CN106399136B - A kind of Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor - Google Patents
A kind of Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor Download PDFInfo
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Abstract
The invention discloses a kind of Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor, and this method is easy to operate, can effectively improve fermentation efficiency while keeping thallus vigor.Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor of the invention is to use molasses, urea, ammonium dihydrogen phosphate to carry out weighing flow feeding as nutrient source using self-priming reactor and carry out high density fermentation to Saccharomyces cerevisiae.
Description
Technical field
The present invention relates to a kind of methods of Saccharomyces cerevisiae High Density Cultivation, more specifically to a kind of using self-priming anti-
The Saccharomyces cerevisiae batch fermentation process in high density for answering device, belongs to bio-fermentation engineering field.
Background technique
Yeast is a kind of unicellular microorganism, and amino acid content is high, contains protein 50% or so, is rich in B family vitamin, also
There is the costly physiological activator of enzyme system and diversified economy abundant.Yeast is a kind of microorganism for having very much application value,
In terms of food industry, it is widely used as excellent leavening and the nutrition of the food such as mankind's bread as basic food, steamed bun, biscuit cake
Agent；In terms of medical industry, yeast is by as adjuvant treatment medicament or as culture medium；In basic research field, yeast is micro-
Biologist, geneticist carry out the optimal mode material of basic research；In addition, the fat extracted from yeast, can be used for
Soap manufacture, it may also be used for flotation tungsten ore, and the protein extracted from yeast can be used as feed.
In the production process of yeast, bioreactor has highly important effect, in occupation of the core of entire technique
Position.The structure of reactor, mode of operation and operating condition have considerable influence to the production of fermented product.Yeast hair
The efficient oxygen supply and energy conservation of ferment device, it is of crucial importance for yeast fermenting and producing.Bioreactor currently used for yeast production
Mostly bubbling style reactor and tradition machinery stirred reactor.For two type of bubbling style reactor and mechanical stir-reactor
The traditional reactor of type, the equipment for requiring a supply compressed air, such as air compressor machine or roots blower.First is that power consumption
It is larger, second is that needing to remove the water mist and mist of oil carried secretly in compressed air in air purification process, need a set of complex
Air cleaning system, it is thus possible to because air supply system in oil, water separation be not thorough, lead to microbiological contamination, influence yeast
Production.In addition, mechanically stirred reactor heat and mass transfer performance is preferable, solid particle suspension is good, therefore mechanic whirl-nett reaction
Device is widely used in chemical industry, but in view of its gas Utilization ability is insufficient, needs to increase gas compressor
Unreacted recyclegas is handled, operating cost is increased.Self-priming reactor is produced for Saccharomyces cerevisiae, production effect
Rate is high, good in economic efficiency.In self-priming reactor, high speed rotor makes Gas-Liquid Dispersion, reduces liquid-film resistance, is sucked by rotor
Air forms small bubble, keeps gas-liquid contact more uniform, and gas-liquid contact surface is constantly updated, and increases dissolved oxygen, improves oxygen
Utilization rate promotes the effect of mass transmitting of microbial metabolism, strengthens heat exchange.Self-priming reactor can use stirring turbine or liquid
Vacuum caused by the jet-action of flowing is from air is sucked, so not needing roots blower or air compressor machine.Therefore energy consumption also phase
To smaller.The speed of agitator of self-priming reactor is higher than traditional reactor, and power of agitator is also higher, but due to not needing pneumatics
The total output consumption of machine, self-priming reactor reduces about 30% instead.Since Saccharomyces cerevisiae possesses very strong tolerance shearing force, because
This self-priming reactor, which is applied to Saccharomyces cerevisiae production, can preferably play its advantage, embody yield height, high-quality advantage.
High density fermentation is always one of main direction of studying of fermentation arts, it is considered that yeast dry weight 30g/L with
Upper is high density fermentation, and the factor of limitation yeast high density fermentation is mainly that nutrition supply is not suitable for, and forms substrate suppression,
Produce accumulation and the influence of fermentation liquid rheological properties etc. of inhibitory substance.Yeast in aerobic fermentation, due to for hypoxgia,
Specific growth rate is excessive or carbon source supply amount is higher than the assimilable rate of yeast institute, will the metabolic by-products such as producing and ethanol, to bacterium
Body growth generates inhibition.In addition, the generation of a large amount of foams uses defoaming agent with unreasonable in high density large scale fermentation
It will lead to fermentation failure.In order to solve the problem of that high concentration substrate inhibition needs to develop new method to solve to exist.
Summary of the invention
Present invention aim to address above-mentioned problems of the prior art and deficiency, provide a kind of using self-priming anti-
The Saccharomyces cerevisiae batch fermentation process in high density of device is answered, this method is easy to operate, can effectively mention while keeping thallus vigor
High fermentation efficiency.
The present invention is achieved by the following technical solutions:
Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor of the invention is to utilize self-priming reaction
Device uses molasses, urea, ammonium dihydrogen phosphate to carry out weighing flow feeding as nutrient source and carries out high density hair to Saccharomyces cerevisiae
Ferment.
The above-mentioned Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor of the invention comprising following step
It is rapid:
The processing of molasses: sugar content mass fraction is diluted to 25%~30% with distilled water by cane molasses, uses sulfuric acid
PH to 4.8 is adjusted, heating is boiled to 95 DEG C, and divulge information 30min or more, then is kept the temperature 70~80 DEG C and stood 8~12h, and preceding use is used
The sterilizing of autoclave wet process, 121 DEG C of sterilising temp, sterilization time 20min；
The processing of urea: urea is diluted to the solution of leaded wastewater 10g/100mL；
The processing of ammonium dihydrogen phosphate: ammonium dihydrogen phosphate is diluted to P2O5Count the solution of P content 10g/100mL；
The addition of vitamin and biotin: vitamin B is added after carrying out real disappear to self-priming reactor10.2g, dimension life
Plain B50.2g, vitamin B60.2g, biotin 0.1g, additive amount is on the basis of self-priming reactor bottom water 12.5L；
The preparation of strain liquid: malt juice liquid medium, pH 4.8, with shaking flask culture, shaking flask liquid amount 25% are used
~30%, inoculum concentration 10%, shaking table culture 30 DEG C of cultivation temperature, revolving speed 200rpm, cultivates 10~12h, prepares seed liquor；
The high density fermentation of Saccharomyces cerevisiae: fedbatch culture feed supplement high density is carried out to Saccharomyces cerevisiae using self-priming reactor
Fermentation.
The above-mentioned Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor of the invention, further skill
Art scheme be in the process of high-density fermentation of the Saccharomyces cerevisiae molasses stream dosage according to current yeast in self-priming reactor
The 2.3 of dry weight add sugaring honey amount doubly, and molasses additive amount is reduced 50%~60% by 2h before fermentation ends, until fermentation knot
Beam, which stops flowing, to be added, and flow of urea dosage is 80g/h, stops stream after 20h and adds, and ammonium dihydrogen phosphate stream dosage is 30g/h, is stopped after 16h
Stream adds, and 25g MgSO is added in every 8h4, it is added 3 times altogether.Further technical solution is the high density hair of the Saccharomyces cerevisiae
The control of residual sugar content is within 1.5% during ferment, and alcoholic strength content controls between 0.7%~1.0%, when molten in reactor
Sugaring honey, urea and ammonium dihydrogen phosphate are flowed when oxygen rapid increase, fermentation middle and later periods DO control is 20%~30%.
The above-mentioned Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor of the invention, further skill
Art scheme can also be in the process of high-density fermentation of the Saccharomyces cerevisiae control of 0~16h fermentation temperature at 31 DEG C, 16~
21h control is at 32 DEG C, and 22~controlled for 24 hours at 33 DEG C.
The above-mentioned Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor of the invention, further skill
Art scheme can also be the preceding 8h pH control of its in the process of high-density fermentation of the Saccharomyces cerevisiae 4.3~4.8；In fermentation
Phase pH controls 4.5~4.8；2h pH control is adjusted in 5.0~5.2, pH by sulfuric acid and sodium carbonate liquor before fermentation ends.
The above-mentioned Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor of the invention, further skill
Art scheme can also be that the initial speed of self-priming reactor in the process of high-density fermentation of the Saccharomyces cerevisiae is 600rpm,
Revolving speed is increased 100rpm by 16h before fermenting, every 3h, and self-priming revolving speed is 1000rpm when 16h, and every 2h increases revolving speed after 16h
100rpm, until fermentation ends.
The above-mentioned Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor of the invention, further skill
Art scheme can also be in the process of high-density fermentation of the Saccharomyces cerevisiae adds defoaming agent according to the foam of generation in right amount.
The above-mentioned Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor of the invention, further skill
Art scheme can also be that the self-priming reactor is the self-priming reactor of 50L.
Compared with prior art the invention has the following advantages:
High density fermentation is carried out using Saccharomyces cerevisiae, feed supplement accurately controls bottoms stream dosage per hour by weighing, and reduces
For yeast because of anaerobic respiration problem caused by residual sugar too high levels, the used cane molasses that ferment are a kind of in fermentation process
The byproduct of sugar industry saves fermentation costs.By using this technique, after 24~26h fermentation, yeast weight in wet base can be stablized
260g/L or so is maintained, yeast dry weight has reached the standard of high density fermentation, while yeast milk fermentation up to 70g/L or so
Power can maintain 650mL/h or more, and the quality for the drying-granulating link in baker's active dried yeast downstream provides guarantee.Pass through two
Grade fermentation, shortens whole fermentation period, current adding substrate type is few in fermentation process, and the entire every h of fermenting step need to only change sugar
Sweet additive amount can effectively improve yeast weight in wet base, simplify fermentation associated operating steps, reduce the excessive bring of culture technique
Living contaminants chance is connected easy to operate between grade, ensure that the fermenting power of yeast.
Detailed description of the invention
Fig. 1 is yeast weight in wet base-time plot in multiple batches of fermenting experiment in the embodiment of the present invention 1.
Fig. 2 is ventilation quantity-time plot in 1 fermentation process of embodiment.
Fig. 3 is residual sugar-time plot in 1 fermentation process of embodiment.
Fig. 4 is molasses additive amount-time plot in 1 fermentation process of embodiment.
Fig. 5 is multilevel alcoholic strength controlling curve in fermentation process in the embodiment of the present invention 2.
Fig. 6 is the signal that Saccharomyces cerevisiae weight in wet base changes with alcoholic strength controlled level in fermentation process in the embodiment of the present invention 2
Figure.
Specific embodiment
The Saccharomyces cerevisiae strain used in embodiment is saccharomyces cerevisiae (CICC 1532), is mainly used as Saccharomyces cerevisiae
Production, self-priming reactor are the self-priming reactor of 50L；The processing method of molasses, urea, ammonium dihydrogen phosphate in embodiment is such as
Under:
The processing of molasses: sugar content mass fraction is diluted to 25%~30% with distilled water by cane molasses, uses sulfuric acid
PH to 4.8 is adjusted, heating is boiled to 95 DEG C, and divulge information 30min or more, then is kept the temperature 70~80 DEG C and stood 8~12h, and preceding use is used
The sterilizing of autoclave wet process, 121 DEG C of sterilising temp, sterilization time 20min；
The processing of urea: urea is diluted to the solution of leaded wastewater 10g/100mL；
The processing of ammonium dihydrogen phosphate: ammonium dihydrogen phosphate is diluted to P2O5Count the solution of P content 10g/100mL.
Embodiment 1
(1) with 1000mL shaking flask with malt juice liquid medium (pH4.8) culture, shaking flask liquid amount 25%~30% is connect
Kind amount 10%, shaking table culture 30 DEG C of cultivation temperature, revolving speed 200rpm, cultivate 10~12h, prepare seed liquor 2.5L；
(2) above-mentioned seed liquor is inoculated into containing 12.5L bottom water (vitamin B10.2g, vitamin B50.2g, vitamin
B60.2g, biotin 0.1g) the self-priming reactor of 50L in, be inoculated with eve calibration dissolved oxygen electrode DO be 100%；
(3) molasses stream dosage adds sugaring honey amount doubly according to the 2.3 of yeast dry weight quality current in self-priming reactor, fermentation
Terminate preceding 2h, molasses additive amount is reduced 50%~60%, until fermentation ends stop stream and add, flow of urea dosage is 8g/h, 20h
Stop flowing afterwards adding, ammonium dihydrogen phosphate stream dosage is 3g/h, stops stream after 16h and adds, every 8h addition 25g MgSO4, it is added 3 times altogether；
(4) control fermentation process in ventilation quantity be preceding 9h, with 300L/h be initial ventilation quantity, be gradually increased to 2400L/h,
Until 1h before fermentation ends, is reduced to 1800L/h；
(5) control of 0~16h fermentation temperature is at 31 DEG C in fermentation process, and 16~21h control is at 32 DEG C, and 22~control for 24 hours
33℃；
(6) 4.3~4.8, ferment middle is controlled before 4.5~4.8, fermentation ends for preceding 8h in fermentation process, pH control
2h is controlled 5.0~5.2；
(7) fermentation 26h stops fermenting, every 2h measurement yeast weight in wet base in fermentation process, centrifuge speed 10000r/min, from
Heart time 10min.
(8) more batch weight are in the above way carried out to test again, verifies the stability of experimental result in this experimental method.
Embodiment 2
(1) with 1000mL shaking flask with malt juice liquid medium (pH4.8) culture, shaking flask liquid amount 25%~30% is connect
Kind amount 10%, shaking table culture 30 DEG C of cultivation temperature, revolving speed 200rpm, cultivate 10~12h, prepare seed liquor 2.5L；
(2) above-mentioned seed liquor is inoculated into containing 12.5L bottom water (vitamin B10.2g, vitamin B50.2g, vitamin
B60.2g, biotin 0.1g) the self-priming reactor of 50L in, be inoculated with eve calibration dissolved oxygen electrode DO be 100%；
(3) molasses stream dosage adds sugaring honey amount doubly according to the 2.3 of yeast dry weight quality current in self-priming reactor, fermentation
Terminate preceding 2h, molasses additive amount is reduced 50%~60%, until fermentation ends stop stream and add, flow of urea dosage is 8g/h, 20h
Stop flowing afterwards adding, ammonium dihydrogen phosphate stream dosage is 3g/h, stops stream after 16h and adds, every 8h addition 25g MgSO4, it is added 3 times altogether；
(4) control fermentation process in ventilation quantity be preceding 9h, with 300L/h be initial ventilation quantity, be gradually increased to 2400L/h,
Until 1h before fermentation ends, is reduced to 1800L/h；
(5) control of 0~16h fermentation temperature is at 31 DEG C in fermentation process, and 16~21h control is at 32 DEG C, and 22~control for 24 hours
33℃；；
(6) 4.3~4.8, ferment middle is controlled before 4.5~4.8, fermentation ends for preceding 8h in fermentation process, pH control
2h is controlled 5.0~5.2；
(7) fermentation 26h stops fermenting, every 2h measurement yeast weight in wet base in fermentation process, centrifuge speed 10000r/min, from
Heart time 10min.
(8) five batch weight are carried out to test again, controlled after 12h in fermentation process alcoholic strength be divided into maintain 4%, 3%, 2%,
1.5%, on 1.0%, 0.7% horizontal line, growth curve of yeast is compared.Fig. 6 is Saccharomyces cerevisiae weight in wet base in fermentation process with wine
The schematic diagram of precision controlling level variation can be seen from the chart can there there be alcoholic strength 0.7%~1.0% in control fermentation process
Effect improves maximum yeast weight in wet base in 260g/L or so.
Claims (7)
1. a kind of Saccharomyces cerevisiae batch fermentation process in high density using self-priming reactor, it is characterised in that utilize self-priming anti-
Device is answered, uses molasses, urea, ammonium dihydrogen phosphate to carry out weighing flow feeding as nutrient source and high density hair is carried out to Saccharomyces cerevisiae
Ferment；The fermentation process the following steps are included:
The processing of molasses: sugar content mass fraction is diluted to 25%~30% with distilled water by cane molasses, is adjusted with sulfuric acid
PH4.8, heating are boiled to 95 DEG C, and divulge information 30min or more, then are kept the temperature 70~80 DEG C and stood 8~12h, use autoclave before use
Wet process sterilizing, 121 DEG C of sterilising temp, sterilization time 20min or more；
The processing of urea: urea is diluted to the solution of leaded wastewater 10g/100mL；
The processing of ammonium dihydrogen phosphate: ammonium dihydrogen phosphate is diluted to P2O5Count the solution of P content 10g/100mL；
The addition of vitamin and biotin: vitamin B is added after carrying out real disappear to self-priming reactor10.2g, vitamin
B50.2g, vitamin B60.2g, biotin 0.1g, additive amount is on the basis of self-priming reactor bottom water 12.5L；
The preparation of strain liquid: using malt juice liquid medium, pH 4.8, with shaking flask culture, shaking flask liquid amount 25%~
30%, inoculum concentration 10%, shaking table culture 30 DEG C of cultivation temperature, revolving speed 200rpm, cultivates 10~12h, prepares seed liquor；
The high density fermentation of Saccharomyces cerevisiae: fedbatch culture feed supplement high density is carried out to Saccharomyces cerevisiae using self-priming reactor and is sent out
Ferment；Molasses stream dosage is according to yeast dry weight matter current in self-priming reactor in the process of high-density fermentation of the Saccharomyces cerevisiae
The 2.3 of amount add sugaring honey amount doubly, and molasses additive amount is reduced 50%~60% by 2h before fermentation ends, until fermentation ends stop
It flows and adds, flow of urea dosage is 80g/h, and stopping, which is flowed, after 20h adds, and ammonium dihydrogen phosphate stream dosage is 30g/h, and stopping, which is flowed, after 16h adds, often
25gMgSO is added in 8h4, it is added 3 times altogether.
2. fermentation process in high density according to claim 1, it is characterised in that the high density fermentation of the Saccharomyces cerevisiae
The control of residual sugar content is within 1.5% in the process, and alcoholic strength content controls between 0.7%~1.0%, when self-priming reactor
Sugaring honey, urea and ammonium dihydrogen phosphate are flowed when middle dissolved oxygen rapid increase, fermentation middle and later periods DO control is 20%~30%.
3. fermentation process in high density according to claim 1, it is characterised in that the high density fermentation of the Saccharomyces cerevisiae
The control of 0~16h fermentation temperature is at 31 DEG C in the process, and 16~21h control is at 32 DEG C, and 22~controlled for 24 hours at 33 DEG C.
4. fermentation process in high density according to claim 1, it is characterised in that the high density fermentation of the Saccharomyces cerevisiae
Its preceding 8h pH control is 4.3~4.8 in the process；Ferment middle pH is controlled 4.5~4.8；2h pH control exists before fermentation ends
5.0~5.2, pH are adjusted by sulfuric acid and sodium carbonate liquor.
5. fermentation process in high density according to claim 1, it is characterised in that the high density fermentation of the Saccharomyces cerevisiae
The initial speed of self-priming reactor is 600rpm in the process, and revolving speed is increased 100rpm, every 2h after 16h by 16h before fermenting, every 3h
Revolving speed is increased into 100rpm, until fermentation ends.
6. fermentation process in high density according to claim 1, it is characterised in that the high density fermentation of the Saccharomyces cerevisiae
Defoaming agent is added according to the foam of generation in right amount in the process.
7. fermentation process in high density according to claim 1, it is characterised in that the reactor is the self-priming reaction of 50L
Device.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102191301A (en) * | 2011-03-31 | 2011-09-21 | 南通秋之友生物科技有限公司 | Method for producing ribonucleic acid by high-density fermentation high-nuclear Candida lipolytica |
| CN103413066A (en) * | 2013-08-28 | 2013-11-27 | 南京工业大学 | Forecasting method of self-suction reactor amplification fermentation culture yeast |
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2016
- 2016-11-16 CN CN201611024612.XA patent/CN106399136B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102191301A (en) * | 2011-03-31 | 2011-09-21 | 南通秋之友生物科技有限公司 | Method for producing ribonucleic acid by high-density fermentation high-nuclear Candida lipolytica |
| CN103413066A (en) * | 2013-08-28 | 2013-11-27 | 南京工业大学 | Forecasting method of self-suction reactor amplification fermentation culture yeast |
Non-Patent Citations (2)
| Title |
|---|
| Economic production of baker"s yeast using a new Saccharomyces cerevisiae isolate;Ehab Ragheb EI-Helow et al.;《Biotechnology&Biotechnological equipment》;20150501;第29卷(第4期);第705-713页 * |
| 固定化酵母发酵生产燃料乙醇研究进展;周天行等;《科技信息》;20111231;第3-4页 * |
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