Difunctional active peptide with ACE (angiotensin converting enzyme) and DPP-IV (dipeptidyl peptidase-IV) inhibitory activity as well as preparation method and application thereof

Abstract

The invention discloses a bifunctional active peptide with ACE (angiotensin converting enzyme) and DPP-IV (dipeptidyl peptidase-IV) inhibitory activity as well as a preparation method and application of the bifunctional active peptide, and belongs to the technical field of active peptide biology. The difunctional novel active peptide with ACE and DPP-IV inhibitory activity is prepared and identified from sea cucumber enzymatic hydrolysate, not only can inhibit ACE activity, but also has DPP-IV inhibitory activity, and the difunctional novel active peptide has application value in preparation of products with blood pressure and blood sugar reducing effects. The active peptide provided by the invention can significantly inhibit the activity of ACE, has DPP-IV inhibitory activity, has the characteristics of simple structure, safety, strong activity and the like, can exert nutritional and health-care effects, is expected to provide effective active ingredients for developing food and products having no side effects and having the effects of reducing blood pressure and blood sugar or having the effects of reducing blood pressure and blood sugar and the like, and has a broad application prospect. Wide application prospects are realized.

Description

Technical Field

[0001] The present invention relates to the field of active peptide biotechnology, and in particular to a bifunctional active peptide having both ACE and DPP-IV inhibitory activities, and a preparation method and application thereof. Background Art

[0002] Hypertension and type 2 diabetes are both serious diseases worldwide. Angiotensin-converting enzyme (ACE) and dipeptidyl peptidase IV (DPP-IV) play an important role in the pathogenesis of the two diseases. ACE catalyzes the conversion of angiotensin I to angiotensin II through the RAS system in the blood pressure regulation system, and can also regulate the KKS system to degrade bradykinin, thereby promoting blood pressure. Therefore, inhibiting ACE activity is considered an important treatment for lowering blood pressure. DPP-IV is an enzyme widely expressed in the intestine and liver, which is involved in the degradation of a variety of bioactive peptides in the body. The main substrates of DPP-IV include glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), both of which play a vital role in blood sugar regulation. By inhibiting the activity of DPP-IV, the degradation of GLP-1 and GIP is reduced, their concentration in the body is increased, and they can work for a longer time, promote insulin secretion, inhibit glucagon secretion, and effectively lower blood sugar levels. Therefore, inhibiting DPP-IV activity is considered an important treatment for lowering blood sugar. Although chemically synthesized ACE inhibitors (such as captopril and enalapril) and DPP-IV inhibitors (such as sitagliptin and vildagliptin) have shown good effects in lowering blood pressure and blood sugar in clinical practice, their therapeutic properties bring certain potential side effects, which in turn limits their long-term use. In contrast, functional peptides with ACE and DPP-IV inhibitory activity derived from natural proteins have become a more ideal choice due to their high safety and low side effects.

[0003] As a traditional marine organism rich in various nutrients, sea cucumbers are high in protein and low in fat. They have multiple functions such as enhancing immunity, promoting growth and development, and regulating blood lipids. Its nutrients are mainly collagen, sulfated polysaccharides, sea cucumber saponins, rich trace elements, vitamins, and minerals. In particular, sea cucumbers are rich in up to 50% protein and are a high-quality source of protein. Protein is an important component of body tissue repair and regeneration, and is of great significance for maintaining life activities. The rich protein resources in sea cucumbers provide great possibilities for generating functional products after hydrolysis. In addition, sea cucumbers, as a valuable traditional tonic food, have long been considered safe and reliable. Its processing byproducts such as sea cucumber intestines and eggs have also been proven to contain a large amount of protein resources, which are important raw materials for the development of functional products. And using these raw materials to explore functional products can not only facilitate the development of active ingredients, but also greatly increase the high-value utilization of sea cucumber byproducts. Existing studies have shown that the enzymatic hydrolyzate obtained by enzymolysis of sea cucumbers has certain ACE inhibitory activity and DPP-IV inhibitory activity, and multifunctional peptides that can inhibit both ACE and DPP-IV can be mined from them. This not only helps to develop high value-added functional foods, but also provides new auxiliary treatment methods for natural interventions in hypertension and type 2 diabetes. Summary of the invention

[0004] The purpose of the present invention is to provide a bifunctional active peptide having both ACE and DPP-IV inhibitory activities and a preparation method and application thereof, so as to solve the problems existing in the above-mentioned prior art.

[0005] To achieve the above object, the present invention provides the following solutions: One of the technical solutions of the present invention is a bifunctional active peptide having both ACE and DPP-IV inhibitory activities, and its amino acid sequence is shown in SEQ ID NO.1.

[0006] The second technical solution of the present invention is a method for preparing a bifunctional active peptide having both ACE and DPP-IV inhibitory activities, comprising the following steps: (1) enzymatically hydrolyzing sea cucumber intestinal ovum with aminopeptidase and proline endonuclease to obtain sea cucumber intestinal ovum enzymatic hydrolysis products; (2) After the sea cucumber intestinal egg hydrolysate is subjected to ultrafiltration and desalting treatment, the peptide sequences in the sea cucumber intestinal egg hydrolysate are identified by LC-MS / MS liquid chromatography-mass spectrometry to construct a peptide database of the sea cucumber intestinal egg hydrolysate, and obtain an enzymatic hydrolysate containing the bifunctional active peptide having both ACE and DPP-IV inhibitory activity; (3) Screen and identify bifunctional active peptides with both ACE and DPP-IV inhibitory activities.

[0007] The third technical solution of the present invention is the use of the bifunctional active peptide having both ACE and DPP-IV inhibitory activity in the preparation of ACE inhibitors and / or DPP-IV inhibitors.

[0008] A fourth technical solution of the present invention is an ACE inhibitor, comprising the bifunctional active peptide having both ACE and DPP-IV inhibitory activities.

[0009] A fifth technical solution of the present invention is a DPP-IV inhibitor, comprising the bifunctional active peptide having both ACE and DPP-IV inhibitory activities.

[0010] The sixth technical solution of the present invention is the use of the bifunctional active peptide having both ACE and DPP-IV inhibitory activity in the preparation of drugs for lowering blood pressure and / or blood sugar.

[0011] The seventh technical solution of the present invention is a drug for lowering blood pressure and / or blood sugar, comprising the bifunctional active peptide having both ACE and DPP-IV inhibitory activity.

[0012] Based on the above technical solution, the present invention has the following technical effects: The present invention prepares and identifies a bifunctional novel active peptide having both ACE and DPP-IV inhibitory activity from sea cucumber enzymatic hydrolysate, which can not only inhibit the activity of ACE, but also has DPP-IV inhibitory activity, and relieves hypertension and type 2 diabetes at the same time, and its bifunctionality makes it have application value in the preparation of blood pressure lowering and blood sugar lowering efficacy products. The active peptide involved in the present invention can significantly inhibit the activity of ACE, and at the same time has DPP-IV inhibitory activity, has the characteristics of simple structure, safety, strong activity, etc., can play the role of nutrition and health care, and is expected to provide effective active ingredients for the development of food and blood pressure lowering, blood sugar lowering or blood pressure lowering and blood sugar lowering efficacy products without side effects, and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0014] Figure 1 This is the amino acid structure and sequence secondary mass spectrometry identification diagram of a new bifunctional active peptide with both ACE and DPP-IV inhibitory activity.

[0015] Figure 2This is a graph showing the inhibition rate of ACE at different concentrations of FPGGPP, a new bifunctional active peptide with both ACE and DPP-IV inhibitory activity.

[0016] Figure 3 This is a schematic diagram of the three-dimensional conformation of the docking of FPGGPP, a new bifunctional active peptide with both ACE and DPP-IV inhibitory activity, with ACE.

[0017] Figure 4 This is a two-dimensional schematic diagram of the docking force between FPGGPP, a new bifunctional active peptide with both ACE and DPP-IV inhibitory activity, and ACE.

[0018] Figure 5 This is a graph showing the inhibition rate of FPGGPP, a new bifunctional active peptide with both ACE and DPP-IV inhibitory activity, on DPP-IV at different concentrations.

[0019] Figure 6 This is a schematic diagram of the three-dimensional conformation of the docking of FPGGPP, a new bifunctional active peptide with both ACE and DPP-IV inhibitory activity, with DPP-IV.

[0020] Figure 7 for Figure 6 Enlarged view of the boxed part.

[0021] Figure 8 This is a two-dimensional schematic diagram of the docking force between FPGGPP, a new bifunctional active peptide with both ACE and DPP-IV inhibitory activity, and DPP-IV. DETAILED DESCRIPTION

[0022] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0023] It should be understood that the terms described in the present invention are only for describing special embodiments and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. Each smaller range between the intermediate value in any stated value or stated range and any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0024] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0025] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to those skilled in the art. The present application description and examples are exemplary only.

[0026] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0027] The technical solutions described in the present invention, unless otherwise specified, are all conventional solutions in the art, and the reagents or raw materials used, unless otherwise specified, are purchased from commercial channels or have been disclosed.

[0028] The embodiment of the present invention provides a bifunctional active peptide having both ACE and DPP-IV inhibitory activities, and its amino acid sequence is shown in SEQ ID NO.1.

[0029] The present invention also provides a method for preparing the bifunctional active peptide having both ACE and DPP-IV inhibitory activities, comprising the following steps: (1) enzymatically hydrolyzing sea cucumber intestinal ovum with aminopeptidase and proline endonuclease to obtain sea cucumber intestinal ovum enzymatic hydrolysis products; (2) After the sea cucumber intestinal egg hydrolysate is subjected to ultrafiltration and desalting treatment, the peptide sequences in the sea cucumber intestinal egg hydrolysate are identified by LC-MS / MS liquid chromatography-mass spectrometry to construct a peptide database of the sea cucumber intestinal egg hydrolysate, and obtain an enzymatic hydrolysate containing the bifunctional active peptide having both ACE and DPP-IV inhibitory activity; (3) Screen and identify bifunctional active peptides with both ACE and DPP-IV inhibitory activities.

[0030] In some specific embodiments, the enzymatic hydrolysis conditions are: the amount of the aminopeptidase used is 600 U / g, the amount of the proline endonuclease used is 1200 U / g, The dosage of the sea cucumber intestinal egg substrate is 8% (v / v), The enzymatic hydrolysis was carried out at 55℃ for 24 h, and the pH value was 7.5.

[0031] In some specific embodiments, the screening method is: identifying the molecular weight of peptides in the sea cucumber intestinal egg enzymatic hydrolysate, and screening peptide sequences with a molecular weight less than 1 kDa to form a peptide database of sea cucumber intestinal egg enzymatic hydrolysate; The DeltaM tool, PeptideRanker functional activity predictor, and Toixinpred safety and toxicity evaluation predictor were further used to predict the ACE and DPP-IV inhibitory activity and function of the peptides, and based on the functional scores, the bifunctional active peptides with both ACE and DPP-IV inhibitory activity were screened.

[0032] The embodiments of the present invention also provide the use of the bifunctional active peptide having both ACE and DPP-IV inhibitory activities in the preparation of ACE inhibitors and / or DPP-IV inhibitors.

[0033] The embodiment of the present invention further provides an ACE inhibitor, comprising the bifunctional active peptide having both ACE and DPP-IV inhibitory activities.

[0034] The embodiment of the present invention further provides a DPP-IV inhibitor, comprising the bifunctional active peptide having both ACE and DPP-IV inhibitory activities.

[0035] The embodiments of the present invention also provide the use of the bifunctional active peptide having both ACE and DPP-IV inhibitory activities in the preparation of drugs for lowering blood pressure and / or blood sugar.

[0036] The embodiment of the present invention also provides a drug for lowering blood pressure and / or blood sugar, comprising the bifunctional active peptide having both ACE and DPP-IV inhibitory activities.

[0037] The novel bifunctional active peptide with both ACE and DPP-IV inhibitory activity provided by the present invention can not only significantly inhibit the activity of ACE, but also effectively inhibit the activity of DPP-IV, play a role in preventing or even alleviating diseases such as hypertension and hyperglycemia, and has important significance for the actual production and theoretical research of multifunctional active peptides.

[0038] Example 1 Preparation of active peptides from sea cucumber intestinal eggs After pretreatment such as washing and homogenization, the sea cucumber intestinal eggs were subjected to composite enzymatic hydrolysis using aminopeptidase and proline endonuclease. The dosage of aminopeptidase was 600 U / g, the dosage of proline endonuclease was 1200 U / g, the dosage of sea cucumber intestinal eggs substrate was 8% (v / v), the enzymatic hydrolysis conditions were 55℃ for 24 h, and the pH value of the enzymatic hydrolysis system was continuously monitored and adjusted to 7.5 during the enzymatic hydrolysis process. The final product of sea cucumber intestinal eggs enzymatic hydrolysis was ultrafiltered and desalted using a solid phase extraction cartridge (SEP-PAK C18 solid phase extraction cartridge, Waters, USA) to prepare sea cucumber intestinal eggs hydrolysate containing active peptides. LC-MS / MS liquid chromatography-mass spectrometry was further used to determine the molecular weight and sequence of each peptide prepared in the sea cucumber intestinal eggs enzymatic hydrolysate, and peptides with a molecular weight less than 1 kDa were selected to form a peptide database of sea cucumber intestinal eggs enzymatic hydrolysate.

[0039] Example 2 Screening and mining of active peptide FPGGPP with ACE inhibitory activity According to the LC-MS / MS liquid phase mass spectrometry method, the peptide molecular weight and sequence of the obtained sea cucumber intestinal egg enzymatic hydrolysate were identified, and the peptide sequences with a molecular weight of less than 1 kDa were selected to form a peptide database, totaling 1876 entries; the DeltaM tool, PeptideRanker functional activity predictor and Toixinpred safety and toxicity evaluation predictor were further used to predict the ACE and DPP-IV inhibitory activity and function of the peptides, and based on the functional score, active peptides with potential ACE and DPP-IV inhibitory activity were screened, and their secondary mass spectrometry structures were analyzed to obtain the amino acid sequence and structure of the active peptide.

[0040] The results are as follows Figure 1 As shown, the secondary mass spectrum of the screened active peptide shows that the amino acid sequence and arrangement of the active peptide are SEQ ID NO.1: FPGGPP.

[0041] Example 3 Verification of ACE inhibitory activity of active peptide FPGGPP The ACE inhibitory activity of the active peptide FPGGPP was determined using a 96-well plate using a visible spectrophotometer. The ACE inhibition rate was determined using 1 mM N-[3-(2-furyl)acryloyl]-L-phenylalanylglycylglycine (FAPPG) as a substrate; 0.1 U / mL angiotensin converting enzyme (ACE); HEPES as a buffer matrix (1.901 g HEPES reagent, 1.755 g NaCl reagent, dissolved in an appropriate amount of deionized water, and then adjusted to pH 8.3 with NaOH solution, and the volume was fixed to 100 mL for standby use).

[0042] The determination steps are as follows: add active peptide FPGGPP samples of different concentrations, as well as FAPPG substrate, ACE and buffer, etc. to a 96-well plate in sequence; use an enzyme-labeled instrument to measure the absorbance of the sample system at 340 nm and record it; place the sample system in a 37°C shaker for 30 min, and use an enzyme-labeled instrument to measure the absorbance of the sample system at 340 nm after the reaction; the ACE inhibition rate of the sample is determined by calculating the change in absorbance before and after the reaction.

[0043] The calculation formula is: ACE inhibition rate of sample (%) = 1-change in sample absorbance value / change in blank absorbance value.

[0044] The results are as follows Figure 2 As shown in the results, when the concentration of the active peptide FPGGPP is 10 μg / mL, the inhibition rate of the active peptide FPGGPP on ACE is 18.4%; when the concentration of the active peptide FPGGPP is 25 μg / mL, the inhibition rate of ACE is 50.02%; when the concentration of the active peptide FPGGPP is 50 μg / mL, the inhibition rate of ACE is 97.4%; when the concentration of the active peptide FPGGPP is 100 μg / mL, the inhibition rate of the active peptide FPGGPP is 103.2%; when the concentration of the active peptide FPGGPP continues to increase, the inhibition rate of ACE can reach 100%. According to the results of the ACE inhibitory activity assay, when the concentration of the active peptide FPGGPP is greater than 50 μg / mL, it can completely inhibit ACE. The above results show that the active peptide FPGGPP has an effective ACE inhibitory effect.

[0045] Example 4 Molecular simulation docking of active peptide FPGGPP and ACE The active peptide FPGGPP was docked with ACE by molecular simulation to further determine the mechanism of action of the active peptide when it exerts its ACE inhibitory activity. The three-dimensional conformational diagram of the active peptide FPGGPP was drawn using Discovery studio software, and molecular simulation docking was performed with the catalytic active center of ACE (PDB: 1o8a), and the interaction force between the active peptide FPGGPP and the key amino acid residues in the active center of ACE was analyzed based on the docking results.

[0046] The results are as follows Figure 3As shown in the figure, the active peptide FPGGPP can bind tightly to the active center of ACE and can be simulated and docked within the normal analysis time. The active peptide FPGGPP interacts with multiple amino acid residues in the active center of ACE, and the main amino acid residues are Phe527, Try520, Val518, Phe457, Glu411, His387, Glu384, Gln281, Trp279, Ala354, Ser355, Ala356, Glu376, His410, His353, Zn 2+ .

[0047] The results of the interaction analysis between the active peptide FPGGPP and the amino acid residues in the active center of ACE are as follows: Figure 4 The active peptide FPGGPP forms 4 hydrogen bonds with ACE, 5 interactions between alkyl groups, 1 π-π interaction, 4 carbon-hydrogen interactions, and Zn 2+ Combination effect.

[0048] In summary, the active peptide FPGGPP can bind to ACE mainly through hydrogen bonds, hydrophobic interactions and Zn 2+ The binding effect interacts with the active center residues of ACE and inhibits the activity of ACE. That is, from the perspective of molecular simulation docking, it is further proved that the active peptide FPGGPP has significant ACE inhibitory activity.

[0049] Example 5 Verification of DPP-IV inhibitory activity of active peptide FPGGPP The DPP-IV inhibitory activity of the active peptide FPGGPP was determined using a 96-well plate. The DPP-IV inhibition rate was determined using 1.6 mM Gly-Pro-pNA as substrate, 0.1 U / mL DPP-IV, and Tris-HCl as the buffer matrix (100 mM, pH 8.0).

[0050] The determination steps are as follows: add different concentrations of active peptide FPGGPP samples, as well as Gly-Pro-pNA substrate, DPP-IV and buffer solution into a 96-well plate in sequence; incubate the sample system at 37°C for 60 min and terminate the reaction, use an enzyme reader to measure the absorbance of the sample system at 504 nm and record it, and obtain the determined DPP-IV inhibition rate according to the calculation formula.

[0051] The calculation formula is: ; Among them, A, B, C, and D are the absorbance values ​​of the sample group, control group, sample blank group, and blank group at 504 nm, respectively.

[0052] The results are as follows Figure 5 As shown, when the concentration of the active peptide FPGGPP is 100 μg / mL, the inhibition rate of the active peptide FPGGPP on DPP-IV is 19.96%; when the concentration of the active peptide FPGGPP is 200 μg / mL, the inhibition rate of DPP-IV is 52.08%; when the concentration of the active peptide FPGGPP is 400 μg / mL, the inhibition rate of DPP-IV is 82.87%; when the concentration of the active peptide FPGGPP is 800 μg / mL, the inhibition rate of the active peptide FPGGPP on DPP-IV is 98.25%. According to the results of the DPP-IV inhibitory activity assay, the active peptide FPGGPP has effective DPP-IV inhibitory activity.

[0053] Example 6 Molecular simulation docking of active peptide FPGGPP and DPP-IV The active peptide FPGGPP was docked with DPP-IV by molecular simulation to further determine the mechanism of action of the active peptide when it exerts its DPP-IV inhibitory activity. The three-dimensional conformational diagram of the active peptide FPGGPP was drawn using Discovery studio software, and molecular simulation docking was performed with the catalytic active center of DPP-IV (PDB: 2QT9), and the interaction force between the active peptide FPGGPP and the key amino acid residues in the active center of DPP-IV was analyzed based on the docking results.

[0054] The results are as follows Figure 6 and Figure 7 As shown, the active peptide FPGGPP can bind tightly to the active center of DPP-IV and can be simulated and docked within the normal analysis time. The active peptide FPGGPP interacts with multiple amino acid residues in the active center of DPP-IV, and the main amino acid residues are Arg471, Ser458, Tyr456, Val558, Arg560, and Val459.

[0055] The results of the interaction force analysis between the active peptide FPGGPP and the amino acid residues in the active center of DPP-IV are as follows Figure 8 As shown, the active peptide FPGGPP forms 4 hydrogen bond forces, 4 Van der Waals forces and 1 salt bridge with DPP-IV.

[0056] In summary, the active peptide FPGGPP can bind to DPP-IV, and mainly interacts with the active center residues of DPP-IV through hydrogen bonds, Wandwala forces and salt bridges to inhibit the activity of DPP-IV. That is, from the perspective of molecular simulation docking, it is further proved that the active peptide FPGGPP has significant DPP-IV inhibitory activity.

[0057] In summary, the present invention prepares and identifies a novel bifunctional active peptide FPGGPP with both ACE and DPP-IV inhibitory activity from sea cucumber intestines and eggs by a composite enzyme hydrolysis method. The present invention obtains its amino acid sequence and structure by performing secondary mass spectrometry analysis on the active peptide, verifies the inhibitory activity of the active peptide FPGGPP on ACE and DPP-IV respectively, and docks the active peptide FPGGPP with the active centers of ACE and DPP-IV respectively by molecular simulation docking to illustrate its inhibitory mechanism on ACE and DPP-IV activity.

[0058] The present invention provides a novel bifunctional active peptide FPGGPP having both ACE and DPP-IV inhibitory activities, which has significant ACE inhibitory activity and DPP-IV inhibitory activity.

[0059] Obviously, the above embodiments of the present invention are merely examples for clearly illustrating the present invention, and are not intended to limit the implementation methods of the present invention. For ordinary technical users in the relevant field, other different forms of changes or modifications can be made on the basis of the above description. It is not necessary and impossible to list all the implementation methods here. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the protection scope of the claims of the present invention.

Claims

1. A bifunctional active peptide having both ACE and DPP-IV inhibitory activity, characterized in that: Its amino acid sequence is shown in SEQ ID NO.

1.

2. The method for preparing a bifunctional active peptide having both ACE and DPP-IV inhibitory activity as claimed in claim 1, characterized in that: The following steps are involved: (1) enzymatically hydrolyzing sea cucumber intestinal ovum with aminopeptidase and proline endonuclease to obtain sea cucumber intestinal ovum enzymatic hydrolysis products; (2) After the sea cucumber intestinal egg hydrolysate is subjected to ultrafiltration and desalting treatment, the peptide sequences in the sea cucumber intestinal egg hydrolysate are identified by LC-MS / MS liquid chromatography-mass spectrometry to construct a peptide database of the sea cucumber intestinal egg hydrolysate, and obtain an enzymatic hydrolysate containing the bifunctional active peptide having both ACE and DPP-IV inhibitory activity; (3) Screen and identify bifunctional active peptides with both ACE and DPP-IV inhibitory activities.

3. The preparation method according to claim 2, characterized in that: The enzymolysis conditions are as follows: the aminopeptidase is used in an amount of 600 U / g, the proline endonuclease is used in an amount of 1200 U / g, The dosage of the sea cucumber intestinal egg substrate is 8% (v / v), The enzymatic hydrolysis was carried out at 55℃ for 24 h, and the pH value was 7.

5.

4. The preparation method according to claim 2, characterized in that: The screening method comprises: identifying the molecular weight of peptides in the sea cucumber intestinal egg enzymatic hydrolysate, and screening peptide sequences with a molecular weight less than 1 kDa to form a peptide database of sea cucumber intestinal egg enzymatic hydrolysate; The DeltaM tool, PeptideRanker functional activity predictor, and Toixinpred safety and toxicity evaluation predictor were further used to predict the ACE and DPP-IV inhibitory activity and function of the peptides, and based on the functional scores, the bifunctional active peptides with both ACE and DPP-IV inhibitory activity were screened.

5. Use of the bifunctional active peptide having both ACE and DPP-IV inhibitory activities as claimed in claim 1 in the preparation of ACE inhibitors and / or DPP-IV inhibitors.

6. An ACE inhibitor, characterized in that The invention comprises the bifunctional active peptide having both ACE and DPP-IV inhibitory activities as claimed in claim 1.

7. A DPP-IV inhibitor, characterized in that The invention comprises the bifunctional active peptide having both ACE and DPP-IV inhibitory activities as claimed in claim 1.

8. Use of the bifunctional active peptide having both ACE and DPP-IV inhibitory activities as claimed in claim 1 in the preparation of drugs for lowering blood pressure and / or blood sugar.

9. A drug for lowering blood pressure and / or blood sugar, characterized in that: The invention comprises the bifunctional active peptide having both ACE and DPP-IV inhibitory activities as claimed in claim 1.

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