A method for the use of a beer yeast in the control of a fermentation process

Abstract

The present application relates to the beer brewing technical field, especially to a kind of beer yeast in fermentation process control application method, comprising: beer yeast strain is successively enlarged culture, monitors live bacteria number and mortality calculates yeast activity index, determines expansion culture eligibility, when unqualified, based on expansion culture level determines processing strategy;Expansion culture qualified yeast liquid is inoculated into wort after low-temperature starvation treatment, monitors dissolved oxygen consumption rate and germination rate calculates inoculation adaptability index, based on its change rate determines inoculation eligibility, when unqualified, based on liquid surface foam half-life and foam characteristic morphology determines unqualified type and adjustment strategy;Inoculation is qualified, and main fermentation is carried out, and after main fermentation, diacetyl reduction is carried out;Yeast mud is recovered, monitors cell viability and mortality calculates yeast reuse index, with reuse threshold value comparison determines whether the current batch of yeast can be used for the next batch fermentation and whether to adjust inoculation amount.The present application improves the stability of beer fermentation process.

Description

Technical Field

[0001] This invention relates to the field of beer brewing technology, and in particular to a method for applying beer yeast in fermentation process control. Background Technology

[0002] Beer fermentation is a biological process in which yeast converts fermentable sugars in wort into ethanol, carbon dioxide, and flavor compounds. The physiological state and metabolic activity of yeast directly determine fermentation efficiency, batch stability, and beer flavor quality. Currently, existing technologies still present many yeast-related challenges in the application and control of beer yeast.

[0003] Factors in malt can cause yeast to flocculate prematurely, leading to significant sedimentation before sugar utilization. This results in slow fermentation, higher sugar content, and incomplete fermentation. Both excessive and insufficient flocculation can affect fermentation integrity and flavor quality. In industrial production, yeast is often reused for multiple generations. Continuous reuse leads to a gradual deterioration of cell physiological state, manifested as prolonged stagnation, fluctuating fermentation degree, and decreased activity. In severe cases, it can lead to degeneration, autolysis, and increased risk of contamination. Improper recycling and processing can further exacerbate batch-to-batch instability.

[0004] Regarding metabolite control, excessive diacetyl produces a stale, rancid flavor, which is particularly pronounced when fermentation temperature is increased or the cycle is shortened; excessive higher alcohols introduce off-flavors. Existing isothermal or simple staged fermentation processes struggle to balance yeast reproduction and flavor metabolism, often leading to flavor imbalances. For active dry yeast, the survival rate varies significantly among different strains after drying. During the drying process, yeast faces multiple forms of damage, including oxidation and osmotic pressure, resulting in cell structure damage. Due to a lack of research on the stress resistance mechanisms of characteristic flavor strains, the performance of dried yeast prepared from them differs considerably from that of commercially available products.

[0005] In summary, this paper proposes an application method that can effectively regulate the entire process of brewer's yeast from expansion to multi-generation reuse, in order to solve problems such as yeast flocculation, vitality decline, metabolic control, drying damage, activation lag, and lack of intergenerational feedback in existing technologies. This method aims to shorten the fermentation cycle, stabilize beer flavor and quality, and adapt to multi-generation industrial production, which is a technical problem that urgently needs to be solved by those skilled in the art. Summary of the Invention

[0006] Therefore, this invention provides a method for applying beer yeast in fermentation process control to overcome the problems of poor batch stability, poor flavor consistency, and difficulty in accurately controlling the fermentation cycle caused by large fluctuations in yeast physiological activity, rough control, and lack of intergenerational feedback regulation in the prior art.

[0007] To achieve the above objectives, the present invention provides a method for applying brewer's yeast in fermentation process control, comprising: Step S1: The brewer's yeast strain is cultured step by step under preset conditions to obtain expanded yeast liquid. During the expansion process, samples are taken step by step to monitor the number of viable yeast cells and the mortality rate to calculate the yeast viability index. The pass rate of yeast expansion is determined based on the comparison between the yeast viability index and the preset viability threshold. If the pass rate is not met, a treatment strategy is determined based on the current expansion level. Step S2: The qualified yeast liquid is subjected to low-temperature starvation treatment. The starved yeast is inoculated into the wort. The dissolved oxygen consumption rate and yeast germination rate are monitored at preset time intervals to calculate the inoculation adaptability index. The inoculation is qualified based on the rate of change of the inoculation adaptability index. If it is unqualified, the unqualified type is determined based on the half-life of the liquid surface foam and the characteristic morphology of the foam, and the adjustment strategy is determined according to the unqualified type. Step S3: After successful inoculation, carry out primary fermentation within the preset fermentation temperature range, monitor the appearance sugar content until the appearance sugar content drops to the preset primary fermentation endpoint threshold, and end the primary fermentation. Step S4: After the main fermentation is completed, the temperature of the fermentation broth is raised to the preset reduction temperature range at a preset heating rate to carry out diacetyl reduction. The reduction ends when the diacetyl content is less than the preset qualified threshold. Step S5: Cool the reduced fermentation broth to allow it to settle, recover the yeast sludge settled at the bottom of the tank, monitor the cell viability and mortality of the recovered yeast to calculate the yeast reuse index, and determine whether the current batch of yeast can be used for the next batch of fermentation based on the comparison result between the yeast reuse index and the preset reuse threshold, and whether the inoculation amount for the next batch should be adjusted if it is available.

[0008] Furthermore, the preset conditions in step S1 are as follows: the expansion temperature is 20-24℃, the expansion medium is wort with a concentration of 8-12°P, the dissolved oxygen content during the expansion process is controlled at 6-10 mg / L, the inoculation ratio for step-by-step expansion is 1:5-1:20, and the expansion time for each step is 24-48 hours.

[0009] Furthermore, in step S1, based on the condition that the yeast activity index is less than a preset activity threshold, the yeast expansion is determined to be unqualified, and a processing strategy is determined based on the current expansion level. The propagation levels are, in order: Erlenmeyer flask level, Karl Casner level, seed tank level, and propagation tank level.

[0010] Furthermore, if the current expansion level is Erlenmeyer flask level, the treatment strategy is to discard the current batch of yeast and start expansion again from the preserved strain. If the current expansion level is the Karl flask level, the handling strategy is to discard the current batch of yeast and start expansion again from the Erlenmeyer flask level. If the current expansion level is seed tank level, the treatment strategy is to extend the expansion time by 2-6 hours and supplement aeration, and re-monitor the yeast activity index. If it is still unqualified, it should be discarded. If the current expansion level is expansion tank level, the treatment strategy is as follows: extend the expansion time by 2-6 hours and supplement aeration, re-monitor the yeast activity index, and if it is still unqualified, discard it and trace back to the quality of the previous level seed tank level yeast.

[0011] Furthermore, in step S2, the low-temperature starvation treatment is a carbon source starvation treatment, the temperature of the low-temperature starvation treatment is 2-10℃, and the treatment time is 6-48 hours; During the treatment, the yeast liquid is intermittently stirred at a speed of 30-100 rpm, once every 2-4 hours, for 5-15 minutes each time.

[0012] Furthermore, in step S2, the process of determining whether the vaccination is qualified is as follows: The first vaccination adaptability index and the second vaccination adaptability index were calculated at the first and second preset time points after vaccination, respectively, and the rate of change between the two was calculated. When the rate of change is greater than or equal to a preset rate of change threshold, and the second vaccination adaptability index is greater than or equal to a preset adaptability threshold, the vaccination is deemed qualified. When the rate of change is less than a preset rate of change threshold, or the second inoculation adaptability index is less than a preset adaptability threshold, the inoculation is deemed unqualified, and the unqualified type is determined based on the half-life of the liquid surface foam and the characteristic morphology of the foam. The inoculation adaptability index is determined by the dissolved oxygen consumption rate and the yeast germination rate.

[0013] Furthermore, the specific process of determining the non-compliance type based on the half-life and characteristic morphology of the liquid surface foam, and then determining the adjustment strategy according to the non-compliance type, is as follows: Measure the foam half-life on the liquid surface within a preset time period after inoculation and observe the characteristic morphology of the foam; The foam half-life is compared with a preset half-life threshold, and the foam morphology is compared with a preset morphology feature. If the foam half-life is less than the first preset half-life threshold, it is judged as an unqualified type of excessively fast settling, and the adjustment strategy is to start bottom pulse ventilation or forced bottom ventilation. If the foam half-life is greater than the second preset half-life threshold and the foam characteristic morphology is viscous and stringy, it is judged as an abnormal viscosity type and is unqualified. The adjustment strategy is to switch to pulse stirring mode. If the foam half-life is greater than the second preset half-life threshold and the foam characteristic morphology is non-sticky and stringy, it is judged as unqualified due to foam runaway. The adjustment strategy is to reduce the tank pressure and start mechanical defoaming.

[0014] Furthermore, the specific process for determining whether the current batch of yeast can be used for the next batch of fermentation is as follows: Monitor the cell viability and mortality rate of recycled yeast, and calculate the yeast reuse index based on the ratio of cell viability to mortality rate; The yeast reuse index is compared with the preset reuse threshold. If the yeast reuse index is greater than or equal to the preset reuse threshold, it is determined that the current batch of yeast can be used for the next batch of fermentation, and it is determined whether the inoculation amount for the next batch should be adjusted. If the yeast reuse index is less than the preset reuse threshold, then the current batch of yeast is determined not to be reused.

[0015] Furthermore, the inoculation amount for the next batch is adjusted based on the condition that the difference in the yeast reuse index is greater than a preset difference, wherein the difference in the yeast reuse index is the difference between the yeast reuse index and the preset reuse threshold.

[0016] Furthermore, the difference between the next batch inoculation amount and the yeast reuse index was positively correlated.

[0017] Compared with the prior art, the beneficial effects of the present invention are as follows: 1. This invention monitors yeast viable cell count and mortality rate at each stage of the expansion process, calculates the yeast viability index, and adopts differentiated treatment strategies based on the current expansion level. For yeast failing at earlier stages, it is discarded and the expansion is restarted. For yeast failing at later stages, the expansion time is first extended and aeration is supplemented; if the yeast still fails, the previous stage's quality is traced back. This tiered treatment strategy prevents substandard yeast from entering subsequent fermentation processes and reduces the time and material waste caused by frequently restarting expansion due to slight fluctuations in early stages.

[0018] 2. This invention introduces a low-temperature starvation treatment during the inoculation stage and determines inoculation compliance by measuring the rate of change in the inoculation adaptability index. Unlike traditional methods that rely on chemical testing or simple feeding, this invention further distinguishes between three types of non-compliance based on foam half-life and foam morphology: excessively rapid settling, abnormal viscosity, and uncontrolled foaming. For each type, purely physical adjustment strategies are employed, including bottom pulse aeration or forced bottom aeration, pulse stirring mode, reducing tank pressure, and initiating mechanical defoaming. This approach reduces reliance on chemical defoamers or additional yeast, while enabling rapid identification and targeted intervention of the causes of non-compliance.

[0019] 3. This invention does not involve intermediate intervention during the primary fermentation stage. Instead, it controls the end time of primary fermentation solely by monitoring the apparent sugar content until it drops to a preset endpoint threshold. This simplified control logic avoids uncontrollable factors introduced by frequent adjustments to fermentation temperature, nutrient supplementation, or yeast addition, reducing operational complexity and providing a relatively stable initial state of the fermentation broth for the subsequent diacetyl reduction stage.

[0020] 4. This invention recovers yeast sludge after diacetyl reduction, calculates a yeast reuse index based on the ratio of cell viability to cell death, and compares this index with a preset reuse threshold to determine whether the current batch of yeast can be used for the next batch of fermentation. If usable, the adjustment range of the inoculum amount is further determined based on the difference between the yeast reuse index and the reuse threshold, and the adjustment amount is positively correlated with the difference. This closed-loop feedback mechanism achieves quantitative transfer and compensation of yeast quality across batches, improving the consistency and predictability of fermentation processes across multiple batches. Attached Figure Description

[0021] Figure 1 This is a flowchart illustrating the application method of brewer's yeast in fermentation process control according to an embodiment of the present invention; Figure 2 This is a flowchart illustrating the process of determining whether an inoculation is successful according to an embodiment of the present invention. Figure 3 This is a flowchart illustrating how to determine whether the current batch of yeast can be used for the next batch of fermentation, as described in an embodiment of the present invention. Detailed Implementation

[0022] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments.

[0023] Those skilled in the art will understand that the method for determining the above-mentioned parameters for a single item in this invention can be as follows: selecting the value with the highest proportion based on the data distribution as the preset standard parameter; using weighted summation to use the obtained value as the preset standard parameter; substituting each historical data into a specific formula and using the value obtained by the formula as the preset standard parameter; or other selection methods, as long as the present invention can clearly define different specific situations in the single item determination process through the obtained values.

[0024] Please see Figure 1 As shown, the method for applying brewer's yeast in fermentation process control according to an embodiment of the present invention includes: Step S1: The brewer's yeast strain is cultured step by step under preset conditions to obtain expanded yeast liquid. During the expansion process, samples are taken step by step to monitor the number of viable yeast cells and the mortality rate to calculate the yeast viability index. The pass rate of yeast expansion is determined based on the comparison between the yeast viability index and the preset viability threshold. If the pass rate is not met, a treatment strategy is determined based on the current expansion level. Step S2: The qualified yeast liquid is subjected to low-temperature starvation treatment. The starved yeast is inoculated into the wort. The dissolved oxygen consumption rate and yeast germination rate are monitored at preset time intervals to calculate the inoculation adaptability index. The inoculation is qualified based on the rate of change of the inoculation adaptability index. If it is unqualified, the unqualified type is determined based on the half-life of the liquid surface foam and the characteristic morphology of the foam, and the adjustment strategy is determined according to the unqualified type. Step S3: After successful inoculation, carry out primary fermentation within the preset fermentation temperature range, monitor the appearance sugar content until the appearance sugar content drops to the preset primary fermentation endpoint threshold, and end the primary fermentation. Step S4: After the main fermentation is completed, the temperature of the fermentation broth is raised to the preset reduction temperature range at a preset heating rate to carry out diacetyl reduction. The reduction ends when the diacetyl content is less than the preset qualified threshold. Step S5: Cool the reduced fermentation broth to allow it to settle, recover the yeast sludge settled at the bottom of the tank, monitor the cell viability and mortality of the recovered yeast to calculate the yeast reuse index, and determine whether the current batch of yeast can be used for the next batch of fermentation based on the comparison result between the yeast reuse index and the preset reuse threshold, and whether the inoculation amount for the next batch should be adjusted if it is available.

[0025] Specifically, the preset conditions in step S1 are: the expansion temperature is 20-24℃, the expansion medium is wort with 8-12°P, the dissolved oxygen content during the expansion process is controlled at 6-10mg / L, the inoculation ratio for step-by-step expansion is 1:5-1:20, and the expansion time for each step is 24-48 hours.

[0026] In this embodiment of the invention, the yeast viable cell count and mortality rate are monitored using methylene blue staining combined with a hemocytometer. The specific procedure is as follows: a yeast sample is mixed with 0.1% methylene blue staining solution in an equal ratio and allowed to stand at room temperature for 3-5 minutes, allowing the live cells to become colorless and the dead cells to become blue. The stained sample is then added to a hemocytometer, and the total number of cells and the number of blue dead cells within a specified number of squares are counted under an optical microscope. The yeast viability index is calculated based on the ratio of viable cells to total cells.

[0027] Specifically, in step S1, based on the condition that the yeast activity index is less than a preset activity threshold, the yeast expansion is determined to be unqualified, and a processing strategy is determined based on the current expansion level. The expansion levels are, in order: Erlenmeyer flask level, Karl flask level, seed tank level, and expansion tank level; The yeast viability index = viable cell count / (viable cell count + dead cell count). In this embodiment of the invention, the preset vigor threshold for the Erlenmeyer flask and Karl argument flask levels is 0.9, and the preset vigor threshold for the seed tank and propagation tank levels is 0.85.

[0028] Specifically, if the current expansion level is Erlenmeyer flask level, the treatment strategy is to discard the current batch of yeast and start expansion again from the preserved strain. If the current expansion level is the Karl flask level, the handling strategy is to discard the current batch of yeast and start expansion again from the Erlenmeyer flask level. If the current expansion level is seed tank level, the treatment strategy is to extend the expansion time by 2-6 hours and supplement aeration, and re-monitor the yeast activity index. If it is still unqualified, it should be discarded. If the current expansion level is expansion tank level, the treatment strategy is as follows: extend the expansion time by 2-6 hours and supplement aeration, re-monitor the yeast activity index, and if it is still unqualified, discard it and trace back to the quality of the previous level seed tank level yeast.

[0029] Specifically, the process of tracing the quality of the seed tank-grade yeast at the next higher level is as follows: retrieve the monitoring records of the seed tank-grade yeast used to inoculate the current expansion tank, including its yeast viability index, viable cell count, mortality rate, and culture condition parameters, and compare them with preset standards to determine whether the non-compliance originates from quality fluctuations in the previous seed tank; if an abnormality is found in the quality of the seed tank-grade yeast at the next higher level, discard that seed tank-grade yeast and start expansion again from the next higher level.

[0030] Understandably, for early expansion stages such as Erlenmeyer flasks and Carbohydrate tanks, the expansion volume is small and the operating cost is low. However, the yeast quality at this stage directly affects the expansion results of all subsequent stages. Once an early stage fails to meet standards, attempting remedial measures not only fails to guarantee successful expansion but may also amplify the quality defects to subsequent stages, resulting in greater waste of materials and time. Therefore, discarding the yeast and starting expansion from the previous stage or from a preserved strain is the most economical and reliable approach. For later expansion stages such as seed tanks and expansion tanks, the expansion volume is large and the cost is high. Since the yeast at this stage already has a certain biomass, a simple decrease in activity can often be recovered by extending the expansion time and supplementing aeration. Therefore, prioritizing remedial measures rather than direct discarding can reduce production costs. Furthermore, failures at the expansion tank stage must be traced back to the quality of the yeast in the previous seed tank stage. This is because the yeast in the expansion tank stage originates directly from the seed tank stage. If the yeast in the expansion tank stage fails to meet standards and the seed tank stage itself has quality fluctuations, even if the expansion tank stage is re-expanded, using the same batch of seed tank yeast will still result in repeated failures. The problem can only be fundamentally solved by tracing back and discarding problematic seed tanks, and starting anew from a higher level of cultivation.

[0031] Specifically, in step S2, the low-temperature starvation treatment is a carbon source starvation treatment, the temperature of the low-temperature starvation treatment is 2-10℃, and the treatment time is 6-48 hours; During the treatment, the yeast liquid is intermittently stirred at a speed of 30-100 rpm, once every 2-4 hours, for 5-15 minutes each time.

[0032] In this embodiment of the invention, the dissolved oxygen consumption rate is continuously monitored using an online fluorescent dissolved oxygen sensor. After inoculation, the sensor probe is inserted into the fermenter, and the dissolved oxygen concentration change curve over time is recorded in real time. The decrease in dissolved oxygen per unit time is calculated in mg / L·h. Yeast budding rate is monitored using a combination of microscopy and hemocytometer. Yeast samples are placed on a glass slide, and multiple fields of view are randomly selected under a 400x optical microscope. The number of budding cells and the total number of cells are counted, with at least 500 cells counted for each sample. The budding rate is calculated as a percentage of the proportion of budding cells to the total number of cells.

[0033] Specifically, the inoculation adaptability index is determined by the dissolved oxygen consumption rate and the yeast germination rate. The inoculation adaptability index = (dissolved oxygen consumption rate / preset standard consumption rate) × (yeast germination rate / preset standard germination rate). In this embodiment of the invention, the preset standard consumption rate is 1.0 mg / L·h and the preset standard germination rate is 20%.

[0034] Please see Figure 2 As shown, in step S2, the process of determining whether the vaccination is qualified is as follows: The first vaccination adaptability index and the second vaccination adaptability index were calculated at the first and second preset time points after vaccination, respectively. The rate of change between the two was calculated as follows: rate of change = (second vaccination adaptability index - first vaccination adaptability index) / first vaccination adaptability index × 100%; When the rate of change is greater than or equal to a preset rate of change threshold, and the second vaccination adaptability index is greater than or equal to a preset adaptability threshold, the vaccination is deemed qualified. When the rate of change is less than a preset rate of change threshold, or the second inoculation adaptability index is less than a preset adaptability threshold, the inoculation is deemed unqualified, and the unqualified type is determined based on the half-life of the liquid surface foam and the characteristic morphology of the foam. In this embodiment of the invention, the first preset time point is 2 hours after vaccination, the second preset time point is 6 hours after vaccination, the preset change rate is 30%, and the preset adaptability threshold is 0.5.

[0035] Understandably, the inoculation adaptability index reflects the overall adaptability of yeast to the wort environment; a higher value indicates stronger respiratory metabolic and reproductive activity. The rate of change reflects the yeast's growth trend within the adaptation window. A rate of change reaching a preset threshold indicates that the yeast has achieved significant metabolic activation within a preset time interval, proving its successful adaptation to the wort environment. A second adaptability index reaching a preset threshold ensures that the yeast has reached a sufficient metabolic level after adaptation to support the subsequent main fermentation process. Both conditions must be met simultaneously for inoculation to be considered successful. If either condition is not met, it indicates that the yeast has problems such as slow adaptation or insufficient metabolic activity, requiring further classification and targeted adjustments using foam characteristics.

[0036] Specifically, the process of determining the non-compliance type based on the half-life and characteristic morphology of the liquid surface foam, and then determining the adjustment strategy based on the non-compliance type, is as follows: Measure the foam half-life on the liquid surface within a preset time period after inoculation and observe the characteristic morphology of the foam; The foam half-life is compared with a preset half-life threshold, and the foam morphology is compared with a preset morphology feature. If the foam half-life is less than the first preset half-life threshold, it is judged as an unqualified type of excessively fast settling, and the adjustment strategy is to start bottom pulse ventilation or forced bottom ventilation. If the foam half-life is greater than the second preset half-life threshold and the foam characteristic morphology is viscous and stringy, it is judged as an abnormal viscosity type and is unqualified. The adjustment strategy is to switch to pulse stirring mode. If the foam half-life is greater than the second preset half-life threshold and the foam characteristic morphology is non-sticky and stringy, it is judged as unqualified due to foam runaway. The adjustment strategy is to reduce the tank pressure and start mechanical defoaming.

[0037] In this embodiment of the invention, the method for measuring the foam half-life is as follows: Within a preset time period after inoculation, the stirring and aeration of the fermenter are stopped. The initial height of the foam layer on the liquid surface is measured using a ruler attached next to the sight glass. Simultaneously, a timer is started to observe the decrease in foam layer height over time. Timing is stopped when the foam layer height decreases to half of its initial height, and the elapsed time is recorded as the foam half-life, measured in seconds. A shorter foam half-life indicates less stable foam, while a longer foam half-life indicates more stable foam.

[0038] The method for observing the characteristic morphology of foam is as follows: while measuring the half-life of the foam, visually observe the appearance of the foam through the sight glass of the fermenter, or gently touch the foam layer with a glass rod and lift it upwards. If the foam is fine and moist, and can form continuous filaments without breaking after being lifted, it is judged as viscous and stringy; if the foam is coarse and dry, and there is no stringing phenomenon after being lifted or the filaments break quickly, it is judged as non-viscous and stringy.

[0039] Bottom pulse ventilation is an intermittent ventilation mode, introducing compressed air for 1 second every 30 seconds, repeated 10 times; forced bottom ventilation is a continuous ventilation mode, continuously ventilating for 15 minutes at 1.5-2.0 times the normal ventilation volume.

[0040] In this embodiment of the invention, the preset time period is preferably 10-20 minutes after vaccination, the first preset half-life threshold is 15 seconds, and the second preset half-life threshold is 90 seconds.

[0041] It is understandable that foam is a direct product of yeast metabolism, and its behavior is closely related to the physiological state of the yeast and the physicochemical properties of the wort. When yeast flocculates and settles prematurely, reduced gas production leads to the rapid disappearance of the foam layer and a significantly shortened foam half-life. Bottom pulse aeration or forced bottom aeration is needed to resuspend the settled yeast. When wort viscosity is abnormally high, bubbles become trapped in the viscous liquid and are difficult to break, resulting in viscous, stringy foam with a significantly prolonged half-life. Pulsed stirring is required to break up the viscous structure with high shear force. When yeast metabolism is excessive or the wort protein content is abnormal, large amounts of stable and persistent foam are produced, with a significantly prolonged half-life, but the foam is coarse, dry, and lacks viscous, stringy characteristics. In this case, reducing tank pressure and initiating mechanical defoaming can effectively control the problem. The foam characteristics of these three types of non-compliance are distinct and easily distinguishable. Diagnosis can be completed simply by observation through a sight glass and basic timing, without the need for chemical testing or complex instruments, enabling rapid localization and targeted intervention of the causes of inoculation failure.

[0042] In this embodiment of the invention, after successful inoculation, the fermentation temperature is controlled within a preset fermentation temperature range for primary fermentation. No intermediate intervention is performed during primary fermentation; only the apparent sugar content is monitored at preset time intervals until the apparent sugar content drops to a preset primary fermentation endpoint threshold, at which point the primary fermentation is terminated. The apparent sugar content is monitored every 12 hours.

[0043] The apparent sugar content was monitored using a handheld saccharimeter or an online densitometer. The procedure was aseptically performed by taking a sample from the sampling port of the fermenter, adding the sample drop onto the prism surface of the saccharimeter, closing the cover, and then reading the apparent sugar content in °P. The apparent sugar content reflects the remaining amount of fermentable sugars in the fermentation broth, and its rate of decrease indicates the fermentation activity of the yeast.

[0044] In this embodiment of the invention, the preset fermentation temperature range is 10-14℃, preferably 12℃. The preset primary fermentation endpoint threshold is 4.0°P. When the apparent sugar content decreases from the initial value (usually 10-14°P) to 4.0°P, it indicates that more than 90% of the fermentable sugars in the fermentation broth have been metabolized by the yeast, and the primary fermentation stage ends.

[0045] It is understandable that this invention does not involve intermediate intervention during the main fermentation stage because, after the expansion, grading, and quality control in step S1 and the inoculation foam diagnosis in step S2, the yeast inoculated into the wort is already in a qualified physiological state and can autonomously complete the main fermentation process under preset temperature conditions. This simplified control logic avoids uncontrollable factors introduced by frequent adjustments to fermentation temperature, supplementation of nutrients, or addition of yeast, reduces operational complexity, and provides a relatively stable initial state of the fermentation broth for the subsequent diacetyl reduction stage.

[0046] In this embodiment of the invention, after the primary fermentation is completed, the temperature of the fermentation broth is raised to a preset reduction temperature range at a preset heating rate for diacetyl reduction. During the reduction process, diacetyl content is sampled and monitored at preset time intervals. Reduction ends when the diacetyl content is less than a preset acceptable threshold. Diacetyl content is monitored every 6 hours.

[0047] The diacetyl content was monitored using gas chromatography or spectrophotometry. The specific procedure was as follows: a fermentation broth sample was distilled, then o-phenylenediamine was added as a colorimetric reagent, and the absorbance was measured at 335 nm. The diacetyl content was calculated by comparing it with a standard curve, and the unit was mg / L. Diacetyl is an important flavor compound in beer; when its content exceeds a certain threshold, it produces an unpleasant stale taste and needs to be metabolized to below the threshold through a reduction phase.

[0048] In this embodiment of the invention, the preset heating rate is 0.5-1.0℃ / h, preferably 0.5℃ / h. The preset reduction temperature range is 12-16℃, preferably 14℃. The preset acceptable threshold is 0.10mg / L. When the diacetyl content is below 0.10mg / L, it indicates that the reduction stage is complete, and subsequent cooling and sedimentation treatment can be carried out.

[0049] Understandably, after the primary fermentation, the fermentation broth still contains a certain amount of diacetyl precursor (α-acetolactic acid), which needs to be reduced by yeast metabolism to acetoin and 2,3-butanediol at a relatively high temperature. A slow heating rate of 0.5℃ / h avoids thermal shock to the yeast from sudden temperature changes, ensuring the stability of the reduction process. Maintaining the reduction temperature at around 14℃ maintains the yeast's metabolic activity without causing autolysis or the production of undesirable byproducts due to excessively high temperatures.

[0050] Specifically, after the diacetyl reduction is completed, the temperature of the fermentation broth is reduced to 0-4℃ at a cooling rate of 0.5-1.0℃ / h, and kept at this temperature for 24-48 hours to allow the yeast sludge to settle fully to the bottom of the tank.

[0051] Please see Figure 3 As shown, the specific process for determining whether the current batch of yeast can be used for the next batch of fermentation is as follows: Monitor the cell viability and mortality rate of recycled yeast, and calculate the yeast reuse index based on the ratio of cell viability to mortality rate. Yeast reuse index = cell viability / (cell viability + mortality rate). The yeast reuse index is compared with the preset reuse threshold. If the yeast reuse index is greater than or equal to the preset reuse threshold, it is determined that the current batch of yeast can be used for the next batch of fermentation, and it is determined whether the inoculation amount for the next batch should be adjusted. If the yeast reuse index is less than the preset reuse threshold, then the current batch of yeast is determined not to be reused. In this embodiment of the invention, the preset reuse threshold value is 0.85.

[0052] In this embodiment of the invention, cell viability and mortality were monitored using methylene blue staining combined with a hemocytometer. The specific procedure was as follows: a sample of recovered yeast sludge was mixed with 0.1% methylene blue staining solution in an equal ratio and allowed to stand at room temperature for 3-5 minutes, allowing the dye to become colorless on live cells and stain the dead cells blue. The stained sample was then added to a hemocytometer, and the total number of cells and the number of blue dead cells within a specified number of squares were counted under an optical microscope. Cell viability was calculated as the proportion of live cells to the total number of cells, and mortality was calculated as the proportion of dead cells to the total number of cells.

[0053] Specifically, the inoculation amount for the next batch is adjusted based on the condition that the difference in the yeast reuse index is greater than a preset difference. The difference in the yeast reuse index is the difference between the yeast reuse index and the preset reuse threshold. In this embodiment of the invention, the preset difference is 0.05.

[0054] Specifically, the next batch inoculation amount is positively correlated with the difference in yeast reuse index. If the difference in yeast reuse index is less than the first preset difference threshold, the next batch inoculation amount is increased to the corresponding value using the first inoculation adjustment coefficient. If the yeast reuse index difference is greater than or equal to the first preset difference threshold and less than the second preset difference threshold, then the second inoculation adjustment coefficient is used to increase the inoculation amount of the next batch to the corresponding value. If the difference in yeast reuse index is greater than or equal to the second preset difference threshold, the third inoculation adjustment coefficient will be used to increase the inoculation amount of the next batch to the corresponding value. In this embodiment of the invention, the preset reuse threshold is 0.85, the first preset difference threshold is 0.05, the second preset difference threshold is 0.10, the first vaccination adjustment coefficient is 1.05, the second vaccination adjustment coefficient is 1.10, and the third vaccination adjustment coefficient is 1.20.

[0055] Example 1 In this embodiment, the brewer's yeast fermentation process is controlled according to the following steps: Expansion stage: The brewer's yeast strain was inoculated from the preservation slant onto Erlenmeyer flask-grade medium at 22℃. The medium was 10°P wort with dissolved oxygen content controlled at 8 mg / L. Samples were taken for monitoring after 36 hours of incubation; the viable count was 2.5 × 10⁻⁶. 8 With a cell / mL count and a mortality rate of 2%, the yeast viability index was calculated to be 0.98. The preset viability threshold for the Erlenmeyer flask stage was 0.9, indicating successful expansion. Propagation was then performed step-by-step to the Karl von Carbide tank stage, seed tank stage, and expansion tank stage. At each stage, the viability index was greater than the corresponding threshold, ultimately yielding the expanded yeast culture.

[0056] Inoculation stage: The successfully propagated yeast culture was subjected to low-temperature starvation treatment at 4°C for 24 hours, with stirring every 3 hours for 10 minutes each time at a stirring speed of 60 rpm. The starved yeast was then inoculated into the wort at an inoculation rate of 1.0 × 10⁻⁶. 7 Species / mL. Dissolved oxygen consumption was monitored at 2 and 6 hours post-inoculation: at 2 hours, the dissolved oxygen consumption rate was 0.3 mg / L·h, with a germination rate of 12%; at 6 hours, the dissolved oxygen consumption rate was 0.4 mg / L·h, with a germination rate of 15%. The inoculation adaptability index was calculated to be 0.18 at 2 hours and 0.30 at 6 hours, with a change rate of 66.7%. The preset change rate threshold was 30%, and the preset adaptability threshold was 0.5. Since the second adaptability index of 0.30 was less than 0.5, the inoculation was deemed unqualified. The foam half-life was measured 10-20 minutes post-inoculation; a value of 12 seconds (less than 15 seconds) indicated excessively rapid settling and was deemed unqualified. Forced bottom ventilation was then implemented, with continuous ventilation at 1.8 times the normal ventilation rate for 15 minutes. After ventilation, the foam half-life returned to 25 seconds.

[0057] Primary fermentation stage: Primary fermentation was carried out at 12°C, with an initial apparent sugar content of 12°P, monitored every 12 hours. The fermentation process was relatively stable, and on day 6, the apparent sugar content decreased to 4.0°P, reaching the endpoint threshold, thus ending the primary fermentation. The primary fermentation cycle was 6 days.

[0058] Diacetyl reduction stage: Reduction was carried out by increasing the temperature to 14℃ at a rate of 0.5℃ / h, with diacetyl content monitored every 6 hours. On the 4th day, the diacetyl content decreased to 0.10 mg / L, reaching the acceptable threshold, and the reduction was terminated. The reduction cycle was 4 days.

[0059] Yeast reuse stage: After cooling and settling, the yeast sludge was recovered. Cell viability was monitored at 88%, and mortality was 10%. The yeast reuse index was calculated to be 0.88. The preset reuse threshold was 0.85. If the reuse index difference of 0.03 was less than the preset difference threshold of 0.05, it was determined that this batch of yeast could be used for the next batch of fermentation, but the inoculum amount was not adjusted and inoculated according to the standard inoculum amount.

[0060] In this embodiment, the main fermentation cycle was 6 days, the reduction cycle was 4 days, and the total fermentation cycle was 10 days. The recovered yeast activity was 88%, which was close to usable. The fermentation process was generally satisfactory, but the cycle was long and the yeast activity decline was quite significant.

[0061] Example 2 This embodiment is based on Embodiment 1, but the inoculation amount has been optimized and adjusted according to the difference in the reuse index. The other conditions are the same as in Embodiment 1.

[0062] Expansion stage: Same as in Example 1, obtain qualified yeast liquid for expansion.

[0063] Inoculation stage: Same as in Example 1, if the sedimentation is too fast, it is deemed unqualified and forced bottom ventilation is performed.

[0064] Primary fermentation stage: Primary fermentation was carried out at 12°C, with an initial apparent sugar content of 12°P, monitored every 12 hours. The fermentation process accelerated significantly, and by day 5, the apparent sugar content had decreased to 4.0°P, reaching the endpoint threshold, thus ending the primary fermentation. The primary fermentation cycle was 5 days, 1 day shorter than in Example 1.

[0065] Diacetyl reduction stage: Reduction was carried out by increasing the temperature to 14°C at a rate of 0.5°C / h, with diacetyl content monitored every 6 hours. On the 3rd day, the diacetyl content dropped to 0.08 mg / L, below the acceptable threshold of 0.10 mg / L, at which point the reduction was terminated. The reduction cycle was 3 days, 1 day shorter than in Example 1.

[0066] Yeast reuse stage: After cooling and settling, the yeast sludge was recovered. Cell viability was monitored at 94%, mortality at 5%, and the yeast reuse index was calculated to be 0.94. The preset reuse threshold was 0.85. Since the reuse index difference of 0.09 was greater than the preset difference threshold of 0.05, this batch of yeast was determined to be usable for the next batch of fermentation. According to the tiered adjustment strategy, the reuse index difference of 0.09 fell between the first preset difference threshold of 0.05 and the second preset difference threshold of 0.10. Therefore, using a second inoculation adjustment coefficient of 1.10, the inoculation amount for the next batch was adjusted to 1.10 times the standard inoculation amount.

[0067] In this embodiment, the main fermentation cycle was 5 days, the reduction cycle was 3 days, and the total fermentation cycle was 8 days, which was 2 days shorter than that of Example 1. The recovered yeast activity was 94%, which was significantly higher than that of Example 1.

[0068] Example 3 This embodiment, based on embodiment 2, further increases the inoculation adjustment coefficient to its maximum value, while keeping the other conditions the same.

[0069] Expansion stage: Same as in Example 1, obtain qualified yeast liquid for expansion.

[0070] Inoculation phase: Similar to Example 1, the type that settles too quickly is deemed unqualified, and forced bottom ventilation is performed. In addition, the ventilation volume is increased to 2.2 times the normal ventilation volume, and the ventilation time is extended to 20 minutes.

[0071] Primary fermentation stage: Primary fermentation was carried out at 12°C, with an initial apparent sugar content of 12°P, monitored every 12 hours. The fermentation process was too rapid; the apparent sugar content dropped to 4.0°P on the 4th day, ending the primary fermentation. The primary fermentation cycle was 4 days, a further reduction of 1 day compared to Example 2. However, monitoring revealed that the highest temperature during fermentation reached 14.5°C, exceeding the upper limit of the preset fermentation temperature range, indicating that yeast metabolism was too vigorous.

[0072] Diacetyl reduction stage: Reduction was carried out by increasing the temperature to 14°C at a rate of 0.5°C / h, with diacetyl content monitored every 6 hours. On the 3rd day, the diacetyl content decreased to 0.09 mg / L, reaching the acceptable threshold, and the reduction was terminated. The reduction cycle was the same as in Example 2.

[0073] Yeast reuse stage: After cooling and settling, the yeast sludge was recovered. Cell viability was monitored at 82%, and the mortality rate was 15%. The calculated yeast reuse index was 0.82, which is less than the preset reuse threshold of 0.85. Therefore, this batch of yeast was deemed unusable. Analysis revealed that excessive inoculation and over-aeration led to overly vigorous yeast metabolism and exceeded fermentation temperature limits. Under high temperature and high osmotic pressure stress, yeast viability significantly decreased, and cell mortality increased.

[0074] In this embodiment, the main fermentation cycle is 4 days, the reduction cycle is 3 days, and the total fermentation cycle is 7 days. Although the cycle is the shortest, the recovered yeast activity is only 82%, which is below the reuse threshold and cannot be used for the next batch of fermentation. Excessive adjustment causes the yeast to be subjected to heat stress and osmotic pressure stress. Although the single-batch fermentation efficiency is the highest, the reuse value of the yeast is sacrificed. From the perspective of multi-batch continuous production, the overall benefits are not as good as in Example 2.

[0075] Those skilled in the art should understand that the discussion of any of the above embodiments is merely exemplary and is not intended to imply that the scope of the invention (including the claims) is limited to these examples; within the framework of the invention, the technical features of the above embodiments or different embodiments can also be combined, the steps can be implemented in any order, and there are many other variations of the different aspects of the invention as described above, which are not provided in the details for the sake of brevity.

[0076] This invention is intended to cover all such substitutions, modifications, and variations that fall within the broad scope of the appended claims. Therefore, any omissions, modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this invention should be included within the scope of protection of this invention.

Claims

1. A method for applying brewer's yeast in fermentation process control, characterized in that, include: Step S1: The brewer's yeast strain is cultured step by step under preset conditions to obtain expanded yeast liquid. During the expansion process, samples are taken step by step to monitor the number of viable yeast cells and the mortality rate to calculate the yeast viability index. The pass rate of yeast expansion is determined based on the comparison between the yeast viability index and the preset viability threshold. If the pass rate is not met, a treatment strategy is determined based on the current expansion level. Step S2: The qualified yeast liquid is subjected to low-temperature starvation treatment. The starved yeast is inoculated into the wort. The dissolved oxygen consumption rate and yeast germination rate are monitored at preset time intervals to calculate the inoculation adaptability index. The inoculation is qualified based on the rate of change of the inoculation adaptability index. If it is unqualified, the unqualified type is determined based on the half-life of the liquid surface foam and the characteristic morphology of the foam, and the adjustment strategy is determined according to the unqualified type. Step S3: After successful inoculation, carry out primary fermentation within the preset fermentation temperature range, monitor the appearance sugar content until the appearance sugar content drops to the preset primary fermentation endpoint threshold, and end the primary fermentation. Step S4: After the main fermentation is completed, the temperature of the fermentation broth is raised to the preset reduction temperature range at a preset heating rate to carry out diacetyl reduction. The reduction ends when the diacetyl content is less than the preset qualified threshold. Step S5: Cool the reduced fermentation broth to allow it to settle, recover the yeast sludge settled at the bottom of the tank, monitor the cell viability and mortality of the recovered yeast to calculate the yeast reuse index, and determine whether the current batch of yeast can be used for the next batch of fermentation based on the comparison result between the yeast reuse index and the preset reuse threshold, and whether the inoculation amount for the next batch should be adjusted if it is available.

2. The method for applying brewer's yeast in fermentation process control according to claim 1, characterized in that, The preset conditions in step S1 are as follows: the expansion temperature is 20-24℃, the expansion medium is wort with 8-12°P, the dissolved oxygen content during the expansion process is controlled at 6-10mg / L, the inoculation ratio for step-by-step expansion is 1:5-1:20, and the expansion time for each step is 24-48 hours.

3. The method for applying brewer's yeast in fermentation process control according to claim 2, characterized in that, In step S1, based on the condition that the yeast activity index is less than the preset activity threshold, the yeast expansion is determined to be unqualified, and a treatment strategy is determined based on the current expansion level. The propagation levels are, in order: Erlenmeyer flask level, Karl Casner level, seed tank level, and propagation tank level.

4. The method for applying brewer's yeast in fermentation process control according to claim 3, characterized in that, If the current expansion level is Erlenmeyer flask level, the treatment strategy is to discard the current batch of yeast and start expansion again from the preserved strain. If the current expansion level is the Karl flask level, the handling strategy is to discard the current batch of yeast and start expansion again from the Erlenmeyer flask level. If the current expansion level is seed tank level, the treatment strategy is to extend the expansion time by 2-6 hours and supplement aeration, re-monitor the yeast activity index, and discard it if it is still unqualified. If the current expansion level is expansion tank level, the treatment strategy is as follows: extend the expansion time by 2-6 hours and supplement aeration, re-monitor the yeast activity index, and if it is still unqualified, discard it and trace back to the quality of the previous level seed tank level yeast.

5. The method for applying brewer's yeast in fermentation process control according to claim 4, characterized in that, In step S2, the low-temperature starvation treatment is a carbon source starvation treatment, and the temperature of the low-temperature starvation treatment is 2-10℃, and the treatment time is 6-48 hours. During the treatment, the yeast liquid is intermittently stirred at a speed of 30-100 rpm, once every 2-4 hours, for 5-15 minutes each time.

6. The method for applying brewer's yeast in fermentation process control according to claim 5, characterized in that, In step S2, the process of determining whether the vaccination is qualified is as follows: The first vaccination adaptability index and the second vaccination adaptability index were calculated at the first and second preset time points after vaccination, respectively, and the rate of change between the two was calculated. When the rate of change is greater than or equal to a preset rate of change threshold, and the second vaccination adaptability index is greater than or equal to a preset adaptability threshold, the vaccination is deemed qualified. When the rate of change is less than a preset rate of change threshold, or the second inoculation adaptability index is less than a preset adaptability threshold, the inoculation is deemed unqualified, and the unqualified type is determined based on the half-life of the liquid surface foam and the characteristic morphology of the foam. The inoculation adaptability index is determined by the dissolved oxygen consumption rate and the yeast germination rate.

7. The method for applying brewer's yeast in fermentation process control according to claim 6, characterized in that, The specific process for determining non-compliance types based on the half-life and characteristic morphology of surface foam, and then determining adjustment strategies based on the non-compliance types, is as follows: Measure the foam half-life on the liquid surface within a preset time period after inoculation and observe the characteristic morphology of the foam; The foam half-life is compared with a preset half-life threshold, and the foam morphology is compared with a preset morphology feature. If the foam half-life is less than the first preset half-life threshold, it is judged as an unqualified type of excessively fast settling, and the adjustment strategy is to start bottom pulse ventilation or forced bottom ventilation. If the foam half-life is greater than the second preset half-life threshold and the foam characteristic morphology is viscous and stringy, it is judged as an abnormal viscosity type and is unqualified. The adjustment strategy is to switch to pulse stirring mode. If the foam half-life is greater than the second preset half-life threshold and the foam characteristic morphology is non-sticky and stringy, it is judged as unqualified due to foam runaway. The adjustment strategy is to reduce the tank pressure and start mechanical defoaming.

8. The method for applying brewer's yeast in fermentation process control according to claim 7, characterized in that, The specific process for determining whether the current batch of yeast can be used for the next batch of fermentation is as follows: Monitor the cell viability and mortality rate of recycled yeast, and calculate the yeast reuse index based on the ratio of cell viability to mortality rate; The yeast reuse index is compared with the preset reuse threshold. If the yeast reuse index is greater than or equal to the preset reuse threshold, it is determined that the current batch of yeast can be used for the next batch of fermentation, and it is determined whether the inoculation amount for the next batch should be adjusted. If the yeast reuse index is less than the preset reuse threshold, then the current batch of yeast is determined not to be reused.

9. The method for applying brewer's yeast in fermentation process control according to claim 8, characterized in that, The inoculation amount for the next batch is adjusted based on the condition that the difference in the yeast reuse index is greater than a preset difference. The difference in the yeast reuse index is the difference between the yeast reuse index and the preset reuse threshold.

10. The method for applying brewer's yeast in fermentation process control according to claim 9, characterized in that, The difference between the next batch inoculation amount and the yeast reuse index is positively correlated.

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