Liquor brewing method based on multi-strain synergistic fermentation

Abstract

The application discloses a liquor brewing method based on multi-strain synergistic fermentation and belongs to the technical field of liquor brewing. The method comprises the steps of raw material pretreatment, multi-strain system construction and carrier loading, synergistic fermentation and post-treatment. After being crushed and sieved, grains are subjected to cooking and sterilization to obtain a fermentation substrate. Gluing red yeast, bacillus subtilis and aspergillus niger are proportioned and loaded on a composite protective carrier of trehalose, chitosan, sodium carboxymethyl cellulose and porous starch microspheres to prepare a load type composite strain. After inoculation, the fermentation is controlled in sections at different temperatures, the pH value is dynamically regulated, and nutrients and composite enzyme preparations are supplemented. The liquor product is obtained through three-stage distillation and decolorization. Through the multi-strain synergistic and carrier protection technology, the method solves the problems of single flavor of the traditional process, long fermentation period, unstable batch, etc., significantly increases the types of volatile flavoring substances, reduces the content of harmful impurities, improves the utilization rate of raw materials, and makes the liquor body aroma coordinated and the taste mellow.

Description

Technical Field

[0001] This invention belongs to the field of liquor brewing technology, specifically relating to a liquor brewing method based on multi-strain synergistic fermentation. Background Technology

[0002] Traditional baijiu fermentation has long relied on natural koji making or single-strain inoculation, presenting multiple technological bottlenecks that urgently need to be overcome. From the perspective of strain application, the metabolic pathways of single strains are relatively limited, synthesizing only a limited number of flavor precursors. This results in a thin aroma profile and insufficient flavor complexity in the finished baijiu, failing to meet consumers' demands for diverse tastes. Simultaneously, single strains are less adaptable to the fermentation environment, easily affected by factors such as temperature fluctuations, pH changes, inhibition by crude fiber in raw materials, and competition from other microorganisms. This leads to decreased metabolic activity, slowed proliferation rates, and even premature inactivation, directly resulting in low fermentation efficiency. Traditional fermentation processes typically have a long cycle of 1-3 months, severely restricting production capacity increases. From the perspective of quality stability, the lack of a systematic strain protection mechanism results in inconsistent survival rates of strains in the complex fermentation system. Significant differences in flavor substance content, alcohol content, and physicochemical indicators exist between different batches of products, leading to poor batch-to-batch consistency and posing a significant challenge to enterprise quality control. Furthermore, in traditional fermentation processes, nutrients such as starch and cellulose in the raw materials are not fully degraded. This not only results in low raw material utilization but may also lead to abnormal viscosity in the fermentation system due to the accumulation of undegraded components. This further affects microbial metabolism and product diffusion, ultimately resulting in insufficient total ester content and the proportion of key flavor substances in the finished baijiu. Moreover, the content of harmful impurities such as methanol and fusel oils is difficult to control effectively, affecting product quality and posing potential safety risks. These problems have long plagued the baijiu brewing industry, becoming core technological pain points restricting its high-quality development.

[0003] CN107475012A discloses a method for producing light-aroma baijiu through multi-strain enhanced fermentation using Daqu (a type of starter culture). This method utilizes fresh mash from Fenjiu (a type of Chinese liquor) to isolate and screen high-ester-producing *Saccharomyces cerevisiae*, amylase-producing *Rhizopus oryzae*, acid- and alcohol-resistant *Monascus fuliginosus*, and heat-resistant *Bacillus subtilis* with protease and amylase activity. The mash is prepared into brewing yeast starter, Rhizopus oryzae starter, Monascus fuciformis starter, and Bacillus subtilis starter culture. Then, based on the weight of the mash, 1%-2% brewing yeast starter, 5%-7% Rhizopus oryzae starter, and 3%-5% starter culture are inoculated for large-scale fermentation. Then, 0.5%-1% Monascus fuciformis starter and 1%-2% Bacillus subtilis starter culture are inoculated for pile fermentation. Finally, 1%-2% brewing yeast starter, 5%-7% Rhizopus oryzae starter, and 3%-5% starter culture are inoculated into the pile fermented mash for secondary fermentation.

[0004] CN110564546A discloses a method for brewing wine using microbial strains, including the following steps: S1, Raw material pretreatment: Select glutinous sorghum and glutinous corn, and then select aged glutinous rice that has been stored for two years. Crush the glutinous sorghum, glutinous corn, and aged glutinous rice separately until the particle size is less than 20 mesh, and then mix the crushed glutinous sorghum, glutinous corn, and aged glutinous rice evenly; S2, Raw material soaking: Put the evenly mixed raw materials from step S1 into a soaking tank, add water at room temperature and soak for 1-2 days, stirring once every 6 hours; S3, Steaming: Pour out the soaking water, rinse the raw materials with clean water, drain, and put them into a steamer for steaming. First, bring the water in the steamer to a boil over high heat and maintain for 30 minutes, then adjust to low heat and maintain for 30 minutes. Close the steamer, scatter the raw materials in the steamer, pile them up again, add warm water, and continue to steam over low heat for 30-40 minutes. 0 min; S4, Preliminary Fermentation: After steaming, the raw materials are ventilated and cooled, then Rhizopus and Monascus purpureus are added and stirred evenly. The mixture is then placed in a ceramic jar for preliminary fermentation at a temperature of 45-50℃ for 2-3 hours; S5, Cellar Fermentation: The mixture is placed in a cellar for fermentation at a temperature of 34-38℃ for 7 days. The bottom of the cellar is lined with cellar mud, and microbial strains are added during the preparation of the cellar mud; S6, Post-processing: After fermentation, the wine and lees are separated using a filter press. The wine is then filtered through a filter cloth to remove impurities. The wine is then blended, heated for sterilization, and allowed to cool naturally before being sealed in wine jars and placed in a dry, cool place. The microbial strains include highly active brewing yeast, aroma-producing yeast, Bacillus subtilis, and lactic acid bacteria. The cellar mud also contains a substrate that provides nutrients for the microbial strains.

[0005] CN117285995A discloses a method for preparing a special starter culture for baijiu (Chinese liquor), comprising the following steps: (1) different Bacillus, mold, yeast, and thermophilic actinomycetes are cultured separately in slant tubes to obtain seed liquids, and then mixed in proportion to obtain mixed seed liquids of Bacillus, mold, yeast, and thermophilic actinomycetes; (2) wheat is crushed into coarse powder resembling plum petals, requiring the coarse powder to contain 65% epidermis and 35% powder, without hard lumps, and then water of 30-35% of the wheat mass is added, and stirred evenly to obtain wheat slurry. (3) Sterilize at 50-60℃ for 120 minutes; (4) Mix the mixed seed liquid of Bacillus, mold, yeast and thermophilic actinomycetes in proportion, add 6-7% of the wheat slurry volume to the wheat slurry, and then add 3% of the wheat slurry volume to the plant extract, mix well, put into a 10cm×10cm×3cm mold, and press into an inner embryo; (5) Mix the mixed seed liquid of Bacillus, mold and yeast in proportion, add 6-7% of the wheat slurry volume to the wheat slurry, and then add 3% of the wheat slurry volume to the plant extract, mix well, put into a 10cm×10cm×3cm mold, and press into an inner embryo; Add plant extracts, mix well to obtain a mixed slurry, spread it on the bottom of a 30cm×20cm×6cm mold with a thickness of 10cm, press it down, put the inner embryo in the center, add the mixed slurry, and press it into a koji embryo; (5) Place the pressed koji embryo for 2-3 hours, and after the surface is slightly dry and hardened, transfer it to the koji room for cultivation. Stack the koji embryos in a staggered pattern of three horizontal and three vertical, with the upper and lower layers perpendicular to each other, for a total of 6 layers. Separate the adjacent koji embryos with straw; (6) Turn the koji each time the core temperature reaches 60-62℃. In summer, turn the koji embryos twice. The process involves three turnings of the koji during winter. When the temperature of the middle layer of the koji embryo drops to room temperature after the turnings are completed, the koji production is finished, and high-temperature koji is obtained. The Bacillus species include Bacillus subtilis, Bacillus cereus, and Bacillus licheniformis, with a volume ratio of (1-1.2):(1-1.5):(1.5-2). The molds include Aspergillus niger, Monascus purpureus, Rhizopus oryzae, Aspergillus glaucus, and Trichoderma reesei, with a volume ratio of (1.2-1.5):(1-1.5):(1.5-1.7):(1.2-1.5):(1-1.5).

[0006] CN118440791A discloses a method for producing a high-yield yeast starter, comprising the following steps: S1, selecting high-quality grains as raw materials; S2, pre-treatment of raw materials, screening and cleaning all raw materials; S3, crushing raw materials, finely crushing wheat bran, corn, barley, and peas; S4, mixing and moistening raw materials, mixing the crushed wheat bran, corn, barley, and peas with rice husks according to the raw material ratio, and moistening the rice husks and the crushed wheat bran, corn, barley, and peas; S5, steaming, steaming the mixed raw materials at high temperature; S6, cultivating the saccharification and fermentation agent. Specific microbial strains are added to the raw materials after steaming; S7, fermentation and aging, the yeast gradually matures after a certain period of fermentation and aging; S8, finished product and storage, the matured yeast is dried, packaged and stored; the microbial strains added in S6 mainly include molds, yeasts and bacteria, and the molds, yeasts and bacteria mainly include high-ester-producing Monascus red mold, high-saccharifying enzyme activity Rhizopus, high-fermentation-capacity Saccharomyces cerevisiae, high-ethyl acetate-producing yeast, Nematomys, Geotrichum candida, high-amylase-activity Bacillus subtilis and Bacillus licheniformis, which are used to improve the efficiency of the fermentation process and the alcohol yield of the yeast.

[0007] CN120682892A discloses a method for preparing a mellow, sauce-flavored baijiu, as follows: Step 1, Raw material preparation: Take sorghum, crush it to a crushing degree of 15-25%, moisten it with fermentation water at 80-95℃ for 3-8 hours, add baijiu lees accounting for 4-8% of the sorghum mass, mix well, and steam together. After cooling indoors, sprinkle with cooling water at 80-95℃ to make up the difference, where the fermentation water accounts for 40-60% of the feed mass and the cooling water accounts for 5-10% of the feed mass, to obtain the sorghum material for the first fermentation stage; Step 2, Preparation of starter culture: Use commercially available starter culture, which is pulverized into sauce-flavored starter powder before use; Step 3, Feeding the first fermentation stage: Spread the sorghum material for the first fermentation stage, add sauce-flavored starter powder and enzyme catalyst, mix well, and then pile it up for fermentation. When the temperature at the top of the pile reaches 45-50℃, the mash inside the pile has a sweet and alcoholic aroma, and the moisture content is 35%. ~40% is fermented for 20-60 days to obtain Xia Sha mash; Step 4, Adding Xia Sha: Add an equal amount of Xia Sha sorghum to Xia Sha mash and steam it together to extract the liquor. After extracting the liquor, spread the mash out to cool, add soy sauce koji powder, functional agent and synergist, mix well and pile it up again, and then put it into the pit for fermentation for 20-60 days to obtain Xia Sha mash; Step 5, Distillation: Take out the Xia Sha mash and mix it with 10-30% of the mass of steamed rice husks, cover the plate and distill the liquor, control the distillation pressure at 0.005-0.02 MPa, the distillation temperature at 32-38℃, cut off the head and tail, and then let it age for 3-9 months to obtain mellow soy sauce aroma type baijiu; The enzyme catalyst is prepared by crushing Bacillus subtilis, Lactobacillus plantarum and brewing yeast evenly in a weight ratio of 0.5-2:1-3:3-5. Summary of the Invention

[0008] The purpose of this invention is to provide a method for brewing baijiu (Chinese liquor) based on the synergistic fermentation of multiple microorganisms, so as to solve the problems mentioned in the background art.

[0009] This invention provides a method for brewing baijiu (Chinese liquor) based on multi-strain synergistic fermentation, which includes the following steps:

[0010] S1. Raw material pretreatment: After the grain is crushed and sieved, the resulting brewing raw material is mixed evenly with water and cooked at a constant temperature. After cooking, it is cooled and sterilized to obtain a sterile fermentation substrate.

[0011] S2. Construction of multi-strain systems and vector loading:

[0012] S201. Using Rhodotorula mucilaginosa (CGMCC 2.167) as the core fermentation strain, Bacillus subtilis (CGMCC 1.1086) as the auxiliary control strain, and Aspergillus niger (CGMCC 3.795) as the flavoring strain, Rhodotorula mucilaginosa, Bacillus subtilis, and Aspergillus niger were activated separately to obtain Rhodotorula mucilaginosa culture, Bacillus subtilis culture, and Aspergillus niger culture.

[0013] S202. Trehalose, chitosan and sodium carboxymethyl cellulose are mixed in a mass ratio of 2~4.5:1.5~3:0.5~2, and water is added to prepare a mixed solution with a mass concentration of 4~6%. The mixed solution is then mixed with porous starch microspheres in a mass ratio of 1:3~5. After adsorption at 2~4℃, the mixture is freeze-dried to obtain a protective carrier.

[0014] S203. Mix the three bacterial cultures with the protective carrier at a mass ratio of 1:4~6, shake to adsorb, and then mix the three loaded bacterial cultures at a mass ratio of 6:1~4:1~4 to obtain a loaded composite bacterial culture.

[0015] S3. Co-fermentation: Inoculate the loaded compound microbial strain into the aseptic fermentation substrate at 0.5-1.5% of the substrate mass, and seal for staged fermentation: First stage: constant temperature fermentation at 35-37.5℃ for 3-4 days; Second stage: after the first stage, cool down to 20-23.5℃ for 7-10 days; the total fermentation time for the first and second stages is 11-13 days.

[0016] During fermentation, on the third day of fermentation, 0.1-1.5% of glucose by weight of the brewing raw materials is added; on the fifth day of fermentation, 0.2-0.4% of a compound enzyme preparation by weight of the brewing raw materials is added. The compound enzyme preparation is a mixture of cellulase and hemicellulase in a mass ratio of 12-14:1-3, or a mixture of cellulase, hemicellulase and pectinase in a mass ratio of 12-14:1-3:2-4; on the seventh day, 0.03-0.07% of yeast extract by weight of the brewing raw materials is added.

[0017] During fermentation, the pH was maintained at 4.0-4.2 in the first stage and 3.6-3.8 in the second stage. The dissolved oxygen content of the fermentation system was kept at 0.5-1.0 mg / L. Fermentation was terminated when the residual sugar content was ≤0.5 g / L and the pH value was stable.

[0018] S4. Post-processing: After fermentation, the liquor is filtered, distilled and decolorized to obtain the finished liquor.

[0019] In the above-mentioned method for brewing baijiu based on multi-strain synergistic fermentation, in step S1, the grains are sorghum: wheat: corn in a mass ratio of 4~6:2.5~3.5:1.5~2.5.

[0020] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, step S1 involves sieving through a 40-mesh standard sieve and collecting the material passing through the sieve.

[0021] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, in step S1, the mass ratio of brewing raw materials to water is 1:1.0~1.5.

[0022] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, step S1 involves constant temperature steaming at 100~110℃ for 20~40 minutes.

[0023] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, in step S1, the cooling is carried out by air cooling, with a cooling rate ≥5℃ / min, cooling to 30~38℃.

[0024] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, the sterilization in step S1 is steam sterilization at 110~130℃ for 20~30 minutes.

[0025] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, the activation operation of *Rhodotorula glutinis* in step S201 is as follows: *Rhodotorula glutinis* is activated at a constant temperature of 24-30℃ for 25-30 minutes using YM medium until the bacterial concentration is ≥1×10⁻⁶. 8 CFU / mL was used to obtain Rhodotorula glutinis culture.

[0026] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, the activation operation of Bacillus subtilis in step S201 is as follows: Bacillus subtilis is activated using LB medium at 30~38℃ for 18~22 min until the bacterial concentration is ≥1×10⁻⁶. 9 CFU / mL was used to obtain Bacillus subtilis bacterial suspension.

[0027] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, the activation operation of Aspergillus niger in step S201 is as follows: Aspergillus niger is cultured in PDA medium at 24~33℃ for 64~72h until the bacterial concentration is ≥1×10⁻⁶. 9 CFU / mL was used to obtain Aspergillus niger solution.

[0028] In the above-mentioned method for brewing baijiu based on multi-strain synergistic fermentation, in step S202, the porous starch microspheres have a particle size of 5nm~100μm.

[0029] Preferably, in the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, the porous starch microspheres in step S202 have a particle size of 50~80μm or 10~30nm.

[0030] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, the adsorption time in step S202 is 2-3 hours.

[0031] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, in step S203, the oscillation adsorption rate is 120~150 r / min, the time is 1~2 h, and the strain adsorption efficiency is ≥92%.

[0032] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, in the first stage of step S3, the stirring rate is controlled at 80~100 r / min for the first two days, and then reduced to 55~65 r / min for the next 1~2 days; in the second stage, the stirring rate is controlled at 40~50 r / min, and an intermittent bidirectional stirring mode is adopted.

[0033] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, the following operation is added in step S3: during the fermentation process, on the 6th day of fermentation, 0.02~0.04% of the mass of the brewing raw materials is added with L-glutamate sodium.

[0034] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, in step S3, the activity of the pectinase is 20,000 to 40,000 U / g.

[0035] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, in step S3, the residual sugar content and pH value are measured every 4 to 8 hours during the fermentation process, and the pH value is dynamically adjusted by adding 0.05 to 0.1 mol / L citric acid aqueous solution.

[0036] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, in step S4, the distillation adopts a three-stage distillation: the first stage is distilled at 65~75℃ for 1~1.5h to remove low-boiling-point impurities; the second stage is distilled at 75~80℃ for 3~5h to extract the target fraction with an alcohol content of 52~55%vol; and the third stage is distilled at 80~85℃ for 0.5~1h to recover the residual liquid.

[0037] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, in step S4, the filtration is pressure filtration, and the filter cloth pore size is 5~10μm.

[0038] In the above-mentioned baijiu brewing method based on multi-strain synergistic fermentation, in step S4, the decolorization involves adding 0.5-1% of the target fraction mass of 100-200 mesh activated carbon to the target fraction, adsorbing and decolorizing at 15-20℃ for 24-36 hours, and then filtering through a precision filter.

[0039] In this invention, Rhodotorula mucilaginosa CGMCC 2.167, Bacillus subtilis CGMCC 1.1086, and Aspergillus niger CGMCC 3.795 are all commercially available and were purchased from the China General Microbiological Culture Collection Center (CGMCC).

[0040] The beneficial effects of this invention are:

[0041] This invention constructs a three-layered microbial symbiotic network consisting of core fermentation bacteria, auxiliary regulatory bacteria, and flavor bacteria, forming a functionally complementary metabolic closed loop. *Rhodotorula glutinis*, as the core fermentation bacteria, efficiently converts starch into alcohol. *Bacillus subtilis* secretes cellulase and protease, which degrade crude fiber and large protein molecules in the raw materials, providing ample nutrition for *Rhodotorula glutinis* and optimizing the microenvironment of the fermentation system. *Aspergillus niger* synthesizes esterases, promoting the formation of esters. The synergistic effect of these three factors increases the variety of volatile flavor compounds by more than 30% compared to traditional processes, significantly enhancing the content of key flavor components such as alcohols and esters, resulting in a more harmonious aroma and a richer, more mellow taste.

[0042] This protective carrier, composed of trehalose, chitosan, sodium carboxymethyl cellulose, and porous starch microspheres, possesses adsorption, slow-release, and protective functions. Trehalose protects the cell membrane structure of the microorganisms from environmental stress, while the cationic properties of chitosan enhance the binding force between the microorganisms and the carrier. The porous starch microspheres with a particle size of 5nm~100μm provide stable colonization sites for the microorganisms, effectively avoiding the fermentation failure problem caused by premature inactivation of the microorganisms in traditional processes. Simultaneously, the segmented fermentation process allows for precise control of pH and temperature, fully leveraging the synergistic fermentation effects of each microorganism, shortening the fermentation cycle, and achieving thorough degradation of the raw material nutrients. The combination of three-stage distillation and low-temperature activated carbon decolorization effectively removes low-boiling-point impurities and harmful substances. Attached Figure Description

[0043] Figure 1 This is a process flow diagram of the present invention. Detailed Implementation

[0044] Specifically, this invention provides a method for brewing baijiu (Chinese liquor) based on multi-strain synergistic fermentation, which includes the following steps:

[0045] S1. Raw Material Pretreatment: Mix sorghum, wheat, and corn in a mass ratio of 4~6:2.5~3.5:1.5~2.5, pulverize using a universal grinder, and pass through a 40-mesh standard sieve. Collect the sieved material to ensure uniform particle size, facilitating subsequent cooking and gelatinization. Add 1.0~1.5 times the mass of deionized water to the sieved brewing raw materials, stir well, and transfer to a cooking tank. Cook at a constant temperature of 100~110℃ for 20~40 minutes to fully gelatinize the starch in the raw materials and break down the cell wall structure. After cooking, rapidly cool to 30~38℃ using air cooling, with a cooling rate controlled at ≥5℃ / min to prevent material clumping from affecting subsequent fermentation. After cooling, transfer the material to a fermentation tank and sterilize with steam at 110~130℃ for 20~30 minutes to kill miscellaneous bacteria in the raw materials, reduce the risk of fermentation contamination, and obtain a sterile fermentation substrate.

[0046] S2. Construction of multi-strain systems and vector loading:

[0047] S201 uses *Rhodotorula mucilaginosa* (CGMCC 2.167) as the core fermentation strain, *Bacillus subtilis* (CGMCC 1.1086) as the auxiliary control strain, and *Aspergillus niger* (CGMCC 3.795) as the flavoring strain. The *Rhodotorula mucilaginosa* is activated on YM medium at 24–30°C for 25–30 min until the bacterial concentration is ≥1×10⁻⁶. 8The concentration of CFU / mL was adjusted to ensure high alcohol production capacity, resulting in Rhodotorula glutinis culture. Bacillus subtilis was activated on LB medium at 30–38°C for 18–22 min until the bacterial concentration was ≥1×10⁻⁶. 9 CFU / mL was used to enhance its enzyme production activity, resulting in Bacillus subtilis bacterial suspension; Aspergillus niger was cultured on PDA medium at 24-33℃ for 64-72 h until the bacterial concentration was ≥1×10⁻⁶. 9 CFU / mL was used to enhance its esterase secretion ability, resulting in Aspergillus niger solution;

[0048] S202. Trehalose, chitosan, and sodium carboxymethyl cellulose are mixed in a mass ratio of 2-4.5:1.5-3:0.5-2, and deionized water is added to prepare a mixed solution with a mass concentration of 4-6%. The mixed solution is then mixed with porous starch microspheres with a particle size of 5nm-100μm (preferably 50-80μm or 10-30nm) in a mass ratio of 1:3-5. The mixture is adsorbed at 2-4℃ for 2-3 hours, and then freeze-dried to obtain a protective carrier. This carrier can effectively retain the active substances of the strain and resist the stress of the fermentation environment.

[0049] S203. The activated bacterial cultures of the three strains are mixed with the protective carrier at a mass ratio of 1:4~6. The cultures are shaken and adsorbed for 1~2 hours at a shaking rate of 120~150 r / min. The bacterial adsorption efficiency is ≥92%. The three loaded bacterial cultures are then mixed at a mass ratio of 6:1~4:1~4 to obtain a loaded composite bacterial culture.

[0050] S3. Co-fermentation: The loaded compound microbial strain is inoculated into the aseptic fermentation substrate at 0.5-1.5% of the substrate mass. The fermentation is carried out in stages in a sealed fermenter: Stage 1: Constant temperature fermentation at 35-37.5℃ for 3-4 days. For the first 2 days, the stirring rate is controlled at 80-100 r / min to promote uniform diffusion and rapid proliferation of the microbial strain in the substrate. For the next 1-2 days, the stirring rate is reduced to 55-65 r / min to reduce excessive nutrient consumption. Stage 2: After the first stage, the temperature is lowered to 20-23.5℃ for 7-10 days of low-temperature fermentation. The stirring rate is adjusted to 40-50 r / min, using an intermittent bidirectional stirring mode to promote diffusion of microbial metabolites and full contact with the substrate. The total fermentation time for both stages is 11-13 days.

[0051] During fermentation, on the third day, 0.1-1.5% of the weight of the brewing raw materials (i.e., the total weight of sorghum, wheat, and corn) of glucose is added to provide a carbon source for microbial metabolism. On the fifth day, 0.2-0.4% of the weight of the brewing raw materials of a compound enzyme preparation is added. The compound enzyme preparation is a mixture of cellulase and hemicellulase in a mass ratio of 12-14:1-3, or a mixture of cellulase, hemicellulase, and pectinase (enzyme activity 20,000-40,000 U / g) in a mass ratio of 12-14:1-3:2-4. This synergistically degrades crude fiber in the raw materials with Bacillus subtilis and promotes the degradation of pectin. On the seventh day, 0.03%-0.07% of the weight of the brewing raw materials of yeast extract is added to supplement nitrogen source and growth factors.

[0052] The following steps can be added during the fermentation process: On the 6th day of fermentation, add 0.02%~0.04% of the weight of the brewing raw materials with L-glutamate to promote the synthesis of γ-aminobutyric acid by Aspergillus niger;

[0053] During fermentation, the residual sugar content and pH value were measured every 4-8 hours. The pH was dynamically adjusted by adding 0.05-0.1 mol / L citric acid aqueous solution. The pH was maintained at 4.0-4.2 in the first stage and 3.6-3.8 in the second stage. The dissolved oxygen content of the fermentation system was kept at 0.5-1.0 mg / L. Fermentation was terminated when the residual sugar content was ≤0.5 g / L and the pH value was stable.

[0054] S4. Post-processing: After fermentation, the fermentation broth is pressure filtered using a plate and frame filter press with a filter cloth pore size of 5-10 μm to remove fermentation residues and insoluble impurities. The clarified filtrate is collected and distilled in three stages: the first stage is distilled at 65-75℃ for 1-1.5 hours to remove low-boiling-point impurities; the second stage is distilled at 75-80℃ for 3-5 hours to obtain the target fraction with an alcohol content of 52-55% vol; and the third stage is distilled at 80-85℃ for 0.5-1 hours to recover the residual liquid. The target fraction is transferred to a decolorization tank, and 0.5-1% of the target fraction mass of 100-200 mesh activated carbon is added for adsorption and decolorization at 15-20℃ for 24-36 hours. Subsequently, it is filtered through a precision filter to remove activated carbon particles and trace impurities, yielding the finished liquor.

[0055] The present invention will be further described in detail below through embodiments, but the scope of protection of the present invention shall not be limited to the scope of the embodiments described herein. Where specific techniques or conditions are not specified in the embodiments, they shall be performed in accordance with the techniques or conditions described in the literature in the art or according to the product instructions. Reagents or instruments whose manufacturers are not specified are all conventional products that can be obtained commercially.

[0056] YM liquid culture medium formula: 10g glucose, 5g peptone, 3g yeast extract, 3g malt extract, 1000mL distilled water; LB liquid culture medium formula: 10g tryptone, 5g yeast extract, 10g sodium chloride, 1000mL distilled water; PDA liquid culture medium formula: 200g potato (peeled and diced, boiled in 1000mL distilled water for 30min, filtered and the filtrate is collected), 20g glucose, distilled water to 1000mL.

[0057] Example 1

[0058] This embodiment follows Figure 1 The flowchart shown illustrates the following steps in the brewing of baijiu:

[0059] S1. Raw Material Pretreatment: Mix 50kg of sorghum, 30kg of wheat, and 20kg of corn, then grind them using a universal grinder and pass them through a 40-mesh standard sieve. Collect the sieve-passing material to ensure uniform particle size, which facilitates subsequent cooking and gelatinization. Add 120kg of deionized water to the ground and sieved brewing raw materials, stir well, and then transfer them to a cooking tank. Cook at a constant temperature of 105℃ for 30 minutes to fully gelatinize the starch in the raw materials and break down the cell wall structure. After cooking, rapidly cool the raw materials to 35℃ using air cooling, with the cooling rate controlled at ≥5℃ / min to prevent material clumping from affecting subsequent fermentation. After cooling, transfer the material to a fermentation tank and sterilize it with steam at 121℃ for 20 minutes to kill any miscellaneous bacteria in the raw materials, reduce the risk of fermentation contamination, and obtain a sterile fermentation substrate.

[0060] S2. Construction of multi-strain systems and vector loading:

[0061] S201 uses *Rhodotorula mucilaginosa* (CGMCC 2.167) as the core fermentation strain, *Bacillus subtilis* (CGMCC 1.1086) as the auxiliary control strain, and *Aspergillus niger* (CGMCC 3.795) as the flavoring strain. The *Rhodotorula mucilaginosa* was activated on YM medium at 28°C for 25-30 minutes until the bacterial concentration was ≥1×10⁻⁶. 8 The concentration of CFU / mL was adjusted to ensure high alcohol production capacity, resulting in Rhodotorula glutinis culture. Bacillus subtilis was activated on LB medium at 37°C for 18-22 min until the bacterial concentration was ≥1×10⁻⁶. 9 CFU / mL was used to enhance its enzyme production activity, resulting in Bacillus subtilis bacterial suspension; Aspergillus niger was cultured on PDA medium at 30℃ for 72 h until the bacterial concentration was ≥1×10⁻⁶. 9 CFU / mL was used to enhance its esterase secretion ability, resulting in Aspergillus niger solution;

[0062] S202. Trehalose, chitosan, and sodium carboxymethyl cellulose were mixed in a mass ratio of 3:2:1, and deionized water was added to prepare a 5% mass concentration mixed solution. The mixed solution was then mixed with porous starch microspheres with a particle size of 60-70 μm in a mass ratio of 1:4. The mixture was adsorbed at 4℃ for 2 hours and then freeze-dried to obtain a protective carrier. This carrier can effectively lock in the active substances of the strain and resist the stress of the fermentation environment.

[0063] S203. The activated bacterial cultures of the three strains are mixed with the protective carrier at a mass ratio of 1:5. The cultures are shaken and adsorbed for 1 hour at a shaking rate of 120~150 r / min. The adsorption efficiency of the strains is ≥92%. The three loaded strains after adsorption are then mixed at a mass ratio of 6:2:2 to obtain a loaded composite strain.

[0064] S3. Co-fermentation: The loaded compound strain is inoculated into the sterilized sterile fermentation substrate at 0.8% of the mass of the sterile fermentation substrate, and the fermentation tank is sealed for staged fermentation.

[0065] Phase 1: Ferment at a constant temperature of 36℃ for 4 days. For the first 2 days, the stirring rate is controlled at 90r / min to promote the uniform diffusion and rapid proliferation of the inoculum in the substrate. For the last 2 days, the stirring rate is reduced to 60r / min to reduce excessive consumption of nutrients.

[0066] Second stage: Cool down to 22℃ for 8 days of low-temperature fermentation, adjust the stirring rate to 45r / min, and adopt intermittent bidirectional stirring mode to promote the diffusion of bacterial metabolites and full contact with the substrate;

[0067] During fermentation, 0.1% glucose by weight of the brewing raw materials was added on the 3rd day to provide a carbon source for microbial metabolism; on the 5th day, 0.3% compound enzyme preparation by weight of the brewing raw materials was added. The compound enzyme preparation was prepared by mixing cellulase and hemicellulase at a mass ratio of 13:2, which synergistically degrades crude fiber in the raw materials by Bacillus subtilis; on the 7th day, 0.05% yeast extract by weight of the brewing raw materials was added to supplement nitrogen source and growth factors; during fermentation, the residual sugar content and pH value were measured every 6 hours, and dynamically regulated by adding 0.1 mol / L citric acid solution. The pH was maintained at 4.0~4.2 for the first 4 days (i.e., the first stage), and at 3.6~3.8 for the next 8 days (i.e., the second stage), while maintaining the dissolved oxygen content of the fermentation system at 0.5~1.0 mg / L. Fermentation was terminated when the residual sugar content was ≤0.5 g / L and the pH value was stable.

[0068] S4. Post-processing: After fermentation, the fermentation broth is pressure filtered using a plate and frame filter press with a filter cloth pore size of 10μm to remove fermentation residues and insoluble impurities. The clarified filtrate is collected and distilled in three stages: the first stage is distilled at 70℃ for 1 hour to remove low-boiling-point impurities; the second stage is distilled at 75℃ for 3 hours to obtain the target fraction with an alcohol content of 52~55% vol; and the third stage is distilled at 80℃ for 0.5 hours to recover the residual liquid. The target fraction is transferred to a decolorization tank, and 0.5% of the target fraction mass of 100~200 mesh activated carbon is added for adsorption and decolorization at 15~20℃ for 24 hours. Then, it is filtered through a precision filter to remove activated carbon particles and trace impurities to obtain the finished liquor.

[0069] Example 2

[0070] Based on the method of Example 1, with everything else remaining unchanged, the co-fermentation parameters were adjusted:

[0071] The first stage of fermentation was carried out at a temperature of 37℃ for 3 days. The stirring rate was controlled at 90 r / min for the first 2 days and then reduced to 60 r / min for the last day. The second stage of fermentation was carried out at a temperature of 21℃ for 9 days and at a stirring rate of 45 r / min, using an intermittent bidirectional stirring mode.

[0072] On the 5th day of fermentation, an additional 0.05% pectinase (enzyme activity 30,000 U / g) was added to the compound enzyme preparation to promote the degradation of pectin in the raw materials.

[0073] Example 3

[0074] Based on the method of Example 1, with everything else remaining unchanged, the protective carrier and fermentation regulation were optimized:

[0075] The particle size of porous starch microspheres in the protective carrier was adjusted to 15~20nm to enhance the enrichment capacity of Aspergillus niger metabolites;

[0076] On day 6 of fermentation, an additional 0.03% of L-glutamate was added to promote the synthesis of γ-aminobutyric acid by Aspergillus niger; the second stage of fermentation lasted for 7 days.

[0077] Table 1 Comparison of Baijiu Indicators in Examples 1-3

[0078]

[0079] As mentioned above, traditional processes rely on natural koji making or a single microbial strain, resulting in a limited variety of volatile flavor compounds and a thin aroma profile. Example 1 of this invention, however, utilizes a multi-strain synergistic fermentation system, composite protective carrier technology, and a segmented process design. Compared to traditional baijiu brewing processes and existing related technologies, the fermentation methods are significantly different, leading to a marked improvement in baijiu quality. Example 1 employs a three-layer symbiotic system of Rhodotorula glutinis, Bacillus subtilis, and Aspergillus niger. The three components complement each other metabolically, forming a closed loop. This increases the variety of volatile flavor compounds by more than 30% compared to traditional processes, and raises the content of key flavor components such as alcohols and esters by 25%–40%. The resulting baijiu has a fuller, more harmonious aroma and a mellow, unblemished taste.

[0080] Traditional fermentation processes typically take 1-3 months. Example 1, through carrier protection technology to improve strain stability and combined with segmented temperature-controlled fermentation, reduces the total cycle to only 12 days, shortening it by more than 60% compared to traditional processes and significantly improving production efficiency. In traditional processes, nutrients such as starch and cellulose are not fully degraded, resulting in a raw material utilization rate of only 65%-75%. Example 1, through the synergistic effect of compound enzyme preparations and compound strain preparations, especially Bacillus subtilis, achieves a starch conversion rate of ≥90% and a crude fiber degradation rate of ≥85% in the raw materials, increasing the raw material utilization rate to 88%-92%. The yield of baijiu (Chinese liquor) is increased by 10%-15% compared to traditional processes, reducing production costs. Example 1 utilizes a multi-strain synergistic fermentation system and composite protective carrier technology to reduce methanol and fusel oil content. Further reductions in methanol and fusel oil content are achieved through a three-stage distillation and activated carbon decolorization process. Methanol content is reduced to 0.0015 g / 100 mL, and fusel oil content to 0.010 g / 100 mL, both significantly lower than national standards (methanol ≤ 0.04 g / 100 mL, fusel oil ≤ 0.2 g / 100 mL), resulting in higher product safety. Traditional processes suffer from large fluctuations in strain survival rates, leading to variations in flavor substance content and alcohol content between different batches of products exceeding 15%. The composite protective carrier in Example 1 achieves a strain adsorption efficiency of ≥ 92%, ensuring stable strain activity during fermentation and reducing the batch-to-batch variation of key indicators to ≤ 5%, thus addressing the industry pain point of batch instability inherent in traditional processes.

[0081] Example 2 adds pectinase to the compound enzyme preparation of Example 1 to further enhance the degradation of pectin in the raw materials. Pectinase degrades pectin in the raw materials to generate intermediate products such as galacturonic acid, providing substrates for Aspergillus niger to synthesize esterifying enzymes, promoting the formation of higher esters. The variety of volatile flavor compounds increases by approximately 14% compared to Example 1, resulting in a richer fruit and cellar aroma and a smoother taste. Simultaneously, pectin degradation avoids the increased viscosity caused by pectin accumulation in traditional fermentation, improving the fluidity of the fermentation broth by 30%, facilitating smoother diffusion of microbial metabolites, reducing residual sugar to below 0.3 g / L, clarifying the fermentation endpoint, and further optimizing batch-to-batch consistency. The thorough degradation of pectin reduces methanol precursors, lowering methanol content by 5%–10% compared to Example 1, and simultaneously reducing fusel oil content by 10%–15%, resulting in higher product purity.

[0082] Example 3 adjusted the particle size of porous starch microspheres to 15-20 nm and supplemented them with monosodium glutamate (MSG) on day 6 of fermentation, achieving the following breakthrough improvements: the porous starch microspheres had a larger specific surface area, increasing their adsorption capacity for Aspergillus niger mycelium by 30% and promoting the secretion of esterase and γ-aminobutyric acid (GABA) synthase; MSG, as a precursor, further induced Aspergillus niger to synthesize GABA, resulting in a GABA content ≥0.32 mg / 100 mL in the finished product, endowing the baijiu with functional health benefits. The presence of GABA neutralized the spiciness of the baijiu, making the taste smoother and more mellow; at the same time, the adsorption-slow release effect of the nanoscale carrier on flavor substances made it easier to separate esters and alcohols during distillation, increasing the proportion of flavor components in the target fraction by about 8%, resulting in a more lasting aroma and extending the aroma time in the empty glass by 2-3 hours. The nanocarrier improved the contact efficiency between the strain and the substrate, shortening the second-stage low-temperature fermentation time from 8 days to 7 days and reducing the total fermentation cycle to 11 days, which is 8% to 10% shorter than Example 1. This further improved production efficiency, while maintaining the raw material utilization rate at over 90%.

[0083] Examples 4-9: Effects of strain, carrier ratio, and fermentation temperature

[0084] Based on Example 1, this experiment investigated the effects of three key process parameters—the mass ratio of the loaded microbial strain, the ratio of the carrier, and the fermentation temperature—on four indicators (indicators 1 to 4) through six examples (Examples 4 to 9). The experimental results are shown in Table 2.

[0085] Table 2 Test Results of Key Process Parameters

[0086]

[0087] Table 2 shows the following effects on index 1 (alcohol content): Carrier ratio (+1.3) > Fermentation temperature (+1.0) > Loaded microbial strain mass ratio (+0.8), indicating that the carrier ratio has the most significant effect on improving index 1. Regarding index 2 (total ester content): Carrier ratio (+0.3) > Loaded microbial strain mass ratio / Fermentation temperature (+0.2), with the carrier ratio showing a slightly higher improvement. For index 3 (total types of volatile flavor compounds): Loaded microbial strain mass ratio (+2) = Carrier ratio (+2) > Fermentation temperature (+1), with the former two having a more significant promoting effect on index 3. Regarding index 4 (methanol content): Carrier ratio (-0.0003) > Loaded microbial strain mass ratio (-0.0002) > Fermentation temperature (-0.0001), with the carrier ratio showing the most significant effect on reducing index 4.

[0088] As shown in Table 2, the effect of parameter levels on the indicators is consistent: higher levels of the three types of parameters (higher proportion of strains, higher carrier ratio, or higher temperature) significantly increased indicators 1-3 and decreased indicator 4. This indicates that increasing parameter levels may be beneficial to fermentation efficiency or product accumulation, while reducing the content of by-products (or inhibitors).

[0089] Optimal parameter direction: The carrier ratio has the most comprehensive impact on all indicators, especially in improving indicators 1 and 3 and reducing indicator 4. The mass ratio of the loaded microorganisms is comparable to the carrier ratio in improving indicator 3 (such as product yield) and can be considered a secondary optimization direction. The fermentation temperature has a relatively small overall impact on the indicators, but the two-stage temperature control (37℃→23℃) can still stably improve key indicators. After comprehensive analysis, the optimal process conditions are: a mass ratio of loaded microorganisms of 6:3:3, a carrier ratio of 4:2.5:1.5, and a fermentation temperature of 37℃ / 23℃ (two-stage).

[0090] Comparative Examples 1-2

[0091] This invention investigated the necessity of composite protective carriers and temperature and pH control during fermentation. Comparative Example 1 was completely identical to Example 1 except that the protective carrier in step S202 was not added and the activated bacterial culture was directly inoculated. Comparative Example 2 was completely identical to Example 1 except that the temperature and pH were not controlled during the fermentation process and natural fermentation temperature and pH were used. The experimental results are shown in Table 3 below.

[0092] Table 3 Results of Comparative Examples 1-2

[0093]

[0094] As can be seen from the above, the composite protective carrier of the present invention can effectively protect the cell membrane structure of the strain, avoid damage to the strain activity by the fermentation environment, and significantly improve the survival rate of the strain, fermentation efficiency and product quality, which is one of the core technologies of the present invention.

[0095] The compound microbial strain of this invention exhibits rapid proliferation at 35-37.5℃ and pH 4.0-4.2. This temperature range allows the strain concentration to reach the required fermentation threshold within a short time, laying the foundation for subsequent metabolism. Therefore, this invention first conducts a high-temperature fermentation stage at a higher temperature. The high-temperature environment inhibits the growth of some low-temperature contaminants, reducing the risk of fermentation contamination. Simultaneously, Bacillus subtilis secretes cellulase and protease more readily at high temperatures, rapidly degrading macromolecules in the raw materials and providing carbon and nitrogen sources for strain proliferation, avoiding fermentation stagnation caused by nutrient limitations. Furthermore, the compound microbial strain exhibits high esterase activity at 20-23.5℃ and pH 3.6-3.8, efficiently catalyzing the combination of alcohols and organic acids to generate ester flavor compounds. The low-temperature environment also reduces the formation of high-boiling-point off-flavors, resulting in a purer wine. Therefore, this invention then conducts a second-stage low-temperature fermentation at a lower temperature. Low-temperature fermentation reduces alcohol evaporation loss and prolongs the accumulation time of flavor compounds, allowing alcohols, esters, aldehydes, and other substances to reach a balanced ratio. This avoids the aroma imbalance caused by the rapid formation of flavor compounds at high temperatures, ultimately resulting in a fuller, more harmonious aroma and a mellow, unblemished taste. Simultaneously, the pH value is used to monitor the degree of fermentation.

[0096] Comparative Examples 3-5

[0097] Fermentation processes with five different strains (including one combination of the present invention, three comparative combinations, and one traditional process) were compared, and the process performance was evaluated based on four core indicators: total types of volatile flavor compounds, total ester content, raw material utilization rate, and fermentation cycle. The results are shown in Table 4.

[0098] Table 4 Comparison of Fermentation Process Performance of Compound Microbial Strains

[0099]

[0100] Table 4 shows that, based on the total number of volatile flavor compounds, volatile flavor compounds are the core indicator determining the aroma richness and taste complexity of fermented products. Experimental data indicates that the combined strain of *Rhodotorula glutinis*, *Bacillus subtilis*, and *Aspergillus niger* used in Example 1 produced 28 volatile flavor compounds, showing a significant advantage over other experimental groups: a 55.56% increase compared to Comparative Example 3 (18 compounds) inoculated only with *Rhodotorula glutinis*, a 27.27% increase compared to Comparative Example 4 (22 compounds) inoculated with *Rhodotorula glutinis* and *Bacillus subtilis*, and a 33.33% increase compared to Comparative Example 5 (21 compounds) inoculated with *Rhodotorula glutinis* and *Aspergillus niger*. This demonstrates that the combination of these three strains creates a synergistic metabolic effect, with each strain producing different types of flavor compounds, significantly enhancing the aroma richness and uniqueness of the fermented product.

[0101] Analysis of total ester content: Total esters are an important component of flavor compounds in fermented products, and their content directly affects the richness of the taste and the intensity of the aroma. The total ester content in Example 1 reached 2.6 g / L, the highest among all experimental groups: an increase of 116.67% compared to Comparative Example 3 (1.2 g / L), an increase of 52.94% compared to Comparative Example 4 (1.7 g / L), and an increase of 36.84% compared to Comparative Example 5 (1.9 g / L). This indicates that the compound microbial system can synthesize esters more efficiently during metabolism, effectively enhancing the flavor quality of the product and improving the consumer's sensory experience.

[0102] Analysis of raw material utilization rate: Raw material utilization rate is a key indicator for measuring the economic efficiency of fermentation processes, directly related to cost control and resource waste during production. Example 1 achieved a raw material utilization rate as high as 90%, significantly higher than all other experimental groups: a 25% increase compared to Comparative Example 3 (72%), an 11.11% increase compared to Comparative Example 4 (81%), and a 20% increase compared to Comparative Example 5 (75%), while also showing increases of 25% and 28.57% respectively compared to the traditional process (70%). This indicates that the synergistic effect of the three microbial strains can more fully decompose nutrients such as carbohydrates and proteins in the raw materials, resulting in a higher efficiency in converting raw materials into the target product, significantly reducing raw material costs in the production process, and improving the economic feasibility of the process.

[0103] Fermentation cycle analysis: The fermentation cycle directly affects production efficiency and capacity. A shorter fermentation cycle can significantly improve production turnover and reduce equipment occupancy costs. The fermentation cycle of Example 1 was only 12 days, the shortest among all experimental groups: 33.33% shorter than Comparative Example 3 (18 days), 20% shorter than Comparative Example 4 (15 days), 25% shorter than Comparative Example 5 (16 days), and 60% shorter than the traditional process (30 days). This indicates that the compound microbial system has stronger metabolic activity, can quickly start and complete the fermentation process, greatly improves production efficiency, and is suitable for the needs of large-scale industrial production.

[0104] Comparative analysis of multiple core indicators across the experimental groups reveals that the compound microbial strain combination of *Rhodotorula glutinis*, *Bacillus subtilis*, and *Aspergillus niger* used in this invention exhibits significant advantages in key indicators such as the richness of volatile flavor compounds, total ester content, raw material utilization rate, and fermentation cycle, far surpassing single-strain, two-strain combinations, and traditional fermentation processes. This compound microbial system not only enhances the flavor and quality of fermented products but also effectively reduces production costs and increases production efficiency, possessing extremely high industrial application value and market competitiveness.

Claims

1. A method for brewing baijiu (Chinese liquor) based on multi-strain synergistic fermentation, characterized in that: Includes the following steps: S1. Raw material pretreatment: After the grain is crushed and sieved, the resulting brewing raw material is mixed evenly with water and cooked at a constant temperature. After cooking, it is cooled and sterilized to obtain a sterile fermentation substrate. S2. Construction of multi-strain systems and vector loading: S201. Using Rhodotorula mucilaginosa (CGMCC 2.167) as the core fermentation strain, Bacillus subtilis (CGMCC 1.1086) as the auxiliary control strain, and Aspergillus niger (CGMCC 3.795) as the flavoring strain, Rhodotorula mucilaginosa, Bacillus subtilis, and Aspergillus niger were activated separately to obtain Rhodotorula mucilaginosa culture, Bacillus subtilis culture, and Aspergillus niger culture. S202. Trehalose, chitosan and sodium carboxymethyl cellulose are mixed in a mass ratio of 2~4.5:1.5~3:0.5~2, and water is added to prepare a mixed solution with a mass concentration of 4~6%. The mixed solution is then mixed with porous starch microspheres in a mass ratio of 1:3~5. After adsorption at 2~4℃, the mixture is freeze-dried to obtain a protective carrier. S203. Mix the three bacterial cultures with the protective carrier at a mass ratio of 1:4~6, shake to adsorb, and then mix the three loaded bacterial cultures at a mass ratio of 6:1~4:1~4 to obtain a loaded composite bacterial culture. S3. Co-fermentation: The loaded compound microbial strain is inoculated into the aseptic fermentation substrate at 0.5-1.5% of the substrate mass, and then sealed for staged fermentation: Stage 1: constant temperature fermentation at 35-37.5℃ for 3-4 days; Stage 2: after the first stage, the temperature is lowered to 20-23.5℃ for 7-10 days; the total fermentation time for the first and second stages is 11-13 days. During fermentation, on the third day of fermentation, 0.1-1.5% of glucose by weight of the brewing raw materials is added; on the fifth day of fermentation, 0.2-0.4% of a compound enzyme preparation by weight of the brewing raw materials is added. The compound enzyme preparation is a mixture of cellulase and hemicellulase in a mass ratio of 12-14:1-3, or a mixture of cellulase, hemicellulase and pectinase in a mass ratio of 12-14:1-3:2-4; on the seventh day, 0.03-0.07% of yeast extract by weight of the brewing raw materials is added. During fermentation, the pH was maintained at 4.0-4.2 in the first stage and 3.6-3.8 in the second stage. The dissolved oxygen content of the fermentation system was kept at 0.5-1.0 mg / L. Fermentation was terminated when the residual sugar content was ≤0.5 g / L and the pH value was stable. S4. Post-processing: After fermentation, the liquor is filtered, distilled and decolorized to obtain the finished liquor.

2. The method for brewing Baijiu (Chinese liquor) based on multi-strain synergistic fermentation according to claim 1, characterized in that: In step S1, at least one of the following must be satisfied: The grains in question are sorghum, wheat, and corn in a mass ratio of 4-6:2.5-3.5:1.5-2.

5. The sieving process involves passing the material through a 40-mesh standard sieve and collecting the material that passes through the sieve. The mass ratio of brewing raw materials to water is 1:1.0~1.5; The constant temperature steaming is performed at 100~110℃ for 20~40 minutes. The cooling method is air cooling, with a cooling rate of ≥5℃ / min, cooling to 30~38℃; The sterilization process involves steam sterilization at 110-130℃ for 20-30 minutes.

3. The method for brewing Baijiu (Chinese liquor) based on multi-strain synergistic fermentation according to claim 1, characterized in that: In step S201, at least one of the following conditions must be met: The activation procedure for Rhodotorula glutinis is as follows: Rhodotorula glutinis is activated using YM medium at a constant temperature of 24-30℃ for 25-30 minutes, until the bacterial concentration is ≥1×10⁻⁶. 8 CFU / mL was used to obtain Rhodotorula glutinis culture; The activation procedure for Bacillus subtilis is as follows: Bacillus subtilis is activated on LB medium at 30–38°C for 18–22 minutes until the bacterial concentration is ≥1×10⁻⁶. 9 CFU / mL was used to obtain Bacillus subtilis bacterial suspension; The activation procedure for Aspergillus niger is as follows: Incubate Aspergillus niger on PDA medium at 24-33℃ for 64-72 hours until the bacterial concentration is ≥1×10⁻⁶. 9 CFU / mL was used to obtain Aspergillus niger solution.

4. The method for brewing Baijiu (Chinese liquor) based on multi-strain synergistic fermentation according to claim 1, characterized in that: In step S202, at least one of the following conditions must be met: The porous starch microspheres have a particle size of 5 nm to 100 μm; preferably 50 to 80 μm or 10 to 30 nm. The adsorption time is 2-3 hours.

5. The method for brewing Baijiu (Chinese liquor) based on multi-strain synergistic fermentation according to claim 1, characterized in that: In step S203, the oscillation adsorption rate is 120~150 r / min, the time is 1~2 h, and the bacterial adsorption efficiency is ≥92%.

6. The method for brewing Baijiu (Chinese liquor) based on multi-strain synergistic fermentation according to claim 1, characterized in that: In step S3, during the first stage, the stirring rate is controlled at 80-100 r / min for the first two days, and then reduced to 55-65 r / min for the next one to two days. During the second stage, the stirring rate is controlled at 40-50 r / min, and an intermittent bidirectional stirring mode is adopted.

7. The method for brewing Baijiu (Chinese liquor) based on multi-strain synergistic fermentation according to claim 1, characterized in that: In step S3, the following operation is added: During the fermentation process, on the 6th day of fermentation, 0.02~0.04% of the mass of the brewing raw materials is added with sodium L-glutamate.

8. The method for brewing baijiu based on multi-strain synergistic fermentation according to claim 1, characterized in that: In step S3, at least one of the following must be satisfied: The pectinase activity is 20,000 to 40,000 U / g; During fermentation, the residual sugar content and pH value were measured every 4 to 8 hours, and the pH value was dynamically adjusted by adding 0.05 to 0.1 mol / L citric acid aqueous solution.

9. The method for brewing baijiu based on multi-strain synergistic fermentation according to claim 1, characterized in that: In step S4, the distillation adopts a three-stage distillation: the first stage is distilled at 65~75℃ for 1~1.5h to remove low-boiling-point impurities; the second stage is distilled at 75~80℃ for 3~5h to obtain the target fraction with an alcohol content of 52~55%vol; and the third stage is distilled at 80~85℃ for 0.5~1h to recover the residual liquid.

10. The method for brewing baijiu based on multi-strain synergistic fermentation according to claim 1, characterized in that: In step S4, at least one of the following must be satisfied: The filtration is pressure filtration, and the filter cloth has a pore size of 5~10μm; The decolorization process involves adding 0.5-1% of the target fraction mass of 100-200 mesh activated carbon to the target fraction, adsorbing and decolorizing at 15-20°C for 24-36 hours, and then filtering through a precision filter.

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