Method for improving salty taste and antioxidant activity of agaricus bisporus taste-active peptide
Through two-step enzymatic and ultrasonic synchronous enzymatic technology, the problem of mass transfer of substrates in enzymatic agaricus bisporus was solved, the salty taste and antioxidant activity of the taste peptide were improved, and healthy and delicious taste peptide powder production was achieved.
Patent Information
- Application Number
- CN202510678801.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-05-26
- Publication Date
- 2025-08-12
AI Technical Summary
In the existing Agaricus bisporus enzymatic technology, there are problems such as limited substrate mass transfer and insufficient enzyme-substrate effect, which leads to insufficient saltiness of the taste peptide and poor antioxidant activity.
Two-step enzymatic decomposition combined with ultrasonic synchronous enzymatic decomposition technology is used. First, neutral protease is used for enzymatic decomposition and synchronous sonication is used for enzymatic decomposition, and ultrasonic is introduced during each enzymatic decomposition process, followed by enzymatic decomposition, separation, concentration and drying to form a odor peptide powder.
It significantly improves the salt-enhancing ability and antioxidant activity of the Agaricus bisporus flavor peptide, enhances the delicious characteristics of the product, meets the needs of healthy diet, and improves the utilization rate of raw materials.
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Abstract
Description
Technical Field
[0001] The invention relates to the technical field of food processing, in particular to a method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide powder. Background Art
[0002] With the rise of awareness of healthy eating, people are gradually realizing the health problems caused by excessive salt intake. Based on this situation, the search for ways to "reduce salt without reducing saltiness" has received widespread attention around the world. Finding healthy sodium substitutes has become a research hotspot. Therefore, the development of flavor peptides with salty and umami flavors has great social significance.
[0003] The protein content of Agaricus bisporus, a dried form, can reach over 40%, containing all essential amino acids (including arginine and glutamic acid). It is also rich in soluble sugars and dietary fiber, and contains antioxidants such as ergothioneine and polyphenols. Currently, Agaricus bisporus is primarily consumed fresh or canned, and its high-quality protein resources have yet to be fully exploited. Developing flavor peptide seasonings using Agaricus bisporus as a natural ingredient would not only enrich the existing seasoning portfolio but also help replace some synthetic seasonings, achieving the health goals of reducing salt and sodium.
[0004] Traditional enzymatic hydrolysis technology for Agaricus bisporus often faces bottleneck problems such as limited substrate mass transfer and insufficient enzyme-substrate interaction, resulting in insufficient saltiness of the resulting flavor peptides and poor product activity. Summary of the Invention
[0005] Purpose of the invention: The present invention aims to provide a method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptides by ultrasound-synchronized enzymatic hydrolysis technology.
[0006] Technical solution: The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide powder of the present invention comprises the following steps:
[0007] S1 pretreatment: washing, slicing, drying and crushing the fresh Agaricus bisporus;
[0008] S2: First step of enzymatic hydrolysis: Agaricus bisporus powder is evenly mixed with water, and neutral protease is added for enzymatic hydrolysis, and ultrasonication is performed simultaneously during the enzymatic hydrolysis process;
[0009] S3 enzyme inactivation treatment;
[0010] S4 second step enzymatic hydrolysis: flavor protease is added for enzymatic hydrolysis, and ultrasonication is performed simultaneously during the enzymatic hydrolysis process;
[0011] S5 enzyme inactivation treatment;
[0012] S6 separation and purification removes water-insoluble substances;
[0013] S7 concentration;
[0014] S8 is dried and crushed to obtain flavor peptide powder.
[0015] Preferably, in step S1, the drying is performed until the moisture content is lower than 13%.
[0016] Preferably, in step S1, the pulverization is performed by passing through an 80-100 mesh sieve.
[0017] Preferably, in step S2, the enzymatic hydrolysis process is carried out synchronously by performing ultrasonic treatment in the first 5 to 15 minutes after the start of enzymatic hydrolysis, with an ultrasonic frequency of 20 kHz and an ultrasonic power of 200 to 350 W. In the first step of enzymatic hydrolysis, the neutral protease acts on the peptide bonds within the protein or polypeptide chain, cutting the long chain into smaller polypeptide fragments. The ultrasonic synchronous treatment helps promote substrate dispersion and the unfolding of the internal structure of the protein, thereby accelerating the enzymatic reaction.
[0018] Preferably, in step S2, the amount of neutral protease added is 0.8-1.0% of the mass of Agaricus bisporus powder. The amount of enzyme used affects the efficiency of protein hydrolysis, which can be expressed as a degree of hydrolysis index. If the amount is insufficient, the protein cannot be fully hydrolyzed, the degree of hydrolysis is low, and the content and variety of the resulting flavor peptides will be relatively small; if the amount of enzyme added is too high, it may cause excessive protein hydrolysis, releasing more small peptides and amino acids with a bitter taste, so it is necessary to control the amount of enzyme added appropriately.
[0019] Preferably, in step S2, the enzymatic hydrolysis conditions are: pH = 6-7, temperature 45-60° C., and time 3-5 hours. In the first enzymatic hydrolysis process, the Agaricus bisporus protein is preliminarily hydrolyzed into macromolecular peptides.
[0020] Preferably, in step S2, the solid-liquid ratio during the enzymatic hydrolysis process is 1:10 to 1:25.
[0021] Preferably, in step S4, the enzymatic hydrolysis process is performed simultaneously with ultrasonic treatment for the first 5 to 15 minutes after the start of enzymatic hydrolysis, with an ultrasonic frequency of 20 kHz and an ultrasonic power of 200 to 350 W. During the second enzymatic hydrolysis step, the flavor protease has both endo- and exo-enzyme activities, capable of hydrolyzing peptide bonds within proteins to generate small peptides and gradually cleaving amino acids from the ends of peptide chains to reduce the accumulation of bitter peptides. The simultaneous ultrasonic treatment can promote more thorough protein hydrolysis and generate more flavor peptides.
[0022] Preferably, in step S4, the amount of flavor protease added is 1.0-1.2% of the mass of Agaricus bisporus powder.
[0023] Preferably, in step S4, the enzymatic hydrolysis conditions are: pH = 6-7, temperature 45-60° C., and time 0.5-1.5 h.
[0024] Preferably, in step S3 or S5, the enzyme inactivation treatment is: inactivating the enzyme in a water bath after enzymolysis at a temperature of 80-100° C. for 5-10 minutes, followed by cooling after completion.
[0025] Step S6 removes the water-insoluble parts, including some insoluble fibers in the mushroom powder.
[0026] Preferably, in step S6, the separation and purification of water-insoluble substances primarily removes the water-insoluble portion, including some insoluble fibers in the mushroom powder. Preferably, the separation and purification of water-insoluble substances is performed by ultrafiltration after centrifugation, at a centrifugal speed of 4000-5000 rpm for 10-20 minutes, and ultrafiltration of the supernatant using an ultrafiltration membrane with a pore size of 0.002-0.003 μm.
[0027] Preferably, in step S7, the concentration is performed by vacuum low-temperature concentration at a concentration temperature of 40-50° C., and the concentration is performed until the solid content reaches 20-50%.
[0028] Preferably, in step S8, the drying is: pre-freezing the liquid obtained in step S7, and then performing vacuum freeze drying in a freeze dryer.
[0029] Invention Mechanism: This invention utilizes two-step enzymatic hydrolysis and ultrasonic synchronous enzymatic hydrolysis to ensure full contact between the enzyme and substrate, promoting the enzymatic reaction and significantly increasing the degree of hydrolysis of Agaricus bisporus protein and raw material utilization. Ultrasonic synchronous enzymatic hydrolysis technology, through the synergistic effect of cavitation and mechanical vibration, alters the protein's spatial conformation, promoting the production of more small-molecule peptides, enhancing the salt-enhancing capacity of flavor peptides, and improving flavor profile, resulting in a delicious, salt-reducing, and healthy experience.
[0030] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages: (1) The present invention improves the salt-enhancing ability and antioxidant activity of Agaricus bisporus flavor peptides through two-step enzymatic hydrolysis combined with simultaneous ultrasound during the two-step enzymatic hydrolysis process, and has the characteristics of being delicious, salt-reducing and healthy. The improvement of the antioxidant activity of the flavor peptides makes up for the defect of loss of biological activity in the current production of flavor peptides, is more in line with the needs of a healthy diet, and helps to improve the competitiveness of the product; (2) The introduction of ultrasound allows the enzyme and the substrate to fully contact, promotes the enzymatic hydrolysis reaction, significantly improves the hydrolysis degree of Agaricus bisporus protein, and increases the utilization rate of raw materials. BRIEF DESCRIPTION OF THE DRAWINGS
[0031] Figure 1 The bar graph of hydrolysis degree of Agaricus bisporus hydrolysate after different ultrasonic treatments;
[0032] Figure 2 The bar graph shows the peptide content of Agaricus bisporus hydrolysate treated with different ultrasound treatments;
[0033] Figure 3 This is the effect of different ultrasonic treatments on the scavenging rate of DPPH free radicals of Agaricus bisporus flavor peptide;
[0034] Figure 4 This is the effect of different ultrasonic treatments on the free radical scavenging rate of Agaricus bisporus flavor peptide ABTS;
[0035] Figure 5 This is a graph showing the effects of different ultrasonic treatments on the scavenging rate of hydroxyl radicals in Agaricus bisporus flavor peptides. DETAILED DESCRIPTION
[0036] The technical solution of the present invention will be further described below in conjunction with embodiments.
[0037] Example 1
[0038] The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptides of the present invention comprises the following steps:
[0039] S1 Pretreatment: Wash fresh Agaricus bisporus, slice and dry until the moisture content is less than 13%, grind and pass through an 80-mesh sieve;
[0040] S2: Enzymatic hydrolysis: Agaricus bisporus powder and water were uniformly mixed at a solid-liquid ratio of 1:25, and edible alkali was added to adjust the pH to 7.0. 0.9% neutral protease (based on the weight of mushroom powder) was added, and enzymatic hydrolysis was carried out at 50°C for 3.5 hours. Ultrasonic treatment was performed within the first 10 minutes after the start of enzymatic hydrolysis at a frequency of 20 kHz and a power of 300 W.
[0041] S3 enzyme inactivation treatment: After the first step of enzymatic hydrolysis, the enzyme is inactivated in a water bath at 90°C for 5 minutes, followed by cooling.
[0042] Step S4: Enzymatic hydrolysis: add edible alkali to adjust the pH to 7.0, add 1.1% flavor protease (based on the weight of mushroom powder), and perform enzymatic hydrolysis at 50°C for 1 hour. Ultrasonic treatment is performed simultaneously within the first 10 minutes after the start of enzymatic hydrolysis at a frequency of 20 kHz and a power of 300 W.
[0043] S5 enzyme inactivation treatment: After the second step of enzymatic hydrolysis, the enzyme is inactivated in a water bath at 90°C for 5 minutes, followed by cooling.
[0044] S6 separation and purification: including centrifugation and ultrafiltration, the centrifugal speed is 4000 rpm, the centrifugal time is 20 min, the supernatant is taken for ultrafiltration, and the pore size of the ultrafiltration membrane is 0.002 μm.
[0045] S7 vacuum concentration: vacuum low-temperature concentration is performed at 45°C until the solid content reaches 50%;
[0046] S8 vacuum freeze drying: pre-freeze the liquid obtained in step S7 at -80°C, and then perform vacuum freeze drying in a freeze dryer. After drying, crush it into a powder without lumps to obtain the flavor peptide powder.
[0047] Example 2
[0048] The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptides of the present invention comprises the following steps:
[0049] S1 Pretreatment: Wash fresh Agaricus bisporus, slice and dry until the moisture content is less than 13%, grind and pass through an 80-mesh sieve;
[0050] S2: Enzymatic hydrolysis: Agaricus bisporus powder and water were uniformly mixed at a solid-liquid ratio of 1:20, and edible alkali was added to adjust the pH to 7.0. 0.9% neutral protease (based on the weight of mushroom powder) was added, and enzymatic hydrolysis was carried out at 55°C for 3.5 hours. Ultrasonic treatment was performed simultaneously within the first 8 minutes after the start of enzymatic hydrolysis at an ultrasonic frequency of 18 kHz and a power of 300 W.
[0051] S3 enzyme inactivation treatment: After the first step of enzymatic hydrolysis, the enzyme is inactivated in a water bath at 90°C for 10 minutes, followed by cooling.
[0052] Step S4: Enzymatic hydrolysis: add edible alkali to adjust the pH to 7.0, add 1.1% flavor protease (based on the weight of mushroom powder), and perform enzymatic hydrolysis at 55°C for 1 hour. Ultrasonic treatment is performed simultaneously within the first 8 minutes after the start of enzymatic hydrolysis at a frequency of 18 kHz and a power of 300 W.
[0053] S5 enzyme inactivation treatment: After the second step of enzymatic hydrolysis, the enzyme is inactivated in a water bath at 90°C for 10 minutes, and then cooled;
[0054] S6 separation and purification: including centrifugation and ultrafiltration, the centrifugal speed is 4000 rpm, the centrifugal time is 15 min, the supernatant is taken for ultrafiltration, and the pore size of the ultrafiltration membrane is 0.002 μm.
[0055] S7 vacuum concentration: vacuum low-temperature concentration is performed at 50°C until the solid content reaches 50%;
[0056] S8 Vacuum freeze drying: pre-freeze the liquid obtained in (7) at -80°C, and then freeze dry it in a freeze dryer. After drying, crush it into a powder without lumps to obtain the flavor peptide powder.
[0057] Example 3
[0058] The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptides of the present invention comprises the following steps:
[0059] S1 Pretreatment: Wash fresh Agaricus bisporus, slice and dry until the moisture content is less than 13%, grind and pass through an 80-mesh sieve;
[0060] S2: Enzymatic hydrolysis: Agaricus bisporus powder and water were uniformly mixed at a solid-liquid ratio of 1:15, and edible alkali was added to adjust the pH to 7.0. 0.9% neutral protease (based on the weight of mushroom powder) was added, and enzymatic hydrolysis was carried out at 55°C for 4 hours. Ultrasonic treatment was performed simultaneously within the first 12 minutes after the start of enzymatic hydrolysis at an ultrasonic frequency of 18 kHz and a power of 300 W.
[0061] S3 enzyme inactivation treatment: After the first step of enzymatic hydrolysis, the enzyme is inactivated in a water bath at 90°C for 10 minutes, followed by cooling.
[0062] Step S4: Enzymatic hydrolysis: add edible alkali to adjust the pH to 7.0, add 1.1% flavor protease (based on the weight of mushroom powder), and perform enzymatic hydrolysis at 55°C for 1.5 hours. Ultrasonic treatment is performed simultaneously within the first 12 minutes after the start of enzymatic hydrolysis at a frequency of 20 kHz and a power of 200 W.
[0063] S5 enzyme inactivation treatment: After the second step of enzymatic hydrolysis, the enzyme is inactivated in a water bath at 90°C for 10 minutes, and then cooled;
[0064] S6 separation and purification: including centrifugation and ultrafiltration, the centrifugal speed is 4000 rpm, the centrifugal time is 20 min, the supernatant is taken for ultrafiltration, and the pore size of the ultrafiltration membrane is 0.002 μm.
[0065] S7 vacuum concentration: vacuum low-temperature concentration is performed at 40°C until the solid content reaches 50%;
[0066] S8 vacuum freeze drying: pre-freeze the liquid obtained in step S7 at -80°C, and then perform vacuum freeze drying in a freeze dryer. After drying, crush it into a powder without lumps to obtain the flavor peptide powder.
[0067] Comparative Example 1
[0068] The difference between Comparative Example 1 and Example 1 is that steps S2 and S4 are not subjected to ultrasonic treatment, and the remaining steps are the same as those in Example 1.
[0069] Comparative Example 2
[0070] The difference between Comparative Example 2 and Example 1 is that, in step S2, Agaricus bisporus powder and water are uniformly mixed at a solid-liquid ratio of 1:25, edible alkali is added to adjust the pH to 7.0, ultrasonic treatment is first performed for 20 minutes at a frequency of 20 kHz and a power of 300 W, and then 0.9% neutral protease (based on the mass of mushroom powder) is added and enzymatic hydrolysis is carried out at 50° C. for 3.5 hours. In step S4, ultrasonic treatment is not performed, and the remaining steps are the same as in Example 1.
[0071] Comparative Example 3
[0072] The difference between Comparative Example 3 and Example 1 is that, in step S2, Agaricus bisporus powder and water are uniformly mixed at a solid-liquid ratio of 1:25, edible alkali is added to adjust the pH to 7.0, 0.9% neutral protease (based on the mass of mushroom powder) is added, and enzymatic hydrolysis is carried out at 50° C. for 3.5 hours. Ultrasonic treatment is simultaneously performed within the first 20 minutes after the start of enzymatic hydrolysis, with an ultrasonic frequency of 20 kHz and a power of 300 W. Ultrasonic treatment is not performed in step S4, and the remaining steps are the same as in Example 1.
[0073] Comparative Example 4
[0074] The difference between Comparative Example 4 and Example 1 is that ultrasonic treatment is not performed in step S2, and step S4 is to add edible alkali to adjust the pH to 7.0, add 1.1% flavor protease (based on the mass of mushroom powder), and enzymatic hydrolysis at 50° C. for 1 hour. Ultrasonic treatment is simultaneously performed within the first 20 minutes after the start of enzymatic hydrolysis, with an ultrasonic frequency of 20 kHz and a power of 300 W. The remaining steps are the same as in Example 1.
[0075] Performance Testing
[0076] 1. Effects of different ultrasonic treatments on the hydrolysis degree and total soluble solids (TSS) of Agaricus bisporus hydrolysate
[0077] Examples 1 to 3 and Comparative Examples 1 to 4 were selected to determine the effects of different ultrasonic methods on the degree of hydrolysis of the enzymatic hydrolyzate. The specific method was as follows: using formaldehyde titration, 5% of the total weight of the enzymatic hydrolysis supernatant was taken, 60 mL of pure water was added, the pH was adjusted to 8.2 with a 0.05 mol / L sodium hydroxide standard solution, 10 mL of formaldehyde was added, and the pH was adjusted to 9.2 with a 0.05 mol / L sodium hydroxide standard solution. The difference in sodium hydroxide consumption between the two titrations was recorded as V1. Water was used as a blank control, and the sodium hydroxide consumption V2 was recorded. The degree of hydrolysis was calculated by the following formula:
[0078]
[0079] Where:
[0080] DH——degree of hydrolysis, unit %;
[0081] C is the molar concentration of sodium hydroxide used for titration, in mol / L;
[0082] m——mass of raw materials, unit: g;
[0083] a——The percentage of protein in the raw material (dry basis), unit %.
[0084] Total soluble solids were read using a handheld refractometer.
[0085] The test results are as follows Figure 1 shown.
[0086] from Figure 1 It can be seen that the hydrolysis degrees of Agaricus bisporus protein in Examples 1-3 and Comparative Examples 1-4 were 45.32%, 44.73%, 45.81%, 36.46%, 41.27%, 43.20%, and 43.78%, respectively. Compared with Comparative Example 1 (without ultrasound treatment), ultrasound treatment significantly increased the hydrolysis degree of Agaricus bisporus protein, with Example 3 having the highest hydrolysis degree, followed by Example 1, which increased by 29.19% and 24.30%, respectively, compared to Comparative Example 1 (without ultrasound treatment). The total soluble solids content showed a trend consistent with the hydrolysis degree. This test shows that ultrasound-synchronized enzymatic hydrolysis significantly promotes protein hydrolysis efficiency.
[0087] 2. Effects of different ultrasonic treatments on the peptide content of Agaricus bisporus hydrolysate
[0088] Examples 1 to 3 and comparative examples 1 to 4 were selected to determine the effects of different ultrasonic methods on the peptide content of the hydrolyzate. The specific method is as follows: the hydrolyzate was mixed with trichloroacetic acid solution (10%, w / v) at a ratio of 1:1 (v / v). After standing for 30 minutes, centrifuged at 4000r / min for 10 minutes, and the supernatant was taken to determine the peptide content using the biuret method. 1mL of the diluted sample was thoroughly shaken with 4mL of the biuret reagent, and the mixture was placed in a water bath at 25°C for 30 minutes. The absorbance at 540nm was measured, and bovine serum albumin was used as a standard curve. The test results are as follows: Figure 2 shown.
[0089] from Figure 2 As can be seen, the peptide contents of the Agaricus bisporus hydrolysates of Examples 1-3 and Comparative Examples 1-4 were 7.40 mg / mL, 7.13 mg / mL, 7.41 mg / mL, 5.01 mg / mL, 5.35 mg / mL, 7.04 mg / mL, and 6.96 mg / mL, respectively. Compared with Comparative Example 1 (without ultrasound), ultrasound treatment further promoted protein hydrolysis and peptide release. The peptide contents of Examples 1 and 3 were significantly higher than those of the other treatments, increasing by 47.70% and 47.90%, respectively, compared to Comparative Example 1 (without ultrasound).
[0090] 3. Effects of different ultrasound methods on the saltiness intensity and salt-enhancing effect of flavor peptides
[0091] Examples 1 to 3 and Comparative Examples 1 to 4 were selected to compare the effects of different ultrasonic methods on the saltiness intensity and salt-enhancing effect of flavor peptides. The sensory evaluation panel consisted of 20 sensory evaluators aged 20 to 30 (10 males and 10 females). The specific evaluation method is as follows:
[0092] (1) Evaluation of saltiness intensity
[0093] A 0.35% NaCl standard solution and a 1% (w / v) sample solution were prepared. A 10-point scoring system was used, with 0 indicating no salty taste and 10 indicating a strong salty taste. The standard solution was assigned a score of 5. Sensory evaluation was conducted at room temperature (20-25°C). Each sample was in contact with the mouth for 10 seconds and then expelled. Between samples, the mouth was rinsed with pure water or a biscuit to remove any residual taste. At least 30 seconds were allowed between samples.
[0094] (2) Evaluation of saltiness-enhancing effect
[0095] A 0.35% (w / v) salt solution was used as the standard solution, and samples were added at a 1% concentration to determine their saltiness. A 10-point scoring system was used, with 0 indicating no saltiness and 10 indicating the most pronounced saltiness. The standard solution was assigned a score of 5.
[0096] Table 1 Evaluation results of saltiness intensity and saltiness enhancement effect of Agaricus bisporus flavor peptides treated with different ultrasound
[0097] Serial number Saltiness intensity (points) Saltiness effect (points) Example 1 4.86 7.94 Example 2 4.74 7.42 Example 3 4.80 7.58 Comparative Example 1 3.36 6.64 Comparative Example 2 4.40 6.46 Comparative Example 3 4.57 6.14 Comparative Example 4 4.73 7.36
[0098] As shown in Table 1, the saltiness intensity of Examples 1-3 and Comparative Examples 2-4, which underwent ultrasonic treatment, increased to varying degrees compared to Comparative Example 1 (non-ultrasonicated group). Example 1, which employed simultaneous ultrasonic treatment for both enzymatic hydrolysis steps, had the highest saltiness intensity. In terms of saltiness enhancement, Example 1 achieved the best saltiness enhancement effect in a 0.35% salt solution, increasing saltiness by 19.58% compared to Comparative Example 1. This result demonstrates that the introduction of simultaneous ultrasonic treatment during both enzymatic hydrolysis steps is the most effective in enhancing the saltiness and saltiness enhancement of flavor peptides.
[0099] 4. Effect of Synchronous Ultrasonic Enzymatic Hydrolysis on the Antioxidant Activity of Flavor Peptides
[0100] Examples 1 to 3 and comparative examples 1 to 4 were selected to compare the effects of ultrasonic synchronous enzymatic hydrolysis on the antioxidant activity of flavor peptides. The specific method is as follows:
[0101] (1) DPPH free radical scavenging rate
[0102] Prepare a 3 mg / mL sample solution in distilled water. Mix 0.5 mL of the sample solution with 0.5 mL of DPPH solution (0.2 mmol / L, dissolved in anhydrous ethanol). Incubate at room temperature in the dark for 30 minutes, and then measure the absorbance of the mixture at 517 nm. Ethanol serves as a blank control, and glutathione serves as a positive control. The DPPH free radical scavenging rate is calculated as follows:
[0103]
[0104] Where:
[0105] P——DPPH free radical scavenging rate;
[0106] As——absorbance of the mixture of the test solution and DPPH solution;
[0107] Ac——absorbance of the mixture of the test solution and anhydrous ethanol;
[0108] Ab——Absorbance of the mixture of DPPH solution and sample solvent.
[0109] The test results are as follows Figure 3 shown.
[0110] (2) ABTS free radical scavenging rate
[0111] Mix the 7.4 mmol / L ABTS stock solution with an equal volume of 2.45 mmol / L potassium persulfate and incubate in the dark at room temperature for 12-16 hours. Before use, dilute the above ABTS solution with phosphate buffer solution (5.0 mmol / L, pH 7.4) to an absorbance of 0.7000 ± 0.020 at 734 nm. Then, add 200 μL of the diluted ABTS solution to 10 μL of the sample with a concentration of 3 mg / mL, incubate at room temperature for 10 minutes, and then measure the absorbance at 734 nm. Use the sample solvent instead of the sample as a blank control, and glutathione as a positive control. The ABTS free radical scavenging rate is calculated as follows:
[0112]
[0113] Where:
[0114] X——ABTS free radical scavenging rate;
[0115] Ac——absorbance of the mixture of ABTS solution and sample solvent;
[0116] As——Absorbance of the mixture of the test solution and ABTS solution.
[0117] The test results are as follows Figure 4 shown.
[0118] (3) Hydroxyl radical scavenging rate
[0119] Prepare a 3.0 mg / mL sample solution with distilled water, a 6.0 mmol / L FeSO4 solution, and a 6.0 mmol / L salicylic acid-ethanol solution. Add 0.5 mL of the prepared FeSO4 solution, 0.5 mL of the salicylic acid-ethanol solution, and 0.5 mL of the sample solution in that order. Finally, add 0.5 mL of a 0.01% H2O2 solution to initiate the reaction. Incubate the mixture at 37°C for 30 minutes, then measure the absorbance of 200 μL of the mixed solution at 510 nm. The hydroxyl radical scavenging rate is calculated as follows:
[0120]
[0121] Where:
[0122] Y——hydroxyl radical scavenging rate;
[0123] As——absorbance of sample solution mixture;
[0124] Ac - absorbance of solution without H2O2 (replaced with an equal amount of water);
[0125] Ab – absorbance of solution without sample (replaced with an equal amount of water).
[0126] The test results are as follows Figure 5 shown.
[0127] Depend on Figures 3-5 It can be seen that ultrasonic treatment further improves the antioxidant activity of the enzymatic hydrolysate. When the concentration is 3 mg / mL, the DPPH free radical scavenging rate, ABTS free radical scavenging rate, and hydroxyl free radical scavenging rate of Example 1 are 78.81%, 58.49%, and 55.74%, respectively, which are increased by 15.09%, 32.36%, and 33.70% respectively compared with Comparative Example 1. This shows that the process significantly enhances the antioxidant activity of flavor peptides, which helps to slow down the fat oxidation and deterioration process in food, thereby extending the shelf life of food and maintaining its flavor and nutrition.
[0128] The above technical effects verify the comprehensive advantages of the technical solution of the present invention in enhancing the flavor characteristics and functional activity of flavor peptides, and provide technical support for the development of natural food additives with both flavor enhancement and health properties.
Claims
1. A method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide, characterized in that: The following steps are involved: S1 pretreatment: washing, slicing, drying, and crushing fresh Agaricus bisporus to obtain Agaricus bisporus powder; S2: First step of enzymatic hydrolysis: Agaricus bisporus powder is evenly mixed with water, and neutral protease is added for enzymatic hydrolysis, and ultrasonication is performed simultaneously during the enzymatic hydrolysis process; S3 enzyme inactivation treatment; S4 second step enzymatic hydrolysis: flavor protease is added for enzymatic hydrolysis, and ultrasonication is performed simultaneously during the enzymatic hydrolysis process; S5 enzyme inactivation treatment; S6 separation and purification removes water-insoluble substances; S7 concentration; S8 is dried and crushed to obtain flavor peptide powder.
2. The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide according to claim 1, characterized in that: In steps S2 and S4, the ultrasonic treatment is performed synchronously during the enzymatic hydrolysis process, that is, the ultrasonic treatment is performed synchronously within the first 5 to 15 minutes after the start of the enzymatic hydrolysis.
3. The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide according to claim 2, characterized in that: The frequency of the ultrasound is 18-20 kHz, and the power of the ultrasound is 200-350 W.
4. The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide according to claim 1, characterized in that: In step S4, the amount of flavor protease added is 1.0-1.2% of the mass of Agaricus bisporus powder.
5. The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide according to claim 1, characterized in that: In step S4, the enzymatic hydrolysis conditions are: pH = 6-7, temperature 45-60° C., and time 0.5-1.5 h.
6. The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide according to claim 1, characterized in that: In step S2, the amount of the neutral protease added is 0.8-1.0% of the mass of the Agaricus bisporus powder.
7. The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide according to claim 1, characterized in that: In step S2, the enzymatic hydrolysis conditions are: pH = 6-7, temperature 45-60° C., and time 3-5 h.
8. The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide according to claim 1, characterized in that: In steps S3 and S5, the enzyme inactivation treatment is: inactivating the enzyme in a water bath after enzymolysis at a temperature of 80-100°C.
9. The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide according to claim 1, characterized in that: In step S6, the method for separating and refining the water-insoluble substances is: centrifugation followed by ultrafiltration, the centrifugal speed is 4000-5000 rpm, the centrifugal time is 10-20 min, and the supernatant is ultrafiltered, the pore size of the ultrafiltration membrane is 0.002-0.003 μm.
10. The method for improving the saltiness and antioxidant activity of Agaricus bisporus flavor peptide according to claim 1, characterized in that: In step S7, the concentration is performed by vacuum low-temperature concentration at a concentration temperature of 40-50° C., and the concentration is performed until the solid content reaches 20-50%.