown in Figure 2.
  • ISP1, ISP3, ISP4, ISP5, calcium malate, Gao No.1, and nutrient agar were used in 7 kinds of medium. After culturing at 28 ° C for 7-10 days, observe the colonies, mycelia, spores, and pigments. The situation occurs, and the results are shown in Table 1 and Figure 3.
  • ISP9 inorganic nitrogen source
  • Catalase test, pH test and temperature test all use ISP2 medium.
  • the catalase test results are shown in Table 4, the pH test results are shown in Table 5, and the temperature test results are shown in Table 6.
  • Carbon source growing situation Carbon source growing situation
  • Inorganic nitrogen source growing situation D-glucose 4 Salicin 2 Ammonium sulfate + D-raffinose 2 D-lactose 2 Potassium nitrate - D-xylose 2
  • Galactose 3 D-sorbitol 2 Inositol 2 L-arabinose 2 Mannitol 3
  • Degradation Degradation concentration result Degradation Degradation concentration result Adenine 0.5% 2,- Casein 1.0% 4,- Guanine 0.5% 4,- Tyrosine 1.0% 4, + Xanthine 0.4% 4,- Tween-40 1.0% 2,- Xylan 0.4% 4,- Tween-60 1.0% 2,- Hypoxanthine 0.4% 4, + Tween-80 1.0% 2,-
  • the 16S rDNA sequence (SEQ ID NO: 3) measured by strain HS-HY-153 (CGMCC No. 14806) was submitted to NCBI and related sequences in GenBank were compared for BLAST. Highly sensitive model strains).
  • Streptomyces cirratus NRRL B-3250 (T) AY999794 99.45 Streptomyces spororaveus LMG 20313 (T) AJ781370 99.45 Streptomyces nojiriensis LMG 20094 (T) AJ781355 99.45 Streptomyces vinaceus NBRC 13425 (T) AB184394 99.44 Streptomyces sporoverrucosus NBRC 15458 (T) AB184684 99.44 Streptomyces goshikiensis NBRC 12868 (T) AB184204 99.44 Streptomyces colombiensis NRRL B-1990 (T) DQ026646 99.38 Streptomyces lavendulae subsp.lavendulae NRRL B-2774 (T) JOEW01000098 99.38 Streptomyces avidinii NBRC 13429 (T) AB184395 99.17
  • strain HS-HY-153 (CGMCC No. 14806) was sequenced and compared with the sequences of related species and genus in the GenBank database by homologous sequence BLAST. It was found to be related to Streptomyces xanthophaeus and tendril Streptomyces cirratus, Streptomyces spororaveus, Streptomyces nojiriensis all have a homology of 99.45%. At the same time, the strain HS-HY-153 (CGMCC No. 14806) has apparent characteristics. It was found that the strain and Streptomyces sp. Classification-related parameters were very close. Therefore, the strain HS-HY-153 (CGMCC No. 14806) was identified as a Streptomyces sp. Strain.
  • US7608420B and EP0444503A disclose the fermentation units of Streptomyces hygroscopicus C39280-450-9 (ATCC 53730) for producing staurosporine are 392 mg / L and 130 mg / L, respectively, and the fermentation of HS-HY-153 (CGMCC No. 14806) of the present invention The unit is as high as 1540mg / L.
  • US4107297 discloses the morphological, cultural, and physiological and biochemical characteristics of Streptomyces sp. AM-2282 (later named Streptomyces staurosporeus nov.sp. NRRL11184).
  • the bacteria HS-HY-153 (CGMCC No. 14806) and Streptomyces of the present invention The cultural characteristics of sp.AM-2282 are shown in Table 10, and the physiological and biochemical characteristics of Streptomyces sp.AM-2282 are shown in Table 11. It can be seen from Tables 10 and 11 that the bacteria HS-HY-153 (CGMCC No. 14806) and Streptomyces sp. AM-2282 of the present invention have physiological and biochemical characteristics such as culture characteristics, milk coagulation, milk mash, cellulose utilization All are different.
  • strain H4138 for the cultural characteristics of strain H4138 is shown in Table 13, and the physiological and biochemical characteristics of strain H4138 are shown in Table 14; as can be seen from Tables 13 and 14, HS-HY of the present invention -153 (CGMCC No. 14806) and strain H4138 are different in physiological and biochemical characteristics such as culture characteristics, hydrogen sulfide production, milk coagulation, milk mash, and cellulose utilization.
  • the strain HS-HY-153 (CGMCC No. 14806) of the present invention Belongs to Streptomyces sp. Strain, and is different from other known staurosporin-producing bacteria, so the streptomyces HS-HY-153 (CGMCC No. 14806) of the present invention is a brand new astrospore Streptozotocin-producing bacteria.
  • Slant medium formula yeast extraction powder 4.0g / L, malt extract 10.0g / L, glucose 4.0g / L, agar 20.0g / L, the balance is purified water, and the pH is adjusted with 6mol / L NaOH solution 7.2-7.4; 250mL eggplant-shaped bottle, filled with 60mL, sterilized at 121 ° C for 20min, cooled to about 55 ° C, and placed in a 37 ° C incubator for 1-2 days after cooling and solidification, inoculated with Streptomyces HS -HY-153 (CGMCC No. 14806) spores or mycelia to oblique plane, after culturing at 28 ⁇ 1 °C for 7-10 days, the spores mature.
  • CGMCC No. 14806 Streptomyces HS -HY-153
  • Seed medium formula yeast powder 7.5g / L, glucose 8.0g / L, corn starch 10.0g / L, glycerin 20.0g / L, peanut cake powder 20.0g / L, calcium carbonate 2.0g / L, the balance is tap water Use a 6mol / L NaOH solution to adjust the pH to 7.2-7.4; 250mL triangular shake flask with a capacity of 25mL and sterilize at 121 ° C for 20min.
  • the spores obtained in step (1) were inoculated into the seed medium at 10 7 -10 8 cfu / mL, and cultured at 28 ⁇ 1 ° C and shaking at 250 rpm for 22-24 hours. At this time, the pH of the culture solution was 6.8-7.2, and the concentration of mycelium 8-10% (volume percentage) to obtain a seed liquid.
  • Formulation of fermentation medium glycerin 50.0g / L, glucose 5.0g / L, peanut cake powder 20.0g / L, yeast powder 5.0g / L, calcium carbonate 3.0g / L, the balance is tap water, using 6mol / L NaOH solution Adjust the pH to 7.4-7.6; 250mL triangular shake flask with a volume of 20mL and sterilize at 121 °C for 20min.
  • the seed liquid obtained in step (2) is inserted with an inoculation amount of 5-10% (volume percentage).
  • the culture was shaken at 28 ⁇ 1 ° C and 250 rpm for 168 hours to obtain a fermentation broth.
  • the astrosporin content in the fermentation broth was measured by HPLC method, and it was found to be 1310 ⁇ g / mL.
  • Fermentation medium formula: glycerol 70.0g / L, glucose 5.0g / L, peanut cake powder 40.0g / L, yeast powder 5.0g / L, magnesium sulfate 1.0g / L, calcium carbonate 3.0g / L, the balance is tap water Use a 6mol / L NaOH solution to adjust the pH to 7.4-7.6; a 250mL triangular shake flask with a volume of 20mL and sterilize at 121 ° C for 20min.
  • the seed liquid obtained in step (2) is inserted with an inoculation amount of 5-10% (volume percentage).
  • the culture was shaken at 28 ⁇ 1 ° C and 250 rpm for 168 hours to obtain a fermentation broth.
  • the astrosporin content in the fermentation broth was measured by HPLC method, and it was found to be 1380 ⁇ g / mL.
  • Fermentation medium formula: glycerol 80.0g / L, peanut cake powder 40.0g / L, yeast powder 5.0g / L, wheat germ flakes 20.0g / L, magnesium sulfate 1.0g / L, calcium carbonate 3.0g / L, subsulfate Iron 5.0 ⁇ 10 -2 g / L, zinc sulfate 2.0 ⁇ 10 -3 g / L, the balance is tap water, adjust the pH to 7.4-7.6 with 6mol / L NaOH solution; 250mL triangle shake flask, 20mL, Sterilize at 121 ° C for 20min.
  • the seed liquid obtained in step (2) is inserted with an inoculation amount of 5-10% (volume percentage).
  • the culture was shaken at 28 ⁇ 1 ° C and 250 rpm for 168 hours to obtain a fermentation broth.
  • the astrosporin content in the fermentation broth was measured by HPLC method, and it was found to be 1480 ⁇ g / mL.
  • Formulation of fermentation medium glycerin 90.0g / L, glucose 10.0g / L, peanut cake powder 50.0g / L, yeast powder 10.0g / L, wheat germ flakes 10.0g / L, magnesium sulfate 1.0g / L, calcium carbonate 4.0 g / L, ferrous sulfate 5.0 ⁇ 10 -2 g / L, zinc sulfate 5.0 ⁇ 10 -5 g / L, the balance is tap water, adjust the pH to 7.4-7.6 with 6mol / L NaOH solution; 250mL triangle shake 20ml bottle, sterilize at 121 °C for 20min.
  • the seed liquid obtained in step (2) is inserted with an inoculation amount of 5-10% (volume percentage).
  • the culture was shaken at 28 ⁇ 1 ° C and 250 rpm for 168 hours to obtain a fermentation broth.
  • the astrosporin content in the fermentation broth was measured by HPLC method, and it was found to be 1505 ⁇ g / mL.
  • the formula of the seed liquid medium in the seed tank is the same as the seed medium in step (2) in Example 4;
  • step (2) Put 10L of seed medium into a 15L seed tank, sterilize it with steam, and sterilize it at 121 ° C for 20min. After cooling to 28 ° C, insert 200mL of the first-stage seed solution prepared in step (2). The stirring speed was 200 rpm, the aeration was 1.0 vvm, and the culture was performed at 28 ⁇ 1 ° C. for 24 hours. At this time, the seed solution had a pH of 7.0-7.4 and a hyphae concentration of 12-15% (volume percentage) to obtain a seed tank seed solution.
  • the formulation of the fermentation medium is the same as the fermentation medium in step (3) in Example 6;
  • the fermentation tank has a volume of 50L and a feed volume of 30L. It is sterilized with steam at 121 ° C for 20min. After cooling to 28 ° C, it is connected to 3L of the seed tank seed solution prepared in step (3).
  • the stirring rotation speed is 200-400 rpm (the rotation speed is gradually increased from 200 rpm to 320 rpm in the first 3 days), the aeration volume is 0.8-1.0 vvm, and the culture is performed at 28 ° C. for 168 hours to obtain a fermentation tank fermentation broth.
  • the astrosporin content in the fermentation broth was measured by HPLC method to be 1540 ⁇ g / mL.
  • Example 8 30 L of the fermentation broth obtained in Example 8 was centrifuged (4000 rpm, 15 min) to obtain mycelia, and the mycelia was extracted twice with acetone to obtain an acetone extract. The acetone extract was concentrated to dryness in vacuo and extracted with ethyl acetate to obtain an extract containing staurosporine. After the sample was extracted with silica gel, the sample was loaded on a silica gel column and eluted with a chloroform / methanol gradient of 99: 1, 98: 2, 97: 3, 96: 4 (V: V). The eluted fractions were collected in stages and detected by TLC.
  • Example 4-7 The fermentation broth obtained in Example 4-7 was processed according to the same isolation, purification, and structure identification steps as described above, and structural data consistent with the above were obtained, confirming that the target component in the fermentation broth was staurosporine.
  • the target peak collected by HPLC in Examples 4-8 was identified by the above structure, and it was also confirmed that the target was staurosporin.

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Abstract

L'invention concerne une souche de streptomyces et son application, et un procédé de préparation de staurosporine par culture de la souche. La souche est nommée streptomyces sp. HS-HY-153, et son numéro de conservation est CGMCC NO. 14806.
PCT/CN2019/089782 2018-06-04 2019-06-03 Souche de streptomyces et son application dans la production de staurosporine Ceased WO2019233367A1 (fr)

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CN108676757B (zh) * 2018-06-04 2020-08-11 浙江海正药业股份有限公司 一株链霉菌属菌株及其产星孢菌素的应用
KR102235779B1 (ko) * 2019-06-19 2021-04-01 국립해양생물자원관 해수로부터 분리된 스트렙토마이세스 속의 snc087 균주, 동 균주를 이용한 스타우로스포린의 생산방법, 동 균주의 배양방법 및 동 균주의 순수배양액
CN110642872B (zh) * 2019-11-18 2020-12-22 湖北宏中药业股份有限公司 一种星孢菌素提取方法
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CN111296417B (zh) * 2020-04-01 2021-04-20 广东省农业科学院植物保护研究所 一种星·甲维悬浮剂及其制备方法与应用
CN111793618A (zh) * 2020-08-05 2020-10-20 广东新达瑞生物科技有限公司 一种星孢菌素产生菌的选育及制备方法
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