A compound microbial inoculant and fermentation method for preparing pickled Chinese cabbage by soybean water fermentation

Abstract

The application provides a compound microbial inoculum for preparing pickled Chinese cabbage through fermentation of bean water and a fermentation method, and belongs to the technical field of fermented food technology.The compound microbial inoculum for preparing pickled Chinese cabbage through fermentation of bean water comprises the following bacterial species in percentage by mass: 40-60% of Lactobacillus plantarum, 20-35% of Lactobacillus acidophilus, 10-25% of Saccharomyces cerevisiae and 5-15% of Bacillus acidophilus.The pre-processed bean water is used as a fermentation substrate, and the compound microbial inoculum is inoculated to prepare pickled Chinese cabbage through fermentation, so that the fermentation period is shortened, the fermentation quality of the pickled Chinese cabbage is improved, the safety of the pickled Chinese cabbage is ensured, the pickled Chinese cabbage is endowed with special bean aroma, and the resource utilization of the bean water is realized.

Description

Technical Field

[0001] This invention belongs to the field of fermented food technology, and particularly relates to a compound microbial agent and fermentation method for preparing sauerkraut by fermenting soybean water. Background Technology

[0002] Traditional sauerkraut typically uses water or low-concentration brine as the fermentation substrate, relying on lactic acid bacteria in the natural environment for natural fermentation. While this method is inexpensive, it has several significant drawbacks: traditional sauerkraut primarily relies on the vegetables' own components and salt content, lacking exogenous functional active substances, thus limiting its nutritional value. The complex microbial community during natural fermentation makes it susceptible to contamination by other microorganisms, leading to large fluctuations in nitrite content, a long fermentation cycle, and poor flavor stability.

[0003] In the soybean deep processing industry, soybean water, as a byproduct of the tofu forming process, has an annual output of millions of tons. Soybean water is rich in active ingredients such as soybean oligosaccharides, small molecule peptides, and isoflavone glycosides; however, it is currently mainly used for high-COD wastewater treatment, resulting in extremely low resource utilization and causing serious environmental burden and economic losses. In recent years, there have been no reports of using soybean water directly as a fermentation substrate for sauerkraut preparation. Furthermore, existing sauerkraut fermentation technologies focus primarily on improving microbial agents (such as shortening the fermentation cycle or reducing nitrite levels), neglecting to enhance the functionality of sauerkraut through substrate innovation.

[0004] Therefore, developing a highly efficient fermentation process based on soybean water to fully utilize its functional components and simultaneously realize the resource utilization of soybean water and the upgrading of sauerkraut quality has become a technical challenge that the industry urgently needs to overcome. Summary of the Invention

[0005] In view of this, the purpose of this invention is to provide a compound microbial agent and fermentation method for preparing sauerkraut by fermenting soybean water, so as to realize the resource utilization of soybean water and improve the fermentation quality of sauerkraut.

[0006] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0007] A compound microbial agent for fermenting soybean water to prepare sauerkraut comprises the following microbial strains in weight percentage: 40-60% Lactobacillus plantarum, 20-35% Lactobacillus acidophilus, 10-25% Saccharomyces cerevisiae, and 5-15% Bacillus acidophilus.

[0008] Preferably, the preparation method includes the following steps: (1) liquid culture of Lactobacillus plantarum, Lactobacillus acidophilus, Saccharomyces cerevisiae, and Bacillus acidophilus respectively until the viable count is ≥1×10⁻⁶. 9 CFU / mL; (2) Mix the bacterial solution obtained in step (1) in proportion and add a freeze-drying protectant; (3) Freeze-dry the mixture to make a powder, vacuum package it and store it at 4-10℃.

[0009] The present invention also provides the application of the compound microbial agent in the preparation of sauerkraut.

[0010] The present invention also provides a method for preparing sauerkraut by fermenting soybean water, comprising the following steps: (1) sterilizing soybean water, adjusting the pH to 5.0-6.5, and the salt concentration to 2-3%; the soybean water is the waste liquid after coagulation and molding in the tofu production process; (2) inoculating the soybean water with the compound bacterial agent and activating and culturing for 1-2 hours; (3) mixing the pre-pickled and dehydrated vegetables with the soybean water in step (2) and fermenting to prepare sauerkraut.

[0011] Preferably, the temperature for sterilizing the soybean water is 80-90℃ and the time is 10-30 minutes; the pH is adjusted using food-grade sodium bicarbonate or citric acid.

[0012] Preferably, the inoculation amount of the compound microbial agent is 0.1-0.5% of the weight of the soybean water.

[0013] Preferably, the activation culture temperature is 25-30℃.

[0014] Preferably, the vegetables and bean water are mixed at a mass ratio of 1:1-2, and the vegetables are compacted until they are completely submerged.

[0015] Preferably, the fermentation is carried out in stages with controlled temperature; fermentation at 20-28℃ for 3-5 days, fermentation at 15-20℃ for 5-15 days, until the pH is ≤4.2.

[0016] The present invention also provides sauerkraut prepared by the fermentation method.

[0017] Compared with the prior art, the present invention has the following beneficial effects:

[0018] The compound microbial agent of this invention has synergistic functions, which can specifically improve fermentation efficiency and quality. By optimizing the ratio of microbial strains, this invention achieves a synergistic effect of acid production, flavor substance generation, and macromolecular degradation, significantly shortening the fermentation cycle and ensuring that the final pH of fermentation is ≤4.2, thus inhibiting contamination by other microorganisms.

[0019] This invention enables the resource utilization of soybean water and imparts functional components to sauerkraut. It transforms soybean water from tofu production into a fermentation substrate, replacing traditional brine, thus reducing wastewater discharge and lowering raw material costs. The soybean water is rich in soybean oligosaccharides (such as stachyose), small molecule peptides, isoflavone glycosides, and other active ingredients. After fermentation, it significantly enhances the nutritional value and functionality of sauerkraut and imparts a unique soybean aroma. Detailed Implementation

[0020] This invention provides a compound microbial agent for preparing sauerkraut through fermentation of soybean water, comprising the following microbial strains in the following weight percentages: 40-60% *Lactobacillus plantarum*, 20-35% *Lactobacillus acidophilus*, 10-25% *Saccharomyces cerevisiae*, and 5-15% *Bacillus acidophilus*; more preferably, 45-55% *Lactobacillus plantarum*, 22-30% *Lactobacillus acidophilus*, 12-20% *Saccharomyces cerevisiae*, and 8-12% *Bacillus acidophilus*; even more preferably, 50% *Lactobacillus plantarum*, 25% *Lactobacillus acidophilus*, 15% *Saccharomyces cerevisiae*, and 10% *Bacillus acidophilus*. This invention does not specifically limit the source of the microbial strains; commonly used strains in this technical field can be selected. As one possible implementation method, the *Lactobacillus plantarum* of this invention is designated CGMCC No. 31144; *Lactobacillus acidophilus* is designated CICC 6093; *Saccharomyces cerevisiae* is designated CICC 1406; and *Bacillus sp.* is designated Bacillus sicca CD92-3.

[0021] In the compound microbial agent of this invention, *Lactobacillus plantarum* utilizes the soluble sugars in soybean water to rapidly generate lactic acid in the early stages of fermentation, lowering the pH of the fermentation environment, inhibiting the proliferation of putrefactive bacteria, and also metabolizing volatile substances such as ethyl acetate and diacetyl, establishing the "fresh and fragrant" base of the sauerkraut. *Lactobacillus acidophilus* continuously produces acid in the middle and later stages of fermentation, further lowering the pH of the fermentation environment, completely inhibiting other bacteria, and simultaneously degrading anti-nutritional factors such as phytic acid and tannins, improving the nutrient utilization rate of the soybean water. Its secreted extracellular polysaccharides also enhance the viscoelasticity of the sauerkraut, alleviating the problem of excessive softening. *Saccharomyces cerevisiae* can generate higher alcohols and esters, giving the sauerkraut layers of "fruity" and "wine" aromas, and can also consume residual oxygen, promoting anaerobic fermentation dominated by lactic acid bacteria. Its autolysis releases amino acids, enhancing the umami flavor. Bacillus acidophilus can secrete proteases and cellulases, which, under acidic conditions, break down soybean globulin and cellulose in soybean water into small peptides and soluble sugars, providing rapid nutrition for lactic acid bacteria and yeast. It can also directionally degrade intercellular matrix by secreting pectinase, maintaining the crisp texture of the vegetables. This invention's compound microbial agent, through complementary functions (acidification, flavor enhancement, and degradation), achieves highly efficient and targeted fermentation of soybean water, combining quality improvement with safety and controllability.

[0022] In this invention, the preferred method for preparing the above-mentioned compound microbial agent includes the following steps:

[0023] (1) Liquid culture of *Lactobacillus plantarum*, *Lactobacillus acidophilus*, *Saccharomyces cerevisiae*, and *Bacillus acidophilus* was carried out to a viable count ≥ 1 × 10⁻⁶. 9CFU / mL. This invention does not impose specific limitations on the propagation methods for each strain; commonly used propagation methods in this technical field are acceptable. As one possible implementation method, the propagation method for *Lactobacillus plantarum* CGMCC No. 31144 refers to the information in CN202411131656.7; the propagation methods for *Lactobacillus acidophilus* CICC 6093 and *Saccharomyces cerevisiae* CICC 1406 refer to the information provided on the official website of the China Industrial Microbial Culture Collection Center; and the propagation method for *Bacillus acidophilus* CD92-3 refers to the information in "Screening and Identification of High-Protein-Producing Bacillus in the Fermentation Process of Shanxi Aged Vinegar".

[0024] (2) Mix the bacterial solution obtained in step (1) in a certain proportion and add a freeze-drying protectant. The freeze-drying protectant preferably includes 15% skim milk powder and 8% trehalose, and more preferably, the volume ratio of the freeze-drying protectant to the bacterial solution is 1:1. As one possible implementation, the skim milk powder and trehalose are dissolved in sterile physiological saline at their respective concentrations, mixed with the bacterial solution at a 1:1 (v / v) ratio, and allowed to stand at 25°C for 30 minutes to equilibrate.

[0025] (3) The mixture is freeze-dried to form a powder, vacuum-packed, and stored at 4-10℃. The freeze-drying conditions are further preferably -40℃ for 2 hours, -25℃ for 12 hours, and 25℃ for 6 hours, more preferably dried until the moisture content of the bacterial agent is ≤3%. The pulverization is further preferably performed using low-temperature airflow pulverization to a powder particle size of 80-100 mesh.

[0026] This invention provides the application of the above-mentioned compound microbial agent in the preparation of sauerkraut.

[0027] This invention also provides a method for preparing sauerkraut using soybean water fermentation, comprising the following steps:

[0028] (1) Sterilize the soybean water, adjust the pH to 5.0-6.5, and the salt concentration to 2-3%. The soybean water mentioned in this invention is the waste liquid after coagulation and molding in the tofu production process. As a by-product of tofu production, soybean water still contains small molecule peptides, free amino acids, oligosaccharides, potassium, magnesium, B vitamins, and other components, and has high nutritional value.

[0029] The preferred sterilization temperature for soybean whey in this invention is 80-90℃, more preferably 82-88℃, and even more preferably 85℃; the preferred sterilization time is 10-30 min, more preferably 15-25 min, and even more preferably 20 min. The pH is preferably adjusted using food-grade sodium bicarbonate or citric acid, and even more preferably to a pH of 6.0. The salt concentration is preferably adjusted using food-grade NaCl or diluted with water, and even more preferably to a salt concentration of 2.5%. High-temperature sterilization preserves nutrients, completely inactivating endogenous lipoxygenases and ureases in soybean whey, preventing off-flavors from subsequent fermentation. Since the initial pH of the soybean whey is unsuitable for fermentation (gypsum waste liquid is nearly neutral, while brine and GDL waste liquid are slightly acidic), food-grade sodium bicarbonate or citric acid is used to neutralize it to pH 6.0, simulating the slightly acidic environment of the initial stage of natural fermentation, promoting lactic acid bacteria to initiate glycolysis while inhibiting the proliferation of putrefactive bacteria. A suitable salt concentration, in addition to inhibiting Gram-negative bacteria, also provides additional benefits. + It also binds with pectin in the cell walls of vegetables, maintaining the crispness of the sauerkraut.

[0030] (2) Inoculate the above-mentioned compound microbial agent into soybean water and activate it for 1-2 hours, more preferably 1.5 hours. The preferred inoculation amount of the compound microbial agent is 0.1-0.5% of the mass of soybean water, more preferably 0.2-0.4%, and even more preferably 0.3%. The preferred activation culture temperature is 25-30℃, more preferably 28℃.

[0031] In this invention, glucose is preferably added simultaneously as an additional carbon source when inoculating the bacterial agent. More preferably, the amount of glucose added is 0.5-1.5% of the weight of the soybean water, and more preferably 1%, to facilitate fermentation.

[0032] (3) Mix the pre-pickled and dehydrated vegetables with the soybean water from step (2) and ferment to prepare sauerkraut.

[0033] The preferred pre-pickled dehydration method of this invention includes pickling in an 8-12% NaCl solution at room temperature for 8-12 hours, more preferably in a 10% NaCl solution for 10 hours; preferably after pickling, the pressure is maintained at 0.1 MPa for 1 hour to drain free water, and then maintained at 0.3 MPa for 0.5 hours to drain bound water, until the vegetable moisture content is 60-65% and the texture is firm. This invention does not have specific limitations on the source of the vegetables, including but not limited to leafy vegetables such as Chinese cabbage, mustard greens, and kale.

[0034] The present invention preferably uses a mixture of vegetables and soybean water at a mass ratio of 1:1-2, more preferably a mass ratio of 1:1.5, and compacts the vegetables until they are completely submerged. The fermentation is preferably carried out in stages with controlled temperature: fermenting at 20-28℃ for 3-5 days, fermenting at 15-20℃ for 5-15 days, more preferably fermenting at 25℃ for 3-5 days, and fermenting at 18℃ for 5-15 days, until the pH reaches ≤4.2.

[0035] This invention employs segmented temperature-controlled fermentation. The first stage, fermentation at 20-28℃, accelerates the glycolysis metabolism of the inoculant, rapidly consuming soluble sugars such as glucose and sucrose in the soybean water to generate lactic acid. This causes the pH of the fermentation system to rapidly drop from 5.0-6.5 to below 4.5 within 3-5 days, creating a highly acidic environment that inhibits the proliferation of Gram-negative bacteria such as Escherichia coli and Salmonella. At this temperature, Bacillus acidophilus exhibits enhanced activity, and its secreted proteases rapidly decompose soybean globulin in the soybean water into small peptides, providing a nitrogen source for lactic acid bacteria. Simultaneously, the generated soluble sugars can be utilized by Saccharomyces cerevisiae, promoting its proliferation and consuming oxygen in the system, creating an anaerobic environment that further inhibits aerobic putrefactive bacteria. The high temperature accelerates acid production by the dominant lactic acid bacteria, causing the fermentation environment to enter a low pH state earlier during the peak of nitrite formation, inhibiting the enzyme activity of nitrite-reducing bacteria and reducing nitrite formation. The second stage of fermentation at 15-20℃ slows down the acid production rate of lactic acid bacteria, promotes the metabolism of brewer's yeast to produce higher alcohols and esters, and imparts aroma to the sauerkraut. In a low-acid environment, *Lactobacillus acidophilus* secretes extracellular polysaccharides (such as glucan and galactan), enhancing the viscoelasticity of the sauerkraut and preventing excessive softening. The secreted pectinase continues to work during this stage, directionally degrading pectin in the intercellular matrix of the vegetables, maintaining their crisp texture. Low temperatures also inhibit the degradation of small-molecule active ingredients in the soybean water, while slowing down the excessive consumption of amino acids by lactic acid bacteria, increasing the content of free amino acids and enhancing the umami flavor of the sauerkraut. Segmented temperature-controlled fermentation can shorten the fermentation cycle, improve fermentation quality, and reduce energy consumption.

[0036] The present invention also provides a sauerkraut product obtained by the above fermentation method.

[0037] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0038] Example 1

[0039] A compound microbial agent for fermenting soybean water to prepare sauerkraut, the preparation method is as follows:

[0040] (1) Liquid culture was performed on Lactobacillus plantarum, Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus acidophilus respectively until the viable count was ≥1×10⁻⁶. 9 CFU / mL;

[0041] (2) Mix the bacterial solution obtained in step (1) in the following proportions: 50% Lactobacillus plantarum, 25% Lactobacillus acidophilus, 15% Saccharomyces cerevisiae and 10% Bacillus acidophilus.

[0042] (3) Dissolve skim milk powder and trehalose in sterile physiological saline (15% skim milk powder + 8% trehalose), mix with the bacterial solution in step (2) at a ratio of 1:1 (v / v), and let stand at 25°C for 30 minutes to equilibrate.

[0043] (4) Freeze-dry the mixture until the water content is ≤3%, then pulverize it with low-temperature airflow to make 80-mesh powder, vacuum package it and store it at 4-10℃.

[0044] Example 2

[0045] A compound microbial agent for fermenting soybean water to prepare sauerkraut, which differs from Example 1 in that:

[0046] The formula contains 40% Lactobacillus plantarum, 35% Lactobacillus acidophilus, 20% Saccharomyces cerevisiae, and 5% Bacillus acidophilus.

[0047] Example 3

[0048] A compound microbial agent for fermenting soybean water to prepare sauerkraut, which differs from Example 1 in that:

[0049] The formula contains 60% Lactobacillus plantarum, 20% Lactobacillus acidophilus, 15% Saccharomyces cerevisiae, and 5% Bacillus acidophilus.

[0050] Example 4

[0051] A method for preparing sauerkraut using soybean water fermentation, the steps of which are as follows:

[0052] (1) Sterilize the soybean water (waste liquid after coagulation and molding in the tofu production process) at 85℃ for 20 minutes, adjust the pH to 6.0 and the salt concentration to 2.5%;

[0053] (2) Inoculate the compound microbial agent of Example 1 into soybean water at an inoculation rate of 0.3% and activate and culture at 28°C for 1.5 h;

[0054] (3) Using Chinese cabbage as raw material, pickle it in 10% NaCl solution for 10 hours, then squeeze it to dehydrate until the water content of the vegetables is 60%. Mix the vegetables with bean water at a mass ratio of 1:1.5 and press the vegetables until they are completely submerged.

[0055] (4) Ferment in a fermenter at 25℃ for 4 days, at 18℃ for 10 days, until pH ≤ 4.2.

[0056] Example 5

[0057] A method for preparing sauerkraut using soybean water fermentation, the steps of which are as follows:

[0058] (1) Sterilize the soybean water (waste liquid after coagulation and molding in the tofu production process) at 80℃ for 30 minutes, adjust the pH to 5.0 and the salt concentration to 2%;

[0059] (2) Inoculate the compound microbial agent of Example 1 into soybean water at an inoculation amount of 0.1% and activate and culture at 25°C for 2 hours;

[0060] (3) Using mustard greens as raw material, pickle them in 8% NaCl solution for 12 hours, then squeeze them to dehydrate until the water content of the vegetables is 65%. Mix the vegetables with bean water at a mass ratio of 1:1 and press the vegetables until they are completely submerged.

[0061] (4) Ferment in a fermenter at 28℃ for 3 days, then at 15℃ for 15 days, until the pH is ≤4.2.

[0062] Example 6

[0063] A method for preparing sauerkraut using soybean water fermentation, the steps of which are as follows:

[0064] (1) Sterilize the soybean water (waste liquid after coagulation and molding in the tofu production process) at 90℃ for 10 minutes, adjust the pH to 6.5 and the salt concentration to 3%;

[0065] (2) Inoculate the compound microbial agent of Example 1 into soybean water at an inoculation amount of 0.5% and activate and culture at 30°C for 1 hour;

[0066] (3) Using cabbage as raw material, pickle it in 12% NaCl solution for 8 hours, then squeeze it to dehydrate until the water content of the vegetables is 63%. Mix the vegetables with bean water at a mass ratio of 1:2 and press the vegetables until they are completely submerged.

[0067] (4) Ferment in a fermenter, ferment at 20℃ for 5 days, ferment at 15℃ for 12 days, until pH≤4.2.

[0068] Experimental Example 1

[0069] 1. Experimental Design

[0070] (1) Materials and Equipment:

[0071] Sauerkraut ingredients: Fresh Chinese cabbage (variety: Qiuguan), cut into 5×5cm pieces, blanch in boiling water for 2 minutes, then cool and drain.

[0072] Soybean water: taken from a fully automated tofu production line (pH 5.5, COD 22000mg / L, total solids 3.8%).

[0073] Traditional brine: 3% NaCl solution (pH 5.5).

[0074] Bacterial strain: Commercially available Lactobacillus plantarum powder (live count ≥ 1 × 10⁻⁶) 9 CFU / g).

[0075] Equipment: pH meter, HPLC (high performance liquid chromatograph), COD analyzer, spectrophotometer, constant temperature incubator.

[0076] (2) The experimental groups are shown in Table 1.

[0077] Table 1 Fermentation substrate test grouping

[0078]

[0079] Pre-treated soybean water: Sterilize at 80℃ for 20 minutes, adjust pH to 5.5, and salt concentration to 3%.

[0080] (3) Detection indicators and methods:

[0081] pH and nitrite content were monitored in real time on days 0, 3, 7, 10, and 15 of fermentation. After fermentation, the isoflavone aglycone content, COD removal rate, and nitrite content of the fermented sauerkraut were measured. After fermentation, the fermented sauerkraut was collected and blindly evaluated by 10 people based on four indicators: acidity, crispness, and off-flavor.

[0082] The sensory evaluation criteria are as follows:

[0083] Acidity: 1-3 points for bland or no acidity, 4-6 points for moderate and acceptable acidity, 7-10 points for distinct and mild acidity without irritation;

[0084] Crispness: Soft or mushy or with broken fibers 1-3 points, slightly soft but still chewable 4-6 points, crisp and hard texture, no fibrous feel when chewing 7-10 points;

[0085] Odor: No odor 0 points, slight bitter / mold odor 1-3 points, obvious odor but acceptable 4-6 points, severe odor unacceptable 7-10 points; bitterness +1 point / grade, moldy odor +2 points / grade, rotten odor is directly judged as unqualified.

[0086] 2. Test Results:

[0087] (1) Acid production rate (pH change)

[0088] As shown in Table 2, using soybean water as the fermentation substrate for preparing sauerkraut resulted in a significantly faster acid production rate than traditional brine, and the pH was even lower after 15 days.

[0089] Table 2 pH changes in sauerkraut prepared with different fermentation substrates

[0090]

[0091] Note: * indicates that the difference between group E and group C is significant (p < 0.05), the same applies below.

[0092] (2) Retention of functional components and safety

[0093] As shown in Table 3, using soybean water as the fermentation substrate for preparing sauerkraut can retain isoflavone aglycones and significantly reduce COD, while traditional brine has no functional components. The nitrite content in the soybean water group was also significantly lower than that in the brine group.

[0094] Table 3 Functional components and safety of sauerkraut prepared using different fermentation substrates

[0095]

[0096] (3) Sensory rating

[0097] As shown in Table 4, using soybean water as the fermentation substrate for preparing sauerkraut results in a higher sensory evaluation, no unpleasant odors, and a unique soybean aroma.

[0098] Table 4 Sensory scores of sauerkraut prepared with different fermentation substrates

[0099] Grouping acidity crispness Odor Bean flavor Group C scores (1-10) 7.2±0.8 6.5±1.0 2.5±0.5(bitter) none Group E Scores (1-10) 8.5±0.6* 7.8±0.7* 0.5±0.2* have

[0100] In summary, using soybean brine as the fermentation substrate for preparing sauerkraut is superior to traditional brine in terms of acid production rate, retention of functional components, safety, and sensory evaluation.

[0101] Experimental Example 2

[0102] Although Experiment 1 shows that using soybean water as a fermentation substrate for sauerkraut preparation has certain advantages compared to traditional brine, a single Lactobacillus plantarum cannot fully utilize the components of soybean water. Further development of compound bacterial agents and determination of optimal fermentation conditions are needed.

[0103] 1. Experimental Design

[0104] (1) Materials and Equipment

[0105] Soybean water: Pretreatment same as in Experiment 1 (pH 5.5, 3% NaCl, sterilized at 80℃ for 20 min).

[0106] Microbial strains: Lactobacillus plantarum CGMCC NO.31144; Lactobacillus acidophilus CICC 6093; Saccharomyces cerevisiae CICC 1406; Bacillus acidophilus CD92-3.

[0107] Reagents: Glucose (food grade), freeze-drying protectant (trehalose + skim milk).

[0108] Equipment: constant temperature shaker, gas chromatography-mass spectrometry (GC-MS), fully automated amino acid analyzer, high-throughput sequencer.

[0109] (2) The experimental groups are shown in Table 5.

[0110] Table 5. Experimental Grouping of Compound Microbial Agents

[0111]

[0112] Note: The inoculum amount for all groups is 0.2% of the soybean water mass, and the viable count of the inoculum is ≥1×10⁻⁶. 9 CFU / g.

[0113] (3) Detection indicators and methods

[0114] Protease activity: determined by the Folin-phenol method on day 3 of fermentation.

[0115] Oligosaccharide retention rate: After fermentation, the retention rate was determined by LC-MS / MS.

[0116] Ester content: determined by GC-MS after fermentation.

[0117] pH value: pH was monitored throughout the fermentation cycle.

[0118] Energy consumption cost: The electricity consumption for constant temperature and segmented temperature control is recorded by the meter throughout the fermentation cycle.

[0119] 2. Test Results

[0120] (1) Protease activity and functional components

[0121] As shown in Table 6, the compound microbial agent in the E2 / E3 group increased protease activity to over 820 U / g, which means that the compound microbial agent can effectively decompose soybean protein in soybean water into small molecule peptides. The E2 / E3 group significantly retained oligosaccharides, and the supplementation of glucose further promoted yeast ester production.

[0122] Table 6. Functional component conversion efficiency under different fermentation inoculants and fermentation conditions

[0123]

[0124]

[0125] Note: * indicates a significant difference between group E and group C (p < 0.05), ** indicates an extremely significant difference between group E and group C (p < 0.01), and the same applies below.

[0126] (2) Process optimization effect

[0127] As shown in Table 7, compound microbial agents combined with segmented temperature-controlled fermentation can significantly shorten the fermentation cycle and save energy. Supplementing with carbon sources can further shorten the fermentation cycle.

[0128] Table 7. Processing effects of different fermentation agents

[0129] index Group C Group E1 Group E2 Group E3 Fermentation cycle 18.5±1.2 16.0±1.0* 12.3±0.8** 10.5±0.5** Energy consumption (kWh / ton) 85.2±3.5 82.0±3.0 68.5±2.8** 65.3±105**

[0130] Experimental Example 3

[0131] 1. Experimental Design

[0132] (1) Materials and Equipment:

[0133] The pickled cabbage of this invention (Group E) was prepared under the optimal conditions of Experiment Example 2 (Group E3: compound microbial agent + segmented temperature control + 1% glucose).

[0134] Traditional pickled cabbage (Group C): Natural fermentation in 3% brine for 15 days (same as Experiment 1, Group C).

[0135] Commercially available pickled cabbage (Group M): A certain brand of pickled cabbage (ingredients: cabbage, water, salt, lactic acid bacteria).

[0136] Main equipment: Texture analyzer (TA.XT Plus), amino acid analyzer, pathogen detection kit (Salmonella / Staphylococcus aureus).

[0137] (2) Detection indicators and methods:

[0138] Sensory evaluation was the same as in Example 1; the hardness and elasticity of the sauerkraut were determined by texture analyzer; the content of isoflavone aglycones was determined by HPLC; the retention of soybean oligosaccharides was quantitatively determined by LC-MS / MS; the total peptide content was determined by BCA method; the residual nitrite was determined according to GB 5009.33; and pathogenic bacteria were detected according to the GB 4789 series.

[0139] 2. Test Results

[0140] (1) Sensory rating and textural characteristics

[0141] As shown in Table 8, the pickled bean curd in group E has a prominent aroma and no unpleasant odor, and its hardness and elasticity are better than those of groups C and M.

[0142] Table 8 Sensory scores and textural characteristics of different pickled vegetables

[0143]

[0144]

[0145] Note: * indicates a significant difference between group E and group C (p < 0.05).

[0146] (2) Functional components and nutrients

[0147] As shown in Table 9, group E contains isoflavone aglycones and stachyose, and its total peptide content is 4 times that of group C.

[0148] Table 9 Functional components and nutritional information of different pickled vegetables

[0149] index Group C Group M Group E Isoflavone aglycones (mg / 100g) Not detected Not detected 52.3±3.1 Stachyose retention (mg / g) Not detected Not detected 8.4±0.5 Total peptide content (mg / g) 1.2±0.1 1.5±0.2 4.8±0.3*

[0150] (3) Security

[0151] As shown in Table 10, the nitrite content in group E was only 40% of that in group C, and there was no pathogenic bacteria contamination.

[0152] Table 10 Safety of Different Pickled Vegetables

[0153] index Group C Group M Group E Nitrite (mg / kg) 24.3±1.5 18.5±1.2 9.8±0.6* Salmonella detected Positive (1 / 3 of samples) Not detected Not detected Staphylococcus aureus detected Positive (1 / 3 of samples) Not detected Not detected

[0154] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A method for preparing isoflavone aglycone sauerkraut using soybean water fermentation, characterized in that, Includes the following steps: (1) Sterilize the soybean water, adjust the pH to 5.0-6.5 and the salt concentration to 2-3%; the soybean water is the waste liquid after coagulation and molding in the tofu production process; (2) Inoculate the soybean water with a compound microbial agent at a rate of 0.2% of the soybean water mass and activate the culture for 1-2 hours. The compound microbial agent consists of the following microbial strains in the following mass percentages: 50% Lactobacillus plantarum CGMCC NO. 31144, 25% Lactobacillus acidophilus CICC6093, 15% Saccharomyces cerevisiae CICC 1406 and 10% Bacillus sicca CD92-3. Glucose is added simultaneously as an additional carbon source during inoculation, at a rate of 1% of the soybean water mass. (3) Mix the pre-pickled and dehydrated vegetables with the soybean water from step (2) and ferment to prepare isoflavone aglycone sauerkraut; the fermentation adopts segmented temperature-controlled fermentation; ferment at 20-28℃ for 3-5 days, ferment at 15-20℃ for 5-15 days, and ferment until pH≤4.

2.

2. The method according to claim 1, characterized in that, The preparation method of the compound microbial agent includes the following steps: (1) Lactobacillus plantarum CGMCC NO. 31144, Lactobacillus acidophilus CICC 6093, Saccharomyces cerevisiae CICC 1406, and Bacillus sicca CD92-3 were cultured in liquid to a viable count ≥1×10⁻⁶. 9 CFU / mL; (2) Mix the bacterial solution obtained in step (1) in proportion and add freeze-drying protectant; (3) Freeze-dry the mixture to make powder, vacuum package it and store it at 4-10℃.

3. The method according to claim 1, characterized in that, The temperature for sterilizing the soybean water is 80-90℃, and the time is 10-30 minutes; the pH is adjusted using food-grade sodium bicarbonate or citric acid.

4. The method according to claim 1, characterized in that, The activation culture temperature is 25-30℃.

5. The method according to claim 1, characterized in that, The vegetables and bean water are mixed at a mass ratio of 1:1-2, and the vegetables are compacted until they are completely submerged.

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