ACE inhibitory peptide, formula goat milk powder for middle-aged and elderly people and preparation method of formula goat milk powder

Abstract

The invention provides ACE (Angiotensin Converting Enzyme) inhibitory peptide, middle-aged and elderly formula goat milk powder and a preparation method thereof, goat milk is taken as a raw material, the goat milk is sterilized and cooled, compound protease consisting of three proteases is inoculated for constant-temperature enzymolysis, and the ACE inhibitory peptide is obtained through ultrafiltration, preparative chromatographic separation and freeze drying. And uniformly mixing the active peptide with a nutrition enhancer and goat milk powder required by the middle-aged and elderly people to obtain the active peptide goat milk powder for the middle-aged and elderly people. Or after the goat milk is subjected to enzymolysis by compound protease, directly adding a nutrition enhancer required by the middle-aged and elderly people, and then concentrating and spray-drying to obtain the formula goat milk powder for the middle-aged and elderly people. The ACE inhibitory peptide prepared by the invention can be used as a food functional factor, and the prepared formula goat milk powder for middle-aged and elderly people contains the ACE inhibitory peptide and has the function of assisting in lowering blood pressure.

Description

Technical Field

[0001] The invention belongs to the technical field of enzyme engineering, and in particular to an ACE inhibitory peptide, middle-aged and elderly formula goat milk powder and a preparation method thereof. Background Art

[0002] Hypertension is one of the most common chronic diseases among middle-aged and elderly people. Among the middle-aged and elderly population, the proportion of people suffering from hypertension is higher than that of the overall adult population, and with the acceleration of the aging process of the population, this proportion is still increasing year by year. For middle-aged and elderly patients with hypertension, drug therapy is a common means of control, mainly including ACE inhibitors, calcium channel blockers, diuretics, beta-blockers, etc. Although these drugs are effective in controlling blood pressure, they are also accompanied by a series of side effects, such as dry cough, hypotension, renal damage, hyperkalemia, tachycardia, and abnormal glucose metabolism, which affect the quality of life of patients to a certain extent.

[0003] In recent years, a more natural, safe and effective way to lower blood pressure has gradually attracted the attention of the medical and nutritional communities, that is, foods containing ACE inhibitory peptides. ACE inhibitory peptides are a class of peptides with special biological activity. They can inhibit the activity of angiotensin converting enzyme (ACE), thereby blocking the production of angiotensin II and achieving the effect of lowering blood pressure. Unlike chemically synthesized drugs, ACE inhibitory peptides are derived from the hydrolyzate of natural food proteins, so they have no toxic side effects and are very suitable as part of the daily diet of middle-aged and elderly people to prevent and relieve hypertension.

[0004] Studies have shown that long-term intake of foods containing ACE inhibitory peptides can not only effectively lower the blood pressure level of hypertensive patients, but also help improve cardiovascular health, providing new ideas for the treatment of hypertensive patients. In order to better apply ACE inhibitory peptides to daily diet, researchers have tried to add them to common foods such as milk powder and dairy products, which not only facilitates the daily consumption of middle-aged and elderly people, but also improves their compliance with healthy foods.

[0005] However, there are still some problems in the preparation process of ACE inhibitory peptides, such as fewer hydrolysis sites of single-enzyme or double-enzyme enzymatic protein, lower content of generated ACE inhibitory peptides, and low ACE inhibition rate of the final product, which affects its antihypertensive function. Therefore, how to improve the ACE inhibition rate of enzymatic hydrolysate has become a hot spot and difficulty in current research. Summary of the invention

[0006] In view of the above-mentioned existing problems, the present invention provides an ACE inhibitory peptide, a middle-aged and elderly formula goat milk powder and a preparation method thereof. The ACE inhibitory peptide is obtained by multi-enzyme synergistic enzymolysis of goat milk. The ACE inhibitory peptide can provide food functional factors for the production of antihypertensive peptide foods. The middle-aged and elderly formula goat milk powder with added ACE inhibitory peptide can directly provide ACE inhibitory peptide to consumers through consumption, which can not only meet the needs of middle-aged and elderly people for hypertension control, but also reduce costs through simplified production processes, thereby providing the market with more competitive functional goat milk powder products.

[0007] To achieve the above object, the present invention provides the following scheme: an ACE inhibitory peptide, the amino acid sequence of the ACE inhibitory peptide is shown in SEQ ID NO: 1 to SEQ ID NO: 16.

[0008] Furthermore, the ACE inhibitory peptide is derived from β-casein, αs1-casein, lactoferrin, β-lactoglobulin, α-lactalbumin, αs2-casein and κ-casein of goat milk.

[0009] Furthermore, the ACE inhibitory activity IC 50 It is 0.15±0.013 mg / mL.

[0010] The present invention also provides a method for preparing an ACE inhibitory peptide, the specific steps of which are as follows: In terms of mass fraction, 2.35‰ to 4‰ of the composite protease is added to goat milk or reconstituted goat milk; Perform enzymolysis at a constant temperature of 45°C to 60°C for 2.4h to 4.0h, inactivate the enzyme, centrifuge and take the supernatant to obtain goat milk enzymolysis solution, which contains ACE inhibitory peptides; The enzymatic hydrolysate with a molecular weight Mw <1 kDa was taken, and the collected solution from 30min to 39min of the preparative chromatography was collected and freeze-dried to obtain an ACE inhibitory peptide.

[0011] Furthermore, the composite protease is protease X, protease C and protease N, the addition ratio of protease X is 10%~53%, the addition ratio of protease C is 10%~45%, and the addition ratio of protease N is 34%~80%, wherein protease X is a high enzyme activity concentrated neutral protease with an enzyme activity of 150,000 U / g; protease C is protease Protamex 1.6 with an enzyme activity of 1.6 AU-N / g; protease N is a neutral protease with an enzyme activity of 0.8AU / g.

[0012] Furthermore, the ACE inhibition rate of the goat milk enzymatic hydrolysate is 90% to 96%.

[0013] The present invention also provides a middle-aged and elderly formula goat milk powder, with a total amount of 1kg, and the raw materials include 0.3g~0.6g of the above-mentioned ACE inhibitory peptide, 3g~8g of goat colostrum powder, 0.50g~1.0g of yeast β-glucan, 0.2g~0.5g of phosphatidylserine, 0.5g~1.0g of probiotic powder, 20g~50g of dietary fiber, and the rest is goat milk powder; The above raw materials are mixed to obtain the middle-aged and elderly formula goat milk powder.

[0014] Furthermore, the probiotics include one or more of Lactobacillus acidophilus, Bifidobacterium animalis subspecies lactis, Lactobacillus plantarum, Lactobacillus paracasei, and Lactobacillus rhamnosus, and the number of viable bacteria is ≥1.0×10 11 CFU / g; The dietary fiber is one or more of polydextran, isomaltooligosaccharide and inulin.

[0015] The present invention also provides a middle-aged and elderly formula goat milk powder, the raw materials of which include goat milk enzymatic hydrolysate, Acer truncatum seed oil, N-acetylneuraminic acid, phosphatidylserine and dietary fiber, the raw materials are mixed, concentrated to a solid content of 45% to 52%, and then spray-dried to obtain the middle-aged and elderly formula goat milk powder; The goat milk enzymolysate is the above-mentioned goat milk enzymolysate.

[0016] Furthermore, the goat milk enzymatic hydrolysate is added with 0.0125%~0.25% (w / v) Acer truncatum seed oil, 0.0125%~0.20% (w / v) N-acetylneuraminic acid, 0.0125%~0.25% (w / v) phosphatidylserine, and 0.25%~0.50% (w / v) dietary fiber; The dietary fiber is one or more of polydextran, isomaltooligosaccharide and inulin.

[0017] The present invention discloses the following technical effects: The present invention provides a goat milk-derived ACE inhibitory peptide. The amino acid sequence of the prepared ACE inhibitory peptide is determined by separation, purification and liquid chromatography-mass spectrometry to ensure its functionality and consistency, thereby providing an ACE inhibitory peptide functional factor with a clear composition for the research and development of antihypertensive peptide foods.

[0018] The present invention provides a method for preparing ACE inhibitory peptides by multi-enzyme synergistic enzymolysis of goat milk. The method increases the hydrolysis sites of milk protein through the synergistic action of multiple enzymes, significantly improves the ACE inhibition rate in goat milk enzymolysate, and overcomes the disadvantage of low efficiency of single-enzyme or double-enzyme enzymolysis in the prior art.

[0019] The present invention also provides a method for preparing goat milk powder with formula for middle-aged and elderly people, which directly utilizes enzymatically hydrolyzed goat milk containing ACE inhibitory peptides and adds nutritional enhancers required by middle-aged and elderly people to produce functional formula milk powder for middle-aged and elderly people, thereby eliminating traditional separation and purification steps, reducing production costs, simplifying the process flow, retaining all ACE inhibitory peptides, and avoiding the loss of ACE inhibitory peptides caused by separation and purification. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 Effects of different proteases on ACE inhibition rate and hydrolysis degree of goat milk; Figure 2 Effects of protease combinations on ACE inhibition rate and hydrolysis degree of goat milk; Figure 3 Response surface diagram of the effect of enzymatic hydrolysis temperature and time on the ACE inhibition rate of goat milk; Figure 4 Response surface diagram of the effects of enzymatic hydrolysis temperature and enzyme addition amount on the ACE inhibition rate of goat milk; Figure 5 Response surface diagram of the effects of enzymatic hydrolysis time and enzyme addition amount on ACE inhibition rate of goat milk; Figure 6 Preparative chromatogram of the fraction less than 1K from ultrafiltrated goat milk; Figure 7 Secondary mass spectrum of ACE inhibitory peptide FPEVF; Figure 8 MS / MS of ACE inhibitory peptide FVVAPFPE; Fig. 9 MS / MS spectrum of ACE inhibitory peptide LNFL; Fig.10 MS / MS spectrum of ACE inhibitory peptide YVRYL; Fig.11 Secondary mass spectrum of ACE inhibitory peptide INYW; Fig.12 MS / MS of ACE inhibitory peptide LFQIN; Fig.13 Secondary mass spectrum of ACE inhibitory peptide ELNVVGET; Fig.14 MS / MS spectrum of ACE inhibitory peptide IMGVPK; Fig.15 MS / MS spectrum of ACE inhibitory peptide LHLPLPLV; Fig.16 MS / MS spectrum of ACE inhibitory peptide LQPEIM; Fig.17 Secondary mass spectrum of ACE inhibitory peptide MPIQAF; Fig.18Secondary mass spectrum of ACE inhibitory peptide VMFPPQS; Fig.19 MS / MS spectrum of ACE inhibitory peptide ILLQK; Fig. 20 MS / MS spectrum of ACE inhibitory peptide QFLPYP; Fig.21 MS / MS spectrum of ACE inhibitory peptide YIPIQY; Fig. 22 MS / MS spectrum of ACE inhibitory peptide FVRIPS. DETAILED DESCRIPTION

[0021] In order to further understand the present invention, the present invention is described below in conjunction with the accompanying drawings and embodiments. These descriptions are only to further explain the features and advantages of the present invention and are not intended to limit the claims of the present invention.

[0022] The present invention provides an ACE inhibitory peptide, wherein goat milk contains β-casein, α s1 -Casein, lactoferrin, β-lactoglobulin, α-lactalbumin, α s2 -casein and κ-casein, the above proteins in goat milk were hydrolyzed by composite protease, and then ultrafiltration, preparative chromatography separation and freeze-drying were used to obtain freeze-dried peptide powder. The ACE inhibitory activity IC 50 It is 0.15±0.013 mg / mL.

[0023] The ACE inhibitory peptide powder contains 16 peptides, and the amino acid sequences of the 16 peptides are shown in SEQ ID NO: 1 to SEQ ID NO: 16, specifically: FPEVF, FVVAPFPE, LNFL, YVRYL, INYW, LFQIN, ELNVVGET, IMGVPK, LHLPLPLV, LQPEIM, MPIQAF, VMFPPQS, ILLQK, QFLPYP, YIPIQY, FVRIPS; The present invention also provides a method for preparing an ACE inhibitory peptide, the specific steps of which are as follows: S1: Add 2.35‰~4‰ of compound protease to sterilized and cooled goat milk; S2 is enzymatically hydrolyzed at 45°C to 60°C for 2.4 to 4 hours, the enzyme is inactivated, and the supernatant is obtained by centrifugation, which is the goat milk enzymatic hydrolyzate containing ACE inhibitory peptides; S3 ultrafilters the enzymatic hydrolysate, and takes the enzymatic hydrolysate with a molecular weight Mw <1k Da, and further collects the component III with a high ACE inhibition rate through preparative chromatography, and freeze-dries it to obtain the ACE inhibitory peptide.

[0024] The composite protease is protease X, protease C and protease N, the addition ratio of protease X is 10%-53%, the addition ratio of protease C is 10%-45%, and the addition ratio of protease N is 34%-80%, wherein protease X is a highly active concentrated neutral protease with an enzyme activity of 150,000 U / g; protease C is protease Protamex 1.6 with an enzyme activity of 1.6 AU-N / g; protease N is a neutral protease with an enzyme activity of 0.8AU / g; The ACE inhibition rate of the goat milk enzymatic hydrolysate is 90% to 96%.

[0025] A goat milk powder formulated for middle-aged and elderly people, with a total weight of 1kg, comprising 0.3g-0.6g of the above-mentioned ACE inhibitory peptide powder, 3g-8g of goat colostrum powder, 0.50g-1.0g of yeast β-glucan, 0.2g-0.5g of phosphatidylserine, 0.5g-1.0g of probiotic powder, 20g-50g of dietary fiber, and the rest being goat milk powder.

[0026] The number of viable bacteria of one or more of Lactobacillus acidophilus, Bifidobacterium animalis subsp. lactis, Lactobacillus plantarum, Lactobacillus paracasei, and Lactobacillus rhamnosus is ≥ 1.0×10 11 CFU / g; The dietary fiber is one or more of polyglucan, isomaltooligosaccharide and inulin; The present invention also provides a wet method for preparing goat milk powder for middle-aged and elderly people, and the specific steps are as follows: S1: In terms of mass fraction, add 2.35‰~4‰ composite protease to goat milk or reconstituted goat milk, perform enzymolysis at 45℃~60℃ for 2.4 h~4 h, inactivate the enzyme, centrifuge and take the supernatant as goat milk enzymolysate; S2 is prepared by adding 0.0125%~0.25% (w / v) Acer truncatum seed oil, 0.0125%~0.20% (w / v) N-acetylneuraminic acid, 0.0125%~0.25% (w / v) phosphatidylserine, and 0.25%~0.50% (w / v) dietary fiber to goat milk enzymatic hydrolysate according to the needs of the middle-aged and elderly people. The solution is then concentrated to a solid content of 45%~52% and spray-dried to obtain goat milk powder formulated for the middle-aged and elderly.

[0027] Preferably, the spray drying conditions are: an inlet air temperature of 170°C to 190°C, an outlet air temperature of 100°C to 110°C, and a liquid flow rate of 6ml / min to 8ml / min.

[0028] Preferably, the dietary fiber is one or more of polyglucan, isomaltooligosaccharide and inulin; Obtaining the optimal parameters of the above-mentioned method for preparing a bioactive peptide powder: 1. Effects of different proteases on ACE inhibition rate and hydrolysis degree of goat milk Bromelain (A), protease P (B), protease X (C), papain (D), alkaline protease (E), composite protease (F), flavor protease (G) and protease N (H) in Table 1 were added to the enzyme-inactivated and cooled goat milk at an addition amount of 2‰ (w / v), and the milk was enzymatically hydrolyzed at 50°C for 3 h. After boiling and inactivating the enzyme, the samples were centrifuged and the ACE inhibition rate and hydrolysis degree were determined. The results are shown in Figure 2. Figure 1 As shown: Table 1 Sources and optimal conditions of different proteases

[0029] Depend on Figure 1 It can be seen that there are great differences in ACE inhibition rate and hydrolysis degree of goat milk after hydrolysis by different proteases. The ACE inhibition rate ranges from 41.53% to 74.62%, and the ACE inhibition rate of proteases from high to low is ranked as H (protease N) > D (protease P) > F (protease C) > B (protease X) > A (bromelain) > C (papain) > E (alkaline protease) > G (flavor protease); the hydrolysis degree ranges from 1.46% to 16.88%, and the hydrolysis degree of proteases from high to low is ranked as E (alkaline protease) > F (protease C) > C (papain) > B (protease X) > H (protease N) > D (protease P) > A (bromelain) > G (flavor protease). There was no positive correlation between the ACE inhibition rate and the degree of protein hydrolysis of goat milk hydrolysates of 8 proteases. The hydrolysis degree of E (alkaline protease) was the highest, but its ACE inhibition rate ranked second to last. The hydrolysis degree of D (protease P) was not high, but its ACE inhibition rate ranked second. This shows that different proteases have different hydrolysis sites on milk protein, and the hydrolysis sites of a single protease are limited. Therefore, 4 proteases with ACE inhibition rates higher than 70% - H (protease N), D (protease P), F (protease C) and B (protease X) were selected in combination to improve the ACE inhibition rate of goat milk hydrolysate.

[0030] 2. Effect of protease combination on ACE inhibition rate and hydrolysis degree of goat milk enzymatic hydrolysate H (Protease N), D (Protease P), F (Protease C) and B (Protease X) were hydrolyzed with two enzymes, three enzymes and four enzymes. The total addition amount was 2‰ (w / v). When two enzymes were compounded, each accounted for 1‰ (w / v), when three enzymes were compounded, each accounted for 1 / 3, and when four enzymes were compounded, each accounted for 0.5‰ (w / v). The enzymes were hydrolyzed at 50℃ for 3h, and then the enzymes were boiled and inactivated, and the samples were centrifuged to determine their ACE inhibition rate and hydrolysis degree. The results are as follows Figure 2 shown.

[0031] Depend on Figure 2It can be seen that the hydrolysis degree of goat milk hydrolyzed by the four protease combinations ranged from 7.26% to 11.15%, and the ACE inhibition rate of goat milk hydrolyzed by the four protease combinations ranged from 64.3% to 81.33%. The protease combinations were ranked from high to low in terms of ACE inhibition rate as BFH>FH>BH>BDFH>DF>BF>>DFH>BD>BDF>BDH, that is, the combinations with the highest ACE inhibition rate were protease X (B), protease C (F) and protease N (H), whose ACE inhibition rates were significantly higher than those of other protease combinations (p<0.05), and also significantly higher than those of single enzyme hydrolysis, indicating that a suitable protease combination can improve the ACE inhibition rate of goat milk, but the more enzyme types, the better. Too many enzyme types may further hydrolyze active ACE inhibitory peptides into inactive peptides. The protease BFH combination was selected for further testing. 3. Effect of protease ratio on ACE inhibition rate of goat milk enzymatic hydrolysate Protease X (B), protease C (F) and protease N (H) were compounded, with a total addition amount of 2‰ (w / v). The addition amount of each protease accounted for 10%~80% of the total protease. The ratio was optimized by mixed test design (Table 2). The enzyme was hydrolyzed at 50℃ for 3h, and then the enzyme was boiled to inactivate the enzyme and then centrifuged to determine the antioxidant capacity and ACE inhibition rate. The results are shown in Table 2.

[0032] Table 2 Mixing test design and results for optimizing protease ratio

[0033] Design~expert 13 was used for quadratic regression, and the model was extremely significant (p=0.0003), and the lack of fit term was not significant (p=0.8024). The ACE inhibition rate was optimized according to the regression model. When the ACE inhibition rate was set to >85%, the required protease X addition ratio range was 10%~53%, the protease C addition ratio range was 10%~45%, and the protease N addition ratio range was 34%~80%. 4 Optimization of the process for preparing ACE inhibitory peptides by enzymatic hydrolysis of goat milk with compound protease Taking ACE inhibition rate as an indicator, three factors including hydrolysis temperature, time and compound enzyme addition were selected to optimize the process of preparing ACE inhibitory peptides by compound protease hydrolysis of goat milk. The hydrolysis temperature was set at 40℃~60℃, the time was 2h~4h, the compound enzyme addition was 2‰~4‰ (protease X: protease C: protease N = 10%: 10%: 80%), and the Box-Behnken experimental design was used to optimize the compound enzyme hydrolysis process. The experimental design and results are shown in Table 3.

[0034] Table 3 Box-Behnken test design and results of compound protease hydrolysis process

[0035] Design-expert 13 was used to perform regression analysis on the response values, and the quadratic regression equation was obtained: Y=94.22+6.01A+4.50B+2.57C+0.89AB-0.46AC+2.34BC-8.63A 2 -2.18B 2 -5.65C 2 ; Within the experimental group, the ACE inhibition rate was 70.86%~96.12%. The ACE inhibition rate was set to 90%~96%, the required enzymatic hydrolysis temperature range was 45℃~60℃, the compound enzyme addition amount range was 2.35‰~4‰, and the enzymatic hydrolysis time range was 2.4h~4.0h.

[0036] Preferably, Figure 3~Figure 5 As shown in the figure, the ACE inhibition rate is required to reach the maximum value. The experimental design software predicts that the optimal process parameters for preparing ACE inhibitory peptides by enzymatic hydrolysis of goat milk with compound protease are: compound enzyme addition amount 2.816‰, enzymatic hydrolysis at 55℃ for 3.493 h, and ACE inhibition rate of 96.142%. By rounding off the enzymatic hydrolysis conditions and conducting a verification test, the ACE inhibition rate of enzymatic hydrolyzed goat milk was 96.32±1.25% when the compound enzyme addition amount was 2.8‰ and the enzymatic hydrolysis was 55℃ for 3.5 h, which was not significantly different from the predicted value, indicating that the model is effective and feasible.

[0037] 5. Preparation of ACE inhibitory peptide powder and peptide spectrum identification 2.8‰ composite protease (Protease X:Protease C:Protease N = 10%:10%:80%) was added to sterilized and cooled goat milk, and enzymolysis was carried out at 55℃ for 3.5h. Then, the goat milk enzymolysis solution was ultrafiltrated and separated using ultrafiltration membranes with molecular weight cutoffs of 5K and 1K to obtain three components (I:>5k Da, II:1~5k Da and III:<1k Da). After vacuum freeze-drying, the IC values ​​of ACE inhibition were determined. 50 As shown in Table 4, the ACE inhibition rate of each component is closely related to its relative molecular weight. The ACE inhibition rate increases with the decrease of molecular weight, which is consistent with the fact that most of the molecular weights of ACE inhibitory peptides reported so far are lower than 1K Da. The IC value of ACE inhibition rate of component III is 50 The smallest value indicates that its ACE inhibition rate is the highest, so the enzymatic hydrolysate component with a content less than 1K Da is selected for further separation.

[0038] Table 4 IC of ACE inhibitory activity of each ultrafiltration fraction 50 value

[0039] As shown in Table 5, component III was further separated and purified by preparative chromatography ( Figure 6 ), five components were obtained (I was the collected solution from 4 min to 25 min, II was the collected solution from 25 min to 30 min, III was the collected solution from 30 min to 39 min, IV was the collected solution from 39 min to 48 min, and V was the collected solution from 48 min to 58 min), which were freeze-dried and then the IC value of ACE inhibition rate was determined. 50 The results show that component III has the strongest ACE inhibitory activity, and its ACE inhibitory activity IC 50 The peptide spectrum was identified by HPLC-MS / MS. The results are shown in Table 6 and Figure 7 to Figure 22 .

[0040] Table 5 IC values ​​of ACE inhibition rates of the components collected by preparative chromatography 50 value

[0041] Table 6 Amino acid sequences of ACE inhibitory peptides and their locations in goat milk proteins

[0042] From Table 6 and Figure 7 to Figure 22 It can be seen that the ACE inhibitory peptide powder of component III obtained by preparative chromatography contains 16 peptides, the number of amino acids ranges from 4 to 8, and the molecular weight ranges from 505.29Da to 1904.47Da, which are mainly derived from α s1 -Casein, α s2 -Casein, β-casein, α-lactalbumin, β-lactoglobulin, κ-casein and lactoferrin.

[0043] 4. Preparation of goat milk powder for middle-aged and elderly people 4.1 Dry preparation Component III obtained by preparative chromatography separation is freeze-dried to obtain ACE inhibitory peptide powder, yeast β-glucan and goat colostrum powder for enhancing immunity, probiotics for regulating intestinal flora balance, phosphatidylserine for improving memory and preventing Alzheimer's disease, dietary fiber for preventing constipation and ACE inhibitory peptide powder are added to the goat milk powder formula, and the mixture is evenly mixed by a three-dimensional mixer to obtain middle-aged and elderly formula goat milk powder. The ACE inhibitory peptide prepared by the present invention is used as a food functional factor, and the prepared middle-aged and elderly formula goat milk powder contains ACE inhibitory peptide and has the function of assisting in lowering blood pressure.

[0044] The total amount of the above-mentioned middle-aged and elderly formula goat milk powder is 1kg, of which the raw materials include 0.3g~0.6g of the above-mentioned ACE inhibitory peptide powder, 3g~8g of goat colostrum powder, 0.50g~1.0g of yeast β-glucan, 0.2g~0.5g of phosphatidylserine, 0.5g~1.0g of probiotic powder, 20g~50g of dietary fiber, and the rest is goat milk powder.

[0045] Preferably, the probiotics are one or more of Lactobacillus acidophilus, Bifidobacterium animalis subsp. lactis, Lactobacillus plantarum, Lactobacillus paracasei, and Lactobacillus rhamnosus, and the number of viable bacteria is ≥1.0×10 11 CFU / g; Preferably, the dietary fiber is one or more of polyglucan, isomaltooligosaccharide and inulin; 4.2 Wet preparation Adding 0.0125%-0.25% (w / v) Acer truncatum seed oil, 0.0125%-0.20% (w / v) N-acetylneuraminic acid, 0.0125%-0.25% (w / v) phosphatidylserine, and 0.25%-0.50% (w / v) dietary fiber (one or more of polyglucan, oligomaltosaccharide, and inulin) to goat milk enzymatic hydrolysate, and then concentrating to 45%-52% solid content and spray drying (spray drying conditions: inlet air temperature 170°C-190°C, outlet air temperature 100°C-110°C, flow rate 6 mL / min-8 mL / min) to obtain middle-aged and elderly formula goat milk powder; The calculation method of the relevant parameters of the above-mentioned preparation method of ACE inhibitory peptide is as follows: 1. ACE inhibitory activity assay Table 7 ACE inhibition rate detection method

[0046] The sample addition reaction was carried out in the order shown in Table 7. Finally, 1.7 mL of ethyl acetate was added and shaken for 15 seconds. After stratification, 1 mL of the ethyl acetate layer was taken into a test tube and evaporated in an oven at 100°C. After cooling to room temperature, 3 mL of deionized water was added. After sufficient dissolution, the absorbance was measured at a wavelength of 228 nm. The ACE inhibition rate was calculated according to the following formula: ACE inhibition rate (%) = (1) Where: A is the reaction of sample with ACE and HHL at the same time; B is the reaction of ACE with HHL; C is the blank control.

[0047] 2. Determination of hydrolysis degree The degree of hydrolysis of goat milk protein was measured by the pH-Stat method of Adler-Nissen with some modifications. This method is based on the fact that when protein is hydrolyzed by enzymes, it will absorb or release protons. The amount of protonation is related to the pH of the reaction. The corresponding degree of hydrolysis can be calculated based on the amount of sodium hydroxide or hydrochloric acid added to maintain the original reaction balance.

[0048] Where: (2) B: volume of sodium hydroxide (mL); Mb: sodium hydroxide concentration (mol / L), the titration concentration of sample sodium hydroxide is 0.1 mol / L; 1 / α: the reciprocal of the degree of dissociation of protein hydrolysis; Mp: protein mass (g); h tot : The amount of peptide bonds contained in each gram of goat milk protein (mmol).

[0049] Since goat milk contains multiple proteins such as casein and whey protein, the ratio of whey protein to casein in goat milk is 25:75, so the 1 / α of goat milk casein and whey protein is converted to 2.2675 according to the corresponding ratio. tot is 8.8 mmol, so the h tot is 8.35 mmol.

Claims

1. An ACE inhibitory peptide, characterized in that The amino acid sequence of the ACE inhibitory peptide is shown in SEQ ID NO: 1 to SEQ ID NO:

16.

2. An ACE inhibitory peptide according to claim 1, characterized in that: The ACE inhibitory peptide is derived from β-casein, α s1 -Casein, lactoferrin, β-lactoglobulin, α-lactalbumin, α s2 -Casein and kappa-casein.

3. An ACE inhibitory peptide according to claim 1, characterized in that: ACE inhibitory activity IC of the ACE inhibitory peptide 50 It is 0.15±0.013 mg / mL.

4. The method for preparing an ACE inhibitory peptide according to claim 1, characterized in that: The specific steps are as follows: In terms of mass fraction, 2.35‰ to 4‰ of the composite protease is added to goat milk or reconstituted goat milk; Perform enzymolysis at a constant temperature of 45°C to 60°C for 2.4h to 4.0h, inactivate the enzyme, centrifuge and take the supernatant to obtain goat milk enzymolysis solution, which contains ACE inhibitory peptides; The enzymatic hydrolysate with a molecular weight Mw <1 kDa was taken, and the collected solution from 30min to 39min of the preparative chromatography was collected and freeze-dried to obtain an ACE inhibitory peptide.

5. The method for preparing an ACE inhibitory peptide according to claim 4, characterized in that: The composite protease comprises protease X, protease C and protease N, wherein the addition ratio of protease X is 10%-53%, the addition ratio of protease C is 10%-45%, and the addition ratio of protease N is 34%-80%. Protease X is a highly active concentrated neutral protease with an enzyme activity of 150,000 U / g; protease C is protease Protamex 1.6 with an enzyme activity of 1.6 AU-N / g; and protease N is a neutral protease with an enzyme activity of 0.8 AU / g.

6. The method for preparing an ACE inhibitory peptide according to claim 4, characterized in that: The ACE inhibition rate of the goat milk enzymatic hydrolysate is 90% to 96%.

7. A goat milk powder for middle-aged and elderly people, characterized in that: The total amount is 1 kg, and the raw materials include 0.3 g to 0.6 g of the ACE inhibitory peptide according to any one of claims 1 to 3 or the ACE inhibitory peptide prepared by the preparation method according to any one of claims 4 to 6, 3 g to 8 g of goat colostrum powder, 0.50 g to 1.0 g of yeast β-glucan, 0.2 g to 0.5 g of phosphatidylserine, 0.5 g to 1.0 g of probiotic powder, 20 g to 50 g of dietary fiber, and the rest is goat milk powder; The above raw materials are mixed to obtain the middle-aged and elderly formula goat milk powder.

8. A middle-aged and elderly formula goat milk powder according to claim 7, characterized in that: The probiotics include one or more of Lactobacillus acidophilus, Bifidobacterium animalis subspecies lactis, Lactobacillus plantarum, Lactobacillus paracasei, and Lactobacillus rhamnosus, and the number of viable bacteria is ≥1.0×10 11 CFU / g; The dietary fiber is one or more of polydextran, isomaltooligosaccharide and inulin.

9. A goat milk powder for middle-aged and elderly people, characterized in that: The raw materials include goat milk enzymatic hydrolysate, Acer truncatum seed oil, N-acetylneuraminic acid, phosphatidylserine and dietary fiber. The raw materials are mixed, concentrated to a solid content of 45% to 52%, and then spray-dried to obtain a middle-aged and elderly formula goat milk powder. The goat milk enzymatic hydrolysate is the goat milk enzymatic hydrolysate in claim 4.

10. A middle-aged and elderly formula goat milk powder according to claim 9, characterized in that: Goat milk enzymatic hydrolysate is added with 0.0125%~0.25% (w / v) Acer truncatum seed oil, 0.0125%~0.20% (w / v) N-acetylneuraminic acid, 0.0125%~0.25% (w / v) phosphatidylserine, and 0.25%~0.50% (w / v) dietary fiber; The dietary fiber is one or more of polydextran, isomaltooligosaccharide and inulin.

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