A taste peptide, a preparation method and a method for adjusting the taste of instant fish sauce

Abstract

This invention discloses a flavor peptide, its preparation method, and a method for adjusting the flavor of quick-brewed fish sauce, belonging to the field of biotechnology. The flavor peptide is extracted from fermented soybean paste. The flavor peptide is used to adjust the flavor of quick-brewed fish sauce: using freshwater fish and / or its processing by-products as raw materials, processing them into a meat paste, adding enzymes for enzymatic hydrolysis, inactivating the enzymes, adding the flavor peptide, inoculating with microbial agents for fermentation, and finally filtering and sterilizing to obtain the flavor-adjusted quick-brewed fish sauce. This method significantly improves the umami and thickness of quick-brewed fish sauce and harmonizes the taste and flavor of the fish sauce.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, specifically to a flavor peptide and its preparation method, as well as a method for adjusting the flavor of quick-brewed fish sauce. Background Technology

[0002] Flavor peptides are a class of small molecule peptides with flavor-enhancing properties, typically with a relative molecular mass of 500-1500 Da. They are generally extracted from natural foods or synthesized using amino acids. The flavor-enhancing effect of flavor peptides mainly depends on the amino acid sequence, peptide chain length, and arrangement structure. Different peptides produce different taste sensations and flavor intensities. When used in conjunction with umami and acidulants, they can enhance the original flavor, strengthen the umami and richness of food, improve the flavor intensity, and increase the nutritional value of food. In addition to their flavor-enhancing effects, as small molecule peptides, they are absorbed and utilized by the human body more efficiently than large protein molecules and free amino acids. They also possess excellent nutritional properties, solving the problem of insufficient flavor richness from single seasoning bases and contributing to improved food quality. Therefore, peptides have significant market potential as a flavor base or excipient in the food industry.

[0003] Pixian Doubanjiang (fermented broad bean paste) is spicy, fragrant, thick, and glossy with a reddish-brown color. With a long history, it is known as the "soul of Sichuan cuisine" and is one of the core seasonings of Sichuan cuisine. The unique components and flavor of Pixian Doubanjiang are formed through the metabolic processes of microorganisms and the interactions between various chemical components. Currently, most research focuses on volatile components and amino acids, with few reports on the isolation of flavor peptides from Doubanjiang. Summary of the Invention

[0004] The purpose of this invention is to provide a flavor peptide that enhances both saltiness and umami, and can be used to adjust the flavor of quick-brewed fish sauce, significantly improving its umami and richness, and harmonizing its taste and flavor.

[0005] This invention is achieved through the following technical solution:

[0006] A flavor peptide with the following amino acid sequence: Asp-Ile-Tyr-Asn-Pro-Gln-Ala-Gly-Arg.

[0007] Preferably, the flavor peptides mentioned above are extracted from fermented soybean paste.

[0008] The preparation method of this flavor peptide includes the following steps:

[0009] S1. After beating the fermented soybean paste into a paste, crude peptide freeze-dried powder was obtained by water extraction and defatting with petroleum ether.

[0010] S2. Ultrafiltration is performed on the crude peptide lyophilized powder to separate and retain ultrafiltration components with a molecular weight cutoff of less than 5 kDa;

[0011] S3. The ultrafiltration fraction was subjected to gel chromatography.

[0012] S4. The components obtained after chromatography were separated and purified by liquid chromatography-mass spectrometry to obtain flavor peptides.

[0013] Furthermore, in the preparation method of the flavor peptide,

[0014] In step S1, the fermented soybean paste is added to ultrapure water at a mass ratio of 1:3~5, shaken in a water bath at 37~40℃ for 1~2 hours, centrifuged at 4℃ and 8000 r / min for 20~30 minutes, filtered through a 0.22μm microporous membrane, and then the filtrate is degreased by adding petroleum ether at a volume ratio of 1:1~2. The lower filtrate is then freeze-dried to obtain freeze-dried powder.

[0015] In step S2, the lyophilized powder is dissolved in distilled water, filtered through a 0.22 μm microporous membrane, and then ultrafiltration is used to separate and retain ultrafiltration components with a molecular weight cutoff of less than 5 kDa. The ultrafiltration components are then freeze-dried and stored at -18°C.

[0016] In step S3, the freeze-dried ultrafiltration fraction is prepared into a solution of 20-30 mg / ml and subjected to chromatography using Sephdex G-15 dextran gel.

[0017] In step S4, the ultrafiltration components with the strongest flavor and richness obtained after chromatography are separated and identified by liquid chromatography-mass spectrometry, and flavor peptides are obtained by solid-phase synthesis.

[0018] Furthermore, to ensure the separation effect of peptides, in step S3, the sample loading volume of Sephdex G-15 dextran gel chromatography is 2~4 ml.

[0019] A method for adjusting the flavor of quick-brewed fish sauce using flavor peptides includes the following steps:

[0020] a. Using freshwater fish and / or their processing byproducts as raw materials, processing them into meat paste;

[0021] b. After adding neutral protease to the meat paste for enzymatic hydrolysis, flavor protease is added for further enzymatic hydrolysis. After the enzymatic hydrolysis is completed, enzyme inactivation treatment is performed to obtain the enzymatically hydrolyzed raw material.

[0022] c. Add the flavor peptide as described in claim 1 to the enzymatically hydrolyzed raw material, then inoculate with microbial agents and carry out incubation fermentation to obtain fermented raw material;

[0023] d. Then filter and sterilize the fermented raw materials to obtain the flavor-adjusted quick-brewed fish sauce.

[0024] Furthermore, in step b, the amount of neutral protease added is 2-3% of the meat sauce mass, and the enzymatic hydrolysis process is carried out by stirring at 120-140 rpm and under constant temperature conditions of 45-50℃ for 12-16 hours.

[0025] Furthermore, in step b, the amount of flavor protease added is 2-3% of the meat sauce mass, and the enzymatic hydrolysis is carried out at a rotation speed of 120-140 rpm and a constant temperature of 50-55°C for 10-12 hours.

[0026] Furthermore, in step c, the amount of flavor peptide added is 0.05~0.1% of the mass of the enzymatically hydrolyzed raw material.

[0027] Furthermore, in step c, the microbial agent includes *Lactobacillus plantarum* (…). Lactiplantibacillus plantarum SR2, viable count 10 6 ~10 7 The inoculum concentration was CFU / mL, and the inoculum amount was 5-10% of the fish sauce raw material mass. The Lactobacillus plantarum SR2 strain was a newly isolated strain and was deposited at the Guangdong Provincial Microbial Culture Collection Center on August 11, 2022, with accession number GDMCC No. 62431.

[0028] Furthermore, in step c, the heat preservation fermentation process involves static fermentation at 35-40℃ for 60-90 days, followed by heating at 90-100℃ for 10-15 minutes to terminate the fermentation.

[0029] Compared with the prior art, the present invention has the following advantages and beneficial effects:

[0030] (1) In this invention, a polypeptide with salty flavor and umami enhancement effect is provided, and its amino acid sequence is disclosed. At the same time, a method for obtaining the polypeptide is provided. The polypeptide can be obtained according to a specific preparation method. The polypeptide can be used to adjust the flavor of quick-brewed fish sauce, significantly improve the umami and thickness of quick-brewed fish sauce, and coordinate the taste and flavor of fish sauce.

[0031] (2) In this invention, a method for regulating the flavor of quick-brewed fish sauce using flavor peptides is proposed, which utilizes microbial agents, including Lactobacillus plantarum (…). Lactiplantibacillus plantarum SR2, the Lactobacillus plantarum strain SR2 is a newly isolated strain in the laboratory. The quick-brewed fish sauce fermented by this strain has better umami and richness. Attached Figure Description

[0032] Figure 1 The chromatograms are those of ultrafiltration fractions with molecular weights below 5 kDa after separation by Sephadex G-15 gel chromatography.

[0033] Figure 2 The results of the taste dilution analysis of ultrafiltration component F and chromatography components F1, F2, and F3 are presented.

[0034] Figure 3 This is the secondary mass spectrum of the DIYNPQAGR sequence.

[0035] Figure 4 The image shows the results of electronic tongue assays for the polypeptide DIYNPQAGR. Detailed Implementation

[0036] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments and accompanying drawings. It should be understood that these descriptions are merely exemplary and not intended to limit the scope of the invention. Furthermore, descriptions of well-known structures and techniques are omitted in the following description to avoid unnecessarily obscuring the concept of the invention. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer are followed. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0037] This invention provides a flavor peptide with the amino acid sequence shown in SEQ ID NO: 1. The flavor peptide is extracted from fermented soybean paste.

[0038] This invention provides a method for preparing flavor peptides, wherein the amino acid sequence of the flavor peptides in this embodiment is the same as that shown in SEQ ID NO: 1 above, and includes the following steps:

[0039] Step 1: Grind the fermented soybean paste into a paste; add ultrapure water to the ground fermented soybean paste at a mass ratio of 1:3~5, shake in a water bath at 37~40℃ for 1~2 hours, centrifuge at 4℃ and 8000 r / min for 20~30 minutes, then filter through a 0.22μm microporous membrane and collect the filtrate; dispense the filtrate into a separatory funnel and add petroleum ether at a volume ratio of 1:1~2 for defatting, then collect the lower layer of filtrate and freeze-dry to obtain freeze-dried powder;

[0040] Step 2: Dissolve the lyophilized powder in distilled water, filter it through a 0.22μm microporous membrane, then use ultrafiltration to separate and retain the ultrafiltration component F with a molecular weight cutoff of less than 5kDa, freeze-dry it and store it at -18℃.

[0041] Step 3: Prepare an ultrafiltration fraction F solution of 20-30 mg / ml and filter it through a 0.22 μm aqueous microporous membrane, then perform chromatography using Sephdex G-15 dextran gel.

[0042] Step 4: The components with the strongest flavor and richness obtained after chromatography were separated and identified by liquid chromatography-mass spectrometry. Flavor peptides were then obtained through solid-phase synthesis, and their amino acid sequences are as follows:

[0043] The polypeptide Asp-Ile-Tyr-Asn-Pro-Gln-Ala-Gly-Arg (abbreviated as DIYNPQAGR).

[0044] In step 3, the sample loading volume of Sephadex G-15 gel chromatography is 2-4 ml, the elution flow rate is 1-2 ml / min, and the detection wavelength is 220 nm to ensure the separation effect of peptides.

[0045] In the following specific embodiments of the present invention, the ultrafiltration components are subjected to Sephadex G-15 gel chromatography to select the components with the strongest flavor and richness (evaluated by human sensory perception), and then further separated and identified by liquid chromatography-mass spectrometry.

[0046] The human sensory evaluation uses taste dilution analysis (TDA) to determine the dilution factor, or TD value, of each sample. The higher the TD value, the stronger the taste characteristics of the sample.

[0047] The TD value is determined as follows: The component to be tested is dissolved in an equal proportion of pure water and diluted 1:1 (volume ratio). A three-point test method is used to evaluate each dilution level until the taste is no longer perceptible at a certain dilution factor. This dilution factor is the taste dilution factor (TD) of the solution; the higher the TD value, the stronger the taste of the component. Sensory evaluation is conducted by eight sensory evaluators (4 men and 4 women, aged 25 to 30), who are trained before the experiment.

[0048] This invention proposes a method for regulating the flavor of quick-brewed fish sauce using flavor peptides, comprising the following steps:

[0049] Step 1: Use freshwater fish (in actual production, freshwater fish and / or their processing by-products can also be used) as raw material and process them into meat paste.

[0050] Step 2: Add neutral protease to the meat paste for enzymatic hydrolysis, the amount of which is 2-3% of the raw meat. Hydrolyze for 12-16 hours at a speed of 120-140 rpm and a constant temperature of 45-50℃. Then add flavor protease for enzymatic hydrolysis, the amount of which is 2-3% of the raw meat. Hydrolyze for 10-12 hours at a speed of 120-140 rpm and a constant temperature of 50-55℃. After the enzymatic hydrolysis is completed, perform enzyme inactivation treatment to obtain the enzymatically hydrolyzed raw material.

[0051] Step 3: Add flavor peptides with the amino acid sequence of Example 1 to the enzymatic hydrolysate, the amount of which is 0.05~0.1% of the enzymatic hydrolysate; then inoculate with microbial agents for fermentation, the amount of which is 5~10% of the fish sauce raw material mass, and keep fermented at 35~40℃ for 30~40 days, then heat at 90~100℃ for 10~15 minutes to stop fermentation, and filter to obtain fermented raw material;

[0052] Step 4: Use eight layers of gauze to filter out large particulate impurities from the fermentation raw material obtained in Step 4, and then filter it through a microporous membrane with a pore size of 0.45μm. Sterilize the filtrate at 121~126℃ for 15~20min, and seal it in a can under aseptic conditions.

[0053] Preferably, the microbial agent is composed of Lactobacillus plantarum (… Lactiplantibacillus plantarum Composed of SR2, with a viable count of 10. 6 ~10 7 The concentration of CFU / mL is preferred, as stated by the *Lactobacillus plantarum* (…). Lactiplantibacillus plantarum SR2 is a newly isolated strain obtained in the laboratory and was deposited at the Guangdong Provincial Microbial Culture Collection Center on August 11, 2022, with accession number GDMCC No. 62431. *Lactobacillus plantarum* ( Lactiplantibacillus plantarum The gene sequence of SR2 is shown in SEQ ID NO: 2.

[0054] Example 1

[0055] A method for the extraction, separation, and purification of flavor peptides, the specific steps of which are as follows:

[0056] Step 1: Use a blender to blend the fermented soybean paste.

[0057] Step 2: Add ultrapure water to the fermented soybean paste at a ratio of 1:4, shake in a 40℃ water bath for 1 hour, centrifuge at 4℃ and 8000 r / min for 20 minutes, filter with a 0.22μm microporous membrane, dispense the filtrate into a separatory funnel, add petroleum ether at a volume ratio of 1:1 for defatting, and then collect the lower filtrate for freeze-drying to obtain freeze-dried powder.

[0058] Step 3: Dissolve the lyophilized powder in distilled water, filter it through a 0.22μm microporous membrane, and then use ultrafiltration to separate and retain the ultrafiltration component F with a molecular weight cutoff of less than 5kDa. After freeze-drying, store it at -18℃.

[0059] Step 4: Prepare a 20 mg / ml solution of ultrafiltration fraction F and filter it through a 0.22 μm aqueous microporous membrane. Perform chromatography using Sephdex G-15 dextran gel chromatography with double-distilled water as the eluent. The chromatography method is as follows: elution flow rate 1 ml / min, detection wavelength 220 nm, and sample loading volume 4 ml. The chromatographic results are shown below. Figure 1The horizontal axis represents the elution time (min), and the vertical axis represents the detection wavelength of 220nm.

[0060] like Figure 1 The three separated components obtained by the chromatography collection and elution process were labeled as F1, F2, and F3, respectively, and then freeze-dried and stored at -20°C for later use.

[0061] Step 5: The F fraction obtained after chromatography, as well as fractions F1, F2, and F3, were subjected to artificial sensory evaluation using taste dilution analysis (TDA). The results are shown in [Figure 5]. Figure 2 The horizontal axis represents each component, and the vertical axis represents the flavor intensity dilution factor of each component.

[0062] like Figure 2 Components F and F2 have the highest TD values ​​and the strongest taste within the same range.

[0063] The results of the artificial sensory evaluation of the taste characteristics of ultrafiltration component F and chromatography components F1, F2, and F3 are shown in Table 1. As can be seen from Table 1, the taste descriptions of chromatography component F2 and component F are the most similar, with obvious fermentation flavor and rich taste.

[0064] Table 1 Sensory evaluation results

[0065]

[0066] comprehensive Figure 2 Based on Table 1, it can be concluded that the F2 component has the most intense peptide flavor and is most similar to the ultrafiltration component F. Therefore, the F2 component was selected for the next step of separation and purification.

[0067] Step 6: The F2 ultrafiltration fraction was separated and purified using liquid chromatography-mass spectrometry (LC-MS). LC conditions: C18 column (75 μm x 25 cm, Thermo, USA); mobile phase A: 2% ACN with 0.1% formic acid; mobile phase B: 80% ACN with 0.1% formic acid; gradient elution: 0-1 min (0-5% B), 1-41 min (5%-23% B), 41-51 min (23%-29%); flow rate: 300 nL / min; sample loading volume: 10 μL.

[0068] Mass spectrometry conditions were as follows: MS scan range (m / z) 200-2000, acquisition mode DDA; Top 20 (selecting the 20 strongest signals from the precursor ions for secondary fragmentation); primary mass spectrometry resolution 70000, AGC target 3e6, maximum injection time 20 ms, fragmentation mode HCD; secondary mass spectrometry resolution 17500, AGC target 1e5, maximum injection time 50 ms, fixed first mass: 100 m / z; minimum AGC target 1e3, intensity threshold 2e4, dynamic exclusion time 18 s.

[0069] The amino acid sequence of the isolated polypeptide was determined to be:

[0070] Asp-Ile-Tyr-Asn-Pro-Gln-Ala-Gly-Arg (DIYNPQAGR).

[0071] like Figure 3 The image shows the secondary mass spectrum of this component. The table below contains information about this polypeptide:

[0072] Table 2 Peptide Information

[0073]

[0074] Furthermore, the characteristics of the isolated flavor peptides were further investigated.

[0075] (1) Flavor threshold and characteristics of polypeptides

[0076] Prior to the experiment, the evaluation team members were trained. The reference standards for umami, sweetness, saltiness, bitterness, and sourness were monosodium glutamate solution (0.35%, w / v), sucrose solution (1%, w / v), sodium chloride (0.35%, w / v), L-isoleucine solution (0.25%, w / v), and citric acid solution (0.08%, w / v), respectively, in order to achieve more accurate sensory evaluation results.

[0077] The obtained peptide, with an amino acid sequence as shown in SEQ ID NO: 1, was prepared into a 1% synthetic peptide solution. Following the triangulation method, it was progressively diluted 1:1, with each dilution level evaluated using three measurements until it was just distinguishable from two blank controls (pure water). The sample concentration at this point was the taste threshold. The stimulus threshold for each evaluation group was the average of the results from all evaluators. The results are shown in Table 3. Simultaneously, group members were required to verbally describe their taste experiences, including umami, sweetness, sourness, saltiness, and bitterness.

[0078] Table 3. Taste characteristics and threshold values ​​of synthetic peptide aqueous solutions

[0079]

[0080] The results in the table show that the taste threshold of the DIYNPQAGR sequence is low, at 0.32 mg / mL, which is lower than the umami standard of 3.5 mg / mL sodium glutamate solution, indicating a good taste effect. It also presents a pleasant taste sensation and a harmonious mouthfeel, suggesting it may be a salty peptide.

[0081] (2) Electronic tongue verification of peptide flavor characteristics

[0082] The synthetic peptide was dissolved in purified water to obtain two solutions with concentrations of 1 mg / mL and 0.5 mg / mL. The solutions were analyzed using a taste analysis system (model: TS-5000Z) from INSENT Corporation, Japan, with a mixture of 30 mM potassium chloride and 0.3 mM tartaric acid as a reference solution. The results are as follows: Figure 4 .

[0083] (3) Umami-enhancing effect of peptides

[0084] Using the umami flavor of a 0.35% monosodium glutamate (MSG) solution as a sensory standard with a score of 5, the polypeptide obtained in Example 1 was added to the MSG solution, resulting in a polypeptide concentration of 1 mg / mL in the 0.35% MSG solution. The umami flavor enhancement effect of the sample was then evaluated, and the final results were averaged. The results are shown in Table 5.

[0085] Table 4. Umami-enhancing effects of peptides

[0086]

[0087] As shown in Table 4, the DIYNPQAGR sequence has a significant umami enhancement effect.

[0088] Example 2

[0089] This embodiment describes a method for adjusting the flavor of quick-brewed fish sauce using flavor peptides, including the following steps:

[0090] Step 1: Use freshwater fish as raw material and process it into a paste.

[0091] Step 2: Add neutral protease to the meat paste for enzymatic hydrolysis at a rate of 2% of the raw meat. Hydrolyze at 120 rpm and 50°C for 14 hours. Then add flavor protease for enzymatic hydrolysis at a rate of 3% of the raw meat. Hydrolyze at 120 rpm and 55°C for 10 hours. After hydrolysis, perform enzyme inactivation treatment to obtain the hydrolyzed raw material.

[0092] Step 3: Add polypeptides to the enzymatic hydrolysate at a rate of 5% of the raw material's mass; then inoculate with microbial agents for fermentation at a rate of 5% of the fish sauce raw material's mass. Ferment at 37°C for 90 days, then heat at 100°C for 15 minutes to terminate the fermentation; filter to obtain the fermented raw material.

[0093] Step 4: Use eight layers of gauze to filter out large particulate impurities from the fermentation raw materials, then filter through a microporous membrane with a pore size of 0.45μm. Sterilize the filtrate at 121℃ for 20 minutes, and seal and bottle it under aseptic conditions.

[0094] In this embodiment, in step 3, the microbial agent is *Lactobacillus plantarum* (…). Lactiplantibacillus plantarum SR2, viable count 10 7 CFU / mL.

[0095] In this embodiment, the polypeptide in step 3 has the following amino acid sequence as obtained in Example 1:

[0096] A polypeptide of Asp-Ile-Tyr-Asn-Pro-Gln-Ala-Gly-Arg.

[0097] Comparative Example 1

[0098] Replace the polypeptide in step 3 of Example 2 with pure water, and the remaining steps are the same as in Example 3.

[0099] Comparative Example 2

[0100] Compared with Example 2, the difference in this comparative example is that the microbial agent in step 3 is replaced with Lactobacillus plantarum CICC 21804, which was purchased from the China Industrial Microbial Culture Collection Center. The remaining steps are exactly the same as in Example 2, and the flavor-adjusted quick-brewed fish sauce is obtained.

[0101] The flavor of the quick-brewed fish sauce obtained in Example 2 and Comparative Examples 1-2 was investigated.

[0102] 1. Physicochemical properties

[0103] The total soluble nitrogen, amino acid nitrogen, and volatile basic nitrogen content of the fish sauce products prepared in Example 2 and the comparative example were determined at different fermentation stages.

[0104] Measurement method:

[0105] The determination of total soluble nitrogen shall be performed according to the Kjeldahl method in GB / T 5009.5—2016;

[0106] The determination of amino acid nitrogen content shall be in accordance with GB5009.235-2016;

[0107] The determination of volatile basic nitrogen content shall be in accordance with GB5009.228-2016.

[0108] The measurement results are shown in Table 5.

[0109] Table 5. Results of physicochemical index determination for Example 2 and the comparative example.

[0110]

[0111] The results in the table above show that the total soluble nitrogen and amino acid nitrogen content in the fish sauce of Example 2 are significantly higher than those in the comparative group; while the total soluble nitrogen and amino acid nitrogen content in the fish sauce of Comparative Example 2 are both higher than those in Comparative Example 1. Furthermore, the fish sauce of Example 2 can reach the standard of Grade 1 fish sauce (total soluble nitrogen ≥ 1.2 g / 100 mL, amino acid nitrogen ≥ 0.9 g / 100 mL) after 2 months of fermentation, while the fish sauce of Comparative Example 2 requires 3 months to reach the standard of Grade 2 fish sauce (total soluble nitrogen ≥ 0.87 g / 100 mL, amino acid nitrogen ≥ 0.65 g / 100 mL), and Comparative Example 1 requires even longer. The experimental data demonstrate that the flavor peptides and *Lactobacillus plantarum* SR2 described in this invention help shorten the brewing time of fish sauce, and the finished product can far exceed the Grade 1 fish sauce standard in the industry.

[0112] 2. Sensory evaluation

[0113] Evaluation method: A scoring system was used to evaluate and score the caramel flavor, meat flavor, umami flavor, sour flavor, bitter flavor, fishy flavor, ammonia flavor, and putrid flavor of the fish sauce prepared in Example 2 and Comparative Example 1. "0" points represent that the flavor cannot be perceived, and "10" points represent that the flavor is very strong.

[0114] The test results are shown in Table 6.

[0115] Table 6 Sensory evaluation results of Example 2 and Comparative Example 1

[0116]

[0117] Compared to Comparative Example 1, Examples 2 and 2 exhibit more prominent caramel, meat, and umami flavors, while unpleasant fishy, ​​ammonia, and putrid odors are almost imperceptible. Compared to the instant fish sauce without added peptides, the fish sauce of Example 2 demonstrates superior sensory characteristics. The main difference between Comparative Example 2 and Example 2 lies in the unpleasant odors such as bitterness, fishiness, ammonia, and putridity. It is evident that the peptides described in this invention can significantly enhance the umami, meat, and caramel flavors of the instant fish sauce, while Lactobacillus plantarum SR2 can effectively reduce the impact of unpleasant odors.

[0118] 3. Determination of volatile flavor compounds by GC-MS

[0119] The types and quantities of volatile aroma compounds were determined using GCMS, and the results are shown in Table 7.

[0120] Table 7 Comparison of the types and quantities of volatile aroma compounds in finished fish sauce (unit: species)

[0121]

[0122] As shown in Table 5, the content of volatile aroma compounds in the fish sauce of Example 2 was significantly increased compared to Comparative Example 1. The changes in aldehydes, hydrocarbons, and furans were the most pronounced, resulting in a pleasant aroma and greatly enhancing the flavor of the quick-brewed fish sauce. Furthermore, the test results of Comparative Example 2 fell between those of Example 2 and Comparative Example 1, indicating that the combination of *Lactobacillus plantarum* SR2 and the flavor peptides further promoted the formation of volatile flavor compounds, contributing to the harmonization of the flavor in the quick-brewed fish sauce.

[0123] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any way. Any simple modifications or equivalent changes made to the above embodiments based on the technical essence of the present invention shall fall within the protection scope of the present invention.

Claims

1. A flavor peptide, characterized in that: Its amino acid sequence is: Asp-Ile-Tyr-Asn-Pro-Gln-Ala-Gly-Arg.

2. A method for regulating the flavor of quick-brewed fish sauce using flavor peptides, characterized in that, Includes the following steps: a. Using freshwater fish as raw material, processing it into meat paste; b. After adding neutral protease to the meat paste for enzymatic hydrolysis, flavor protease is added for further enzymatic hydrolysis. After the enzymatic hydrolysis is completed, enzyme inactivation treatment is performed to obtain the enzymatically hydrolyzed raw material. c. Add the flavor peptide as described in claim 1 to the enzymatic hydrolysate, then inoculate with *Lactobacillus plantarum* SR2 and incubate for fermentation to obtain the fermentation feedstock with a viable count of 10-1. 6 ~10 7 The inoculation amount was 5-10% of the fish sauce raw material mass, and the *Lactobacillus plantarum* SR2 strain was a newly isolated strain, which was deposited at the Guangdong Provincial Microbial Culture Collection Center on August 11, 2022, with accession number GDMCC No. 62431. d. Then filter and sterilize the fermented raw materials to obtain the flavor-adjusted quick-brewed fish sauce.

3. The method for adjusting the flavor of quick-brewed fish sauce with flavor-enhancing peptides according to claim 2, characterized in that: In step b, the amount of neutral protease added is 2-3% of the meat sauce mass, and the enzymatic hydrolysis process is carried out by stirring at 120-140 rpm and under constant temperature of 45-50℃ for 12-16 hours.

4. The method for adjusting the flavor of quick-brewed fish sauce with flavor-enhancing peptides according to claim 2, characterized in that: In step b, the amount of flavor protease added is 2-3% of the meat sauce mass, and the enzymatic hydrolysis is carried out at a rotation speed of 120-140 rpm and a constant temperature of 50-55℃ for 10-12 hours.

5. The method for adjusting the flavor of quick-brewed fish sauce with flavor-enhancing peptides according to claim 2, characterized in that: In step c, the amount of flavor peptide added is 0.05~0.1% of the mass of the enzymatically hydrolyzed raw material.

6. The method for regulating the flavor of quick-brewed fish sauce with flavor-enhancing peptides according to claim 2, characterized in that: In step c, the heat preservation fermentation process involves static fermentation at 35-40℃ for 60-90 days, followed by heating at 90-100℃ for 10-15 minutes to terminate the fermentation.

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