CN106047967A - Culture medium for Streptomyces actuosus fermentation production of nosiheptide, and culture method - Google Patents
Culture medium for Streptomyces actuosus fermentation production of nosiheptide, and culture method Download PDFInfo
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Abstract
The present invention relates to a culture medium for Streptomyces actuosus fermentation production of nosiheptide, and a culture method. According to the present invention, the seed culture medium for Streptomyces actuosus fermentation production of nosiheptide, and the carbon source, the nitrogen source and optimal compatibility ratio in the fermentation culture medium are determined; and with the culture medium formula and the fermentation control process of the present invention, the nosiheptide fermentation unit can be increased, the fermentation cost can be reduced, the influence of the environment on the raw material source and the auxiliary material source can be minimized, the sufficient supply can be ensured, and the stable and efficient production of the nosiheptide can be achieved.
Description
Technical field
The invention belongs to fermentation technical field, particularly relate to the culture medium of a kind of S.actuosus fermenting and producing nosiheptide
And cultural method.
Background technology
Nosiheptide, also known as nosiheptide, is a kind of antibiotic, uses S.actuosus fermentation pattern to produce, and current nosiheptide is
Being put into European Pharmacopoeia and Japan's feed safety method, many countries and regions are all using nosiheptide.The Ministry of Agriculture of China is by that
Western peptide is classified as the feed additive that can add for a long time in feedstuff.
At present, Related domestic documents reports nosiheptide production technology, and its fermentation pattern is respectively second order fermentation or three grades
Fermentation.There is the problem that
1 industrialized great production fermentation level is relatively low, and its fermentation level maintains about 1500-1800ug/L.
The 2 domestic complete fermentation technology documents of nosiheptide that lack are reported, and relevant document report is based on test,
Technical support effectively can not be provided for big production model.
3 nosiheptide big production fermentation period is long, and typically at about 240h, energy consumption is high.
4 nosiheptide fermentation comprehensive production costs are higher, and product lacks the market competitiveness.
Summary of the invention
The purpose of the present invention is that the defect overcoming above-mentioned prior art, it is provided that a kind of fermentation technology is simple, effectively carries
High fermentation unit, reduces the supplementary material impact on fermentation unit the most to greatest extent, reduces production cost, and supplementary material source
The most affected by environment, it is ensured that it is in liberal supply, it is achieved nosiheptide is stable, the S.actuosus fermenting and producing nosiheptide of efficient production
Culture medium.
It is a further object of the present invention to provide the cultural method utilizing above-mentioned culture medium to produce nosiheptide.
The technical scheme taked for achieving the above object is:
The culture medium of a kind of S.actuosus fermenting and producing nosiheptide, it is characterised in that include seed culture medium and fermentation culture
Base, wherein
Consisting of of described seed culture medium:
Bextran 45～8g/L, glycogen 8～12g/L, beerwort 6～10ml/L, fish flour 9～13g/L, cottonseed meal 11～15g/
L, Zein powder 13～17g/L, ammonium sulfate 0.04～0.08g/L, precipitated calcium carbonate 1～5g/L；
Consisting of of described fermentation medium:
Glucosan 6～10g/L, glycogen 10～14g/L, beerwort 8～12ml/L, fish flour 11～15g/L, cottonseed meal 13～
17g/L, Zein powder 14～18g/L, pupa albumen peptone 7～11g/L, Sargassum powder 4～8g/L, ammonium sulfate 0.06～0.8g/
L, precipitated calcium carbonate 1～5g/L, potassium dihydrogen phosphate 0.05～0.09g/L, magnesium sulfate 0.03～0.07g/L, nickel sulfate 0.006～
0.01g/L, sodium chloride 0.02～0.06g/L, polyether modified silicon oil 0.5～0.9ml/L, liquid alkane 0.3～0.7ml/L.
Described liquid alkane refers to n-dodecane and the mixture of n-tridecane oil, and wherein the content of n-dodecane is 85-
90%。
A kind of cultural method utilizing above-mentioned culture medium fermenting and producing nosiheptide, it is characterised in that its processing step is:
1) seed culture: first by seed culture medium sterilizing and be cooled to 20～25 DEG C, and use filtrated air pressurize, then at fire
Under flame protection, by the most cultured S.actuosus mother's bottle fermentation liquid according to 2～3L/m3Inoculum concentration access seed culture medium
In cultivate, to cell concentration 26～30%, incubation time 40～45h subcultivation；
2) fermentation culture: first by fermentation medium sterilizing, is cooled to 20～25 DEG C, and uses filtrated air pressurize, then will cultivate
Good seed liquor moves into fermentation medium and carries out fermentation culture, to cell concentration 38～42%, fermentation period less than 220h, change
Learn titer ＞ 2300ug/ml and terminate fermentation.
Prescription after described seed culture medium sterilizing is: amino nitrogen ＞ 30mg/L, molten phosphorus 100～140ug/ml, fat
Fat 6～10g/L；
Prescription after described fermentation medium sterilizing is: amino nitrogen ＞ 50mg/L, molten phosphorus 40～80ug/ml, fat 12～
16g/L。
Described cultured S.actuosus mother's bottle fermentation liquid prescription is: cell concentration 20～30%；Microscopy is without miscellaneous
Bacterium.
Described seed culture condition is: tank pressure 0.03～0.06MPa；Tank temperature 28～29 DEG C；Air mass flow: 30～50m3/
h；Speed of agitator 60～80r/min.
Described fermentation culture conditions is:
The initial pH of a culture medium: after fermentation medium sterilizing, pH controls 5～7；
B temperature: cultivation temperature 28～29 DEG C,
C speed of agitator: rotating speed controls at 100-120r/min,
D Stress control: tank pressure 0.05～0.06MPa；
E pH controls: in sweat, pH controls 5～7；
F air mass flow: 300～400m3/ h,
H dissolved oxygen controls:
Before fermentation in 80h, do not control dissolved oxygen,
81～160h, dissolved oxygen controls more than 40%,
161h～fermentation ends: dissolved oxygen controls more than 30%.
Using stream addition to carry out feed supplement in described sweat, feed supplement includes mending cane molasses, moisturizing, benefit ammonium sulfate, its
In
A mends cane molasses technology controlling and process:
Filling into cane molasses at fermentation 90h, 150h respectively, its dosage is the 0.2-0.3% of fermentating liquid volume；
B moisturizing technology controlling and process:
Before fermentation, 80h need not carry out moisturizing,
Fermentation time, 81～100h, when cell concentration is higher than 45%, adds aquesterilisa, it is desirable to controls fermentation liquid cell concentration and exists
40～45%, detect fermentation liquid cell concentration every 6～8h,
Fermentation time, in 101h～fermentation ends, when cell concentration is higher than 42%, adds aquesterilisa, it is desirable to control fermentation liquid thalline
Concentration, 38～42%, detects fermentation liquid cell concentration every 6～8h；
C mends ammonium sulfate:
Fill into the ammonium sulfate of 10% when fermenting 80h and 120h, its consumption controls at the 0.01～0.03% of fermentating liquid volume.
The technical advantage of the present invention is embodied in:
1 present invention confirms carbon source in the seed culture medium of S.actuosus fermenting and producing nosiheptide, fermentation medium, nitrogen source
With optimal proportion compatibility.
2 present invention have carried out detailed elaboration to the technique of seed culture, fermentation culture, it is thus identified that nosiheptide is optimal
Fermentation technology and control parameter.By fermenting and producing nosiheptide of the present invention, its fermentation unit reaches more than 2300ug/ml, same to state
Interior technology is compared, and fermentation unit improves more than 27%；Fermentation period controls within 220h, reduces fermentation energy consumption.
3 present invention are applicable to industrially scalable fermenting and producing nosiheptide.
Specific implementation method
It is explained the present invention below, it should be understood that example is for illustrating rather than the present invention with example
Restriction.The scope of the present invention is determined according to claims with core content.
The strain of following embodiment selects S.actuosus: the strain source of production and application is female bottle fermentation liquid.Female bottle is sent out
Ferment liquid prescription is: cell concentration 20～30%；Microscopy is without miscellaneous bacteria.
Embodiment 1
Seed culture: culture volume 10m3。
Gentran 40 kg, glycogen 80kg, beerwort 60L, fish flour 90kg, cottonseed meal 110 kg, Zein powder
130kg, ammonium sulfate 0.4kg, precipitated calcium carbonate 10kg；
First by seed culture medium sterilizing and be cooled to 20 DEG C, and filtrated air pressurize is used, the quality after seed culture medium sterilizing:
Amino nitrogen 32mg/L, molten phosphorus 101ug/ml, fat 6.2g/L；Then, under flame is protected, strepto-is enlivened by the most cultured
Starter bottle fermentation liquid accesses in seed culture medium according to the inoculum concentration of 20L and cultivates.Seed culture condition is: tank pressure 0.03～
0.06MPa；Tank temperature 28～29 DEG C；Air mass flow: 30m3/h；Speed of agitator 60r/min, to cell concentration 26%, incubation time
40h subcultivation.
Fermentation culture: culture volume 100m3。
Dextran 60 0kg, glycogen 1000kg, beerwort 800L, fish flour 1100kg, cottonseed meal 1300kg, zein
Powder 1400kg, pupa albumen peptone 700kg, Sargassum powder 400kg, ammonium sulfate 6kg, precipitated calcium carbonate 100kg, potassium dihydrogen phosphate 5kg,
Magnesium sulfate 3kg, nickel sulfate 0.6kg, sodium chloride 2kg, polyether modified silicon oil 50L, liquid alkane 30L.
First by fermentation medium sterilizing, it is cooled to 20 DEG C, and uses filtrated air pressurize, the quality after fermentation medium sterilizing
For: amino nitrogen 53mg/L, molten phosphorus 42ug/ml, fat 12g/L.Then cultured seed liquor immigration fermentation medium is carried out
Fermentation culture.
Fermentation culture conditions is:
The initial pH of a culture medium: after fermentation medium sterilizing, pH controls 5.0；
B temperature: cultivation temperature 28～29 DEG C,
C speed of agitator: rotating speed controls at 100r/min,
D Stress control: tank pressure 0.05～0.06MPa；
E pH controls: in sweat, pH controls 5～7；
F air mass flow: 300m3/ h,
H dissolved oxygen controls:
Before fermentation in 80h, do not control dissolved oxygen,
81～160h, dissolved oxygen controls more than 40%,
161～fermentation ends: dissolved oxygen controls more than 30%.
Fermentation culture terminates, cell concentration 38%, fermentation period 210h, chemical titer 2308ug/ml.
Embodiment 2
Seed culture: culture volume 10m3。
Glucosan 50kg, glycogen 90kg, beerwort 70L, fish flour 100kg, cottonseed meal 140 kg, Zein powder
140kg, ammonium sulfate 0.5kg, precipitated calcium carbonate 20kg；
First by seed culture medium sterilizing and be cooled to 22 DEG C, and filtrated air pressurize is used, the quality after seed culture medium sterilizing:
Amino nitrogen 35mg/L, molten phosphorus 111ug/ml, lipase 37 g/L；Then under flame is protected, by the most cultured S.actuosus
Female bottle fermentation liquid accesses in seed culture medium according to the inoculum concentration of 22L and cultivates.Seed culture condition is: tank pressure 0.03～
0.06MPa；Tank temperature 28～29 DEG C；Air mass flow: 35m3/h；Speed of agitator 65r/min, to cell concentration 27%, incubation time
41h subcultivation.
Fermentation culture: culture volume 100m3。
Macrodex 0kg, glycogen 1100kg, beerwort 900L, fish flour 1200kg, cottonseed meal 1400kg, zein
Powder 1500kg, pupa albumen peptone 800kg, Sargassum powder 500kg, ammonium sulfate 7kg, precipitated calcium carbonate 200kg, potassium dihydrogen phosphate 6kg,
Magnesium sulfate 4kg, nickel sulfate 0.7kg, sodium chloride 3kg, polyether modified silicon oil 60L, liquid alkane 40L.
First by fermentation medium sterilizing, it is cooled to 22 DEG C, and uses filtrated air pressurize, the quality after fermentation medium sterilizing
For: amino nitrogen 57mg/L, molten phosphorus 51ug/ml, fat 13g/L.Then cultured seed liquor immigration fermentation medium is carried out
Fermentation culture.
Fermentation culture conditions is:
The initial pH of a culture medium: after fermentation medium sterilizing, pH controls 5.5；
B temperature: cultivation temperature 28～29 DEG C,
C speed of agitator: rotating speed controls at 105r/min,
D Stress control: tank pressure 0.05～0.06MPa；
E pH controls: in sweat, pH controls 5～7；
F air mass flow: 320m3/ h,
H dissolved oxygen controls:
Before fermentation in 80h, do not control dissolved oxygen,
81～160h, dissolved oxygen controls more than 40%,
161～fermentation ends: dissolved oxygen controls more than 30%.
Fermentation culture terminates, cell concentration 39%, fermentation period 212h, chemical titer 2374ug/ml.
Embodiment 3
Seed culture: culture volume 10m3。
Dextran 60 kg, glycogen 100kg, beerwort 80L, fish flour 110kg, cottonseed meal 150 kg, Zein powder
150kg, ammonium sulfate 0.6kg, precipitated calcium carbonate 30kg；
First by seed culture medium sterilizing and be cooled to 23 DEG C, and filtrated air pressurize is used, the quality after seed culture medium sterilizing:
Amino nitrogen 39mg/L, molten phosphorus 122ug/ml, fat 8g/L；Then under flame is protected, by the most cultured S.actuosus
Female bottle fermentation liquid accesses in seed culture medium according to the inoculum concentration of 25L and cultivates.Seed culture condition is: tank pressure 0.03～
0.06MPa；Tank temperature 28～29 DEG C；Air mass flow: 40m3/h；Speed of agitator 70r/min, to cell concentration 28%, incubation time
42h subcultivation.
Fermentation culture: culture volume 100m3。
Dextran 8 00kg, glycogen 1200kg, beerwort 1000L, fish flour 1300kg, cottonseed meal 1500kg, zein
Powder 1600kg, pupa albumen peptone 900kg, Sargassum powder 600kg, ammonium sulfate 8kg, precipitated calcium carbonate 300kg, potassium dihydrogen phosphate 7kg,
Magnesium sulfate 5kg, nickel sulfate 0.8kg, sodium chloride 4kg, polyether modified silicon oil 70L, liquid alkane 50L.
First by fermentation medium sterilizing, it is cooled to 23 DEG C, and uses filtrated air pressurize, the quality after fermentation medium sterilizing
For: amino nitrogen 62mg/L, molten phosphorus 60ug/ml, fat 14g/L.Then cultured seed liquor immigration fermentation medium is carried out
Fermentation culture.
Fermentation culture conditions is:
The initial pH of a culture medium: after fermentation medium sterilizing, pH controls 6；
B temperature: cultivation temperature 28～29 DEG C,
C speed of agitator: rotating speed controls at 110r/min,
D Stress control: tank pressure 0.05～0.06MPa；
E pH controls: in sweat, pH controls 5～7；
F air mass flow: 350m3/ h,
H dissolved oxygen controls:
Before fermentation in 80h, do not control dissolved oxygen,
81～160h, dissolved oxygen controls more than 40%,
161～fermentation ends: dissolved oxygen controls more than 30%.
Fermentation culture terminates, cell concentration 40%, fermentation period 215h, chemical titer 2468ug/ml.
Embodiment 4
Seed culture: culture volume 10m3。
Macrodex kg, glycogen 110kg, beerwort 90L, fish flour 120kg, cottonseed meal 160 kg, Zein powder
160kg, ammonium sulfate 0.7kg, precipitated calcium carbonate 40kg.
First by seed culture medium sterilizing and be cooled to 24 DEG C, and filtrated air pressurize is used, after seed culture medium sterilizing
Quality: amino nitrogen 43mg/L, molten phosphorus 131ug/ml, fat 9g/L；Then, under flame is protected, chain is enlivened by the most cultured
Mould starter bottle fermentation liquid accesses in seed culture medium according to the inoculum concentration of 28L and cultivates.Seed culture condition is: tank pressure 0.03
～0.06MPa；Tank temperature 28～29 DEG C；Air mass flow: 45m3/h；Speed of agitator 75r/min, to cell concentration 29%, incubation time
43h subcultivation.
Fermentation culture: culture volume 100m3。
Glucosan 900kg, glycogen 1300kg, beerwort 1100L, fish flour 1400kg, cottonseed meal 1600kg, zein
Powder 1700kg, pupa albumen peptone 1000kg, Sargassum powder 700kg, ammonium sulfate 9kg, precipitated calcium carbonate 400kg, potassium dihydrogen phosphate
8kg, magnesium sulfate 6kg, nickel sulfate 0.9kg, sodium chloride 5kg, polyether modified silicon oil 80L, liquid alkane 60L.
First by fermentation medium sterilizing, it is cooled to 24 DEG C, and uses filtrated air pressurize, the quality after fermentation medium sterilizing
For: amino nitrogen 66mg/L, molten phosphorus 71ug/ml, fat 15g/L.Then cultured seed liquor immigration fermentation medium is carried out
Fermentation culture.
Fermentation culture conditions is:
The initial pH of a culture medium: after fermentation medium sterilizing, pH controls 6.5；
B temperature: cultivation temperature 28～29 DEG C,
C speed of agitator: rotating speed controls at 115r/min,
D Stress control: tank pressure 0.05～0.06MPa；
E pH controls: in sweat, pH controls 5～7；
F air mass flow: 370m3/ h,
H dissolved oxygen controls:
Before fermentation in 80h, do not control dissolved oxygen,
81～160h, dissolved oxygen controls more than 40%,
161～fermentation ends: dissolved oxygen controls more than 30%.
Fermentation culture terminates, cell concentration 41%, fermentation period 217h, chemical titer 2390ug/ml.
Embodiment 5
Seed culture: culture volume 10m3。
Dextran 8 0kg, glycogen 120kg, beerwort 100L, fish flour 130kg, cottonseed meal 150 kg, Zein powder
170kg, ammonium sulfate 0.8kg, precipitated calcium carbonate 50kg.
First by seed culture medium sterilizing and be cooled to 25 DEG C, and filtrated air pressurize is used, after seed culture medium sterilizing
Quality: amino nitrogen 47mg/L, molten phosphorus 140ug/ml, fat 10g/L；Then under flame is protected, by the most cultured active
Strepto-starter bottle fermentation liquid accesses in seed culture medium according to the inoculum concentration of 30L and cultivates.Seed culture condition is: tank pressure
0.03～0.06MPa；Tank temperature 28～29 DEG C；Air mass flow: 50m3/h；Speed of agitator 80r/min, to cell concentration 30%, cultivates
Time 44h subcultivation.
Fermentation culture: culture volume 100m3。
Glucosan 1000kg, glycogen 1400kg, beerwort 1200L, fish flour 1500kg, cottonseed meal 1700kg, Semen Maydis egg
White lead 1800kg, pupa albumen peptone 1100kg, Sargassum powder 800kg, ammonium sulfate 10kg, precipitated calcium carbonate 500kg, potassium dihydrogen phosphate
9kg, magnesium sulfate 7kg, nickel sulfate 1kg, sodium chloride 6kg, polyether modified silicon oil 90L, liquid alkane 70L.
First by fermentation medium sterilizing, it is cooled to 25 DEG C, and uses filtrated air pressurize, the quality after fermentation medium sterilizing
For: amino nitrogen 70mg/L, molten phosphorus 80ug/ml, fat 16g/L.Then cultured seed liquor immigration fermentation medium is carried out
Fermentation culture.
Fermentation culture conditions is:
The initial pH of a culture medium: after fermentation medium sterilizing, pH controls 7
B temperature: cultivation temperature 28～29 DEG C,
C speed of agitator: rotating speed controls at 120r/min,
D Stress control: tank pressure 0.05～0.06MPa；
E pH controls: in sweat, pH controls 5～7；
F air mass flow: 400m3/ h,
H dissolved oxygen controls:
Before fermentation in 80h, do not control dissolved oxygen,
81～160h, dissolved oxygen controls more than 40%,
161～fermentation ends: dissolved oxygen controls more than 30%.
Fermentation culture terminates, cell concentration 42%, fermentation period 220h, chemical titer 2327ug/ml.
In the sweat of above-described embodiment 1-5, according to detection case, using stream addition to carry out feed supplement, feed supplement includes mending
Cane molasses, moisturizing, benefit ammonium sulfate.
A mends cane molasses technology controlling and process:
Filling into cane molasses at 90h, 150h respectively, its dosage is the 0.2-0.3% of fermentating liquid volume.
B moisturizing technology controlling and process:
Before fermentation, 80h need not carry out moisturizing,
Fermentation time, 81～100h, when cell concentration is higher than 45%, adds aquesterilisa, it is desirable to controls fermentation liquid cell concentration and exists
40～45%, detect fermentation liquid cell concentration every 6～8h,
Fermentation time, in 101h～fermentation ends, when cell concentration is higher than 42%, adds aquesterilisa, it is desirable to control fermentation liquid thalline
Concentration, 38～42%, detects fermentation liquid cell concentration every 6～8h；
C mends ammonium sulfate:
Fill into the ammonium sulfate of 10% when fermenting 80h and 120h, its consumption controls at the 0.01～0.03% of fermentating liquid volume.
Comparative example's 1(three grade fermemtation pattern)
First order seed is cultivated: culture volume is 1m3
Soluble starch 7kg, glucose 10kg, analysis for soybean powder 12kg, peptone 11kg, Semen Maydis pulp 15kg, ammonium sulfate 0.05kg, light
Matter calcium carbonate 4kg.
First by primary-seed medium sterilizing and be cooled to 25 DEG C, and filtrated air pressurize is used, seed culture medium sterilizing
After quality: amino nitrogen 3.4mg/L, molten phosphorus 12ug/ml, fat 0.9g/L；Then under flame is protected, by the most cultured
S.actuosus mother's bottle fermentation liquid accesses in seed culture medium according to the inoculum concentration of 2L and cultivates.Seed culture condition is: tank
Pressure 0.03～0.06MPa；Tank temperature 28～29 DEG C；Air mass flow: 10m3/h；Speed of agitator 80r/min, to cell concentration 20%, training
Support time 40h subcultivation.
Secondary seed is cultivated: culture volume 10m3。
Soluble starch 80kg, glucose 120kg, analysis for soybean powder 130kg, peptone 150 kg, Semen Maydis pulp 170kg, yeast
Cream 120kg, 130 ammonium sulfate 0.8kg, precipitated calcium carbonate 50kg.
First by seed culture medium sterilizing and be cooled to 25 DEG C, and filtrated air pressurize is used, after seed culture medium sterilizing
Quality: amino nitrogen 42mg/L, molten phosphorus 129ug/ml, fat 8g/L；Then under flame is protected, by the most cultured one-level kind
Sub-liquid accesses in secondary seed medium and cultivates.Secondary seed condition of culture is: tank pressure 0.03～0.06MPa；Tank temperature 28
～29 DEG C；Air mass flow: 50m3/h；Speed of agitator 80r/min, to cell concentration 30%, incubation time 45h subcultivation.
Fermentation culture: culture volume 100m3。
Solubility 1200kg, glucose 1600kg, analysis for soybean powder 1700kg, peptone 900kg, Semen Maydis pulp 1400kg, yeast
Cream 1100kg, glycine 100kg, ammonium sulfate 9kg, precipitated calcium carbonate 500kg, potassium dihydrogen phosphate 9kg, magnesium sulfate 7kg, sulphuric acid are sub-
Ferrum 6kg, manganese sulfate 1kg, sodium chloride 6kg.
Fermentation culture conditions is:
The initial pH of a culture medium: after fermentation medium sterilizing, pH controls at 6-6.5,
B temperature: cultivation temperature 28～29 DEG C,
C speed of agitator: rotating speed controls at 120r/min,
D Stress control: tank pressure 0.05～0.06MPa；
E pH controls: in sweat, pH controls 5～7；
F air mass flow: 500m3/ h,
Fermentation culture terminates, cell concentration 38%, fermentation period 240h, chemical titer 1735ug/ml.
Comparative example's 2(second order fermentation pattern)
Seed culture: culture volume 10m3。
Liquefying starch 100kg, glucose 100kg, lactose 60kg, soybean cake powder 120kg, peptone 130 kg, Semen Maydis pulp
160kg, dextrin 120kg, ammonium sulfate 0.9kg, precipitated calcium carbonate 40kg.
First by seed culture medium sterilizing and be cooled to 25 DEG C, and filtrated air pressurize is used, after seed culture medium sterilizing
Quality: amino nitrogen 36mg/L, molten phosphorus 131ug/ml, fat 9g/L；Then, under flame is protected, chain is enlivened by the most cultured
Mould starter bottle fermentation liquid accesses in seed culture medium according to the inoculum concentration of 30L and cultivates.Seed culture condition is: tank pressure 0.03
～0.06MPa；Tank temperature 28～29 DEG C；Air mass flow: 40m3/h；Speed of agitator 80r/min, to cell concentration 35%, incubation time
48h subcultivation.
Fermentation culture: culture volume 100m3。
Liquefying starch 1100kg, glucose 1300kg, lactose 400kg, soybean cake powder 1400kg, peptone 1000kg, jade
Rice & peanut milk 1300kg, dextrin 1300kg, glutamic acid 80kg, glycine 100kg, cysteine 75kg, ammonium sulfate 8kg, lightweight carbonic acid
Calcium 400kg, potassium dihydrogen phosphate 10kg, magnesium sulfate 6kg, ferrous sulfate 7kg, manganese sulfate 0.8kg, sodium chloride 5kg.
Fermentation culture conditions is:
The initial pH of a culture medium: after fermentation medium sterilizing, pH controls at 6-6.5
B temperature: cultivation temperature 28～29 DEG C,
C speed of agitator: rotating speed controls at 120r/min,
D Stress control: tank pressure 0.05～0.06MPa；
E pH controls: in sweat, pH controls 5～7；
F air mass flow: 500m3/ h,
Fermentation culture terminates, cell concentration 40%, fermentation period 240h, chemical titer 1513ug/ml.
Claims (8)
1. the culture medium of a S.actuosus fermenting and producing nosiheptide, it is characterised in that include seed culture medium and fermentation culture
Base, wherein
Consisting of of described seed culture medium:
Bextran 45～8g/L, glycogen 8～12g/L, beerwort 6～10ml/L, fish flour 9～13g/L, cottonseed meal 11～15g/
L, Zein powder 13～17g/L, ammonium sulfate 0.04～0.08g/L, precipitated calcium carbonate 1～5g/L；
Consisting of of described fermentation medium:
Glucosan 6～10g/L, glycogen 10～14g/L, beerwort 8～12ml/L, fish flour 11～15g/L, cottonseed meal 13～
17g/L, Zein powder 14～18g/L, pupa albumen peptone 7～11g/L, Sargassum powder 4～8g/L, ammonium sulfate 0.06～0.8g/
L, precipitated calcium carbonate 1～5g/L, potassium dihydrogen phosphate 0.05～0.09g/L, magnesium sulfate 0.03～0.07g/L, nickel sulfate 0.006～
0.01g/L, sodium chloride 0.02～0.06g/L, polyether modified silicon oil 0.5～0.9ml/L, liquid alkane 0.3～0.7ml/L.
2. according to the culture medium described in claim 1, it is characterised in that described liquid alkane refers to n-dodecane and n-tridecane
The mixture of oil, wherein the content of n-dodecane is 85-90%.
3. one kind utilizes the cultural method of culture medium fermenting and producing nosiheptide described in claim 1 or 2, it is characterised in that its technique
Step is:
1) seed culture: first by seed culture medium sterilizing and be cooled to 20～25 DEG C, and use filtrated air pressurize, then at fire
Under flame protection, by the most cultured S.actuosus mother's bottle fermentation liquid according to 2～3L/m3Inoculum concentration access seed culture medium
In cultivate, to cell concentration 26～30%, incubation time 40～45h subcultivation；
2) fermentation culture: first by fermentation medium sterilizing, is cooled to 20～25 DEG C, and uses filtrated air pressurize, then will cultivate
Good seed liquor moves into fermentation medium and carries out fermentation culture, to cell concentration 38～42%, fermentation period less than 220h, change
Learn titer ＞ 2300ug/ml and terminate fermentation.
4. according to the cultural method described in claim 3, it is characterised in that the prescription after described seed culture medium sterilizing is:
Amino nitrogen ＞ 30mg/L, molten phosphorus 100～140ug/ml, fat 6～10g/L；
Prescription after described fermentation medium sterilizing is: amino nitrogen ＞ 50mg/L, molten phosphorus 40～80ug/ml, fat 12～
16g/L。
5. according to the cultural method described in claim 3, it is characterised in that described cultured S.actuosus mother's bottle fermentation liquid
Prescription is: cell concentration 20～30%；Microscopy is without miscellaneous bacteria.
6. according to the cultural method described in claim 3, it is characterised in that described seed culture condition is: tank pressure 0.03～
0.06MPa；Tank temperature 28～29 DEG C；Air mass flow: 30～50m3/h；Speed of agitator 60～80r/min.
7. according to the cultural method described in claim 3, it is characterised in that described fermentation culture conditions is:
The initial pH of a culture medium: after fermentation medium sterilizing, pH controls 5～7；
B temperature: cultivation temperature 28～29 DEG C,
C speed of agitator: rotating speed controls at 100-120r/min,
D Stress control: tank pressure 0.05～0.06MPa；
E pH controls: in sweat, pH controls 5～7；
F air mass flow: 300～400m3/ h,
H dissolved oxygen controls:
Before fermentation in 80h, do not control dissolved oxygen,
81～160h, dissolved oxygen controls more than 40%,
161h～fermentation ends: dissolved oxygen controls more than 30%.
8. according to the cultural method described in claim 3, it is characterised in that described sweat uses stream addition carry out feed supplement,
Feed supplement includes mending cane molasses, moisturizing, benefit ammonium sulfate, wherein
A mends cane molasses technology controlling and process:
Filling into cane molasses at fermentation 90h, 150h respectively, its dosage is the 0.2-0.3% of fermentating liquid volume；
B moisturizing technology controlling and process:
Before fermentation, 80h need not carry out moisturizing,
Fermentation time, 81～100h, when cell concentration is higher than 45%, adds aquesterilisa, it is desirable to controls fermentation liquid cell concentration and exists
40～45%, detect fermentation liquid cell concentration every 6～8h,
Fermentation time, in 101h～fermentation ends, when cell concentration is higher than 42%, adds aquesterilisa, it is desirable to control fermentation liquid thalline
Concentration, 38～42%, detects fermentation liquid cell concentration every 6～8h；
C mends ammonium sulfate:
Fill into the ammonium sulfate of 10% when fermenting 80h and 120h, its consumption controls at the 0.01～0.03% of fermentating liquid volume.
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