A low-salt and fresh-tasting processing method for stink catfish based on functional complementary lactic acid bacteria synergistic fermentation

Abstract

The present application belongs to the technical field of aquatic product fermentation processing, and particularly relates to a low-salt flavor-enhanced stink catfish processing method based on functional complementary lactic acid bacteria synergistic fermentation. The method separates endogenous lactic acid bacteria from stink catfish, evaluates the salt tolerance and metabolic characteristics of the lactic acid bacteria during implementation, and optimally selects and constructs a functional complementary lactic acid bacteria combination. Through synergistic fermentation, the degradation of protein and lipid in fish meat is promoted, and flavor substances beneficial to umami formation are generated, so that the umami and overall quality of stink catfish are improved under the condition of significantly reducing the amount of salt. Compared with high-salt natural fermentation and low-salt natural fermentation, the stink catfish prepared by the method of the present application has higher umami response, more optimal texture characteristics, and lower pH value and water activity. The method of the present application is beneficial to improving the stability and safety of the product, and is suitable for the standardized and large-scale production of low-salt stink catfish.

Description

Technical Field

[0001] This invention belongs to the field of aquatic product fermentation and processing technology, and in particular relates to a low-salt, flavor-enhancing fermented mandarin fish processing method based on the synergistic fermentation of functionally complementary lactic acid bacteria. Background Technology

[0002] Stinky mandarin fish is a traditional aquatic product processed through natural fermentation. It is widely loved by consumers for its unique sensory characteristics: a pungent smell, a fragrant taste, and meat resembling garlic cloves. The fermentation process relies on the microbial community naturally present in the raw fish, undergoing complex biochemical reactions under specific salinity and temperature conditions to create its distinctive flavor and texture. For a long time, stinky mandarin fish production has primarily relied on natural fermentation, a process significantly influenced by environmental factors, resulting in a long fermentation cycle and insufficient product quality stability.

[0003] In traditional stinky mandarin fish processing, increasing the amount of salt is typically used to inhibit the growth of spoilage microorganisms and ensure the safety of the fermentation process. However, while a high-salt environment improves product safety, it also leads to excessive sodium intake, which is inconsistent with the current trend of low-sodium, healthy diets. Furthermore, higher salinity may inhibit the metabolic activity of some beneficial microorganisms, affecting the formation of flavor compounds and limiting the improvement of stinky mandarin fish product quality and industrial upgrading.

[0004] To reduce the amount of salt used in fermented mandarin fish products, studies have attempted to improve fermentation under low-salt conditions by controlling processing conditions or introducing fermenting microorganisms. The flavor of aquatic products, especially umami, mainly originates from the degradation and transformation of macromolecules such as proteins and lipids during fermentation, forming flavor compounds such as free amino acids, flavor nucleotides, organic acids, and small peptides. However, under low-salt conditions, the fermentation of fermented mandarin fish often suffers from insufficient fermentation kinetics, incomplete flavor formation, and textural degradation, making it difficult to maintain the traditional flavor characteristics of the product while reducing salinity.

[0005] Lactic acid bacteria, a class of functional microorganisms widely used in fermented foods, have attracted attention in the processing of fermented meat and aquatic products due to their acid-producing, enzyme-producing, and inhibitory effects on the growth of other microorganisms. Related studies have shown that lactic acid bacteria can promote the degradation of proteins and lipids through the action of proteases and lipases, thereby affecting the flavor and texture of the product. However, the application of lactic acid bacteria in the processing of stinky mandarin fish still has certain limitations in current technologies. For example, the use of natural enrichment methods or single-strain fermentation makes it difficult to simultaneously ensure fermentation safety, flavor formation, and product stability; some fermentation strains lack adaptability to the complex fermentation system of stinky mandarin fish and struggle to stably perform their metabolic functions in low-salt environments.

[0006] Therefore, under the premise of ensuring the safety of fermented mandarin fish, how to effectively promote the formation of umami substances under low-salt conditions, improve product texture, and enhance the controllability and stability of the fermentation process remains a pressing technical problem to be solved in the current processing of fermented mandarin fish.

[0007] The information disclosed in this background section is only intended to enhance the understanding of the background technology of this application and should not be construed as an admission or in any way implying that the information constitutes prior art known to those skilled in the art. Summary of the Invention

[0008] To address the problems of high dependence on high-salt environments, insufficient fermentation stability under low-salt conditions, and decreased umami and textural quality in existing stinky mandarin fish processing techniques, this invention provides a low-salt umami-enhancing stinky mandarin fish processing method based on the synergistic fermentation of functionally complementary lactic acid bacteria. Through the rational selection and synergistic utilization of fermentation microorganisms, the method achieves a synergistic improvement in the flavor and quality of stinky mandarin fish while reducing salt usage, thereby meeting the demands of healthy, standardized, and industrialized production.

[0009] This invention proposes a low-salt, flavor-enhancing fermented mandarin fish processing method based on the synergistic fermentation of functionally complementary lactic acid bacteria, comprising the following steps: (1) The raw material of stinky mandarin fish is processed under low-salt conditions, wherein the low-salt conditions are 2-4 wt% saline solution; (2) Inoculate the raw material of the stinky mandarin fish with a combination of lactic acid bacteria consisting of at least one strain of Lactobacillus plantarum and at least one strain of Pediococcus pentosaceus; (3) Fermentation was carried out under anaerobic conditions at 10-20 ℃ to obtain low-salt, flavor-enhancing stinky mandarin fish; Among them, the *Lactobacillus plantarum* and the *Pediococcus pentosaceus* have complementary functions in regulating the fermentation microenvironment during metabolic behavior and fermentation. They can synergistically promote the generation of umami-related substances in stinky mandarin fish under low-salt conditions, thereby maintaining or improving the umami quality of the product while reducing the amount of salt used.

[0010] Preferably, the low-salt condition is a 3 wt% saline solution.

[0011] Preferably, both *Lactobacillus plantarum* and *Pediococcus pentosaceus* are endogenous strains isolated from mandarin fish.

[0012] Furthermore, the *Lactobacillus plantarum* and / or *Pediococcus pentosaceus* exhibit metabolic activities that contribute to protein and / or lipid degradation. These metabolic activities can be provided by a single strain or enhanced or manifested through interstrain interactions during co-fermentation.

[0013] Preferably, the inoculation mass ratio of *Lactobacillus plantarum* to *Pediococcus pentosus* is 1:1.

[0014] Furthermore, the *Lactobacillus plantarum* and *Pediococcus pentosaceus* exhibit stage-specific differences in growth and metabolic activity, thereby synergistically promoting the generation of umami precursor substances.

[0015] Preferably, the fermentation is carried out under anaerobic conditions at 15 °C for 8 days.

[0016] The *Lactobacillus plantarum* used in this invention Lactiplantibacillus plantarum LZR-1 and Pediococcus pentosaceus Pediococcus pentosaceus Both LWT-1 samples were deposited on January 8, 2026, at the China Center for Type Culture Collection (CCTCC), located at No. 299 Bayi Road, Wuchang District, Wuhan City, Hubei Province, with accession numbers CCTCC NO: M 2026036 and CCTCC NO: M 2026037, respectively.

[0017] Based on the same inventive concept, this invention provides a lactic acid bacteria combination for fermenting low-salt fermented mandarin fish, the combination including Lactobacillus plantarum. Lactiplantibacillus plantarum LZR-1 and Pediococcus pentosaceus Pediococcus pentosaceus LWT-1, wherein the Lactobacillus plantarum and Pediococcus pentosaceus have a synergistic flavor-enhancing effect under 3 wt% saline conditions.

[0018] Furthermore, both *Lactobacillus plantarum* and *Pediococcus pentosaceus* are endogenous strains isolated from mandarin fish.

[0019] In the early stages of fermentation, *Lactobacillus plantarum* can rapidly colonize and metabolize carbohydrates, promoting acidification of the fermentation system, thereby inhibiting the growth of spoilage microorganisms and improving the accessibility of proteins and lipids in the fish matrix. Furthermore, in a relatively stable low-salt, low-pH environment, *Pediococcus pentosaceus* exhibits more fully utilized protease and lipase activities, further promoting the degradation of macromolecules and the generation of umami precursors. Although some *Pediococcus pentosaceus* strains themselves possess protease and lipase activities, under single-inoculation conditions, their efficient conversion of umami substances is limited by the fermentation microenvironment and the accumulation of metabolites. However, through synergistic fermentation with *Lactobacillus plantarum*, the overall fermentation efficiency and flavor-building effect can be significantly improved, demonstrating a significantly superior low-salt umami enhancement effect compared to single-strain fermentation.

[0020] Compared with the prior art, the present invention has the following beneficial effects: (1) This invention introduces specific functionally complementary endogenous lactic acid bacteria for synergistic fermentation. Under conditions where the salt concentration is significantly lower than that of traditional fermented mandarin fish (6 wt% brine), the fermentation process can still be stably completed, avoiding the common problems of spoilage, off-flavors, and quality deterioration under low-salt conditions. Under low-salt conditions (3 wt% brine), the umami and saltiness responses of the synergistic fermentation group (LWT-1 LZR-1 3%) are significantly better than those of the high-salt natural fermentation group and the low-salt natural fermentation group. This proves that this invention can enhance umami while reducing salt, overcoming the technical difficulty of balancing safety and flavor quality in low-salt fermentation in the prior art. This result breaks through the long-standing technical prejudice in the field that "less salt leads to spoilage, and spoilage leads to a lack of freshness." Its technical effect is not something that those skilled in the art could reasonably expect without experimental inspiration. (2) The *Lactobacillus plantarum* and *Pediococcus pentosaceus* used in this invention have significant complementary metabolic functions. Their synergistic effect can promote the targeted degradation of proteins and lipids in fish meat, generating more free amino acids, umami peptides, and flavor precursors that are beneficial to the formation of umami. The fermented mandarin fish made with this mixed strain has better texture indicators such as cohesion, elasticity, chewiness, and hardness than single-strain inoculation or random mixed inoculation of multiple strains, indicating that this invention effectively improves the texture and taste quality of the product while enhancing umami. (3) This invention enables the fermentation system to rapidly produce acid and form a competitive advantage under low-salt conditions (3 wt% saline), effectively inhibiting the growth of spoilage microorganisms by directional inoculation with endogenous lactic acid bacteria. The pH and water activity of the synergistic fermentation group (LWT-1 LZR-13%) are lower than those of the natural fermentation group, which is beneficial to improving the microbial stability and storage safety of the product. The strains used in this invention have a clear source and high safety, and the process conditions are controllable. It can achieve a synergistic improvement in flavor and safety under the premise of reducing salinity, making it suitable for the standardized, large-scale, and low-salt production of stinky mandarin fish. Attached Figure Description

[0021] To more clearly illustrate the technical solution of the present invention, the accompanying drawings used in the description of the specific embodiments will be briefly introduced below. Obviously, the following description is only a part of the embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0022] Figure 1 This is a Gram staining image of the lactic acid bacteria used in this invention.

[0023] Figure 2 To investigate the effect of salt concentration on the survival rate of different strains during fermentation, measurements were taken at (a) 12 h, (b) 24 h, (c) 36 h, and (d) 48 h.

[0024] Figure 3 Electronic tongue taste maps of stinky mandarin fish under different fermentation methods: (a) PCA map of electronic tongue test; (b) radar map of electronic tongue of stinky mandarin fish.

[0025] Figure 4 Texture analysis of stinky mandarin fish fermented under different methods.

[0026] Figure 5 The determination of (a) pH, (b) moisture content and (c) water activity of stinky mandarin fish under different fermentation methods: Note: If the same indicator is labeled with different letters in different groups, it indicates that the difference is statistically significant (* indicates...). P <0.05, ** indicates P <0.01, *** indicates P <0.001, **** indicates P <0.0001). Detailed Implementation

[0027] This invention proposes a low-salt, flavor-enhancing method for processing stinky mandarin fish based on synergistic fermentation of functionally complementary lactic acid bacteria. To facilitate understanding of this invention by those skilled in the art, the specific embodiments of this invention are described below in conjunction with the accompanying drawings.

[0028] In this invention, unless otherwise specified, the equipment and raw materials used are commercially available or commonly used in the art. The methods in the following embodiments, unless otherwise specified, are conventional methods in the art. Unless otherwise defined, all technical and scientific terms used in this specification have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0029] Example 1: Isolation, purification, and characterization of endogenous bacterial strains To screen for lactic acid bacteria suitable for low-salt fermentation of mandarin fish and with complementary metabolic functions, endogenous strains were isolated from the mandarin fish itself, and then the salt tolerance and enzyme production capacity of the isolated endogenous strains were characterized.

[0030] 1.1 Experimental Methods 1.1.1 Isolation, purification and identification of endogenous strains Thaw the stinky mandarin fish sample and leave it at room temperature for 12 hours. Then, open the packaging in the sterile area of ​​the laminar flow hood and take 30 g of sample for a 10-fold serial dilution. First, mix 5.00 g of minced meat with 45.0 mL of sterile physiological saline to prepare a decimal dilution solution (10... -1 Up to 10 -6100 μL of the diluted solution was spread onto plates, and the lactic acid bacteria were cultured in MRS broth at 37.0 °C for 2–3 days. After single colonies grew, colonies were picked for microscopic examination to observe the morphology of the bacteria. Impure strains were further purified by streak plating using the original medium. After purification, the strains were numbered and further identified by 16S rRNA gene sequencing. The isolated strains were stored in 50% (v / v) glycerol at -80 °C for later use.

[0031] 1.1.2 Characterization of endogenous strains (1) Salt tolerance test The survival ability and salt tolerance of bacterial strains in mandarin fish are key indicators for strain screening. In this experiment, a culture medium with a salt concentration of 1% to 6% was selected as the conditions for detecting the salt tolerance of the strains.

[0032] MRS broth culture media with NaCl mass fractions (wt%) of 1%, 2%, 3%, 4%, 5%, and 6% were prepared, and 10 mL of each was dispensed into test tubes. The media were then autoclaved. After cooling, the different bacterial strains were prepared into 1×10⁻⁶ culture media. 8 CFU / mL bacterial suspensions were inoculated into culture media with different salt concentrations at a volume of 200 μL and incubated at 37 ℃ for 48 h. The OD values ​​at 600 nm were measured using a microplate reader at 6, 12, 36, and 48 h. Corresponding culture media were used as negative controls, and each treatment was performed in triplicate.

[0033] (2) Enzyme production capacity assay Lipase production capacity assay: The test strain was activated and cultured in the appropriate medium using neutral red oil medium, and then inoculated onto neutral red agar plates and incubated at 15 °C for 3 days. Observe whether the colony color deepens and whether discoloration occurs around the colony.

[0034] Protease production capacity assay: Prepare 3% skim milk medium, inoculate the test strain into protein screening plate medium, and incubate at 37 ℃ for 24 h. Observe whether a protein hydrolysis clear zone appears around the colony.

[0035] 1.2 Experimental Results and Analysis 1.2.1 Isolation, purification and identification of endogenous strains Three strains of lactic acid bacteria on the edible list were identified by 16S rRNA gene sequencing: one strain of *Pediococcus pentosaceus* and two strains of *Lactobacillus plantarum*, named *Pediococcus pentosaceus* LWT-1, *Lactobacillus plantarum* LZR-1, and *Lactobacillus plantarum* LZR-2, respectively. All three are Gram-positive bacteria. Figure 1 ).

[0036] 1.2.2 Characterization of endogenous strains (1) Salt tolerance like Figure 2 As shown, the OD values ​​of all three tested strains decreased significantly with increasing culture medium salt concentration. However, with prolonged culture time, the OD values ​​of the strains increased to some extent under all salt concentration conditions, and the increase trend was relatively slow with no obvious fluctuations. All three tested strains showed strong salt tolerance under salt concentration conditions of 1% to 3%. Among them, the OD values ​​of Lactobacillus plantarum LZR-1 and LZR-2 were significantly higher than those of Lactobacillus pentosus LWT-1, and the salt tolerance of LZR-1 and LZR-2 was better than that of LWT-1.

[0037] (2) Enzyme production capacity Proteases and lipases can degrade macromolecular nutrients (proteins and lipids) to generate key flavor precursors such as alcohols, esters, and ketones. These not only help improve product flavor but also, through the synergistic effect of lactic acid bacteria acid production, inhibit the growth of spoilage bacteria, thereby enhancing product safety and quality. Table 1 shows that both *Lactobacillus plantarum* and *Pediococcus pentosaceus* produce lipases, with LZR-1 and LZR-2 exhibiting stronger lipase-producing capabilities. However, in skim milk culture medium, a clear zone of proteolytic activity was only observed around *Pediococcus pentosaceus* LWT-1, while it was not observed in either of the two *Lactobacillus plantarum* strains.

[0038] Table 1. Identification of the ability of strains to produce lipase and protease

[0039] Note: "++" indicates a strong ability to produce related enzymes, "+" indicates the production of related enzymes, and "-" indicates no enzyme production.

[0040] The results showed that Pediococcus pentosaceus LWT-1 had a strong protein hydrolysis ability, while Lactobacillus plantarum LZR-1 and LZR-2 had a strong fat hydrolysis ability and salt tolerance, providing a strain basis for the subsequent construction of a functionally complementary synergistic fermentation system.

[0041] To verify the effects of different inoculation methods on the flavor and quality of stinky mandarin fish under low-salt conditions, the following Example 2 and Comparative Examples 1-5 were designed.

[0042] Example 2: LWT-1 and LZR-1 mixed inoculation group (LWT-1 LZR-1 3%) Fresh mandarin fish were slaughtered, cleaned, and neatly arranged before being placed in a special fermentation device. Dry-fried Sichuan peppercorns were sprinkled on top as a flavoring agent. The fish were submerged in brine, and 1% (by weight of the total raw material) of freeze-dried bacterial powder was inoculated and mixed thoroughly. 40% of the fish's own weight was applied to the fish, which was then covered and fermented anaerobically at 15°C for 8 days. Collected samples were stored in dry ice and transported back to the laboratory within 24 hours for storage at -20°C.

[0043] The saline solution used was 3% by mass, and the lyophilized bacterial powder used for inoculation was a 1:1 mixture of LWT-1 and LZR-1 by mass. These conditions were used to verify the effect of synergistic fermentation of functionally complementary lactic acid bacteria under low-salt conditions on the flavor and quality of stinky mandarin fish.

[0044] Comparative Example 1: Natural fermentation group (NF 6%) The difference between Comparative Example 1 and Example 2 is that the saline solution used is a 6% saline solution by mass, and no lyophilized bacterial powder is inoculated.

[0045] Comparative Example 2: Low-salt natural fermentation group (NF 3%) The difference between Comparative Example 2 and Example 2 is that no lyophilized bacterial powder was inoculated.

[0046] Comparative Example 3: LWT-1 single inoculation group (LWT-1 3%) The difference between Comparative Example 3 and Example 2 is that the lyophilized bacterial powder used for inoculation was only LWT-1.

[0047] Comparative Example 4: LWT-1 and LZR-2 co-inoculation group (LWT-1 LZR-2 3%) The difference between Comparative Example 4 and Example 2 is that the lyophilized bacterial powder used for inoculation was a mixture of LWT-1 and LZR-2 in a mass ratio of 1:1.

[0048] Comparative Example 5: Mixed inoculation group of LWT-1, LZR-1, and LZR-2 bacteria (LWT-1, LZR-1, LZR-2, 3%) The difference between Comparative Example 5 and Example 2 is that the lyophilized bacterial powder used for inoculation is LWT-1, LZR-1 and LZR-2, which are mixed in a mass ratio of 1:1:1.

[0049] Example 3 Characterization of the product after fermentation 3.1 Experimental Methods 3.1.1 Flavor Characterization 50 g of minced meat samples were incubated in a water bath at 40 ℃ for 60 min, and distilled water at 40 ℃ was added at a ratio of 1:5. The samples were then centrifuged at 10,000 g for 10 min at room temperature, and the supernatant was collected for electronic tongue testing. The test used an SA-402B system (INSENT, Japan), equipped with sensors for bitterness (COO), astringency (AE1), sourness (CAO), saltiness (CTO), and umami (AAE), as well as two reference electrodes. The negative electrode cleaning solution was 100 mM HCl and 30% ethanol, the positive electrode cleaning solution was 10 mM KOH, 100 mM KCl, and 30% ethanol, and the reference solution was 30 mM KCl and 0.3 mM tartaric acid. The sensors were calibrated and balanced before testing. The test was repeated four times, and the average of the remaining three results was taken after removing the first set of data.

[0050] 3.1.2 Physicochemical property characterization (1) Texture determination Texture characteristics of the stinky mandarin fish were analyzed using the texture apparatus multifaceted analysis (TPA) test. The fish meat from the back was cut into 1 cm pieces. 3 The small cubic blocks were used. The probe was cylindrical, with a downward speed of 1 mm / s, a return speed of 5 mm / s, a test interval of 5 s, a downward displacement of 30 mm, a compression ratio of 35%, and an initiation force of 5 g. Each sample was tested three times, and the average value was used as the measured value of each parameter.

[0051] The main parameters measured include: cohesiveness (i.e., internal cohesion), elasticity, adhesiveness, chewiness, hardness, and adhesion.

[0052] (2) pH measurement Take 5 g of fish meat tissue from the abdomen and back, remove the skin, and place it in a new sealed homogenizing bag. Then add 10 mL of ultrapure water to the sealed bag to homogenize the fish meat tissue, and measure the pH value using a pH meter.

[0053] (3) Moisture content determination Moisture content was determined by direct drying method, in accordance with national standard GB5009.3-2010.

[0054] The specific steps are as follows: First, weigh an appropriate amount of stinky mandarin fish sample (approximately 5 g) and record the initial mass (m1). Place the sample in a pre-weighed desiccator and dry it in a 105 ℃ oven until constant weight. After drying, remove the desiccator, cool it to room temperature, weigh it, and record the dried mass (m2). The moisture content is calculated using the following formula: X=(m1-m2) / m1×100 In the formula, X represents the moisture content in grams per 100 grams (g / 100g); m1 represents the initial mass of the sample in grams; and m2 represents the mass of the dried sample in grams.

[0055] (4) Water activity determination After crushing the sample, spread it evenly in the sample box, ensuring it completely covers the bottom. Open the lid and place the sample box in a water activity meter. Measure at 25±0.1 ℃. Once the reading stabilizes, the water activity of the sample can be directly read from the display screen. Perform three parallel measurements.

[0056] 3.2 Experimental Results and Analysis 3.2.1 Flavor Characterization Electronic tongues are an important tool for evaluating the umami characteristics of food. Principal component analysis of fermented mandarin fish from different fermentation groups is shown below. Figure 3 As shown in Figure a, the total value of principal component 1 (PC1) and principal component 2 (PC2) reaches 87.83%, indicating that these two components can well explain the relationship between the umami flavor of the fermented mandarin fish from different groups. NF 6% and LWT-1 LZR-1 LZR-2 3% differ significantly from other groups. The electronic tongue evaluation of fermented mandarin fish from different strains is as follows: Figure 3 As shown in b. Compared with other fermentation treatment groups, the LWT-1 and LZR-1 mixed inoculation group (LWT-1 LZR-1 3%) had the highest umami and saltiness scores and the lowest sourness score, showing a more ideal overall taste. This indicates that the fermented mandarin fish fermented with LWT-1 and LZR-1 strains at a salt concentration of 3% had higher umami and saltiness than the naturally fermented mandarin fish fermented with salt concentrations of 6% and 3% and the mandarin fish fermented with other strains at a salt concentration of 3%, and had a better overall taste. This shows that the fermentation effect was better, and the fermentation with LWT-1 and LZR-1 strains achieved the effect of enhancing the umami and reducing the salt in the fermented mandarin fish.

[0057] 3.2.2 Physicochemical characterization (1) Texture Cohesiveness reflects the ability of fish meat to resist damage and maintain its integrity during chewing, and also reflects the strength of intercellular binding forces, but its nature is opposite to that reflected by adhesiveness; elasticity reflects the degree of deformation under external force and the degree of recovery after the force is removed. The elasticity of fish meat is related to the strength of intermuscular binding forces. The greater the binding force (the less damage to muscle tissue), the greater the elasticity; adhesiveness is used to describe the flowability parameters of semi-solid foods under a certain force; chewiness is a comprehensive quality evaluation index, simulating the energy required for fish meat samples to reach a stable state when chewed and swallowed (i.e., "bite force"), and is the result of the combined effects of muscle hardness, intercellular cohesiveness, and elasticity; hardness is expressed as the softness or hardness felt by the human body, reflecting the force required to make food reach a certain deformation, that is, the internal binding force of food to maintain its shape; adhesiveness reflects the internal binding force or surface stickiness of fish meat, affecting its processing performance and texture, and is usually closely related to the gelation ability and tissue structure of muscle proteins.

[0058] Differences in texture among six different fermentation groups of stinky mandarin fish, such as Figure 4 As shown. By Figure 4 It can be seen that the LWT-1 LZR-1 3% sample has the greatest cohesiveness, elasticity, chewiness and hardness, indicating that the internal structure of the LWT-1 LZR-1 3% sample is more compact and stable, with strong compressive strength and good elastic recovery performance. It requires more energy during chewing and exhibits the characteristics of a relatively compact texture, good elasticity and complete structure.

[0059] (2) pH Physicochemical properties of six groups of fermented mandarin fish were tested. pH values ​​were measured after different fermentation groups of mandarin fish were treated according to the specified procedures. Each sample was tested in triplicate. The results are as follows: Figure 5 As shown in Figure a, during the fermentation of stinky mandarin fish, a moderate decrease in pH not only helps inhibit the growth of spoilage microorganisms and improve product safety, but also affects protein degradation and flavor compound formation. Among the six fermentation groups, LWT-1 and LZR-1 with a pH of 3% had the lowest pH value, indicating that stinky mandarin fish fermented with LWT-1 and LZR-1 strains had the strongest ability to inhibit the growth of spoilage microorganisms.

[0060] (3) Moisture content Moisture content generally reflects the water-holding capacity, tenderness, and freshness of fish meat. For example... Figure 5 As shown in b, there was no significant difference in the moisture content of the six groups of fermented mandarin fish, indicating that under the same fermentation conditions, different salinity values ​​and strains had little impact on the overall water retention of the samples.

[0061] (4) Water activity Water activity (Aw) is an important parameter for fish meat quality control. Figure 5As shown in Figure c, the water activity of LWT-1 and LZR-1 was at its lowest value of 3%, but there was no significant difference among the four groups of fermented mandarin fish using these strains. This study indicates that more free water participating in protein hydrolysis leads to a decrease in water activity. Furthermore, microbial growth is closely related to water activity; lower water activity may be more conducive to the control of contaminating microorganisms.

[0062] In summary, the LWT-1 LZR-1 3% group achieved the highest scores in umami and saltiness, and exhibited greater cohesion, elasticity, chewiness, and firmness, while also having lower pH and water activity values. This indicates that the co-fermentation of LWT-1 and LZR-1 strains can enhance the overall umami perception by promoting the production of umami substances, achieving a "flavor enhancement and salt reduction" effect while lowering salinity; simultaneously, the lower pH and water activity contribute to improved product stability. Therefore, the LWT-1 and LZR-1 strains were preserved. The 16S rRNA gene sequence of the LWT-1 strain is shown in SEQ ID NO.1, and the 16S rRNA gene sequence of the LZR-1 strain is shown in SEQ ID NO.2.

[0063] The embodiments of the present invention described above do not constitute a limitation on the scope of protection of the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the scope of protection of the claims of the present invention.

Claims

1. A method for processing low-salt, flavor-enhancing fermented mandarin fish based on synergistic fermentation of functionally complementary lactic acid bacteria, characterized in that, Includes the following steps: (1) The raw material of stinky mandarin fish is processed under low-salt conditions, wherein the low-salt conditions are 2-4 wt% saline solution; (2) Inoculate the raw material of the stinky mandarin fish with a combination of lactic acid bacteria consisting of Lactobacillus plantarum and Pediococcus pentosaceus; The *Lactobacillus plantarum* is Lactiplantibacillus plantarum LZR-1 was deposited at the China Center for Type Culture Collection on January 8, 2026, with accession number CCTCC NO: M 2026036; The Pediococcus pentosaceus is Pediococcus pentosaceus LWT-1 was deposited at the China Center for Type Culture Collection on January 8, 2026, with accession number CCTCC NO: M 2026037; The inoculation mass ratio of *Lactobacillus plantarum* to *Pediococcus pentosus* was 1:

1. (3) Fermentation was carried out under anaerobic conditions at 10-20 ℃ to obtain low-salt stinky mandarin fish.

2. The method according to claim 1, characterized in that: The low-salt condition is a 3 wt% saline solution.

3. The method according to claim 1 or 2, characterized in that: Both *Lactobacillus plantarum* and *Pediococcus pentosus* are endogenous strains isolated from mandarin fish.

4. The method according to claim 3, characterized in that: The *Lactobacillus plantarum* and / or *Pediococcus pentosus* exhibit metabolic activities that contribute to the degradation of proteins and / or lipids.

5. The method according to claim 1, characterized in that: The fermentation was carried out for 8 days under anaerobic conditions at 15 °C.

6. A lactic acid bacteria combination for fermenting low-salt fermented mandarin fish, characterized in that: This combination consists of Lactobacillus plantarum Lactiplantibacillus plantarum LZR-1 and Pediococcus pentosaceus Pediococcus pentosaceus LWT-1 is composed of Lactobacillus plantarum and Pediococcus pentosus in an inoculation mass ratio of 1:1; The *Lactobacillus plantarum* is Lactiplantibacillus plantarum LZR-1 was deposited at the China Center for Type Culture Collection on January 8, 2026, with accession number CCTCC NO: M 2026036; The Pediococcus pentosaceus is Pediococcus pentosaceus LWT-1 was deposited at the China Center for Type Culture Collection on January 8, 2026, with accession number CCTCC NO: M 2026037.

7. The lactic acid bacteria combination according to claim 6, characterized in that: The *Lactobacillus plantarum* and *Pediococcus pentosus* strains are endogenous strains isolated from mandarin fish.

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