Method for high-yield production of gamma-aminobutyric acid by mixed fermentation

Abstract

This invention belongs to the fields of fermentation engineering and enzyme engineering technology, specifically relating to a method for high-yield γ-aminobutyric acid (GABA) production through mixed fermentation. Specifically, this invention employs a mixed fermentation method, promoting the growth of *Lactobacillus hessei* through the growth of *Bifidobacterium*, achieving a synergistic effect between the two strains and thus significantly increasing GABA yield. Experimental results show that compared with traditional single-strain fermentation, the mixed fermentation method can increase GABA yield by 2-3 times and shorten the fermentation cycle by more than 50%, demonstrating significant technical advantages and economic benefits. Therefore, the mixed fermentation method proposed in the above technical solution provides an efficient and environmentally friendly new approach for high-yield GABA production, possessing significant application value and market prospects.

Description

Technical Field

[0001] This invention belongs to the fields of fermentation engineering and enzyme engineering technology, and specifically relates to a method for high-yield γ-aminobutyric acid through mixed fermentation. Background Technology

[0002] The information disclosed in the background section of this invention is intended only to enhance the understanding of the overall background of the invention and is not necessarily to be construed as an admission or in any way implying that such information constitutes prior art known to those skilled in the art.

[0003] Gamma-aminobutyric acid (GABA) is an important non-protein amino acid widely found in animals, plants, and microorganisms, possessing a variety of biological activities. In the central nervous system, GABA acts as a major inhibitory neurotransmitter, participating in the regulation of neuronal excitability and playing a crucial role in maintaining the normal function of the nervous system. Furthermore, GABA also exhibits physiological effects such as anti-anxiety, blood pressure reduction, sleep improvement, and promotion of brain function recovery, thus showing broad application prospects in the pharmaceutical, food, and functional beverage industries. Currently, GABA production methods mainly include chemical synthesis and bio-fermentation. While chemical synthesis is a mature process, it suffers from high costs, significant environmental pollution, and low product purity. In contrast, bio-fermentation, due to its environmental friendliness, low cost, and high product purity, is gradually becoming the mainstream method for GABA production.

[0004] In biological fermentation, Lactobacillus hesitantus ( Lactobacillus hilgardii Lactobacillus hessler is one of the most commonly used strains. It possesses a unique metabolic pathway, directly converting glutamate to γ-aminobutyric acid (GABA) via transamination. This biotransformation process is not only highly efficient but also requires no complex chemical reagents, thus it is widely used in GABA production. However, despite the significant advantages of Lactobacillus hessler in GABA production, traditional single-strain fermentation methods still have some obvious limitations. First, single-strain fermentation often results in low GABA yields. This is mainly because the growth of a single strain is greatly affected by environmental factors, such as pH, temperature, and nutrient concentration; changes in these factors significantly affect strain growth and GABA production. Second, single-strain fermentation has a long fermentation cycle, typically requiring a considerable amount of time to reach the desired GABA yield. This not only increases production costs but also limits the improvement of production efficiency. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides a method for high-yield γ-aminobutyric acid (GABA) production through mixed fermentation. This invention screened and obtained a strain of *Bifidobacterium lactis*, which, when mixed with *Lactobacillus hessieri* (a GABA-producing bacterium), significantly increased the GABA production of *Lactobacillus hessieri*. Experiments have shown that *Bifidobacterium lactis* can grow rapidly during fermentation, producing metabolites such as lactic acid. These metabolites not only lower the pH of the fermentation broth and inhibit the growth of harmful bacteria, but also promote the growth of *Lactobacillus hessieri* and GABA synthesis through competitive inhibition and metabolic regulation. Furthermore, *Bifidobacterium lactis* can further enhance GABA production by producing enzymes such as glutamate decarboxylase (GAD). Based on these research findings, this invention is thus completed.

[0006] To achieve the above-mentioned technical objectives, the present invention relates to the following technical solutions:

[0007] The first aspect of the present invention provides a strain of Bifidobacterium lactis ( Bifidobacterium lactis ga-2, deposited at the China Center for Type Culture Collection (address: Wuhan University, Luojia Mountain, Wuchang, Wuhan, Hubei Province), on May 26, 2025, with accession number CCTCC NO: M 20251181.

[0008] A second aspect of the present invention provides the use of the above-mentioned Bifidobacterium lactis ga-2 in any one or more of the following:

[0009] (a) Promotes the growth of Lactobacillus schrei;

[0010] (b) Promotes the production of γ-aminobutyric acid by Lactobacillus hesitantus.

[0011] A third aspect of the present invention provides a microbial composition comprising the aforementioned Bifidobacterium lactis ga-2 and Lactobacillus hesitantii.

[0012] A fourth aspect of the present invention provides the use of the above-described microbial composition in the production of γ-aminobutyric acid.

[0013] A fifth aspect of the present invention provides a method for producing γ-aminobutyric acid by fermentation, the method comprising:

[0014] The activated Bifidobacterium lactis ga-2 was mixed with Lactobacillus hessier and inoculated into a culture medium, and sodium glutamate was added for fermentation culture to produce γ-aminobutyric acid.

[0015] Furthermore, the method includes:

[0016] S1. Select single colonies of Bifidobacterium lactis ga-2 and Lactobacillus spp. respectively into the first culture medium for a first activation culture, and then select single colonies of the activated Bifidobacterium lactis ga-2 and Lactobacillus spp. respectively into the second culture medium for a second activation culture.

[0017] S2. After the secondary activated Lactobacillus hirsutus is inoculated into the third culture medium for fermentation culture for a period of time, Bifidobacterium lactis is inoculated, and sodium glutamate is added in a fed-batch manner to continue fermentation culture to produce γ-aminobutyric acid.

[0018] The beneficial technical effects of one or more of the above technical solutions are as follows:

[0019] The above-mentioned technical solution employs a mixed fermentation method, promoting the growth of *Lactobacillus hesperidin* through the growth of *Bifidobacterium*, achieving a synergistic effect between the two strains and thus significantly increasing GABA yield. Experimental results show that compared with traditional single-strain fermentation, the mixed fermentation method can increase GABA yield by 2-3 times and shorten the fermentation cycle by more than 50%, demonstrating significant technical advantages and economic benefits. Therefore, the mixed fermentation method proposed in this technical solution provides an efficient and environmentally friendly new approach for high-yield GABA production, possessing significant application value and market prospects. Detailed Implementation

[0020] It should be noted that the following detailed descriptions are exemplary and intended to provide further illustration of the invention. Unless otherwise specified, all technical and scientific terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0021] It should be noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the exemplary embodiments according to the present invention. As used herein, the singular form is intended to include the plural form as well, unless the context clearly indicates otherwise. Furthermore, it should be understood that when the terms "comprising" and / or "including" are used in this specification, they indicate the presence of features, steps, operations, devices, components, and / or combinations thereof.

[0022] In a typical specific embodiment of the present invention, a strain of Bifidobacterium lactis ( Bifidobacterium lactis ga-2, deposited at the China Center for Type Culture Collection (address: Wuhan University, Luojia Mountain, Wuchang, Wuhan, Hubei Province), on May 26, 2025, with accession number CCTCC NO: M 20251181.

[0023] A second aspect of the present invention provides the use of the above-mentioned Bifidobacterium lactis ga-2 in any one or more of the following:

[0024] (a) Promotes the growth of Lactobacillus schrei;

[0025] (b) Promotes the production of γ-aminobutyric acid by Lactobacillus hesitantus.

[0026] A third aspect of the present invention provides a microbial composition comprising the aforementioned Bifidobacterium lactis ga-2 and Lactobacillus hesitantii.

[0027] A fourth aspect of the present invention provides the use of the above-described microbial composition in the production of γ-aminobutyric acid.

[0028] A fifth aspect of the present invention provides a method for producing γ-aminobutyric acid by fermentation, the method comprising:

[0029] The activated Bifidobacterium lactis ga-2 was mixed with Lactobacillus hessier and inoculated into a culture medium, and sodium glutamate was added for fermentation culture to produce γ-aminobutyric acid.

[0030] Furthermore, the method includes:

[0031] S1. Select single colonies of Bifidobacterium lactis ga-2 and Lactobacillus spp. respectively into the first culture medium for a first activation culture, and then select single colonies of the activated Bifidobacterium lactis ga-2 and Lactobacillus spp. respectively into the second culture medium for a second activation culture.

[0032] S2. After the secondary activated Lactobacillus hirsutus is inoculated into the third culture medium for fermentation culture for a period of time, Bifidobacterium lactis is inoculated, and sodium glutamate is added in a fed-batch manner to continue fermentation culture to produce γ-aminobutyric acid.

[0033] In step S1, the first culture medium has the following composition:

[0034] Each 1000 mL contains 10 g peptone, 5 g beef meal, 4 g yeast powder, 2 g glucose, 1 mL Tween 80, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, 0.05 g manganese sulfate, and 15-20 g agar.

[0035] The composition of the second culture medium is as follows:

[0036] Each 1000 mL contains 10 g peptone, 5 g beef meal, 4 g yeast powder, 2 g glucose, 2 g fructooligosaccharides, 1 mL Tween 80, 2 g monosodium glutamate, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, and 0.05 g manganese sulfate.

[0037] In step S2, the composition of the third culture medium is as follows:

[0038] Each 1000 mL contains 10 g peptone, 5 g beef meal, 5 g soybean peptides, 5 g wheat peptides, 4 g yeast powder, 2 g glucose, 2 g fructooligosaccharides, 1 mL Tween 80, 2 g monosodium glutamate, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, and 0.05 g manganese sulfate.

[0039] The specific conditions for the fermentation culture for a period of time are: 30-37 ℃, 80-120 r / min, culture for 2-4 h, and the OD600 value reaches 1.0-1.5.

[0040] In step S2, the inoculation amount of both Lactobacillus schrei and Bifidobacterium lactis is 2%-3% (v / v).

[0041] The specific conditions for adding monosodium glutamate by the fed-batch method are as follows: the feeding rate of monosodium glutamate is 5-20 g / L;

[0042] The conditions for continued fermentation culture are: 30-37 ℃, 20-80 r / min, culture for 10-30 h.

[0043] Furthermore, the method includes:

[0044] Separately selected preserved *Lactobacillus hessei* and *Bifidobacterium lactis* strains were streaked onto solid culture medium (prepared by adding 1.5-2% agar to the first culture medium) for activation culture. *Lactobacillus hessei* was cultured at 30℃ for 18-24 h, and *Bifidobacterium lactis* for 18-24 h, respectively. After culturing, single colonies were inoculated into a second culture medium for static liquid culture. *Lactobacillus hessei* and *Bifidobacterium lactis* were cultured at 30℃ for 18-24 h, respectively, to allow the strains to recover maximum growth activity. 2%-3% ( v / v Inoculate *Lactobacillus hesseri* into the third culture medium at an inoculation rate of 37 °C and 100 r / min for approximately 2-4 h. Measure the OD600 value of *Lactobacillus hesseri* in the culture medium. When the OD600 value reaches 1.0-1.5, add 3%-5% (…) v / v Bifidobacterium lactis was inoculated into the third culture medium at an inoculation rate of 10 g / L. At this time, monosodium glutamate was added by fed-batch feeding. Fermentation was carried out at 37 °C and 50 r / min for 24 h.

[0045] The first culture medium has the following composition:

[0046] Each 1000 mL contains 10 g peptone, 5 g beef meal, 4 g yeast powder, 2 g glucose, 1 mL Tween 80, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, and 0.05 g manganese sulfate.

[0047] The composition of the second culture medium is as follows:

[0048] Each 1000 mL contains 10 g peptone, 5 g beef meal, 4 g yeast powder, 2 g glucose, 2 g fructooligosaccharides, 1 mL Tween 80, 2 g monosodium glutamate, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, and 0.05 g manganese sulfate.

[0049] The composition of the third culture medium is as follows:

[0050] Each 1000 mL contains 10 g peptone, 5 g beef meal, 5 g soybean peptides, 5 g wheat peptides, 4 g yeast powder, 2 g glucose, 2 g fructooligosaccharides, 1 mL Tween 80, 2 g monosodium glutamate, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, and 0.05 g manganese sulfate.

[0051] The following examples further illustrate the present invention, but do not constitute a limitation thereof. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the invention. In the examples, *Lactobacillus sherry* CGMCC NO. 26755 and *Lactobacillus sherry* CGMCC NO. 26138 have been disclosed in Chinese patents CN116814468A and CN116286513A, respectively, and *Lactobacillus plantarum* GDMCC NO. 64769 has been disclosed in Chinese patent CN118497086A.

[0052] The solid culture medium containing the first culture medium has the following composition:

[0053] Each 1000 mL contains 10 g peptone, 5 g beef meal, 4 g yeast powder, 2 g glucose, 1 mL Tween 80, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, 0.05 g manganese sulfate, and 20 g agar.

[0054] The composition of the second culture medium is as follows:

[0055] Each 1000 mL contains 10 g peptone, 5 g beef meal, 4 g yeast powder, 2 g glucose, 2 g fructooligosaccharides, 1 mL Tween 80, 2 g monosodium glutamate, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, and 0.05 g manganese sulfate.

[0056] The composition of the third culture medium is as follows:

[0057] Each 1000 mL contains 10 g peptone, 5 g beef meal, 5 g soybean peptides, 5 g wheat peptides, 4 g yeast powder, 2 g glucose, 2 g fructooligosaccharides, 1 mL Tween 80, 2 g monosodium glutamate, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, and 0.05 g manganese sulfate.

[0058] Example 1: Mixed fermentation of Lactobacillus hesitant CGMCC NO.26755 and Bifidobacterium lactis

[0059] Preserved *Lactobacillus hessei* CGMCC NO.26755 and *Bifidobacterium lactis* strain ga-2 were separately inoculated onto solid culture medium containing the first culture medium and streak-enhanced for activation (incubated at 30℃ for approximately 24 h). Single colonies of each strain were then picked and incubated statically in liquid culture medium containing the second culture medium (incubated at 30℃ for approximately 24 h) to allow the strains to regain maximum growth activity. In the experimental group, *Lactobacillus hessei* was inoculated into the third culture medium at a 2% inoculum and cultured at 37℃, 100 r / min for approximately 3 h. Then, monosodium glutamate (MSG) was added at a feed rate of 10 g / L and fermented at 37℃, 50 r / min for 24 h. In the control group 1, *Bifidobacterium lactis* was inoculated into the third culture medium at a 2% inoculum and cultured at 37℃, 100 r / min for approximately 3 h. Then, monosodium glutamate (MSG) was added at a feed rate of 10 g / L and fermented at 37℃, 50 r / min for 24 h. Control group 2: *Lactobacillus hessei* was inoculated into the third culture medium at a 2% inoculum and cultured at 37 °C and 100 r / min for approximately 3 h. The OD of *Lactobacillus hessei* in the culture medium was then measured. 600 Once the OD600 value reached 1.0, *Bifidobacterium lactis* was inoculated into the third culture medium at a 3% inoculum size. Monosodium glutamate (MSG) was then added in a fed-batch manner at a rate of 10 g / L. Fermentation was carried out at 37 °C and 50 r / min for 24 h. Samples were taken every 12 hours to measure GABA production and the OD600 value of the bacterial culture.

[0060] The results showed that Bifidobacterium lactis alone did not produce GABA, and Lactobacillus schrei alone produced low amounts of GABA. After 24 hours of mixed fermentation, the GABA yield reached 16.3 g / L, which was 1.4 times higher than that of single-strain fermentation (6.8 g / L). The cell growth (OD600) increased by 88%, and the fermentation cycle was shortened to 24 hours (single-strain fermentation requires 48 hours to achieve a similar yield).

[0061] Table 1. Determination of GABA yield and OD value in different fermentation groups

[0062]

[0063] Example 2: Mixed fermentation of Lactobacillus hesitant CGMCC NO.26138 and Bifidobacterium lactis

[0064] Preserved *Lactobacillus hessei* CGMCC NO.26138 and *Bifidobacterium lactis* strain ga-2 were separately inoculated onto solid medium containing the first culture medium and streak-enhanced for activation (incubated at 30℃ for approximately 24 h). Single colonies of each strain were then picked and incubated statically in liquid medium containing the second culture medium (incubated at 30℃ for approximately 24 h) to allow the strains to regain maximum growth activity. In the experimental group, *Lactobacillus hessei* was inoculated into the third medium at a 2% inoculum and cultured at 37℃, 100 r / min for approximately 3 h. Then, monosodium glutamate (MSG) was added at a feed rate of 10 g / L and fermented at 37℃, 50 r / min for 24 h. In the control group 1, *Bifidobacterium lactis* was inoculated into the third medium at a 2% inoculum and cultured at 37℃, 100 r / min for approximately 3 h. Then, monosodium glutamate (MSG) was added at a feed rate of 10 g / L and fermented at 37℃, 50 r / min for 24 h. Control group 2: *Lactobacillus hessei* was inoculated into the third culture medium at a 2% inoculum and cultured at 37 °C and 100 r / min for approximately 3 h. The OD of *Lactobacillus plantarum* in the culture medium was then measured. 600 Once the OD600 value reached 1.0, *Bifidobacterium lactis* was inoculated into the third culture medium at a 3% inoculum size. Monosodium glutamate (MSG) was then added in a fed-batch manner at a rate of 10 g / L. Fermentation was carried out at 37 °C and 50 r / min for 24 h. Samples were taken every 12 hours to measure GABA production and the OD600 value of the bacterial culture.

[0065] The results showed that Bifidobacterium lactis alone did not produce GABA, and Lactobacillus schrei alone produced low amounts of GABA. After 24 hours of mixed fermentation, the GABA yield reached 36.3 g / L, which was 93% higher than that of single-strain fermentation (18.8 g / L); the cell growth (OD600) increased by 125%, and the fermentation cycle was shortened to 24 hours (single-strain fermentation requires 48 hours to achieve a similar yield).

[0066] Table 2. Determination of GABA yield and OD value in different fermentation groups

[0067]

[0068] Example 3: Mixed fermentation of Lactobacillus plantarum GDMCC NO.64769 and Bifidobacterium lactis

[0069] Preserved *Lactobacillus plantarum* GDMCC NO.64769 and *Bifidobacterium lactis* strain ga-2 were separately inoculated onto solid culture medium containing the first culture medium and streak-enhanced for activation (incubated at 30℃ for approximately 24 h). Single colonies of each strain were then picked and incubated statically in liquid culture medium containing the second culture medium (incubated at 30℃ for approximately 24 h) to allow the strains to recover maximum growth activity. In the experimental group, *Lactobacillus plantarum* was inoculated into the third culture medium at a 2% inoculum size and cultured at 37℃, 100 r / min for approximately 3 h. Then, monosodium glutamate (MSG) was added at a feed rate of 10 g / L and fermented at 37℃, 50 r / min for 24 h. In the control group (Case 1), *Bifidobacterium lactis* was inoculated into the third culture medium at a 2% inoculum size and cultured at 37℃, 100 r / min for approximately 3 h. Then, monosodium glutamate (MSG) was added at a feed rate of 10 g / L and fermented at 37℃, 50 r / min for 24 h. Control group 2: *Lactobacillus plantarum* was inoculated into the third culture medium at an inoculum size of 2%, and cultured at 37 °C and 100 r / min for approximately 3 h. The OD value of *Lactobacillus plantarum* in the culture medium was then measured. 600 Once the OD600 value reached 1.0, *Bifidobacterium lactis* was inoculated into the third culture medium at a 3% inoculum size. Monosodium glutamate (MSG) was then added in a fed-batch manner at a rate of 10 g / L. Fermentation was carried out at 37 °C and 50 r / min for 24 h. Samples were taken every 12 hours to measure GABA production and the OD600 value of the bacterial culture.

[0070] Table 3. Determination of GABA yield and OD value in different fermentation groups

[0071]

[0072] The results showed that Bifidobacterium lactis cultured alone did not produce GABA, and the combination of another GABA-producing Lactobacillus plantarum with Bifidobacterium lactis in this application had no promoting effect (including promoting growth and promoting GABA production).

[0073] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A strain of Bifidobacterium lactis ( Bifidobacterium lactis The application of ga-2 in promoting the production of γ-aminobutyric acid by Lactobacillus hesit, said Bifidobacterium lactis ( Bifidobacterium lactis ga-2 is deposited at the China Center for Type Culture Collection on May 26, 2025, with accession number CCTCC NO: M 20251181.

2. The application of a microbial composition in promoting the production of γ-aminobutyric acid, characterized in that, The microbial composition comprises Bifidobacterium lactis ga-2 and Lactobacillus schrei as described in claim 1.

3. A method for promoting the production of γ-aminobutyric acid, characterized in that, The method includes: The activated Bifidobacterium lactis ga-2 of claim 1 was mixed with Lactobacillus hessier and inoculated into a culture medium, and sodium glutamate was added as a substrate for fermentation culture to produce γ-aminobutyric acid.

4. The method as described in claim 3, characterized in that, The method includes: S1. Select single colonies of Bifidobacterium lactis ga-2 and Lactobacillus spp. respectively into the first culture medium for a first activation culture, and then select single colonies of the activated Bifidobacterium lactis ga-2 and Lactobacillus spp. respectively into the second culture medium for a second activation culture. S2. After the secondary activated Lactobacillus hirsutus is inoculated into the third culture medium for fermentation culture for a period of time, Bifidobacterium lactis is inoculated, and sodium glutamate is added in a fed-batch manner to continue fermentation culture to produce γ-aminobutyric acid.

5. The method as described in claim 4, characterized in that, In step S1 The composition of the first culture medium is as follows: Each 1000 mL contains 10 g peptone, 5 g beef meal, 4 g yeast powder, 2 g glucose, 1 mL Tween 80, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, 0.05 g manganese sulfate, and 15-20 g agar. The composition of the second culture medium is as follows: Each 1000 mL contains 10 g peptone, 5 g beef meal, 4 g yeast powder, 2 g glucose, 2 g fructooligosaccharides, 1 mL Tween 80, 2 g monosodium glutamate, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, and 0.05 g manganese sulfate.

6. The method as described in claim 4, characterized in that, In step S2, the composition of the third culture medium is as follows: Each 1000 mL contains 10 g peptone, 5 g beef meal, 5 g soybean peptides, 5 g wheat peptides, 4 g yeast powder, 2 g glucose, 2 g fructooligosaccharides, 1 mL Tween 80, 2 g monosodium glutamate, 2 g dipotassium hydrogen phosphate, 5 g sodium acetate, 2 g triammonium citrate, 0.2 g magnesium sulfate, and 0.05 g manganese sulfate.

7. The method as described in claim 4, characterized in that, In step S2, the specific conditions for fermentation culture for a period of time are: 30-37 ℃, 80-120 r / min, culture for 2-4 h, and the OD600 value reaches 1.0-1.

5.

8. The method as described in claim 4, characterized in that, In step S2, the inoculation amount of both Lactobacillus schrei and Bifidobacterium lactis is 2%-3% (v / v). The specific conditions for adding monosodium glutamate by the fed-batch method are as follows: the feeding rate of monosodium glutamate is 5-20 g / L; The conditions for continued fermentation culture are: 30-37 ℃, 20-80 r / min, culture for 10-30 h.

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