An active oligopeptide for promoting GLP-1 secretion in the intestine, and a preparation method and application thereof

Abstract

The present application relates to a kind of active oligopeptide for promoting intestinal GLP-1 secretion and its preparation method and application.Active oligopeptide for promoting intestinal GLP-1 secretion is LPVDVLA active oligopeptide, and amino acid sequence is: Leu-Pro-Val-Asp-Val-Leu-Ala.The active oligopeptide of the present application can significantly and long time stimulate intestinal endocrine cell to secrete GLP-1.Compared with prior art, the active oligopeptide of the present application can be artificially synthesized using chemical solid-phase synthesis method, and can also be obtained by enzymatic oat protein separation and purification;Meanwhile, the active oligopeptide has the advantages of safe and non-toxic side effects, simple preparation process, can be realized by conventional enzymolysis and chromatographic purification, easy scale production and the like.The oligopeptide described in the present application can be used for preparing T2DM functional food or drug research and development, to provide new scheme for diabetes intervention, and has wide application prospect.

Description

Technical Field

[0001] This invention relates to the field of bioactive peptide technology in the field of biotechnology, and in particular to an active oligopeptide that promotes the secretion of GLP-1 in the intestine, its preparation method and application. Background Technology

[0002] Diabetes has become a global public health problem. Many chronic metabolic diseases, including cardiovascular disease and non-alcoholic fatty liver disease, are associated with diabetes, with type 2 diabetes (T2DM) accounting for over 90%. Insulin resistance caused by damage to pancreatic β cells is the core cause of T2DM. Therefore, restoring the body's insulin secretion function is a key approach to prevent, slow, or treat T2DM. Glucagon-like peptide-1 (GLP-1), an incretin hormone secreted by intestinal L cells, can promote insulin secretion in a glucose-dependent manner, while also having physiological functions such as delaying gastric emptying and regulating appetite, playing a central role in glycemic regulation in T2DM. Existing research has shown that GLP-1 secretion is regulated by dietary factors, and food-derived peptides are one of the main dietary factors regulating GLP-1 secretion. However, endogenous GLP-1 has significant drawbacks: its half-life in vivo is only 2-3 minutes, and its degradation product GLP-1(9-36) can also act as a GLP-1 receptor antagonist, further hindering the function of active GLP-1. Furthermore, patients with type 2 diabetes mellitus (T2DM) generally exhibit a weakened incretin effect, with postprandial GLP-1 secretion reduced by 40-50% compared to healthy individuals, exacerbating insulin insufficiency and glycemic dysregulation. Therefore, promoting intestinal GLP-1 secretion through diet has become a potentially important strategy for improving or alleviating T2DM and related chronic diseases.

[0003] Food-derived bioactive peptides are a common type of functional dietary factor. They are easily digested and absorbed by the human body, have high safety for consumption, low toxicity and side effects, and have clear sources. They have broad application prospects in the fields of functional foods and metabolic disease intervention.

[0004] Patent CN115353551B discloses the application of an oat-derived oligopeptide or an enzymatic hydrolysate containing an oat-derived oligopeptide in the preparation of products that promote the secretion of GLP-1 in the intestine. This oat-derived oligopeptide is DVNNNANQLEPR, with the amino acid sequence: Asp-Val-Asn-Asn-Ala-Asn-Gln-Leu-Glu-Pro-Arg. This patent discloses a polypeptide derived from oat protein that promotes the secretion of GLP-1 in the intestine; however, the polypeptide fragment in this patent is relatively long, and long polypeptides are generally not conducive to absorption.

[0005] Patent CN121045334B discloses a method for preparing oat bioactive peptides that promote GLP-1 secretion and their applications. The oat bioactive peptides contain any one or more of the peptides NDQRGEII, KQGDVIALPA, PFVQQQQ, and PSEQYQPYPEQQEPFVQ. This patent discloses different peptides derived from oat protein that promote intestinal GLP-1 secretion. However, the disclosed data shows that the GLP-1 level in the intestinal cell supernatant is only about 7-8 pmol / mL, with limited improvement compared to the blank control group. Therefore, further screening from oat protein for bioactive peptides that can target and efficiently promote intestinal GLP-1 secretion, especially short-chain peptides, is of significant value. This is important for preventing and improving type 2 diabetes mellitus (T2DM) and related chronic metabolic diseases, and for reducing the hepatic metabolic burden caused by traditional hypoglycemic drugs. Summary of the Invention

[0006] Given the current lack of short-chain bioactive peptides that can efficiently promote the secretion of GLP-1 in the intestine in the existing technology, this invention provides an active oligopeptide that promotes the secretion of GLP-1 in the intestine, its preparation method, and its application.

[0007] The objective of this invention can be achieved through the following technical solutions: In a first aspect, the present invention provides an active oligopeptide that promotes the secretion of GLP-1 in the intestine, the active oligopeptide being LPVDVLA with the amino acid sequence: Leu-Pro-Val-Asp-Val-Leu-Ala.

[0008] This application research found that the LPVDVLA active oligopeptide exists in oat protein, derived from oat 12S seedstorage globulin 1 (Uniprot protein accession number: P12615). Therefore, the LPVDVLA active oligopeptide provided in this application can be prepared from oat protein.

[0009] In a second aspect, the present invention provides a polynucleotide encoding the said active oligopeptide.

[0010] In a third aspect, the present invention provides a method for preparing the active oligopeptide, which is obtained by enzymatic hydrolysis of oat protein followed by separation and purification, or by direct artificial synthesis through genetic engineering, or by direct chemical synthesis.

[0011] The artificial synthesis of the active oligopeptides through genetic engineering is a technical solution that can be achieved by those skilled in the art. For example, it can be based on DNA recombination technology, using a suitable DNA template to control the sequence synthesis of the oligopeptides.

[0012] One possible method for directly obtaining oat protein from oat protein through separation and purification is to use conventional biological techniques such as enzymatic hydrolysis, separation, and purification based on the given amino acid sequence of the active oligopeptide to obtain the active oligopeptide from oats.

[0013] The above-mentioned active oligopeptides were prepared by chemical synthesis using the traditional solid-phase synthesis method.

[0014] In a fourth aspect, the present invention provides a method for preparing an enzymatic hydrolysis product containing the active oligopeptide, wherein oat protein is enzymatically hydrolyzed, and the method includes the following steps: enzymatically hydrolyzing oat protein with a flavor protease to obtain an oat protein hydrolysis product containing the active oligopeptide, which is the enzymatic hydrolysis product containing the active oligopeptide.

[0015] In one embodiment of the present invention, the method for enzymatically hydrolyzing oat protein with flavored protease includes the following steps: 1) Extracting oat protein from oats; 2) The oat protein is hydrolyzed with the flavor protease to obtain the oat protease hydrolysate of the active oligopeptide.

[0016] In a more specific embodiment of the present invention, the method for extracting oat protein from oats includes the following steps: Oat flour was ground and defatted, then added to distilled water. The temperature was adjusted to 45 ℃ and the pH to 4.5. The mixture was pretreated with cellulase and saccharifying enzyme for 0.5-2 h. After cooling to room temperature, the pH was adjusted to 11 for protein extraction. After extraction, the supernatant was collected by centrifugation. The pH of the supernatant was adjusted to 4.5, and the mixture was allowed to stand before centrifugation and washing with water. Finally, the oat protein was obtained by drying.

[0017] In one embodiment of the present invention, the mass ratio of the flavor protease to the oat protein is 1:20100.

[0018] In one embodiment of the present invention, the flavor protease is selected as a complex flavor protease extracted by Aspergillus oryzae fermentation. The flavor protease (complex flavor protease) is a complex protease preparation extracted by Aspergillus oryzae fermentation, and its core characteristic is that it simultaneously contains endopeptidase and exopeptidase (aminopeptidase, carboxypeptidase).

[0019] In one embodiment of the present invention, the enzymatic hydrolysis conditions of the flavor protease are 50°C for 1-4 h.

[0020] In one embodiment of the present invention, after obtaining the oat protein hydrolysate in step 2), the following steps are further included: The oat protein hydrolysate was separated using a YMC ODS C18 column. Deionized water, 10% ethanol aqueous solution, 30% ethanol aqueous solution, 50% ethanol aqueous solution, 70% ethanol aqueous solution and anhydrous ethanol were used as eluents in sequence at a certain flow rate for elution and chromatographic purification. Different components were collected. Then, the effect of different components on the secretion of GLP-1 by enteroendocrine cells was detected. The component that stimulated the highest activity of GLP-1 secretion was selected as the target component, which is the enzymatic hydrolysate containing the active oligopeptide.

[0021] In a fifth aspect, the present invention provides an enzymatic hydrolysis product containing the active oligopeptide prepared by the above-described preparation method.

[0022] In a sixth aspect, the present invention provides the use of the said active oligopeptide and the enzymatic hydrolysis product containing the said active oligopeptide in the preparation of a product having at least one of the following functions: 1) and 4). 1) Promotes GLP-1 secretion; 2) Reduce the degree of damage to the liver and pancreas; 3) Improves glucose and lipid metabolism disorders; 4) Helps lower blood sugar.

[0023] The active oligopeptide and the enzymatic hydrolysate containing the active oligopeptide have the function of promoting GLP-1 secretion, which can reduce the degree of damage to the liver and pancreas, improve glucose and lipid metabolism disorders, and help lower blood sugar.

[0024] In a seventh aspect, the present invention provides a product comprising the active oligopeptide, or an enzymatic hydrolysis product containing the active oligopeptide, the product having at least one of the following functions: 1) and 4). 1) Promotes GLP-1 secretion; 2) Reduce the degree of damage to the liver and pancreas; 3) Improves glucose and lipid metabolism disorders; 4) Helps lower blood sugar.

[0025] The products include food and pharmaceuticals, and the food includes functional foods or health foods.

[0026] Compared with the prior art, the present invention has the following beneficial effects: Addressing the challenges in treating type 2 diabetes mellitus (T2DM) such as the short duration of endogenous GLP-1 in vivo, gastrointestinal adverse reactions of artificial GLP-1 inhibitors, and the increased liver burden of traditional hypoglycemic drugs, this invention screened and obtained an oat-derived GLP-1 secretion-promoting active oligopeptide as a novel peptide sequence, which has not been reported before. A single oral administration can stimulate the body to produce GLP-1 for a prolonged period of up to 2 hours; it can also significantly and for a long time stimulate intestinal endocrine cells to secrete GLP-1.

[0027] The active oligopeptides provided in this application are short-chain peptides, which are easily absorbed. Furthermore, they are derived from food-grade oat protein, making them safe and free of toxic side effects.

[0028] Furthermore, the active oligopeptides provided in this application can be artificially synthesized using chemical solid-phase synthesis methods, or obtained by enzymatic hydrolysis of oat protein followed by separation and purification. The preparation process is simple, can be achieved through conventional enzymatic hydrolysis and chromatographic purification, and is easy to scale up for production.

[0029] The active oligopeptides provided in this application can reduce the degree of damage to the liver and pancreas, improve glucose and lipid metabolism disorders, and help lower blood sugar.

[0030] The oligopeptides described in this invention can be used in the research and development of functional foods or drugs for type 2 diabetes mellitus (T2DM), providing new solutions for the high-value utilization of oat resources and diabetes intervention, with broad application prospects. Attached Figure Description

[0031] Figure 1 The effect of LPVDVLA active oligopeptide on STC-1 cell viability was investigated, with C representing the control group (without LPVDVLA active oligopeptide).

[0032] Figure 2 The effect of LPVDVLA active oligopeptide on GLP-1 secretion in STC-1 cells was investigated, with C representing the control group (without LPVDVLA active oligopeptide).

[0033] Figure 3 The effect of oat protein hydrolysate on GLP-1 secretion in STC1 cells was investigated, with C representing the control group (without oat protein hydrolysate).

[0034] Figure 4 The effect of oat protein hydrolysate on GLP-1 secretion by mouse intestinal endocrine cells.

[0035] Figure 5 This is a chromatogram of oat protein hydrolysate separated by a YMC ODS C18 column.

[0036] Figure 6 The effects of oat protein hydrolysate and isolated components on GLP-1 secretion in STC-1 cells.

[0037] Figure 7 MS / MS map and sequence analysis of LPVDVLA active oligopeptide in isolated component F1. Detailed Implementation

[0038] The present invention will now be described in detail with reference to the accompanying drawings and specific embodiments. These embodiments are based on the technical solution of the present invention and provide detailed implementation methods and specific operating procedures. However, the scope of protection of the present invention is not limited to the following embodiments.

[0039] Example 1 Artificial synthesis of LPVDVLA active oligopeptides and evaluation of their GLP-1 secretion-promoting activity I. Synthesis of LPVDVLA Active Oligopeptides The active oligopeptide Leu-Pro-Val-Asp-Val-Leu-Ala (LPVDVLA) was synthesized by Hefei Hesheng Biotechnology Co., Ltd. using a peptide solid-phase synthesis method. The purity of the synthesized peptide was verified to be greater than 95% by high performance liquid chromatography and mass spectrometry.

[0040] II. Effects of LPVDVLA on STC1 cell viability and GLP-1 secretion (1) Culture of STC-1 cells STC1 cells were cultured in DMEM medium containing 10% fetal bovine serum (FBS), 1% non-essential amino acids (NEAA), 100 U / mL penicillin, and 0.1 mg / mL streptomycin. Cells were incubated at 37°C in a cell culture incubator containing 5% CO2 and passaged by trypsin digestion when they reached 80-90% confluence.

[0041] (2) Cell viability assay Cell viability was determined using the CCK-8 colorimetric method. Cells were inoculated at 5 × 10⁶ cells / mL. 3 Cells / well were seeded at a density of 96-well plates and cultured in a 5% CO2 incubator for 24 h, after which the culture medium was changed. The experimental group received LPVDVLA active oligopeptide solution at a final concentration of 5 mM, while the control group received an equal volume of D-Hank's solution. Each group had 6 parallel wells. After culturing for another 2 h, 10 μL of CCK-8 reagent was added to each well, and the absorbance was measured at 450 nm using a microplate reader after 2 h of reaction. Cell viability was calculated using the following formula: (Experimental group OD / Control group OD × 100%). Results are as follows: Figure 1 As shown, the activity of the oligopeptide did not significantly change the viability of STC1 cells compared to the control group (C) at different test concentrations, indicating that the LPVDVLA active oligopeptide is non-toxic to cells.

[0042] (3) Determination of hormone secretion levels in STC1 cells STC1 cells were cultured in 24-well plates at a rate of 1.25 × 10⁶ cells / well. 5Cells were seeded at a density of [number] cells. When the cells reached 80%-90% confluence, they were washed twice with Hank's buffer to remove the culture medium. Peptide solutions were added to the STC1 cells to final molar concentrations of 0, 0.2, 2, and 5 mM, respectively. Cells were incubated at 37°C for 2 hours. After incubation, the cells were centrifuged at 1000 g for 20 minutes, and the supernatant was collected. GLP-1 levels were determined using a GLP-1 detection kit from the Japan Immunobiology Institute.

[0043] The total protein content of the supernatant determined using the Beyotime Biotechnology BCA kit was used as a reference to reduce errors caused by operations between different batches. The calculation formula is as follows: C GLP-1 / (C BCA ×100)(×10 7 ), where C GLP-1 The concentration of GLP-1 in the supernatant is expressed as pg / mL, C BCA This represents the total protein concentration measured in the supernatant minus the concentration of the added peptide LPVDVLA (mg / mL).

[0044] The effect of LPVDVLA active oligopeptide on GLP-1 secretion in STC1 cells is as follows: Figure 2 , Figure 2 The LPVDVLA concentration in this solution is 5 mM. Figure 2 In the control group (C), no LPVDVLA was added (using the same amount of Hank's buffer). The results show that the LPVDVLA active oligopeptide significantly increased GLP-1 secretion. Numerous studies have confirmed that GLP-1 is an important member of the brain-gut peptide group. After binding to a specific receptor, it promotes insulin secretion and inhibits glucagon secretion in a glucose-dependent manner, and can also improve pancreatic β-cell function and maintain metabolic homeostasis. Increasing intestinal GLP-1 secretion can improve insulin sensitivity and simultaneously inhibit glucagon secretion. The LPVDVLA active oligopeptide described in this application can significantly promote GLP-1 secretion by intestinal endocrine cells STC1. Therefore, these peptides play an important role in reducing liver and pancreatic damage and improving glucose and lipid metabolism disorders.

[0045] Example 2 Preparation and Activity Evaluation of Oat Protein Hydrolysate (1) Preparation of oat protein hydrolysate Oat flour was milled and passed through an 80-sieve, then defatted with hexane. The defatted oat flour was soaked in distilled water at a mass ratio of 1:12 (oat flour:water) in a beaker. The pH was adjusted to 4.5 with 1 mol / L HCl, and the mixture was treated with saccharifying enzyme and cellulase (1% of the oat flour mass fraction) at 45°C for 0.5–2 h. The pH was then adjusted to 11.0 with 1 mol / L NaOH, and the mixture was stirred with a magnetic stirrer for 2 h before centrifugation to collect the supernatant. The pH of the supernatant was adjusted to its isoelectric point (pH 4.5) with 1 mol / L HCl, allowed to stand for 1 h, centrifuged, and the precipitate was washed with water until neutral. The precipitate was reconstituted with a small amount of distilled water and freeze-dried to obtain oat protein, which was stored at 4°C for later use.

[0046] 1 g of freeze-dried protein powder (i.e., the oat protein obtained above) was dissolved in 20 mL of an aqueous solution containing 40 mg of freshly prepared flavor protease (a complex flavor protease extracted by Aspergillus oryzae fermentation, commercially known as Novozymes Flavorzyme). The pH of the solution was adjusted to 7.0, and the mixture was incubated at 50 °C for 3 h, yielding a hydrolysate of 12%. NaOH solution (0.5 mol / L) was added dropwise at intervals to adjust the pH to the optimal pH of 7.5 for the relevant flavor protease. After incubation, the supernatant was collected by centrifugation and freeze-dried to obtain the oat protein hydrolysate (named Fla12).

[0047] (2) Activity evaluation The oat protein hydrolysate was prepared into a 5 mg / mL solution using Hank's buffer. The effect of the hydrolysate on GLP-1 secretion by STC1 cells was determined using the method described above. The results are shown below. Figure 3 The results show that oat protein hydrolysate can significantly stimulate STC1 cells to secrete GLP-1.

[0048] Furthermore, the effects of oat protein hydrolysate on hormone secretion by intestinal endocrine cells in mice were evaluated at the animal level. After a one-week acclimatization period, ICR mice were randomly divided into two groups (n=20 per group). The control group received saline via gavage; the oat protein hydrolysate group received oat protein hydrolysate (1.0 g / kg body weight) via gavage. Following gavage, blood was collected by enucleation at 0, 30, 60, 90, and 120 minutes. The blood samples were placed in centrifuge tubes containing EDTA (final concentration 1.8 mg / mL blood) and a protease inhibitor (final concentration 1 μL / mL blood). After centrifugation, the supernatant was collected, and serum GLP-1 levels were measured using ELISA. Figure 4As shown, the serum GLP-1 level in the saline group remained at around 8 pmol / mL during this period; while oral administration of oat protein hydrolysate significantly increased the GLP-1 level in mice, reaching approximately 25 pmol / mL in the blood at 60 min. Even at 2 hours, the GLP-1 level in the mouse blood was still significantly higher than that in the control group, indicating that a single oral administration of oat protein hydrolysate can stimulate the body to produce GLP-1 for a prolonged period.

[0049] Example 3: Preparation of active oligopeptides from oat protein that promote GLP-1 secretion Flavor enzyme hydrolysate (Fla12) was separated and purified using a YMC ODS C18 column (1.6 × 25 cm). The C18 packing material was swollen with ethanol to form a homogenate, and then slowly packed using a wet packing technique to ensure a uniform column bed free of bubbles and cracks. During separation, the mobile phase level was always 2–3 cm above the top of the column packing. Ultrapure water was used as the initial mobile phase, and the sample was loaded after equilibration to a baseline of 4 column volumes. The oat protein hydrolysate sample (100 mg / mL) was filtered through a 0.45 μm microporous membrane and centrifuged at 4 °C and 10,000 g for 5 min to completely remove particulate matter. Sample separation was performed using a C18 reversed-phase chromatography column at a constant flow rate of 2 mL / min. The gradient elution program was as follows: 0% ethanol solution was used initially for 0-60 minutes; 10% ethanol solution was used for 61-100 minutes; 30% ethanol solution was used for 101-160 minutes; 50% ethanol solution was used for 161-220 minutes; 70% ethanol solution was used for 221-280 minutes; and finally, 100% ethanol solution was used for elution from 281-340 minutes. The absorbance signal at 220 nm was monitored in real time using an HD-A chromatography system during the elution process. The separation chromatogram of the oat protein hydrolysate is shown below. Figure 5 As can be seen, the C18 column separated the protease hydrolysate into four peptide fractions. The collected eluent was rapidly transferred to -80°C for pre-freezing, and then freeze-dried to remove the solvent, yielding a lyophilized powder. The cytotoxicity and secretagogue activity of each fraction (5 mg / mL) were evaluated using the STC-1 cell model system. Figure 6 As shown, among the four peptide fractions, fraction F1 exhibits the best ability to stimulate STC1 cells to secrete GLP-1. Therefore, fraction F1 is a highly active oligopeptide that promotes GLP-1 secretion.

[0050] Example 4 Identification of LPVDVLA bioactive oligopeptides in oat protein The peptide sequences in fraction F1 were identified using mass spectrometry. The sample was dissolved in distilled water to prepare a 1 mg / mL sample. Separation was performed using a reversed-phase column (150 μm id. × 150 mm, packed with Acclaim PepMap RPLC C18, 1.9 μm). Mobile phase A was 0.1% formic acid aqueous solution, and mobile phase B was 0.1% formic acid / 80% acetonitrile solution. Gradient elution was performed at a flow rate of 600 nL / min. The separation gradients were: 0.2 min, 48% B; 2.45 min, 8.40% B; 4.555 min, 4.060% B; 5.556 min, 6.095% B; 5.666 min, 95% B. The ion source for mass spectrometry was electrospray ionization (ESI), positive ion scan mode, ionization voltage 2200 V, and capillary temperature 270 °C. Level 1 mass spectrometry parameter settings: scan range 100-2000 m / z, maximum resolution 70000, automatic gain parameter 3000000. Level 2 mass spectrometry parameter settings: scan range 50-2000 m / z, maximum resolution 17500, automatic gain parameter 100000.

[0051] Mass spectrometry analysis revealed that the main ion peak in fraction F1 was at m / z = 362.7, z = 2. Further secondary mass spectrometry analysis of the molecular ion peak was performed; the secondary mass spectrum of this molecular ion peak is shown below. Figure 7 Through database matching and manual analysis, the peptide corresponding to the ion peak is Leu-Pro-Val-Asp-Val-Leu-Ala (LPVDVLA).

[0052] Oat protein is mainly a globulin, and the amino acid sequence of oat 12S seed storage globulin 1 is shown in SEQ ID NO.1. It can be found that the LPVDVLA active oligopeptide is derived from oat 12S seed storage globulin 1 (Uniprot protein accession number: P12615), and the LPVDVLA active oligopeptide can be prepared from oat protein.

[0053] In this invention, the LPVDVLA active oligopeptide is located at position 462468 of the amino acid sequence shown in SEQ ID NO.1.

[0054] The amino acid sequence shown in SEQ ID NO.1 is as follows: MHERAVWIFL CDLGGQNMPF EGSIPVEIGN LVNLFSLGME 40 GLRGCKFDRL QAFEPLRQVR SQAGITEYFD EQNEQFRCAG 80 VSVIRRVIE PQGLLLPQYH NAPGLVYILQ GRGFTGLTFPG 120 CPATFQQQFQ QFDQARFAQG QSKSQNLKDE HQRVHHIKQG 160 DVVALPAGIV HWCYNDGDAP IVAVYVFDVN NNANQLEPRQ 200 KEFLLAGNNK REQQFGQNIF SGFSVQLLSE ALGISQQAAQ 240 KIQSQNDQRG EIIRVSQGLQ FLKPFVSQQG PVEHQAYQPI 280 QSQQEQSTQY QVGQSPQYQE GQSTQYQSGQ SWDQSFNGLE 320 ENFCSLEARQNIENPKRADT YNPRAGRITH LNSKNFPTLN 360 LVQMSATRVN LYQNAILSPY WNINAHSVMH MIQGRARVQV 400 VNNHGQTVFND ILRRGQLLII PQHYVVLKKA EREGCQYIS 440 FKTTPNSMVS YIAGKTSILR ALPVDVLANA YRISRQESQN 480 LKNNRGEEFG AFTPKFAQTG SQSYQDEGES SSTEKASE 518 The above description of the embodiments is provided to enable those skilled in the art to understand and use the invention. It will be apparent to those skilled in the art that various modifications can be made to these embodiments, and the general principles described herein can be applied to other embodiments without inventive effort. Therefore, the present invention is not limited to the above embodiments, and any improvements and modifications made by those skilled in the art based on the disclosure of the present invention without departing from the scope of the invention should be within the protection scope of the present invention.

Claims

1. An active oligopeptide that promotes the secretion of GLP-1 in the intestine, characterized in that, The active oligopeptide is LPVDVLA, with the amino acid sequence Leu-Pro-Val-Asp-Val-Leu-Ala.

2. A polynucleotide encoding the active oligopeptide of claim 1.

3. The method for preparing the active oligopeptide according to claim 1, characterized in that, It can be obtained directly through chemical synthesis, artificially synthesized through genetic engineering, or obtained from oat protein through enzymatic hydrolysis and purification.

4. A method for preparing an enzymatic hydrolysis product containing the active oligopeptide of claim 1, characterized in that, Includes the following steps: Oat protein is hydrolyzed with a flavor protease to obtain an oat protein hydrolysate, which is an enzymatic hydrolysate containing the active oligopeptide described in claim 1.

5. The preparation method according to claim 4, characterized in that, The method for extracting oat protein from oats includes the following steps: Oat flour was ground and defatted, then added to distilled water. The temperature was set to 45 ℃ and the pH was adjusted to 4.

5. The mixture was pretreated with cellulase and saccharifying enzyme for 0.5-2 h. After cooling to room temperature, the pH was adjusted to 11 for protein extraction. After extraction, the supernatant was collected by centrifugation. The pH of the supernatant was adjusted to 4.5, and the mixture was allowed to stand before centrifugation and washing with water. Finally, the oat protein was obtained by drying.

6. The preparation method according to claim 4, characterized in that, The mass ratio of the flavor protease to the oat protein is 1:20-100; The enzymatic hydrolysis conditions for the flavor protease are 50℃ for 1-4 h. The flavor protease was selected as a compound flavor protease extracted by Aspergillus oryzae fermentation.

7. The preparation method according to claim 4, characterized in that, After obtaining the oat protein hydrolysate, the following steps are also included: The oat protein hydrolysate was separated using a YMC ODS C18 column, with deionized water, 10% ethanol aqueous solution, 30% ethanol aqueous solution, 50% ethanol aqueous solution, 70% ethanol aqueous solution and anhydrous ethanol as eluents, eluted and purified by chromatography at a certain flow rate, and different components were collected. Then, the effect of different components on the secretion of GLP-1 by intestinal endocrine cells was detected, and the component that stimulated the highest activity of GLP-1 secretion was selected as the target component, which is the enzymatic hydrolysate containing the active oligopeptide of claim 1.

8. An enzymatic hydrolysis product containing the active oligopeptide of claim 1, prepared by any one of the preparation methods of claims 4-7.

9. The use of the active oligopeptide of claim 1, and the enzymatic hydrolysis product containing the active oligopeptide of claim 1, in the preparation of a product having at least one of the following functions: Promotes GLP-1 secretion; Reduce the degree of damage to the liver and pancreas; Improve glucose and lipid metabolism disorders; To prevent or help lower blood sugar.

10. A product characterized in that, The product contains the active oligopeptide of claim 1, or the enzymatic hydrolysis product of claim 8, and the product has at least one of the following functions: Promotes GLP-1 secretion; Reduce the degree of damage to the liver and pancreas; Improve glucose and lipid metabolism disorders; It helps lower blood sugar; The products include food, functional food / health food, and pharmaceuticals.

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