CN105779535A - Culture medium for fermenting and producing enramycin and fermentation method - Google Patents
Culture medium for fermenting and producing enramycin and fermentation method Download PDFInfo
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- CN105779535A CN105779535A CN201610320834.XA CN201610320834A CN105779535A CN 105779535 A CN105779535 A CN 105779535A CN 201610320834 A CN201610320834 A CN 201610320834A CN 105779535 A CN105779535 A CN 105779535A
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/04—Linear peptides containing only normal peptide links
- C07K7/08—Linear peptides containing only normal peptide links having 12 to 20 amino acids
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- 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
Abstract
The invention discloses a culture medium for fermenting and producing enramycin and a fermentation method. The culture medium is prepared from a shake flask culture medium, a seed culture medium and a fermentation culture medium, wherein the shake flask culture medium further is prepared from the following components: 4.0wt% of dextrin, 3.0wt% of maize slurry, 2wt% of soyabean cake powder, 0.2wt% of ammonium chloride, 1.5wt% of light calcium carbonate, 0.06wt% of glycerophosphoryl ethanolamine and the balance of water. The seed culture medium further is prepared from the following components: 3.0wt% of dextrin, 3.0wt% of maize slurry, 2wt% of soyabean cake powder, 0.2wt% of ammonium chloride, 2.5wt% of light calcium carbonate, 0.06wt% of glycerophosphoryl ethanolamine and the balance of water. The fermentation culture medium further is prepared from the following components: 2-6wt% of dextrin, 0.5-2wt% of maize slurry, 1-2wt% of maize flour, 1-3wt% of soyabean cake powder, 1-2wt% of sunflower seed oil, 0.01wt% of magnesium sulfate, 0.01wt% of ferrous sulfate, 0.2-0.5wt% of ammonium chloride, 1-2wt% of light calcium carbonate, 0.07wt% of glycerophosphoryl ethanolamine and the balance of water. The culture medium fermentation process is simple to control, free of material supplement and short in period; the culture medium is low in production cost and is especially suitable for large-scale production.
Description
Technical field
The invention belongs to technical field of microbial fermentation, be specifically related to culture medium and the fermentation process of a kind of producing enramycin by fermentation.
Background technology
Enramycin (Enramycin) has another name called Enramycin, enramycin, enramycin, enramycin, it is a kind of ring type polypeptide class antibiotic, it is made up of 13 different types of 17 amino acid moleculars and fatty acid molecule, amino acid molecular composition ring type polypeptide, fatty acid molecule is connected on the aspartic acid of end, difference according to terminal aliphatic molecule is divided into enramycin A and enramycin B, enramycin is the mixture of both materials, it is that actinomycetes (StreptomycesefungicidiousNO.B-5477) the antifungal streptomycete fermentation separated from the soil in western palace city, Japan Hyogo Prefecture by military field (Takeda) pharmaceutical industries Co., Ltd. of Japan research worker in 1966 produces.Gram positive bacteria is had very strong inhibitory action by enramycin, the main synthesis hindering bacteria cell wall.Its sensitivity has various gram-positive coccis, bacillus subtilis, anthrax bacillus, clostridium tetani, bacillus botulinus, the bacillus perfringens etc. such as staphylococcus aureus.Life-time service enramycin medicine is not likely to produce drug resistance, and intestinal flora distribution can be changed, be conducive to the absorption of Parenteral Nutrition composition, can promoting the increase of the weight of animals and improve the utilization rate of feedstuff, therefore many countries recommend these products as the antibiotic of poultry and growth promoter in the world.1993, the Ministry of Agriculture of China ratified this medicine and registers in China.2005, domestic production enterprise and Schering Plough animal health-care product company limited of the U.S. (ScheringPloughAnimalHealthCorp.) started co-production enramycin premix.Owing to enramycin has excellent growth promotion and improves the effect of efficiency of feed utilization, therefore recommended as antibiotic growth promoter by many countries in the world.China granted patent CN102174624B, by sweat moisturizing, improves fermentation middle and late stage dissolved oxygen level, and final fermentation level is less than 1000u/ml.The basestocks such as China granted patent CN201010609213.6 and CN101899490B all employ glucose, and sweat creates glucose effect, and early stage mycelial growth is too fast, bacterium is dense too high, and late growing stage is obstructed, and fermentation period extends, power and the cost of raw material are all higher, uneconomical.China granted patent CN102787153A and CN104195203A etc. sweat respectively in fill into inorganic in a large number and Organic Ingredients or the nutritional labeling such as sugar and aminoacid, operating process is complicated, add microbiological contamination probability, the a large amount of materials filled into, causing feed liquid thickness after feed supplement, sweat requires big ventilation and big power of agitator or dilute, and utilization rate is low, improve cost, effect is unsatisfactory.
Summary of the invention
For solving the problems referred to above existed in prior art and practical situation, the invention provides the culture medium of a kind of producing enramycin by fermentation.This culture medium includes Shake flask medium, seed culture medium and fermentation medium, described Shake flask medium farther includes: dextrin 4.0wt%, Semen Maydis pulp 3.0wt%, soybean cake powder 2wt%, ammonium chloride 0.2wt%, precipitated calcium carbonate 1.5wt%, bubble enemy 0.06wt%, water surplus；
Described seed culture medium farther includes: dextrin 3.0wt%, Semen Maydis pulp 3.0wt%, soybean cake powder 2wt%, ammonium chloride 0.2wt%, precipitated calcium carbonate 2.5wt%, bubble enemy 0.06wt%, water surplus；
Described fermentation medium farther includes: dextrin 2～6wt%, Semen Maydis pulp 0.5～2wt%, Semen Maydis powder 1～2wt%, soybean cake powder 1～3wt%, Oleum Helianthi 1～2wt%, sulphuric acid U.S. 0.01wt%, ferrous sulfate 0.01wt%, ammonium chloride 0.2～0.5wt%, precipitated calcium carbonate 1～2wt%, bubble enemy 0.07wt%, water surplus.
Preferably, the pH of described Shake flask medium is 6.8-7.2.
Preferably, the pH of described seed culture medium is 6.8-7.5.
Preferably, the PH of described fermentation medium is 6.8-7.8.
Another aspect of the present invention, it is provided that a kind of method of producing enramycin by fermentation, the seed liquor of cabicidin streptomycete is seeded in above-mentioned culture medium and carries out liquid fermentation, after fermentation culture 180-200 hour, extracts enramycin from fermented liquid.
Preferably, the temperature of producing enramycin by fermentation is 26～28 DEG C, and speed of agitator is 30-100r/min, and ventilation is 0.2-1.0vvm.
Preferably, in fermentation culture process, pH is 6.8-7.2.
Beneficial effect:
Culture medium fermentation processes provided by the invention is simple, and without feed supplement, the cycle is short, and the fermentation period of 192 hours can make enramycin fermentation level reach 9000～10000u/ml, and production cost is low, is particularly suitable for large-scale production.
Detailed description of the invention
Technical solution of the present invention is described in detail below.
Embodiment one
Take cold preservation and be seeded in sterilized Shake flask medium in-80 cabicidin streptomycete cryovial bacterial strains spent in refrigerator by the inoculum concentration of 1%, cultivation temperature 26～28 DEG C, when rotating speed 220r/min, cultivate 30h；By cultured seed liquor by the inoculum concentration of 0.2%, it is seeded to the 1.0m equipped with 400L culture medium by pressure reduction inocalation method3In first class seed pot, cultivate after 2 days for 28 DEG C, move into equipped with 57m3The 75m of fermentation medium3In fermentation tank, speed of agitator 0 ~ 50h is 30 ~ 60r/min, 50 ~ 192h is 60 ~ 100r/min；Ventilation starts as 0.2vvm, and sweat steps up with the decline of dissolved oxygen, brings up to 1.0vvm, temperature 26～28 DEG C, cultivate and put tank in 8 days after 60 hours, and fermentation level is 9100u/ml, puts tank volume 52 tons.
Shake flask medium: dextrin 4.0wt%, Semen Maydis pulp 3.0wt%, soybean cake powder 2wt%, ammonium chloride 0.2wt%, precipitated calcium carbonate 1.5wt%, bubble enemy 0.06wt%, water surplus, pH7.0.
Seed tank culture base: dextrin 3.0wt%, Semen Maydis pulp 3.0wt%, soybean cake powder 2wt%, ammonium chloride 0.2wt%, precipitated calcium carbonate 2.5wt%, bubble enemy 0.06wt%, water surplus, pH6.9.
Fermentation tank culture medium: dextrin 4wt%, Semen Maydis pulp 2wt%, Semen Maydis powder 2wt%, soybean cake powder 3wt%, Oleum Helianthi 1.5wt%, sulphuric acid U.S. 0.01wt%, ferrous sulfate 0.01wt%, ammonium chloride 0.4wt%, precipitated calcium carbonate 1.5wt%, bubble enemy 0.07wt%, water surplus, pH6.8.
Example two
Take cold preservation and be seeded in sterilized Shake flask medium in-80 cabicidin streptomycete cryovial bacterial strains spent in refrigerator by the inoculum concentration of 1%, cultivation temperature 26～28 DEG C, when rotating speed 220r/min, cultivate 30h；By cultured seed liquor by the inoculum concentration of 0.2%, it is seeded to the 1.0m equipped with 400L culture medium by pressure reduction inocalation method3In first class seed pot, cultivate after 2 days for 28 DEG C, move into equipped with 56.4m3The 75m of fermentation medium3In fermentation tank, speed of agitator 0 ~ 50h is 20 ~ 60r/min, 50 ~ 192h is 60 ~ 100r/min；Ventilation starts as 0.3vvm, and sweat steps up with the decline of dissolved oxygen, brings up to 1.0vvm, temperature 26～28 DEG C after 50 hours, and cultivating 200h fermentation level is 9450u/ml, puts tank volume 51 tons.
Shake flask medium: dextrin 4.0wt%, Semen Maydis pulp 3.0wt%, soybean cake powder 2wt%, ammonium chloride 0.2wt%, precipitated calcium carbonate 1.5wt%, bubble enemy 0.06wt%, water surplus, pH6.8
Seed tank culture base: dextrin 3.0wt%, Semen Maydis pulp 3.0wt%, soybean cake powder 2wt%, ammonium chloride 0.2wt%, precipitated calcium carbonate 2.5wt%, bubble enemy 0.06wt%, water surplus, pH6.8.
Fermentation tank culture medium: dextrin 6wt%, Semen Maydis pulp 1.5wt%, Semen Maydis powder 1.5wt%, soybean cake powder 2wt%, Oleum Helianthi 2wt%, sulphuric acid U.S. 0.01wt%, ferrous sulfate 0.01wt%, ammonium chloride 0.5wt%, precipitated calcium carbonate 2wt%, bubble enemy 0.07wt%, water surplus, pH7.4.
Example three
Take cold preservation and be seeded in sterilized Shake flask medium in-80 cabicidin streptomycete cryovial bacterial strains spent in refrigerator by the inoculum concentration of 1%, cultivation temperature 26～28 DEG C, when rotating speed 220r/min, cultivate 30h；By cultured seed liquor by the inoculum concentration of 0.2%, it is seeded to the 1.0m equipped with 350L culture medium by pressure reduction inocalation method3In first class seed pot, cultivate after 2 days for 28 DEG C, move into equipped with 53m3The 75m of fermentation medium3In fermentation tank, speed of agitator 0 ~ 50h is 30 ~ 60r/min, 50 ~ 192h is 60 ~ 90r/min；Ventilation starts for 0.3vvm, to step up with the decline of sweat dissolved oxygen, bring up to 1.0vvm, temperature 26～28 DEG C, cultivate and put tank in 8 days after 80 hours, and fermentation level is 9670u/ml, puts tank volume 49.5 tons.Shake flask medium: dextrin 4.0wt%, Semen Maydis pulp 3.0wt%, soybean cake powder 2wt%, ammonium chloride 0.2wt%, precipitated calcium carbonate 1.5wt%, bubble enemy 0.06wt%, water surplus, pH7.1
Seed tank culture base: dextrin 3.0wt%, Semen Maydis pulp 3.0wt%, soybean cake powder 2wt%, ammonium chloride 0.2wt%, precipitated calcium carbonate 2.5wt%, bubble enemy 0.06wt%, water surplus, pH7.2.
Fermentation tank culture medium: dextrin 5.5wt%, Semen Maydis pulp 2wt%, Semen Maydis powder 2wt%, soybean cake powder 1wt%, Oleum Helianthi 1wt%, sulphuric acid U.S. 0.01wt%, ferrous sulfate 0.01wt%, ammonium chloride 0.4wt%, precipitated calcium carbonate 2wt%, bubble enemy 0.07wt%, water surplus, pH7.0.
The beneficial effects of the present invention is without feed supplement, basic components employs dextrin cleverly, not only effectively reduces feed liquid denseness, and avoids the glucose effect that the microbial decomposition metabolism caused because of high concentration glucose in formula is obstructed.In addition, without glucose in basic components of the present invention, utilize the feature of the enzyme of cabicidin streptomycete metabolism growth process generation hydrolyzable dextrin and Semen Maydis powder, the reducing sugar that can make sweat without feed supplement is stable 1～2%, mycelia is made to keep higher more stable physiologically active, bacterium is dense is maintained at 45～50% for a long time, ensure that effective picked-up of mycelial vigor and oxygen, promoted thalline by primary metabolite to secondary metabolites enramycin produce excessive, improve enramycin fermentation level, shorten fermentation period, reduce raw material and power cost, it is a kind of efficient, economical, simple and practical production technology, it is particularly suitable for big production.
Last it is noted that the foregoing is only the preferred embodiments of the present invention, it is not limited to the present invention, although the present invention being described in detail with reference to previous embodiment, for a person skilled in the art, technical scheme described in foregoing embodiments still can be modified by it, or wherein portion of techniques feature carries out equivalent replacement.All within the spirit and principles in the present invention, any amendment of making, equivalent replacement, improvement etc., should be included within protection scope of the present invention.
Claims (7)
1. a culture medium for producing enramycin by fermentation, including Shake flask medium, seed culture medium and fermentation medium, it is characterised in that:
Described Shake flask medium farther includes: dextrin 4.0wt%, Semen Maydis pulp 3.0wt%, soybean cake powder 2wt%, ammonium chloride 0.2wt%, precipitated calcium carbonate 1.5wt%, bubble enemy 0.06wt%, water surplus；
Described seed culture medium farther includes: dextrin 3.0wt%, Semen Maydis pulp 3.0wt%, soybean cake powder 2wt%, ammonium chloride 0.2wt%, precipitated calcium carbonate 2.5wt%, bubble enemy 0.06wt%, water surplus；
Described fermentation medium farther includes: dextrin 2～6wt%, Semen Maydis pulp 0.5～2wt%, Semen Maydis powder 1～2wt%, soybean cake powder 1～3wt%, Oleum Helianthi 1～2wt%, sulphuric acid U.S. 0.01wt%, ferrous sulfate 0.01wt%, ammonium chloride 0.2～0.5wt%, precipitated calcium carbonate 1～2wt%, bubble enemy 0.07wt%, water surplus.
2. culture medium according to claim 1, it is characterised in that the pH of described Shake flask medium is 6.8-7.2.
3. culture medium according to claim 1, it is characterised in that the pH of described seed culture medium is 6.8-7.5.
4. culture medium according to claim 1, it is characterised in that the PH of described fermentation medium is 6.8-7.8.
5. the method for a producing enramycin by fermentation, it is characterized in that, the seed liquor of cabicidin streptomycete is seeded in the culture medium described in Claims 1-4 any one and carries out liquid fermentation, after fermentation culture 180-200 hour, extracts enramycin from fermented liquid.
6. method according to claim 5, it is characterised in that the temperature of producing enramycin by fermentation is 26～28 DEG C, speed of agitator is 30-100r/min, and ventilation is 0.2-1.0vvm.
7. method according to claim 6, it is characterised in that in fermentation culture process, pH is 6.8-7.2.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109628532A (en) * | 2018-12-30 | 2019-04-16 | 广东容大生物股份有限公司 | Fermentation medium and its application |
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| CN102787153A (en) * | 2012-09-07 | 2012-11-21 | 山东胜利生物工程有限公司 | Method for producing enramycin by microbial fermentation supplement feed |
| CN102943101A (en) * | 2012-11-19 | 2013-02-27 | 安徽丰原发酵技术工程研究有限公司 | Method for producing enramycin by fermenting |
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| CN104419740A (en) * | 2013-09-03 | 2015-03-18 | 河南天方药业股份有限公司 | Method for fermenting enramycin |
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- 2016-05-16 CN CN201610320834.XA patent/CN105779535A/en active Pending
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| CN101899490A (en) * | 2010-07-14 | 2010-12-01 | 山东胜利股份有限公司 | Method for producing enramycin by using microbial fermentation |
| CN102787153A (en) * | 2012-09-07 | 2012-11-21 | 山东胜利生物工程有限公司 | Method for producing enramycin by microbial fermentation supplement feed |
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| CN104419740A (en) * | 2013-09-03 | 2015-03-18 | 河南天方药业股份有限公司 | Method for fermenting enramycin |
| CN104131054A (en) * | 2014-06-17 | 2014-11-05 | 安徽丰原发酵技术工程研究有限公司 | Fermentation culture medium and fermentation method for improving enramycin yield |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN109628532A (en) * | 2018-12-30 | 2019-04-16 | 广东容大生物股份有限公司 | Fermentation medium and its application |
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Inventor after: Sun Mei Inventor after: Chen Jianhui Inventor after: Lin Panjiao Inventor after: Fu Wenli Inventor after: Shi Mei Inventor after: Zhou Ping Inventor after: Zhao Fei Inventor after: Wang Yu Inventor after: Kou Wei Inventor before: Sun Mei Inventor before: Chen Jianhui Inventor before: Lin Panjiao Inventor before: Fu Wenli Inventor before: Zhou Ping Inventor before: Zhao Fei Inventor before: Wang Yu Inventor before: Kou Wei |
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Application publication date: 20160720 |