CN103695511A - Method for producing daptomycin by fermentation - Google Patents
Method for producing daptomycin by fermentation Download PDFInfo
- Publication number
- CN103695511A CN103695511A CN201310644852.XA CN201310644852A CN103695511A CN 103695511 A CN103695511 A CN 103695511A CN 201310644852 A CN201310644852 A CN 201310644852A CN 103695511 A CN103695511 A CN 103695511A
- Authority
- CN
- China
- Prior art keywords
- fermentation
- daptomycin
- seed liquor
- streptomyces roseosporus
- capraldehyde
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention provides a method for producing daptomycin by fermentation, and belongs to the field of production of daptomycin. In the method for synthesis of daptomycin by fermentation, the parameters such as the concentration and the inoculum size of a culture seed solution and pH of fermentation medium are optimized, the growth activity and the fermentation activity of streptomyces roseosporus are maintained, the production of the daptomycin is improved; in the fermentation process, a decanal solution is fed to the fermentation medium, and the adding time and the adding amount are optimized, so that the inhibition concentration of the daptomycin can be improved, and the synthetic production of the daptomycin is further improved.
Description
Technical field
The invention belongs to the production field of daptomycin, relate in particular to a kind of method of producing daptomycin by fermentation.
Background technology
Over nearly 10 years, the trend of Gram-positive drug-fast bacteria infection is in continuous rising.The difficult point for the treatment of clinically bacterium infection at present mainly concentrates on methicillin-resistant staphylococcus aureus (MRSA), the faecalis of vancomycin resistance (VRE), the golden Portugal bacterium (GISA) of glycopeptide class sensitivity etc., they are also the very high pathogenic agent of spreading rate profit lethality rate in Hospitals at Present, although the glycopeptide antibiotics such as vancomycin are treated this class infection and have been obtained certain curative effect, but in recent years, found clinically the resistant organism of vancomycin, and this trend is increasing gradually.Therefore the research of antimicrobial agent new antibiotic becomes again heat subject.Daptomycin all has good sterilization effect to above resistant organism, and preparation medication is convenient, and toxic side effect is little, becomes the best substitute of " pathogenic bacteria last line of defense---vancomycin "; What have more meaning is, daptomycin is as first product of cyclic lipopeptide microbiotic family, its chemical structure and mechanism of action are different from the microbiotic of existing all categories, be over nearly 40 years after mouthful oxazolidinones microbiotic, be applied to clinical unique new texture classification microbiotic, and daptomycin seldom causes generation resistance strain isolated in clinical trial.
Daptomycin (Daptomycin) is a kind of cyclic lipopeptide microbiotic obtaining from Streptomyces roseosporus (Streptomyces roseosporus) separation of fermentative broth, and its chemical structure is suc as formula shown in (I).From formula (I), can find out, daptomycin is comprised of 13 amino acid and 1 longer chain fatty acid, and wherein 10 amino acid form an amino acid ring, separately have 3 amino acid to be arranged in wire, and L-tryptophane (L-Trp) is endways upper, connects 1 n-capric acid.
Daptomycin not only has novel chemical structure, its binding mode is also different from the microbiotic of any existing listing: daptomycin can be upset cytolemma to amino acid whose transhipment, thereby hinder the biosynthesizing of bacteria cell wall peptidoglycan, change the character of cytoplasmic membrane; In addition, it can also be by destroying the cytolemma of bacterium, its content is leaked and reaches the object of sterilization.Daptomycin, by destroy bacterial cell membrane function from many aspects, can kill rapidly gram-positive microorganism, and make bacterium be difficult to its produce resistance (A.Raja et al., Nature Rev.Drug Discov., 2003,2,943-944).Daptomycin, except acting on most of clinical relevant gram-positive microorganisms, the more important thing is, it also presents powerful activity to the resistance isolated strains of X-1497, vancomycin and linwzolid etc. in testing in vitro.
The synthetic method of daptomycin has complete synthesis, the molecular design of chemistry and three kinds of techniques of biosynthesizing, and wherein the first two is planted due to complex technical process and the low commercial value that there is no of productive rate.At present, in industry, the ordinary method of production daptomycin is mainly to make by Streptomyces roseosporus strain fermentation.
During the synthetic daptomycin of fermentation method, due to the feedback inhibition in microbial metabolism, when the daptomycin producing when fermentation reaches finite concentration, will suppress the further synthetic of daptomycin, make the concentration of daptomycin of fermentative Production not high.
Summary of the invention
For addressing the above problem, the invention provides a kind of method of producing daptomycin by fermentation, adopt following technical scheme:
A method for producing daptomycin by fermentation, is characterized in that comprising following content:
A. prepare Streptomyces roseosporus seed liquor;
B. inoculation: the seed liquor obtaining in step a is inoculated in fermention medium, and inoculum size is 10%;
C. fermentation culture: the postvaccinal substratum obtaining in step b is cultivated and fermented, and fermentation culture temperature is 30～32 ℃;
D. add capraldehyde solution: after fermentation culture 18h, to stream in fermented liquid, add capraldehyde solution, keeping the concentration of capraldehyde in fermented liquid is 0.1～2mmol/L, until fermentation ends;
E. fermentation ends separation: after fermentation culture 48～168h, finish fermentation, the fermented liquid obtaining is extracted to separation, obtain described daptomycin.
Preferably, the Streptomyces roseosporus seed liquor of preparing described in step a comprises following content: the Streptomyces roseosporus bacterial strain of preservation is transferred on slant medium, cultivate 120h for 30 ℃, then be inoculated in liquid nutrient medium, in the shaking table of 30 ℃, cultivate 36h, obtain described Streptomyces roseosporus seed liquor.
Preferably, described slant medium comprises the component of following weight part: 20 parts of Zulkovsky starches, 0.5 part of dipotassium hydrogen phosphate, 0.5 part of magnesium sulfate heptahydrate, 0.01 part of green vitriol, 1 part, saltpetre, 20 parts, agar, 1000 parts of deionized waters.
Preferably, pH=6.5 ~ 7.0 of the fermention medium described in step b.
Beneficial effect of the present invention is as follows:
In the method for the synthetic daptomycin of fermentation provided by the invention, the parameters such as bacterial classification seed liquor concentration, inoculum size, fermention medium pH are optimized, maintain growth activity and the fermentation activity of Streptomyces roseosporus, improve the turnout of daptomycin; During the fermentation, to stream in fermention medium, add capraldehyde solution, and its interpolation time and addition are optimized, can improve the inhibition concentration of daptomycin, further improve the synthetic output of daptomycin.
Embodiment
Below in conjunction with specific embodiment, the present invention is described in detail.Following examples will contribute to those skilled in the art further to understand the present invention, but not limit in any form the present invention.It should be pointed out that to those skilled in the art, without departing from the inventive concept of the premise, can also make some distortion and improvement.These all belong to protection scope of the present invention.
embodiment 1
Stream adds a method for capraldehyde producing daptomycin by fermentation, comprises following content:
A. cultivate Streptomyces roseosporus seed liquor;
B. inoculation: the seed liquor obtaining in step a is inoculated in fermention medium, and inoculum size is 10%;
C. fermentation culture: the postvaccinal substratum obtaining in step b is cultivated and fermented, and fermentation culture temperature is 30～32 ℃;
D. add capraldehyde solution: after fermentation culture 18h, to stream in fermented liquid, adding mass concentration is 6.0% capraldehyde solution (solvent is Witconol 2301), the speed that stream adds is: in every liter of fermented liquid with the flow velocity of 1.0～2.0ml/h toward feed supplement in fermentor tank, until fermentation ends;
E. fermentation ends separation: after fermentation culture 48～168h, finish fermentation, the fermented liquid obtaining is extracted to separation, obtain described daptomycin.
embodiment 2
Stream adds a method for capraldehyde producing daptomycin by fermentation, comprises following content:
A. cultivate Streptomyces roseosporus seed liquor: the Streptomyces roseosporus bacterial strain of preservation is transferred on slant medium, cultivate 120h for 30 ℃, be then inoculated in liquid nutrient medium, in the shaking table of 30 ℃, cultivate 36h, obtain described Streptomyces roseosporus seed liquor;
Wherein, the compound method of described slant medium is as follows: Zulkovsky starch 20g, dipotassium hydrogen phosphate 0.5g, magnesium sulfate heptahydrate 0.5g, green vitriol 0.01g, saltpetre 1g are added in 1L deionized water, regulate pH=7.2, and then add agar 20g, through steam sterilizing, then contra bevel is dull and stereotyped, obtains slant medium;
B. inoculation: the seed liquor obtaining in step a is inoculated in fermention medium (pH=6.5 ~ 7.0 of fermention medium), and inoculum size is 10%;
C. fermentation culture: the postvaccinal substratum obtaining in step b is cultivated and fermented, and fermentation culture temperature is 30～32 ℃;
D. add halfcystine: after fermentation culture 18h, to stream in fermented liquid, adding mass concentration is 6.0% capraldehyde solution (solvent is ethanol), the speed that stream adds is: in every liter of fermented liquid with the flow velocity of 1.0～2.0ml/h toward feed supplement in fermentor tank, until fermentation ends;
E. fermentation ends separation: after fermentation culture 48～120h, finish fermentation, the fermented liquid obtaining is extracted to separation, obtain described daptomycin.
Claims (4)
1. a method for producing daptomycin by fermentation, is characterized in that comprising following content:
Prepare Streptomyces roseosporus seed liquor;
Inoculation: the seed liquor obtaining in step a is inoculated in fermention medium, and inoculum size is 10%;
Fermentation culture: the postvaccinal substratum obtaining in step b is cultivated and fermented, and fermentation culture temperature is 30～32 ℃;
D. add capraldehyde solution: after fermentation culture 18h, to stream in fermented liquid, add capraldehyde solution, keeping the concentration of capraldehyde in fermented liquid is 0.1～2mmol/L, until fermentation ends;
E. fermentation ends separation: after fermentation culture 48～168h, finish fermentation, the fermented liquid obtaining is extracted to separation, obtain described daptomycin.
2. the method for producing daptomycin by fermentation according to claim 1, it is characterized in that, the Streptomyces roseosporus seed liquor of preparing described in step a comprises following content: the Streptomyces roseosporus bacterial strain of preservation is transferred on slant medium, cultivate 120h for 30 ℃, then be inoculated in liquid nutrient medium, in the shaking table of 30 ℃, cultivate 36h, obtain described Streptomyces roseosporus seed liquor.
3. the method for producing daptomycin by fermentation according to claim 2, it is characterized in that, described slant medium comprises the component of following weight part: 20 parts of Zulkovsky starches, 0.5 part of dipotassium hydrogen phosphate, 0.5 part of magnesium sulfate heptahydrate, 0.01 part of green vitriol, 1 part, saltpetre, 20 parts, agar, 1000 parts of deionized waters.
4. the method for producing daptomycin by fermentation according to claim 1, is characterized in that, pH=6.5 ~ 7.0 of the fermention medium described in step b.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310644852.XA CN103695511A (en) | 2013-12-05 | 2013-12-05 | Method for producing daptomycin by fermentation |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310644852.XA CN103695511A (en) | 2013-12-05 | 2013-12-05 | Method for producing daptomycin by fermentation |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN103695511A true CN103695511A (en) | 2014-04-02 |
Family
ID=50357174
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310644852.XA Pending CN103695511A (en) | 2013-12-05 | 2013-12-05 | Method for producing daptomycin by fermentation |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103695511A (en) |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20100047873A1 (en) * | 2008-08-01 | 2010-02-25 | Antibioticos S.P.A | Process for the production of daptomycin |
| CN101928677A (en) * | 2009-06-26 | 2010-12-29 | 上海来益生物药物研究开发中心有限责任公司 | Producing strain of lipopeptide compound A21978C and application thereof |
| CN102796680A (en) * | 2012-07-04 | 2012-11-28 | 鲁南新时代生物技术有限公司 | Streptomyces roseosporus and method for producing daptomycin by utilizing combined precursor |
-
2013
- 2013-12-05 CN CN201310644852.XA patent/CN103695511A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20100047873A1 (en) * | 2008-08-01 | 2010-02-25 | Antibioticos S.P.A | Process for the production of daptomycin |
| CN101928677A (en) * | 2009-06-26 | 2010-12-29 | 上海来益生物药物研究开发中心有限责任公司 | Producing strain of lipopeptide compound A21978C and application thereof |
| CN102796680A (en) * | 2012-07-04 | 2012-11-28 | 鲁南新时代生物技术有限公司 | Streptomyces roseosporus and method for producing daptomycin by utilizing combined precursor |
Non-Patent Citations (1)
| Title |
|---|
| 韩培等: "达托霉素生物合成研究进展及临床研究", 《中国医药工业杂志》 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103329907B (en) | A kind of preparation method of fen second mycin biological bactericide | |
| CN102363754A (en) | Clear liquid fermentation medium for clostridium butyricum and fermentation culture method thereof | |
| CN103343150B (en) | A kind of preparation method of azophenlyene-1-formyl hydrazine biological bactericide | |
| CN101503708B (en) | Cultivation fermentation method of Bacillus coagulans antimycotics active substance | |
| CN104593303B (en) | A kind of liquid complex micro organism fungicide and its production method | |
| CN103060405B (en) | Fermentation technique of A40926 | |
| CN1724649A (en) | Entomopathogenic nematode symbiotic bacteria fermentation process and application of fermentation liquor thereof | |
| CN101884394B (en) | Composite microbial feed additive for chicken and preparation method thereof | |
| CN101194917B (en) | Grifola frondosa oral liquid rich in gastrodine, organic selenium and method for preparing the same | |
| CN101333547A (en) | Method for producing phenyl-lactic acid by controlling pH value, feeding and fermenting | |
| CN103103135B (en) | Aspergillus oryzae strain and method for producing fungal alpha-amylase by liquid-state fermentation of aspergillus oryzae strain | |
| CN103992977A (en) | Method for culturing bacillus subtilis through high-density fermentation | |
| CN103966271A (en) | Method for producing DHA through fermentation | |
| CN101880703B (en) | Method for fermenting daptomycin by adding caprate | |
| CN103710291B (en) | The method of one strain bacillus megaterium Z2013513 and production phenyl-lactic acid thereof | |
| CN105420143A (en) | Acetobacter orientalis and method for producing astragalus polysaccharide through same | |
| CN108251334A (en) | The microorganism mixed bacterial and fermentation process of a kind of fermenting lactic acid | |
| CN102634553B (en) | Hainanmycin fermentation method | |
| CN103695510A (en) | Method for producing Daptomycin by fed-batch methyl oleate fermentation | |
| CN102703357A (en) | Application of polyaspartic acid as agricultural bacillus spray drying protective agent | |
| CN101861796B (en) | Method for culturing amanita pantherina by using waste distillage after fermentation of coloured rice | |
| CN103695511A (en) | Method for producing daptomycin by fermentation | |
| CN103695496A (en) | Method for producing tacrolimus by fermentation | |
| CN107815431A (en) | A kind of method with water-soluble raw material culture bacillus subtilis | |
| CN103436472A (en) | Composite micro-ecological preparation for improving pond water quality |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20140402 |
|
| RJ01 | Rejection of invention patent application after publication |