Method for naturally extracting freshness of soy sauce based on enriched acylamino acid

Abstract

The invention discloses a method for naturally extracting freshness of soy sauce based on enriched acid acyl amino acid, which comprises the following steps: mixing soybean meal powder and wheat germ powder into water, cooking at high temperature to obtain fermentation liquor, inoculating zygosaccharomyces rouxii, and performing sterile fermentation and freeze-drying to obtain yeast extract powder; adding yeast extract powder into the raw soy sauce, and continuing to ferment for 1-5 days under the same conditions to obtain zinc-rich and coenzyme A-rich soy sauce; adding reverse hydrolase and forward acyltransferase into the zinc-rich and coenzyme A-rich soy sauce, treating at 45-60 DEG C for 2 hours, and treating at 70-75 DEG C for 2 hours to obtain enzymatic soy sauce; and adding yeast extract powder into the enzymatic soy sauce, and heating and sterilizing to obtain the cooked soy sauce.

Description

Technical Field

[0001] The invention belongs to the technical field of soy sauce brewing, and particularly relates to a method for naturally enhancing the flavor of soy sauce based on enrichment of acid acylamino acids. Background Art

[0002] Reduced-salt soy sauce is a popular choice for consumers seeking health benefits. However, the fermentation of traditional high-salt soy sauce often results in a deterioration in flavor. For example, after salt reduction, excessive metabolism of lactic acid bacteria leads to the accumulation of organic acids, which acidifies the sauce. Furthermore, natural spoilage bacteria colonize the sauce, producing alkaloids, biogenic amines, and other bittering agents.

[0003] To address the flavor challenges of reduced-salt soy sauce, researchers in the field have sought to engineer the bacteria that produce it to resemble traditional, high-salt soy sauce. However, the significant difference in salt content makes replicating the bacterial flora and flavor profile extremely challenging. Nitrogen-rich substrates, flavor additives (based on flavor-mouthfeel synergy), or emulsifiers have been used to enhance the flavor of reduced-salt soy sauce, but clean label requirements and cost constraints have limited their practical application. Taste adjustment requires a strategy based on the unique properties of reduced-salt soy sauce, rather than blindly approximating and replicating the parameters of traditional, high-salt soy sauce.

[0004] Therefore, there is an urgent need in the art for a method for improving the taste of reduced-salt soy sauce that is effective, low-cost, does not require the introduction of additional chemical reagents, and is consistent with the characteristics and production process of the original reduced-salt soy sauce. Summary of the Invention

[0005] The purpose of this section is to summarize some aspects of embodiments of the present invention and to briefly introduce some preferred embodiments.

[0006] In view of the above problems and / or the problems existing in the prior art, the present invention is proposed.

[0007] Therefore, the purpose of the present invention is to overcome the deficiencies in the prior art and provide a method for naturally enhancing the flavor of soy sauce based on enrichment of acid acylamino acids.

[0008] In order to solve the above technical problems, the present invention provides the following technical solution: a method for naturally enhancing the flavor of soy sauce based on enrichment of acid acylamino acids, comprising: Mix soybeans, roasted wheat, pumpkin seed powder, and chia seed powder, steam at high temperature, cool until inoculated with Aspergillus oryzae, and turn the mixture over 8 and 24 hours after inoculation until magnesium-rich Daqu is obtained 36 hours later. Mixing magnesium-rich Daqu with salt water to obtain a mash, which is naturally fermented at 30°C with stirring once a day for 60 to 80 days, and then inoculated with Lactobacillus plantarum and Zygosaccharomyces rouxii. Fermentation is continued under the same conditions for 10 to 30 days to obtain raw soy sauce. Soybean meal powder and wheat germ powder were mixed in water and cooked at high temperature to obtain a mixture, which was then inoculated with Zygosaccharomyces rouxii and sterilized at high density at 30°C for 2-5 days. The fermentation liquid was crushed at 2.0 kBar for 20 minutes and freeze-dried at -90°C to obtain yeast extract powder. Adding yeast extract powder to raw soy sauce and continuing fermentation under the same conditions for 1 to 5 days to obtain zinc-rich and coenzyme A-rich soy sauce; After adding reverse hydrolase to zinc-rich and coenzyme A-rich soy sauce and treating it at 45-60℃ for 2 h, forward acyltransferase was added and treated at 70-75℃ for 2 h to obtain enzymatic soy sauce. Yeast extract powder is added to the enzymatic soy sauce, and the temperature is raised to 90-100°C and sterilized for 30-60 minutes to obtain cooked soy sauce.

[0009] As a preferred embodiment of the method of the present invention, soybeans, roasted wheat, pumpkin seed powder and chia seed powder are mixed, wherein, by weight, the soybeans are 5 to 10 parts, the roasted wheat is 1 to 5 parts, the pumpkin seed powder is 0.1 to 1 part, and the chia seed powder is 0.1 to 1 part.

[0010] As a preferred embodiment of the method of the present invention, the high-temperature cooking and cooling are followed by inoculation with Aspergillus oryzae, wherein the cooking temperature is 121° C., the pressure is 0.1 MPa, the time is 20 min, the Aspergillus oryzae is Aspergillus oryzae 3.042, and the inoculation amount of Aspergillus oryzae relative to the mass percentage of the material after high-temperature cooking is 0.3%wt.

[0011] As a preferred embodiment of the method of the present invention, the magnesium-rich Daqu is mixed with brine, wherein, by weight, the magnesium-rich Daqu is 1 to 2 parts, the brine is 3 to 5 parts, and the concentration of the brine is 10% w:v.

[0012] As a preferred embodiment of the method of the present invention, wherein: the inoculation of Lactobacillus plantarum and Zygosaccharomyces rouxii, wherein the inoculation amount of Lactobacillus plantarum is 10 4 ~10 6 CFU / mL mash, the inoculum size of Zygosaccharomyces rouxii was 10 4 ~10 6 CFU / mL mash.

[0013] As a preferred embodiment of the method of the present invention, the soybean meal powder and the wheat germ powder are mixed into water and cooked at high temperature, wherein the soybean meal powder is 1 to 10 parts, the wheat germ powder is 1 to 10 parts, the water is 50 to 100 parts, the high temperature cooking temperature is 121°C, the pressure is 0.1 MPa, and the time is 20 minutes; the inoculation of Zygosaccharomyces rouxii, wherein the inoculation amount is 10 8 CFU / mL fermentation broth.

[0014] As a preferred embodiment of the method of the present invention, wherein: the yeast extract powder is added to the raw soy sauce, and the added amount of the yeast extract powder is 0.1-1%wt.

[0015] As a preferred embodiment of the method of the present invention, the reverse hydrolase is composed of papain, thermolysin, fungal acid protease, etc. in a mass ratio, and the amount of the reverse hydrolase added is 0.01-0.1%wt; The forward acyltransferase is composed of acyltransferase and transglutaminase in a certain weight ratio, and the added amount of the forward acyltransferase is 0.01-0.1%wt.

[0016] As a preferred embodiment of the method of the present invention, wherein: yeast extract powder is added to the enzymatic soy sauce, wherein the amount of yeast extract powder added is 0.01-0.1%wt.

[0017] Beneficial effects of the present invention: (1) In the early stage of fermentation, the natural fermentation substrate is enriched with magnesium ions to catalyze the active acylation of amino acids mediated by the microbial enzyme system; the chia seed polysaccharide gel has a nutrient-sustaining effect, which prevents the produced acid-acylamino acids from being used as nutrients; the catalyst pre-fermentation of the invention increases the level of β-citrol-L-glutamate by 0.029 mM.

[0018] (2) The strains selected for bio-enrichment in the present invention are the original soy sauce strains Lactobacillus plantarum and Zygosaccharomyces rouxii, and the organic acids metabolized by them and the amino acids overflowed are important substrates of acid-acylamino acids; when the biomass of Lactobacillus plantarum is much higher than that of Zygosaccharomyces rouxii, the co-fermentation process enriches 0.052 mM and 0.105 mM more β-citrol-L-glutamate and N-lactol-L-phenylalanine, respectively, than the control group without artificial intervention.

[0019] (3) In the late fermentation stage, the present invention supplements the natural fermentation catalytic factors zinc and coenzyme A from food and yeast sources to promote the efficiency of enzymatic and thermal synthesis of acylamino acids during the post-processing of soy sauce, while protecting the generated acylamino acids from being reused as nutrients by the fermentation bacteria. This process focuses on increasing N-lactyl-L-phenylalanine by 0.094 mM.

[0020] (4) The enzymes selected for enzymatic synthesis in the present invention are composed of a forward acyltransferase and a reverse hydrolase. The forward acyltransferase transfers the acyl group directly to the amino acid, while the reverse hydrolase mediates the reverse reaction of peptide bond cleavage, which is beneficial to the formation of carbon-nitrogen amide bonds. The forward acyltransferase and the reverse hydrolase preferentially catalyze the synthesis of β-citrate-L-glutamate and N-lactate-L-phenylalanine, respectively. These two types of enzymes increase the content of β-citrate-L-glutamate and N-lactate-L-phenylalanine by 0.061 mM and 0.112 mM, respectively. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following briefly introduces the drawings required for describing the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. Those skilled in the art can also derive other drawings based on these drawings without inventive effort. Among them: Figure 1 Figure 2 shows the differences in the content of various acylamino acids (left) and the sensory evaluation of soy sauce (right) in the examples of the present invention. Lac, Ace, Cit, and Pro represent lactoyl, acetyl, citric acid, and propionic acid, respectively. Amino acid groups are represented by three letters.

[0022] Figure 2 This is a sensory evaluation difference diagram of Examples 1, 2, 4, and 5 of the present invention. DETAILED DESCRIPTION

[0023] In order to make the above-mentioned objects, features and advantages of the present invention more obvious and easy to understand, the specific implementation methods of the present invention are described in detail below in conjunction with the embodiments of the specification.

[0024] Raw materials used in the present invention: Pumpkin seed powder: Jiangxi Haisiyi Food Co., Ltd. Chia seed powder: Xi'an Dongfeng Biotechnology Co., Ltd. Soybean meal powder: Shandong Tangzheng Biotechnology Co., Ltd.: Wheat germ powder: Henan Kunhua Biotechnology Co., Ltd. Aspergillus oryzae Shanghai Brewing 3.042: Culture Collection Center of Tianjin University of Science and Technology; Lactobacillus plantarum CS3 (CCTCCNO: M2021532), Zygosaccharomyces rouxii GT-C8 (CCTCC NO: M2024426): China Center for Type Culture Collection; Acyltransferase (also known as acylase) A114166 enzyme activity ≥30000 u / g, transglutaminase T777729 enzyme activity 120 u / g: Shanghai Aladdin Biochemical Technology Co., Ltd.; Thermolysin S10236 enzyme activity ≥50 u / mg protein: Shanghai Yuanye Biotechnology Co., Ltd.; Papain, enzyme activity ≥10 u / mg protein, acid protease, enzyme activity ≥10 u / mg protein: Nanning Dongheng Huadao Biotechnology Co., Ltd. Other raw materials are commercially available unless otherwise specified.

[0025] Example 1 This embodiment provides a method for enriching acid acylamino acids to improve the taste of reduced-salt soy sauce, the main steps comprising: (1) Catalytic factor pre-fermentation By weight, 8 parts of soybeans, 2 parts of roasted wheat, 0.6 parts of pumpkin seed powder, and 0.4 parts of chia seed powder were mixed and steamed at 121°C and 0.1 MPa for 20 min. After cooling to 40°C, 0.3%wt Aspergillus oryzae 3.042 was inoculated. The material was turned over 8 and 24 h after inoculation, and magnesium-rich Daqu was obtained 36 h later.

[0026] (2) Bioaccumulation in brewing By weight, 1 part of magnesium-enriched Daqu was mixed with 3 parts of 10% (w: v) brine to form a mash, which was naturally fermented at 30°C with stirring once a day for 70 days. Vaccination 10 6 CFU / mL mash of Lactobacillus plantarum CS3 (CCTCCNO: M2021532) and 10 5 CFU / mL mash was fermented with Zygosaccharomyces rouxii GT-C8 (CCTCC NO: M 2024426) under the same conditions for 20 days to obtain raw soy sauce.

[0027] (3) Catalytic factor replenishment By weight, 5 parts of soybean meal powder and 10 parts of wheat germ powder were mixed into 100 parts of water and steamed at 121℃ and 0.1 MPa for 20 minutes. 8 CFU / mL fermentation broth of Zygosaccharomyces rouxii GT-C8 (CCTCC NO: M 2024426) was aseptically fermented at high density at 30°C for 3 days. The fermentation broth was crushed at 2.0 kBar for 20 min and freeze-dried at -90°C to obtain yeast extract powder. 0.1% wt yeast extract powder was added to raw soy sauce and fermented under the same conditions for another 2 days to obtain zinc-rich and CoA-rich soy sauce.

[0028] (4) Enzymatic enrichment after brewing The zinc-rich and coenzyme A-rich soy sauce was further supplemented with 0.05% wt of a reverse hydrolase composed of papain, thermolysin, and fungal acid protease in equal proportions, and 0.1% wt of a forward acyltransferase composed of acyltransferase and transglutaminase in equal proportions; after treatment at 48°C for 2 h and then at 72°C for 2 h, the enzymatic soy sauce was obtained.

[0029] (5) Spontaneous enrichment by heat sterilization Continue to add 0.5%wt yeast extract powder, heat to 95℃ and sterilize for 30 minutes to obtain the final cooked soy sauce.

[0030] Example 2 (Ordinary Reduced-Salt Soy Sauce Not Enriched with Acid Acylamino Acids) This embodiment provides a method for preparing salt-reduced soy sauce without enrichment of acid acylamino acids, comprising: 8 parts of soybeans and 2 parts of fried wheat were mixed by weight, steamed at 121°C and 0.1 MPa for 20 min, cooled to 40°C and inoculated with 0.3%wt Aspergillus oryzae 3.042. The material was turned over 8 and 24 h after inoculation, and Daqu was obtained 36 h later.

[0031] By weight, 1 part of Daqu is mixed with 3 parts of 10% (w: v) brine to form a mash, which is naturally fermented at 30°C for 90 days with stirring once a day to obtain raw soy sauce.

[0032] The raw soy sauce is heated to 95°C and sterilized for 30 min to obtain the final cooked soy sauce.

[0033] Example 3 (Ordinary high-salt soy sauce not enriched with acid acylamino acids) Compared with Example 2, the concentration of the brine used in this example is 18% (w: v).

[0034] Other preparation process conditions are the same as those in Example 2.

[0035] Acylamino acid content was determined using ultra-performance liquid chromatography-orbitrap tandem mass spectrometry (UPLC-Orbitrap-MS / MS, Waters, Manchester, UK). The aqueous fractions were separated using an HSS T3 column (100 mm × 2.1 mm, 1.7 μm particles) on a Waters Acquity UPLC system.

[0036] The mobile phase was a linear gradient of 0.1% formic acid in water (A) and acetonitrile (B): 0 min, 1% B; 11 min, 40% B; 13 min, 70% B; 15-18 min, 99% B; 19-22 min, 1% B.

[0037] Parameter setting: flow rate 400 μL·min-1 ; Column temperature 40 ℃; Injection volume 3 μL.

[0038] Mass spectra were acquired in positive and negative ion modes using a heated electrospray ionization (HESI) source. Full MS scans (100–1500 m / z, 70,000 full width half maximum (FWHM) at 200 m / z) and data-dependent MS / MS acquisition were performed. A mixed standard solution was serially diluted (1–128 mM, 2ⁿ gradient) to generate concentration-response curves under the same conditions for quantitative analysis.

[0039] Sensory evaluation: The sensory evaluation test procedures for soy sauce in each example refer to T / CNFIA 213-2024 Guidelines for Sensory Evaluation of Condiments (Soy Sauce); The standard taste solutions included citric acid (0.18, 0.36, 0.72 g / L, sour), sucrose (3.6, 7.2, 14.4 g / L, sweet), caffeine (0.09, 0.18, 0.36 g / L, bitter), sodium chloride (0.5, 1.0, 2.0 g / L, salty), monosodium glutamate (0.07, 0.14, 0.28 g / L, umami), and glutathione (0.5, 1.0, 2.0 g / L, strong), and the concentration gradient corresponded to the sensory intensity score (1, 5, 9).

[0040] The acid acylamino acid enrichment effect of Examples 1 to 3 is as follows: Figure 1 As shown on the left, β-citrol-L-glutamic acid and N-lactoyl-L-phenylalanine, characteristic amino acids and amino acids in reduced-salt soy sauce, are increased by 0.093 mM and 0.147 mM, respectively, significantly exceeding levels in traditional high-salt soy sauce. This enhancement of these two amino acids and amino acids stems from the accumulation of citric and lactic acids during salt-reduced fermentation, which in turn enhances the dehydration condensation reaction between the carboxyl groups of the acids and the amino groups of the amino acids.

[0041] The invention mentions two steps of "pre-fermentation and post-fermentation" to enrich catalytic factors, and three steps of "biocatalysis, enzymatic catalysis, and thermal catalysis" in situ catalysis steps, all of which are aimed at promoting the forward progress of the above-mentioned dehydration condensation reaction.

[0042] Sensory evaluation of the soy sauces of Examples 1 to 3 revealed that the invention made the umami and kokumi of ordinary reduced-salt soy sauce more intense (around 9 points), while the sourness (6.3 points dropped to 4 points) and bitterness (3.8 points dropped to 2 points) were significantly weakened.

[0043] Example 4 (Conventional Reduced-Salt Soy Sauce with Addition of β-Citroyl-L-Glutamic Acid) Compared with Example 2, the preparation process conditions of this example are the same as those of Example 2. However, 0.093 mM β-citrol-L-glutamic acid is added to the final cooked soy sauce.

[0044] Example 5 (Conventional Reduced-Salt Soy Sauce with Addition of N-Lactoyl-L-Phenylalanine) Compared with Example 2, the preparation process conditions of this example are the same as those of Example 2. However, 0.147 mM N-lactyl-L-phenylalanine is added to the final cooked soy sauce.

[0045] Figure 2 β-Citroyl-L-glutamate (left) and N-lactoyl-L-phenylalanine (right) have been shown to act as flavor modifiers in conventional reduced-salt soy sauce. While both enhance umami and thickness, β-Citroyl-L-glutamate is more effective at reducing acidity, while N-lactoyl-L-phenylalanine is more effective at reducing bitterness. These compounds carry groups similar to sour and bitter components, competing for binding with the corresponding taste receptors. However, due to their acid-acyl modification, they are unable to activate the receptors.

[0046] Example 6 (No enrichment of catalytic factors in the pre-fermentation) Compared with Example 1, the pre-fermentation steps in this example are as follows: 8 parts of soybeans and 2 parts of fried wheat were mixed by weight, steamed at 121°C and 0.1 MPa for 20 min, cooled to 40°C and inoculated with 0.3%wt Aspergillus oryzae 3.042. The material was turned over 8 and 24 h after inoculation, and Daqu was obtained 36 h later.

[0047] Other process conditions are the same as those in Example 1.

[0048] Example 7 (Post-fermentation process without catalyst) Compared with Example 1, this example does not perform catalytic factor supplementation, but directly uses raw soy sauce for the subsequent post-brewing enzymatic enrichment step.

[0049] Other process conditions are the same as those in Example 1.

[0050] Example 8 (Lactobacillus + Lu Shi co-fermentation not used) Compared with Example 1, this example does not perform bio-enrichment: By weight, 1 part of magnesium-enriched Daqu is mixed with 3 parts of 10% (w: v) brine to form a mash, which is naturally fermented at 30°C for 90 days with stirring once a day to obtain raw soy sauce.

[0051] Other process conditions are the same as those in Example 1.

[0052] Example 9 (Lactobacillus:Roche is 1:1) Compared with Example 1, the method of bioaccumulation in brewing in this example is as follows: By weight, 1 part of magnesium-enriched Daqu was mixed with 3 parts of 10% (w: v) brine to form a mash, which was naturally fermented at 30°C with stirring once a day for 70 days. Vaccination 10 5 CFU / mL mash of Lactobacillus plantarum CS3 (CCTCCNO: M2021532) and 10 5 CFU / mL mash was fermented with Zygosaccharomyces rouxii GT-C8 (CCTCC NO: M 2024426) under the same conditions for 20 days to obtain raw soy sauce.

[0053] Other process conditions are the same as those in Example 1.

[0054] Example 10 (Lactobacillus:Roche is 1:10) Compared with Example 1, the method of bioaccumulation in brewing in this example is as follows: By weight, 1 part of magnesium-rich Daqu was mixed with 3 parts of 10% (w: v) brine to form a mash, which was naturally fermented at 30°C with stirring once a day for 70 days. 5 CFU / mL mash of Lactobacillus plantarum CS3 (CCTCCNO: M2021532) and 10 6 CFU / mL mash was fermented with Zygosaccharomyces rouxii GT-C8 (CCTCC NO: M 2024426) under the same conditions for 20 days to obtain raw soy sauce.

[0055] Other process conditions are the same as those in Example 1.

[0056] Example 11 (without reverse hydrolase catalysis) Compared with Example 1, the steps of enzymatic enrichment after brewing in this example are as follows: Zinc-enriched and CoA-enriched soy sauce was supplemented with 0.1% wt of a forward acyltransferase consisting of acyltransferase and transglutaminase in equal weight ratios. The mixture was treated at 48°C for 2 h and then at 72°C for 2 h to obtain enzymatic soy sauce.

[0057] Other process conditions are the same as those in Example 1.

[0058] Example 12 (without using forward acyltransferase) Compared with Example 1, the steps of enzymatic enrichment after brewing in this example are as follows: Zinc-enriched and coenzyme A-enriched soy sauce was supplemented with 0.05% wt of a reverse hydrolase consisting of papain, thermolysin, and fungal acid protease in equal weight ratios. After treatment at 48°C for 2 h and then at 72°C for 2 h, enzymatic soy sauce was obtained.

[0059] Other process conditions are the same as those in Example 1.

[0060] Table 1 Differences in the contents of β-citrol-L-glutamic acid and N-lactrol-L-phenylalanine in Examples 1, 6-12 Comparison of Examples 1-2 and 6-7 shows that the two catalyst enrichment steps are beneficial to the production of two acylamino acids. Among them, the pre-fermentation of pumpkin seed powder and chia seeds is beneficial to the enrichment of magnesium ions. Magnesium ions can stabilize the catalytic conformation based on the chemical properties of Lewis acids, which is beneficial to the acyl transfer forward reaction of β-citrol-L-glutamate. Whether in biological metabolism or enzymatic reaction, the catalyst pre-fermentation increased β-citrol-L-glutamate by 0.029 mM. The catalyst post-fermentation is designed to obtain zinc ion catalysts from soybean meal and wheat germ powder, as well as yeast extract as substrates for acylamino acid synthesis, while protecting the generated acylamino acids from being reused as nutrients by fermentation bacteria. This process focuses on increasing N-lactyl-L-phenylalanine by 0.094 mM.

[0061] Comparison of Examples 1-2 and 8-10 shows that co-fermentation with a significantly higher Lactobacillus plantarum biomass than Zygosaccharomyces rouxii facilitates the enrichment of both acid-acylamino acids. This is because acylation of organic acids onto amino acids requires a sufficient concentration of organic acid, which in turn requires a larger population of organic acid-metabolizing bacteria. This process enriched 0.052 mM more β-citrol-L-glutamate and 0.105 mM more N-lactoyl-L-phenylalanine, respectively, than natural fermentation without human intervention.

[0062] Comparison of Examples 1-2 and 11-12 shows that the forward acyltransferase and reverse hydrolase preferentially catalyze the synthesis of β-citrol-L-glutamate and N-lactrol-L-phenylalanine, respectively, and these two types of enzymes increase the contents of β-citrol-L-glutamate and N-lactrol-L-phenylalanine by 0.061 mM and 0.112 mM, respectively.

[0063] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solutions of the present invention may be modified or replaced by equivalents without departing from the spirit and scope of the technical solutions of the present invention, and all of these should be included in the scope of the present invention.

Claims

1. A method for naturally enhancing the flavor of soy sauce based on enrichment of acid acylamino acids, characterized in that: include, Mix soybeans, roasted wheat, pumpkin seed powder, and chia seed powder, steam at high temperature, cool until inoculated with Aspergillus oryzae, and turn the mixture over 8 and 24 hours after inoculation until magnesium-rich Daqu is obtained 36 hours later. Mixing magnesium-rich daqu with salt water to obtain a mash, which is naturally fermented at 30°C with stirring once a day for 60 to 80 days, and then inoculated with Lactobacillus plantarum and Zygosaccharomyces rouxii. Fermentation is continued under the same conditions for 10 to 30 days to obtain raw soy sauce. Soybean meal powder and wheat germ powder were mixed in water and cooked at high temperature to obtain a mixture, which was then inoculated with Zygosaccharomyces rouxii and sterilized at high density at 30°C for 2-5 days. The fermentation liquid was crushed at 2.0 kBar for 20 minutes and freeze-dried at -90°C to obtain yeast extract powder. Adding yeast extract powder to raw soy sauce and continuing fermentation under the same conditions for 1 to 5 days to obtain zinc-rich and coenzyme A-rich soy sauce; After adding reverse hydrolase to zinc-rich and coenzyme A-rich soy sauce and treating it at 45-60℃ for 2 h, forward acyltransferase was added and treated at 70-75℃ for 2 h to obtain enzymatic soy sauce. Yeast extract powder is added to the enzymatic soy sauce, and the temperature is raised to 90-100°C and sterilized for 30-60 minutes to obtain cooked soy sauce.

2. The method according to claim 1, wherein: The soybeans, fried wheat, pumpkin seed powder and chia seed powder are mixed, wherein, by weight, the soybeans are 5 to 10 parts, the fried wheat is 1 to 5 parts, the pumpkin seed powder is 0.1 to 1 part, and the chia seed powder is 0.1 to 1 part.

3. The method according to claim 1 or 2, wherein: The high-temperature cooking and cooling are followed by inoculation with Aspergillus oryzae, wherein the cooking temperature is 121° C., the pressure is 0.1 MPa, the time is 20 min, the Aspergillus oryzae is Aspergillus oryzae 3.042, and the inoculation amount of Aspergillus oryzae is 0.3%wt relative to the mass percentage of the material after high-temperature cooking.

4. The method according to claim 3, wherein: The magnesium-rich Daqu is mixed with brine, wherein, by weight, the magnesium-rich Daqu is 1 to 2 parts, the brine is 3 to 5 parts, and the concentration of the brine is 10% w: v.

5. The method according to claim 1 or 4, wherein: The inoculation amount of plant lactobacillus and zygosaccharomyces rouxii is 10 4 ~10 6 CFU / mL mash, the inoculum size of Zygosaccharomyces rouxii was 10 4 ~10 6 CFU / mL mash.

6. The method according to claim 1, wherein: The soybean meal powder and wheat germ powder are mixed into water and cooked at high temperature, wherein the soybean meal powder is 1-10 parts, the wheat germ powder is 1-10 parts, the water is 50-100 parts, the high temperature cooking temperature is 121°C, the pressure is 0.1 MPa, and the time is 20 minutes; the inoculation of Zygosaccharomyces rouxii, wherein the inoculation amount is 10 8 CFU / mL fermentation broth.

7. The method according to claim 1 or 6, wherein: The yeast extract powder is added to the raw soy sauce, and the added amount of the yeast extract powder is 0.1-1%wt.

8. The method according to claim 1, wherein: The reverse hydrolase is composed of papain, thermolysin, fungal acid protease and the like in a mass ratio, and the addition amount of the reverse hydrolase is 0.01-0.1%wt; The forward acyltransferase is composed of acyltransferase and transglutaminase in a certain weight ratio, and the added amount of the forward acyltransferase is 0.01-0.1%wt.

9. The method according to claim 1 or 8, wherein: Yeast extract powder is added to the enzymatic soy sauce, wherein the amount of yeast extract powder added is 0.01-0.1%wt.

10. The method according to claim 1, wherein: The enriched taste-modulating acylamino acids include β-citrol-L-glutamic acid and N-lactyl-L-phenylalanine.

Images