Rice-pea protein compound as well as preparation method and application thereof

Abstract

The invention provides a rice-pea protein compound as well as a preparation method and application thereof, and belongs to the technical field of protein compound preparation. The preparation method comprises the following steps: mixing rice meal with water, adjusting the pH value to 10-13, and sequentially carrying out first stirring treatment and first centrifugation to obtain rice meal supernate; mixing the defatted pea meal with water, adjusting the pH value to 8-11, and sequentially performing second stirring treatment and second centrifugation to obtain pea meal supernate; mixing the rice protein and the pea protein, adjusting the pH value to 12-13, carrying out third stirring treatment, then adjusting the pH value to 6.5-7.5, carrying out fourth stirring treatment, then carrying out standing and third centrifugation in sequence, and carrying out dialysis and freeze drying on the obtained mixed supernatant in sequence to obtain the rice-pea protein compound. According to the method disclosed by the invention, the rice protein can be modified safely, greenly and harmlessly; the rice-pea protein compound has high solubility, and the amino acid composition is balanced.

Description

Technical Field

[0001] The invention relates to the technical field of protein modification and preparation, and in particular to a rice-pea protein complex and a preparation method and application thereof. Background Art

[0002] Rice protein is rich in sulfur-containing amino acids, easy to digest in the gastrointestinal tract, and has the advantages of low allergenicity and high bioavailability. However, it lacks lysine, and the low solubility of rice protein in water greatly limits its application in food. It is very important to use modification methods to improve the functional properties of protein. Common protein modification methods include physical modification, chemical modification, and enzyme modification. Physical modification requires high equipment and high cost, making it difficult to achieve industrial production; the chemical modification process is difficult to control, and may produce undesirable by-products, causing food safety problems and large nutritional losses; the hydrolysis process of enzymatic modification will produce bitter peptides, affecting the flavor of the food. Summary of the invention

[0003] The purpose of the present invention is to provide a rice-pea protein complex and a preparation method and application thereof. The method provided by the present invention can achieve the modification of rice protein in a safe, green and harmless manner; the rice-pea protein complex prepared by the method of the present invention has high solubility and balanced amino acid composition.

[0004] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0005] The present invention provides a method for preparing a rice-pea protein complex, comprising the following steps:

[0006] Mixing rice flour with water to obtain a rice flour suspension; adjusting the pH value of the rice flour suspension to 10-13, and sequentially performing a first stirring treatment and a first centrifugation to obtain a rice flour supernatant;

[0007] Mixing defatted pea flour with water to obtain a pea flour suspension; adjusting the pH value of the pea flour suspension to 8-11, and sequentially performing a second stirring treatment and a second centrifugation to obtain a pea flour supernatant;

[0008] The rice flour supernatant and the pea flour supernatant are mixed to obtain a first mixed solution; the pH value of the first mixed solution is adjusted to 12-13, and a third stirring process is performed to obtain a second mixed solution; the pH value of the second mixed solution is adjusted to 6.5-7.5, and a fourth stirring process is performed, and then standing and a third centrifugation are performed in sequence to obtain a mixed supernatant;

[0009] The mixed supernatant is dialyzed and freeze-dried in sequence to obtain the rice-pea protein complex.

[0010] Preferably, the particle size of the rice flour is ≤178 μm; and the solid-liquid ratio of rice flour to water in the rice flour suspension is 1 g:8-12 mL.

[0011] Preferably, the defatted pea flour is obtained by defatting pea flour with n-hexane; the particle size of the pea flour is ≤178 μm; and the solid-liquid ratio of defatted pea flour to water in the pea flour suspension is 1 g:8-12 mL.

[0012] Preferably, the time of the first stirring treatment and the second stirring treatment is independently 3 to 5 hours; the speed of the first centrifugation and the second centrifugation is independently 6000 to 8000 rpm, and the time is independently 15 to 20 minutes.

[0013] Preferably, the mass ratio of rice protein to pea protein in the first mixed solution is 9:1 to 1:9.

[0014] Preferably, the third stirring treatment lasts for 1 to 3 hours.

[0015] Preferably, the fourth stirring treatment is performed for 0.5 to 1.5 hours; the standing time is 12 to 24 hours; the third centrifugation is performed at a speed of 7000 to 8000 rpm and for 15 to 20 minutes.

[0016] The present invention provides a rice-pea protein complex prepared by the preparation method described in the above technical scheme.

[0017] Preferably, the solubility of the rice-pea protein complex is 35-90%, and the particle size is 50-200 nm.

[0018] The present invention provides application of the rice-pea protein complex described in the above technical solution in food.

[0019] The invention provides a method for preparing a rice-pea protein complex. The method comprises the following steps: mixing rice flour with water to obtain a rice flour suspension; adjusting the pH value of the rice flour suspension to 10-13, sequentially performing a first stirring treatment and a first centrifugation to obtain a rice flour supernatant; mixing defatted pea flour with water to obtain a pea flour suspension; adjusting the pH value of the pea flour suspension to 8-11, sequentially performing a second stirring treatment and a second centrifugation to obtain a pea flour supernatant; mixing the rice flour supernatant with the pea flour supernatant to obtain a first mixed solution; adjusting the pH value of the first mixed solution to 12-13, performing a third stirring treatment to obtain a second mixed solution; adjusting the pH value of the second mixed solution to 6.5-7.5, performing a fourth stirring treatment, and then sequentially performing a standing treatment and a third centrifugation to obtain a mixed supernatant; and sequentially performing dialysis and freeze drying on the mixed supernatant to obtain the rice-pea protein complex. The present invention adopts the method of compounding pea protein with rice protein to improve the nutrition and functional properties of rice protein, which is a safe, green and harmless protein modification method. Specifically, the present invention adopts a "one-step method" to prepare a rice-pea protein complex, and there is no need to extract rice protein and pea protein separately before compounding, and the preparation process is simple, the pollution to the environment is small, the energy consumption is low, and it is suitable for large-scale production. At the same time, the rice-pea protein complex prepared by the method of the present invention has a higher solubility and a balanced amino acid composition, and can be used as a high-quality plant protein source in the food industry, solves the problem of low water solubility of rice protein and poor digestibility of pea protein, helps to expand the application field of rice protein and pea protein, improves the utilization rate of protein, and increases the economic benefits of enterprises. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 The process flow chart of preparing rice-pea protein complex in the embodiment of the present invention;

[0021] Figure 2 The particle size distribution diagrams of the A1 complex in Example 1, the B1 mixture in Comparative Example 1, the rice protein (B# rice protein), and the pea protein (B# pea protein);

[0022] Figure 3 The particle size distribution diagrams of the A2 complex in Example 2, the B2 mixture in Comparative Example 2, the rice protein (B# rice protein), and the pea protein (B# pea protein);

[0023] Figure 4 The particle size distribution diagrams of the A3 complex in Example 3, the B3 mixture in Comparative Example 3, the rice protein (B# rice protein), and the pea protein (B# pea protein);

[0024] Figure 5The particle size distribution diagrams of the A4 complex in Example 4, the B4 mixture in Comparative Example 4, the rice protein (B# rice protein), and the pea protein (B# pea protein);

[0025] Figure 6 It is the particle size distribution diagram of the A5 complex in Example 5, the B5 mixture in Comparative Example 5, and the rice protein (B# rice protein) and the pea protein (B# pea protein). DETAILED DESCRIPTION

[0026] The present invention provides a method for preparing a rice-pea protein complex, comprising the following steps:

[0027] Mixing rice flour with water to obtain a rice flour suspension; adjusting the pH value of the rice flour suspension to 10-13, and sequentially performing a first stirring treatment and a first centrifugation to obtain a rice flour supernatant;

[0028] Mixing defatted pea flour with water to obtain a pea flour suspension; adjusting the pH value of the pea flour suspension to 8-11, and sequentially performing a second stirring treatment and a second centrifugation to obtain a pea flour supernatant;

[0029] The rice flour supernatant and the pea flour supernatant are mixed to obtain a first mixed solution; the pH value of the first mixed solution is adjusted to 12-13, and a third stirring process is performed to obtain a second mixed solution; the pH value of the second mixed solution is adjusted to 6.5-7.5, and a fourth stirring process is performed, and then standing and a third centrifugation are performed in sequence to obtain a mixed supernatant;

[0030] The mixed supernatant is dialyzed and freeze-dried in sequence to obtain the rice-pea protein complex.

[0031] Pea protein is rich in amino acids, including lysine, arginine, phenylalanine, leucine and isoleucine. Pea protein is cheap and has no problems such as genetic modification, but the inherent beany smell, low digestibility and lack of sulfur-containing amino acids (methionine, cysteine) of pea protein make it a challenging task to add pea protein to food. The nutritional and functional properties of pea protein and rice protein can be complementary, and the combination of the two proteins can solve the shortcomings of each of them. In the related art, a pH cycle method is used to prepare a plant protein complex. The principle is to adjust the pH value away from neutrality so that the protein structure has the opportunity to curl randomly, and then slowly restore the pH value to neutrality. The non-covalent action allows the proteins from two different sources to form a common intermolecular main chain structure. However, the above method usually requires the extraction of the corresponding proteins separately, and then the extracted proteins are compounded to prepare the complex. The whole process is very complicated and requires repeated acid and alkali adjustment. A large amount of acid and alkali will pollute the environment. The cost is high in industry and it is difficult to achieve industrial application. The present invention adopts a method of compounding pea protein with rice protein to improve the nutritional and functional properties of rice protein, which is a green and harmless protein modification method. Specifically, the present invention adopts a "one-step method" to prepare a rice-pea protein complex, and there is no need to extract rice protein and pea protein separately before compounding. The preparation process is simple, the pollution to the environment is small, the energy consumption is low, and it is suitable for large-scale production. The method of the present invention is described in detail below.

[0032] In the present invention, unless otherwise specified, the raw materials used are commercially available products well known to those skilled in the art or are prepared by methods well known to those skilled in the art.

[0033] The present invention mixes rice flour with water to obtain a rice flour suspension; adjusts the pH value of the rice flour suspension to 10-13, and sequentially performs a first stirring treatment and a first centrifugation to obtain a rice flour supernatant. As an embodiment of the present invention, the rice flour can be obtained by milling rice, and the particle size of the rice flour can be ≤178 μm; the solid-liquid ratio of rice flour to water in the rice flour suspension can be 1g:8-12mL, further can be 1g:9-11mL, specifically can be 1g:10mL. As an embodiment of the present invention, the reagent used to adjust the rice flour suspension can be a NaOH solution, and the concentration of the NaOH solution can be 0.8-1.2mol / L, specifically can be 1mol / L; the present invention adjusts the pH value of the rice flour suspension to 10-13, specifically can be 10, 10.5, 11, 11.5, 12, 12.5 or 13. As an embodiment of the present invention, the time of the first stirring treatment can be 3 to 5 hours, specifically 3 hours, 3.5 hours, 4 hours, 4.5 hours or 5 hours; the speed of the first centrifugation can be 6000 to 8000 rpm, specifically 6000 rpm, 6500 rpm, 7000 rpm, 7500 rpm or 8000 rpm; the time can be 15 to 20 minutes. The present invention adjusts the pH value of the rice flour suspension to 10 to 13, which can fully dissolve the rice protein in the rice flour; in the embodiment of the present invention, the first stirring treatment is carried out under the above conditions, which can accelerate the dissolution rate of the rice protein, which is conducive to the full dissolution of the rice protein in the rice flour. As an embodiment of the present invention, the protein concentration in the rice flour supernatant can be 5 to 10 mg / mL, specifically 5 mg / mL, 6 mg / mL, 7 mg / mL, 8 mg / mL, 9 mg / mL or 10 mg / mL.

[0034] The present invention mixes defatted pea flour with water to obtain a pea flour suspension; adjusts the pH value of the pea flour suspension to 8-11, and sequentially performs a second stirring treatment and a second centrifugation to obtain a pea flour supernatant. As an embodiment of the present invention, the defatted pea flour can be obtained by defatting pea flour with n-hexane; the pea flour can be obtained by grinding peas, and the particle size of the pea flour can be ≤178μm. As an embodiment of the present invention, the defatting conditions include: the solid-liquid ratio of pea flour to n-hexane can be 1g:5-10mL, specifically 1g:5mL, 1g:6mL, 1g:7mL, 1g:8mL, 1g:9mL or 1g:10mL; the time can be 3-4h, specifically 3h, 3.5h or 4h; the temperature can be room temperature; the defatting can be carried out under stirring conditions. As an embodiment of the present invention, after the defatting treatment, the process further comprises: filtering the liquid obtained after the defatting treatment, collecting the filter cake and placing it in a fume hood for air drying to obtain the defatted pea powder.

[0035] As an embodiment of the present invention, the liquid ratio of defatted pea flour to water in the pea flour suspension can be 1g:8-12mL, further can be 1g:9-11mL, specifically can be 1g:10mL. As an embodiment of the present invention, the reagent used to adjust the pea flour suspension can be a NaOH solution, and the concentration of the NaOH solution can be 0.8-1.2mol / L, specifically can be 1mol / L; the present invention adjusts the pH value of the pea flour suspension to 8-11, specifically can be 8, 8.5, 9, 9.5, 10, 10.5 or 11. As an embodiment of the present invention, the time of the second stirring treatment can be 3-5h, specifically can be 3h, 3.5h, 4h, 4.5h or 5h; the speed of the second centrifugation can be 6000-8000rpm, specifically can be 6000rpm, 6500rpm, 7000rpm, 7500rpm or 8000rpm; the time can be 15-20min. The present invention adjusts the pH value of the pea flour suspension to 8-11, which can fully dissolve the pea protein in the defatted pea flour; in the embodiment of the present invention, the second stirring treatment is performed under the above conditions, which can accelerate the dissolution rate of the pea protein, which is conducive to the full dissolution of the pea protein in the defatted pea flour. As an embodiment of the present invention, the protein concentration in the pea flour supernatant can be 22-26 mg / mL, specifically 22 mg / mL, 23 mg / mL, 24 mg / mL, 25 mg / mL or 26 mg / mL.

[0036] After obtaining the rice flour supernatant and the pea flour supernatant, the present invention mixes the rice flour supernatant with the pea flour supernatant to obtain a first mixed solution. As one embodiment of the present invention, the mass ratio of rice protein to pea protein in the first mixed solution can be 9:1 to 1:9, specifically 9:1, 8:2, 7:3, 6:4, 5:5, 4:6, 3:7, 2:8 or 1:9. The present invention can dilute the pea flour supernatant with water to the same protein concentration as the rice flour supernatant, and then mix the rice flour supernatant with the pea flour dilution obtained after dilution in a volume ratio of 9:1 to 1:9 to obtain a first mixed solution. As one embodiment of the present invention, the total concentration of rice protein and pea protein in the first mixed solution can be 5 to 10 mg / mL, specifically 5 mg / mL, 6 mg / mL, 7 mg / mL, 8 mg / mL, 9 mg / mL or 10 mg / mL. In the embodiment of the present invention, limiting the mass ratio of rice protein to pea protein to the above range is conducive to making the rice-pea protein complex have a higher solubility, especially when the mass ratio of rice protein to pea protein is 1:9, a rice-pea protein complex with higher solubility can be obtained.

[0037] After obtaining the first mixed solution, the present invention adjusts the pH value of the first mixed solution to 12-13, performs a third stirring treatment, and obtains a second mixed solution. As one embodiment of the present invention, the reagent used to adjust the first mixed solution may be a NaOH solution, and the concentration of the NaOH solution may be 0.8-1.2 mol / L, specifically 1 mol / L; the present invention adjusts the pH value of the first mixed solution to 12-13, specifically 12, 12.3, 12.5, 12.8 or 13. As one embodiment of the present invention, the time of the third stirring treatment may be 1-3h, specifically 1h, 1.5h, 2h, 2.5h or 3h. The present invention adjusts the pH value of the first mixed solution to 12-13, which can open the protein structure; in the embodiment of the present invention, the third stirring treatment is performed under the above conditions, which can increase the reaction area between rice protein and pea protein, which is beneficial to promote the interaction between the two.

[0038] After obtaining the second mixed solution, the present invention adjusts the pH value of the second mixed solution to 6.5-7.5, performs a fourth stirring treatment, and then sequentially performs standing and a third centrifugation to obtain a mixed supernatant. As an embodiment of the present invention, the reagent used to adjust the second mixed solution can be an HCl solution, and the concentration of the HCl solution can be 0.08-0.12 mol / L, specifically 0.1 mol / L; the present invention adjusts the pH value of the second mixed solution to 6.5-7.5, specifically 6.5, 6.8, 7, 7.2 or 7.5. As an embodiment of the present invention, the time of the fourth stirring treatment can be 0.5-1.5h, specifically 0.5h, 1h or 1.5h; the standing time can be 12-24h, specifically 12h, 14h, 16h, 18h, 20h, 22h or 24h. As an embodiment of the present invention, the rotation speed of the third centrifugation can be 7000-8000 rpm, specifically 7000 rpm, 7500 rpm or 8000 rpm; the time can be 15-20 min. The present invention adjusts the pH value of the second mixed solution to 6.5-7.5, which can refold the protein with the opened structure; in the embodiment of the present invention, the fourth stirring treatment is carried out under the above conditions, which can increase the reaction area between the rice protein and the pea protein, which is conducive to promoting the interaction between the two.

[0039] After obtaining the mixed supernatant, the present invention dialyzes and freeze-dries the mixed supernatant in sequence to obtain the rice-pea protein complex. As one embodiment of the present invention, the molecular weight cutoff of the dialysis bag used for dialysis can be specifically 3400Da; the dialysate used for dialysis can be water; the dialysis time can be 20 to 28 hours, specifically 24 hours. As one embodiment of the present invention, the freeze-drying conditions can include: the cold trap temperature can be -70 to -60°C, specifically -65°C; the vacuum pressure can be 0.04 to 0.06Pa, specifically 0.05Pa; the freeze-drying time can be 45 to 50 hours, specifically 48 hours.

[0040] As an embodiment of the present invention, during the stirring process involved in preparing the rice-pea protein complex by the above method, the stirring rate can be independently 700-900 rpm, specifically 800 rpm.

[0041] The present invention utilizes chemical auxiliary means (adjusting pH value) to compound rice protein and pea protein together to form a new protein structure, thereby improving the nutrition and functional characteristics of rice protein and pea protein. The rice-pea protein complex prepared by the inventive method has higher solubility and balanced amino acid composition, and can be used as a high-quality plant protein source in the food industry, solves the problem of low water solubility of rice protein and poor digestibility of pea protein, helps to expand the application field of rice protein and pea protein, improves the utilization rate of protein, and increases the economic benefits of enterprises. Moreover, the inventive method has the characteristics of simple extraction, green environmental protection, and high safety.

[0042] The present invention provides a rice-pea protein complex prepared by the preparation method described in the above technical solution. As an embodiment of the present invention, the solubility of the rice-pea protein complex can be 35-90%, specifically 35%, 38%, 45%, 51%, 55%, 58%, 62%, 67%, 75%, 81% or 90%; the particle size (specifically the average particle size) of the rice-pea protein complex can be 50-200nm, further 75-150nm, further 89-143nm, further 101-125nm, and further 110-115nm.

[0043] The present invention provides application of the rice-pea protein complex described in the above technical solution in food.

[0044] The present invention uses rice and peas as raw materials, and by adjusting the pH value, the solubility of rice protein, the digestibility of pea protein and other functional properties are significantly improved, providing a scientific idea and theoretical basis for the further development and utilization of rice protein and pea protein. The preparation method of the present invention has the characteristics of simple operation, high safety, and green environmental protection, and can improve the resource utilization rate of rice and peas and expand the economic benefits of enterprises. The rice-pea protein complex prepared by the present invention has good functional properties, and provides a scientific idea for the rice-pea protein complex as a high-quality plant protein source to produce high-value-added products with excellent functions.

[0045] Figure 1 The process flow chart for preparing the rice-pea protein complex in the embodiment of the present invention is shown below. The technical scheme in the present invention will be clearly and completely described in combination with the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0046] The rice used in the following experiments are all raw materials from the same batch, and the peas used are all raw materials from the same batch.

[0047] The magnetic stirring rate in the following experiments was 800 rpm.

[0048] In the following experiments, the Kjeldahl method was used to determine protein purity, and the conversion factor was 6.25.

[0049] The solubility determination method of the rice-pea protein complex prepared in the following experiment is as follows:

[0050] The protein complex sample was dissolved in ultrapure water to prepare a protein suspension with a concentration of 1 mg / mL. After stirring for 1 hour to fully disperse it, it was centrifuged at 10,000 rpm for 15 minutes. The supernatant was taken to determine the protein concentration using the BCA method;

[0051] The protein solubility was calculated by the following formula:

[0052] Solubility (%) = c·V / ρ·m;

[0053] Wherein, c is the protein concentration in the supernatant (mg / mL), V is the volume of the protein suspension (mL), ρ is the protein purity of the protein complex sample (%), and m is the mass of the protein complex sample used to prepare the protein suspension (mg).

[0054] The particle size determination method of the rice-pea protein complex prepared in the following experiment is as follows:

[0055] The protein complex sample was dissolved in ultrapure water to prepare a protein suspension with a concentration of 1 mg / mL. After stirring for 1 hour to fully disperse it, it was centrifuged at 10,000 rpm for 15 minutes. The supernatant was taken and the protein particle size was measured using a particle size measuring instrument.

[0056] Example 1

[0057] (1) Grinding rice and peas respectively to collect rice flour and pea flour with a particle size of ≤178 μm; mixing the pea flour with n-hexane at a solid-liquid ratio of 1 g:5 mL, defatting the mixture at room temperature (25° C.) under magnetic stirring for 3 h, filtering the mixture, and air-drying the filter cake in a fume hood to obtain defatted pea flour;

[0058] (2) The rice flour and defatted pea flour were respectively mixed with ultrapure water at a solid-liquid ratio of 1 g: 10 mL to obtain a rice flour suspension and a pea flour suspension, and the pH value of the rice flour suspension was adjusted to 12 with a 1 mol / L NaOH solution, and the pH value of the pea flour suspension was adjusted to 9 with a 1 mol / L NaOH solution, and the mixtures were magnetically stirred for 3 h, and then centrifuged at 6000 rpm for 15 min, respectively, to collect the rice flour supernatant (protein concentration was 8 mg / mL) and the pea flour supernatant (protein concentration was 24 mg / mL);

[0059] (3) The pea flour supernatant was diluted with ultrapure water to the same protein concentration as the rice flour supernatant (protein concentration of 8 mg / mL), the rice flour supernatant and the diluted pea flour dilution were mixed at a volume ratio of 9:1, the pH value of the mixed solution was adjusted to 12 with a 1 mol / L NaOH solution, and magnetic stirring was performed for 1 hour. Then, the pH value of the system was adjusted to 7 with a 0.1 mol / L HCl solution, and magnetic stirring was performed for 0.5 hour and then allowed to stand for 12 hours. Then, the mixture was centrifuged at 7000 rpm for 20 minutes, the supernatant was collected, and the supernatant was dialyzed for 24 hours using a dialysis bag with a molecular weight cutoff of 3400 Da and ultrapure water as a dialysate to remove salt, and then freeze-dried for 48 hours at a cold trap temperature of -65°C and a vacuum pressure of 0.05 Pa to obtain a rice-pea protein complex (denoted as A1 complex).

[0060] Example 2

[0061] The method of Example 1 was used for operation, except that the rice flour supernatant and the diluted pea flour dilution were mixed in a volume ratio of 7:3, and the obtained rice-pea protein complex was recorded as A2 complex.

[0062] Example 3

[0063] The method of Example 1 was used for operation, except that the rice flour supernatant and the diluted pea flour dilution were mixed at a volume ratio of 5:5, and the obtained rice-pea protein complex was recorded as A3 complex.

[0064] Example 4

[0065] The method of Example 1 was used for operation, except that the rice flour supernatant and the diluted pea flour dilution were mixed at a volume ratio of 3:7, and the obtained rice-pea protein complex was recorded as A4 complex.

[0066] Example 5

[0067] The method of Example 1 was used for operation, except that the rice flour supernatant and the diluted pea flour dilution were mixed at a volume ratio of 1:9, and the obtained rice-pea protein complex was recorded as A5 complex.

[0068] Comparative Examples 1 to 5

[0069] The preparation of the rice protein-pea protein mixture comprises the following steps:

[0070] (1) Grinding rice to collect rice flour with a particle size of ≤178 μm; mixing the rice flour with ultrapure water at a solid-liquid ratio of 1 g:10 mL to obtain a rice flour suspension; adjusting the pH value of the rice flour suspension to 12 with a 1 mol / L NaOH solution, magnetically stirring for 3 h, and then centrifuging at 6000 rpm for 15 min to collect the rice flour supernatant; adjusting the pH value of the rice flour supernatant to 12 with a 1 mol / L NaOH solution, magnetically stirring for 3 h, and then centrifuging at 6000 rpm for 15 min to collect the rice flour supernatant; Magnetic stirring was carried out for 2h, the pH value of the obtained system was adjusted to 7 by the HCl solution of 0.1mol / L using concentration afterwards, left standstill for 18h after magnetic agitation 1h, centrifugal for 15min under 8000rpm condition afterwards, supernatant was collected, the dialysis bag adopting molecular weight cut-off was 3400Da, with ultrapure water as the dialysate that described supernatant was dialyzed for 24h to remove salt, afterwards in cold trap temperature, be-65 ℃ and vacuum pressure be freeze drying 48h under 0.05Pa condition, obtain rice protein (being designated as B# rice protein);

[0071] (2) Grinding peas to collect pea flour with a particle size of ≤178 μm, mixing the pea flour with n-hexane at a solid-liquid ratio of 1 g:5 mL, defatting for 3 h at room temperature (25°C) and under magnetic stirring, then filtering, and air-drying the filter cake in a fume hood to obtain defatted pea flour; mixing the defatted pea flour with ultrapure water at a solid-liquid ratio of 1 g:10 mL to obtain a pea flour suspension, adjusting the pH value of the pea flour suspension to 9 with a NaOH solution with a concentration of 1 mol / L, magnetically stirring for 3 h, and then centrifuging at 6000 rpm for 20 min to collect the pea flour suspension. clear liquid; the pH value of the supernatant was adjusted to 12 with a NaOH solution having a concentration of 1 mol / L, and magnetically stirred for 2 h, and then the pH value of the obtained system was adjusted to 7 with a HCl solution having a concentration of 0.1 mol / L, and magnetically stirred for 1 h and then allowed to stand for 18 h, and then centrifuged at 8000 rpm for 15 min, the supernatant was collected, and a dialysis bag with a molecular weight cutoff of 3400 Da was used to dialyze the supernatant for 24 h with ultrapure water as a dialysate to remove salt, and then freeze-dried for 48 h at a cold trap temperature of -65°C and a vacuum pressure of 0.05 Pa to obtain pea protein (denoted as B# pea protein);

[0072] (3) The rice protein and the pea protein are mixed in a mass ratio of 9:1, 7:3, 5:5, 3:7, and 1:9 to obtain rice protein-pea protein mixtures, which are respectively recorded as B1 to B5 mixtures.

[0073] Comparative Examples 6 to 10

[0074] The rice-pea protein complex is prepared by a pH cycle method, comprising the following steps:

[0075] (1) grinding rice to collect rice flour with a particle size of ≤178 μm; mixing the rice flour with ultrapure water at a solid-liquid ratio of 1 g:10 mL to obtain a rice flour suspension; adjusting the pH value of the rice flour suspension to 12 with a 1 mol / L NaOH solution, stirring magnetically for 3 h, and then centrifuging at 6000 rpm for 15 min to collect a rice flour supernatant; adjusting the pH value of the rice flour supernatant to 4.5 (isoelectric point) with a 1 mol / L HCl solution, and then centrifuging at 8000 rpm for 10 min, collecting a precipitate and washing it with distilled water for 4 times, and then freeze-drying it at a cold trap temperature of -65°C and a vacuum pressure of 0.05 Pa for 48 h to obtain rice protein (denoted as C# rice protein);

[0076] (2) Grinding peas to collect pea flour with a particle size of ≤178 μm, mixing the pea flour with n-hexane at a solid-liquid ratio of 1 g:5 mL, defatting for 3 h at room temperature (25° C.) and under magnetic stirring, filtering, and air-drying the filter cake in a fume hood to obtain defatted pea flour; mixing the defatted pea flour with ultrapure water at a solid-liquid ratio of 1 g:10 mL to obtain a pea flour suspension, and adjusting the pH of the pea flour suspension using a 1 mol / L NaOH solution. The pH value was adjusted to 9, magnetic stirring was performed for 3 hours, and then centrifuged at 6000 rpm for 20 minutes to collect the pea powder supernatant; the pH value of the pea powder supernatant was adjusted to 4.5 (isoelectric point) using a 1 mol / L HCl solution, and then centrifuged at 8000 rpm for 10 minutes, the precipitate was collected and washed with distilled water for 4 times, and then freeze-dried at a cold trap temperature of -65°C and a vacuum pressure of 0.05 Pa for 48 hours to obtain pea protein (denoted as C# pea protein);

[0077] (3) The rice protein and pea protein are mixed in a mass ratio of 9:1, 7:3, 5:5, 3:7, and 1:9, respectively, and then ultrapure water is added to obtain a suspension with a total concentration of rice protein and pea protein of 0.01 g / mL; the pH value of the suspension is adjusted to 12 using a 1 mol / L NaOH solution, and magnetic stirring is performed for 1 hour, and then the pH value of the obtained system is adjusted to 7 using a 0.1 mol / L HCl solution, and magnetic stirring is performed for 0.5 hour, and then centrifuged at 8000 rpm for 15 minutes, the supernatant is collected, and a dialysis bag with a molecular weight cutoff of 3400 Da is used to dialyze the supernatant for 24 hours with ultrapure water as a dialysate to remove salt, and then freeze-dried for 48 hours at a cold trap temperature of -65°C and a vacuum pressure of 0.05 Pa to obtain a rice-pea protein complex, which is respectively recorded as C1 to C5 complexes.

[0078] Test Example 1 Determination of solubility

[0079] The solubility of the complexes prepared in Examples 1 to 5 (A1 to A5 complexes), the rice protein (B# rice protein), pea protein (B# pea protein) and the mixture (B1 to B5 mixture) prepared in Comparative Examples 1 to 5, and the complexes prepared in Comparative Examples 6 to 10 (C1 to C5 complexes) were tested. The results are shown in Table 1, where RP represents rice protein, PP represents pea protein, and the ratio of the two is a mass ratio; the solubility data in Table 1 are expressed as mean ± standard deviation; a, b, c, and d in Table 1 represent significant differences (p < 0.05) in solubility between protein samples of different compositions. As shown in Table 1, there is a significant difference in solubility between the uncompounded rice protein-pea protein mixture and the rice-pea protein complex in Examples 1 to 5; compared with the rice protein-pea protein mixture, the solubility of the rice-pea protein complex in Examples 1 to 5 increased by 4.44%, 11.84%, 12.12%, 15.01%, and 23.22%, respectively; compared with the rice-pea protein complex prepared by the pH circulation method, the solubility of the rice-pea protein complex in Examples 1 to 5 increased by 1.3%, 2.6%, 1.9%, 2.8%, 3.2%, 4.6%, 5.9%, 6.8%, 7.9%, 8.8%, 9.7%, 10.9%, 11.1%, 12.2%, 13.0%, 14.1%, 15.2%, 16.3%, 17.4%, 18.9%, 19.8%, 20.9%, 21.8%, 22.9%, 34.7%, 35.9%, 36.8%, 37.9%, 38. The solubility of the compound increased by 2.12%, 2.39%, 2.37%, 3.95%, and 13.48% respectively; compared with rice protein (B# rice protein), the solubility of the rice-pea protein complex in Examples 1 to 5 increased by 7.51%, 21.03%, 27.44%, 36.45%, and 50.78% respectively; compared with pea protein (B# pea protein), the solubility of the rice-pea protein complex in Examples 4 to 5 increased by 5.82% and 20.15% respectively. It is shown that compared with simply mixing the two proteins, the solubility of the rice-pea protein complex after compounding by the method of the present invention is greatly improved; the solubility of the rice-pea protein complex prepared by the present invention is comparable to that of the rice-pea protein complex prepared by the existing pH circulation method, and even reaches a better level. At the same time, the method of the present invention can improve the disadvantage of poor solubility of rice protein and improve the solubility of pea protein to a certain extent after compounding pea protein with rice protein.

[0080] Table 1 Solubility of rice protein, pea protein, complex and mixture in Examples and Comparative Examples (%)

[0081] Protein composition Examples 1 to 5 Comparative Examples 1 to 5 Comparative Examples 6 to 10 RP / <![CDATA[30.15±2.41 d ]]> / RP:PP=9:1 <![CDATA[37.66±2.61 d ]]> <![CDATA[33.22±1.97 d ]]> <![CDATA[35.54±2.23 d ]]> RP:PP=7:3 <![CDATA[51.18±2.83 c ]]> <![CDATA[39.34±1.46 cd ]]> <![CDATA[48.79±2.02 c ]]> RP:PP=5:5 <![CDATA[57.59±1.64 bc ]]> <![CDATA[45.47±1.78 bc ]]> <![CDATA[55.22±1.29 bc ]]> RP:PP=3:7 <![CDATA[66.60±4.21 b ]]> <![CDATA[51.59±2.66 ab ]]> <![CDATA[62.65±1.93 ab ]]> RP:PP=1:9 <![CDATA[80.93±4.68 a ]]> <![CDATA[57.71±3.71 a ]]> <![CDATA[67.45±2.35 a ]]> PP / <![CDATA[60.78±4.26 ac ]]> /

[0082] Test Example 2 Determination of Particle Size

[0083] The particle sizes of the composites prepared in Examples 1 to 5 (A1 to A5 composites), the rice protein (B# rice protein), the pea protein (B# pea protein) and the mixture (B1 to B5 mixture) prepared in Comparative Examples 1 to 5 were measured. Figure 2The particle size distribution diagrams of the A1 complex in Example 1, the B1 mixture in Comparative Example 1, the rice protein (B# rice protein), and the pea protein (B# pea protein), are shown in FIG. Figure 3 The particle size distribution diagrams of the A2 complex in Example 2, the B2 mixture in Comparative Example 2, the rice protein (B# rice protein), and the pea protein (B# pea protein), are shown in FIG. Figure 4 The particle size distribution diagrams of the A3 complex in Example 3, the B3 mixture in Comparative Example 3, the rice protein (B# rice protein), and the pea protein (B# pea protein), are shown in FIG. Figure 5 The particle size distribution diagrams of the A4 complex in Example 4, the B4 mixture in Comparative Example 4, the rice protein (B# rice protein), and the pea protein (B# pea protein), are shown in FIG. Figure 6 The particle size distribution diagrams of the A5 complex in Example 5, the B5 mixture in Comparative Example 5, the rice protein (B# rice protein), and the pea protein (B# pea protein) are shown. The results show that the particle size distribution of the rice-pea protein complex in Examples 1 to 5 is more concentrated, the particle size distribution of the rice protein-pea protein mixture is more dispersed, and the particle size distribution of the rice-pea protein complex in Examples 1 to 5 is between rice protein and pea protein. This indicates that the rice protein and pea protein in the complex prepared by the present invention interact with each other, resulting in a new protein complex structure.

[0084] The average particle sizes of rice protein (B# rice protein), pea protein (B# pea protein) and the complexes (A1-A5 complexes) prepared in Examples 1-5 are shown in Table 2, where the average particle size data are expressed as mean ± standard deviation; a, b, c, d, e, and f in Table 2 represent significant differences in average particle size between different protein sources (p < 0.05). As can be seen from Table 2, the particle size of the rice-pea protein complex in Examples 1-5 is between the particle sizes of rice protein (B# rice protein) and pea protein (B# pea protein), and it is speculated that this is the direct reason for the increased solubility of the rice-pea protein complex, because the smaller the protein particle size, the easier it is to disperse in water, and thus more soluble in water.

[0085] Table 2 Average particle size of rice protein and pea protein in the composites of the examples and the comparative examples

[0086] Protein Source Average particle size (nm) Rice protein(B#rice protein) <![CDATA[150.07±4.75 a ]]> Example 1 <![CDATA[142.77±1.72 b ]]> Example 2 <![CDATA[114.93±1.18 c ]]> Example 3 <![CDATA[109.53±2.12 c ]]> Example 4 <![CDATA[100.94±1.44 d ]]> Example 5 <![CDATA[89.45±1.02 e ]]> Pea protein (B# pea protein) <![CDATA[78.24±0.32 f ]]>

[0087] Embodiments 6 to 10

[0088] The methods of Examples 1 to 5 were used respectively, except that in step (3), when a 0.1 mol / L HCl solution was used to adjust the pH value of the system, the pH value was adjusted from 7 to 6.5.

[0089] Examples 11 to 15

[0090] The methods of Examples 1 to 5 were used respectively, except that in step (3), when a 0.1 mol / L HCl solution was used to adjust the pH value of the system, the pH value was adjusted from 7 to 7.5.

[0091] The solubility of the complexes in Examples 6 to 15 was measured and compared with the solubility of the complexes in Examples 1 to 5. The results are shown in Table 3. In Table 3, RP represents rice protein and PP represents pea protein; the solubility data in Table 3 are expressed as mean ± standard deviation; a, b, c, and d in Table 3 represent significant differences in solubility between different protein samples (p < 0.05). As shown in Table 3, the solubility of the rice-pea protein complex prepared by adjusting the pH value to 7 for the second time after mixing the two supernatants in step (3) is significantly higher than the solubility of the rice-pea protein complex prepared by adjusting the pH value to 6.5 or 7.5. Therefore, the present invention preferably applies the second adjustment of the pH value to 7 after mixing the two supernatants in step (3) to the preparation of the rice-pea protein complex.

[0092] Table 3 Effect of different pH values ​​on the solubility (%) of rice-pea protein complexes with different protein ratios

[0093]

[0094] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for preparing a rice-pea protein complex, comprising the following steps: Mixing rice flour with water to obtain a rice flour suspension; adjusting the pH value of the rice flour suspension to 10-13, and sequentially performing a first stirring treatment and a first centrifugation to obtain a rice flour supernatant; Mixing defatted pea flour with water to obtain a pea flour suspension; adjusting the pH value of the pea flour suspension to 8-11, and sequentially performing a second stirring treatment and a second centrifugation to obtain a pea flour supernatant; Mixing the rice flour supernatant with the pea flour supernatant to obtain a first mixed solution; The pH value of the first mixed solution is adjusted to 12-13, and a third stirring process is performed to obtain a second mixed solution; the pH value of the second mixed solution is adjusted to 6.5-7.5, and a fourth stirring process is performed, and then standing and a third centrifugation are performed in sequence to obtain a mixed supernatant; The mixed supernatant is dialyzed and freeze-dried in sequence to obtain the rice-pea protein complex.

2. The preparation method according to claim 1, characterized in that: The particle size of the rice flour is ≤178 μm; the material-liquid ratio of the rice flour to water in the rice flour suspension is 1 g:8-12 mL.

3. The preparation method according to claim 1, characterized in that: The defatted pea flour is obtained by defatting pea flour with n-hexane; the particle size of the pea flour is ≤178 μm; and the solid-liquid ratio of the defatted pea flour to water in the pea flour suspension is 1 g:8-12 mL.

4. The preparation method according to any one of claims 1 to 3, characterized in that: The time of the first stirring treatment and the second stirring treatment is independently 3 to 5 hours; the speed of the first centrifugation and the second centrifugation is independently 6000 to 8000 rpm, and the time is independently 15 to 20 minutes.

5. The preparation method according to claim 1, characterized in that: The mass ratio of rice protein to pea protein in the first mixed solution is 9:1 to 1:

9.

6. The preparation method according to claim 1, characterized in that: The third stirring treatment lasts for 1 to 3 hours.

7. The preparation method according to claim 1, characterized in that: The fourth stirring treatment time is 0.5 to 1.5 hours; the standing time is 12 to 24 hours; the third centrifugation speed is 7000 to 8000 rpm, and the time is 15 to 20 minutes.

8. The rice-pea protein complex prepared by the preparation method according to any one of claims 1 to 7.

9. The rice-pea protein complex according to claim 8, characterized in that The solubility of the rice-pea protein complex is 35-90%, and the particle size is 50-200nm.

10. Use of the rice-pea protein complex according to claim 8 or 9 in food.

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