Preparation method and application of sunflower seed protein

Abstract

The invention relates to the technical field of food processing, in particular to a preparation method and application of sunflower seed protein. The preparation method comprises the following steps: mixing sunflower seed meal with an aqueous two-phase solvent, and then carrying out constant-temperature extraction, dialysis, alkali extraction and acid precipitation treatment; the aqueous two-phase solvent is a mixed solution of ammonium sulfate and an ethanol water solution. According to the preparation method, the content of chlorogenic acid in the sunflower seed meal can be reduced, the defect that browning is serious in the sunflower seed protein extraction process is overcome, the quality of the sunflower seed protein is improved, further application of the sunflower seed protein in the field of food processing is promoted, and therefore the economic value of bulk crops is increased.

Description

Technical Field

[0001] The invention relates to the technical field of food processing, and in particular to a preparation method and application of sunflower seed protein. Background Art

[0002] As one of the four major oil crops in the world, the output of sunflower seeds is increasing year by year. Compared with other plant proteins, sunflower seed protein has a balanced amino acid composition, good digestibility (90%) and low allergenicity. It is a high-quality and low-cost high-quality protein (see Food Hydrocolloids, 2024, 147: 109329). At present, sunflower meal protein is mainly used in the feed processing industry due to severe denaturation caused by high-temperature pressing, and is rarely used in food. In order to achieve high-value utilization of sunflower meal protein, domestic and foreign scholars and entrepreneurs have begun to pay attention to the application of low-temperature sunflower meal in food. However, since sunflower meal is rich in polyphenols, especially high content of chlorogenic acid, it is easy to interact with protein to produce browning and bad flavor, which greatly limits its deep processing application.

[0003] Chlorogenic acid (CGA) has strong polarity and is prone to intramolecular group and force migration under high temperature, light and high pH conditions, thereby isomerizing to form three main polyphenol isomers: 5-caffeoylquinic acid (5-CQA), 3-caffeoylquinic acid (3-CQA) and 4-caffeoylquinic acid (4-CQA). During the polyphenol removal process of sunflower meal, CGA and its isomers inevitably interact with sunflower seed protein to form green pigments (see Food Research International, 2022, 162: 111996). Protein greening not only reduces the digestibility of protein, but also affects the solubility of protein and has a significant negative impact on sensory properties. The greening of sunflower seed protein is particularly significant during its preparation process. Under acidic or neutral conditions, the active hydroxyl groups of CGA cannot undergo free radical reactions, which has little effect on sunflower seed protein. Under alkaline preparation conditions, the covalent and non-covalent interactions between the hydroxyl groups in CGA and amino acid groups such as lysine, methionine, and arginine can quickly combine to cause the greening of sunflower seed protein. In addition, in the processing and utilization of sunflower seed protein, high pH or high temperature baking will promote the oxidation of CGA, resulting in green pigment precipitation in bread and biscuit products with added sunflower seed protein powder, which is also an important factor limiting the deep processing and utilization of sunflower seed protein (see Food Science, 2024, 45(18): 290-298.). Therefore, it is of great significance to efficiently extract and remove chlorogenic acid from sunflower seed meal and prepare high-quality sunflower seed protein.

[0004] Therefore, how to provide a method for efficiently removing chlorogenic acid from sunflower seed meal and preparing high-quality sunflower seed protein has become a technical problem that urgently needs to be solved in this field. Summary of the invention

[0005] In order to solve the above technical problems, the present invention provides a method for preparing sunflower seed protein, comprising: mixing sunflower seed meal with a two-phase aqueous solvent to obtain a sunflower seed meal dispersion; then subjecting the mixture to constant temperature extraction, dialysis, alkali extraction and acid precipitation treatment to obtain sunflower seed protein; the two-phase aqueous solvent is a mixture of ammonium sulfate and ethanol aqueous solution.

[0006] The present invention finds through a large number of exploratory experiments on two-phase aqueous solvents that the two-phase aqueous solvent prepared by selecting ammonium sulfate and ethanol is very suitable for the extraction process of sunflower seed meal protein, and can efficiently remove chlorogenic acid from sunflower seed meal and obtain high-quality sunflower seed protein.

[0007] Preferably, the mass percentage of ethanol in the aqueous two-phase solvent is 30% to 38%.

[0008] Preferably, the mass percentage of ethanol in the aqueous two-phase solvent is 32% to 36%.

[0009] Preferably, the mass percentage of ammonium sulfate in the aqueous two-phase solvent is 12% to 20%.

[0010] Preferably, the mass percentage of ammonium sulfate in the aqueous two-phase solvent is 14% to 18%.

[0011] Preferably, the weight ratio of sunflower seed meal to the aqueous two-phase solvent is 1:(5-30); preferably 1:(5-15); more preferably 1:(8-12).

[0012] Preferably, the constant temperature extraction comprises: stirring at 30-50° C. and 300-350 rpm for 20-40 min, and centrifuging to obtain dephenolized sunflower seed meal; And / or, the dialysis comprises: dialysis for 24 to 72 hours using a dialysis membrane with a pore size of 8000 to 10000Da; And / or, the alkaline extraction comprises: mixing the dialyzed dephenolized sunflower seed meal with a sodium hydroxide solution, adjusting the pH to 9-10, and then extracting; And / or, the acid precipitation treatment includes: mixing the supernatant after alkali extraction with a hydrochloric acid solution, adjusting the pH to 4.0-5.0 to precipitate protein, taking the precipitate and adjusting it to neutral, and drying to obtain sunflower seed protein.

[0013] Preferably, during the alkaline extraction process, the weight ratio of the dialyzed dephenolized sunflower seed meal to the sodium hydroxide solution is 1:(5-30); and / or, the concentration of the sodium hydroxide solution is 0.5-1.5 mol / L; and / or, the extraction is performed at a rotation speed of 300-350 rpm for 90-150 min; and / or, after the extraction, the supernatant is obtained by centrifugation at 3000-6000 r / min for 10-20 min; And / or, during the acid precipitation process, the concentration of the hydrochloric acid solution is 0.5-1.5 mol / L; and / or, after adjusting the pH to 4.0-5.0, the protein is precipitated by centrifugation at 3000-6000 r / min for 10-20 min.

[0014] Preferably, the chlorogenic acid removal rate is above 91% (e.g., 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, etc.); and / or the purity of sunflower seed protein is above 84% (e.g., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 93%, 95%, 97%, 98%, 99%, etc.).

[0015] Most preferably, the present invention provides a method for preparing sunflower seed protein, comprising: (1) mixing sunflower seed meal with an aqueous two-phase solvent in a weight ratio of 1:(8-12) to prepare a sunflower seed meal dispersion; the aqueous two-phase solvent is a mixture of ammonium sulfate and ethanol aqueous solution, the mass percentage of ethanol in the aqueous two-phase solvent is 32%-36%, and the mass percentage of ammonium sulfate in the aqueous two-phase solvent is 14%-18%; (2) The sunflower seed meal dispersion was stirred at 35-45°C and 330-350 rpm for 20-40 min; then, it was dialyzed using a 8000-10000Da pore size dialysis membrane for 40-50 h, the dialyzed dephenolized sunflower seed meal was mixed with 0.5-1.5 mol / L sodium hydroxide solution at a weight ratio of 1:(5-15), the pH was adjusted to 9-10, and then, it was extracted at 300-350 rpm for 110-130 min; after extraction, it was centrifuged at 4000-5000 r / min for 10-20 min to obtain a supernatant; the supernatant was mixed with 0.5-1.5 mol / L hydrochloric acid solution, the pH was adjusted to 4.0-5.0, and then, it was centrifuged at 4000-5000 r / min for 10-20 min to precipitate protein, the precipitate was adjusted to neutral, and dried to obtain sunflower seed protein.

[0016] Furthermore, the present invention provides application of the preparation method in sunflower seed protein processing.

[0017] Compared with the prior art, the present invention has the following beneficial effects: The present invention provides a method for removing chlorogenic acid from sunflower seed meal by using a two-phase aqueous solvent and preparing high-quality sunflower seed protein, which can reduce the chlorogenic acid content in sunflower seed meal, overcome the defect of serious browning in the sunflower seed protein extraction process, improve the quality of sunflower seed protein, promote its further application in the field of food processing, and thus improve the economic value of bulk crops; specifically, it has the following advantages: (1) The raw material used in the present invention is sunflower seed meal, which is obtained by pressing sunflower seeds at a suitable temperature or low temperature. It is abundant in source, of good quality and at a low price.

[0018] (2) The biggest innovation of the sunflower seed protein of the present invention is "green preparation, high quality, and high purity". Compared with traditional dephenolization and protein preparation, it greatly saves costs and is environmentally friendly and pollution-free in many aspects such as solvent selection, processing and manufacturing, and product quality, injecting new products and technologies into the sunflower seed processing market.

[0019] (3) The sunflower seed protein of the present invention has outstanding advantages over similar products in terms of protein purity, protein quality, protein solubility, etc.

[0020] (4) The two-phase aqueous solvent of the present invention is an immiscible aqueous solution system formed under certain conditions by dissolving two or more water-soluble compounds in water at appropriate concentrations. It is an excellent green extractant. It has the advantages of safety and non-toxicity, mild operating conditions, simple process equipment, low interfacial tension, easy process scale-up, low energy consumption and high productivity, and is very suitable for the separation and extraction of polyphenols in sunflower seed meal. DETAILED DESCRIPTION

[0021] In order to make the purpose, technical solution and advantages of the present invention clearer, the technical solution of the present invention will be described clearly and completely below. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. 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.

[0022] If no specific techniques or conditions are specified in the examples, all are carried out according to conventional methods or the techniques or conditions described in the literature in the field, or according to the product instructions. If no manufacturer is specified for the reagents and instruments used, all are conventional products that can be purchased through regular channels.

[0023] Example 1

[0024] This embodiment provides a method for preparing sunflower seed protein, the steps are as follows: Take 10 parts by weight of sunflower seed meal, add 100 parts by weight of an aqueous two-phase solvent, the aqueous two-phase solvent is a mixture of ammonium sulfate and ethanol aqueous solution; the mass percentage of ethanol in the aqueous two-phase solvent is 34%, the mass percentage of ammonium sulfate in the aqueous two-phase solvent is 16%, and the balance is water. The mixture was mixed evenly to obtain a sunflower seed meal dispersion, the temperature was maintained at 40°C, and the mixture was reacted and extracted at a speed of 350 rpm for 30 min. The sunflower seed meal dephenolized by the two-phase solvent was dialyzed using an 8000Da dialysis membrane for 48 h. 10 parts by weight of dephenolized sunflower seed meal and 100 parts by weight of a 1.0 mol / L sodium hydroxide solution were taken to adjust the pH to 9.5. The mixture was reacted and extracted at 350 rpm for 120 min. After the reaction was completed, the mixture was centrifuged at 4500 r / min for 15 min to obtain the supernatant. A 1.0 mol / L hydrochloric acid solution was added to the alkaline extraction solution to adjust the pH to 4.5. After the reaction was completed, the mixture was centrifuged at 4500 r / min for 15 min to obtain the precipitate. The precipitate was adjusted to neutral and dried to obtain sunflower seed protein.

[0025] Example 2

[0026] This embodiment provides a method for preparing sunflower seed protein, and the steps are only different from those in Embodiment 1: Take 10 parts by weight of sunflower seed meal, add 100 parts by weight of a two-phase aqueous solvent, the two-phase aqueous solvent is a mixture of ammonium sulfate and ethanol aqueous solution; the mass percentage of ethanol in the two-phase aqueous solvent is 30%, the mass percentage of ammonium sulfate in the two-phase aqueous solvent is 20%, and the balance is water. Mix well to obtain a sunflower seed meal dispersion.

[0027] Example 3

[0028] This embodiment provides a method for preparing sunflower seed protein, and the steps are only different from those in Embodiment 1: Take 10 parts by weight of sunflower seed meal, add 100 parts by weight of a two-phase aqueous solvent, the two-phase aqueous solvent is a mixture of ammonium sulfate and ethanol aqueous solution; the mass percentage of ethanol in the two-phase aqueous solvent is 38%, the mass percentage of ammonium sulfate in the two-phase aqueous solvent is 12%, and the balance is water. Mix well to obtain a sunflower seed meal dispersion.

[0029] Example 4

[0030] This embodiment provides a method for preparing sunflower seed protein, and the steps are only different from those in Embodiment 1: Take 10 parts by weight of sunflower seed meal, add 100 parts by weight of a two-phase aqueous solvent, the two-phase aqueous solvent is a mixture of ammonium sulfate and ethanol water solution; the mass percentage of ethanol in the two-phase aqueous solvent is 34%, the mass percentage of ammonium sulfate in the two-phase aqueous solvent is 12%, and the balance is water. Mix well to obtain a sunflower seed meal dispersion.

[0031] Example 5

[0032] This embodiment provides a method for preparing sunflower seed protein, and the steps are only different from those in Embodiment 1: Take 10 parts by weight of sunflower seed meal, add 100 parts by weight of a two-phase aqueous solvent, the two-phase aqueous solvent is a mixture of ammonium sulfate and ethanol aqueous solution; the mass percentage of ethanol in the two-phase aqueous solvent is 38%, the mass percentage of ammonium sulfate in the two-phase aqueous solvent is 20%, and the balance is water. Mix well to obtain a sunflower seed meal dispersion.

[0033] Comparative Example 1

[0034] This comparative example provides a method for preparing sunflower seed protein, the steps of which are different from those of Example 1 only in that ammonium sulfate is not added.

[0035] Comparative Example 2

[0036] This comparative example provides a method for preparing sunflower seed protein, and the steps are only different from those of Example 1 in that the ethanol aqueous solution is replaced by a methanol aqueous solution of the same concentration.

[0037] Comparative Example 3

[0038] This comparative example provides a method for preparing sunflower seed protein, and the steps are only different from those in Example 1 in that ammonium sulfate is replaced by sodium carbonate of the same concentration.

[0039] Test example

[0040] This test example tests the purity and chlorogenic acid removal rate of the sunflower seed protein prepared in the above examples and comparative examples. Among them, the purity of sunflower seed protein is determined according to the first method in GB 5009.5-2016 "National Food Safety Standard Determination of Protein in Food". The UPLC method is used to detect the chlorogenic acid content in the sunflower seed meal raw material and sunflower seed protein, and the chlorogenic acid removal rate of the sunflower seed meal is calculated; the calculation formula for the chlorogenic acid removal rate of the sunflower seed meal is: Chlorogenic acid removal rate = (1-chlorogenic acid content in sunflower seed meal after dephenolization / total chlorogenic acid in sunflower seed meal) × 100%; the UPLC method refers to "UPLC-MS-MS Analysis of 12 Polyphenolic Compounds in Fresh Tobacco Leaves".

[0041] The test results are shown in Table 1.

[0042] Table 1

[0043] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or make equivalent replacements for some of the technical features therein. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for preparing sunflower seed protein, characterized in that: include: The sunflower seed meal is mixed with a two-phase aqueous solvent to obtain a sunflower seed meal dispersion; then the sunflower seed protein is obtained through constant temperature extraction, dialysis, alkali extraction and acid precipitation treatment; the two-phase aqueous solvent is a mixture of ammonium sulfate and ethanol aqueous solution.

2. The preparation method according to claim 1, characterized in that: The mass percentage of ethanol in the aqueous two-phase solvent is 30% to 38%.

3. The preparation method according to claim 2, characterized in that: The mass percentage of ethanol in the aqueous two-phase solvent is 32% to 36%.

4. The preparation method according to claim 1, characterized in that: The mass percentage of ammonium sulfate in the aqueous two-phase solvent is 12% to 20%.

5. The preparation method according to claim 4, characterized in that: The mass percentage of ammonium sulfate in the aqueous two-phase solvent is 14% to 18%.

6. The preparation method according to claim 1, characterized in that: The weight ratio of sunflower seed meal to the aqueous two-phase solvent is 1:(5-30); preferably 1:(5-15); more preferably 1:(8-12).

7. The preparation method according to claim 1, characterized in that: The constant temperature extraction comprises: stirring at 30-50°C and 300-350 rpm for 20-40 min; And / or, the dialysis comprises: dialysis for 24 to 72 hours using a dialysis membrane with a pore size of 8000 to 10000Da; And / or, the alkaline extraction comprises: mixing the dialyzed dephenolized sunflower seed meal with a sodium hydroxide solution, adjusting the pH to 9-10, and then extracting; And / or, the acid precipitation treatment includes: mixing the supernatant after alkali extraction with a hydrochloric acid solution, adjusting the pH to 4.0-5.0 to precipitate protein, taking the precipitate and adjusting it to neutral, and drying to obtain sunflower seed protein.

8. The preparation method according to claim 7, characterized in that: During the alkaline extraction process, the weight ratio of the dialyzed dephenolized sunflower seed meal to the sodium hydroxide solution is 1:(5-30); and / or, the concentration of the sodium hydroxide solution is 0.5-1.5 mol / L; and / or, the extraction is performed at a rotation speed of 300-350 rpm for 90-150 min; and / or, after the extraction, the supernatant is obtained by centrifugation at 3000-6000 r / min for 10-20 min; And / or, during the acid precipitation process, the concentration of the hydrochloric acid solution is 0.5-1.5 mol / L; and / or, after adjusting the pH to 4.0-5.0, the protein is precipitated by centrifugation at 3000-6000 r / min for 10-20 min.

9. The preparation method according to any one of claims 1 to 8, characterized in that The chlorogenic acid removal rate is greater than 91%; and / or the purity of the sunflower seed protein is greater than 84%.

10. Use of the preparation method according to any one of claims 1 to 9 in sunflower seed protein processing.

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