A wheat peptide with hypoglycemic effect, its preparation method and application

Abstract

The present invention discloses a wheat peptide with hypoglycemic effect, its preparation method and application, belonging to the field of biomedical technology. The present invention provides a bioactive peptide LPQ or LPQF with hypoglycemic effect. Through animal model experiments, it is verified that the bioactive peptide can effectively reduce blood glucose levels, improve related metabolic indexes, and has no obvious side effects. The present invention also provides a wheat peptide rich in oligopeptides containing the peptide segment LPQ. Through animal model experiments, it has a significant effect in reducing blood glucose. The present invention provides a safe and effective alternative for the development of diabetes treatment drugs, and has good market prospects and application potential.

Description

Technical Field

[0001] The present invention relates to the technical field of biomedicine, and in particular to a wheat peptide with blood sugar lowering effect, and a preparation method and application thereof. Background Art

[0002] Diabetes mellitus (DM) is a chronic disease characterized by hyperglycemia, caused by absolute or relative insulin deficiency and utilization disorders. The disease is mainly divided into three types: type 1, type 2 and gestational diabetes. The cause is mainly attributed to the combined effects of genetic and environmental factors, including decreased insulin secretion caused by islet cell dysfunction, or the body's insensitivity to insulin, or both, which prevents glucose in the blood from being effectively utilized and stored. The symptoms of diabetes are mainly manifested as "three mores and one less", namely, polydipsia, polyuria, polyphagia and weight loss, which not only affects the patient's quality of life, but also increases the risk of serious complications such as cardiovascular disease and kidney disease, bringing a heavy economic burden to patients and society.

[0003] In this context, the search for safe and effective blood sugar-lowering ingredients has received increasing attention. Although traditional drug treatments are effective, they are often accompanied by side effects and drug resistance, which makes patients face difficulties in choosing treatment options. Therefore, the development of blood sugar-lowering products based on natural ingredients has become a trend.

[0004] Bioactive peptides are a class of peptide compounds that are beneficial to the life activities of biological organisms or have physiological effects. They are composed of two or more amino acids connected by peptide bonds and have a variety of biological functions. Compared with proteins, bioactive peptides have the advantages of simple spatial structure, high stability, low immunogenicity or no immunogenicity. In recent years, domestic and foreign scholars have continuously isolated polypeptide substances with hypoglycemic function from natural animals and plants and human bodies, and studied their structure and mechanism of action, opening up a new path for the prevention and treatment of diabetes.

[0005] For example, patent document CN119192288A discloses the use of multiple enzymes to enzymolyze defatted peanut meal, and screened out four bioactive peptides with antioxidant and hypoglycemic functions from the enzymolysis products, with amino acid sequences of AFPKFR, PFPIK, APPFDPNKPK, and FATPVPLPK. Patent document CN119019492A discloses the use of neutral protease to enzymolyze spirulina, and screened out bioactive peptides with DPP-IV inhibitory activity from the enzymolysis products, with amino acid sequences of IASY and IAGIDE.

[0006] Wheat peptides are prepared from wheat protein (gluten) through processes such as biological enzymolysis, refining, and spray drying. Patent document CN114990180A discloses that gluten is enzymolyzed successively with alkaline protease, compound enzyme, and flavor protease, decolorized and deodorized after inactivating the enzyme, and the enzymolysis solution is subjected to 0.25μm membrane filtration and 10nm membrane filtration successively, and then dried to obtain wheat peptides, which have the effect of assisting in reducing blood sugar. Therefore, by analyzing the peptide map of wheat peptides and screening out high-efficiency peptide segments related to the blood sugar-lowering effect, it will provide a basis for the development of new blood sugar-lowering preparations. Summary of the Invention

[0007] The purpose of the present invention is to provide a natural small molecule bioactive peptide with a blood sugar-lowering effect and apply it to the development of related blood sugar-lowering products.

[0008] To achieve the above purpose, the present invention adopts the following technical solutions:

[0009] The present invention successively performs biological enzymolysis on wheat gluten with alkaline protease, neutral protease, and flavor protease, then adsorbs the enzymolysis solution with ZGC108DQ gel strong acid cation exchange resin, and obtains clarified wheat peptides after desorbing with hydrochloric acid solution. The polypeptide sequences of the clarified wheat peptides before and after gastrointestinal simulated digestion are analyzed by LC-MS / MS peptide map analysis technology, and it is found that the clarified wheat peptides are rich in oligopeptides with the LPQF peptide segment (amino acid sequence: Leu-Pro-Gln-Phe), and the peptide segment LPQF is generated after gastrointestinal digestion. The molecular docking technology is used to explore the interaction between the peptide segment and dipeptidyl peptidase-IV (DPP-IV). The results show that both the peptide segments LPQ and LPQF have high molecular docking energies with DPP-IV. Further, the tripeptide LPQ and the tetrapeptide LPQF are artificially synthesized, and functional verification finds that these peptide segments have DPP-IV inhibitory activity and blood sugar-lowering functions.

[0010] Therefore, the present invention provides a bioactive peptide with a blood sugar-lowering effect, and the amino acid sequence of the bioactive peptide is Leu-Pro-Gln or Leu-Pro-Gln-Phe.

[0011] The present invention also provides a method for preparing the bioactive peptide. The bioactive peptide can be prepared by solid-phase synthesis method. Specifically, the Fmoc solid-phase synthesis strategy is adopted, using Fmoc-protected amino acids as raw materials, and Wang resin is selected as the solid-phase carrier. Glutamine, proline, and leucine residues are introduced successively to extend the peptide chain from the C-terminus to the N-terminus to solid-phase synthesize the tripeptide LPQ, or phenylalanine, glutamine, proline, and leucine residues are introduced successively to extend the peptide chain from the C-terminus to the N-terminus to solid-phase synthesize the tetrapeptide LPQF.

[0012] The present invention also provides the use of the bioactive peptide in the preparation of a drug for preventing or treating diabetes or a food for assisting in reducing blood sugar. The research of the present invention shows that the bioactive peptide has a blood sugar lowering effect. In in vitro experiments, the bioactive peptide exhibits significant DPP-IV inhibitory activity; in a hyperglycemic zebrafish model, the blood sugar lowering effect after the intervention of the bioactive peptide is better than that of the positive control metformin. Therefore, it can be applied to the development of related products for reducing blood sugar.

[0013] Furthermore, the diabetes is type 2 diabetes. The manifestations of the diabetes include hyperglycemia and insulin resistance.

[0014] Another object of the present invention is to provide a wheat peptide having a blood sugar lowering effect. The wheat peptide is rich in oligopeptides containing the peptide segment LPQ, and the amino acid sequence of the peptide segment LPQ is Leu-Pro-Gln;

[0015] The preparation method of the wheat peptide includes:

[0016] (1) Enzymatic hydrolysis: Mix wheat gluten and water at a mass ratio of 1:15 - 30, adjust the pH to 8.0 ± 0.2, add alkaline protease accounting for 0.5% of the total weight of gluten and hydrolyze for 15 - 30 min; then shear at a rotation speed of 10000 - 15000 r / min for 15 - 30 min; then add alkaline protease accounting for 0.5% - 3.0% of the total weight of gluten and neutral protease accounting for 0.5% - 2.0% of the total weight of gluten and continue to hydrolyze for 4 - 6 h. After completion, add flavor protease accounting for 0.1% - 1.5% of the total weight of gluten and act for 15 - 30 min;

[0017] (2) Separation: Centrifuge the enzymatic hydrolysate at a rotation speed of 4000 - 6000 r / min and take the supernatant. Add strongly acidic cation resin to the supernatant, adjust the pH to 3.0, stir and adsorb, then filter. Take the filter cake and redissolve it in a hydrochloric acid solution with a mass concentration of 3% - 5% for desorption. Take the supernatant and obtain the wheat peptide through nanofiltration, concentration, sterilization, and drying.

[0018] The wheat peptides provided by the present invention are rich in oligopeptides with a specific amino acid sequence Leu - Pro - Gln. Through systematic research and experimental verification, it is proved that they have a significant effect on lowering blood sugar. In a hyperglycemic mouse model induced by a high - fat diet, after the intervention of the wheat peptides, the blood sugar level can be effectively reduced, and related metabolic indexes can be improved, the trend of impaired glucose tolerance can be improved, the insulin resistance index can be reduced, and the body weight growth rate and fat index can be significantly down - regulated. The research results show that the active components of the wheat peptides can play a role through multiple pathways such as regulating insulin secretion, improving insulin sensitivity, and down - regulating inflammatory factors, so as to achieve a comprehensive blood - sugar - lowering effect. Therefore, it can be applied to the development of related products for lowering blood sugar.

[0019] Furthermore, the oligopeptide containing the peptide segment LPQ is at least one of the peptide segments with amino acid sequences LPQFEELLNR (SEQ ID NO.2), LPQFEELR (SEQ ID NO.3), LPQFAELR (SEQ ID NO.4), LPQFEALR (SEQ ID NO.5), LPQFEELNR (SEQ ID NO.6), LPQFEF (SEQ ID NO.7), LPQQL (SEQ ID NO.8).

[0020] The present invention provides the application of the wheat peptides described above in the preparation of drugs for preventing or treating diabetes or foods for assisting in lowering blood sugar. The wheat peptides play a blood - sugar - lowering role after gastrointestinal digestion.

[0021] The present invention also provides a pharmaceutical composition for preventing or treating diabetes. The pharmaceutical composition includes a bioactive peptide with an amino acid sequence of Leu - Pro - Gln or Leu - Pro - Gln - Phe in an effective dose, or a wheat peptide rich in oligopeptides containing the peptide segment LPQ.

[0022] In the pharmaceutical composition provided by the present invention, the tripeptide LPQ or the tetrapeptide LPQF or the wheat peptide rich in oligopeptides containing the peptide segment LPQ can be used as the only active ingredient for lowering blood sugar, or can be compounded with other active ingredients with blood - sugar - lowering functions.

[0023] Furthermore, the pharmaceutical composition also includes a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier is any preparation or carrier medium that can deliver the active substance in an effective dose of the present invention, does not interfere with the biological activity of the active substance, and has no toxic or side effects on the host or subject.

[0024] Furthermore, the pharmaceutically acceptable carrier includes one or more of the following: fillers, wetting agents, disintegrants, binders, or lubricants.

[0025] The present invention uses the tripeptide LPQ, or the tetrapeptide LPQF, or a wheat peptide rich in polypeptides containing the peptide segment LPQ as the main active ingredient, adds a pharmaceutically acceptable carrier, and is prepared into a preparation according to the preparation method recorded in pharmaceutics. Further, the dosage form of the pharmaceutical composition may be, but is not limited to, an oral preparation. Specifically, the dosage form may be, but is not limited to, oral liquid, capsule, microcapsule powder, tablet, granule or emulsion.

[0026] The present invention also provides a health food for assisting in reducing blood sugar, which includes a bioactive peptide with an amino acid sequence of Leu-Pro-Gln or Leu-Pro-Gln-Phe as the active ingredient, or a wheat peptide rich in oligopeptides containing the peptide segment LPQ, and a food-grade excipient acceptable in food science.

[0027] The food-grade excipient acceptable in food science is a food-grade excipient that can deliver the active substance in an effective dose of the present invention without interfering with the biological activity of the active substance.

[0028] Further, the dosage form of the health food is beverage, oral liquid, capsule, microcapsule powder, tablet, granule or emulsion.

[0029] The beneficial effects of the present invention are as follows:

[0030] (1) The present invention provides a bioactive peptide LPQ or LPQF with a blood sugar-lowering effect. Through animal model experiments, it is verified that the bioactive peptide can effectively reduce the blood sugar level, improve related metabolic indexes, and has no obvious side effects. Therefore, the bioactive peptide can be applied to the preparation of drugs or foods for reducing blood sugar.

[0031] (2) The present invention provides a wheat peptide rich in oligopeptides containing the peptide segment LPQ. Through animal model experiments, it has a significant effect in reducing blood sugar. The present invention provides a safe and effective alternative for the development of diabetes treatment drugs, and has good market prospects and application potential. Description of the Drawings

[0032] Figure 1It is the secondary mass spectrum of the peptide segment LPQF. In the figure, the vertical coordinate Intensity(%) represents relative abundance; L / P / Q / F represents the tetrapeptide sequence; y1 represents the first fragment ion generated by the cleavage of the C-terminus of the peptide segment; b2 represents the second fragment ion generated by the cleavage of the N-terminus of the peptide segment; y2 represents the second fragment ion generated by the cleavage of the C-terminus of the peptide segment; y2-H2O represents the second fragment ion after dehydration generated by the cleavage of the C-terminus of the peptide segment; y2-NH3 represents the second fragment ion after deamination generated by the cleavage of the C-terminus of the peptide segment; b3-H2O represents the third fragment ion after dehydration generated by the cleavage of the N-terminus of the peptide segment; y3-NH3 represents the third fragment ion after deamination generated by the cleavage of the C-terminus of the peptide segment; bMax represents the maximum value at the N-terminus; yMax represents the maximum value at the C-terminus; Error(Da) represents the deviation.

[0033] Figure 2 It is the secondary mass spectrum of the peptide segment LPQFE. In the figure, the vertical coordinate Intensity(%) represents relative abundance; L / P / Q / F / E represents the pentapeptide sequence; y1 represents the first fragment ion generated by the cleavage of the C-terminus of the peptide segment; b2 represents the second fragment ion generated by the cleavage of the N-terminus of the peptide segment; y2 represents the second fragment ion generated by the cleavage of the C-terminus of the peptide segment; b3 represents the third fragment ion generated by the cleavage of the N-terminus of the peptide segment; y3 represents the third fragment ion generated by the cleavage of the C-terminus of the peptide segment; y3-H2O represents the third fragment ion after dehydration generated by the cleavage of the C-terminus of the peptide segment; y3-NH3 represents the third fragment ion after deamination generated by the cleavage of the C-terminus of the peptide segment; b4 represents the fourth fragment ion generated by the cleavage of the N-terminus of the peptide segment; y4 represents the fourth fragment ion generated by the cleavage of the C-terminus of the peptide segment; pre[1+] represents the charge [1+]; bMax represents the maximum value at the N-terminus; yMax represents the maximum value at the C-terminus; Error(Da) represents the deviation.

[0034] Figure 3 It is the secondary mass spectrum of the peptide segment LPQQL. In the figure, the vertical coordinate Intensity(%) represents relative abundance; L / P / Q / Q / L represents the pentapeptide sequence; y2-H2O represents the second fragment ion after dehydration generated by the cleavage of the C-terminus of the peptide segment; y2-NH3 represents the second fragment ion after deamination generated by the cleavage of the C-terminus of the peptide segment; b3 represents the third fragment ion generated by the cleavage of the N-terminus of the peptide segment; y4 represents the fourth fragment ion generated by the cleavage of the C-terminus of the peptide segment; pre[1+] represents the charge [1+]; bMax represents the maximum value at the N-terminus; yMax represents the maximum value at the C-terminus; Error(Da) represents the deviation.

[0035] Figure 4It is the second mass spectrometry diagram of the peptide segment LPQFEELLNR. In the diagram, the vertical coordinate Intensity(%) represents relative abundance; L / P / Q / F / E / E / L / L / N / R represents the decapeptide sequence; b2 represents the second fragment ion generated by the cleavage at the N-terminus of the peptide segment; y2 represents the second fragment ion generated by the cleavage at the C-terminus of the peptide segment; b3 represents the third fragment ion generated by the cleavage at the N-terminus of the peptide segment; y3 represents the third fragment ion generated by the cleavage at the C-terminus of the peptide segment; y5 represents the fifth fragment ion generated by the cleavage at the C-terminus of the peptide segment; y6 represents the sixth fragment ion generated by the cleavage at the C-terminus of the peptide segment; y7 represents the seventh fragment ion generated by the cleavage at the C-terminus of the peptide segment; y9-NH3 represents the ninth deaminated fragment ion generated by the cleavage at the C-terminus of the peptide segment; y9 represents the ninth fragment ion generated by the cleavage at the C-terminus of the peptide segment; bMax represents the maximum value at the N-terminus; yMax represents the maximum value at the C-terminus; Error(Da) represents the deviation.

[0036] Figure 5 It is the second mass spectrometry diagram of the peptide segment LPQFEELR. In the diagram, the vertical coordinate Intensity(%) represents relative abundance; L / P / Q / F / E / E / L / R represents the octapeptide sequence; y1 represents the first fragment ion generated by the cleavage at the C-terminus of the peptide segment; b2 represents the second fragment ion generated by the cleavage at the N-terminus of the peptide segment; y2 represents the second fragment ion generated by the cleavage at the C-terminus of the peptide segment; y3 represents the third fragment ion generated by the cleavage at the C-terminus of the peptide segment; y4 represents the fourth fragment ion generated by the cleavage at the C-terminus of the peptide segment; y5 represents the fifth fragment ion generated by the cleavage at the C-terminus of the peptide segment; y6 represents the sixth fragment ion generated by the cleavage at the C-terminus of the peptide segment; y7 represents the seventh fragment ion generated by the cleavage at the C-terminus of the peptide segment; bMax represents the maximum value at the N-terminus; yMax represents the maximum value at the C-terminus; Error(Da) represents the deviation.

[0037] Figure 6It is the secondary mass spectrometry diagram of the peptide segment LPQFAELR. In the figure, the vertical coordinate Intensity(%) represents relative abundance; L / P / Q / F / A / E / L / R represents the octapeptide sequence; y1 represents the first fragment ion generated by the cleavage of the C-terminus of the peptide segment; b2 represents the second fragment ion generated by the cleavage of the N-terminus of the peptide segment; y2 represents the second fragment ion generated by the cleavage of the C-terminus of the peptide segment; y3 represents the third fragment ion generated by the cleavage of the C-terminus of the peptide segment; y4 represents the fourth fragment ion generated by the cleavage of the C-terminus of the peptide segment; y5 represents the fifth fragment ion generated by the cleavage of the C-terminus of the peptide segment; y6-H2O represents the sixth dehydrated fragment ion generated by the cleavage of the C-terminus of the peptide segment; y6 represents the sixth fragment ion generated by the cleavage of the C-terminus of the peptide segment; y7 represents the seventh fragment ion generated by the cleavage of the C-terminus of the peptide segment; bMax represents the maximum value at the N-terminus; yMax represents the maximum value at the C-terminus; Error(Da) represents the deviation.

[0038] Figure 7 It is the secondary mass spectrometry diagram of the peptide segment LPQFEALR. In the figure, the vertical coordinate Intensity(%) represents relative abundance; L / P / Q / F / EA / L / R represents the octapeptide sequence; y1 represents the first fragment ion generated by the cleavage of the C-terminus of the peptide segment; b2 represents the second fragment ion generated by the cleavage of the N-terminus of the peptide segment; y2 represents the second fragment ion generated by the cleavage of the C-terminus of the peptide segment; b3-H2O represents the third dehydrated fragment ion generated by the cleavage of the N-terminus of the peptide segment; y4 represents the fourth fragment ion generated by the cleavage of the C-terminus of the peptide segment; y5 represents the fifth fragment ion generated by the cleavage of the C-terminus of the peptide segment; y6-NH3 represents the sixth deaminated fragment ion generated by the cleavage of the C-terminus of the peptide segment; y6 represents the sixth fragment ion generated by the cleavage of the C-terminus of the peptide segment; y7 represents the seventh fragment ion generated by the cleavage of the C-terminus of the peptide segment; bMax represents the maximum value at the N-terminus; yMax represents the maximum value at the C-terminus; Error(Da) represents the deviation.

[0039] Figure 8 It is the secondary mass spectrometry diagram of the peptide segment LPQFEELNR. In the figure, the vertical coordinate Intensity(%) represents relative abundance; L / P / Q / F / E / E / L / NR represents the nonapeptide sequence; y2 represents the second fragment ion generated by the cleavage of the C-terminus of the peptide segment; y3 represents the third fragment ion generated by the cleavage of the C-terminus of the peptide segment; y5 represents the fifth fragment ion generated by the cleavage of the C-terminus of the peptide segment; y6 represents the sixth fragment ion generated by the cleavage of the C-terminus of the peptide segment; y8 represents the eighth fragment ion generated by the cleavage of the C-terminus of the peptide segment; bMax represents the maximum value at the N-terminus; yMax represents the maximum value at the C-terminus; Error(Da) represents the deviation.

[0040] Figure 9It is the secondary mass spectrum of the peptide LPQFEF. In the figure, the vertical coordinate Intensity(%) represents relative abundance; L / P / Q / F / E / F represents the hexapeptide sequence; b2 represents the second fragment ion generated by the cleavage at the N-terminus of the peptide; b3 represents the third fragment ion generated by the cleavage at the N-terminus of the peptide; b4 represents the fourth fragment ion generated by the cleavage at the N-terminus of the peptide; b5 represents the fifth fragment ion generated by the cleavage at the N-terminus of the peptide; y5 represents the fifth fragment ion generated by the cleavage at the C-terminus of the peptide; pre[1+] represents the charge [1+]; bMax represents the maximum value at the N-terminus; yMax represents the maximum value at the C-terminus; Error(Da) represents the deviation.

[0041] Figure 10 It is a schematic diagram of the binding of the tripeptide LPQ to DPP-IV.

[0042] Figure 11 It is a schematic diagram of the binding of the tetrapeptide LPQF to DPP-IV.

[0043] Figure 12 It is the DPP-IV inhibition rate of the tripeptide LPQ and the tetrapeptide LPQF.

[0044] Figure 13 It is the hypoglycemic effect of the tripeptide LPQ and the tetrapeptide LPQF in the hyperglycemic zebrafish model. In the figure, the * symbol represents the significant difference compared with the blank control group (NC), * represents p <0.05, **** represents p <0.0001; the # symbol represents the significant difference compared with the model group (MC), ## represents p <0.01, represents p <0.001, # represents p <0.0001.

[0045] Figure 14 It is to compare the DPP-IV inhibition rate (A) and the hypoglycemic effect (B) of LPQ / LPQF and LP. Among them, the letters a, b, and c in A represent the significant differences between groups. The same letters indicate no significant differences between groups, and different letters indicate significant differences between groups. p <0.05; the * symbol in B represents the significant difference compared with the blank control group (NC), ** represents p <0.01, *** represents p <0.001, the # symbol represents the significant difference compared with the model group (MC), ## represents p <0.01.

[0046] Figure 15To compare the DPP-IV inhibition rates (A) and hypoglycemic effects (B) of LPQ / LPQF with those of FP / VP, where the letters a, b, and c in A indicate significant differences between groups, the same letter indicates no significant difference between groups, and different letters indicate significant differences between groups. p <0.05; in B, the * symbol indicates a significant difference compared with the blank control group (NC), and *** indicates p <0.001, and **** indicates p <0.0001, and the # symbol indicates a significant difference compared with the model group (MC), and ## indicates p <0.01.

[0047] Figure 16 To compare the DPP-IV inhibition rates (A) and hypoglycemic effects (B) of LPQ / LPQF with those of LPT, where the letters a, b, and c in A indicate significant differences between groups, the same letter indicates no significant difference between groups, and different letters indicate significant differences between groups. p <0.05; in B, the * symbol indicates a significant difference compared with the blank control group (NC), and *** indicates p <0.001, and **** indicates p <0.0001, and the # symbol indicates a significant difference compared with the model group (MC), and ## indicates p <0.01.

[0048] Figure 17 To compare the hypoglycemic effects of LPQ / LPQF with those of IPI. In the figure, the * symbol indicates a significant difference compared with the blank control group (NC), and **** indicates p <0.0001, and the # symbol indicates a significant difference compared with the model group (MC), and ## indicates p <0.01.

[0049] Figure 18 The hypoglycemic effect of wheat peptides containing the peptide segment LPQ in a hyperglycemic mouse model. Where A is the experimental design, B is the energy intake, C is the body weight, D is the adipocyte trait size, and E is the H&E staining result of epididymal fat; in the figure, the * symbol indicates a significant difference compared with the normal group, and **** indicates p <0.0001, and the # symbol indicates a significant difference compared with the model establishment group, and # indicates p <0.05.

[0050] Figure 19 The hypoglycemic effect of wheat peptides containing the peptide segment LPQ in a hyperglycemic mouse model. Where A is the oral glucose tolerance, B is the area under the blood glucose curve, C is the fasting blood glucose, and D is the insulin resistance index; in the figure, the * symbol indicates a significant difference compared with the normal group, and * indicatesp < 0.05, ** indicates p < 0.01, **** indicates p < 0.0001, # indicates the significant difference compared with the model group, # indicates p < 0.05, ## indicates p < 0.01, indicates p < 0.001. Specific embodiments

[0051] The present invention will be further described below in conjunction with specific embodiments. The following embodiments are only used to illustrate the present invention and are not used to limit the scope of application of the present invention. Without departing from the spirit and essence of the present invention, any modification or replacement of the methods, steps or conditions of the present invention belongs to the scope of the present invention.

[0052] Unless otherwise specified, the test methods used in the following embodiments are all conventional methods; the materials, reagents, etc. used, unless otherwise specified, are reagents and materials that can be obtained from commercial channels.

[0053] Wheat protein (gluten) was purchased from Binzhou Zhongyu Food Co., Ltd.; alkaline protease (derived from Bacillus licheniformis) was purchased from Angel Enzyme Preparation (Yichang) Co., Ltd.; neutral protease (derived from Bacillus subtilis) was purchased from Nanning Pangbo Bioengineering Co., Ltd.; flavor protease was purchased from Angel Enzyme Preparation (Yichang) Co., Ltd.

[0054] The amino acid sequence is from the N-terminus to the C-terminus from left to right.

[0055] Example 1

[0056] Preparation and analysis of clarified wheat peptides

[0057] 1. Preparation of clarified wheat peptides by enzymatic hydrolysis: Put wheat protein (gluten) and water in a ratio of 1:20 into an enzymatic hydrolysis reaction tank, adjust the pH of the feed liquid to 8.0 ± 0.2, add 0.5% of alkaline protease (derived from Bacillus licheniformis) based on the total weight of gluten and hydrolyze for 30 min, then turn on the shearing machine, with a shearing speed of 10,000 r / min and shear for 30 min.

[0058] After the shearing is completed, adjust the pH of the feed liquid to 8.0 ± 0.2, add 2.5% of alkaline protease (derived from Bacillus licheniformis) based on the total weight of gluten and 1.0% of neutral protease (derived from Bacillus subtilis) based on the total weight of gluten and continue to hydrolyze for 4 h. After completion, use 0.8% of flavor protease based on the total weight of gluten and act for 30 min.

[0059] After the enzymatic hydrolysis is completed, separate by a disc centrifuge (rotation speed 6000 r / min) and intercept the supernatant.

[0060] Add ZGC108DQ gel strong acid cation exchange resin to the supernatant after butterfly separation, adjust the pH to 3.0, stir and adsorb for 2 h. After the adsorption is completed, filter through a plate and frame filter press. Dissolve the filter cake in a 3% hydrochloric acid solution by mass for 60 min of desorption. Take the supernatant and obtain wheat peptide (glutamine peptide) powder through nanofiltration, concentration, sterilization, and spray drying.

[0061] 2. Analysis of wheat peptide (glutamine peptide)

[0062] Determine the main components and molecular weight distribution range of the wheat peptide (glutamine peptide) prepared in step 1. The results are shown in Table 1 - Table 3:

[0063] Table 1. Content of main components of glutamine peptide

[0064] Component Content (%) Moisture 4.02 Ash 4.21 Protein Content (on dry basis) 83.12 Degree of Hydrolysis 12.38 Peptide Content (on dry basis) 50.31

[0065] Table 2. Relative molecular weight distribution of glutamine peptide

[0066] Relative Molecular Weight Range, Da Wheat Peptide (Glutamine Peptide), % >10000 2.61 5000-10000 6.60 5000-3000 3.53 3000-2000 3.72 2000-1000 9.89 1000-500 18.62 500-180 38.84 <180 16.20

[0067] Table 3. Amino acid composition of glutamine peptide

[0068]

[0069] Example 2

[0070] Screening of active peptide segments

[0071] In this example, in vitro simulated gastrointestinal digestion of wheat peptide and LC-MS / MS were used to identify peptide segments with correlation before and after digestion. Molecular docking technology was used to explore its specific mechanism of action with dipeptidyl peptidase-IV (DPP-IV), and efficient DPP-IV inhibitory peptides were screened out.

[0072] 1. In vitro simulated digestion: Take 10 mL of the sample solution in Example 1 (pre-warmed in a 37 °C water bath), add 9 mg / mL NaCl solution (physiological saline, pre-warmed in a 37 °C water bath), adjust the pH value to 2 with 1 mol / L HCl, add 4 mL of gastric juice (0.4 g of pepsin dissolved in 0.01 mol / L HCl solution) to the system, and complete the simulation of gastric digestion by shaking at 37 °C in a water bath at 120 r / min for 2 h. Pre-adjust the pH value of the digested sample to 6.5. Pipette 2 mL of the sample after gastric digestion into a centrifuge tube, add 4 mL of intestinal juice (0.02 g of trypsin and 0.12 g of porcine bile salt are respectively dissolved in 10 mL of 0.1 mol / L NaHCO3 solution to obtain 2 mg / mL trypsin solution and 12 mg / mL bile solution, and the pancreatic juice and bile are mixed evenly at a ratio of 1:1), adjust the pH to 7.0, and complete the simulation of intestinal digestion by shaking at 37 °C in a water bath at 120 r / min for 2 h. All experiments are carried out in the dark and anaerobic conditions. The samples after simulated digestion are freeze-dried and stored at -80 °C until analysis.

[0073] 2. Identification of bioactive peptide segments of wheat peptides before and after digestion by LC-MS / MS

[0074] Dissolve the sample prepared in Example 1 and the digested sample prepared in Step 1 above in NH4HCO3 solution, add dithiothreitol solution, and place it in a 56 °C water bath for reduction for 1 h. Subsequently, add iodoacetamide solution and react in the dark for 40 min. After desalting, evaporate the solvent to dryness, and then dissolve the sample with 10 μL of mobile phase A (0.1% formic acid) into a liquid phase injection vial. Subsequently, perform LC-MS / MS analysis.

[0075] Chromatographic conditions: Analytical column (Acclaim PepMap RPLC C18, 150×150 mm, 3 μm); Mobile phase A (0.1% formic acid); Mobile phase B (0.1% formic acid and 80% acetonitrile); Flow rate (600 nL / min). Gradient elution program: 0 - 2 min, 4% B - 8% B; 2 - 45 min, 8% B - 40% B; 45 - 55 min, 40% B - 60% B; 55 - 56 min, 60% B - 95% B; 56 - 66 min, 95% B.

[0076] Mass spectrometry conditions: Full scan MS is performed by Orbitrap for the first - level scan, scan range (100~1500 m / z), resolution (70000), maximum ion introduction time (100 ms), automatic gain control (3×10 6); Use high-energy collision dissociation to fragment the top 20 precursor ions that meet the tandem (MS / MS) fragmentation conditions and scan with Orbitrap, resolution (17500), maximum ion injection time (50 ms), automatic gain control (1×10 5 ). The raw data obtained from mass spectrometry was analyzed using the De novo software of PEAKS Studio for peptide sequence analysis.

[0077] Analysis of the peptide profiles before and after wheat peptide digestion found that multiple peptide segments containing the LPQ sequence were rich both before and after wheat peptide digestion. After sorting, they are shown in Table 4.

[0078] Table 4. LPQ amino acid sequences in wheat peptides

[0079]

[0080] The secondary mass spectrometry diagrams of the above peptide segments are as Figure 1 - Figure 9 shown.

[0081] 3. High molecular docking scores of LPQ / LPQF with DPP-IV

[0082] Dipeptidyl peptidase-IV (DPP-IV) can weaken its hypoglycemic effect by degrading glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP). Inhibiting DPP-IV activity can maintain the activity of these incretin hormones, thereby promoting insulin secretion and inhibiting glucagon release to achieve blood glucose control. DPP-IV inhibitors have become a core class of drugs for treating type 2 diabetes by selectively blocking the activity of this enzyme to enhance the function of the endogenous incretin system. Therefore, in this example, molecular docking of wheat peptide segments with DPP-IV was performed to screen for small peptide segments that may have hypoglycemic effects.

[0083] First, download the crystal structure (1WCY) of DPP-IV from the PDB protein database. After removing water molecules and adding hydrogen atoms to the receptor target through DiscoveryStudio software, define its active center. Perform molecular docking of the peptide segment LPQ / LPQF with dipeptidyl peptidase-IV (DPP-IV).

[0084] The 2D and 3D diagrams of the molecular docking of the tripeptide LPQ with DPP-IV are as Figure 10As shown, analysis of chemical bonds revealed that the tripeptide LPQ binds to DPP-IV mainly through van der Waals forces, hydrogen bonds (conventional hydrogen bonds), and electrostatic interactions (salt bridges), with a docking energy of -32.50 kcal / mol. The tripeptide LPQ forms two van der Waals forces with amino acid residues SER458 and SER460, two hydrogen bonds with ARG471 and VAL459, and two electrostatic interactions with GLU408 and ARG471.

[0085] The 2D and 3D molecular docking diagrams of the tetrapeptide LPQF and DPP-IV are as Figure 11 shown. Analysis of chemical bonds revealed that the tetrapeptide binds to DPP-IV mainly through van der Waals forces, hydrogen bonds (including conventional hydrogen bonds and carbon-hydrogen bonds), hydrophobic interactions (alkyl groups), and electrostatic interactions (including salt bridges and attractive charge), with a docking energy of -52.58 kcal / mol. The tetrapeptide LPQF forms seven van der Waals forces with amino acid residues SER473, TYR456, SER458, GLY406, VAL459, ILE418, and LEU57, three hydrogen bonds with ARG471 and SER59, two hydrophobic interactions with ILE405, and three electrostatic interactions with ARG471 and GLU408.

[0086] From the molecular docking results, it can be seen that the tripeptide LPQ and the tetrapeptide LPQF can bind to DPP-IV, thereby inhibiting DPP-IV and playing a hypoglycemic role.

[0087] The peptide segments LPQ and LPQF were synthesized by Gil Biochemical (Shanghai) Co., Ltd. with a purity ≥ 98% for subsequent functional verification.

[0088] Example 3

[0089] In vitro DPP-IV inhibition rate of LPQ / LPQF

[0090] The DPP-IV inhibition rate of the peptide segments was measured using a DPP-IV inhibitor kit from Cayman (manufacturer). The measurement principle is as follows: This test uses the fluorescent substrate gly - pro - aminomethylcoumarin (AMC) to measure the activity of DPP-IV. DPP cleaves the peptide bond to release the free AMC group, generating fluorescence with an excitation wavelength of 350 - 360 nm and an emission wavelength of 450 - 465 nm.

[0091] The DPP-IV inhibition rates of the tripeptide LPQ and the tetrapeptide LPQF were measured at 1000 µM and 500 µM respectively. The measurement method is as follows:

[0092] (1) 100% initial active wells - Add 30 µL of diluted buffer, 10 µL of DPP-IV, and 10 µL of pure water to three wells.

[0093] (2) Background wells - Add 40 µL of diluted buffer and 10 µL of pure water to three wells.

[0094] (3) Inhibitor wells - Add 30 µL of diluted experimental buffer, 10 µL of diluted DPP-IV, and 10 µL of inhibitor (intervention samples, LPQ and LPQF at concentrations of 1000 µM and 500 µM) to three wells.

[0095] The specific operation is as follows: First, add 30 µL of buffer, 10 µL of DPP-IV enzyme, and 10 µL of sample to a 96-well plate, mix well, add 50 µL of substrate solution to start the reaction. After incubating at 37 °C for 30 min, read the fluorescence at an excitation wavelength of 350 - 360 nm and an emission wavelength of 450 - 465 nm. Record the fluorescence intensity as A sample. Record the result measured using buffer instead of DPP-IV as A blank, and record the result measured using buffer instead of the sample as A control. The calculation formula for the DPP-IV inhibition rate is as follows: Inhibition rate (%) = (A control - A sample) / (A control - A blank) × 100%.

[0096] The results are as Figure 12 shown. When the tripeptide LPQ and the tetrapeptide LPQF are at 1000 µM and 500 µM, their DPP-IV inhibition rates are both higher than 85%, indicating that the tripeptide LPQ and the tetrapeptide LPQF have DPP-IV inhibition potential.

[0097] Example 4

[0098] Hypoglycemic effect of LPQ / LPQF in hyperglycemic zebrafish model

[0099] Use the hyperglycemic zebrafish model to determine the hypoglycemic effects of the tripeptide LPQ and the tetrapeptide LPQF. Four-day-old wild AB strain zebrafish are placed in six-well plates, with 3 wells in parallel for each group and 10 zebrafish in each well. The grouping interventions are as follows:

[0100] (1) Blank control group (NC): System water;

[0101] (2) Model group (MC): 333 μM alloxan + 2.67% glucose + system water;

[0102] (3) Metformin positive control group (Met): 333 μM alloxan + 2.67% glucose + 5 μg / mL metformin + system water;

[0103] (4)Tripeptide LPQ intervention group (LPQ-5, LPQ-1, LPQ-0.1): 333 μM alloxan + 2.67% glucose + 5 / 1 / 0.1 μg / mL tripeptide LPQ + system water.

[0104] (5)Tetrapeptide LPQF intervention group (LPQF-5, LPQF-1, LPQF-0.1): 333 μM alloxan + 2.67% glucose + 5 / 1 / 0.1 μg / mL tetrapeptide LPQF + system water.

[0105] After zebrafish at 4 days old were intervened according to the above intervention protocol for 24 h, they were washed repeatedly 3 times with PBS solution to wash away the sugar solution on the surface of the zebrafish. The zebrafish in the six-well plate were transferred to a 1.5 mL centrifuge tube using a disposable dropper, the excess liquid in the centrifuge tube was aspirated, 0.1 mL of absolute ethanol was added, and after standing in a cool place for 15 min, it was transferred to an oven for 120 min (60 °C) until dried (ground up). Finally, 5 μL of ultrapure water was added to the centrifuge tube, sonicated and shaken evenly for 10 min, and 2 μL was aspirated to measure the glucose value (S) using a glucometer.

[0106] The results were as Figure 13 shown. Compared with the blank control group (NC group), the blood glucose value of the model group (MC group) after modeling with 333 μM alloxan + 2.67% glucose for 24 h increased significantly ( p < 0.0001), indicating that a hyperglycemic model was successfully established. After intervention with the positive drug 5 μg / mL metformin (Met group), there was a significant down-regulation compared with the MC group ( p < 0.01). When the tripeptide LPQ and the tetrapeptide LPQF were intervened at concentrations of 5 μg / mL, 1 μg / mL, and 0.1 μg / mL, the blood glucose value could be significantly down-regulated compared with the MC group ( p < 0.001), and it was down-regulated to have no significant difference from the NC group ( p > 0.05). Moreover, the significance of the difference between LPQ / LPQF and the MC group at the measured doses (5 μg / mL, 1 μg / mL, and 0.1 μg / mL) ( p < 0.001) was stronger than the significance between the positive control metformin and the MC group ( p < 0.05), indicating that the tripeptide LPQ and the tetrapeptide LPQF have hypoglycemic effects.

[0107] Example 5

[0108] In vitro DPP-IV inhibitory rate of dipeptides with P in the second position and blood glucose lowering in zebrafish

[0109] The above research shows that both the tripeptide LPQ and the tetrapeptide LPQF have potential DPP-IV inhibitory effects and hypoglycemic effects. To explore whether further reducing one amino acid still has DPP-IV inhibitory effects and hypoglycemic potential, the peptide segments LP (amino acid sequence: Leu-Pro), FP (amino acid sequence: Phe-Pro), and VP (amino acid sequence: Val-Pro) with a purity ≥ 98% were synthesized by Gil Biochemical (Shanghai) Co., Ltd. for subsequent functional verification experiments.

[0110] 1. LPQ / LPQF is superior to LP

[0111] The DPP-IV inhibition experiment method was the same as that of "in vitro DPP-IV inhibition rate of LPQ / LPQF". The DPP-IV inhibition rates of LPQ / LPQF and LP at 1000 μM and 500 μM were compared respectively.

[0112] The results are as Figure 14 shown in A of p <0.0001).

[0113] The experimental method for exploring hypoglycemic effect in zebrafish model was the same as that of "hypoglycemic effect of LPQ / LPQF in hyperglycemic zebrafish model". The hypoglycemic effects of LPQ / LPQF and LP at a dose of 1 μg / mL were compared.

[0114] The results are as Figure 14 shown in B of p <0.001), and the blood glucose level was decreased to no significant difference compared with the NC group ( p >0.05). However, there was no significant difference between the dipeptide LP and the MC group after intervention, indicating that the tripeptide LPQ and the tetrapeptide LPQF have stronger hypoglycemic effects than the dipeptide LP.

[0115] 2. LPQ / LPQF is superior to FP / VP

[0116] The results of in vitro DPP-IV inhibition rate and zebrafish hypoglycemic effect are as Figure 15 shown. The tripeptide LPQ and the tetrapeptide LPQF have stronger hypoglycemic effects than the dipeptide FP / VP.

[0117] Example 6

[0118] In vitro DPP-IV inhibition rate and zebrafish hypoglycemic effect of other tripeptides with P at the second position

[0119] The above research shows that both the tripeptide LPQ and the tetrapeptide LPQF have potential DPP-IV inhibitory effects and hypoglycemic effects. To explore whether there are still DPP-IV inhibitory effects and hypoglycemic potential after the change of the third amino acid, the peptide segment LPT (amino acid sequence: Leu-Pro-Thr) with a purity ≥ 98% was synthesized by GL Biochem (Shanghai) Ltd. for subsequent functional verification experiments.

[0120] The method for the DPP-IV inhibition experiment was the same as that for the "in vitro DPP-IV inhibition rate of LPQ / LPQF", and the DPP-IV inhibition rates of LPQ / LPQF and LPT at 1000 μM and 500 μM were compared.

[0121] The results are as Figure 16 shown in A of p <0.001, p <0.0001).

[0122] The method for exploring the hypoglycemic effect in the zebrafish model was the same as that for the "hypoglycemic effect of LPQ / LPQF in the hyperglycemic zebrafish model", and the hypoglycemic effects of LPQ / LPQF and LPT at a dose of 1 μg / mL were compared.

[0123] The results are as Figure 16 shown in B of p <0.001), and were decreased to no significant difference from the NC group ( p >0.05). However, there was no significant difference between the tripeptide LPT and the MC group after intervention, indicating that the tripeptide LPQ and the tetrapeptide LPQF have stronger hypoglycemic effects than the tripeptide LPT.

[0124] Example 7

[0125] Comparison of the hypoglycemic effects of LPQ / LPQF and IPI

[0126] IPI has been reported to have DPP-IV inhibitory effects. In this example, the hypoglycemic effects of LPQ / LPQF and IPI were compared. The peptide segment IPI (amino acid sequence: Ile-Pro-Ile) with a purity ≥ 98% was synthesized by GL Biochem (Shanghai) Ltd. for subsequent functional verification experiments.

[0127] The method for exploring the hypoglycemic effect in the zebrafish model was the same as that for the "hypoglycemic effect of LPQ / LPQF in the hyperglycemic zebrafish model", and the hypoglycemic effects of LPQ / LPQF and IPI at a dose of 1 μg / mL were compared.

[0128] The results are as follows Figure 17 shown. When the tripeptide LPQ and the tetrapeptide LPQF were intervened at a concentration of 1 μg / mL, compared with the MC group, they could significantly down-regulate the blood glucose value ( p < 0.01), and the down-regulation was not significantly different from that of the NC group ( p > 0.05). After IPI intervention, the blood glucose could also be significantly down-regulated ( p < 0.01). From the perspective of the significance level, the hypoglycemic effects of the tripeptide LPQ and the tetrapeptide LPQF were equivalent to that of IPI. From the average value, the hypoglycemic effects of the tripeptide LPQ and the tetrapeptide LPQF were better than that of IPI.

[0129] Example 8

[0130] Hypoglycemic effect of wheat peptide (prepared in Example 1) in a hyperglycemic mouse model

[0131] A hyperglycemic mouse model established by a high-fat diet was used to explore whether wheat peptide has hypoglycemic efficacy. The experiment is as follows:

[0132] (1) Mice and materials

[0133] Male C57BL / 6 mice at 6 - 8 weeks of age (certificate number: 20230524Abzz0100018521, ethics batch number: MG20230531082) were purchased from Hangzhou Medical College, and the production license number: SCXK(Zhe)2019 - 0042. The basal diet for mice was purchased from Shenzhen Maohua Biotechnology Co., Ltd. The 45% purified high-fat diet (MD12032) was provided by Jiangsu Medison Biotechnology Co., Ltd.

[0134] (2) Animal grouping and feeding

[0135] 60 male C57BL / 6 mice at 6 - 8 weeks of age were housed in the Animal Experiment Center of Zhejiang University of Technology at a temperature of 23 ± 1 °C with a 12 h day-night cycle. After the mice were purchased, they were fed with the normal diet for one week of adaptation, and then divided into a normal diet normal group and a high-fat diet model group. The normal group was fed with the normal diet, and the high-fat group was fed with a 45% kcal high-fat diet for 8 consecutive weeks. After the eighth week, LPQ / LPQF wheat peptides at high, medium, and low doses were used for intervention for 8 weeks. The specific experimental design is shown in Figure 18 A in. The food intake and body weight of the mice were recorded weekly. After 16 weeks, the mice were fasted for 16 h, blood was collected from the orbital cavity, and tissues such as epididymal fat were taken, frozen quickly with liquid nitrogen, transported with dry ice and stored in a -80 °C refrigerator for standby. The blood samples were centrifuged at 3000 rpm for 10 min to obtain serum. A small part of the epididymal fat tissue was fixed in 4% neutral formaldehyde.

[0136] (3) Hematoxylin-eosin (H&E) staining of fat sections

[0137] After fixation with 4% neutral paraformaldehyde for 24 h, the epididymal fat and liver of mice were dehydrated, embedded in paraffin, sectioned (3 μm in thickness), baked for 2 h, dewaxed and hydrated, stained with H&E, and finally dehydrated and sealed with neutral plastic. Observation and analysis were performed under an optical microscope, and fat sections were photographed in a random field of view. The size of epididymal fat cells in each section was statistically analyzed using Image J software.

[0138] (4) OGTT test for mice

[0139] One day before the test, the mice were fasted for 16 h without water deprivation, gavaged with glucose at 2 g / kg, and the gavage time was recorded. Blood glucose levels were measured at 0 min, 30 min, 60 min, and 120 min using a blood glucose meter. A blood glucose value-time curve was drawn, and the area under the curve of each experimental group was calculated.

[0140] (5) Determination of biochemical related indexes and hormone levels in serum

[0141] The glucose content (fasting blood glucose) in serum was determined using a kit from Nanjing Jiancheng. The insulin content in mouse serum was determined using a GeneMe ELISA kit. The insulin resistance index was calculated based on the fasting blood glucose value and fasting insulin value of the mice, and the calculation formula was as follows:

[0142] Insulin resistance index = fasting blood glucose value of mice (mmol / L) × fasting insulin value of mice (μU / mL) / 22.5.

[0143] (6) Result analysis

[0144] a. Hypoglycemic effect of wheat peptides in hyperglycemic mouse models - body weight and fat

[0145] After the mice with insulin resistance induced by a high-fat diet were intervened with the wheat peptides prepared in Example 1 at doses of 1000 mg / kg (high dose), 250 mg / kg (medium dose), and 50 mg / kg (low dose) for 8 weeks, the food intake, body weight, and epididymal fat of the mice were as Figure 18 shown. As can be seen from B in Figure 18 , there were no significant differences in energy intake among the normal group, the model control group, the high-dose wheat peptide intervention group, the medium-dose wheat peptide intervention group, and the low-dose wheat peptide intervention group ( p > 0.05). As can be seen from C in Figure 18 , in terms of body weight gain, the weight growth rate of the model control group was significantly higher than that of the normal group ( p<0.0001), after intervention with 1000 mg / kg (high dose) and 250 mg / kg (medium dose) of wheat peptide, the body weight growth rate was significantly downregulated compared with the model control group ( p <0.05), indicating that wheat peptides have the effect of reducing weight gain. The results of H&E staining of epididymal fat are as follows Figure 18 As shown in E in Figure 1, the size of fat cells was statistically analyzed, and the results are as follows: Figure 18 As shown in D, it can be seen that the high-fat diet model significantly enlarges the adipocytes ( p <0.0001), and after intervention with 250 mg / kg dose of wheat peptide, the size of adipocytes in the wheat peptide group was significantly reduced compared with the model control group ( p <0.01), indicating that wheat peptide has lipid-lowering effect.

[0146] b. The hypoglycemic effect of wheat peptides in a hyperglycemic mouse model: blood sugar and insulin resistance

[0147] The results of the oral glucose tolerance test (OGTT) are as follows: Figure 19 As shown in A. The blood glucose levels of the four groups of mice fed with high-fat diets were higher than those of the normal group at each time point after oral glucose administration, and the blood glucose levels of the mice in the model control group at 0 min, 30 min, 90 min and 120 min were significantly higher than those of the mice in the model control group. Statistical analysis of the area under the blood glucose curve showed that ( Figure 19 B), 16 weeks of high-fat diet feeding significantly increased the area under the blood glucose curve of mice, and the intake of wheat peptides at doses of 1000 mg / kg, 250 mg / kg and 50 mg / kg significantly improved the trend of impaired glucose tolerance caused by high-fat diet ( p <0.01), indicating that wheat peptides may have a significant effect on alleviating high blood sugar induced by high-fat diet. In addition, from the fasting blood sugar results ( Figure 19 C) It can also be seen that compared with the model control group, the intervention of wheat peptide can significantly adjust the fasting blood glucose value ( p <0.05). In terms of insulin resistance, Figure 19 As shown in D, the insulin resistance index of the model control group mice was significantly higher than that of the normal group ( p <0.05), indicating that mice fed a high-fat diet had already developed significant insulin resistance. Compared with the model control group, wheat peptides at three intervention doses, high, medium and low, could significantly reduce the high-fat diet-induced insulin resistance index ( p <0.05), indicating that wheat peptides can significantly improve insulin resistance induced by a high-fat diet.

Claims

1. A wheat peptide having a blood sugar lowering effect, characterized in that: The preparation method of the wheat peptide comprises: (1) Enzymatic hydrolysis: wheat gluten and water were mixed at a mass ratio of 1:20, the pH was adjusted to 8.0±0.2, and 0.5% alkaline protease based on the total weight of gluten was added for enzymatic hydrolysis for 30 min; then sheared at a speed of 10,000 r / min for 30 min; then 2.5% alkaline protease based on the total weight of gluten and 1.0% neutral protease based on the total weight of gluten were added for further enzymatic hydrolysis for 4 h, and after the end, 0.8% flavor protease based on the total weight of gluten was added for 30 min; (2) Separation: The enzymatic hydrolysate was centrifuged at a speed of 6000 r / min, and the supernatant was taken. Strong acid cationic resin was added to the supernatant, and the pH was adjusted to 3.

0. The mixture was stirred for adsorption and then filtered. The filter cake was redissolved in a 3% hydrochloric acid solution for desorption. The supernatant was nanofiltered, concentrated, sterilized, and dried to obtain the wheat peptide.

2. The wheat peptide according to claim 1, characterized in that The wheat peptide is rich in at least one of the peptide segments having the amino acid sequences of LPQFEELLNR, LPQFEELR, LPQFAELR, LPQFEALR, LPQFEELNR, LPQFEF, and LPQQL.

3. Use of the wheat peptide according to claim 1 or 2 in the preparation of a drug for preventing or treating diabetes or a food for assisting in lowering blood sugar.

4. The use according to claim 3, characterized in that The diabetes is type 2 diabetes.

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