Extraction method of black rice bran protein and protein supplement

Abstract

The invention discloses an extraction method of black rice bran protein and a protein supplement, and relates to the technical field of food processing.The extraction method of the black rice bran protein comprises the following steps that unhulled black rice is husked, milled and smashed, and black rice bran powder is obtained; degreasing with normal hexane to obtain degreased black rice bran; the preparation method comprises the following steps: mixing degreased black rice bran with water, adjusting the pH value, adding one or more of cellulase, saccharifying enzyme, amylase and phytase for enzymolysis, and centrifuging to obtain a black rice bran protein precipitate; stirring and extracting black rice bran protein with alkali liquor, and centrifuging to obtain a black rice bran protein crude extract; adding a hydrogen peroxide solution into the solution, adjusting the pH value, and decolorizing; and adjusting the decolorized black rice bran protein extract to an isoelectric point pH value, centrifuging, and drying the obtained precipitate to obtain the black rice bran protein. The extraction method is simple and effective, the extraction rate is high, and the obtained black rice bran protein is high in purity, good in quality, light in color and complete in protein structure.

Description

Technical Field

[0001] The invention relates to the technical field of food processing, and in particular to a method for extracting black rice bran protein and a protein supplement. Background Art

[0002] Black rice has become a staple food to replace rice because of its rich nutrients and bioactive ingredients, as well as its unique sensory properties. However, black rice tastes worse than white rice and is difficult to digest. Moderate milling can improve the edible quality and color of black rice. Black rice bran is a byproduct of the endosperm of black rice and brown rice during the rice milling process. Compared with ordinary rice bran, black rice bran has higher content of nutrients and bioactive ingredients, more reasonable amino acid composition, and has multiple physiological functions such as antioxidant, anti-inflammatory, and prevention of cardiovascular diseases. It can be used as a functional food raw material with broad application prospects. However, most black rice bran is currently used as livestock and poultry feed or organic fertilizer, and some black rice bran is used to extract anthocyanins to develop sports nutrition products. There are few high-value-added black rice bran products, and the overall deep processing level of black rice bran resources is low.

[0003] Black rice bran protein is a high-quality plant protein with a protein content of about 12-20%, which is higher than ordinary rice bran. Black rice bran protein has the advantages of low allergy, high lysine content, and reasonable amino acid composition, which is comparable to egg and meat protein. It is very suitable as a protein supplement for infants, adolescents or special groups. Therefore, fully developing and utilizing the protein resources of black rice bran and carrying out the development and application research of black rice bran protein products can achieve high-value utilization of black rice bran, reduce the consumption pressure of animal protein, and have important social and economic value.

[0004] At present, the methods used for protein extraction mainly include alkaline method and enzyme method. However, the structural integrity of black rice bran protein extracted by enzyme method is poor, and the purity of black rice bran protein extracted by alkaline method is low. Summary of the invention

[0005] The main purpose of the invention is to provide a method for extracting black rice bran protein and a protein supplement, aiming to solve the problems of low utilization rate of existing black rice bran, low purity of black rice bran protein, dark color and limited application.

[0006] To achieve the above object, the present invention provides a method for extracting black rice bran protein, comprising the following steps:

[0007] S10, hulling black rice to obtain black rice, grinding the black rice layer by layer from the outer layer to the inner layer and sieving the grinding product to obtain black rice bran with a grinding rate of 5 to 10%; crushing the black rice bran to obtain black rice bran powder with a mesh size of 200 meshes;

[0008] S20, mixing the black rice bran powder with n-hexane, stirring, and filtering, repeating 2 to 4 times to obtain defatted black rice bran;

[0009] S30, mixing the defatted black rice bran with water to obtain a mixed solution, adjusting the pH value of the mixed solution to the enzymatic pH, adding one or more of cellulase, saccharifying enzyme, amylase, and phytase to the system to perform an enzymatic reaction; after the enzymatic reaction is completed, washing the precipitate with water, and centrifuging at 4000-10000×g for 15-30 min to obtain a precipitate containing black rice bran protein;

[0010] S40, mixing the precipitate with an alkaline solution, stirring and extracting, centrifuging at 4000-10000×g for 15-30 min, collecting the supernatant, and obtaining a crude extract containing black rice bran protein;

[0011] S50, mixing the crude extract with a hydrogen peroxide solution, adjusting the pH of the system to a decolorization reaction pH, stirring for decolorization, and centrifuging at 4000-10000×g for 15-30 min, collecting the supernatant, and obtaining a decolorized solution;

[0012] S60, mixing the decolorized solution with an acid, adjusting the pH of the system to the isoelectric point of the black rice bran protein, and then centrifuging at 4000-10000×g for 15-30 min, taking the precipitate, and drying to obtain the black rice bran protein.

[0013] In one embodiment, in step S20: the solid-liquid ratio of the black rice bran powder to the n-hexane is 1:3-15; and / or,

[0014] The stirring time is 2 to 8 hours.

[0015] In one embodiment, in step S30: the solid-liquid ratio of the defatted black rice bran to water is 1:10-20; and / or,

[0016] Based on the mass of defatted black rice bran, the added amount of the cellulase, saccharifying enzyme, amylase and phytase is 0.1-1%; and / or,

[0017] The enzymatic hydrolysis temperature is 30-60°C; and / or,

[0018] The enzymatic hydrolysis time is 1 to 6 hours; and / or,

[0019] The pH of the enzymatic hydrolysis is 2.5-8.

[0020] In one embodiment, in step S40: the pH of the alkaline solution is 8 to 12; and / or,

[0021] The stirring time is 1 to 6 hours.

[0022] In one embodiment, in step S50, the amount of hydrogen peroxide solution added is 0.5-8% based on the mass of the crude extract; and / or,

[0023] The decolorization temperature is 20-60°C; and / or,

[0024] The decolorization time is 0.5 to 4 hours; and / or,

[0025] The pH of the decolorization is 6-11.

[0026] The present invention also provides a protein supplement, which comprises black rice bran protein extracted by the above-mentioned black rice bran protein extraction method.

[0027] The technical solution of the present invention has the following beneficial effects:

[0028] (1) The present invention develops a low-cost, high-efficiency method for extracting black rice bran protein, realizes the development of high-quality black rice bran protein resources, develops a new type of plant protein, alleviates the consumption pressure of animal protein, achieves sustainable development, and reduces environmental pollution and resource waste.

[0029] (2) The present invention adopts enzymatic pretreatment assisted alkaline method to extract black rice bran protein, and further decolorizes with hydrogen peroxide solution. The obtained black rice bran protein has a high extraction rate and high purity, and the quality of the obtained black rice bran protein is good, and its protein structure is more complete; the process is simple and effective, and the obtained black rice bran protein product is lighter in color.

[0030] (3) In the process of the present invention, hydrogen peroxide is used for decolorization. Hydrogen peroxide is a cheap, safe and environmentally friendly oxidative decolorizing agent, which effectively removes phenols and other pigments in black rice bran protein. In addition, the treatment method proposed by the present invention is simple to operate, has a short extraction time, and is easy to industrialize.

[0031] The realization of the purpose, functional features and advantages of the present invention will be further explained in conjunction with the embodiments. DETAILED DESCRIPTION

[0032] To make the purpose, technical scheme and advantages of the embodiment of the present invention clearer, the technical scheme in the embodiment of the present invention will be described clearly and completely below. If the specific conditions are not specified in the embodiment, it is carried out according to the normal conditions or the conditions recommended by the manufacturer. If the reagents or instruments used do not specify the manufacturer, they are all conventional products that can be purchased commercially. In addition, the meaning of "and / or" appearing in the full text includes three parallel schemes, taking "A and / or B" as an example, including scheme A, or scheme B, or schemes that A and B meet at the same time. In addition, the technical schemes between the various embodiments can be combined with each other, but must be based on the ability of ordinary technicians in the field to achieve. When the combination of technical solutions is contradictory or cannot be achieved, it should be considered that the combination of such technical solutions does not exist, and is not within the scope of protection required by the present invention. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in the field without making creative work belong to the scope of protection of the present invention.

[0033] The present invention provides a method for extracting black rice bran protein, comprising the following steps:

[0034] S10, hulling black rice to obtain black rice, grinding the black rice layer by layer from the outer layer to the inner layer and sieving the grinding product to obtain black rice bran with a grinding rate of 5 to 10%; crushing the black rice bran to obtain black rice bran powder with a mesh size of 200 meshes;

[0035] S20, mixing the black rice bran powder with n-hexane, stirring, and filtering, repeating 2 to 4 times to obtain defatted black rice bran;

[0036] S30, mixing the defatted black rice bran with water to obtain a mixed solution, adjusting the pH value of the mixed solution to the enzymatic pH, adding one or more of cellulase, saccharifying enzyme, amylase, and phytase to the system to perform an enzymatic reaction; after the enzymatic reaction is completed, washing the precipitate with water, and centrifuging at 4000-10000×g for 15-30 min to obtain a precipitate containing black rice bran protein;

[0037] S40, mixing the precipitate with an alkaline solution, stirring and extracting, centrifuging at 4000-10000×g for 15-30 min, collecting the supernatant, and obtaining a crude extract containing black rice bran protein;

[0038] S50, mixing the crude extract with a hydrogen peroxide solution, adjusting the pH of the system to a decolorization reaction pH, stirring for decolorization, and centrifuging at 4000-10000×g for 15-30 min, collecting the supernatant, and obtaining a decolorized solution;

[0039] S60, mixing the decolorized solution with an acid, adjusting the pH of the system to the isoelectric point of the black rice bran protein, and then centrifuging at 4000-10000×g for 15-30 min, taking the precipitate, and drying to obtain the black rice bran protein.

[0040] In the technical scheme of the present invention, macromolecules such as polysaccharides or anti-nutritional factors in black rice bran can be degraded by enzymatic hydrolysis, so that proteins bound to macromolecules are released, and the structure and physicochemical properties of black rice bran are changed, so that the solubility of protein is improved, thereby improving the extraction efficiency of black rice bran protein; alkaline solution can change the charge property of black rice bran protein, making it easier to dissolve in water, thereby further improving the extraction efficiency of black rice bran protein; hydrogen peroxide can better remove pigment impurities such as polyphenols in the crude extract, thereby improving the purity of black rice bran protein. The extraction method of the present invention is simple and effective, has a high extraction rate of black rice bran protein, and the obtained black rice bran protein has high purity, good quality, light color, and a relatively complete protein structure.

[0041] The novel black rice bran protein extraction technology proposed in the present invention improves the purity of black rice bran protein and properly decolorizes it, optimizes its functional properties, expands the application range of black rice bran protein, and improves its utilization value. The process is simple to operate and provides high-quality protein resources for food additives, daily chemical products, health care products and pharmaceutical industries.

[0042] It should be noted that the cellulase, saccharifying enzyme and amylase in the present invention can degrade polysaccharides well, and the phytase can degrade phytic acid well, so as to reduce the influence of polysaccharides and phytic acid in defatted rice bran on the extraction of black rice bran protein and improve the purity of black rice bran protein.

[0043] In some embodiments of the present invention, in step S20: the solid-liquid ratio of the black rice bran powder to the n-hexane is 1:3-15; and / or the stirring time is 2-8 hours. The solid-liquid ratio within the above range can ensure a high degreasing efficiency and a good degreasing effect. The solid-liquid ratio refers to the ratio of the mass of the black rice bran to the volume of the organic solvent.

[0044] In some embodiments of the present invention, in step S30: the solid-liquid ratio of the defatted black rice bran to water is 1:10-20; and / or, based on the mass of the defatted black rice bran, the addition amount of the cellulase, saccharifying enzyme, amylase, and phytase is 0.1-1%; and / or, the temperature of the enzymolysis is 30-60°C; and / or, the time of the enzymolysis is 1-6h; and / or, the pH of the enzymolysis is 2.5-8. At the same time, controlling the above parameters within a suitable range can ensure a high enzymolysis efficiency and a good enzymolysis effect, and can ensure that the polysaccharides and / or phytic acid in the defatted black rice bran can be fully degraded into small molecules.

[0045] In some embodiments of the present invention, in step S40: the pH of the alkaline solution is 8 to 12; and / or the stirring time is 1 to 6 hours. The above parameters within a suitable range can make the charge properties of the black rice bran protein change faster and more easily dissolve in water.

[0046] In some embodiments of the present invention, in step S50, the amount of hydrogen peroxide solution added is 0.5-8% based on the mass of the crude extract; and / or, the decolorization temperature is 20-60°C; and / or, the decolorization time is 0.5-4h; and / or, the decolorization pH is 6-11. The hydrogen peroxide solution can react well with the phenolic hydroxyl (-OH) in the polyphenols in the crude extract to generate quinone compounds, so that the reaction system changes from colored to colorless. The decolorization temperature, decolorization time and decolorization pH within a reasonable range can ensure a good decolorization effect.

[0047] The present invention also provides a protein supplement, which includes black rice bran protein extracted by the aforementioned black rice bran protein extraction method. Therefore, the protein supplement has all the beneficial effects of the aforementioned black rice bran protein extraction method, which will not be described one by one here.

[0048] The technical solution of the present invention is further described in detail below in conjunction with specific embodiments. It should be understood that the following embodiments are only used to explain the present invention, and are not used to limit the present invention.

[0049] Example 1

[0050] A method for extracting black rice bran protein comprises the following steps:

[0051] (1) Black rice was used as raw material, and the rice was shelled, milled and crushed, and passed through a 200-mesh sieve to obtain fresh black rice bran. Fresh rice bran was mixed with n-hexane at a solid-liquid ratio of 1:10 (w / v), magnetically stirred for 4 hours, and filtered, and repeated 3 times. The precipitate was placed in a fume hood overnight to obtain dry defatted black rice bran.

[0052] (2) Defatted black rice bran and water are mixed uniformly at a solid-liquid ratio of 1:20 (w / v), the pH of the mixed solution is adjusted to 4.5, 0.5% cellulase is added, the mixed solution is placed at 50°C for enzymolysis for 2h, and the precipitate is washed by centrifugation. Subsequently, water is added to the precipitate, the pH of the mixed solution is adjusted to 10, the mixed solution is placed in a 45°C constant temperature water bath and magnetically stirred for 120min, the mixed solution is then centrifuged at 8000×g for 20min, the supernatant is collected, a 30% mass concentration hydrogen peroxide solution is added thereto at a ratio of 1% (v / v), the pH of the mixed solution is adjusted to 10, the mixture is stirred in a 25°C constant temperature water bath for 60min, the mixture is then centrifuged at 8000×g for 20min, the pH of the mixed solution is then adjusted to 4.5, the mixture is allowed to stand at 25°C for 10min, the mixture is centrifuged at 8000×g for 20min to obtain a black rice bran protein precipitate, and the black rice bran protein is obtained by drying.

[0053] Example 2

[0054] A method for extracting black rice bran protein comprises the following steps:

[0055] (1) Black rice was used as raw material, and the black rice was shelled, milled and crushed, and passed through a 200-mesh sieve to obtain fresh black rice bran. The fresh rice bran was mixed with n-hexane at a solid-liquid ratio of 1:12 (w / v), magnetically stirred for 4 hours, and filtered, and the mixture was repeated 3 times. The precipitate was placed in a fume hood overnight to obtain dry defatted black rice bran.

[0056] (2) Defatted black rice bran and water were mixed at a solid-liquid ratio of 1:20 (w / v), the pH of the mixed solution was adjusted to 4.5, 0.5% cellulase and 0.25% saccharifying enzyme were added, the mixture was placed in a water bath at 55°C for 2h, and the precipitate was washed by centrifugation. Water was added to the precipitate, the pH of the mixed solution was adjusted to 10, the mixture was placed in a 45°C constant temperature water bath and magnetically stirred for 120min, the mixed solution was then centrifuged at 10000×g for 20min, the supernatant was collected, a 30% mass concentration of hydrogen peroxide solution was added thereto at a ratio of 1% (v / v), the pH of the mixed solution was adjusted to 8, the mixture was stirred in a 30°C constant temperature water bath for 60min, the mixture was then centrifuged at 8000×g for 20min, the pH of the mixed solution was adjusted to 4.5, the mixture was allowed to stand at 25°C for 10min, the black rice bran protein precipitate was obtained by centrifugation at 8000×g for 20min, and the black rice bran protein was obtained by drying.

[0057] Example 3

[0058] A method for extracting black rice bran protein comprises the following steps:

[0059] (1) Black rice was used as raw material, and the black rice was shelled, milled and crushed, and passed through a 200-mesh sieve to obtain fresh black rice bran. The fresh rice bran was mixed with n-hexane at a solid-liquid ratio of 1:10 (w / v), magnetically stirred for 4 hours, and filtered, and the mixture was repeated 3 times. The precipitate was placed in a fume hood overnight to obtain dry defatted black rice bran.

[0060] (2) Defatted black rice bran and water were mixed at a solid-liquid ratio of 1:20 (w / v), the pH of the mixture was adjusted to 4.5, 0.5% cellulase and 0.25% phytase were added, the mixture was placed in a 55°C water bath for 2h, and the precipitate was washed by centrifugation. Water was added to the precipitate, the pH of the mixture was adjusted to 10, the mixture was placed in a 20°C constant temperature water bath for magnetic stirring for 180min, the mixture was then centrifuged at 8000×g for 20min, the supernatant was collected, a 30% mass concentration of hydrogen peroxide solution was added thereto at a ratio of 1% (v / v), the pH of the mixture was adjusted to 10, the mixture was stirred in a 50°C constant temperature water bath for 80min, the mixture was then centrifuged at 8000×g for 20min, the pH of the mixture was adjusted to 4.5, the mixture was allowed to stand at 25°C for 10min, the mixture was centrifuged at 8000×g for 20min to obtain a black rice bran protein precipitate, and the black rice bran protein was obtained by drying.

[0061] Example 4

[0062] A method for extracting black rice bran protein comprises the following steps:

[0063] (1) Black rice was used as raw material, and the black rice was shelled, milled and crushed, and passed through a 200-mesh sieve to obtain fresh black rice bran. The fresh rice bran was mixed with n-hexane at a solid-liquid ratio of 1:15 (w / v), magnetically stirred for 4 hours, and filtered, and the mixture was repeated 3 times. The precipitate was placed in a fume hood overnight to obtain dry defatted black rice bran.

[0064] (2) Defatted black rice bran and water were mixed at a solid-liquid ratio of 1:20 (w / v), the pH of the mixture was adjusted to 5, 0.5% cellulase and 0.25% amylase were added, the mixture was shaken in a water bath at 50°C for 2h, and the precipitate was washed by centrifugation. Water was added to the precipitate, the pH of the mixture was adjusted to 10, the mixture was placed in a constant temperature water bath at 45°C for magnetic stirring for 120min, the mixture was then centrifuged at 8000×g for 20min, the supernatant was collected, a 30% mass concentration of hydrogen peroxide solution was added thereto at a ratio of 1% (v / v), the pH of the mixture was adjusted to 10, the mixture was stirred in a constant temperature water bath at 25°C for 40min, the mixture was then centrifuged at 8000×g for 20min, the pH of the mixture was adjusted to 4.5, the mixture was allowed to stand at 25°C for 10min, the mixture was centrifuged at 8000×g for 20min to obtain a black rice bran protein precipitate, and the black rice bran protein was obtained by drying.

[0065] Comparative Example 1

[0066] Compared with Example 1, Comparative Example 1 is the same as Example 1 except that enzymatic hydrolysis is not performed and hydrogen peroxide solution is not added.

[0067] Comparative Example 2

[0068] Comparative Example 2 is compared with Example 1, except that the hydrogen peroxide solution is not added, and other aspects are the same as Example 1.

[0069] Performance Testing

[0070] The black rice bran protein extracted in Examples 1 to 4 and Comparative Example 1 was tested for brightness and purity. The brightness of the black rice bran protein was tested using a colorimeter. The content of the black rice bran protein was tested using the Kjeldahl method according to GB5009.5-2010. The protein content was calculated as follows:

[0071] Protein content (100%) = protein mass / sample mass × 100.

[0072] The test results are shown in Table 1.

[0073] The larger the L* value, the whiter the surface of the substance, which means the better the decolorization effect of the protein.

[0074] Table 1 Characterization of black rice bran protein in Examples 1 to 4 and Comparative Example 1

[0075] Group L* Protein content (%) Example 1 29.79±0.32 82.90±0.81 Example 2 31.42±0.19 84.27±0.72 Example 3 31.95±0.22 85.18±0.27 Example 4 30.98±0.14 83.76±0.48 Comparative Example 1 23.36±0.11 44.22±0.53 Comparative Example 2 24.01±0.23 62.13±0.67

[0076] The above are only preferred embodiments of the present invention, and do not limit the patent scope of the present invention. For those skilled in the art, the present invention may have various changes and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the patent protection scope of the present invention.

Claims

1. A method for extracting black rice bran protein, characterized in that, The following steps are involved: S10, hulling black rice to obtain black rice, grinding the black rice layer by layer from the outer layer to the inner layer and sieving the grinding product to obtain black rice bran with a grinding rate of 5 to 10%; crushing the black rice bran to obtain black rice bran powder with a mesh size of 200 meshes; S20, mixing the black rice bran powder with n-hexane, stirring, and filtering, repeating 2 to 4 times to obtain defatted black rice bran; S30, mixing the defatted black rice bran with water to obtain a mixed solution, adjusting the pH value of the mixed solution to the enzymatic pH, adding one or more of cellulase, saccharifying enzyme, amylase, and phytase to the system to perform an enzymatic reaction; after the enzymatic reaction is completed, washing the precipitate with water, and centrifuging at 4000-10000×g for 15-30 min to obtain a precipitate containing black rice bran protein; S40, mixing the precipitate with an alkaline solution, stirring and extracting, centrifuging at 4000-10000×g for 15-30 min, collecting the supernatant, and obtaining a crude extract containing black rice bran protein; S50, mixing the crude extract with a hydrogen peroxide solution, adjusting the pH of the system to a decolorization reaction pH, stirring for decolorization, and centrifuging at 4000-10000×g for 15-30 min, collecting the supernatant, and obtaining a decolorized solution; S60, mixing the decolorized solution with an acid, adjusting the pH of the system to the isoelectric point of the black rice bran protein, and then centrifuging at 4000-10000×g for 15-30 min, taking the precipitate, and drying to obtain the black rice bran protein.

2. The method for extracting black rice bran protein as claimed in claim 1, wherein In step S20: the solid-liquid ratio of the black rice bran powder to the n-hexane is 1:3-15; and / or, The stirring time is 2 to 8 hours.

3. The method for extracting black rice bran protein as claimed in claim 1, wherein In step S30: the solid-liquid ratio of the defatted black rice bran to water is 1:10-20; and / or, Based on the mass of defatted black rice bran, the added amount of the cellulase, saccharifying enzyme, amylase and phytase is 0.1-1%; and / or, The enzymatic hydrolysis temperature is 30-60°C; and / or, The enzymatic hydrolysis time is 1 to 6 hours; and / or, The pH of the enzymatic hydrolysis is 2.5-8.

4. The method for extracting black rice bran protein as claimed in claim 1, wherein In step S40: the pH of the alkaline solution is 8 to 12; and / or, The stirring time is 1 to 6 hours.

5. The method for extracting black rice bran protein as claimed in claim 1, wherein In step S50, the amount of hydrogen peroxide solution added is 0.5-8% based on the mass of the crude extract; and / or, The decolorization temperature is 20-60°C; and / or, The decolorization time is 0.5 to 4 hours; and / or, The pH of the decolorization is 6-11.

6. A protein supplement, characterized in that The protein supplement comprises black rice bran protein extracted by the method for extracting black rice bran protein according to any one of claims 1 to 5.