Soy sauce-derived umami peptide and use thereof

Abstract

This invention discloses a umami peptide derived from soy sauce, belonging to the field of bioactive peptides. The amino acid sequence of the umami peptide is GAAGAAD. This invention identifies and screens a novel umami peptide from soy sauce peptide components eluted with 60% ethanol using virtual screening technology. The synthesized umami peptide, verified by an electronic tongue, exhibits good umami flavor and a synergistic umami-enhancing effect with monosodium glutamate (MSG). The key amino acids and mechanism of action of the umami peptide in binding to the umami receptors T1R1 / T1R3 were also investigated, providing a reference for studying the umami-enhancing mechanism of umami peptides. This peptide has a small molecular weight, is safe, non-toxic, and non-allergenic, and has good water solubility, making it valuable for use as a condiment or food additive in food.

Description

Technical Field

[0001] This invention belongs to the field of bioactive peptides, and specifically relates to a umami peptide. Background Technology

[0002] Umami is considered the "fifth basic taste" after sour, sweet, bitter, and salty. Umami not only enhances the richness and smoothness of food but also reduces sourness and masks unpleasant odors. Research indicates that organic acids, nucleotides, amino acids, and umami peptides are the main components that produce umami. Among them, umami peptides have attracted much attention due to their safety, rich flavor, good processing properties, and low taste threshold.

[0003] Traditional methods for screening umami peptides involve isolation, purification, and identification, followed by sensory evaluation or electronic tongue assessment of the peptide's umami flavor. These methods are time-consuming and expensive, typically limiting the efficiency of identifying and discovering new umami peptides. Recently, machine learning simulation tools based on peptide sequences and amino acid composition, including online tools such as BIO-UWM, iUmami-SCM, Umami_YYDS, UMPred-FRL, and Umami-MRNN, have been widely used for peptide umami screening and prediction. Among them, iUmami-SCM, UMPred-FRL, and Umami-MRNN achieved accuracies of approximately 88.9%, 86.7%, and 93% in predicting umami flavor, respectively. Therefore, virtual screening tools have accelerated the efficiency and accuracy of umami peptide screening. However, research on using virtual tools to screen soy sauce umami peptides has not yet been reported.

[0004] Umami peptides not only possess umami flavor but also enhance it. Previous studies have shown that some peptides have been found to have umami flavor and umami-enhancing effects. For example, peptides Arg-Asp, Asp-Gly-Val, Asp-Arg, Asp-Lys, Ser-Gly-Asp-Ala-Trp, and Asn-Asp-Asp-Gly-Trp possess umami flavor and can also enhance the umami flavor of monosodium glutamate (MSG) solutions. The peptide AEEHVEAVN can significantly enhance the umami intensity of chicken broth. Furthermore, molecular docking and molecular dynamics can be used to explore the mechanisms of umami enhancement. For instance, a novel hypothetical receptor T1R3-MSG complex was constructed, and the umami enhancement mechanism of the peptide was explored through molecular docking with the T1R3-MSG complex. The study showed that when activated by MSG, the T1R3 receptor was found to be in an open conformation. Umami peptides primarily interact with the open state of T1R3 to enhance the umami flavor of MSG.

[0005] Soy sauce is a traditional condiment made from soybeans and wheat through microbial fermentation, and it is loved for its strong and unique umami flavor. Umami peptides are an important component of the umami flavor of soy sauce. In recent years, several polypeptides, such as ALPEEV, LPEEV, AQALQAQAEKQQQ, ISWCFTY, and QISPYRRI, have been identified from soy sauce, exhibiting umami characteristics. However, there are still relatively few studies on the identification and characterization of soy sauce umami peptides. Therefore, further research and exploration of soy sauce umami peptides remains essential and is of great significance for advancing research on soy sauce flavor. Summary of the Invention

[0006] This invention protects a umami peptide with the amino acid sequence GAAGAAD. The bioactive peptide GAAGAAD provided by this invention is safe, non-toxic, and non-allergenic, and possesses excellent umami flavor and synergistic flavor-enhancing effects.

[0007] This invention provides a method for extracting umami peptides from soy sauce, the method comprising the following steps:

[0008] S1: Immerse the dried XAD-16 macroporous resin in a 95% ethanol solution for a period of time, wash the macroporous resin repeatedly with distilled water, and then remove the distilled water from the macroporous resin.

[0009] S2: Mix the washed macroporous resin with soy sauce and fully adsorb it on a constant temperature shaker. Then add distilled water, 20% ethanol, 40% ethanol and 60% ethanol in sequence for elution. After elution, collect the 60% ethanol eluent.

[0010] S3: The collected 60% ethanol eluent was evaporated and concentrated using a vacuum rotary evaporator, and then freeze-dried to obtain a soy sauce umami peptide sample, which was stored in a refrigerator at -20°C for later use.

[0011] Further, the dried XAD-16 macroporous resin described in step S1 is immersed in a 95% ethanol solution for 10 to 12 hours; the macroporous resin is then washed repeatedly with distilled water 5 to 10 times.

[0012] Further, the mass ratio of the dried macroporous resin to the volume of soy sauce in step S2 is (4g~6g):100 mL.

[0013] Furthermore, in step S2, the temperature at which the adsorption is fully performed on the constant temperature shaker is 30℃~40℃, the adsorption time is 10 h~12 h, and the speed of the shaker is 100 rpm~200 rpm.

[0014] Further, in step S2, the volume ratio of distilled water, 20% ethanol, 40% ethanol and 60% ethanol is 300 mL:300 mL:300 mL:300 mL.

[0015] Further, the evaporation and concentration temperature in step S3 is 50℃~55℃; the freeze-drying is as follows: first, freeze-dry in a refrigerator at -80℃ for 16 h~24 h, and then freeze-dry in a freeze dryer at a temperature of -50℃~-45℃ for 45 h~50 h.

[0016] The beneficial effects of this invention are as follows:

[0017] The umami peptide GAAGAAD provided by this invention is a novel umami peptide with characteristics such as non-toxicity, non-allergenicity, and good water solubility. Electronic tongue analysis results show that this peptide possesses umami characteristics, with an umami intensity of 3.03. Adding the peptide to MSG solution can enhance the umami intensity of the MSG solution.

[0018] The umami peptide GAAGAAD provided by this invention can undergo molecular docking with the umami receptor T1R1 / T1R3, and exhibits a lower binding energy with the umami receptor T1R3. The binding energies of the peptide with T1R1 and T1R3 are -7.6 kcal / mol and -8.2 kcal / mol, respectively. Furthermore, the umami peptide GAAGAAD has an even lower binding energy with the umami receptor T1R1 / T1R3-Glu, with binding energies of -8.2 kcal / mol with T1R1 and -8.6 kcal / mol with T1R3, respectively.

[0019] The umami peptides provided by this invention can be widely used in the preparation of seasonings or food additives, providing new umami peptide raw materials for the development of seasonings and food additives, and have important application value. Attached Figure Description

[0020] Figure 1 This is the secondary mass spectrum of the umami peptide GAAGAAD.

[0021] Figure 2 An electronic tongue scoring radar map (including umami, saltiness, bitterness, sourness, aftertaste A, aftertaste B, astringency, and richness) of umami peptide GAAGAAD and its synergistic umami-enhancing effect with MSG.

[0022] Figure 3 Homology modeling and reliability analysis of umami receptors T1R1 / T1R3. (A): Origin model of umami receptors T1R1 / T1R3; (B): Ramachandran diagram of umami receptors T1R1 / T1R3.

[0023] Figure 4This diagram shows the molecular docking of the umami peptide GAAGAAD with the umami receptors T1R1 / T1R3. (A): Interaction between GAAGAAD and T1R1; (B): Interaction between GAAGAAD and T1R3. The left column shows the 3D diagram of the docking between the peptide and T1R1 / T1R3; the right column shows the 2D diagram of the binding sites of the peptide's interaction with T1R1 / T1R3.

[0024] Figure 5 This diagram shows the molecular docking of the umami peptide GAAGAAD with the umami receptors T1R1 / T1R3-Glu. (A): Interaction between GAAGAAD and T1R1-Glu; (B): Interaction between GAAGAAD and T1R3-Glu. The left column shows the 3D diagram of the docking between the peptide and T1R1 / T1R3; the right column shows the 2D diagram of the binding sites of the peptide's interaction with T1R1 / T1R3.

[0025] Figure 6 Figures show the molecular dynamics simulation results for the T1R1-GAAGAAD, T1R3-GAAGAAD, T1R1-Glu-GAAGAAD, and T1R3-Glu-GAAGAAD complexes. (AD): Root Mean Square Deviation (RMSD) results; (EH): Root Mean Square Fluctuation (RMSF) results; (IL): Radius of Rotation (Rg) results. Detailed Implementation

[0026] Various exemplary embodiments of the present invention are now described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, characteristics, and embodiments of the present invention. Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods. Unless otherwise specified, the materials, reagents, etc., used in the following embodiments are commercially available.

[0027] To enable those skilled in the art to better understand the technical solution of the present invention, the technical solution of the present invention will be further described below with reference to the accompanying drawings and embodiments, and the beneficial effects of the present invention will be verified through the following embodiments.

[0028] Example 1: Isolation and Extraction of Soy Sauce Peptides

[0029] First, XAD-16 macroporous resin was activated with 95% ethanol and then dried in an oven at 60℃. 5 g of the dried XAD-16 macroporous resin was weighed and immersed in 95% ethanol for 12 h of activation, then washed with distilled water to remove the ethanol. Subsequently, 100 mL of soy sauce was thoroughly mixed with the resin for 10 h, and the mixture was kept at 30℃ with shaking for 8-10 h (150 r / min). Then, a gradient elution was performed using 300 mL of distilled water, 20% ethanol, 40% ethanol, and 60% ethanol, respectively, and the 60% ethanol eluent was collected. Finally, the eluent was concentrated using a vacuum rotary evaporator and then freeze-dried to obtain the soy sauce peptide sample. The sample was collected and stored at -20℃ for later use.

[0030] Example 2: Identification and Screening of Umami Peptides in Soy Sauce

[0031] 2.1 Identification of soy sauce peptide sequences

[0032] 1 mg of soy sauce peptide sample was weighed and dissolved in 100 µL of formic acid water. The solution was then desalted using a C18 centrifugal column, and the filtrate was collected and dried. Analysis was performed using ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS / MS). The chromatographic conditions were as follows: the sample was dissolved in mobile phase A (0.1% formic acid) and analyzed pre-column on a nanofluidic UPLC system. Mobile phase B (0.1% formic acid-acetonitrile aqueous solution (acetonitrile 80%)) was used at a flow rate of 600 nL / min. Gradient elution was set as follows: 6% ~ 9%, 5 min; 9% ~ 14%, 15 min; 14% ~ 30%, 30 min; 30% ~ 40%, 8 min; 40% ~ 95%, 2 min. In addition, the mass spectrometry conditions were as follows: after peptide separation, data-dependent acquisition mass spectrometry analysis was performed using an Orbitrap mass analyzer for 120 min; detection mode: positive ion; precursor ion scan range: 375–1500 m / z. Primary mass spectrometry resolution: 60,000; automatic gain control (AGC) target: 4 × 10⁻⁶. 5 Maximum injection time: 100 ms; Secondary mass spectrometry resolution: 15,000; Automatic gain: 5 × 10⁻⁶ 4 The maximum injection time for the second stage was 512 ms, and the HCD collision energy was 30%. Finally, the original MS file was analyzed and searched using Thermo ProteomeDiscoverer software in conjunction with the UniProt soybean protein database.

[0033] 2.2 Virtual screening of potential soy sauce umami peptides

[0034] Umami-SCM (https: / / camt.pythonanywhere.com / iUmami-SCM), UMPred-FRL (https: / / pmlabstack.pythonanywhere.com / UMPred-FRL), Umami_YYDS (http: / / tastepeptides-meta.com / Umami_YYDS), and Umami-MRNN (umami-mrnn.herokuapp.com) umami prediction tools were used to predict the umami flavor of the identified peptide sequences, and peptides with all predicted umami flavor were screened out. ToxinPred (https: / / webs.iiitd.edu.in / raghava / toxinpred / ) and AllerTOP (https: / / www.ddg-pharmfac.net / AllerTOP / ) were used to assess the toxicity and allergenicity of the peptides, respectively. Finally, potential umami peptides without toxicity or allergenicity were screened out.

[0035] A total of 159 peptides were identified using UPLC-MS / MS. Peptides with a length of no more than 10 amino acid residues may possess umami flavor. Therefore, online tools such as iUmami-SCM, UMPred-FRL, Umami_YYDS, and Umami-MRNN were used to predict the umami flavor of peptide sequences with an amino acid length of no more than 10, quickly screening for potential umami peptides whose predicted flavor was umami. ToxinPred and AllerTOP tools were used to further predict the toxicity and allergenicity of potential umami peptides. Finally, peptide GAAGAAD was screened as a non-toxic and non-allergenic potential umami peptide, and its mass spectrometry is shown below. Figure 1 As shown. Amino acids Asp (D) and Glu (E) play key roles in umami flavor, and many umami peptides contain one or more D and E residues. The peptide GAAGAAD contains D residues, with a frequency range of 14.29%.

[0036] Example 3: Flavor Characteristics Analysis of Umami Peptides

[0037] 3.1 Evaluation of the umami characteristics of umami peptides

[0038] The umami, salty, sour, and bitter tastes of the synthetic peptides were analyzed using an SA402B taste sensor system (INSENT, Tokyo, Japan). 7 mg of the synthetic peptide was dissolved in 70 mL of distilled water to prepare a solution with a concentration of 0.1 mg / mL. Before testing, the sensor and reference electrode were activated for 24 h. The taste acquisition program was set to 30 s, the aftertaste acquisition time to 30 s, the stabilization time to 30 s, and the washing time to 336 s. Each sample was measured four times, and the data from the last three measurements were analyzed.

[0039] 3.2 Evaluation of the flavor-enhancing effect of umami peptides.

[0040] A 0.1 mg / mL peptide solution was mixed with a 0.1 mg / mL MSG solution, and the flavor-enhancing effect of the peptide was evaluated using an SA402B electronic tongue. The MSG solution (0.1 mg / mL) served as a control group. Each sample was measured four times, and the data from the last three measurements were collected for analysis.

[0041] like Figure 2 As shown, the peptide GAAGAAD exhibits umami flavor with an intensity of 3.03. In addition, GAAGAAD also exhibits bitterness, sourness, astringency, and saltiness. The acidity and astringency of the peptide may be due to acidic amino acid residues such as D and E, while the bitterness may be due to hydrophobic amino acid residues such as Gly, Phe, and Pro. To further investigate whether the peptide has an umami-enhancing effect together with monosodium glutamate (MSG), the umami characteristics of a mixture of peptide solution (1 mg / mL) and MSG solution (1 mg / mL) were evaluated using an electronic tongue. Figure 2 As shown, the addition of peptide GAAGAAD enhanced the umami intensity of MSG solution, with an enhancement rate of 22.31%. Furthermore, peptide GAAGAAD significantly increased the saltiness of the MSG solution while reducing bitterness. These findings indicate that the umami peptide GAAGAAD can not only enhance the umami flavor of food but also reduce salt intake, thus benefiting human health.

[0042] Example 4: Interaction Analysis of Umami Peptides and Umami Receptors

[0043] 4.1 Homology modeling of umami receptors T1R1 / T1R3

[0044] Three-dimensional (3D) structures of the umami receptors T1R1 / T1R3 were constructed using homology modeling. The amino acid sequences of human taste receptors T1R1 (Q7RTX1) and T1R3 (Q7RTX0) were retrieved and downloaded from the UniProtKB database (https: / / www.uniprot.org / ). Using the crystal structure of the killifish taste receptors T1R2a-T1R3 (PDB 5X2M) as a template, homology models T1R1 / T1R3 were constructed using SwissModel (https: / / swissmodel.expasy.org / interactive). The reliability of the homology models was then evaluated using Ramachandran diagrams obtained from the SAVESv 6.0 online tool (https: / / saves.mbi.ucla.edu / ).

[0045] T1R1 / T1R3 are the main umami receptors with high affinity for umami compounds. However, the crystal structures of the T1R1 / T1R3 umami receptors have not been reported. In homology modeling, the similarity between the template sequence and the target sequence is required to exceed 30%. Alignment results from the NCBI BLAST server showed that the sequence similarity of T1R1 and T1R3 to 5X2M was 34.75% and 37.29%, respectively. Therefore, this study constructed a T1R1 / T1R3 homology model based on the target sequences (T1R1 and T1R3) and the template sequence (5X2M) using the Swiss-model online tool. Figure 3 As shown, T1R1 is on the left and T1R3 is on the right. Furthermore, the reliability of the T1R1 / T1R3 homology model was evaluated using the SAVESv6.0 tool. Ramachandran plot analysis showed that all amino acid residues in the T1R1 / T1R3 homology model were located within reasonable regions, with 90.4% in the optimal region, 8.5% in the additionally allowed region, and 1.1% in the maximum allowed region. The proportion of amino acids within reasonable regions exceeded the 90% critical value, demonstrating the rationality of the constructed T1R1 / T1R3 receptor, which can be used for subsequent molecular docking studies.

[0046] 4.2 Interaction analysis of umami peptides with umami receptors T1R1 / T1R3

[0047] The molecular structures of umami peptides were constructed and optimized using ChemDraw (version 20.0) and Chem3D (version 20.0) software. The energy of the peptides was minimized using the MMFF94s force field in Avogadro software (version 1.99) to obtain stable umami peptide structures. The structure of the umami receptors T1R1 / T1R3 was optimized using Auto Dock software (version 1.5.6) by adding nonpolar hydrogen and removing water of crystallization; the peptide structure was further optimized by adding nonpolar hydrogen. Molecular docking of the peptides with T1R1 / T1R3 was performed using Auto DockVina software, and the optimal binding conformation was selected based on the docking energy. The pocket size and principal center coordinates were set as follows: T1R1: Center X = 40.845, Y = 30.808, Z = 43.536; Dimensions X = 40, Y = 40, Z = 40; T1R3: Center X = 49.528, Y = 37.528, Z = 7.194; Dimensions X = 40, Y = 40, Z = 40. For the molecular docking results, Pymol was used for visualization and interaction analysis.

[0048] In this study, to investigate the interaction between umami peptides and T1R1 / T1R3, the peptide GAAGAAD was molecularly docked with the T1R1 / T1R3 receptor. It is generally believed that peptides are more likely to produce umami flavor when the binding energy of molecular docking is in the range of -5 to -15 kcal / mol. The results showed that the binding energies of the peptide GAAGAAD with T1R1 and T1R3 were -7.60 and -8.20 kcal / mol, respectively (Table 1). The docking binding energy of the peptide with T1R3 was lower than that with T1R1, indicating that T1R3 is more likely to form a stable complex with the peptide. This may be because T1R3 exists in an open state, making it easier for the umami peptide to bind. Figure 4 This study investigated the binding sites of T1R1 / T1R3 and the interactions between umami peptides and T1R1 / T1R3. The results showed that umami peptides can enter the binding pocket of the VFT domain located in the T1R1 / T1R3 cavity. Interactions were primarily hydrogen bonds and electrostatic interactions, with hydrogen bonds accounting for a larger proportion. Specifically, the peptides GAAGAAD formed 11 hydrogen bonds with T1R1 and 10 hydrogen bonds with T1R3. Hydrogen bonds are likely a key force in forming a stable peptide-T1R1 / T1R3 complex. Further analysis of the sites on the receptor that generate hydrogen bonds was conducted. By docking the peptides, 8 amino acid residues were found on both T1R1 and T1R3, with hydrophilic amino acid sites accounting for 100% and 76%, respectively. This indicates that hydrophilic amino acid residues at the active sites of T1R1 / T1R3 play a crucial role in recognizing umami peptides.

[0049] Table 1. Binding energy and binding sites of T1R1 / T1R3 with the umami peptide GAAGAAD molecule.

[0050]

[0051] 4.3 Interaction analysis between umami peptides and umami receptors T1R1 / T1R3-Glu

[0052] This study explored the molecular mechanism by which peptides enhance umami flavor through molecular docking with the T1R1 / T1R3-Glu receptor. Glu was molecularly docked with the T1R1 / T1R3 receptor using Auto Dock software. The optimal binding conformation was screened based on binding energy, and the T1R1 / T1R3-Glu receptor complex was constructed. Subsequently, molecular docking of the T1R1 / T1R3-Glu receptor complex with the peptide was performed using Auto Dock Vina software.

[0053] In this study, Glu was first docked with T1R1 / T1R3 to obtain the T1R1 / T1R3-Glu complex. The T1R1 / T1R3-Glu complex was then used as the acceptor for peptide docking. The binding energies of the peptide GAAGAAD to T1R1 / T1R3-Glu were -8.0 and -8.6 kcal / mol, respectively (Table 2). Furthermore, it was found that the binding energy of the peptide to T1R1 / T1R3-Glu was lower than that to T1R1 / T1R3, indicating that the peptide-T1R1 / T1R3-Glu complex is more stable. This may be one of the reasons why peptides enhance umami flavor. The 3D and 2D interactions of the peptide with T1R1 / T1R3-Glu are shown in Figure 2. Figure 5 As shown, similar to T1R1 / T1R3, this peptide can enter the VFT domain and form hydrogen bonds, hydrophobic interactions, and electronic interactions with key amino acid residues at the active site. Among these, hydrogen bonds account for a relatively high proportion; peptide GAAGAAD forms 13 hydrogen bonds with T1R1-Glu and 18 hydrogen bonds with T1R3-Glu. The interaction strength and the number of hydrogen bonds between T1R1 / T1R3-Glu and peptide GAAGAAD are both higher than those between T1R1 / T1R3. This may be the reason why peptide GAAGAAD has an umami-enhancing effect.

[0054] Table 2 Binding energy and binding sites of T1R1 / T1R3-Glu with GAAGAAD molecules

[0055]

[0056] Example 5: Molecular dynamics simulation of umami peptides and umami receptor complexes.

[0057] Molecular dynamics simulations of protein-ligand complexes obtained through molecular docking were performed using GROMACS 2020 software. Proteins and ligands were separated in the docked complexes using Pymol software and exported as separate PDB files. The topology of T1R1 / T1R3 was generated using the Amber99SB force field, while the ligand topology was generated using the GAFF force field. Water was added to the protein-ligand system using the TIP3P water model, and sodium and chloride ions were added to make the system electrically neutral. The protein-ligand system was minimized over 50,000 steps using the conjugate gradient algorithm. The protein-ligand system was then further equilibrated under NVT isothermal conditions above 100 ps and NPT isobaric conditions, and molecular dynamics simulations were performed at ambient temperature and pressure for 100 ns. The root mean square deviation (RMSD), root mean square fluctuation (RMSF), and radius of gyration (Rg) were used to evaluate the dynamic simulation results of the protein-ligand complexes.

[0058] Molecular dynamics simulations (RMSD) of the T1R1 / T1R3-GAAGAAD and T1R1 / T1R3-Glu-GAAGAAD complexes were performed over 100 ns to explore their umami mechanism and umami-enhancing effect. RMSD is commonly used to assess the fluctuation range of the macromolecular backbone during ligand-receptor binding. The smoother the RMSD curve, the more stable the formation of the receptor-ligand complex. In this study, we found that for the T1R1-GAAGAAD, T1R3-GAAGAAD, T1R1-Glu-GAAGAAD, and T1R3-Glu-GAAGAAD complexes, the RMSD fluctuated significantly in the first 40 ns, but then reached equilibrium and remained stable around 0.45, 0.52, 0.47, and 0.6 nm, respectively. Figure 6 AD). Within a reasonable fluctuation range, the RMSD fluctuation values ​​of all complexes were less than 1 nm, indicating that the structure of the complexes was in equilibrium during the simulation. The RMSF curves represent the degree of fluctuation of the receptor amino acid residues during dynamic simulation, reflecting the flexibility and motion intensity of amino acid residues in the receptor protein. The RMSF curves of the T1R1-GAAGAAD, T1R3-GAAGAAD, T1R1-Glu-GAAGAAD, and T1R3-Glu-GAAGAAD complexes all fluctuated within the range of 1 nm. Figure 6EH). The RMSF values ​​of most amino acids were within 0.4 nm. These results indicate that the receptor protein was relatively stable during the simulation. Furthermore, Rg is often used to describe the closeness of complex interactions. Similar to RMSD, the Rg value tended to stabilize after 40 ns of simulation (…). Figure 6 The presence of IL indicates that the umami peptide can form tight complexes with the T1R1 / T1R3 and T1R1 / T1R3-Glu receptors. In summary, GAAGAAD has minimal impact on the overall structural stability of T1R1 / T1R3 and T1R1 / T1R3-Glu, demonstrating the structural stability of the receptors and peptides and the validity of the molecular docking results.

Claims

1. A umami peptide derived from soy sauce, characterized in that, The umami peptide is GAAGAAD, and the amino acid sequence of the peptide GAAGAAD is Gly-Ala-Ala-Gly-Ala-Ala-Asp.

2. The method for preparing umami peptides derived from soy sauce according to claim 1, comprising the following steps: S1: Immerse the dried XAD-16 macroporous resin in a 95% ethanol solution for a period of time, wash the macroporous resin repeatedly with distilled water, and then remove the distilled water from the macroporous resin. S2: Mix the washed macroporous resin with soy sauce and fully adsorb it on a constant temperature shaker. Then add distilled water, 20% ethanol, 40% ethanol and 60% ethanol in sequence for elution. After elution, collect the 60% ethanol eluent. S3: The collected 60% ethanol eluent was evaporated and concentrated using a vacuum rotary evaporator, and then freeze-dried to obtain a soy sauce peptide sample, which was stored in a refrigerator at -20°C for later use.

3. The method for extracting umami peptides from soy sauce according to claim 2, characterized in that, The dried XAD-16 macroporous resin described in step S1 is immersed in a 95% ethanol solution for 10 to 12 hours; the macroporous resin is then washed repeatedly with distilled water 5 to 10 times.

4. The method for extracting umami peptides from soy sauce according to claim 2, characterized in that, The mass ratio of the dried macroporous resin to the volume of soy sauce in step S2 is (4 g ~ 6 g): 100 mL.

5. The method for extracting umami peptides from soy sauce according to claim 2, characterized in that, In step S2, the temperature at which the food is fully adsorbed on the constant temperature shaker is 30℃~40℃, the adsorption time is 10 h~12 h, and the speed of the shaker is 100 rpm~200 rpm.

6. The method for extracting umami peptides from soy sauce according to claim 2, characterized in that, In step S2, the volume ratio of distilled water, 20% ethanol, 40% ethanol, and 60% ethanol is 300 mL:300 mL:300 mL:300 mL.

7. The method for extracting umami peptides from soy sauce according to claim 2, characterized in that, The evaporation and concentration temperature in step S3 is 50℃~55℃; the freeze drying is as follows: first, freeze-dry in a refrigerator at -80℃ for 16 h~24 h, and then freeze-dry in a freeze dryer at a temperature of -50℃~-45℃ for 45 h~50 h.

8. The umami peptide according to claim 1, characterized in that, Umami peptides and monosodium glutamate (MSG) have a synergistic flavor-enhancing effect.

9. The application of the umami peptide according to claim 1 in enhancing the umami flavor of food and preparing seasonings.

10. The application according to claim 9, characterized in that, The umami component of the seasoning includes the umami peptides described in claim 1.

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