Compound microbial agent and application thereof in production of amaranth stinking bean curd brine

Abstract

The application belongs to the technical field of microbial fermentation, and specifically discloses a kind of compound microbial inoculant and its application in the production of amaranth stinky bean curd brine. The inoculant is compounded by Paracaseolyticum, Fermented Lactobacillus Mucosus, Short Fermenting Agent Lactobacillus, Enterococcus Faecalis and Saccharomyces Cerevisiae. During production, the inoculant is inoculated into amaranth brine raw materials at an inoculation amount of 0.5%-1%, and then fermented at 25-30℃ for 30-45 days. The flavor index is regularly monitored and stirred. This process can replace traditional natural fermentation, solve the problems of large batch difference and high contamination rate of miscellaneous bacteria, and realize standardized production. The key flavor substance content of the brine fermented by the inoculant is significantly improved: indole 337.7 μg / L, 3-methylindole 93.2 μg / L, phenol 96.9 μg / L, and 4-ethylphenol 194.9 μg / L. The sensory score of amaranth stinky bean curd produced by the brine reaches 48 points (out of 50 points), and the flavor and process controllability advantage are significant.

Description

Technical Field

[0001] This invention relates to the field of microbial fermentation technology, and in particular to a compound microbial agent and its application in the production of amaranth-flavored stinky tofu brine. Background Technology

[0002] Amaranth stinky tofu is a traditional Chinese fermented soy product. Its unique flavor comes from the fermented brine of amaranth. Its traditional process relies on natural microorganisms, and it is made by long-term open-air fermentation of raw materials and environmental microorganisms at room temperature. The fermentation cycle ranges from several weeks to several years. It is generally believed in the industry that the longer the fermentation time, the richer the flavor and the better the quality of the finished product.

[0003] During natural fermentation, the proteins in the raw materials undergo a series of complex biochemical reactions under the action of enzymes secreted by microorganisms such as lactic acid bacteria and yeast, decomposing into umami amino acids such as glutamic acid and small molecule polypeptides that give the finished product its umami flavor. Some proteins further decompose to produce marker substances such as indole, 3-methylindole, phenol and 4-ethylphenol, ultimately forming a unique sensory characteristic of "smelling bad but tasting good" composed of various flavor substances such as amino acids, alcohols and esters.

[0004] However, the traditional natural fermentation of amaranth stinky tofu brine still has multiple defects and safety hazards: First, the open fermentation microbial community structure is greatly affected by the environment, which can easily lead to the growth of pathogenic bacteria, resulting in large batch quality differences and food safety risks; Second, fermentation relies on experience, and the parameters lack quantitative standards, making it difficult to standardize and scale up production; Third, the microbial community is uncontrollable, which can easily cause acute poisoning, and long-term consumption also poses chronic health risks.

[0005] Currently, there are patented technologies for preparing stinky tofu brine, but these methods all have significant drawbacks and cannot completely overcome the shortcomings of traditional processes. CN114009687A discloses a method for preparing stinky tofu brine, but only mentions the use of bacilli and cocci for fermentation, without specifying the specific strains or providing a microbial preservation number. The lack of core technical information makes this technology lack guidance and cannot be truly applied or reproduced. CN107594460A discloses a method for preparing stinky tofu brine. Although it introduces a fermentation step, its fermenting agent is only a general commercial brewing yeast (such as Angel Yeast), lacking a multi-strain (especially lactic acid bacteria) synergistic system specifically required to form the unique flavor of amaranth stinky tofu. Therefore, it cannot effectively construct the specific flavor profile required for amaranth stinky tofu and ensure quality. CN105613784A discloses a steady-state rapid fermentation method for fermentation liquid of brine for stinky tofu in southern China. The formula introduces animal raw materials such as cuttlefish or snail meat. However, these raw materials are prone to spoilage and deterioration under long fermentation cycles and relatively high fermentation temperatures, resulting in the generation of undesirable metabolites, contamination of the brine, and introduction of new safety risks. As a result, the stability of the fermentation process still has fundamental defects, and new process defects and safety hazards have been brought about.

[0006] In summary, existing technologies suffer from core problems such as missing key information, improper use of fermentation agents, or flawed process design. To address these shortcomings, this invention provides a compound microbial agent with clearly defined functions (composed of specific strains) and a well-defined synergistic ratio (with a definite live bacteria ratio) and its precise and controllable fermentation process, aiming to fundamentally achieve stable, safe, and standardized production of amaranth-flavored stinky tofu brine. Summary of the Invention

[0007] In view of the above-mentioned defects in the existing technology and the lack of dedicated functional microbial agents, the present invention provides a standardized compound microbial agent that can achieve stable and controllable fermentation, and provides its application in the production of amaranth stinky tofu brine.

[0008] To achieve the above objectives, the technical solution adopted by the present invention is as follows: A compound microbial agent for producing brine for amaranth-flavored stinky tofu is composed of Lactobacillus casei LKG-H01, Lactobacillus fermentum LKG-H02, Lactobacillus short starter LKG-B01, Enterococcus faecalis LKG-B02, and Saccharomyces cerevisiae. The Lactobacillus casei LKG-H01 ( Lactobacillus paracasei (), deposited at the China General Microbiological Culture Collection Center, China, on March 16, 2026, with accession number CGMCC NO: 37914; The fermented Lactobacillus mucinus LKG-H02 ( Limosilactobacillus fermentum (), deposited at the China General Microbiological Culture Collection Center, China, on March 16, 2026, with accession number CGMCC NO: 37915; The short fermentation agent Lactobacillus LKG-B01 ( Levilactobacillus brevis (), deposited at the China General Microbiological Culture Collection Center, China, on March 16, 2026, with the biological accession number CGMCCNO: 37911; The Enterococcus faecalis LKG-B02 ( Enterococcus faecium (), deposited at the China General Microbiological Culture Collection Center, China, on March 16, 2026, with accession number CGMCC NO: 37912; The brewing yeast is a commercially available brewing yeast. Preferably, it can be the rice wine-specific yeast from Angel Yeast Co., Ltd.

[0009] Furthermore, the live bacteria ratio of *Lactobacillus casei*, *Lactobacillus fermentum*, *Lactobacillus short starter*, *Enterococcus faecalis*, and *Saccharomyces cerevisiae* is 5~15:3~10:3~10:10~25:15~30. More preferably, the live bacteria ratio is 8:5:5:15:20.

[0010] The compound microbial agent described in this invention can be used to produce brine for amaranth and stinky tofu. Specifically, the compound microbial agent is added to the raw material of amaranth and stinky tofu brine for fermentation. The fermentation termination conditions are: the content of indole in the brine is ≥200μg / L, 3-methylindole is ≥70μg / L, phenol is ≥70μg / L and 4-ethylphenol is ≥100μg / L.

[0011] Furthermore, the compound microbial agent is added to the raw materials at a rate of 0.1~0.3 kg / 100 kg of raw materials, and the initial total viable count after inoculation is 7.5. 10 5 ~1.5 10 6 CFU / g raw material.

[0012] Furthermore, the compound microbial agent is added at a rate of 0.2 kg / 100 kg of raw material, and the initial total viable count is 1.1. 10 6 CFU / g raw material.

[0013] The preparation method of amaranth-flavored stinky tofu brine includes the following steps: (1) Raw material processing: sort, wash and drain the old amaranth stalks until the moisture content is ≤70% and then cut them into sections. Select the sections with a stem diameter ≥1cm for later use; process the winter bamboo shoots and cut them into chunks / slices; wash and chop other vegetables (such as snow cabbage and alfalfa) and prepare the spice packet.

[0014] (2) Salting: Add 3-5 kg ​​of salt to every 100 kg of fresh raw materials, sprinkle them into the fermentation tank in layers and mix them evenly; the preferred amount of salt added is 4 kg.

[0015] (3) Inoculation procedure: Add 0.1-0.3 kg of the above compound microbial agent to every 100 kg of raw material, stir evenly, and then add water to completely submerge the raw material, with the liquid level 5 cm above the raw material; the preferred amount of agent added is 0.2 kg.

[0016] (4) Segmented fermentation: First, ferment in a sealed container at 25-30℃ for 6-7 days, stirring once a day. Then, let it stand for 7-9 days, controlling the pH of the system to 4.5-5.5. After 14-16 days of fermentation, cool down to 15-20℃ and continue fermentation until the total fermentation cycle is 30-45 days. Monitor the salinity daily throughout the fermentation process and maintain it at 3%-5%. Regularly test the content of indole, 3-methylindole, phenol and 4-ethylphenol in the brine. When the content of indole ≥200μg / L, 3-methylindole ≥70μg / L, phenol ≥70μg / L, and 4-ethylphenol ≥100μg / L, and the brine and amaranth stalks meet the maturity standards, the fermentation is complete. The preferred fermentation conditions are: sealed fermentation at 30℃ for 6-7 days, standing for 8-9 days, cooling down to 20℃ to continue fermentation, and maintaining the salinity at 4% throughout the process.

[0017] (5) Filtration and juice extraction: The fermentation product is filtered using a 30-mesh filter to separate and remove vegetable residue and crude brine.

[0018] (6) Sterilization and filling: Sterilize the crude brine at 60-75℃ for 20-30 minutes, then seal and fill it in a clean workshop and cool it to room temperature; the preferred sterilization conditions are 70℃ for 25 minutes.

[0019] (7) Storage of finished product: Store the bottled brine in a cool and dry place at 10-15℃ for 6-12 months; after opening, store in a refrigerator at 0-4℃ and use within 1 month.

[0020] The present invention also provides amaranth stinky tofu brine prepared from the above-mentioned compound microbial agent.

[0021] Compared with the prior art, the outstanding effect of the present invention is as follows: (1) This invention solves the inherent problems of traditional natural fermentation by inoculating a combination of strains with specific functions, thereby achieving standardization, controllability and efficiency in the production of amaranth stinky tofu brine.

[0022] (2) Improved food safety: The five functional strains selected can quickly form a dominant bacterial community, inhibit the growth of harmful bacteria, and reduce the contamination rate; the key indicators of the fermentation process are stable and controllable, avoiding the irregular accumulation of harmful metabolites and significantly improving the safety of the finished product.

[0023] (3) The production process has been optimized: the strain ratio and fermentation parameters are precisely controlled, shortening the traditional 6-12 month fermentation cycle to 30-45 days, and improving efficiency by more than 50%; all key operations are quantified, eliminating the need to rely on experience judgment, breaking through traditional technical barriers, and making it suitable for industrial-scale production.

[0024] (4) Reduce health risks: Targeted generation of signature flavor compounds reduces the excessive production of harmful volatile substances; Salt content is reduced from the traditional 15%-20% to 3%-5%, reducing the risk of cardiovascular disease caused by long-term consumption.

[0025] (5) Improve overall benefits: Standardized fermentation reduces raw material loss and defective products, and lowers production costs; it shortens the fermentation cycle, improves equipment utilization, and further enhances efficiency; it is more adaptable to production sites and the production process is energy-saving and environmentally friendly.

[0026] The following description, in conjunction with the accompanying drawings and specific embodiments, further illustrates the compound microbial agent of the present invention and its application in the production of amaranth-flavored stinky tofu brine.

[0027] Biological Preservation Information

[0028] Lactobacillus casei LKG-H01, classified and named Lactobacillus paracasei It is deposited at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing. The deposit date is March 16, 2026, and the accession number is CGMCC NO: 37914.

[0029] Lactobacillus fermentum LKG-H02, classified and named Limosilactobacillus fermentum It is deposited at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing. The deposit date is March 16, 2026, and the accession number is CGMCC NO: 37915.

[0030] The short-acting starter, Lactobacillus LKG-B01, is classified and named as follows: Levilactobacillus brevis It is deposited at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing. The deposit date is March 16, 2026, and the accession number is CGMCC NO: 37911.

[0031] Enterococcus faecalis LKG-B02, classified and named Enterococcus faecium It is deposited at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing. The deposit date is March 16, 2026, and the accession number is CGMCC NO: 37912. Attached Figure Description

[0032] Figure 1 Sensory evaluation comparison chart of stinky tofu samples made with amaranth brine. Detailed Implementation

[0033] 1. Strains and culture media

[0034] (1) Strain

[0035] The *Lactobacillus casei* LKG-H01, *Lactobacillus fermentum* LKG-H02, *Lactobacillus short starter* LKG-B01, and *Enterococcus faecalis* LKG-B02 involved in this invention are all deposited at the China General Microbiological Culture Collection Center. The brewing yeast is a rice wine-specific yeast purchased commercially from Angel Yeast Co., Ltd.

[0036] (2) Culture medium formulation

[0037] Bacterial culture medium formula: beef extract 3g / L, peptone 5g / L, glucose 2.5g / L, sodium chloride 5g / L.

[0038] The culture medium formula for yeast is: 10 g / L peptone, 10 g / L sucrose, 3 g / L potassium dihydrogen phosphate, and 5 g / L magnesium sulfate heptahydrate.

[0039] 2. Cultivation Methods

[0040] (1) Bacterial culture: Single colonies of *Lactobacillus casei*, *Lactobacillus fermentum*, *Lactobacillus short starter*, and *Enterococcus faecalis* were picked and inoculated into their respective bacterial culture media, and cultured at 37°C and 220 r / min for 24 h. After centrifugation, the colonies were resuspended in a preservative solution containing 100 g / L skim milk powder, and the bacterial concentration was adjusted to 1. 10 8 CFU / mL was used to obtain single bacterial suspensions of each bacterium.

[0041] (2) Yeast culture: Single colonies of Saccharomyces cerevisiae were picked and inoculated into yeast culture medium, and cultured at 30℃ and 150 r / min for 24 h; after the culture was completed, the colonies were centrifuged, and the colony weight was suspended in a protective agent containing 100 g / L skim milk powder, and the colony concentration was adjusted to 1. 10 8 CFU / mL was used to obtain a yeast suspension.

[0042] 3. Detection Method

[0043] (1) Detection of flavor compounds: This invention uses headspace solid-phase microextraction (HS-SPME) combined with gas chromatography-mass spectrometry (GC-MS) to detect the content of characteristic flavor compounds such as indole, 3-methylindole, phenol and 4-ethylphenol in the brine of amaranth fermented tofu. The specific conditions are as follows: ① Standard product preparation: Take 770 μL of n-hexane and put it into a 1.5 mL centrifuge tube. Add appropriate amounts of C10-C25 commercial mixed standard, C26, C27, C28, and C29 n-alkane standards in sequence, vortex and mix well to obtain a mixed standard stock solution of C10-C29 n-alkane (concentration of 50 μg / mL). Dilute the above stock solution to 10 μg / mL and analyze it in the same batch as the test sample.

[0044] ② Sample pretreatment: Take 2 g of amaranth-infused stinky tofu brine sample and place it in a 20 mL headspace vial. Add 2.5 µL of internal standard (naphthalene-d8, concentration 20 µg / mL) and 4 mL of saturated sodium chloride aqueous solution, and immediately seal the headspace vial.

[0045] ③HS-SPME-GC / MS analysis conditions: The analytical instrument was a Thermo Orbitrap Exploris GC gas chromatograph-mass spectrometer.

[0046] SPME conditions: SPME Arrow Fiber (120 μm, Thermo, DVB / Carbon WR / PDMS).

[0047] GC conditions: Samples were injected into the GC-MS system in split mode with a split ratio of 10:1. After separation by a VF-WAXms capillary column (25 m × 0.25 mm × 0.2 µm), the samples were detected by mass spectrometry. The injection port temperature was 240℃, the carrier gas was high-purity helium at a flow rate of 1.0 mL / min, and the septum purge flow rate was 3 mL / min. Temperature program: Initial temperature 40℃, equilibration for 0 min, increased to 120℃ at a rate of 8℃ / min, then increased to 230℃ at a rate of 20℃ / min and held for 4.5 min, for a total run time of 20 min.

[0048] MS conditions: full scan mode, scan range 35-500 m / z, resolution 30000, ion source EI, ion source temperature 250℃, electron energy 70 eV, emission current 50 μA.

[0049] (2) Sensory evaluation methods

[0050] Take 10 mL of amaranth stinky tofu brine and place it in a colorless transparent beaker. Twenty trained professional evaluators will score it from two aspects: aroma (odor, comfort, persistence, and off-odor) and appearance (color, uniformity, and abnormality). The full score is 50 points (see Table 1 and Table 2 for the scoring criteria).

[0051] Table 1. Scoring Criteria for Brine Odor

[0052] Note: If an unusual odor is detected, you must describe the characteristics of the odor in words while giving a score.

[0053] Table 2 Appearance Scoring Criteria for Brine

[0054] Note: If any anomalies are observed, the anomaly characteristics must be described in words while assigning a score.

[0055] Example 1: Preparation of Compound Microbial Agent

[0056] Mix single-strain bacterial suspensions of five strains according to the following ratio (each strain concentration is 1×10⁻⁶). 8 Five groups of compound microbial agents with different ratios were prepared (CFU / mL): One set of compound microbial inoculants: five strains mixed in a volume ratio of 5:3:3:10:15; Two sets of compound microbial inoculants: five strains mixed in a volume ratio of 15:10:10:25:30; Three groups of compound microbial inoculants: five strains mixed in a volume ratio of 10:8:8:20:20; Four groups of compound microbial inoculants: five strains mixed in a volume ratio of 10:10:10:20:25; Five groups of compound microbial inoculants: five strains mixed in a volume ratio of 10:8:8:20:25.

[0057] Example 2: Preparation of brine for amaranth-flavored stinky tofu

[0058] Based on the production environment of Suzhou Jinji Food Co., Ltd., a standardized process was used to prepare the brine for amaranth and stinky tofu, with the specific settings as follows: Option 1: One set of compound microbial inoculant Raw material processing: Sort, wash, and drain the old amaranth stalks until the moisture content is ≤70%, then cut them into sections. Select sections with a stem diameter ≥1cm for later use; process the winter bamboo shoots and cut them into chunks / slices; wash and chop the snow cabbage and alfalfa, and prepare the spice packet. Add salt and mix: Add 4 kg of salt per 100 kg of raw materials, and mix well in layers; Inoculation: Add 0.2 kg of compound microbial agent per 100 kg of raw material, and then add water to submerge the raw material, maintaining the water level 5 cm above the raw material. Segmented fermentation: Seal fermentation at 30℃ for 6-7 days, stirring once a day; then let stand for 7-9 days, controlling the pH of the system at 4.5-5.5; after 14-16 days of fermentation, cool down to 20℃ and continue fermentation until the total fermentation cycle is 30-45 days, maintaining a salinity of 4% throughout; terminate fermentation when indole ≥200μg / L, 3-methylindole ≥70μg / L, phenol ≥70μg / L, and 4-ethylphenol ≥100μg / L are detected in the brine; Post-processing: Filter through a 30-mesh filter to obtain coarse brine, sterilize at 70℃ for 25 minutes, seal and fill, and store at 10-15℃. After opening, refrigerate at 0-4℃ and use within one month to obtain amaranth stinky tofu brine 1.

[0059] Option 2, other ratios of microbial agents

[0060] Based on the previous scheme, simply replace compound microbial agent 1 with compound microbial agents 2-5 respectively to obtain amaranth stinky tofu brine 2-5.

[0061] Following the detection method described above, the contents of indole, 3-methylindole, phenol and 4-ethylphenol in amaranth stinky tofu brine 1-5 were detected respectively. Furthermore, sensory evaluation of amaranth stinky tofu brine 1-5 was conducted, and the results are shown in Table 3.

[0062] As shown in Table 3, the preparation of amaranth stinky tofu brine using the compound microbial agent 1 of this invention can fundamentally overcome the potential problems of traditional processes in terms of food safety, production process, and health risks, and truly achieve standardized production. Among them, the content of indole in amaranth stinky tofu brine 1 can reach 337.7 μg / L, 3-methylindole can reach 93.2 μg / L, phenol can reach 96.9 μg / L, and 4-ethylphenol can reach 194.9 μg / L, with a sensory score of 48 points. All indicators are significantly better than those of compound microbial agents with other ratios, and the improvement effect of other microbial agents is far weaker than that of compound microbial agent 1.

[0063] Comparative Example

[0064] Three comparative examples were set up to verify the beneficial effects of the present invention. Except for the inoculation step, all other operations in the comparative examples were completely consistent with Scheme 1 of Example 2: Comparative Example 1: Brine fermented with a single brewing yeast The inoculation steps were adjusted as follows: only 0.2 kg of brewing yeast (CGMCC No: 4747, the same brewing yeast strain used in Example 1) was added for every 100 kg of raw materials, without adding other strains. Fermentation and post-treatment were completed according to the steps of Scheme 1 in Example 2 to obtain single-strain fermented brine 1.

[0065] The test and sensory evaluation results are shown in Table 3. The flavor substance content of the brine prepared in this comparative example is much lower than that of the amaranth brine 1, the sensory score is lower, the fermentation cycle is extended to 50-60 days, and it is prone to contamination by miscellaneous bacteria, which leads to the deterioration of the brine. It cannot meet the requirements of amaranth stinky tofu brine for flavor and safety.

[0066] Comparative Example 2: Fermentation of brine using a combination of commercial microbial agents

[0067] The inoculation steps were adjusted as follows: 0.2 kg of commercial microbial agent combination (Lactobacillus plantarum CGMCC NO: 1.1856, Saccharomyces cerevisiae CGMCC No: 4747, Bacillus subtilis CGMCC NO: 1.1086, volume ratio 1:1:1) was added for every 100 kg of raw materials, and fermentation and post-treatment were completed according to the steps of Scheme 1 in Example 2 to obtain commercial microbial agent combination fermented brine 2.

[0068] The results of the testing and sensory evaluation are shown in Table 3. The content of flavor substances in the brine prepared in this comparative example was lower than that in the amaranth brine 1, and the sensory score was close to that of the blank control. Moreover, the salinity fluctuated greatly during the fermentation process, the stability of the brine was poor, and the shelf life was only 3-6 months, making it impossible to achieve standardized production.

[0069] Comparative Example 3: Traditional Naturally Fermented Brine

[0070] Without inoculation, the raw materials are processed, salt is added, and water is added to submerge the raw materials before direct sealing and fermentation. The remaining operations are completely consistent with Scheme 1 of Example 2. The traditional naturally fermented brine 3 is obtained by relying on the microorganisms carried by the raw materials for natural fermentation.

[0071] The testing and sensory evaluation results are shown in Table 3. The fermentation cycle of this comparative example was unstable, ranging from 45 to 70 days. The content of flavor substances was low and fluctuated greatly between batches. It was prone to the growth of harmful bacteria, posing a significant food safety hazard. The quality of products from different batches varied greatly, making it impossible to achieve standardized and large-scale production. It was significantly different from the brine prepared by compound microbial agent group 1.

[0072] Table 3. Content of main flavor compounds in amaranth-flavored stinky tofu brine

[0073] Combined with Table 3 and Figure 1 The results show that the amaranth stinky tofu brine prepared by the compound microbial agent of this invention has significantly higher content of characteristic flavor substances (indole, 3-methylindole, phenol, 4-ethylphenol) and sensory scores than the comparative examples, with the amaranth brine prepared by compound microbial agent 1 showing the best results. The comparative examples generally suffered from insufficient flavor, unstable fermentation, and numerous safety hazards, which sufficiently proves that the compound microbial agent of this invention can effectively solve the drawbacks of traditional and conventional microbial agent fermentation, truly achieving standardized and safe production of amaranth stinky tofu brine.

[0074] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A compound microbial inoculant for producing brine for amaranth-flavored stinky tofu, characterized in that: It is composed of Lactobacillus casei LKG-H01, Lactobacillus fermentum LKG-H02, short-starter Lactobacillus LKG-B01, Enterococcus faecalis LKG-B02 and Saccharomyces cerevisiae; The L. caseiformis LKG-H01 ( Lacticaseibacillus paracasei (), deposited at the China General Microbiological Culture Collection Center, China, on March 16, 2026, with the biological accession number CGMCCNO: 37914; The fermented Lactobacillus mucinus LKG-H02 ( Limosilactobacillus fermentum (), deposited at the China General Microbiological Culture Collection Center, China, on March 16, 2026, with accession number CGMCCNO: 37915; The short fermentation agent Lactobacillus LKG-B01 ( Levilactobacillus brevis The specimen is deposited at the China General Microbiological Culture Collection Center (CGMCC) on March 16, 2026, with the accession number CGMCC NO: 37911. The Enterococcus faecalis LKG-B02 ( Enterococcus faecium (), deposited at the China General Microbiological Culture Collection Center, China, on March 16, 2026, with accession number CGMCC NO: 37912.

2. The compound microbial agent according to claim 1, characterized in that: The live cell ratio of the Lactobacillus casei, Lactobacillus fermentum, Lactobacillus short starter, Enterococcus faecalis, and Saccharomyces cerevisiae is 5~15:3~10:3~10:10~25:15~30.

3. The compound microbial agent according to claim 1 or 2, characterized in that: The live bacteria ratio of *Lactobacillus casei*, *Lactobacillus fermentum*, *Lactobacillus short starter*, *Enterococcus faecalis*, and *Saccharomyces cerevisiae* is 8:5:5:15:

20.

4. The application of the compound microbial agent according to any one of claims 1-3 in the production of amaranth stinky tofu brine, characterized in that: The compound microbial agent was added to the raw material of amaranth stinky tofu brine for fermentation. The fermentation termination conditions were: indole content ≥200μg / L, 3-methylindole ≥70μg / L, phenol ≥70μg / L and 4-ethylphenol ≥100μg / L in the brine.

5. The application of the compound microbial agent according to claim 4 in the production of amaranth stinky tofu brine, characterized in that: The compound microbial agent is added to the raw materials at a rate of 0.1~0.3 kg / 100 kg of raw materials, and the initial total viable count after inoculation is 7.5%. 10 5 ~1.5 10 6 CFU / g raw material.

6. The application of the compound microbial agent according to claim 5 in the production of amaranth stinky tofu brine, characterized in that: The compound microbial agent is added at a rate of 0.2 kg / 100 kg of raw material, and the initial total viable count is 1.

1. 10 6 CFU / g raw material.

7. The application of the compound microbial agent according to claim 4 in the production of amaranth stinky tofu brine, characterized in that, The fermentation includes the following steps: (1) Raw material processing and salting: sort, wash and drain the old amaranth stalks until the moisture content is ≤70% and then cut them into sections. Select the sections with a stem diameter ≥1cm for later use; process the winter bamboo shoots and cut them into chunks / slices; wash and chop the remaining vegetables and prepare the spice packet; the remaining vegetables include snow cabbage and alfalfa. (2) Inoculation and fermentation: Add the compound microbial agent, add water to submerge, seal and ferment at 25-30℃ for 6-7 days, stir daily, then let stand to control the pH value to 4.5-5.5, cool down to 15-20℃ when fermentation reaches 14-16 days, maintain salinity of 3%-5% throughout the process, and the total fermentation period is 30-45 days; (3) Post-processing: When the fermentation broth reaches the termination conditions, it is filtered, sterilized and filled.

8. The application of the compound microbial agent according to claim 7 in the production of amaranth stinky tofu brine, characterized in that: In step (1), the amount of salt added is 3-5 kg ​​per 100 kg of fresh raw materials.

9. The application of the compound microbial agent according to claim 8 in the production of amaranth stinky tofu brine, characterized in that: In step (2), the initial fermentation temperature is 25~30℃, the static stage is 7-9 days, the temperature is maintained at 15~20℃ after cooling, and the salinity is 3~5% throughout the fermentation process.

10. Amaranth stinky tofu brine produced using any one of the compound microbial agents according to claims 1-3.

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