Multifunctional pea polypeptide and preparation method thereof

Abstract

The application discloses a multifunctional pea polypeptide and a preparation method thereof, and belongs to the technical field of functional polypeptides. The method comprises the following steps: extracting pea protein isolate by using an ultrasonic-assisted alkali dissolution and acid precipitation method; preparing the pea polypeptide by using a steam explosion technology-assisted enzymatic hydrolysis method; and evaluating the functional activity of the pea polypeptide by using a chemical method. The pea polypeptide prepared by the method has the functions of antioxidation, blood pressure reduction and blood sugar reduction. In the application, the pea polypeptide is obtained by using steam explosion treatment to assist enzymatic hydrolysis, and the above functions of the pea polypeptide are improved, so that the DPPH free radical scavenging capacity is 92%, the ABTS free radical scavenging capacity is 93%, the O2 ‑ free radical scavenging capacity is 98%, the angiotensin transferase ACE activity inhibition rate is 68%, the alpha-glucosidase activity inhibition rate is 75%, the alpha-amylase activity inhibition rate is 71%, and the foundation is laid for the wide application of the pea polypeptide in functional foods or special diet foods.

Description

Technical Field

[0001] This invention belongs to the field of functional polypeptide technology, specifically relating to a multifunctional pea polypeptide and its preparation method. Background Technology

[0002] Peas are an important source of plant protein. Due to their high nutritional value, low price, and ability to be mass-produced, they have a broad market potential. Furthermore, with the continuous improvement of dietary concepts, the deepening of research on alternative proteins, and increasing environmental awareness, the market demand for pea protein isolate is further growing. The bioactive peptides obtained from the enzymatic hydrolysis of pea protein isolate not only have the advantages of easy absorption, safety, and no side effects, but have also been reported to possess various functional activities, such as anti-inflammatory, antioxidant, immunomodulatory, and hypoglycemic effects.

[0003] Currently, enzymatic hydrolysis is widely used in peptide preparation. Increasingly, research utilizes physical modification techniques to assist enzymatic hydrolysis, aiming to improve the yield and functional activity of protein peptides. For example, patent CN112626155B discloses a method for preparing pea peptides. It uses high-pressure steam sterilization as a pretreatment to unfold the structure of pea protein, which then assists in complex enzymatic hydrolysis. The resulting pea peptide powder has a protein content ≥90% and a peptide yield of 73%. However, the high-pressure steam is only used to enhance the subsequent enzymatic hydrolysis reaction; the functional properties of the pea peptides are not investigated. Another example is a bifunctional soybean peptide and its preparation method, patent CN113197316A. This method uses infrared heat treatment to assist enzymatic hydrolysis, obtaining soybean peptides with both depressurizing and antioxidant activities. These peptides exhibit good emulsifying properties and emulsion stability, but their antioxidant performance needs further improvement.

[0004] Therefore, there is an urgent need for a new physical-assisted enzymatic hydrolysis method to prepare pea peptides, in order to further improve the functional properties of pea protein peptides, such as antioxidant, blood pressure lowering, and blood sugar lowering. Summary of the Invention

[0005] The purpose of this invention is to overcome the shortcomings of the existing technology and provide a method for preparing antioxidant pea peptides. This method utilizes a steam explosion-assisted enzymatic hydrolysis method to obtain a higher peptide yield, while simultaneously possessing antioxidant, blood pressure lowering, and blood sugar lowering functional properties.

[0006] To achieve the above objectives, the present invention adopts the following technical solution: a method for preparing antioxidant pea peptides, comprising the following steps:

[0007] S1. Pea protein isolate extraction: Pea raw materials are crushed and sieved and then dissolved in deionized water. The ratio of pea powder to water is 1:15-35 w / v. Ultrasonic technology is used to assist dissolution. The ultrasonic power is 100-500w, the ultrasonic time is 10-30min, and the pH is 8-10. Then, the protein in the pea powder is extracted by alkaline dissolution and acid precipitation. The pea protein isolate powder is obtained by freeze drying.

[0008] S2. Steam explosion treatment: The pea protein isolate obtained in step S1 above is added to the steam explosion cylinder of the steam explosion test bench for steam explosion treatment. The steam explosion time is 20-100s and the steam explosion pressure is 0.4-1.2MPa. The pea protein isolate is then freeze-dried to obtain steam-exploded pea protein isolate.

[0009] S3. Preparation of pea peptides: The steam-exploded pea protein powder obtained in step S2 above was dissolved in deionized water to prepare a substrate concentration of 10% and the pH was adjusted. Plant protease was added to carry out enzymatic hydrolysis. After the reaction was completed, the enzyme was inactivated by high temperature. Ultrafiltration was used to separate the ultrafiltrate with a molecular weight of <3kDa and freeze-drying to obtain pea active peptide powder.

[0010] In step S1, preferably, the ratio of pea flour to water is 1:29w / v, the ultrasonic power is 260w, the ultrasonic time is 27min, and the pH value is 9.45.

[0011] In step S2, preferably, the steam explosion time is 40s and the steam explosion pressure is 1.0MPa.

[0012] In step S1, the pea raw material is the protein-rich Zhongwan No. 9 pea, and the peas are ground into powder and passed through a 40-100 mesh sieve.

[0013] In step S1, the specific steps of the alkali dissolution and acid precipitation method are as follows: heating and ultrasonic treatment, adjusting the pH to 8-10 with 1.0 mol / L sodium hydroxide, stirring the sample solution continuously during the process, centrifuging to collect the supernatant, and then adjusting the pH to 2.5-5.5 with hydrochloric acid. The above steps are repeated 2-3 times, and the precipitate is washed with deionized water until neutral.

[0014] A multifunctional pea polypeptide was prepared according to the preparation method described above.

[0015] The multifunctional pea polypeptide described herein, its DPPH free radical scavenging rate and ABTS were measured. + Free radical scavenging rate, O2 - The free radical scavenging rates reached 92%, 93%, and 98%, respectively.

[0016] The multifunctional pea polypeptide exhibited an angiotensin-converting enzyme (ACE) inhibition rate of 68%.

[0017] The multifunctional pea polypeptide exhibited inhibition rates of 75% and 71% on the measured activities of α-glucosidase and α-amylase, respectively.

[0018] The beneficial effects of this invention are:

[0019] 1) In this invention, steam explosion treatment is used to assist enzymatic hydrolysis to obtain pea protein peptides, which improves the antioxidant activity of the pea peptides, enabling them to achieve a DPPH free radical scavenging capacity of 92%, an ABTS free radical scavenging capacity of 93%, and an O2 free radical scavenging capacity of 93%. - The free radical scavenging capacity is 98%; the angiotensin-converting enzyme (ACE) inhibition rate reaches 68%; and the inhibition rates of α-glucosidase and α-amylase activities are 75% and 71%, respectively; laying the foundation for the widespread application of pea polypeptides in functional foods or special dietary foods.

[0020] 2) Currently, steam explosion technology is more often used in the food industry for the modification and functional activity enhancement of dietary fiber, food polysaccharides, and plant polyphenols. However, there has been no further research on the modification and effects on pea protein and peptides. Therefore, the use of steam explosion technology to assist enzymatic hydrolysis in the preparation of pea protein peptides has created a new application for steam explosion technology and laid a theoretical foundation for the deep processing and comprehensive utilization of pea protein.

[0021] 3) The preparation method of the present invention uses steam explosion treatment to perform flexible thermal processing on proteins, which destroys the intermolecular forces of proteins, allowing them to stretch and helping to expose hydrophobic groups, improve their enzymatic hydrolysis efficiency, and facilitate the release of bioactive substances. It has a positive effect on the prevention and control of chronic non-communicable diseases and has the potential to be applied to pregnant women, the elderly and other patients with chronic diseases. It can be widely used in functional foods or special dietary foods.

[0022] 4) The overall process of the method of the present invention can be summarized as protein extraction and proteolytic hydrolysis. The operation process is simple and easy to implement. The entire process is green and safe and does not cause pollution to the environment. The cost is low and it can realize industrial-scale production, laying the foundation for the development of plant-based food ingredients. Attached Figure Description

[0023] Figure 1 This is a graph showing the effects of material-to-liquid ratio, pH value, ultrasonic time, and ultrasonic power on the extraction rate of pea protein isolate in the single-factor experiment of this invention.

[0024] Figure 2 This is a graph showing the influence of four factors on the response value in the response surface optimization experiment of this invention;

[0025] Figure 3 This is a graph showing the effect of steam explosion pressure and steam explosion time on the yield of pea peptides in the single-factor experiment of this invention.

[0026] Figure 4 The figure shows the effect of two factors on the yield of protein and peptide in the orthogonal optimization experiment.

[0027] Figure 5 The present invention relates to the effects of steam-exploded pea polypeptides on DPPH and ABTS. + O2 - Graph showing the results of free radical scavenging rate determination;

[0028] Figure 6 The graph shows the results of the determination of the inhibition rate of the steam-exploded pea protein polypeptide of the present invention on the activities of angiotensin-transferase (ACE), α-glucosidase, and α-amylase. Detailed Implementation

[0029] The present invention will be further explained and described below with reference to the accompanying drawings and specific embodiments.

[0030] The biological materials used in the following experiments included: alkaline protease (CAS No.: 9014-01-1) and neutral protease (CAS No.: 9068-59-1), purchased from Solarbio, with an enzyme activity of 2 × 10⁻⁶. 5 U / g.

[0031] Pea protein isolate extraction: Place 200g of peas in a grinder and grind for 15-20s to pass the ground peas through a 60-mesh sieve. Mix the pea powder thoroughly with deionized water, then adjust the pH of the solution to 9 with 1.0mol / L NaOH. After sonication, dispense the solution into 50mL centrifuge tubes and centrifuge at 8000×g for 20min at 4℃. Gently remove the centrifuge tubes and collect the supernatant. Adjust the pH to 4.6 with 1.0mol / L HCl, then centrifuge at 10000×g for 10min at 4℃. Collect the precipitate and freeze-dry it.

[0032] Based on the above steps for extracting pea protein isolate, the following experiments were conducted to determine the relevant variable parameters.

[0033] 1. Single-factor experimental design

[0034] 1) Effect of the pea flour to water ratio on the extraction of pea protein isolate

[0035] Using the solid-liquid ratio as a variable (1:15, 1:20, 1:25, 1:30, 1:35 w / v), with other conditions including pH=9, ultrasonic time 20 min, and ultrasonic power 300 W, the effect of the solid-liquid ratio on the extraction of pea protein isolate under different variable levels was determined, and the extraction rate of pea protein isolate was calculated.

[0036] Multiple experiments were conducted using different material-to-liquid ratios as variables to determine the content of pea protein isolate. The experimental results are as follows: Figure 1 As shown in Figure A, a better extraction rate of pea protein isolate can be achieved when the material-to-liquid ratio is 1:25.

[0037] 2) Effect of pH on the extraction of pea protein isolate

[0038] Using pH as a variable (8.0, 8.5, 9.0, 9.5, 10), with other conditions including a material-to-liquid ratio of 1:25, ultrasonic time of 20 min, and ultrasonic power of 300 W, the effect of pH on the extraction of pea protein isolate under different variable levels was determined, and the extraction rate of pea protein isolate was calculated.

[0039] Multiple experiments were conducted using different pH values ​​as variables to determine the content of pea protein isolate. The experimental results are as follows: Figure 1 As shown in Figure B, a better extraction rate of pea protein isolate can be achieved when the pH value is 9.

[0040] 3) Effect of ultrasound time on the extraction of pea protein isolate

[0041] Using ultrasonic time as a variable (10, 15, 20, 25, 30 min), and other conditions such as a solid-liquid ratio of 1:25, pH=9, and ultrasonic power of 300 W, the effect of ultrasonic time on the extraction of pea protein isolate under different variable levels was determined, and the extraction rate of pea protein isolate was calculated.

[0042] Multiple experiments were conducted using different ultrasound times as variables to determine the content of pea protein isolate. The experimental results are as follows: Figure 1 As shown in Figure C, a better extraction rate of pea protein isolate can be achieved when the ultrasonic time is 20 minutes.

[0043] 4) Effect of ultrasonic power on the extraction of pea protein isolate

[0044] Using ultrasonic power as a variable (100, 200, 300, 400, 500 W), a material-to-liquid ratio of 1:25, pH=9, and ultrasonic time of 20 min, the effect of ultrasonic power on the extraction of pea protein isolate under different variable levels was determined, and the extraction rate of pea protein isolate was calculated.

[0045] Multiple experiments were conducted using different ultrasonic powers as variables to determine the content of pea protein isolate. The experimental results are as follows: Figure 1 As shown in Figure D, a better extraction rate of pea protein isolate can be achieved when the ultrasonic power is 300W.

[0046] 2. Response surface optimization experimental design

[0047] Based on the single-factor experiments, a three-factor, three-level Box-Behnken central composite design experiment was conducted, as shown in Table 1 below. The pea protein content extracted from pea protein using ultrasound-assisted alkaline extraction was used as the response value (Y), and the material-to-liquid ratio (A), pH (B), ultrasound time (C), and ultrasound power (D) of pea protein isolate extraction using ultrasound-assisted alkaline pretreatment were used as experimental factors. The effects of these four factors on the response value were analyzed using response surface methodology. The experimental model design results are shown in Table 2 below. The conditions for pea protein isolate extraction using ultrasound-assisted alkaline pretreatment were optimized based on the pea protein isolate content.

[0048] Table 1 shows the levels of factors in response surface methodology experiments.

[0049]

[0050] Table 2 shows the analysis of variance for the Box-Behnken experimental regression model.

[0051]

[0052]

[0053] Experimental results are as follows Figure 2 As shown, the experimental model was fitted using Design Expert software with a quadratic multiple regression. The quadratic multiple regression equation for pea protein isolate content (Y) against material-to-liquid ratio (A), pH (B), ultrasonic time (C), and ultrasonic power (D) is as follows:

[0054] Y=76.10+0.19×A+1.15×B-1.07×C-1.33×D-0.28×A×B+1.97×A×C-2.29×A×D+1.31×B×C+0.34×B×D-0.63×C×D-8.68×A 2 -6.09×B 2 -3.90×C 2 -5.91×D 2 Based on the above quadratic multiple regression fitting equation and the regression model variance analysis table, the 3D response surface plot can be obtained, as shown below. Figure 2 As shown in (AF). Based on the regression equation, the highest predicted material-to-liquid ratio was 1:29, pH = 9.45, ultrasonic time 27 min, and ultrasonic power 260 W. Under these conditions, the pea extract protein content was 81.75%. There was no significant difference between this and the model's predicted value (p>0.05), and the model F-value was 44.17, indicating that the model was significant.

[0055] Steam explosion treatment: Accurately weigh 20g of pea protein isolate and add it to the steam explosion cylinder of the steam explosion test bench. Screw on the piston, set the steam explosion time to 20-100s and the pressure to 0.4-1.2Mpa. High-temperature and high-pressure gas is generated and enters the cylinder through the inlet valve. After 20-100s, the inlet valve is closed, and the pressure is instantly depressurized (0.00875s), completing the explosion of the material. Collect the exploded pea protein isolate and store it at -20℃ for freeze drying.

[0056] Preparation of pea polypeptides: Proteins were separated by dual-enzyme hydrolysis. The plant proteases included two types: alkaline protease and neutral protease. The reaction temperature was 35–65℃, the pH was 5.5–9.5, and the reaction time was 1–6 h.

[0057] Pea protein isolate hydrolysis process: Add 10g of pea protein isolate to 100mL of deionized water (substrate concentration 10%) → adjust the pH of the solution to 8.5 → adjust the temperature to 50℃ → add 3g of alkaline protease → hydrolyze at constant temperature and pH → extract under ultrasonic-assisted extraction for 3h → inactivate enzyme in a 95℃ water bath for 10min → adjust the pH of the solution to 7.0 → adjust the temperature to 40℃ → add 3g of neutral protease → hydrolyze at constant temperature and pH → extract under ultrasonic-assisted extraction for 3.5h → inactivate enzyme in a 95℃ water bath for 10min → centrifuge at 6000r / min for 15min → remove the lower precipitate → collect the supernatant → freeze-dry to obtain pea polypeptide powder.

[0058] Based on the steam explosion treatment steps described above, the following experiments were conducted to determine the relevant variable parameters.

[0059] 3. Single-factor experimental design

[0060] 1) Effect of steam explosion pressure on the yield of peptides prepared from pea protein isolate

[0061] Using steam explosion pressure as a variable (0.4, 0.6, 0.8, 1.0, 1.2 MPa) and a pressure holding time of 60 s, the effect of steam explosion pressure on the content of prepared protein peptides under different variable levels was determined, and the yield of pea protein peptides was calculated.

[0062] Multiple experiments were conducted using different steam explosion pressures as variables to determine the content of pea protein peptides. The experimental results are as follows: Figure 3 As shown in Figure A, a better yield of pea protein peptides can be achieved when the steam explosion pressure is 0.8 MPa.

[0063] 2) Effect of steam explosion time on the yield of peptides prepared from pea protein isolate

[0064] Using the steam explosion time as a variable (20, 40, 60, 80, 100 s) and the steam explosion pressure set at 0.8 MPa, the effect of steam explosion time on the content of prepared protein peptides under different variable levels was determined, and the yield of pea protein peptides was calculated.

[0065] Multiple experiments were conducted using different steam explosion times as variables to determine the content of pea protein peptides. The experimental results are as follows: Figure 3 As shown in B, a better yield of pea protein peptides can be achieved when the steam explosion time is 60s.

[0066] 4. Orthogonal experimental design

[0067] Based on the results of the single-factor experiment, the L9(32) orthogonal array was used to design a two-factor, three-level orthogonal experiment. The factor levels are shown in Table 3 below. The yield of peptides in steam explosion pressure and steam explosion time were used as indicators, and the comprehensive factors were used as indicators for evaluation of the orthogonal experiment. The results of the steam explosion optimization experiment are shown in Table 4 below.

[0068] Table 3 shows the optimization factors for steam explosion.

[0069]

[0070] Table 4 shows the steam explosion optimization experiment.

[0071]

[0072] The results of the orthogonal experiment are shown in Table 4 above. Within the orthogonal range of this experiment, there is a significant difference between the mean comprehensive evaluation values ​​of the two factors, steam explosion pressure and steam explosion time (p<0.05). Figure 4 The yield results of pea protein peptides are shown in the figure. The optimal process conditions for preparing pea peptides are orthogonal group 3, namely, steam explosion pressure of 1.0 MPa and steam explosion time of 40 s.

[0073] Based on the above experimental design and results, the optimal preparation conditions for pea peptides include: a material-to-liquid ratio of 1:29 w / v, pH = 9.45, ultrasonic time of 27 min, ultrasonic power of 260 W, steam explosion pressure of 1.0 MPa, and steam explosion time of 40 s.

[0074] 5. Determination of test results:

[0075] Pea peptides were prepared using the optimal process conditions determined in the above experiments, and the following performance tests were performed on the obtained steam-exploded pea protein peptides.

[0076] 5.1 Determination of absorbance value of free radical scavenging peptides from steam-exploded peas

[0077] 1) Determination of DPPH free radical scavenging capacity:

[0078] A 1:1 mixture of 0.1 mmol / L DPPH-anhydrous ethanol solution and sample solutions of steam-exploded pea peptides (5%, 10%, 15%, 20%) was prepared and reacted at room temperature in the dark for 30 min. The absorbance Ax was measured at 517 nm. A 1:1 mixture of sample and anhydrous ethanol was prepared, and the absorbance A0 was measured. Distilled water was used as a blank control and mixed with 0.1 mmol / L DPPH-anhydrous ethanol solution, and the absorbance A1 was measured. DPPH free radical scavenging rate = [1-(A...] x -A0) / A1]×100%.

[0079] Experimental results are as follows Figure 5 As shown in Figure A, the activity of steam-exploded pea peptides was higher than that of pea peptides at the same concentration, reaching its highest point at a concentration of 20%, with a DPPH free radical scavenging capacity of 92%.

[0080] 2) ABTS + Free radical scavenging ability determination:

[0081] 7 mmol / L ABTS + The stock solution and sample vaporized pea peptide (5%, 10%, 15%, 20%) solutions were mixed 1:1 and reacted at room temperature in the dark for 16 h. The absorbance A1 was measured at a wavelength of 734 nm. The absorbance A2 was measured in the sample solution and phosphate buffered saline (PBS). Distilled water and ABTS were also tested. + Absorbance A0 of free radical solution; ABTS + Free radical scavenging rate = [1 - (A1 - A2) / A0] × 100%.

[0082] Experimental results are as follows Figure 5 As shown in Figure B, the peptide activities of steam-exploded peas were all higher than those of pea protein peptides at the same concentration, reaching the highest point (ABTS) at a concentration of 20%. + The free radical scavenging capacity is 94%.

[0083] 3) O2 - Free radical scavenging ability determination:

[0084] The absorbance (A) of distilled water was measured at a wavelength of 530 nm; O2 - Absorbance A1 of free radical solution was measured; Fe was diluted by 1 volume. 2+ The solution and Tris-HCl buffer were mixed with the sample steam-exploded pea peptides (5%, 10%, 15%, 20%), and the absorbance A2 was measured after incubation for 5 min. - Free radical scavenging rate = [(A2-A1) / (A1-A0)] × 100%.

[0085] Experimental results are as follows Figure 5As shown in C, the peptide activities of steam-exploded peas were all higher than those of peas at the same concentration, reaching their highest point at a concentration of 20% (O2). - The free radical scavenging capacity is 98%. Therefore, the free radical scavenging capacity achieved by the steam-exploded pea peptide preparation technology provided by this invention is more significant.

[0086] 5.2 Determination of absorbance value of steam-exploded pea peptides for lowering blood pressure

[0087] 1) Assay for angiotensin-converting enzyme (ACE) inhibitory activity:

[0088] The absorbance (A) of the steam-exploded pea peptide samples (5%, 10%, 15%, 20%) was measured at 228 nm. Hip-His-Leu substrate was added to the samples, and the mixture was incubated in a 37°C water bath for 5 min. ACE was immediately added to initiate the reaction, and the mixture was kept at this temperature for 30 min. HCl was then added to terminate the reaction. Ethyl acetate was added, the mixture was mixed thoroughly, centrifuged, and dried in a 95°C oven for 30 min. The mixture was then redissolved in deionized water, and the absorbance (A1) was measured. The absorbance (A2) was measured using distilled water. The ACE activity inhibition rate was also measured.

[0089] =(A-A1)×100 / (A-A2)×100%.

[0090] Experimental results are as follows Figure 6 As shown in Figure A, the activity of steam-exploded pea peptides was higher than that of pea peptides at the same concentration, exhibiting the strongest ACE activity inhibition at a concentration of 15%, with an inhibition rate reaching 68%. Therefore, the ACE inhibition effect obtained by the steam-exploded pea peptide preparation technology provided by this invention is more significant.

[0091] 5.3 Determination of absorbance value of steam-exploded pea peptides for lowering blood glucose

[0092] 1) Determination of α-glucosidase inhibitory activity:

[0093] The absorbance (A) of the α-glucosidase solution was measured at 405 nm; the absorbance (A1) of distilled water was measured; for samples of steam-exploded pea peptides (5%, 10%, 15%, 20%), PBS buffer, inhibitor solution, and substrate were added sequentially, and the mixture was incubated at 37°C for 10 min. Then, the enzyme solution was added, and the mixture was thoroughly mixed and incubated at 37°C for 20 min. Finally, Na₂CO₃ solution was added to stop the reaction, and the absorbance (A2) was measured; the absorbance (A3) of samples of steam-exploded pea peptides (5%, 10%, 15%, 20%) was measured; the α-glucosidase inhibition rate =

[0094] [1-(A2-A3) / (A-A1)]×100%.

[0095] Experimental results are as follows Figure 6As shown in B, the activities of steam-exploded pea peptides were all higher than those of pea peptides at the same concentration, and at a concentration of 20%, they exhibited the strongest inhibitory effect on α-glucosidase activity, with an inhibition rate of up to 75%.

[0096] 2) Determination of α-amylase inhibitory activity:

[0097] The samples were steam-exploded pea peptides (5%, 10%, 15%, 20%) and α-amylase (2.0 mg / ml). -1 Dissolve ferulic acid (0-2.5 mg / ml) in a phosphate buffer solution with a pH of 6.9. -1 Mix thoroughly, dissolve in 30% ethanol, and incubate at 37°C for 10 min. Add fully cooked potato starch, incubate at 37°C for 20 min, then add 95% ethanol to inactivate the enzyme. After centrifugation at 4000 rpm for 10 min, collect the supernatant and mix with invertase (15.0 mg / ml). -1 Incubate at 37°C for 30 minutes. Calculate the inhibition rate using the following formula: α-amylase inhibition rate (%) = (1 - A0 / A) × 100% Where: A0 is the absorbance of the control group, and A is the absorbance of the sample group.

[0098] Experimental results are as follows Figure 6 As shown in Figure C, the activity of steam-exploded pea peptides was higher than that of pea peptides at the same concentration. At a concentration of 20%, it exhibited the strongest inhibitory effect on α-amylase activity, with an inhibition rate reaching 71%. Therefore, the steam-exploded pea peptide preparation technology provided by this invention can achieve a more significant inhibition rate of α-glucosidase and α-amylase activities.

[0099] In this invention, steam explosion treatment is used to assist enzymatic hydrolysis to obtain pea peptides, which improves the activity and function of the pea peptides, enabling them to achieve a DPPH free radical scavenging capacity of 92% and an ABTS content of [missing information]. + Free radical scavenging ability is 93%, O2 - The free radical scavenging capacity is 98%; the angiotensin-converting enzyme (ACE) inhibition rate can reach 68%; and the inhibition rates of α-glucosidase and α-amylase activities are 75% and 71%, respectively.

[0100] The above description is only used to illustrate the technical solution of the present invention and is not intended to limit it. Any other modifications or equivalent substitutions made by those skilled in the art to the technical solution of the present invention, as long as they do not depart from the spirit and scope of the technical solution of the present invention, should be covered within the scope of the claims of the present invention.

Claims

1. A method for preparing a multifunctional pea polypeptide, characterized in that: Includes the following steps: S1. Pea protein isolate extraction: Pea raw material is crushed and sieved and then dissolved in deionized water. The ratio of pea powder to water is 1:15~35w / v. Ultrasonic technology is used to assist dissolution. The ultrasonic power is 100~500w, the ultrasonic time is 10~30min, and the pH is 8~10. Then, the protein in the pea powder is extracted by alkaline dissolution and acid precipitation. The pea protein isolate powder is obtained by freeze drying. S2. Steam explosion treatment: The pea protein isolate obtained in step S1 above is added to the steam explosion cylinder of the steam explosion test bench for steam explosion treatment. The steam explosion time is 20~100s and the steam explosion pressure is 0.4~1.2MPa. The pea protein isolate is then freeze-dried to obtain steam-exploded pea protein isolate. S3. Preparation of pea peptides: Dissolve the steam-exploded pea protein isolate obtained in step S2 above in deionized water to prepare a substrate concentration of 10% and adjust the pH. Add alkaline protease and neutral protease respectively for enzymatic hydrolysis. Extract under ultrasonic-assisted extraction. After the reaction, inactivate the enzyme at high temperature, centrifuge at 6000 r / min for 15 min, remove the lower precipitate, collect the supernatant, and freeze-dry to obtain pea active peptide powder.

2. The preparation method according to claim 1, characterized in that: In step S1, the ratio of pea flour to water is 1:29w / v, the ultrasonic power is 260w, the ultrasonic time is 27min, and the pH value is 9.

45.

3. The preparation method according to claim 1, characterized in that: In step S2, the steam explosion time is 40s and the steam explosion pressure is 1.0MPa.

4. The preparation method according to claim 1, characterized in that: In step S1, the pea raw material is the protein-rich Zhongwan No. 9 pea, and the peas are ground into powder and passed through a 40-100 mesh sieve.

5. The preparation method according to claim 1, characterized in that: In step S1, the specific steps of the alkaline dissolution and acid precipitation method are as follows: ultrasonic treatment, adjusting the pH to 8-10 with 1.0 mol / L sodium hydroxide, stirring the sample solution continuously during the process, centrifuging to collect the supernatant, and then adjusting the pH to 2.5-5.5 with hydrochloric acid. The above steps are repeated 2-3 times, and the precipitate is washed with deionized water until neutral.

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