Preparation method and application of soybean protein with optimal calcium-phosphorus ratio

Abstract

The invention discloses a preparation method and application of soybean protein with an optimal calcium-phosphorus ratio. The preparation method comprises the following steps: firstly, adding soybean meal into deionized water, adjusting the pH value to 7.5-8.0 by using a sodium hydroxide solution, stirring, centrifuging, taking supernatant, adjusting the pH value to 4.5 by using a hydrochloric acid solution, standing, centrifuging, and taking precipitate; then stirring and dispersing the precipitate in deionized water, adjusting the pH value to 7 by using a sodium hydroxide solution, and performing pulsed electric field treatment to obtain a premix; adding a compound enzyme I composed of neutral protease, bromelain, trypsin and fig protease into the premix, carrying out first enzymolysis, enzyme deactivation and homogenization treatment, adding a compound enzyme II composed of neutral phytase and alkaline phosphatase, and carrying out second enzymolysis, enzyme deactivation and pulsed electric field treatment to obtain an enzyme treatment product; and finally, adding magnetic chitosan microspheres into an enzyme treatment product, carrying out adsorption treatment and magnetic separation to remove the chitosan microspheres, and carrying out vacuum freeze drying to obtain the soybean protein. The calcium-phosphorus ratio of the obtained soybean protein is the best for human bodies, efficient absorption of calcium by the human bodies is facilitated, and the soybean protein can be used for food and beverage processing and is wide in application prospect.

Description

Technical Field

[0001] The present invention relates to a soybean protein preparation method, in particular to a soybean protein preparation method and application with an optimal calcium-phosphorus ratio, belonging to the technical field of plant protein processing. Background Art

[0002] Plant protein has a protein content of more than 60%, rich in amino acids, no cholesterol, and low in purine, making it an excellent raw material to replace animal protein. The cost of processing and preparing plant protein is relatively low, and it is widely used in the food industry. Soy protein is one of the most common plant proteins, with high yield, complete amino acid composition, and high nutritional value.

[0003] Trace elements play a very important role in maintaining normal physiological functions of the human body. Among them, calcium and phosphorus are two very important trace elements. Calcium and phosphorus are important factors in forming human bones and promoting bone growth and development. The ratio of calcium and phosphorus is fixed. If the content of one element changes, the content of the other element will also be affected. Studies have shown that the calcium-phosphorus ratio of 2:1 is the best calcium-phosphorus ratio for the human body, which is conducive to the absorption and utilization rate of calcium.

[0004] However, the phosphorus content of soy protein is about 800mg / 100g, and the calcium content is about 80mg / 100g. Too high phosphorus content will cause imbalance in calcium-phosphorus ratio, which will lead to calcium loss. Therefore, reducing the phosphorus content in soy protein and increasing the calcium content is of great significance to human health.

[0005] Patent application CN109329556A discloses a soy protein preparation process, which uses soybean cake as the main raw material, adds hydrochloric acid and ethanol aqueous solution during the preparation process, and adds dietary fiber and trace elements required by the human body during the preparation process to finally prepare soy protein. The calcium-phosphorus ratio of the soy protein product prepared in this patent application is not optimal for the human body.

[0006] Patent application CN101779721A discloses a method for preparing soybean protein, which uses two immobilized enzyme membranes, namely, Bacillus licheniformis alkaline protease membrane and transglutaminase membrane, and adopts the basic mode of combining continuous product separation with ultrafiltration separation technology to complete enzyme immobilization, and modifying natural soybean protein by enzymatic hydrolysis and repolymerization. This patent application helps to improve the solubility and dispersibility of soybean protein. The molecular structure integrity of soybean protein is good, but the original calcium-phosphorus ratio in soybean protein is basically retained, and the phosphorus content is too high, which cannot meet the optimal calcium-phosphorus ratio requirement of the human body. Summary of the invention

[0007] The purpose of the present invention is to overcome the deficiencies of the above-mentioned prior art and to provide a method for preparing soybean protein with an optimal calcium-phosphorus ratio and its application, wherein the calcium-phosphorus ratio is optimal for the human body and is conducive to the efficient absorption of calcium by the human body.

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

[0009] 1. A method for preparing soybean protein with an optimal calcium-phosphorus ratio, the specific steps are as follows:

[0010] (1) firstly add soybean meal into deionized water, adjust the pH value to 7.5-8.0 with sodium hydroxide solution, stir, centrifuge and take the supernatant, adjust the pH value to 4.5 with hydrochloric acid solution, let stand, and centrifuge to take the precipitate;

[0011] (2) then stirring and dispersing the precipitate obtained in step (1) in deionized water, adjusting the pH to 7 using a sodium hydroxide solution, and subjecting the mixture to a pulse electric field treatment to obtain a premix;

[0012] (3) adding a composite enzyme I composed of neutral protease, bromelain, trypsin, and ficin to the premix, performing a first enzymatic hydrolysis, inactivating the enzymes, and homogenizing the premix, adding a composite enzyme II composed of neutral phytase and alkaline phosphatase, performing a second enzymatic hydrolysis, inactivating the enzymes, and treating the premix with a pulsed electric field to obtain an enzyme-treated product;

[0013] (4) Finally, magnetic chitosan microspheres are added to the enzyme-treated product, subjected to adsorption treatment, the chitosan microspheres are removed by magnetic separation, and vacuum freeze-dried to obtain the soy protein.

[0014] Preferably, in step (1), the mass ratio of soybean meal to deionized water is 1:12-15.

[0015] Preferably, in step (1), the concentration of the sodium hydroxide solution is 1 to 2 mol / L, and the concentration of the hydrochloric acid solution is 1 to 2 mol / L.

[0016] Preferably, in step (1), the stirring process conditions are: stirring at 300-500 r / min for 50-60 minutes.

[0017] Preferably, in step (1), the mixture is allowed to stand for 3 to 5 hours and then centrifuged at 4°C under the conditions of 6000 to 8000 r / min for 8 to 10 minutes.

[0018] Preferably, in step (2), the mass ratio of precipitate to deionized water is 1:15-20.

[0019] Preferably, in step (2), the concentration of the sodium hydroxide solution is 1 to 2 mol / L.

[0020] Preferably, in step (2), the process conditions of the pulse electric field treatment are: frequency 300-400 Hz, electric field strength 30-40 V / cm, and treatment time 300-500 μs.

[0021] Preferably, in step (3), the mass ratio of the premix, complex enzyme I, and complex enzyme II is 100:0.8-1.2:0.8-1.2.

[0022] Preferably, in step (3), the compound enzyme I is obtained by mixing neutral protease, bromelain, trypsin, ficin and the like.

[0023] Preferably, in step (3), the process conditions for the first enzymatic hydrolysis are: temperature 40-50° C., pH=7-8, and time 50-60 minutes.

[0024] Preferably, in step (3), the process conditions for homogenization treatment are: pressure 80 MPa, time 8 to 10 minutes.

[0025] Preferably, in step (3), the complex enzyme II is obtained by mixing neutral phytase and alkaline phosphatase in a mass ratio of 2:1.

[0026] Preferably, in step (3), the process conditions of the second enzymatic hydrolysis are: temperature 40-45° C., pH=7-8, and time 55-65 minutes; after the second enzymatic hydrolysis is completed, the pH is adjusted to 7 using sodium hydroxide solution.

[0027] Preferably, in step (3), the process conditions of the pulse electric field treatment are: frequency 600-800 Hz, electric field strength 40-50 V / cm, and treatment time 600-800 μs.

[0028] Preferably, in step (4), the mass ratio of the enzyme-treated product to the magnetic chitosan microspheres is 100:1-2.

[0029] Preferably, in step (4), the magnetic chitosan microspheres are prepared by the following method, based on parts by mass:

[0030] (A) First, chitosan is prepared into 24 parts of chitosan acetic acid solution with a mass concentration of 2% using a 1% acetic acid solution, then 0.15 parts of ferrosoferric oxide powder is added, and ultrasonic oscillation is performed at 500W for 30 minutes, and then 45 parts of liquid paraffin and 22 parts of Span-80 are added, and the mixture is stirred to obtain a mixed solution;

[0031] (B) The mixed solution is heated to 55° C., 3 parts of glutaraldehyde are added, and the mixture is stirred at 500 r / min for cross-linking for 3 hours. The solid I is filtered out and dispersed in a 1 mol / L sodium hydroxide solution with a mass that is 4 times the mass of the solid I. The mixture is heated to 67° C. and stirred for 2 hours. The solid II is filtered out and washed with acetone, petroleum ether, anhydrous ethanol, and distilled water in sequence. The mixture is filtered out, dried, and magnetically separated and collected to obtain the magnetic chitosan microspheres.

[0032] Preferably, in step (4), the process conditions for the adsorption treatment are: stirring and adsorption at 300-500 r / min for 20-30 minutes.

[0033] Preferably, in step (4), the process conditions for vacuum freeze drying are: vacuum degree 0.07 to 0.09 MPa, temperature -50 to -55°C, and time 40 to 50 hours.

[0034] 2. A soybean protein with an optimal calcium-phosphorus ratio is obtained by the above-mentioned preparation method.

[0035] 3. Application of the aforementioned soy protein with an optimal calcium-phosphorus ratio in food and beverage processing.

[0036] Beneficial effects of the present invention:

[0037] The method first adds soybean meal to deionized water, adjusts pH to 7.5-8.0 with sodium hydroxide solution, stirs, centrifuges to obtain supernatant, adjusts pH to 4.5 with hydrochloric acid solution, stands, and centrifuges to obtain precipitate; then stirs and disperses the precipitate in deionized water, adjusts pH to 7 with sodium hydroxide solution, and performs pulse electric field treatment to obtain a premix; then adds composite enzyme I composed of neutral protease, bromelain, trypsin, and ficin to the premix, performs a first enzymolysis, inactivates the enzyme, and performs homogenization treatment, adds composite enzyme II composed of neutral phytase and alkaline phosphatase, performs a second enzymolysis, inactivates the enzyme, and performs pulse electric field treatment to obtain an enzyme treatment product; finally, magnetic chitosan microspheres are added to the enzyme treatment product, adsorbed, and the chitosan microspheres are removed by magnetic separation, and vacuum freeze-dried to obtain soybean protein. The soybean protein obtained by the method has an optimal calcium-phosphorus ratio for human body, is beneficial to the efficient absorption of calcium by human body, can be used for food and beverage processing, and has broad application prospects.

[0038] When preparing the premix, pulse electric field treatment is used to break the entanglement of large molecular proteins and release the phosphorus-containing components, providing a good foundation for the subsequent reduction of phosphorus content.

[0039] The invention utilizes compound enzyme I and compound enzyme II to perform enzymatic hydrolysis twice, wherein compound enzyme I is composed of neutral protease, bromelain, trypsin and ficin, and compound enzyme II is composed of neutral phytase and alkaline phosphatase, so as to further release and degrade phosphorus-containing components and reduce phosphorus content.

[0040] After the enzymatic hydrolysis is completed, the present invention further realizes the adsorption and removal of phosphorus by adsorption treatment of magnetic chitosan microspheres. The adsorption treatment time should not be too long to avoid too low phosphorus content. DETAILED DESCRIPTION

[0041] The present invention is further described below in conjunction with embodiments. It should be noted that the following description is only for explaining the present invention and does not limit its contents.

[0042] Embodiment 1:

[0043] A method for preparing soybean protein with an optimal calcium-phosphorus ratio, the specific steps are as follows:

[0044] (1) First, soybean meal was added to deionized water (the mass ratio of soybean meal to deionized water was 1:12), pH was adjusted to 7.5 using 1 mol / L sodium hydroxide solution, stirred at 300 r / min for 50 minutes, centrifuged to obtain the supernatant, pH was adjusted to 4.5 using 1 mol / L hydrochloric acid solution, allowed to stand for 3 hours, and then centrifuged at 4°C. The centrifugal conditions were: centrifugation at 6000 r / min for 8 minutes, and the precipitate was obtained;

[0045] (2) then taking the precipitate obtained in step (1), stirring and dispersing it in deionized water (the mass ratio of the precipitate to the deionized water is 1:15), adjusting the pH to 7 using a 1 mol / L sodium hydroxide solution, and subjecting it to a pulse electric field treatment to obtain a premix;

[0046] (3) adding 0.8 kg of compound enzyme I to 100 kg of the premix, performing the first enzymolysis, inactivating the enzyme, and homogenizing, adding 0.8 kg of compound enzyme II, performing the second enzymolysis, inactivating the enzyme, and treating with a pulse electric field to obtain an enzyme-treated product;

[0047] (4) Finally, magnetic chitosan microspheres are added to the enzyme-treated product, subjected to adsorption treatment, the chitosan microspheres are removed by magnetic separation, and vacuum freeze-dried to obtain the soy protein.

[0048] Wherein, in step (2), the process conditions of the pulse electric field treatment are: frequency 300 Hz, electric field strength 30 V / cm, and treatment time 300 μs.

[0049] In step (3), the compound enzyme I is obtained by mixing neutral protease, bromelain, trypsin, ficin and the like.

[0050] The process conditions of the first enzymatic hydrolysis were: temperature 40°C, pH=7, and time 50 minutes.

[0051] The process conditions of homogenization treatment are: pressure 80MPa, time 8 minutes.

[0052] The complex enzyme II is obtained by mixing neutral phytase and alkaline phosphatase in a mass ratio of 2:1.

[0053] The process conditions of the second enzymatic hydrolysis were: temperature 40° C., pH=7, and time 55 minutes; after the second enzymatic hydrolysis was completed, the pH was adjusted to 7 using sodium hydroxide solution.

[0054] The process conditions of the pulse electric field treatment are: frequency 600 Hz, electric field strength 40 V / cm, and treatment time 600 μs.

[0055] In step (4), the mass ratio of the enzyme-treated product to the magnetic chitosan microspheres is 100:1.

[0056] The magnetic chitosan microspheres are prepared by the following method:

[0057] (A) First, chitosan is prepared into 24 kg of chitosan acetic acid solution with a mass concentration of 2% using a 1% acetic acid solution, then 0.15 kg of ferrosoferric oxide powder is added, and ultrasonic oscillation is performed at 500 W for 30 minutes, and then 45 kg of liquid paraffin and 22 kg of Span-80 are added, and the mixture is stirred to obtain a mixed solution;

[0058] (B) The mixed solution is heated to 55° C., 3 kg of glutaraldehyde is added, and the mixture is stirred at 500 r / min for cross-linking for 3 hours. The solid I is filtered out and dispersed in a 1 mol / L sodium hydroxide solution with a mass that is 4 times the mass of the solid I. The mixture is heated to 67° C. and stirred for 2 hours. The solid II is filtered out and washed with acetone, petroleum ether, anhydrous ethanol, and distilled water in sequence. The mixture is filtered out, dried, and magnetically separated and collected to obtain the magnetic chitosan microspheres.

[0059] The process conditions of adsorption treatment are: stirring and adsorption at 300r / min for 20 minutes.

[0060] The process conditions of vacuum freeze drying are: vacuum degree 0.07MPa, temperature -50°C, and time 40 hours.

[0061] Embodiment 2:

[0062] A method for preparing soybean protein with an optimal calcium-phosphorus ratio, the specific steps are as follows:

[0063] (1) First, soybean meal was added to deionized water (the mass ratio of soybean meal to deionized water was 1:15), the pH was adjusted to 8.0 using 2 mol / L sodium hydroxide solution, stirred at 500 r / min for 60 minutes, and the supernatant was collected by centrifugation. The pH was adjusted to 4.5 using 2 mol / L hydrochloric acid solution, and the supernatant was collected after standing for 5 hours and centrifuged at 4°C. The centrifugation conditions were: centrifugation at 8000 r / min for 10 minutes, and the precipitate was collected;

[0064] (2) then taking the precipitate obtained in step (1) and stirring and dispersing it in deionized water (the mass ratio of the precipitate to the deionized water is 1:20), adjusting the pH to 7 using a 2 mol / L sodium hydroxide solution, and subjecting it to a pulse electric field treatment to obtain a premix;

[0065] (3) adding 1.2 kg of compound enzyme I to 100 kg of the premix, performing the first enzymolysis, inactivating the enzyme, and homogenizing, adding 1.2 kg of compound enzyme II, performing the second enzymolysis, inactivating the enzyme, and treating with a pulse electric field to obtain an enzyme-treated product;

[0066] (4) Finally, magnetic chitosan microspheres are added to the enzyme-treated product, subjected to adsorption treatment, the chitosan microspheres are removed by magnetic separation, and vacuum freeze-dried to obtain the soy protein.

[0067] Wherein, in step (2), the process conditions of the pulse electric field treatment are: frequency 400 Hz, electric field strength 40 V / cm, and treatment time 500 μs.

[0068] In step (3), the compound enzyme I is obtained by mixing neutral protease, bromelain, trypsin, ficin and the like.

[0069] The process conditions of the first enzymatic hydrolysis were: temperature 50°C, pH=8, and time 60 minutes.

[0070] The process conditions of homogenization treatment are: pressure 80MPa, time 10 minutes.

[0071] The complex enzyme II is obtained by mixing neutral phytase and alkaline phosphatase in a mass ratio of 2:1.

[0072] The process conditions of the second enzymatic hydrolysis are: temperature 45°C, pH=8, and time 65 minutes; after the second enzymatic hydrolysis is completed, the pH is adjusted to 7 using sodium hydroxide solution.

[0073] The process conditions of the pulse electric field treatment are: frequency 800 Hz, electric field strength 50 V / cm, and treatment time 800 μs.

[0074] In step (4), the mass ratio of the enzyme-treated product to the magnetic chitosan microspheres is 100:2.

[0075] The magnetic chitosan microspheres are prepared by the following method:

[0076] (A) First, chitosan is prepared into 24 kg of chitosan acetic acid solution with a mass concentration of 2% using a 1% acetic acid solution, then 0.15 kg of ferrosoferric oxide powder is added, and ultrasonic oscillation is performed at 500 W for 30 minutes, and then 45 kg of liquid paraffin and 22 kg of Span-80 are added, and the mixture is stirred to obtain a mixed solution;

[0077] (B) The mixed solution is heated to 55° C., 3 kg of glutaraldehyde is added, and the mixture is stirred at 500 r / min for cross-linking for 3 hours. The solid I is filtered out and dispersed in a 1 mol / L sodium hydroxide solution with a mass that is 4 times the mass of the solid I. The mixture is heated to 67° C. and stirred for 2 hours. The solid II is filtered out and washed with acetone, petroleum ether, anhydrous ethanol, and distilled water in sequence. The mixture is filtered out, dried, and magnetically separated and collected to obtain the magnetic chitosan microspheres.

[0078] The process conditions of adsorption treatment are: stirring and adsorption at 500r / min for 30 minutes.

[0079] The process conditions of vacuum freeze drying are: vacuum degree 0.09 MPa, temperature -55°C, and time 50 hours.

[0080] Embodiment 3:

[0081] A method for preparing soybean protein with an optimal calcium-phosphorus ratio, the specific steps are as follows:

[0082] (1) First, soybean meal was added to deionized water (the mass ratio of soybean meal to deionized water was 1:13), the pH was adjusted to 8.0 using 1 mol / L sodium hydroxide solution, stirred at 400 r / min for 55 minutes, and the supernatant was collected by centrifugation. The pH was adjusted to 4.5 using 1 mol / L hydrochloric acid solution, and the supernatant was collected after standing for 4 hours and centrifuged at 4°C. The centrifugation conditions were: centrifugation at 7000 r / min for 9 minutes, and the precipitate was collected;

[0083] (2) then taking the precipitate obtained in step (1) and stirring and dispersing it in deionized water (the mass ratio of the precipitate to the deionized water is 1:18), adjusting the pH to 7 using a 1 mol / L sodium hydroxide solution, and subjecting it to a pulse electric field treatment to obtain a premix;

[0084] (3) adding 1 kg of compound enzyme I to 100 kg of the premix, performing the first enzymolysis, inactivating the enzyme, and homogenizing, adding 1 kg of compound enzyme II, performing the second enzymolysis, inactivating the enzyme, and treating with a pulse electric field to obtain an enzyme-treated product;

[0085] (4) Finally, magnetic chitosan microspheres are added to the enzyme-treated product, subjected to adsorption treatment, the chitosan microspheres are removed by magnetic separation, and vacuum freeze-dried to obtain the soy protein.

[0086] Wherein, in step (2), the process conditions of the pulse electric field treatment are: frequency 400 Hz, electric field strength 35 V / cm, and treatment time 400 μs.

[0087] In step (3), the compound enzyme I is obtained by mixing neutral protease, bromelain, trypsin, ficin and the like.

[0088] The process conditions of the first enzymatic hydrolysis were: temperature 45°C, pH=7, and time 55 minutes.

[0089] The process conditions of homogenization treatment are: pressure 80MPa, time 9 minutes.

[0090] The complex enzyme II is obtained by mixing neutral phytase and alkaline phosphatase in a mass ratio of 2:1.

[0091] The process conditions of the second enzymatic hydrolysis are: temperature 42° C., pH=7, and time 60 minutes; after the second enzymatic hydrolysis is completed, the pH is adjusted to 7 using sodium hydroxide solution.

[0092] The process conditions of the pulse electric field treatment are: frequency 700 Hz, electric field strength 45 V / cm, and treatment time 700 μs.

[0093] In step (4), the mass ratio of the enzyme-treated product to the magnetic chitosan microspheres is 100:1.5.

[0094] The magnetic chitosan microspheres are prepared by the following method:

[0095] (A) First, chitosan is prepared into 24 kg of chitosan acetic acid solution with a mass concentration of 2% using a 1% acetic acid solution, then 0.15 kg of ferrosoferric oxide powder is added, and ultrasonic oscillation is performed at 500 W for 30 minutes, and then 45 kg of liquid paraffin and 22 kg of Span-80 are added, and the mixture is stirred to obtain a mixed solution;

[0096] (B) The mixed solution is heated to 55° C., 3 kg of glutaraldehyde is added, and the mixture is stirred at 500 r / min for cross-linking for 3 hours. The solid I is filtered out and dispersed in a 1 mol / L sodium hydroxide solution with a mass that is 4 times the mass of the solid I. The mixture is heated to 67° C. and stirred for 2 hours. The solid II is filtered out and washed with acetone, petroleum ether, anhydrous ethanol, and distilled water in sequence. The mixture is filtered out, dried, and magnetically separated and collected to obtain the magnetic chitosan microspheres.

[0097] The process conditions of adsorption treatment are: stirring and adsorption at 400r / min for 25 minutes.

[0098] The process conditions of vacuum freeze drying are: vacuum degree 0.08MPa, temperature -55°C, and time 45 hours.

[0099] Comparative Example 1

[0100] A method for preparing soybean protein, the specific steps are as follows:

[0101] (1) First, soybean meal was added to deionized water (the mass ratio of soybean meal to deionized water was 1:12), pH was adjusted to 7.5 using 1 mol / L sodium hydroxide solution, stirred at 300 r / min for 50 minutes, centrifuged to obtain the supernatant, pH was adjusted to 4.5 using 1 mol / L hydrochloric acid solution, allowed to stand for 3 hours, and then centrifuged at 4°C. The centrifugal conditions were: centrifugation at 6000 r / min for 8 minutes, and the precipitate was obtained;

[0102] (2) then taking the precipitate obtained in step (1), stirring and dispersing it in deionized water (the mass ratio of the precipitate to the deionized water is 1:15), adjusting the pH to 7 using a 1 mol / L sodium hydroxide solution, and subjecting it to a pulse electric field treatment to obtain a premix;

[0103] (3) adding 0.8 kg of compound enzyme I to 100 kg of the premix, performing the first enzymolysis, inactivating the enzyme, and homogenizing, adding 0.8 kg of compound enzyme II, performing the second enzymolysis, inactivating the enzyme, and treating with a pulse electric field to obtain an enzyme-treated product;

[0104] (4) Finally, magnetic chitosan microspheres are added to the enzyme-treated product, subjected to adsorption treatment, the chitosan microspheres are removed by magnetic separation, and vacuum freeze-dried to obtain the soy protein.

[0105] Wherein, in step (2), the process conditions of the pulse electric field treatment are: frequency 300 Hz, electric field strength 30 V / cm, and treatment time 300 μs.

[0106] In step (3), the compound enzyme I is obtained by mixing neutral protease, bromelain, trypsin, ficin and the like.

[0107] The process conditions of the first enzymatic hydrolysis were: temperature 40°C, pH=7, and time 50 minutes.

[0108] The process conditions of homogenization treatment are: pressure 80MPa, time 8 minutes.

[0109] The complex enzyme II is neutral phytase.

[0110] The process conditions of the second enzymatic hydrolysis were: temperature 40° C., pH=7, and time 55 minutes; after the second enzymatic hydrolysis was completed, the pH was adjusted to 7 using sodium hydroxide solution.

[0111] The process conditions of the pulse electric field treatment are: frequency 600 Hz, electric field strength 40 V / cm, and treatment time 600 μs.

[0112] In step (4), the mass ratio of the enzyme-treated product to the magnetic chitosan microspheres is 100:1.

[0113] The magnetic chitosan microspheres are prepared by the following method:

[0114] (A) First, chitosan is prepared into 24 kg of chitosan acetic acid solution with a mass concentration of 2% using a 1% acetic acid solution, then 0.15 kg of ferrosoferric oxide powder is added, and ultrasonic oscillation is performed at 500 W for 30 minutes, and then 45 kg of liquid paraffin and 22 kg of Span-80 are added, and the mixture is stirred to obtain a mixed solution;

[0115] (B) The mixed solution is heated to 55° C., 3 kg of glutaraldehyde is added, and the mixture is stirred at 500 r / min for cross-linking for 3 hours. The solid I is filtered out and dispersed in a 1 mol / L sodium hydroxide solution with a mass that is 4 times the mass of the solid I. The mixture is heated to 67° C. and stirred for 2 hours. The solid II is filtered out and washed with acetone, petroleum ether, anhydrous ethanol, and distilled water in sequence. The mixture is filtered out, dried, and magnetically separated and collected to obtain the magnetic chitosan microspheres.

[0116] The process conditions of adsorption treatment are: stirring and adsorption at 300r / min for 20 minutes.

[0117] The process conditions of vacuum freeze drying are: vacuum degree 0.07MPa, temperature -50°C, and time 40 hours.

[0118] Comparative Example 2

[0119] A method for preparing soybean protein, the specific steps are as follows:

[0120] (1) First, soybean meal was added to deionized water (the mass ratio of soybean meal to deionized water was 1:12), pH was adjusted to 7.5 using 1 mol / L sodium hydroxide solution, stirred at 300 r / min for 50 minutes, centrifuged to obtain the supernatant, pH was adjusted to 4.5 using 1 mol / L hydrochloric acid solution, allowed to stand for 3 hours, and then centrifuged at 4°C. The centrifugal conditions were: centrifugation at 6000 r / min for 8 minutes, and the precipitate was obtained;

[0121] (2) then taking the precipitate obtained in step (1), stirring and dispersing it in deionized water (the mass ratio of the precipitate to the deionized water is 1:15), adjusting the pH to 7 using a 1 mol / L sodium hydroxide solution, and subjecting it to a pulse electric field treatment to obtain a premix;

[0122] (3) Add 0.8 kg of compound enzyme I to 100 kg of the premix, perform the first enzymolysis, inactivate the enzyme, and homogenize, then add 0.8 kg of compound enzyme II, perform the second enzymolysis, inactivate the enzyme, perform pulse electric field treatment, and vacuum freeze-dry to obtain the soy protein.

[0123] Wherein, in step (2), the process conditions of the pulse electric field treatment are: frequency 300 Hz, electric field strength 30 V / cm, and treatment time 300 μs.

[0124] In step (3), the compound enzyme I is obtained by mixing neutral protease, bromelain, trypsin, ficin and the like.

[0125] The process conditions of the first enzymatic hydrolysis were: temperature 40°C, pH=7, and time 50 minutes.

[0126] The process conditions of homogenization treatment are: pressure 80MPa, time 8 minutes.

[0127] The complex enzyme II is obtained by mixing neutral phytase and alkaline phosphatase in a mass ratio of 2:1.

[0128] The process conditions of the second enzymatic hydrolysis were: temperature 40° C., pH=7, and time 55 minutes; after the second enzymatic hydrolysis was completed, the pH was adjusted to 7 using sodium hydroxide solution.

[0129] The process conditions of the pulse electric field treatment are: frequency 600 Hz, electric field strength 40 V / cm, and treatment time 600 μs.

[0130] The process conditions of vacuum freeze drying are: vacuum degree 0.07MPa, temperature -50°C, and time 40 hours.

[0131] Test example

[0132] The calcium content and phosphorus content of the soybean proteins obtained in Examples 1 to 3 and Comparative Examples 1 and 2 were tested respectively. The results are shown in Table 1.

[0133] Calcium content: refer to GB5009.92-2016 Method 1.

[0134] Phosphorus content: refer to GB5009.87-2016 method 2.

[0135] Table 1. Test of calcium and phosphorus content in soy protein

[0136] Calcium content (mg / 100g) Phosphorus content (mg / 100g) Calcium to phosphorus ratio Example 1 80.03 39.42 2.03：1 Example 2 79.98 40.39 1.98：1 Example 3 80.11 39.85 2.01：1 Comparative Example 1 78.23 156.27 0.50：1 Comparative Example 2 79.99 89.18 0.90：1

[0137] As can be seen from Table 1, the calcium-phosphorus ratio of the soybean protein obtained in Examples 1 to 3 is close to 2:1, which is the optimal calcium-phosphorus ratio for the human body.

[0138] In comparative example 1, alkaline phosphatase was omitted from the complex enzyme II, and in comparative example 2, magnetic chitosan microsphere adsorption treatment was omitted. The phosphorus content increased significantly and the calcium-phosphorus ratio decreased significantly, which is not conducive to human absorption. This shows that the two-step enzymatic hydrolysis treatment and the subsequent adsorption treatment work synergistically to reduce the phosphorus content in the product.

[0139] Although the above describes the specific implementation methods of the present invention, it does not limit the scope of protection of the present invention. Based on the technical solution of the present invention, various modifications or variations that can be made by those skilled in the art without creative work are still within the scope of protection of the present invention.

Claims

1. A method for preparing soybean protein with an optimal calcium-phosphorus ratio, characterized in that: The specific steps are as follows: (1) firstly add soybean meal into deionized water, adjust the pH value to 7.5-8.0 with sodium hydroxide solution, stir, centrifuge and take the supernatant, adjust the pH value to 4.5 with hydrochloric acid solution, let stand, and centrifuge to take the precipitate; (2) then stirring and dispersing the precipitate obtained in step (1) in deionized water, adjusting the pH to 7 using a sodium hydroxide solution, and subjecting the mixture to a pulse electric field treatment to obtain a premix; (3) adding a composite enzyme I composed of neutral protease, bromelain, trypsin, and ficin to the premix, performing a first enzymatic hydrolysis, inactivating the enzymes, and homogenizing the premix, adding a composite enzyme II composed of neutral phytase and alkaline phosphatase, performing a second enzymatic hydrolysis, inactivating the enzymes, and treating the premix with a pulsed electric field to obtain an enzyme-treated product; (4) Finally, magnetic chitosan microspheres are added to the enzyme-treated product, subjected to adsorption treatment, the chitosan microspheres are removed by magnetic separation, and vacuum freeze-dried to obtain the soy protein.

2. The preparation method according to claim 1, characterized in that: In step (1), the mass ratio of soybean meal to deionized water is 1:12-15; The concentration of sodium hydroxide solution is 1-2 mol / L, and the concentration of hydrochloric acid solution is 1-2 mol / L; The stirring process conditions are: stirring at 300-500 r / min for 50-60 minutes; After standing for 3 to 5 hours, centrifuge at 4°C at 6000 to 8000 r / min for 8 to 10 minutes.

3. The preparation method according to claim 1, characterized in that: In step (2), the mass ratio of precipitate to deionized water is 1:15-20; The concentration of sodium hydroxide solution is 1-2 mol / L; The process conditions of the pulse electric field treatment are: frequency 300-400 Hz, electric field strength 30-40 V / cm, and treatment time 300-500 μs.

4. The preparation method according to claim 1, characterized in that: In step (3), the mass ratio of the premix, complex enzyme I and complex enzyme II is 100: 0.8-1.2: 0.8-1.

2.

5. The preparation method according to claim 1, characterized in that: In step (3), the compound enzyme I is obtained by mixing neutral protease, bromelain, trypsin, ficin and the like; The process conditions of the first enzymatic hydrolysis are: temperature 40-50°C, pH=7-8, and time 50-60 minutes.

6. The preparation method according to claim 1, characterized in that: In step (3), the process conditions for homogenization treatment are: pressure 80 MPa, time 8 to 10 minutes.

7. The preparation method according to claim 1, characterized in that: In step (3), the complex enzyme II is obtained by mixing neutral phytase and alkaline phosphatase in a mass ratio of 2:1; The process conditions of the second enzymatic hydrolysis are: temperature 40-45°C, pH=7-8, time 55-65 minutes; after the second enzymatic hydrolysis is completed, the pH is adjusted to 7 using sodium hydroxide solution; In step (3), the process conditions of the pulse electric field treatment are: frequency 600-800 Hz, electric field strength 40-50 V / cm, and treatment time 600-800 μs.

8. The preparation method according to claim 1, characterized in that: In step (4), the mass ratio of the enzyme-treated product to the magnetic chitosan microspheres is 100:1-2; The process conditions of adsorption treatment are: stirring and adsorption at 300-500 r / min for 20-30 minutes; The process conditions of vacuum freeze drying are: vacuum degree 0.07~0.09MPa, temperature -50~-55℃, time 40~50 hours.

9. A soy protein with an optimal calcium-phosphorus ratio, characterized in that: The invention is obtained by the preparation method according to any one of claims 1 to 8.

10. Use of the soybean protein with an optimal calcium-phosphorus ratio as claimed in claim 9 in food and beverage processing.