Lactobacillus mucilaginosus LF.3 and application thereof

Abstract

The invention belongs to the technical field of microorganisms, and particularly relates to lactobacillus mucus LF.3 and application thereof. The fermented Lactobacillus mucilaginosus LF.3 fermented source-rich pickled Chinese cabbages provided by the invention have the effects of strong wiredrawing feeling and improvement of chewiness and overall flavor, can endow the source-rich pickled Chinese cabbages with special sour and fragrant flavor, crispness and chewiness, and also can maintain the unique wiredrawing posture of the source-rich pickled Chinese cabbages. The strain and the LP.3 strain are compounded to ferment the rich-source pickled Chinese cabbages, so that the advantage complementation effect can be realized, the wiredrawing state of the product can be kept, the rich fermented sour and fragrant flavor can be endowed, the bright color, good crispness and good chewiness can be ensured, the color, fragrance, taste and crispness of the rich-source pickled Chinese cabbages are remarkably improved, the rich-source pickled Chinese cabbages are comprehensively improved, and the market competitiveness of the rich-source pickled Chinese cabbages is improved. According to the method, the problem that the fermentation batch quality of the rich-source pickled Chinese cabbages is unstable is solved, the food safety risk is reduced, industrialized, standardized and normalized production is easy to realize, and the problems that the fermentation batch quality of the rich-source pickled Chinese cabbages is unstable and industrialization is difficult are solved.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology, and in particular relates to a fermenting Lactobacillus mucinus LF.3 and its applications. Background Technology

[0002] Fuyuan pickled cabbage is a unique delicacy from Fuyuan County, Yunnan Province. Its distinctive flavor and texture are attributed to the local climate and water source. The product is characterized by a short fermentation cycle, long, tangy texture, a thick, smooth consistency, and high acidity. In October 2016, it was recognized as a municipal-level intangible cultural heritage. In recent years, pickled cabbage has become increasingly popular. Fuyuan pickled cabbage is rich in dietary fiber, calcium, phosphorus, iron, and selenium, and is believed to have benefits such as aiding digestion, weight loss, relieving summer heat, and sobering up. Currently, Fuyuan pickled cabbage is mainly produced through natural fermentation in small family workshops. The complex fermentation process and difficulty in controlling the microorganisms result in inconsistent product quality, including dull color, poor fermentation flavor, poor crispness, low safety, and short shelf life. This hinders industrial-scale production and severely restricts the advanced processing of Fuyuan pickled cabbage.

[0003] Currently, domestic and international research on Fuyuan sauerkraut focuses on the application of hazard analysis and critical control point (HACCP) systems, nitrite content control, bacterial flora composition of fermentation systems, screening of lactic acid bacteria, and small-scale application studies of adding exogenous or native lactic acid bacteria. For example, Huang Huifu et al. (Huang Huifu, Xiao Yuxue. Effects of lactic acid bacteria fermentation on the quality and safety of Fuyuan sauerkraut [J]. Food and Machinery, 2019, 35(05):191-194+201.DOI:10.13652 / j.issn.1003-5788.2019.05.032.) found that adding lactic acid bacteria to ferment Fuyuan sauerkraut can reduce the amount of nitrite and pH value in the sauerkraut, increase the total acid content, and reduce the total number of colonies, molds, and yeasts, resulting in a milder product with a more mellow and fragrant sour taste. However, the fermentation characteristics of different strains and species in different regions vary significantly, and purchased strains have the disadvantages of not possessing the unique flavor and quality of Fuyuan sauerkraut and being expensive. Therefore, selecting high-quality local lactic acid bacteria strains from traditionally naturally fermented Fuyuan sauerkraut can not only preserve the original excellent flavor and quality of Fuyuan sauerkraut, but also improve the quality and safety of Fuyuan sauerkraut and promote its standardized production. Summary of the Invention

[0004] Based on the above technical problems, the main objective of this invention is to overcome the shortcomings of the aforementioned background technology. A strain of *Limosilactobacillus fermentum* LF.3 was screened from traditional Fuyuan sauerkraut. This strain can maintain the good stringy texture of Fuyuan sauerkraut, as well as its crispness and chewiness. When combined with strain LP.3 for the fermentation of Fuyuan sauerkraut, it can achieve complementary advantages, maintaining the stringy texture of the product, imparting a rich fermented sour aroma, ensuring its bright color, good crispness, and chewiness, and also inhibiting the growth of other bacteria and yeasts to a certain extent. This solves the problem of unstable batch quality in the fermentation of Fuyuan sauerkraut, improves the problems of insufficient fermentation flavor, uneven color, and poor chewiness, and reduces potential food safety risks.

[0005] To achieve the above objectives, the inventors conducted in-depth research and, through repeated studies and demonstrations, obtained the solution of this invention, as detailed below:

[0006] In a first aspect, embodiments of this application provide a strain of fermenting *Lactobacillus mucinus* (… Limosilactobacillus fermentum Lactobacillus fermentans LF.3 was deposited at the Guangdong Provincial Center for Microbial Culture Collection on April 28, 2025, with accession number GDMCC No. 66236.

[0007] The colonies of Lactobacillus fermentans LF.3 on MRS medium are milky white, round, with neat edges, smooth surface, and moist and glossy. After Gram staining, the strain is observed under a microscope to be Gram-positive, with typical rod-shaped cells, 0.5-0.8 μm in length and 0.2-0.4 μm in width.

[0008] Secondly, the present invention also provides a fermentation agent comprising the above-mentioned Lactobacillus fermentans LF.3, preferably Lactobacillus plantarum LP.3, with accession number CGMCC No. 30203.

[0009] Thirdly, the present invention also provides the application of the above-mentioned fermentation agent in the fermentation of Fuyuan sauerkraut, especially to improve the problems of insufficient fermentation flavor, uneven color and poor chewiness of Fuyuan sauerkraut, and enhance the color, aroma, taste and crispness of Fuyuan sauerkraut, thereby comprehensively improving the quality of Fuyuan sauerkraut.

[0010] Fourthly, the present invention also provides a fermentation method for Fuyuan pickled cabbage, the method comprising the step of fermentation using the above-mentioned fermentation agent.

[0011] Furthermore, the amount of the fermentation agent added is 10. 6 -10 7CFU / mL.

[0012] Furthermore, the ratio of *Lactobacillus mucinus* LF.3 to *Lactobacillus plantarum* LP.3 in the fermentation agent is 1:1.

[0013] Furthermore, the fermentation agent is added at the initial stage of fermentation.

[0014] Furthermore, the fermentation time is 6-8 days.

[0015] Fifthly, the present invention provides a Fuyuan pickled cabbage, which is prepared by the method described in this application.

[0016] One or more technical solutions provided in the embodiments of this application have at least the following technical effects or advantages:

[0017] 1. The fermented Fuyuan sauerkraut provided by the present invention has the effect of strong stringiness, improved chewiness and overall flavor, and no off-flavors or odors. It can give Fuyuan sauerkraut its characteristic sour and fragrant flavor, crispness and chewiness, and can also maintain its unique stringy texture.

[0018] 2. The fermentation of Lactobacillus mucilaginosus strain LF.3 provided by this invention, when combined with LP.3 for the fermentation of Fuyuan sauerkraut, can achieve complementary advantages. It can not only ensure the characteristics of Fuyuan sauerkraut itself, such as strong stringiness, rich and harmonious sour taste, good crispness and refreshing taste, but also solve the problems of insufficient sour aroma, dark color and easy browning of fermented Fuyuan sauerkraut. It can significantly improve the color, aroma, taste and crispness of Fuyuan sauerkraut, and comprehensively improve the quality of Fuyuan sauerkraut.

[0019] 3. The technical solution provided by this invention adopts direct-inoculation pure culture fermentation technology, and exogenously adds fermenting Lactobacillus mucilaginosus LF.3 strain and Lactobacillus plantarum LP.3 compound bacterial agent to realize the compound bacterial agent fermentation of Fuyuan sauerkraut. It can rapidly reproduce in the fermentation system, play a role in nutrient occupation, reduce the growth space of miscellaneous bacteria, inhibit the growth of miscellaneous bacteria, make the fermentation process more controllable, improve the safety and fermentation quality of Fuyuan sauerkraut, reduce food safety risks, and is easy to realize industrialization, standardization and standardized production, solving the problems of unstable batch quality and difficulty in industrialization of Fuyuan sauerkraut fermentation. Attached Figure Description

[0020] Figure 1 A graph showing the change in total acid content in a purebred fermented Fuyuan sauerkraut system;

[0021] Figure 2 A graph showing the pH changes in a purebred fermented sauerkraut system;

[0022] Figure 3 This is a colony morphology diagram of *Lactobacillus mucinus* LF.3.

[0023] Figure 4 Microscopic image of cell morphology of Lactobacillus fermentation LF.3;

[0024] Figure 5 Figure 1 shows the results of live lactic acid bacteria count at different fermentation stages;

[0025] Figure 6 Figure 1 shows the results of yeast viable cell counts at different fermentation stages.

[0026] Figure 7 A graph showing the changes in total acid content in the Fuyuan pickled cabbage system at different fermentation stages;

[0027] Figure 8 A graph showing the pH changes of the Fuyuan pickled cabbage system at different fermentation stages;

[0028] The fermented Lactobacillus mucinus provided by this invention ( Limosilactobacillus fermentum LF.3 was deposited on April 28, 2025, at the Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, Guangdong Province, with accession number GDMCC No. 66236. The strain was received and registered by the collection center on April 28, 2025, and was confirmed to be a viable strain by the collection center on the same day.

[0029] The plant lactobacillus provided by this invention ( Lactiplantibacillus plantarum LP.3 was deposited on March 29, 2024, at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 30203. The strain was received and registered by the collection center on March 29, 2024, and was confirmed to be a viable strain by the collection center on the same day. Detailed Implementation

[0030] To facilitate understanding of the present invention, a more comprehensive description will be given below. However, the present invention can be implemented in many different forms and is not limited to the embodiments described herein. Rather, these embodiments are provided to provide a thorough and complete understanding of the disclosure of the present invention.

[0031] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0032] Unless otherwise specified, the reagents, methods and equipment used in this invention are conventional food-grade reagents, methods and equipment in the art.

[0033] The terminology used herein in the specification of this invention is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. The term "and / or" as used herein includes any and all combinations of one or more of the associated listed items.

[0034] Example 1: Screening of fermented Lactobacillus mucinus LF.3

[0035] 1. Raw material processing:

[0036] Under aseptic conditions, naturally fermented Fuyuan pickled cabbage products with good flavor, uniform color, and no browning were subjected to microscopic examination to confirm the presence of the target microorganism in the material to be isolated. Under aseptic conditions, 25g of naturally fermented Fuyuan pickled cabbage with good flavor was weighed and placed in 225mL of sterile water. The mixture was homogenized for 30 minutes and then diluted using a 10-fold serial dilution method with sterile water. 10... -4 10 -5 10 -6 Three dilution gradients of bacterial suspension were inoculated onto MRS solid medium using the pour culture method, with 1 mL inoculated per dish for each dilution. The dishes were then placed in a 30°C incubator and incubated anaerobically upside down for 48 hours.

[0037] Culture dishes with a growth rate of 30-300 CFU / plate were selected. Colony morphology was labeled and morphological differences were described. Colonies with different morphologies were examined and described under a microscope. Typical lactic acid bacteria were selected for purification. After purification three times, Gram staining was used for microscopic examination to preliminarily determine the purified strains. Glycerol cryovials were prepared and stored at -80℃ in a medical refrigerator for later use. A total of five test microorganisms were isolated and named LF.3, LB.1, LB.2, MB.1, and MB.2, respectively. Patent CN202410718164.1 discloses a strain of *Lactobacillus plantarum* LP.3 isolated from fermented small-leaf mustard greens, which has the advantages of rapid acid production (high malic acid production) and promoting the fermentation process. This strain was also used as a preferred test strain for fermentation testing of sauerkraut in Fuyuan.

[0038] 2. Lactic acid bacteria screening

[0039] Take the glycerol cryopreserved tubes of the 6 selected test bacteria, and inoculate them into 100 mL of liquid culture medium (MRS medium) at an inoculum rate of 0.2% (v / v). Incubate at 37°C for 48 h, and adjust the bacterial concentration to 1×10⁻⁶. 8 CFU / mL is the test bacterial suspension.

[0040] Fresh radishes were peeled to remove any discolored skin and inedible parts. After washing thoroughly, the radishes were shredded. 100g of shredded radish was accurately weighed into an Erlenmeyer flask, and 100mL of distilled water was added. The flasks were sealed. A total of 14 flasks were prepared and divided into two equal portions (7 flasks per portion). One portion was sterilized at 95℃ for 20 minutes. After cooling, the test bacterial suspension was aseptically inoculated at a ratio of 1% of the total system. The total acid content and pH value were measured at fermentation days 1, 3, 5, and 7. The results are shown in the table below. Figure 1-2 Color difference was measured at 3, 5, and 7 days of fermentation, and the results are recorded in Table 1. Sensory evaluation was performed at 7 days of fermentation, and the organic acid content was measured, and the results are recorded in Tables 2-3. Another sample was not sterilized; the test bacterial suspension was directly inoculated at a ratio of 1% of the total system, and the whole texture analysis was performed after 4 days of fermentation, and the results are recorded in Table 4.

[0041] The organic acid content of Fuyuan pickled cabbage was determined according to GB / T 5009.157-2016 "National Food Safety Standard - Determination of Organic Acids in Food". Chromatographic conditions: Welchrom-C18 column (250nm×4.6mm, 5μm), detection wavelength: λ=210nm; mobile phase: 0.1% phosphoric acid solution-methanol (97.5:2.5), column temperature: 40℃, flow rate: 1 mL / min, injection volume: 20µL.

[0042] Depend on Figures 1-2 It can be seen that as the fermentation process progresses, the total acid in each fermentation system continuously accumulates, and the pH continuously decreases. From the initial stage of fermentation to the end of fermentation 7 days later, the CK (fermentation without inoculum) group had the least acid accumulation and the highest pH among all treatment groups. The acid accumulation of Fuyuan sauerkraut fermented with inoculum was higher than that without inoculum, and the pH value was lower in all treatment groups. From the above data, it can be seen that the Fuyuan sauerkraut system without inoculum has a small amount of acid accumulation but limited acid production capacity. Adding inoculum can promote a large amount of acid production in the system and reduce the pH value. Among them, LP.3 is the strain with the strongest acid production capacity and the greatest effect on promoting the fermentation process, while MB.2 is the strain with the weakest acid production capacity. However, there is no significant difference in the pH value within the LP.3, LB.1, LB.2, and LF.3 systems. These four strains can be used as reference strains for Fuyuan sauerkraut fermentation.

[0043] Table 1. Color difference at different stages of purebred fermented Fuyuan sauerkraut

[0044]

[0045] Table 1 shows that as fermentation time increased, the brightness of Fuyuan sauerkraut gradually decreased, darkened, turned blue, and then green. Throughout the fermentation process, the brightness of Fuyuan sauerkraut fermented with the added bacterial strain was higher than that without the added strain. After fermentation, the brightness of each treatment group was in the following order: LP.3 > LB.1 > LB.2 > LF.3 > MB.2 > MB.1 > CK. The best color was achieved with LP.3 fermentation, and the worst with MB.1. In terms of ΔE, compared with CK, the ΔE of the treatments with added bacterial strain, from largest to smallest, was: LP.3 > LB.1 > LB.2 > LF.3 > MB.2 > MB.1. Adding bacterial strain significantly affected the color of Fuyuan sauerkraut. After 7 days of fermentation, compared with the control (CK), LP.3, LB.1, LF.3, MB.2, and MB.1 showed brighter, more yellowish-green colors, with ΔE values ​​of 23.44, 21.13, 13.99, 10.87, and 4.15, respectively; LB.2 showed a brighter, more greenish-blue color, with a ΔE of 18.40. All strains improved the color of Fuyuan sauerkraut during fermentation, with LP.3, LB.1, LF.3, and LB.2 showing the most significant effects and thus serving as candidate strains for pure-culture fermentation of Fuyuan sauerkraut.

[0046] Table 2 Organic Acid Content of Purebred Fermented Fuyuan Sauerkraut

[0047]

[0048] Table 2 shows that after 7 days of fermentation, the organic acid content of the fermented Fuyuan sauerkraut with the added strains was higher than that of the control (CK). The organic acid content also varied among different strains, with lactic acid being the highest and malic acid the lowest. Among them, the Fuyuan sauerkraut fermented with MB.1 had the lowest organic acid content, which was closest to that of the control (CK). The organic acid content of the Fuyuan sauerkraut fermented with LP.3, LB.1, LB.2, and LF.3 all showed significant increases, but there were no significant differences among them. The organic acid contents were: lactic acid 18.55, 18.49, 18.39, and 18.01 mg•100 g⁻¹; citric acid 0.62, 0.62, 0.61, and 0.57 mg•100 g⁻¹; tartaric acid 1.15, 0.56, 1.06, and 1.12 mg•100 g⁻¹; and malic acid 0.21, 0.16, 0.24, and 0.25 mg•100 g⁻¹. LP.3, LB.1, LF.3 and LB.2 are strains with strong organic acid production capabilities and can be used as candidate strains for the fermentation of Fuyuan pickled cabbage.

[0049] Table 3 Sensory Evaluation of Purebred Fermented Fuyuan Sauerkraut

[0050]

[0051] Table 3 shows that after 7 days of fermentation, the Fuyuan sauerkraut fermented with the added strain had a brighter color than CK, consistent with the color difference results. Each treatment of the fermented Fuyuan sauerkraut had its own unique flavor. Among them, LP.3 had the best overall sensory evaluation results, with a bright color, good crispness of the radish shreds, a crisp texture, and a pure sour taste. LF.3 fermented with strain LF.3 had a good stringy texture, but the color and crispness were inferior, and the sour taste was weaker. LB.1, LB.2, MB.2, and MB.1 fermented with mixed and impure flavors, and inferior sour taste and crispness. LB.1 and MB.2 sauerkraut developed a film on the surface, affecting the flavor and shelf life of the product. LB.2, MB.2, and MB.1 fermented sauerkraut had an alcoholic taste, which significantly affected their flavor and taste. CK had the worst sensory quality among all treatments, with a dark color, a sour and rancid taste, and mushy radish shreds. Based on a comprehensive assessment of various sensory indicators, LP.3 significantly enhances the flavor, texture, and color of Fuyuan sauerkraut during fermentation, while LF.3 imparts a unique stringy texture to Fuyuan sauerkraut. Both strains can be considered as candidate strains for pure-culture fermentation of Fuyuan sauerkraut.

[0052] Table 4. Texture analysis of purebred fermented Fuyuan pickled mustard greens

[0053]

[0054] Table 4 shows that the overall textural properties of Fuyuan sauerkraut fermented with different strains and without inoculation differed significantly, and the patterns of each textural property varied. The greatest difference was in hardness, followed by adhesiveness, chewiness, and elasticity, while the smallest differences were in cohesiveness and elasticity. Regarding hardness, there were significant differences among the various fermentation methods, following the order LP.3 > LF.3 > LB.2 > MB.1 > CK > LB.1 > MB.2, with hardness values ​​of 678.64, 643.64, 554.62, 463.60, 424.59, 371.58, and 359.58 g, respectively. These data indicate that adding different strains during fermentation of Fuyuan sauerkraut can increase or decrease the product hardness. LP.3, LF.3, LB.2, and MB.1 can increase the hardness of the sauerkraut, while LB.1 and MB.2 fermentation leads to a decrease in hardness. The chewiness, resilience, and elasticity of Fuyuan sauerkraut from all treatments exhibited consistent patterns. The cohesiveness and adhesiveness showed similar patterns, but differed somewhat from the first three characteristics. Overall, except for hardness, LF.3 and LP.3 showed the best performance in the other characteristics, followed by LB.2, CK, and LB.1, while MB.1 and MB.2 showed the worst. Among the various textural properties of LF.3, cohesiveness was 0.41 g•sec⁻¹, adhesiveness was 264.98, chewiness was 141.87, resilience was 0.45, and elasticity was 0.54, showing significant differences from other treatments. This indicates that LF.3 can significantly improve the textural properties of Fuyuan sauerkraut, increase the internal binding force of the product, and enhance its taste. In contrast, MB.1 and MB.2 fermented Fuyuan sauerkraut, which reduced its textural properties, negatively impacting its overall sensory appeal. Compared to CK, LB.2 and LB.1 altered the textural properties of Fuyuan sauerkraut, but their effects were not as significant as those of LF.3 and LP.3. Therefore, LF.3 and LP.3 can be considered as candidate strains for the fermentation of Fuyuan sauerkraut.

[0055] Based on the physicochemical indicators (total acid and pH), color difference, sensory evaluation, organic acid content, and overall texture analysis in the fermentation tests of the strains, it was found that fermentation with strain LF.3 increased the total acid and organic acid content of the product, produced a stringy texture, enhanced the binding force between radish shreds, and improved the taste, but the sourness was not well-balanced, and the color was slightly darker. Fermentation with strain LP.3 significantly increased the total acid and organic acid content of the product, enhanced the brightness of the color, increased the firmness, and extended the product's shelf life. Since both strains have unique characteristics in fermenting Fuyuan sauerkraut, they were selected to be combined to prepare a compound fermentation starter as the inoculum for fermenting Fuyuan sauerkraut.

[0056] 3. Identification and Preservation of Lactic Acid Bacteria

[0057] A suspension of *Lactobacillus mucilaginosus* LF.3, selected for fermentation testing of Fuyuan sauerkraut, was subjected to streak plating and Gram staining. Colony morphology and cell morphology were observed and described using both the naked eye and microscope. The colony morphology of LF.3 is shown below. Figure 3As shown, the colonies are milky white, round, with neat edges, smooth surfaces, and a moist, glossy appearance. After Gram staining, the morphology of the obtained strain was observed under a 100x oil immersion microscope as follows. Figure 4 As shown, this strain is a Gram-positive bacterium with typical rod-shaped cells, 0.5-0.8 μm in length and 0.2-0.4 μm in width.

[0058] The pure culture of the tested LF.3 strain was sent for sequencing and identified as *Lactobacillus fermentans*. Limosilactobacillus fermentum This strain meets the requirements of the "List of Microbial Strains that Can Be Used in Food" and the "List of Microbial Strains that Can Be Used in Infant Food" issued by the Ministry of Health of my country. It was deposited at the Guangdong Provincial Center for Microbial Culture Collection on April 28, 2025, with the strain accession number GDMCC No. 66236.

[0059] Example 2: Fermentation Test of Mixed Strains

[0060] Preparation of bacterial suspension: 0.6 mL of LF.3 and LP.3 cryopreserved bacterial suspension were added to 300 mL of MRS medium and incubated at 30°C for 48 h to obtain the bacterial suspension. The bacterial concentration was adjusted to 1 × 10⁻⁶. 8 CFU / mL.

[0061] Take 7 kg of fresh radishes, remove the heads and tails, and remove any discolored or mixed-color skin. Wash them thoroughly and cut them into 2cm*2cm*2cm cubes for easy comparison of texture and crispness. Blanch the radishes and place them in a fermentation jar. Add the suspensions of each test strain to cooled boiled water as shown in Table 5, mix well, add the cooled boiled water to the mouth of the jar, seal the mouth with the liquid, mark it, and place it in a dark indoor place for natural fermentation for 7 days. During fermentation, pay attention to maintaining the mouth of the jar. After 7 days of fermentation, take samples to determine the total acidity, and perform sensory and overall texture analysis. The results are shown in Tables 6 and 7.

[0062] Table 5. Inoculum addition details

[0063]

[0064] Table 6 Sensory and total acidity results of mixed strain fermentation tests

[0065]

[0066] Table 6 shows the fermentation test results of mixed strains. The mixed strain fermentation of Fuyuan sauerkraut yielded the best overall sensory quality and highest total acid content, followed by single-strain fermentation, with naturally fermented Fuyuan sauerkraut being the worst. Fuyuan sauerkraut fermented with LP.3 single strain had a brighter color, a richer and more harmonious sour taste, and a higher total acid content than LF.3, but lacked the stringy texture. Fermentation with a mixed strain containing 1% LP.3 and LF.3 resulted in a porcelain-white, bright color, a rich sour aroma, a crisp texture, strong stringiness, and excellent crispness, with the highest total acid content. When the addition of LP.3 and LF.3 was halved, the sensory quality decreased to some extent, and the total acid content also decreased accordingly. Therefore, only when the addition amount of the LP.3 and LF.3 compound microbial agent reaches a certain level can the strains play a dominant role in the system, maximizing their contribution to product quality improvement.

[0067] Table 7. Texture analysis of mixed-culture fermentation

[0068]

[0069] Table 7 shows that there are significant differences in the overall texture characteristics of fermented Fuyuan sauerkraut under uninoculated, single-strain, and mixed-strain inoculation. The most significant differences are in hardness, adhesiveness, and chewiness. The mixed strains LP.3 and LF.3 show better results in all indicators compared to single-strain and uninoculated fermentation. Regarding hardness, the order is LPF-2 > LPF-1 > LP.3 > LF.3 > CK1, with hardness values ​​of 753.63, 725.15, 676.14, 658.64, and 599.63 g, respectively. These data indicate that the addition of mixed strains significantly improves the hardness of Fuyuan sauerkraut. The cohesiveness, adhesiveness, chewiness, resilience, and elasticity of fermented Fuyuan sauerkraut with the addition of 1% LP.3 and LF.3 and 0.5% mixed strains of LP.3 and LF.3 were 0.44 and 0.41, 333.47 and 293.36, 179.66 and 151.84, 0.43 and 0.39, and 0.54 and 0.52, respectively. All these indicators were superior to those of single-strain fermentation and natural fermentation. The mixed fermentation of Fuyuan sauerkraut with LP.3 and LF.3 significantly improved the shortcomings of poor crispness, soft texture, and unpleasant taste. Furthermore, the fermentation process using the mixed strains offered strong controllability, ensuring product quality stability.

[0070] Example 3: Fermentation of Fuyuan Sauerkraut using Mixed Strains in a 200kg Fermentation Tank

[0071] 200 μL of frozen *Lactobacillus plantarum* LP.3 and LF.3 cultures were inoculated into 100 mL of liquid culture medium (MRS medium) and incubated at 37°C for 48 h to obtain primary seed culture. The primary seed culture was then inoculated into liquid culture medium (MRS medium) at a ratio of 1% and incubated at 37°C for 24 h. After centrifugation, the bacterial cells were resuspended in physiological saline, and the bacterial concentration was adjusted to 1 × 10⁻⁶. 8 The CFU / mL solution is then mixed at a 1:1 ratio to form a compound bacterial seed solution.

[0072] Take 100kg of fresh radishes, remove discolored skin and inedible parts, wash thoroughly, shred the radishes, blanch the shreds, and place them in a fermentation jar. Add 2% of the compound microbial agent seed liquid to cooled boiled water and mix well. Add the cooled boiled water to the mouth of the jar, seal the mouth with the liquid, and use uninoculated natural fermentation as a control. Mark the jar and place it in a dark indoor place for natural fermentation for 7 days, while maintaining the jar mouth. Take samples on days 1, 3, 5, and 7 of fermentation to test pH, total acidity, lactic acid bacteria, and viable yeast counts. Record the results. Figure 5-8 Sensory evaluation was conducted after 7 days of fermentation. The evaluation criteria are shown in Table 8, and the scores are recorded in Table 9.

[0073] Depend on Figure 5-6 It can be seen that the number of viable lactic acid bacteria and yeast in Fuyuan sauerkraut was relatively low in the early stage of fermentation. As the fermentation process progressed, the number of viable lactic acid bacteria in Fuyuan sauerkraut fermented with mixed strains increased significantly, while the number of viable yeast decreased. In naturally fermented Fuyuan sauerkraut, the number of viable lactic acid bacteria increased slightly, while the number of viable yeast increased significantly. The lactic acid bacteria and yeast in the fermentation system basically reached stability after 5 days of fermentation. In Fuyuan sauerkraut fermented with a mixture of LP.3 and LF.3, the number of viable lactic acid bacteria increased from 6.34 lg CFU / g to 9.24 lg CFU / g, while the number of viable yeast decreased from 4.24 lg CFU / g to 3.66 lg CFU / g. In naturally fermented (CK2) Fuyuan sauerkraut, the number of viable lactic acid bacteria increased from 6.02 lg CFU / g to 7.01 lg CFU / g, while the number of viable yeast increased from 4.23 lg CFU / g to 7.00 lg CFU / g. In the Fuyuan sauerkraut fermented with LP.3 and LF.3, lactic acid bacteria dominate, competing for nutrients in the system and thus inhibiting the growth and reproduction of yeast. In contrast, the naturally fermented Fuyuan sauerkraut system has a complex and diverse range of microorganisms, with multiple strains competing for nutrients. The number of lactic acid bacteria is limited, resulting in a smaller number of lactic acid bacteria and a large number of yeasts.

[0074] Depend on Figure 7-8It can be seen that as the fermentation process progresses, the total acid content of Fuyuan sauerkraut continuously increases, and with the accumulation of total acid, the pH value continuously decreases. In the Fuyuan sauerkraut inoculated with LP.3 and LF.3 for combined fermentation, the total acid content increased from 0.21% to 0.79%, and the pH value decreased from 3.555 to 2.95; in the naturally fermented Fuyuan sauerkraut, the total acid content increased from 0.1% to 0.35%, and the pH value decreased from 4.21 to 3.525. The Fuyuan sauerkraut system inoculated with LP.3 and LF.3 for combined fermentation exhibits stronger acid production capacity and can rapidly reduce the pH value of the system.

[0075] Table 8 Sensory Evaluation Criteria for Fuyuan Pickled Cabbage

[0076]

[0077] Table 9 Sensory Scores of Fuyuan Pickled Cabbage

[0078]

[0079] Table 9 shows that there were significant differences in color, aroma, taste, crispness, and overall score between LP.3 and LF.3 combined fermentation and natural fermentation of Fuyuan sauerkraut. The scores were 9.05 and 7.30, 8.50 and 5.92, 8.80 and 6.52, 8.46 and 6.55, and 8.75 and 6.64, respectively. After inoculation with LP.3 and LF.3 combined fermentation, the color, aroma, taste, crispness, and overall score of Fuyuan sauerkraut were significantly improved.

[0080] In summary, the combined fermentation of Fuyuan sauerkraut with LP.3 and LF.3 can increase the number of viable lactic acid bacteria in the system, giving them a dominant position. This allows them to seize nutrients and multiply rapidly, producing lactic acid and other organic acids, significantly increasing the total acid content, lowering the pH value, inhibiting the growth of harmful microorganisms such as yeast, promoting the formation of sauerkraut flavor, and significantly improving the color, aroma, taste, and crispness of Fuyuan sauerkraut. This comprehensively enhances the quality of Fuyuan sauerkraut and makes it easy to achieve industrialization, standardization, and regulated production.

[0081] The embodiments described above are merely illustrative of several implementations of the present invention, and while the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the invention patent. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these all fall within the protection scope of the present invention. Therefore, the protection scope of this invention patent should be determined by the appended claims.

Claims

1. A strain of fermenting *Lactobacillus mucilaginosus* ( Limosilactobacillus fermentum LF.3, characterized in that, The preservation number of the fermenting Lactobacillus mucinus LF.3 is GDMCC No. 66236.

2. A fermentation agent, characterized in that, The fermentation agent comprises *Lactobacillus LF.3* as described in claim 1.

3. The fermentation agent as described in claim 2, characterized in that, The fermentation agent also contains *Lactobacillus plantarum* (… Lactiplantibacillus plantarum LP.3, accession number CGMCC No. 30203.

4. The application of the fermentation agent as described in claim 3 in the fermentation of Fuyuan sauerkraut.

5. A fermentation method for Fuyuan pickled cabbage, characterized in that, The method includes the step of fermentation using the fermentation agent according to claim 3.

6. The method as described in claim 5, characterized in that, The amount of fermentation agent added is 10. 6 -10 7 CFU / mL.

7. The method as described in any one of claims 5 or 6, characterized in that, The ratio of *Lactobacillus mucinus* LF.3 to *Lactobacillus plantarum* LP.3 in the fermentation agent is 1:

1.

8. The method as described in any one of claims 5 or 6, characterized in that, The fermentation agent is added at the initial stage of fermentation.

9. The method as described in any one of claims 5 or 6, characterized in that, The fermentation time is 6-8 days.

10. A type of Fuyuan pickled cabbage, characterized in that, The Fuyuan pickled cabbage is prepared by any one of the methods described in claims 5-9.

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