A method for rapidly fermenting high-flavor soy sauce by synergistically enhancing the soy sauce and black bean flavors of high-flavor soy sauce with multiple strains

Abstract

The present invention relates to the technical field of food fermentation and brewing, specifically a method for rapidly fermenting a high-flavor soy sauce by synergizing multiple strains to enhance the flavor of soy sauce and fermented black beans. The method comprises the following steps: adding flavor lactic acid bacteria to the soy sauce in the early stage of fermentation; adding flavor-producing yeast to the soy sauce in the middle stage of fermentation; inoculating the flavor lactic acid bacteria in an amount of 10 3 CFU / g; the inoculum size of aroma-producing yeast was 10 6～ 10 7 The present invention uses the synergistic fermentation of aroma-producing microorganisms and process innovation to further improve the product's umami flavor, enhance the traditional characteristic flavors of soy sauce, such as soy sauce aroma and fermented black bean aroma, and improve product quality, while retaining the high raw material utilization rate and controllable fermentation of the high-salt dilute fermentation process.

Description

Technical Field

[0001] The present invention relates to the technical field of food fermentation and brewing, and specifically to a method for rapidly fermenting high-flavor soy sauce by synergistically enhancing the soy sauce flavor and black bean flavor of soy sauce with multiple strains. Background Art

[0002] Soy sauce originated in my country during the Zhou Dynasty and later spread to Japan, South Korea, Southeast Asia, and other regions, gradually becoming an indispensable condiment in East Asian cuisine. Traditional Chinese soy sauce brewing primarily utilizes a natural sun-dried fermentation process. This involves making a koji from soybeans and flour, mixing it with salt water, and then placing it in large ceramic vats, where it is exposed to the sun and dew for years, turning and drying the fermented mash. This process takes two to three years. This traditional soy sauce boasts a unique flavor, characterized by a rich soy sauce aroma and a prominent fermented black bean flavor, making it highly popular with consumers. However, this natural sun-dried fermentation process is plagued by long production cycles and low efficiency. Furthermore, the long years of manual labor and unregulated fermentation lead to inconsistent production quality and uncontrolled food safety. Consequently, this process is still practiced only by a few small workshops, while the current mainstream soy sauce brewing process is high-salt, dilute fermentation.

[0003] The high-salt, dilute fermentation process is a scientific brewing method developed by incorporating modern brewing equipment and scientific management techniques based on the essence of natural sun-dried fermentation. High-salt, dilute soy sauce products are known for their high umami, rich flavor, and bright red color. However, their aroma of soy sauce and fermented black beans is weak, lacking the characteristic flavor of traditional soy sauce. The characteristic flavor of soy sauce is primarily formed through the fermentation of microorganisms (such as Aspergillus, yeast, and lactic acid bacteria) to hydrolyze the raw materials into a variety of amino acids, peptides, organic acids, and sugars. These are then transformed through complex biochemical reactions to form the characteristic flavor.

[0004] Therefore, the use of beneficial microorganisms for synergistic fermentation combined with brewing process innovation, while retaining the high utilization rate of raw materials and controllable fermentation of high-salt dilute fermentation process, further improves the umami of the product and enhances the traditional characteristic flavors of soy sauce such as soy sauce aroma and black bean aroma, which plays an important role in promoting the high-quality development of my country's soy sauce industry. Summary of the Invention

[0005] The present invention aims to address the technical deficiencies of the aforementioned high-salt, dilute-state fermentation process by providing a rapid, multi-strain synergistic fermentation method for enhancing the flavors of high-umami soy sauce, including the soy sauce and black bean flavors. This technology, through the collaborative fermentation of aroma-producing microorganisms and process innovation, further enhances the umami flavor of the product, while maintaining the high raw material utilization and controllable fermentation characteristics of the high-salt, dilute-state fermentation process. This enhances the traditional soy sauce flavors, such as the soy sauce and black bean flavors, and improves product quality.

[0006] In order to achieve the above-mentioned object of the invention, the specific technical solution of the present invention is as follows:

[0007] A method for rapidly fermenting a high-flavor soy sauce by synergistically enhancing the flavor of soy sauce and fermented black bean sauce with multiple strains, comprising the following steps: adding flavor lactic acid bacteria to ferment the soy sauce in the early stage of soy sauce fermentation; adding flavor-producing yeast to ferment the soy sauce in the middle stage of soy sauce fermentation; inoculating the flavor lactic acid bacteria in an amount of 10 3 CFU / g; the inoculum size of aroma-producing yeast was 10 6～ 10 7 CFU / g.

[0008] Furthermore, in the method of rapidly fermenting a multi-strain synergistic enhancement of the soy sauce and fermented black bean flavors of high-flavor soy sauce, the flavor lactic acid bacteria is Pediococcus acidilactici Pi Niang R036, which is an existing strain (this application does not involve preservation), and has been deposited in the Guangdong Provincial Microbiological Culture Collection Center on March 21, 2019, with a preservation number of CDMCC No: 60617; the aroma-producing yeast is Zygosaccharomyces rouxii Pi Niang J028, which is an existing strain (this application does not involve preservation), and has been deposited in the Guangdong Provincial Microbiological Culture Collection Center on March 21, 2019, with a preservation number of CDMCC No: 60616.

[0009] Furthermore, the above-mentioned method of rapidly fermenting a multi-strain synergistically enhancing the flavor of high-flavor soy sauce and fermented black bean sauce comprises the following specific steps:

[0010] (1) Making fermented black beans and soy sauce koji: soybeans or black beans, the raw materials of fermented black beans, are selected, impurities removed, and soaked, then steamed, cooled, and inoculated with soy sauce koji to make koji and culture to obtain fermented black beans; soybeans, the raw materials of soy sauce, are selected, impurities removed, and soaked, then steamed at high temperature, then immediately cooled, mixed evenly with fried wheat flour, inoculated with soy sauce koji to make koji and culture to obtain soy sauce koji.

[0011] (2) Low-temperature dilute fermentation of fermented black beans and fermentation of soy sauce: the fermented black bean koji is mixed with low-temperature brine, and then transferred to a temperature-controlled closed fermentation tank for low-temperature dilute fermentation; the soy sauce koji is mixed with low-temperature brine, the salt content of the sauce mash is controlled, and then the temperature is varied in stages in a closed heat-insulated fermentation tank to simulate the temperature change of natural fermentation to obtain mature sauce mash;

[0012] (3) Solid-liquid separation: After the low-temperature dilute fermentation of fermented black beans is completed, the koji and brine are separated to obtain solid koji and fermented brine respectively;

[0013] (4) high-temperature solid-state fermentation of fermented black beans: placing the solid fermented black bean curd obtained by separation in step (3) in a sealed heat-insulating fermentation tank for anaerobic fermentation to obtain a finished fermented black bean curd;

[0014] (5) Soaking and pressing: adding the fermented fermented black beans obtained in step (4) to the mature soy sauce mash obtained in step (2), then performing heat preservation and soaking extraction, and finally conveying the material to a press for pressing and extracting oil, thereby obtaining a soy sauce crude oil having both soy sauce and black bean flavors.

[0015] As a better embodiment of the present application, in step (1) of the method for rapidly fermenting a multi-strain synergistically enhancing the soy sauce flavor and fermented black bean flavor of high-flavor soy sauce, the amount of soy sauce koji essence added in the fermented black bean koji is 0.03% based on the dry weight of the raw materials; the culture environment is 28-33°C for 48 hours, and the koji is turned once every 12-18 hours, and finally the surface of the koji is covered with yellow-green mycelium.

[0016] As a preferred embodiment of the present application, in step (1) of the method for rapidly fermenting a multi-strain synergistically enhancing the soy sauce and black bean flavors of high-flavor soy sauce, the amount of soy sauce koji essence added in the soy sauce koji making is 0.03% based on the total weight of soybeans and roasted wheat flour; the culture environment is 28-33°C for 40-45 hours, and the koji is turned over at 12 hours and 24 hours respectively.

[0017] As a preferred embodiment of the present application, in the low-temperature dilute fermentation of step (2) of the method for rapidly fermenting a high-flavor soy sauce with multiple strains of bacteria in a synergistic manner to enhance the soy sauce and fermented black bean flavor, the temperature of the low-temperature brine is 5-10°C, and the brine concentration is 18-19°Bé; the fermented black bean koji and the low-temperature brine are mixed in a mass ratio of 1:1.5-2; the temperature of the low-temperature dilute fermentation is 15-20°C, and the time is 3-5 days, during which the mixture is stirred once every morning and evening, each stirring for 10-20 minutes.

[0018] As a better embodiment of the present application, in the soy sauce fermentation of step (2) of the method for rapidly fermenting high-flavor soy sauce with multiple strains of bacteria synergistically enhancing the soy sauce flavor and fermented black bean flavor, the temperature of the low-temperature brine is 5-10°C, and the brine concentration is 19-20°Bé; the soy sauce koji and the low-temperature brine are mixed in a mass ratio of 1:1.8-2, the salt content of the soy sauce mash is controlled to be 16.5%-18%, and the soy sauce mash temperature is 10-15°C.

[0019] As a preferred embodiment of the present application, in the soy sauce fermentation of step (2) of the method for rapidly fermenting high-flavor soy sauce with multiple strains of bacteria to synergistically enhance the soy sauce flavor and black bean flavor of high-flavor soy sauce, the staged temperature-variable fermentation comprises the following steps:

[0020] In the early stage of fermentation, i.e. 0-30 days: control the temperature of the mash at 15-20℃ for low-temperature pre-fermentation, and the temperature in the center of the fermentation tank must be controlled as much as possible not to exceed 20℃; ventilate and stir for 20-30 minutes every day for 1-2 weeks before fermentation, and stir once every 4-5 days thereafter. The mash should be in a uniform state without material cover or material blocks; inoculate flavor lactic acid bacteria on the 20th to 25th day of fermentation, with an inoculation amount of 103 CFU / g mash;

[0021] In the middle stage of fermentation, i.e. 30-90 days: slowly raise the temperature of the mash to 28-30℃ and keep it warm for 60±1 days. During this period, aerate and stir for 20-30 minutes every 4-5 days. When the pH of the mash is 5.0-5.2, inoculate the aroma-producing yeast with an inoculation amount of 10 6～ 10 7 CFU / g mash, and aeration and stirring for 20-30 minutes every day for 1 week after yeast inoculation;

[0022] In the late fermentation stage, i.e. 90-120 days: continue to increase the fermentation temperature to 40-45℃ for high-temperature post-ripening and aroma-enhancing fermentation, during which time aerate and stir for 20-30 minutes every 5 days; until the beans are melted, the color of the sauce mash is dark brown, the amino acid nitrogen content of the sauce mash is ≥1.0g / 100mL and there is no change for 1 week before and after, a mature sauce mash is obtained.

[0023] As a better embodiment of the present application, in step (4) of the method for rapidly fermenting a multi-strain synergistically enhancing the soy sauce and fermented black bean flavor of high-flavor soy sauce, the temperature for anaerobic fermentation of the solid fermented black bean koji is 45-50°C, and the anaerobic fermentation time is 2-3 months.

[0024] As a better embodiment of the present application, in step (5) of the method for rapidly fermenting a multi-strain synergistically enhancing the soy sauce and fermented black bean flavor of high-flavor soy sauce, the amount of fermented black bean curd product added is 5% to 10% of the mass of the mature soy sauce mash; the temperature of the heat preservation and immersion extraction is 45 to 50°C, and the time is 24 to 48 hours, during which compressed air is passed and stirred for 10 minutes every 6 hours; and the amino acid nitrogen content in the crude oil is ≥1.0g / 100g.

[0025] Compared with the existing technology, the beneficial effects of the present invention are:

[0026] (1) In the present invention, fermentation of fermented black beans in a low-temperature, dilute state is adopted. While suppressing the growth of miscellaneous bacteria at low temperature, the autolysis of Aspergillus oryzae hyphae is promoted, and the release of umami substances such as intracellular biological enzymes (such as glutaminase) and nucleic acids of Aspergillus oryzae is accelerated.

[0027] (2) In the present invention, after the low-temperature, dilute fermentation of fermented black beans is completed, the koji and brine are separated to obtain solid koji and fermentation brine, respectively. The solid koji is used for subsequent fermentation of fermented black beans; while the fermentation brine, rich in various biological enzymes, nucleic acids, and nutrients, is used for subsequent soy sauce fermentation, further improving the fermentation efficiency and product flavor of the soy sauce.

[0028] (3) In the present invention, after solid-liquid separation, high-temperature solid-state fermentation is also adopted to ferment fermented black beans. The high temperature increases the fermentation speed, accelerates the decomposition of substances such as protein, starch, fat in the raw materials to form small molecules such as amino acids, small molecule sugars, fatty acids, and further promotes these small molecules to form the unique black bean flavor and sauce flavor of black beans through esterification reaction, Maillard reaction, etc.

[0029] (5) In the present invention, soy sauce fermentation adopts staged temperature-variable fermentation. In the early stage of fermentation, low temperature is used to inhibit the growth and metabolism of microorganisms to prevent the rapid decrease in the pH of the sauce mash, creating favorable conditions for various biological enzymes to hydrolyze substrates; in the middle stage of fermentation, suitable conditions are created for the growth of beneficial microorganisms through fermentation at appropriate temperature, promoting the synergistic fermentation of flavor lactic acid bacteria and aroma-producing yeasts to form flavor substances such as acids, alcohols, aldehydes and ketones; in the late stage of fermentation, high-temperature post-ripening and flavor-enhancing fermentation is carried out to promote the esterification reaction of acids and alcohol compounds metabolized by lactic acid bacteria, yeasts, etc., thereby enhancing the flavor of soy sauce.

[0030] (6) All stages cooperate with each other and work together to ensure that the amino acid nitrogen content of the prepared crude oil is ≥1.0g / 100g, with a bright red color, a fresh and mellow taste, a strong aroma, and prominent sauce and soy flavors. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 The changes in the physical and chemical indicators of the mash during the fermentation process of different processes in Example 1;

[0032] Figure 2 The free amino acid content of the crude soy sauce fermented by different processes in Example 1;

[0033] Figure 3 The sensory evaluation scores of the soy sauce crude oil fermented by different processes in Example 1;

[0034] Figure 4 The effect of synergistic fermentation on organic acids and free amino acids in high-salt dilute fermented mash in Example 2;

[0035] Figure 5 The effect of synergistic fermentation on the volatile flavor substances in high-salt dilute fermented mash in Example 2;

[0036] Figure 6 The effect of synergistic fermentation on the sensory quality of high-salt dilute fermented mash in Example 2;

[0037] Figure 7 This is the effect of high-temperature post-ripening fermentation on the soy sauce and fermented bean flavors of high-salt dilute soy sauce in Example 3. DETAILED DESCRIPTION

[0038] A method for rapidly fermenting high-flavor soy sauce by synergistically enhancing the soy sauce flavor and black bean flavor of high-flavor soy sauce with multiple strains, comprising the following steps:

[0039] (1) Making fermented black beans. Soybeans or black beans are selected, cleaned, and soaked, then steamed at 105°C for 5-10 minutes. Cool to room temperature, inoculate with 0.03% soy sauce koji (based on the dry weight of the raw material), and incubate at 28-33°C for 48 hours. Turn the koji over once every 12-18 hours until the surface of the koji is covered with yellow-green mycelium. This is the fermented black bean koji.

[0040] (2) Low-temperature dilute fermentation of fermented black beans. Mix fermented black bean koji with low-temperature brine at a mass ratio of 1:1.5-2, so that the koji particles are fully dispersed in the brine. Then transfer the fermented bean koji to a temperature-controlled, sealed fermentation tank and ferment it at a low temperature of 15-20°C for 3-5 days. Stir once a day in the morning and evening, each time for 10-20 minutes. While inhibiting the growth of miscellaneous bacteria by low temperature, it promotes the autolysis of Aspergillus oryzae hyphae and accelerates the release of umami substances such as enzymes (such as glutaminase) and nucleic acids in Aspergillus oryzae cells.

[0041] Furthermore, the temperature of the low-temperature brine is 5-10°C, and the brine concentration is 18-19°Bé.

[0042] (3) Solid-liquid separation. After the low-temperature dilute fermentation of fermented black beans is completed, the koji and brine are separated to obtain solid koji and fermentation brine, respectively. The solid koji is used for subsequent fermentation of fermented black beans; while the fermentation brine, which is rich in various biological enzymes, nucleic acids and nutrients, is used for subsequent soy sauce fermentation, further improving the fermentation efficiency and product flavor of soy sauce.

[0043] (4) High-temperature solid-state fermentation of fermented black beans. The solid fermented black bean curd separated in step (3) is placed in a sealed heat-insulated fermentation tank and anaerobically fermented at a high temperature of 45 to 50° C. for 2 to 3 months until the product has a strong black bean aroma and sauce flavor. This is the finished product of fermented black beans. The high temperature increases the fermentation speed, accelerates the decomposition of substances such as protein, starch, and fat in the raw materials to form small molecules such as amino acids, small molecule sugars, and fatty acids, and further promotes these small molecules to form the unique black bean aroma and sauce flavor of fermented black beans through esterification reactions and Maillard reactions.

[0044] (5) Making koji for soy sauce. Soybeans are selected, cleaned, and soaked, then steamed at 120-130°C for 3-6 minutes. Immediately cool to 35-40°C, mix with roasted wheat flour at a mass ratio of 7:3 or 6:4, and inoculate with 0.03% soy sauce koji essence (based on the total weight of soybeans and roasted wheat flour). The koji is cultured at 28-33°C for 40-45 hours, turning the koji twice every 12-24 hours until the surface of the koji is covered with yellow-green mycelium. This is the finished koji for soy sauce.

[0045] (6) Soy sauce fermentation. The soy sauce koji was mixed with low-temperature brine at a mass ratio of 1:1.8-2. The salt content of the mixed soy sauce mash was controlled at 16.5%-18% and the temperature of the soy sauce mash was controlled at 10-15°C. Then, the soy sauce mash was fermented in a sealed heat-insulated fermentation tank to simulate the temperature changes of natural fermentation.

[0046] Early fermentation stage (0-30 days): Control the temperature of the mash at 15-20℃ for low-temperature pre-fermentation. The temperature in the center of the fermentation tank must be controlled as much as possible not to exceed 20℃. Low temperature is used to inhibit the growth and metabolism of microorganisms to prevent the pH of the mash from dropping rapidly, creating favorable conditions for various biological enzymes to hydrolyze substrates. Aerate and stir for 20-30 minutes every day for 1-2 weeks before fermentation, and stir once every 4-5 days thereafter. The mash should be in a uniform state without any cover or block. On the 20th-25th day of fermentation, inoculate flavor lactic acid bacteria with an inoculation amount of 10 3 CFU / g mash.

[0047] Fermentation mid-term (30-90 days): slowly raise the temperature of the mash to 28-30℃, not exceeding 35℃, and keep it at 28-30℃ for about 60 days, aerating and stirring for 20-30 minutes every 4-5 days. When the pH of the mash is 5.0-5.2, inoculate the aroma-producing yeast with an inoculation amount of 10 6～ 10 7 CFU / g mash, and aeration and stirring for 20-30 minutes daily for one week after yeast inoculation. During this stage, optimal temperature fermentation creates favorable conditions for the growth of beneficial microorganisms, promoting the synergistic fermentation of flavor-producing lactic acid bacteria and aroma-producing yeasts to form flavor substances such as acids, alcohols, aldehydes, and ketones.

[0048] Late fermentation (90-120 days): Continue to raise the fermentation temperature to 40-45°C, but not exceeding 48°C, for high-temperature post-ripening and flavor-enhancing fermentation. This promotes the esterification of acids and alcohols metabolized by lactic acid bacteria and yeasts, enhancing the soy sauce's aroma and flavor. Aerate and stir the soy sauce for 20-30 minutes every five days to prevent the growth of surface film-forming yeast. Mature soy sauce mash is considered mature when the beans have melted, the mash is dark brown, and the amino acid nitrogen content is ≥1.0g / 100mL, with no change for a week.

[0049] Furthermore, the concentration of the low-temperature brine is 19-20° Bé and the temperature is 5-10° C.; and the solid-liquid separation brine rich in various biological enzymes and nutrients obtained by separation in step (3) is added.

[0050] Furthermore, the flavor lactic acid bacteria is Pediococcus acidilactici Pi Niang R036, which was deposited in Guangdong Provincial Microbial Culture Collection Center on March 21, 2019, with the deposit number CDMCC No: 60617.

[0051] Furthermore, the aroma-producing yeast is the Roux combined yeast Pi Niang J028, which was deposited in the Guangdong Provincial Microbial Culture Collection Center on March 21, 2019, with a deposit number of CDMCC No: 60616.

[0052] (7) Soaking and pressing. Add 5% to 10% (m / m) of the fermented fermented black beans obtained in step (4) to the mature mash of step (6), and further heat the mash to 45 to 50°C. Keep warm and soak for 24 to 48 hours, stirring with compressed air for 10 minutes every 6 hours. Then, transfer the mash to a press to extract oil, thereby obtaining crude soy sauce.

[0053] Furthermore, the amino acid nitrogen content of the crude oil is ≥1.0g / 100g, and the crude oil has a bright red color, a fresh and mellow taste, a strong aroma, and prominent sauce and black bean flavors.

[0054] All features disclosed in this specification, or all steps in the disclosed methods or processes, except mutually exclusive features and / or steps, can be combined in any manner.

[0055] Any feature disclosed in this specification (including claims and abstract), unless otherwise stated, may be replaced by other equivalent or similar features. That is, unless otherwise stated, each feature is only an example of a series of equivalent or similar features.

[0056] The detection methods involved in the following examples are:

[0057] (1) Total acid determination: Determined by pH meter point titration method according to GB 12456-2021 “Determination of total acid in foods”.

[0058] (2) Determination of amino acid nitrogen: Determined by the acidometer method in accordance with GB 5009.235-2016 “Determination of amino acid nitrogen in foods”.

[0059] (3) Reducing sugar determination: 3,5-dinitrosalicylic acid method was used for determination.

[0060] (4) Determination of organic acids: Liquid chromatography was used for determination. For the specific method, please refer to the 2023 published paper "Salinity plays a dual role in broad bean paste-meju fermentation".

[0061] (5) Determination of free amino acids: Liquid chromatography was used for determination. For the specific method, please refer to the 2023 published paper "Salinity plays a dual role in broad bean paste-meju fermentation".

[0062] (6) Determination of volatile flavor substances: Headspace solid phase microextraction combined with gas chromatography-mass spectrometry (GC-MS) was used for determination. For specific methods, please refer to the 2024 published paper “Analysis of Flavor Quality Characteristics of Traditional Sichuan Brewed Soy Sauce”.

[0063] (7) Sensory evaluation method:

[0064] Eight evaluators who are in good health (including food allergies), non-smokers, and have normal basic taste and smell sense were selected to form a soy sauce sensory evaluation panel. The panel members include 5 men and 3 women, aged between 20 and 30 years old.

[0065] When smelling the sample, take 20mL of soy sauce sample directly and put it into a 3-digit coded, transparent, odorless, covered tasting cup; when tasting the sample, dilute the soy sauce 1 times with purified water and put it into the corresponding tasting cup. The taster takes 0.5mL of sample into a small spoon through a plastic straw for tasting. All samples are stored in a refrigerator at 4°C and taken out for use about 30 minutes before the test. The samples are randomly provided to the tasters in rounds, with 5 samples per round. The tasting time for each round is within 30 minutes, and there is an interval of 15 minutes between each round. During this period, purified water or odorless biscuits are provided to the tasters to clean their mouths. All samples are tasted 3 times. The sensory evaluation team members conduct olfactory and taste sensory evaluations of the soy sauce respectively, and score them using a 15-point system. The specific scoring basis is shown in the following table:

[0066] Table 1 Sensory evaluation standards for soy sauce

[0067]

[0068] The features and performance of the present invention are further described in detail below with reference to the embodiments.

[0069] Example 1:

[0070] A rapid fermentation process for enhancing the soy sauce and fermented black bean flavors of high-flavor soy sauce

[0071] Experimental group:

[0072] (1) Preparation of fermented black beans. Soybeans are selected, cleaned, and soaked, then steamed at 105°C for 8 minutes, cooled to room temperature, inoculated with 0.03% soy sauce koji (based on the dry weight of soybeans), and cultured at 28-33°C for 48 hours. The koji is turned over once every 12 hours until the surface of the koji is covered with yellow-green mycelium. This is the finished fermented black bean koji.

[0073] (2) Low-temperature dilute fermentation of fermented black beans. Mix the fermented black bean koji with low-temperature brine at a mass ratio of 1:2, so that the koji particles are fully dispersed in the brine. Then transfer the mixture to a temperature-controlled, sealed fermentation tank and ferment it at a low temperature of 15-20°C for 5 days. Stir once every morning and evening for 10-20 minutes each time.

[0074] Furthermore, the temperature of the low-temperature brine is 5°C and the brine concentration is 18°Bé.

[0075] (3) Solid-liquid separation. After the low-temperature dilute fermentation of fermented black beans is completed, the koji and brine are separated to obtain solid koji and fermented brine respectively.

[0076] (4) High-temperature solid-state fermentation of fermented black beans. The solid fermented black bean curd separated in step (3) is placed in a sealed heat-insulated fermentation tank and anaerobically fermented at a high temperature of 45-50° C. for 3 months until the product has a strong black bean aroma and sauce aroma.

[0077] (5) Soy sauce koji making. Soybeans are selected, cleaned, and soaked, then steamed at 125°C for 4 minutes. Immediately cool to 35-40°C, mix with roasted wheat flour at a mass ratio of 6:4, and inoculate with 0.03% soy sauce koji essence (based on the total weight of soybeans and roasted wheat flour). The koji is incubated at 28-33°C for 40 hours, turning the koji once every 12 hours and 18 hours, until the surface of the koji is covered with yellow-green mycelium. This is the soy sauce koji.

[0078] (6) Soy sauce fermentation. The soy sauce koji was mixed with low-temperature brine at a mass ratio of 1:1.8. The salt content of the mixed soy sauce mash was controlled at 17% and the temperature of the soy sauce mash was controlled at around 15°C. Then, the soy sauce mash was fermented in a sealed, heat-insulated fermentation tank to simulate the temperature changes of natural fermentation and perform staged temperature-variable fermentation.

[0079] Early fermentation stage (0-30 days): Control the temperature of the mash at 15-20°C for low-temperature pre-fermentation. The temperature in the center of the fermentation tank must be controlled to be no more than 20°C. Aerate and stir for 20-30 minutes every day for 1-2 weeks before fermentation, and stir once every 4-5 days thereafter. The mash should be uniform without any cover or lumps. On the 20th day of fermentation, inoculate flavor lactic acid bacteria with an inoculation rate of 1×10 3 CFU / g mash.

[0080] Fermentation mid-term (30-90 days): slowly raise the temperature of the mash to 30°C, not exceeding 35°C, and maintain it at 28-30°C for about 60 days, with ventilation and stirring every 4-5 days for 20-30 minutes. When the pH of the mash reaches 5.2, inoculate the aroma-producing yeast with an inoculum size of 1×10 6 CFU / g mash, and the mash needs to be aerated and stirred for 20 to 30 minutes every day for one week after yeast inoculation.

[0081] Late fermentation (90-120 days): Continue to raise the fermentation temperature to 45°C, but not exceeding 48°C. Maintain the temperature at 40-45°C for post-ripening and flavor-enhancing fermentation. This promotes the esterification of acids and alcohols metabolized by lactic acid bacteria and yeasts, enhancing the soy sauce's aroma and flavor. Aerate and stir the soy sauce for 20-30 minutes every five days to prevent the growth of surface film-forming yeast. Mature soy sauce mash is considered mature when the beans have melted, the mash is dark brown, and the amino acid nitrogen content is ≥1.0g / 100mL, with no change for a week.

[0082] Furthermore, the concentration of the low-temperature brine is 19°Bé and the temperature is 10°C; and the solid-liquid separation brine obtained by separation in step (3) is added.

[0083] Furthermore, the flavor lactic acid bacteria is Pediococcus acidilactici Pi Niang R036, which was deposited in Guangdong Provincial Microbial Culture Collection Center on March 21, 2019, with the deposit number CDMCC No: 60617.

[0084] Furthermore, the aroma-producing yeast is the Roux combined yeast Pi Niang J028, which was deposited in the Guangdong Provincial Microbial Culture Collection Center on March 21, 2019, with a deposit number of CDMCC No: 60616.

[0085] (7) Soaking and pressing. Add 5% (m / m) of the fermented fermented black beans obtained in step (4) to the mature mash of step (6). The mash is then heated to 50°C and soaked for 48 hours, with compressed air stirring for 10 minutes every 6 hours. The mash is then transferred to a press to extract the oil, thereby obtaining crude soy sauce.

[0086] Control group:

[0087] Soy sauce was prepared according to the method of steps (5) to (6), and the low-temperature brine added was 19°Bé ordinary brine at 5°C. The fermented mash was further heated to 50°C and kept warm for 48 hours, during which time compressed air was passed through and stirred for 10 minutes every 6 hours. The mash was then transferred to a press to extract the oil, thereby obtaining crude soy sauce.

[0088] The physical and chemical indicators of the experimental group and the control group during the fermentation process of the sauce mash were tracked and tested, and the free amino acid content and sensory quality of the crude oil were analyzed.

[0089] The results show that:

[0090] 1) By Figure 1 It can be seen that the total acid, amino acid nitrogen and reducing sugar in the mash are mainly formed in the early stage of fermentation, and the fermentation speed of the experimental group is faster, and the increasing rate of its physical and chemical indicators is slightly higher than that of the control group; but after 60 days of fermentation, the physical and chemical indicators tend to be stable, and there is no significant difference in the physical and chemical indicators between the experimental group and the control group.

[0091] 2) By Figure 2 The total free amino acid content in the experimental crude oil was 80.79g / kg, a 13.23% increase compared to the control group's 71.35g / kg. Umami amino acids, such as glutamic acid and aspartic acid, saw a significant increase. The glutamic acid and aspartic acid contents in the experimental crude oil were 11.77g / kg and 8.43g / kg, respectively, significantly higher than the control group's 9.46g / kg and 7.25g / kg, representing increases of 24.42% and 16.28%, respectively.

[0092] 3) By Figure 3 The experimental crude oil had a stronger umami flavor, with a sensory evaluation score of 10.92, significantly higher than the control crude oil's 9.09, representing a 20.13% improvement in umami. Furthermore, the experimental crude oil's sauce and soy sauce aroma sensory scores (12.44 and 10.37) were also significantly higher than those of the control crude oil (10.72 and 8.58), representing increases of 16.04% and 20.86%, respectively.

[0093] This result shows that during the high-salt dilute fermentation process of soy sauce, the experimental group's process method not only increases the fermentation speed of soy sauce, increases the content of umami amino acids, and improves the umami taste of the product, but also significantly improves the sauce and fermented black bean flavors of high-salt dilute soy sauce.

[0094] Example 2: Improvement of the quality of high-salt dilute soy sauce by synergistic fermentation of flavor-producing lactic acid bacteria and aroma-producing yeast

[0095] (1) Soy sauce koji making

[0096] After soybeans are selected, cleaned and soaked, they are steamed at 125℃ for 4 minutes, then immediately cooled to 35-40℃, mixed evenly with roasted wheat flour in a mass ratio of 6:4, and inoculated with 0.03% soy sauce koji essence (based on the total weight of soybeans and roasted wheat flour). The koji is cultured in an environment of 28-33℃ for 40 hours, turning the koji once every 12 hours and 18 hours respectively, until the surface of the koji is covered with yellow-green mycelium, which is the soy sauce koji.

[0097] (2) Soy sauce fermentation

[0098] The prepared soy sauce koji is mixed with 19°Bé low-temperature brine at 5°C in a mass ratio of 1:1.8. The salt content of the mixed soy sauce mash system is controlled at 17%, and the soy sauce mash temperature is controlled at around 10°C. Then, segmented temperature-variable fermentation is carried out in a closed insulated fermentation tank to simulate the temperature changes of natural fermentation.

[0099] Early stage of fermentation (0-30 days): Control the temperature of the sauce mash at 15-20℃ for low-temperature pre-fermentation, and the temperature in the center of the fermentation tank should not exceed 20℃ as much as possible; ventilate and stir for 20-30 minutes every day for 1-2 weeks before fermentation, and stir once every 4-5 days thereafter. The sauce mash should be in a uniform state without any cover or lumps.

[0100] Mid-fermentation period (30-90 days): slowly raise the temperature of the mash to 30°C, not exceeding 35°C, and maintain the temperature at 28-30°C for about 60 days, during which time the mash is aerated and stirred for 20-30 minutes every 4-5 days.

[0101] Late fermentation stage (90-120 days): continue to raise the fermentation temperature to 45°C, with a maximum of no more than 48°C, and maintain high-temperature post-ripening and aroma-enhancing fermentation at 40-45°C. During this period, ventilate and stir for 20-30 minutes every 5 days to prevent the growth of film-forming yeast on the surface; until the beans are melted, the color of the mash is dark brown, the amino acid nitrogen content of the mash is ≥1.0g / 100mL and there is no change for 1 week before and after, a mature mash is obtained.

[0102] Soy sauce fermentation was carried out according to the above operation, and 4 experimental groups were set up with the following specific conditions:

[0103] Blank group: no bacteria were inoculated.

[0104] Lactic acid bacteria group: Pediococcus acidilactici Pi Niang R036 (CDMCC No: 60617) was inoculated only in the early stage of fermentation, i.e. on the 25th day, with an inoculation size of 1×10 3 CFU / g mash.

[0105] Yeast group: Only in the middle of fermentation, when the pH of the mash dropped to 5.2, the Lushi combined yeast Pi Niang J028 (CDMCC No: 60616) was inoculated at an inoculation size of 1×10 7 CFU / g mash, and the mash needs to be aerated and stirred for 20 to 30 minutes every day for one week after yeast inoculation.

[0106] Co-fermentation group: On the 25th day of fermentation, Pediococcus acidilactici Pi Niang R036 (CDMCC No: 60617) was inoculated with an inoculum size of 1×10 3 CFU / g mash; and continue fermenting until the pH of the mash drops to 5.2, then inoculate with Lushi combined yeast Pi Niang J028 (CDMCC No: 60616) at an inoculation size of 1×10 7 CFU / g mash, and the mash needs to be aerated and stirred for 20 to 30 minutes every day for one week after yeast inoculation.

[0107] The organic acids, free amino acids and volatile flavor components in the fermented mash were detected and analyzed, and the sensory quality of the mash was evaluated. Figures 4 to 6 .

[0108] The results show:

[0109] (1) In terms of free amino acids, the total amount of free amino acids in the mash of the synergistic fermentation group increased by 19.13% compared with the blank group, especially the umami amino acid aspartic acid content increased by 52%. The total amount of free amino acids in the lactic acid bacteria group and yeast group also increased by 9.15% and 11.34% respectively compared with the blank group. In terms of organic acids, inoculation of Pediococcus acidilactici Pi Niang R036 can significantly increase the content of lactic acid and acetic acid in the mash. The lactic acid content of the synergistic fermentation group and lactic acid bacteria group increased by 170% and 180% respectively compared with the blank group, and the acetic acid content increased by 60.71% and 46.43% respectively.

[0110] (2) The synergistic fermentation of Pediococcus acidilactici Pi Niang R036 and Saccharomyces rouxii Pi Niang J028 significantly increased the content of volatile flavor substances in the mash, especially the content of esters, alcohols and acids, which increased by 134.81%, 13.58% and 129.11% respectively compared with the blank group.

[0111] (3) Compared with the blank group, the overall sensory quality of the mash in the synergistic fermentation group was significantly improved, and the flavor characteristics of the mash, such as umami, sauce aroma, caramel, honey, and fruit aroma, were enhanced. The umami sensory score increased by 35.33%, and the sauce aroma sensory score increased by 22.79%. In addition, compared with the single-strain enhanced fermentation of Pediococcus acidilactici Pi Niang R036 and Saccharomyces rouxii Pi Niang J028, the synergistic fermentation also significantly improved the overall sensory quality of the mash.

[0112] These results indicate that the synergistic fermentation of Pediococcus acidilactici Pi Niang R036 and Saccharomyces rouxii Pi Niang J028 can enhance the umami flavor of soy sauce while strengthening various aroma characteristics such as sauce aroma, honey aroma, and fruit aroma, thereby improving the quality of high-salt dilute soy sauce.

[0113] Example 3: High-temperature post-ripening fermentation enhances the flavor of high-salt dilute soy sauce and fermented black bean sauce

[0114] (1) Soy sauce koji making

[0115] After soybeans are selected, cleaned and soaked, they are steamed at 125℃ for 4 minutes, then immediately cooled to 35-40℃, mixed evenly with roasted wheat flour in a mass ratio of 6:4, and inoculated with 0.03% soy sauce koji essence (based on the total weight of soybeans and roasted wheat flour). The koji is cultured in an environment of 28-33℃ for 40 hours, turning the koji once every 12 hours and 18 hours respectively, until the surface of the koji is covered with yellow-green mycelium, which is the soy sauce koji.

[0116] (2) Soy sauce fermentation

[0117] The prepared soy sauce koji is mixed with 20°Bé low-temperature brine at 5°C in a mass ratio of 1:2, the salt content of the mixed soy sauce mash system is controlled at 18%, and then high-salt dilute fermentation is carried out at different temperatures.

[0118] The mash was prepared according to the above operation, and then fermented under three temperature modes. The specific experimental group designs are as follows:

[0119] High-temperature post-ripening group: In the early fermentation phase, maintain the mash temperature at 15-20°C for 30 days of low-temperature pre-fermentation. The center temperature of the fermenter must be kept below 20°C. Aerate and stir for 20-30 minutes daily for the first 1-2 weeks, and stir every 4-5 days thereafter. Ensure the mash is homogenous and free of any buildup or lumps. In the middle fermentation phase, slowly raise the mash temperature to 30°C, not exceeding 35°C. Maintain the fermentation temperature at 28-30°C for 60 days, aerating and stirring for 20-30 minutes every 4-5 days. In the late fermentation phase, continue raising the fermentation temperature to 45°C and perform high-temperature post-ripening and flavoring at 40-45°C for 30 days. The maximum temperature should not exceed 48°C, and aerate and stir for 20-30 minutes every 5 days to prevent the growth of surface film-forming yeast.

[0120] Room temperature fermentation group: The mash was fermented at room temperature in a closed fermentation tank for 120 days.

[0121] Temperature-adapted fermentation group: During the early fermentation phase, the mash temperature was controlled at 15-20°C for 30 days of low-temperature pre-fermentation. The center temperature of the fermentation tank must be kept below 20°C. During the first 1-2 weeks, aeration and stirring were performed daily for 20-30 minutes, and thereafter every 4-5 days. The mash was required to be uniform and free of any lids or lumps. In the later stages of fermentation, the mash temperature was slowly raised to 30°C, not exceeding 35°C, and maintained at 28-30°C for 90 days. Aeration and stirring were performed every 4-5 days for 20-30 minutes.

[0122] The results showed that high-temperature post-ripening and aroma-enhancing fermentation significantly enhanced the sauce and soy sauce flavors of high-salt, dilute fermented soy sauce. Compared to high-salt, dilute fermentation at room temperature, the sauce and soy sauce flavors increased by 13.97% and 30.47%, respectively. Compared to the mainstream moderate-temperature fermentation process, the sauce and soy sauce flavors increased by 28.24% and 57.73%, respectively.

[0123] The above-described embodiments merely represent specific implementation methods of the present application. While the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of protection of the present application. It should be noted that a person skilled in the art would be able to make numerous variations and improvements without departing from the technical concept of the present application, and all such variations and improvements fall within the scope of protection of the present application.

[0124] This background section is provided to generally present the context of the invention, and the work of the presently named inventors, the work to the extent described in this background section, and aspects of the description in this section that did not constitute prior art at the time of filing are neither explicitly nor implicitly admitted to be prior art to the present invention.

Claims

1. A method for rapidly fermenting high-flavor soy sauce by synergistically enhancing the flavor of soy sauce and black bean sauce with multiple strains, characterized in that: Flavor lactic acid bacteria were added to the soy sauce in the early stage of fermentation; flavor yeast was added to the soy sauce in the middle stage of fermentation; the inoculation amount of flavor lactic acid bacteria was 10 3 CFU / g; the inoculum size of aroma-producing yeast was 10 6 ~10 7 CFU / g; the flavor lactic acid bacteria is Pediococcus acidilactici Pi Niang R036, which is an existing strain and has been deposited in Guangdong Provincial Microbial Culture Collection Center on March 21, 2019, with a deposit number of CDMCC No: 60617; the aroma-producing yeast is Zygosaccharomyces rouxii Pi Niang J028, which is an existing strain and has been deposited in Guangdong Provincial Microbial Culture Collection Center on March 21, 2019, with a deposit number of CDMCC No: 60616; The specific method includes the following steps: (1) Making fermented black beans and soy sauce koji: soybeans or black beans, the raw materials of fermented black beans, are selected, impurities removed, and soaked, then steamed, cooled, and inoculated with soy sauce koji to make koji and culture to obtain fermented black beans; soybeans, the raw materials of soy sauce, are selected, impurities removed, and soaked, then steamed at high temperature, then immediately cooled, mixed evenly with fried wheat flour, inoculated with soy sauce koji to make koji and culture to obtain soy sauce koji; (2) Low-temperature dilute fermentation of fermented black beans and fermentation of soy sauce: Mix the fermented black bean koji with low-temperature brine and then transfer it to a temperature-controlled closed fermentation tank for low-temperature dilute fermentation; mix the soy sauce koji with low-temperature brine to control the salt content of the sauce mash, and then simulate the natural fermentation temperature change in a closed heat-insulated fermentation tank for staged temperature-variable fermentation to obtain mature sauce mash; The temperature of low-temperature dilute fermentation of fermented black beans is 15-20℃ and the time is 3-5 days; The soy sauce staged temperature-variable fermentation process comprises the following steps: In the early stage of fermentation, i.e. 0-30 days: control the temperature of the mash at 15-20℃ for low-temperature pre-fermentation, and the temperature in the center of the fermentation tank must be controlled not to exceed 20℃; aerate and stir for 20-30 minutes every day for 1-2 weeks before fermentation, and stir once every 4-5 days thereafter. The mash should be in a uniform state without any cover or lumps; on the 20th-25th day of fermentation, inoculate flavor lactic acid bacteria with an inoculation amount of 10 3 CFU / g mash; In the middle stage of fermentation, i.e. 30-90 days: slowly raise the temperature of the mash to 28-30℃ and keep it warm for 60 ± 1 days, with ventilation and stirring for 20-30 minutes every 4-5 days; when the pH of the mash is 5.0-5.2, inoculate the aroma-producing yeast with an inoculation rate of 10 6 ~10 7 CFU / g mash, and aeration and stirring for 20-30 min every day for 1 week after yeast inoculation; In the late fermentation period, i.e., 90-120 days: continue to raise the fermentation temperature to 40-45°C for high-temperature post-ripening and aroma-enhancing fermentation, during which aeration and stirring are performed for 20-30 minutes every 5 days. Mature mash is obtained when the beans are melted, the mash is dark brown, and the amino acid nitrogen content is ≥1.0 g / 100 mL and remains unchanged for 1 week. (3) Solid-liquid separation: After the low-temperature dilute fermentation of fermented black beans is completed, the koji and brine are separated to obtain solid koji and fermented brine respectively; (4) High-temperature solid-state fermentation of fermented black beans: placing the solid-state fermented black beans obtained in step (3) in a sealed heat-insulating fermentation tank for anaerobic fermentation to obtain a finished fermented black beans product; the temperature for anaerobic fermentation of the solid-state fermented black beans is 45-50° C., and the anaerobic fermentation time is 2-3 months; (5) Soaking and pressing: adding the fermented fermented black beans obtained in step (4) to the mature mash obtained in step (2), then performing heat preservation and soaking extraction, and finally conveying the material to a press to press and extract oil, thereby obtaining a soy sauce crude oil having both soy sauce and black bean flavors; The addition amount of fermented black beans is 5%~10% of the mass of mature mash; the temperature of heat preservation and soaking extraction is 45~50℃, and the time is 24~48 hours, during which compressed air is stirred for 10 minutes every 6 hours.

2. A method for rapidly fermenting a multi-strain synergistically enhancing the flavor of high-flavor soy sauce and fermented black bean sauce according to claim 1, characterized in that: In step (1), the amount of soy sauce koji added in the fermented black bean koji preparation is 0.03% based on the dry weight of the raw material; the culture environment is 28-33°C for 48 hours, and the koji is turned once every 12-18 hours, and finally the surface of the koji is covered with yellow-green mycelium.

3. A method for rapidly fermenting a multi-species synergistically enhancing the flavor of high-flavor soy sauce and fermented black bean sauce according to claim 1, characterized in that: In step (1), the amount of soy sauce koji added in the soy sauce koji making process is 0.03% based on the total weight of soybeans and roasted wheat flour; the culture environment is 28-33°C for 40-45 hours, and the koji is turned over at 12 hours and 24 hours respectively.

4. A method for rapidly fermenting a multi-strain synergistically enhancing the flavor of high-flavor soy sauce and fermented black bean sauce according to claim 1, characterized in that: In the low-temperature dilute fermentation of fermented black beans in step (2), the temperature of the low-temperature brine is 5-10°C, and the brine concentration is 18-19°Bé; the fermented black bean koji and the low-temperature brine are mixed in a mass ratio of 1:1.5-2; during the low-temperature dilute fermentation of fermented black beans, the fermented black beans are stirred once every morning and evening, each time for 10-20 minutes.

5. A method for rapidly fermenting a multi-strain synergistically enhancing the flavor of high-flavor soy sauce and fermented black bean sauce according to claim 1, characterized in that: In the soy sauce fermentation of step (2), the temperature of the low-temperature brine is 5-10°C, and the brine concentration is 19-20°Bé; the soy sauce koji and the low-temperature brine are mixed in a mass ratio of 1:1.8-2, and the salt content of the soy sauce mash is controlled to be 16.5%-18%, and the temperature of the soy sauce mash is 10-15°C.

6. A method for rapidly fermenting a multi-strain synergistically enhancing the flavor of high-flavor soy sauce and fermented black bean sauce according to claim 1, characterized in that: In the step (5), the amino acid nitrogen content in the crude oil is ≥1.0 g / 100 g.

Images