CN110684680B - Preparation method of high-density yeast fermentation liquor - Google Patents
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Abstract
The invention relates to a preparation method of high-density yeast fermentation liquor, which comprises the steps of mixing corn and bean pulp with water, adding high-temperature amylase, saccharifying enzyme and acidic protein respectively, carrying out thorough enzymolysis, and carrying out solid-liquid separation to obtain solid wet residue and dilute composite nutrient solution. Diluting a part of the dilute composite nutrient solution, inoculating the diluted composite nutrient solution into a yeast seed solution for culturing, and evaporating the rest of the dilute composite nutrient solution under reduced pressure to obtain a concentrated solution. And in the culture process, sufficient dissolved oxygen is controlled by feeding the concentrated composite nutrient solution and the nutrient salt, when the density of the yeast is maximum, feeding is stopped, the yeast fermentation liquid is obtained after the consumption of the fermentable sugar is completely consumed, and the high-density yeast liquid is obtained by standing. The invention is characterized in that different enzyme preparations are enzymolyzed to obtain sugar and amino acid required by yeast growth, and high-density yeast liquid is obtained by a continuous fed-batch fermentation batch discharging mode, so that the yeast content is improved, the consumption of fermentation sterilization steam is low, the production water is low, the discharge amount of tank washing wastewater of a fermentation tank is reduced, and the effects of energy conservation, consumption reduction and clean production are achieved.
Description
Technical Field
The invention belongs to the technical field of microbial fermentation, and particularly relates to a preparation method of high-density yeast fermentation liquor.
Background
The saccharomyces cerevisiae has high nutritive value of mycoprotein, the protein content in yeast dry matter can reach 50 percent, the content of essential amino acid required by animals is high, the content is close to animal protein, and the utilization rate of net protein is far higher than that of plant protein. In addition, the saccharomyces cerevisiae also contains abundant nucleotide, B vitamins, minerals and other physiological active substances. With the development of microbial engineering and fermentation engineering, saccharomyces cerevisiae is more and more widely used for producing high-quality protein feed.
Meanwhile, with the wide application of yeast in feed, the realization of high-density fermentation of yeast in a fermentation tank space with limited volume is a key for maintaining the production scale of yeast products and controlling the cost, so that research on various characteristics of yeast, particularly fermentation characteristics, is necessary.
According to the traditional yeast value-added fermentation method, high-density fermentation of saccharomyces cerevisiae is realized by optimizing culture conditions, and a set of complete fermentation process is formed by integrating and optimizing a plurality of factors and mutual influences thereof, and the process is a work with complex procedure and high comprehensiveness and is not beneficial to the requirement of industrial development. In addition, the raw materials such as glucose and yeast extract are mostly used as reducing sugar and nitrogen source for yeast culture, and the production cost is high. Therefore, according to the utilization mode of the saccharomyces cerevisiae on reducing sugar and nitrogen source, the yeast high-density culture production process which is simple and convenient to operate, suitable for large-scale industrial production, energy-saving and environment-friendly by adopting raw materials with wide sources and low price is needed to be developed.
Disclosure of Invention
The invention aims to utilize cheap raw materials, carry out enzymolysis by different enzyme preparations to obtain sugar and organic nitrogen sources required by yeast growth, obtain high-density yeast liquid in a continuous fed-batch fermentation batch discharging mode, improve the yeast content, have simple operation, small consumption of fermentation sterilization steam and less production water, reduce the discharge of tank washing wastewater of a fermentation tank, and achieve the effects of energy conservation, consumption reduction and clean production.
The technical scheme adopted by the invention is as follows:
a preparation method of high-density yeast fermentation liquor is characterized by comprising the following steps:
preparing a dilute composite nutrient solution and a concentrated composite nutrient solution:
crushing corn and bean pulp, adding water for size mixing, sequentially adding high-temperature amylase, saccharifying enzyme and protease, performing enzymolysis at controlled temperature, and performing enzymolysis completely to obtain liquid enzymolysis liquid; carrying out solid-liquid separation on the liquid enzymolysis liquid to prepare solid wet residues and a dilute composite nutrient solution;
reserving a part of the dilute composite nutrient solution as fermentation substrate sugar, and performing reduced pressure evaporation and concentration on the rest of the dilute composite nutrient solution to obtain a concentrated composite nutrient solution;
② preparation of high-density yeast liquid
In a full-automatic fermentation tank, diluting a part of dilute composite nutrient solution, sterilizing at high temperature, cooling, inoculating yeast seed solution, introducing sufficient sterile air for culture, controlling sufficient dissolved oxygen by respectively feeding the concentrated composite nutrient solution and diammonium hydrogen phosphate solution in a flowing manner in the culture process, stopping feeding when the yeast density is maximum, and preparing yeast fermentation liquor after the fermentable sugar is consumed;
keeping the fermentation tank at positive pressure, standing the yeast fermentation liquor, and discharging the precipitated yeast from the tank bottom after complete sedimentation to obtain high-density yeast liquid; and continuously adding the residual fermentation liquor in the fermentation tank with the concentrated composite nutrient solution and the diammonium hydrogen phosphate solution in a flowing manner to control sufficient dissolved oxygen, performing enrichment culture on the yeast again to the maximum density, standing, discharging high-density yeast liquid, continuously and circularly producing for multiple times, and collecting all the high-density yeast liquid.
Further, the yeast seed liquid is saccharomyces cerevisiae CICC 32280.
Further, the reaction conditions in step (i) are as follows:
the mass ratio of the corn to the soybean meal is 1: 0.01-0.2; adding water to adjust the concentration of the dry matter to 10-40%; the addition amount of the high-temperature amylase is 0.1-1 kg per ton of corn and soybean meal mixture, the pH value of the slurry is controlled to be 5.5-6.5, and the enzymolysis time is 0.5-2 hours;
the addition amount of the saccharifying enzyme is that 0.1-0.5 kg of controlled slurry pH 4.5-5.5 is added to each ton of corn and soybean meal mixture for enzymolysis for 0.5-20 h;
the addition amount of the protease is 0.1-1 kg per ton of the corn and soybean meal mixture, the pH of the slurry is controlled to be 4.5-5.5, and the enzymolysis time is 0.5-20 h;
the concentration of the concentrated dry matter of the dilute composite nutrient solution is 40-65%.
Preferably, the reaction conditions in step (i) are as follows:
the mass ratio of the corn to the soybean meal is 1: 0.05-0.1; adding water to adjust the concentration of the dry matter to be 25-30%; the addition amount of the high-temperature amylase is 0.3-0.5 kg per ton of the corn and soybean meal mixture, the pH value of the slurry is controlled to be 5.8-6.2, and the enzymolysis time is 0.5-1 h;
the addition amount of the saccharifying enzyme is that 0.4-0.5 kg of controlled slurry pH4.8-5.2 is added to each ton of corn and soybean meal mixture for enzymolysis for 5-10 h;
the addition amount of the protease is that 0.5-0.6 kg of controlled slurry pH is added to each ton of corn and soybean meal mixture, the enzymolysis time is 15-20 h;
the concentration of the concentrated dry matter of the dilute composite nutrient solution is 40-45%.
Further, the reaction conditions in step two are as follows:
diluting the dilute composite nutrient solution to a dry matter concentration of 0.01-5%; the mass ratio of diammonium hydrogen phosphate to water in the diammonium hydrogen phosphate solution is 10-50: 50-90; the feeding mass ratio of the concentrated compound nutrient solution to the diammonium hydrogen phosphate solution is 1: 0.001-0.1; the inoculation mass ratio of the yeast seed liquid is 0.2-20%;
when preparing the yeast fermentation liquor: fermentation temperature: 20-40 ℃; fermentation pH: 4.0 to 6.0; the stirring speed of aerobic fermentation is 20-1000 r/min; the ventilation ratio is 1: 0.5-3 v/v; the concentration of dissolved oxygen is 10-100%; and (3) fermentation period: 12-72 h, the number of bacteria at the fermentation end point of the yeast is 0.5-1.20 multiplied by 109cfu/ml；
The yeast culture end marker is: the content of fermentable sugar in the fermentation liquor is 0-0.1%, and the content of dissolved oxygen is 60-100%; standing and settling time is 1-20 h.
Preferably, the reaction conditions in step (ii) are as follows:
diluting the dilute composite nutrient solution to a dry matter concentration of 0.5-2%; the mass ratio of the diammonium hydrogen phosphate to the water in the diammonium hydrogen phosphate solution is 30-40: 60-70 parts of; the feeding mass ratio of the concentrated compound nutrient solution to the diammonium hydrogen phosphate solution is 1: 0.005-0.01; the inoculation mass ratio of the yeast seed liquid is 0.2-20%;
when preparing the yeast fermentation liquor: fermentation temperature: 30-35 ℃; fermentation pH: 4.0 to 4.5; the rotation speed of aerobic fermentation stirring is 100-150 r/min; the ventilation ratio is 1: 0.5-2 v/v; the concentration of dissolved oxygen is 20-40%; and (3) fermentation period: 48-56 h, the number of the bacteria at the end point of the yeast fermentation is 1.15-1.20 multiplied by 109cfu/ml；
The yeast culture end marker is: the content of fermentable sugar in the fermentation liquor is 0-0.05%, and the content of dissolved oxygen is 60-80%; standing and settling time is 4-6 h.
Further, in the step I, a part of the dilute composite nutrient solution accounts for 1-2% of the dilute composite nutrient by mass.
Furthermore, in the second step, the continuous cycle production times are 8-10.
The invention has the beneficial effects that:
1. cheap corn and bean pulp are subjected to enzymolysis through amylase, glucoamylase and protease to form reducing sugar and organic nitrogen source mixed nutrient solution which is suitable for the growth of yeast in required proportion, the reducing sugar and organic nitrogen source required for the growth of yeast are provided, and in addition, diammonium hydrogen phosphate solution is added to effectively replace dipotassium hydrogen phosphate and ammonium sulfate in the traditional process, so that phosphorus source and inorganic ammonia nitrogen required for the growth of yeast can be met, the sulfate ion content of fermentation liquor can be reduced, and the production raw material cost is saved.
2. By means of continuous feeding and concentration of the composite nutrient solution and diammonium hydrogen phosphate solution, the method can effectively control the dissolved oxygen content of the fermentation liquor to be stable, provide reducing sugar, nitrogen source and phosphorus source for sufficient yeast growth, avoid substrate inhibition caused by one-time feeding and reduction of fermentation efficiency caused by oxygen deficiency, obtain the maximum increment speed and the maximum density of the yeast, and further, keep the yeast cells to settle after the fermentation is finished, the high-density yeast liquid is obtained by the bottom batch partial discharge mode, the rest fermentation liquid can be continuously fed and fermented without high-temperature sterilization and cooling inoculation again, the high-density yeast fermentation liquor obtained by the method not only has the yeast content increased to 18 percent, but also has the advantages of simple operation, small consumption of fermentation and sterilization steam, less production water, reduction of the discharge amount of waste water from tank washing of a fermentation tank, and achievement of the effects of energy conservation, consumption reduction and clean production.
Detailed Description
Example 1
A preparation method of high-density yeast fermentation liquor comprises the following specific operation methods:
(1) and preparing the dilute composite nutrient solution and the concentrated composite nutrient solution:
crushing 95kg of corn and 5kg of soybean meal, adding 200kg of water to prepare slurry, adjusting the pH value to 6.0, adding 0.04kg of high-temperature amylase, and carrying out enzymolysis at the high temperature of 90 ℃ for 1 hour; cooling to 55 deg.C, adjusting pH to 5.2, adding 0.04kg saccharifying enzyme, and performing enzymolysis for 5 hr; adjusting the pH value to 4.8, adding 0.06kg of protease, and keeping the temperature for enzymolysis for 15h to obtain liquid enzymolysis liquid;
filtering the liquid enzymolysis liquid to obtain 260kg of filtrate which is the dilute composite nutrient solution, and evaporating and concentrating the dilute composite nutrient solution under reduced pressure to obtain 163kg of concentrated composite nutrient solution with the solid content of 44%.
(2) The production method of the high-density yeast liquid comprises the following steps:
diluting 2kg of dilute composite nutrient solution to 85L in a 200L full-automatic fermentation tank, wherein the concentration of dry matters is 0.5%, heating feed liquid to 90 ℃, heating a tank body above the liquid level to 121 ℃ for sterilization, keeping the temperature for sterilization for 30 minutes, cooling to 35 ℃, inoculating 5L of yeast seed solution, observing the initial volume to be 105L, adjusting the initial ventilation volume to be 1:0.5v/v, stirring the rotation speed to be 100r/min, and improving the rotation speed and the ventilation ratio when the dissolved oxygen is lower than 20%. Culturing for 8h, when dissolved oxygen suddenly rises to above 60%, feeding sterile concentrated composite nutrient solution and diammonium hydrogen phosphate solution by a peristaltic pump, wherein the feeding mass ratio of the concentrated composite nutrient solution to the diammonium hydrogen phosphate is 1:0.005, the pH is controlled to be 4.0-4.5, the dissolved oxygen is controlled to be 20-40%, the stirring speed is increased to 150r/min, the ventilation volume is 1:2v/v, culturing for 56h, the volume of the fermentation broth reaches 160L, and the number of detected yeast bacteria reaches 1.19 multiplied by 109cfu/ml, stopping feeding, and stopping ventilation and stirring until the dissolved oxygen rises to 70 percent. Standing for 4h to allow yeast to naturally settle, discharging 52L of the settled yeast through a discharge valve at the bottom of the tank, and detecting that the dry matter content of the yeast reaches 182 g/L.
Secondly, closing a tank bottom valve, recovering the ventilation quantity of 1:2v/v, stirring at the rotating speed of 150r/min, feeding sterile concentrated composite nutrient solution and diammonium phosphate solution again through a peristaltic pump, controlling the pH to be 4.0-4.5, controlling the dissolved oxygen to be 20% -40%, continuing fermentation culture for 52 hours, stopping feeding when the volume of fermentation liquor reaches 168L, and stopping ventilation and stirring when the dissolved oxygen rises back to 70%. Standing for 6h to allow yeast to naturally settle, discharging 56L of the settled yeast through a discharge valve at the bottom of the tank, and detecting that the dry matter content of the yeast reaches 186 g/L.
Example 2
A preparation method of high-density yeast fermentation liquor comprises the following specific steps:
(1) and preparing the dilute composite nutrient solution and the concentrated composite nutrient solution:
pulverizing 1850kg of semen Maydis and 150kg of soybean meal, adding 4400kg of water to obtain slurry, adjusting pH to 6.0, adding 0.8kg of high temperature amylase, and performing enzymolysis at 90 deg.C for 1 hr; cooling to 55 deg.C, adjusting pH to 5.2, adding 0.8kg saccharifying enzyme, and performing enzymolysis for 5 hr; adjusting the pH value to 4.8, adding 1.2kg of protease, and keeping the temperature for enzymolysis for 15 h; 5416kg of filtrate is obtained by filtering, namely the dilute compound nutrient solution, and the concentrated compound nutrient solution 3520kg is obtained by carrying out reduced pressure evaporation and concentration on the dilute compound nutrient solution, wherein the solid content is 40%.
(2) The production method of the high-density yeast liquid comprises the following steps:
firstly, diluting 25kg of dilute composite nutrient solution to 0.9m3 in a 2m3 fermentation tank, wherein the concentration of dry matters is 0.6%, heating feed liquid to 90 ℃, heating the tank above the liquid level to 121 ℃ for sterilization, keeping the temperature for sterilization for 30 minutes, cooling to 35 ℃, inoculating 100L of yeast seed solution, observing and recording the initial volume of 1.1m3, adjusting the initial ventilation amount to be 1:0.5v/v, stirring the rotation speed to be 100r/min, and increasing the rotation speed and the ventilation ratio when the dissolved oxygen is lower than 20%. Culturing for 6h, when dissolved oxygen suddenly rises to above 60%, feeding sterile concentrated composite nutrient solution and diammonium hydrogen phosphate solution by a peristaltic pump, wherein the feeding mass ratio of the concentrated composite nutrient solution to the diammonium hydrogen phosphate is 1:0.004, the pH is controlled to be 4.0-4.5, the dissolved oxygen is controlled to be 20% -40%, the stirring speed is increased to 150r/min, the ventilation volume is 1:1.8v/v, culturing for 56h, the volume of the fermentation broth reaches 1.54m3, and the number of detected yeast bacteria reaches 1.18 multiplied by 109cfu/ml, stopping feeding, and stopping ventilation and stirring until the dissolved oxygen rises to 70 percent. Standing for 6h to allow yeast to naturally settle, discharging the settled yeast through a tank bottom discharge valve to discharge 0.44m3, and detecting that the dry matter content of the yeast reaches 182 g/L.
Secondly, closing a tank bottom valve, recovering the ventilation quantity to be 1:1.8v/v, stirring at the rotating speed of 150r/min, feeding sterile concentrated composite nutrient solution and diammonium hydrogen phosphate solution by a peristaltic pump again, controlling the pH to be 4.0-4.5, controlling the dissolved oxygen to be 20% -40%, continuing fermentation culture for 50h, stopping feeding when the volume of the fermentation liquor reaches 1.45m3, and stopping ventilation and stirring when the dissolved oxygen rises to 70%. Standing for 6h to allow yeast to naturally settle, discharging the settled yeast through a tank bottom discharge valve to discharge 0.44m3, and detecting that the dry matter content of the yeast reaches 183 g/L.
Thirdly, closing a tank bottom valve, recovering the ventilation quantity to be 1:1.8v/v, stirring at the rotating speed of 150r/min, feeding sterile concentrated composite nutrient solution and diammonium phosphate solution again through a peristaltic pump, controlling the pH to be 4.0-4.5, controlling the dissolved oxygen to be 20% -40%, continuing fermentation culture for 52 hours, stopping feeding when the volume of the fermentation liquor reaches 1.46m3, and stopping ventilation and stirring when the dissolved oxygen is raised back to 70%. Standing for 6h to allow yeast to naturally settle, discharging the settled yeast through a tank bottom discharge valve to discharge 0.46m3, and detecting that the dry matter content of the yeast reaches 184 g/L.
Fourthly, 8 batches of the yeast are repeatedly cultured in the way, the yeast still grows well, the concentration of the yeast reaches more than 180g/L, and the yeast liquid with the high concentration of 3.67m3 is obtained in total.
Comparative example 1
Comparative example 1 is a conventional yeast preparation method, which specifically comprises:
firstly, preparing a fermentation culture medium: 65g/L of industrial glucose, 1.8g/L of urea, 10g/L of industrial corn flour and KH2PO410g/L，MgSO44g/L，ZnSO410mg/L，FeSO412mg/L，MnSO42mg/L。
Preparing a supplementary culture medium: 500g/L glucose solution.
Thirdly, inoculating the saccharomyces cerevisiae seed liquid into a 2m3 fermentation tank for fermentation according to the inoculation amount of 10 percent, controlling the culture temperature to be 30 ℃, controlling the pH to be 5.5, controlling the liquid loading amount to be 0.9m3, controlling the initial rotation speed to be 150r/min, and controlling the initial ventilation amount to be 1:2 v/v. Feeding materials at a rate of 2.4L/h when the fermentation is started to 10h, increasing the stirring speed to 200r/min, adjusting the aeration ratio to 1:2.5v/v, changing the fermentation in the middle and later stages according to dissolved oxygen, slightly increasing the feeding rate if the dissolved oxygen is suddenly increased to more than 30%, controlling the dissolved oxygen to maintain about 30%, ending the fermentation period of 55h, controlling the dry weight of the saccharomyces cerevisiae cells to be 122g/L, and controlling the cell number of the fermentation broth to be 1.05 multiplied by 109cfu/ml。
The yeast cell number and dry matter concentration of fermentation broth of the first, example 1, example 2 and comparative example 1 are shown in table 1 below.
Table 1, example 2 and comparative example 1 fermentation broth yeast cell number and retort yeast dry matter concentration
As can be seen from Table 1, the fermentation mode of controlling dissolved oxygen by feeding and enzymolysis of the composite nutrient solution during fermentation can obtain 1.19X 10 cells at the end of fermentation9cfu/ml is improved by 13% compared with the traditional fermentation mode of example 1, the concentration of dry substances of high-concentration yeast liquid obtained by a standing precipitation batch discharging mode in the fermentation process reaches 186g/L, the supernatant can continue to adopt a feed supplement mode to ferment and culture yeast, the fermentation efficiency is improved by 52% compared with the traditional fermentation mode of example 1, the utilization rate of equipment is greatly improved, the operation is simple, the consumption of fermentation sterilization steam is small, the production water is less, the discharge amount of waste water in tank washing of a fermentation tank is reduced, and the effects of energy conservation, consumption reduction and clean production are achieved.
Claims (6)
1. A preparation method of high-density yeast fermentation liquor is characterized by comprising the following steps:
preparing a dilute composite nutrient solution and a concentrated composite nutrient solution:
crushing corn and bean pulp, adding water for size mixing, sequentially adding high-temperature amylase, saccharifying enzyme and protease, performing temperature-controlled enzymolysis, and performing thorough enzymolysis to obtain liquid enzymolysis liquid; carrying out solid-liquid separation on the liquid enzymolysis liquid to prepare solid wet residues and a dilute composite nutrient solution;
reserving a part of the dilute composite nutrient solution as fermentation substrate sugar, and performing reduced pressure evaporation and concentration on the rest of the dilute composite nutrient solution to obtain a concentrated composite nutrient solution;
wherein the mass ratio of the corn to the soybean meal is 1: 0.01-0.2; adding water to adjust the concentration of the dry matter to 10-40%; the addition amount of the high-temperature amylase is 0.1-1 kg per ton of corn and soybean meal mixture, the pH of the slurry is controlled to be 5.5-6.5, and the enzymolysis time is 0.5-2 hours;
the addition amount of the saccharifying enzyme is 0.1-0.5 kg of saccharifying enzyme added to each ton of corn and soybean meal mixture, and the enzymolysis time of the slurry is controlled to be 0.5-20 h, wherein the pH value of the slurry is 4.5-5.5;
the addition amount of the protease is 0.1-1 kg per ton of the corn and soybean meal mixture, the pH of the slurry is controlled to be 4.5-5.5, and the enzymolysis time is 0.5-20 hours;
the concentration of the concentrated dry matter of the dilute composite nutrient solution is 40-65%;
secondly, preparing high-density yeast liquid:
in a full-automatic fermentation tank, diluting a part of dilute composite nutrient solution, sterilizing at high temperature, cooling, and inoculating a yeast seed solution, wherein the yeast seed solution is saccharomyces cerevisiae, introducing sufficient sterile air for culturing, controlling dissolved oxygen to be not less than 20%, when the dissolved oxygen suddenly rises to be more than 60%, feeding sterile concentrated composite nutrient solution and diammonium hydrogen phosphate solution by a peristaltic pump to control sufficient dissolved oxygen, the feeding mass ratio of the concentrated composite nutrient solution to diammonium hydrogen phosphate is 1:0.004, controlling the pH to be 4.0-4.5, when the yeast density is maximum, stopping feeding, and stopping ventilation and stirring when the dissolved oxygen rises to 70%;
keeping the fermentation tank at positive pressure, standing the yeast fermentation liquor, and discharging the precipitated yeast from the tank bottom after complete sedimentation to obtain high-density yeast liquid; continuously adding the residual fermentation liquor in the fermentation tank in a flowing manner to control sufficient dissolved oxygen in a manner of respectively adding the concentrated composite nutrient solution and the diammonium hydrogen phosphate solution in a flowing manner, stopping adding in a flowing manner after the yeast is enriched and cultured again to the maximum density, and stopping ventilation and stirring when the dissolved oxygen is raised back to 70%; standing, discharging high-density yeast liquid, continuously and circularly producing for multiple times, and collecting all the high-density yeast liquid;
wherein, the method for controlling sufficient dissolved oxygen comprises the following steps: controlling the dissolved oxygen to be 20-40%, and increasing the stirring speed to 150 r/min;
wherein, the dilute composite nutrient solution is diluted to a dry matter concentration of 0.01-5%; the mass ratio of the diammonium hydrogen phosphate to the water in the diammonium hydrogen phosphate solution is 10-50: 50-90; the feeding mass ratio of the concentrated compound nutrient solution to the diammonium hydrogen phosphate solution is 1: 0.001-0.1; the inoculation mass ratio of the yeast seed liquid is 0.2-20%;
when preparing the yeast fermentation liquor: fermentation temperature: 20-40 ℃; fermentation pH: 4.0 to 6.0; the stirring speed of aerobic fermentation is 20-1000 rMin; the ventilation ratio is 1: 0.5-3 v/v; the concentration of dissolved oxygen is 10-100%; and (3) fermentation period: 12 to 72 hours, the number of the bacteria at the end point of the yeast fermentation is 0.5 to 1.20 multiplied by 109 cfu/ml；
The yeast culture end marker is: the content of fermentable sugar in the fermentation liquor is 0-0.1%, and the content of dissolved oxygen is 60-100%; standing and settling for 1-20 h.
2. The method according to claim 1, wherein the method comprises the steps of: the yeast seed liquid is Saccharomyces cerevisiae CICC 32280.
3. The method for preparing a high-density yeast fermentation broth according to claim 1, wherein the reaction conditions in step (i) are as follows:
the mass ratio of the corn to the soybean meal is 1: 0.05-0.1; adding water to adjust the concentration of dry matters to be 25-30%; the addition amount of the high-temperature amylase is 0.3-0.5 kg per ton of the corn and soybean meal mixture, the pH of the slurry is controlled to be 5.8-6.2, and the enzymolysis time is 0.5-1 h;
the addition amount of the saccharifying enzyme is that 0.4-0.5 kg of saccharifying enzyme is added to each ton of corn and soybean meal mixture to control the pH of the slurry to be 4.8-5.2, and the enzymolysis time is 5-10 hours;
the addition amount of the protease is that 0.5-0.6 kg of protease is added into each ton of corn and soybean meal mixture to control the pH of the slurry to be 4.5-5.5, and the enzymolysis time is 15-20 h;
the concentration of the concentrated dry matter of the dilute composite nutrient solution is 40-45%.
4. The method for preparing high-density yeast fermentation broth according to claim 1, wherein the reaction conditions in step (ii) are as follows:
diluting the dilute composite nutrient solution to a dry matter concentration of 0.5-2%; the mass ratio of the diammonium hydrogen phosphate to the water in the diammonium hydrogen phosphate solution is 30-40: 60-70 parts of; the feeding mass ratio of the concentrated compound nutrient solution to the diammonium hydrogen phosphate solution is 1: 0.005-0.01; the inoculation mass ratio of the yeast seed liquid is 0.2-20%;
when preparing the yeast fermentation liquor: fermentation temperature: 30-35 ℃; fermentation pH: 4.0 to 4.5; aerobic fermentation stirring speed100 to 150 r/min; the ventilation ratio is 1: 0.5-2 v/v; the concentration of dissolved oxygen is 20-40%; and (3) fermentation period: 48-56 h, the number of the bacteria at the end point of the yeast fermentation is 1.15-1.20 multiplied by 109 cfu/ml；
The yeast culture end marker is: the content of fermentable sugar in the fermentation liquor is 0-0.05%, and the content of dissolved oxygen is 60-80%; standing and settling for 4-6 h.
5. The method for preparing the high-density yeast fermentation liquor according to claim 1, wherein in the step (i), the mass percentage of a part of the dilute composite nutrient solution in the total dilute composite nutrient solution is 1-2%.
6. The method for preparing high-density yeast fermentation liquor according to claim 1, wherein the continuous cycle production times in the step (II) are 8-10.
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Citations (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001085794A1 (en) * | 2000-05-10 | 2001-11-15 | Mogam Biotechnology Research Institute | Method of preparation for recombinant guamerin and pharmaceutical compositions containing the recombinant guamerin for wound healing |
| CN101429537A (en) * | 2008-12-14 | 2009-05-13 | 甘肃正生生物科技有限公司 | Method for high-density fermentation production of reductive glutathione with saccharomyces cerevisiae |
| CN101570735A (en) * | 2009-06-12 | 2009-11-04 | 河南工业大学 | Method for producing flour yeast by using corn flour as raw material |
| CN101979649A (en) * | 2010-10-13 | 2011-02-23 | 山东华辰生物科技有限公司 | Method for controlling Pichia pastoris to ferment to produce antibacterial peptide by utilizing dissolved oxygen parameters |
| CN102311927A (en) * | 2011-08-19 | 2012-01-11 | 华南理工大学 | Medium and method for high density fermentation of saccharomyces cerevisiae |
| CN104403953A (en) * | 2014-11-07 | 2015-03-11 | 湖南农业大学 | High-density fermentation culture medium formula for saccharomyces cerevisiae for feed and applications thereof |
| CN105018361A (en) * | 2015-07-13 | 2015-11-04 | 江南大学 | Method for high-density fermentation cultivation of brewer's yeast |
| CN108220175A (en) * | 2016-12-12 | 2018-06-29 | 安琪酵母股份有限公司 | Saccharomyces cerevisiae high-density cultivation method and its pH regulation and control methods |
| CN109321477A (en) * | 2018-11-20 | 2019-02-12 | 江南大学 | The method of one primary yeast High Density Cultivation |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2010053636A1 (en) * | 2008-11-06 | 2010-05-14 | Tate & Lyle Technology Ltd | Enhanced production and purification of a natural high intensity sweetener |
-
2019
- 2019-11-22 CN CN201911155830.0A patent/CN110684680B/en active Active
Patent Citations (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001085794A1 (en) * | 2000-05-10 | 2001-11-15 | Mogam Biotechnology Research Institute | Method of preparation for recombinant guamerin and pharmaceutical compositions containing the recombinant guamerin for wound healing |
| CN101429537A (en) * | 2008-12-14 | 2009-05-13 | 甘肃正生生物科技有限公司 | Method for high-density fermentation production of reductive glutathione with saccharomyces cerevisiae |
| CN101570735A (en) * | 2009-06-12 | 2009-11-04 | 河南工业大学 | Method for producing flour yeast by using corn flour as raw material |
| CN101979649A (en) * | 2010-10-13 | 2011-02-23 | 山东华辰生物科技有限公司 | Method for controlling Pichia pastoris to ferment to produce antibacterial peptide by utilizing dissolved oxygen parameters |
| CN102311927A (en) * | 2011-08-19 | 2012-01-11 | 华南理工大学 | Medium and method for high density fermentation of saccharomyces cerevisiae |
| CN104403953A (en) * | 2014-11-07 | 2015-03-11 | 湖南农业大学 | High-density fermentation culture medium formula for saccharomyces cerevisiae for feed and applications thereof |
| CN105018361A (en) * | 2015-07-13 | 2015-11-04 | 江南大学 | Method for high-density fermentation cultivation of brewer's yeast |
| CN108220175A (en) * | 2016-12-12 | 2018-06-29 | 安琪酵母股份有限公司 | Saccharomyces cerevisiae high-density cultivation method and its pH regulation and control methods |
| CN109321477A (en) * | 2018-11-20 | 2019-02-12 | 江南大学 | The method of one primary yeast High Density Cultivation |
Non-Patent Citations (3)
| Title |
|---|
| High-cell-density fermentation for ergosterol production by Saccharomyces cerevisiae;FeiShang et al.;《JOURNAL OF BIOSCIENCE AND BIOENGINEERING》;20061231;第101卷(第1期);第38-41页 * |
| The impact of oxygen on the final alcohol content of wine fermented by a mixed starter culture;Pilar Morales et al.;《Appl Microbiol Biotechnol》;20150113;第99卷;第3993-4003页 * |
| 不同补料方式对酿酒酵母高密度发酵的影响;荣博涵 等;《中国酿造》;20151231;第34卷(第2期);第72-75页 * |
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