Tartary buckwheat protein compound as well as preparation method and application thereof

Abstract

The invention discloses a tartary buckwheat protein compound as well as a preparation method and application thereof, and relates to the technical field of preparation of Pickering emulsion stabilizers. The method comprises the following steps: diluting tartary buckwheat protein hydrogel with distilled water, and adjusting the pH value to prepare protein hydrogel; mixing quercetin with an ethanol solution, and adjusting the pH value to prepare a quercetin solution; and mixing the protein hydrogel with a quercetin solution, heating in a water bath, carrying out ultrasonic emulsification, and freeze-drying to obtain the tartary buckwheat protein compound. The invention provides a non-toxic, harmless and pollution-free natural emulsifier and stabilizer, simultaneously prepares a Pickering emulsion capable of loading functional substances and nutritional ingredients, and provides reference for screening research of the natural emulsifier, basic research of tartary buckwheat protein, development of tartary buckwheat products and protection of nutritional ingredients easy to photodecompose and oxidize. The problem that in the prior art, the effect of protein serving as a Pickering emulsion stabilizer is poor is solved.

Description

Technical Field

[0001] The invention relates to the technical field of preparation of a Pickering emulsion stabilizer, and in particular to a tartary buckwheat protein complex and a preparation method and application thereof. Background Art

[0002] Tartary buckwheat is a plant of the genus Fagopyrum in the family Rhizoma Coptidis. It is a cereal crop that contains seven major nutrients at the same time. It is mainly grown in Yunnan, Sichuan, Guizhou, Hunan, Hubei and other regions. There are many kinds of amino acids in tartary buckwheat protein, and the proportions are relatively balanced. The 8 essential amino acids for the human body are found in high levels in tartary buckwheat protein.

[0003] Pickering emulsion is an emulsion prepared by replacing emulsifiers with solid particles. Pickering emulsion has better advantages than traditional emulsions. First, the solid particles are irreversibly adsorbed and anchored on the surface of the emulsion droplets, forming a dense interface film on the adsorption surface. Through the interface exclusion effect, the droplets are prevented from contacting each other and interacting with each other. At the same time, the solid particles make the oil-water system interact with each other to form a three-dimensional network structure. This three-dimensional structure increases the viscosity of the emulsion, reduces the Brownian motion of the particles, and makes the emulsion very stable. Second, Pickering emulsion has no surfactant, thus reducing the toxicity, environmental pollution and foaming caused by surfactants, achieving a non-toxic, harmless and pollution-free state. Third, Pickering emulsion uses solid particles to stabilize the emulsion, reducing the amount of traditional emulsifiers and reducing production costs. Fourth, Pickering emulsion can be used as a carrier of bioactive substances, which can well encapsulate and transport active substances, while also masking unpleasant odors and improving the antioxidant capacity of active substances.

[0004] Protein is composed of 20 different amino acids. The diversity of amino acid composition and arrangement order leads to different higher-level structures, spatial conformations and functional properties of proteins, which give proteins unique functional properties, including emulsification, solubility, water retention and foaming properties, and can be used as stabilizers for Pickering emulsions. Although natural proteins have good functional properties, their application still cannot meet the various needs of the food industry due to the limitations and variability of their structure. Therefore, it is necessary to obtain the desired protein surface functional properties through appropriate modification, modification and preparation of complexes, so as to increase the application of proteins in the food industry and obtain better Pickering emulsion stabilizers.

[0005] β-carotene, as an important member of the carotenoid family, is widely distributed in nature and is one of the most common natural pigments in nature. β-carotene has high pharmacological and nutritional value and has been widely used in medicine, health products, food additives and cosmetics. Insufficient intake can cause symptoms such as night blindness, dry mucous membranes, dry eyes and myopia; it can increase the chances of cancer, cataracts, cardiovascular, reproductive system, urinary system diseases and respiratory infections. However, β-carotene is unstable in oxygen, heat and light, which limits its effective application and requires a packaging system to improve its stability. Summary of the invention

[0006] In order to solve the above technical problems, the purpose of the present invention is to provide a tartary buckwheat protein complex and a preparation method and application thereof, so as to solve the problem that the Pickering emulsion stabilizer in the prior art has a poor effect.

[0007] The technical solution of the present invention to solve the above technical problem is as follows: a method for preparing a tartary buckwheat protein complex is provided, comprising the following steps:

[0008] (1) diluting the buckwheat protein hydrogel with distilled water and adjusting the pH value to prepare the protein hydrogel;

[0009] (2) mixing quercetin and an ethanol solution and adjusting the pH value to prepare a quercetin solution;

[0010] (3) mixing the protein hydrogel prepared in step (1) and the quercetin solution prepared in step (2), heating in a water bath, ultrasonic emulsification, and freeze drying to prepare a buckwheat protein complex.

[0011] On the basis of the above technical solution, the present invention can also be improved as follows:

[0012] Further, in step (1), the tartary buckwheat protein hydrogel is prepared by the following method:

[0013] S1: dissolving tartary buckwheat flour with PBS buffer, stirring and centrifuging to obtain a first supernatant, adding ammonium sulfate, stirring and centrifuging to obtain a second supernatant, and then continuing to add ammonium sulfate, stirring and centrifuging to obtain a precipitate; re-dissolving with water, and then dialyzing, adjusting the pH value and freeze-drying to obtain tartary buckwheat protein;

[0014] S2: dissolving the tartary buckwheat protein obtained in step S1 in PBS buffer to obtain a tartary buckwheat protein solution, stirring for 1.5-2.5 hours, and then placing it at 3-6°C for 10-15 hours, adjusting the pH value, and obtaining a hydrated tartary buckwheat protein solution; centrifuging, heating the supernatant at 75-90°C, and then treating it in an ice water bath, diluting, and obtaining a tartary buckwheat protein hydrogel.

[0015] Further, in step S1, the mass volume ratio of tartary buckwheat flour and PBS buffer solution is 1 g:9-11 mL.

[0016] Furthermore, in step S1, the concentration of the PBS buffer is 0.008-0.012 mol / L, and the pH value is 6.5-7.5.

[0017] Further, in step S1, after the tartary buckwheat flour is dissolved in PBS buffer, it is stirred for 1.5-2.5 hours and centrifuged at 3000-6000 r / min for 25-35 minutes.

[0018] Further, in step S1, the concentration of ammonium sulfate in the first supernatant is 30-50wt%.

[0019] Further, in step S1, the concentration of ammonium sulfate in the second supernatant is 70-90wt%.

[0020] Further, in step S1, after ammonium sulfate is added to the first supernatant and the second supernatant, the mixture is stirred for 50-70 minutes and centrifuged at 9000-11000 r / min for 10-20 minutes.

[0021] Further, in step S1, dialyze for 40-50 hours at 3-6°C using a 3000-4000Da dialysis bag.

[0022] Further, in step S1, the pH value is adjusted to 9.

[0023] Further, in step S2, the concentration of the tartary buckwheat protein solution is 4-8wt%.

[0024] Further, in step S2, the pH value is 8.

[0025] Furthermore, in step (1), the concentration of the tartary buckwheat protein hydrogel is 0.015-0.025 g / mL.

[0026] Furthermore, in step (1), the concentration of the tartary buckwheat protein hydrogel is 0.02 g / mL.

[0027] Furthermore, in step (1), the pH value is adjusted to 8.5-9.5.

[0028] Furthermore, in step (1), the pH value is adjusted to 9.

[0029] Furthermore, in step (1), the pH value is adjusted using NaOH at a concentration of 0.1 mol / L.

[0030] Furthermore, in step (2), the concentration of the ethanol solution is 50-70 vt%.

[0031] Furthermore, in step (2), the concentration of the ethanol solution is 60 vt%.

[0032] Furthermore, in step (2), the pH value is adjusted using NaOH at a concentration of 0.1 mol / L.

[0033] Furthermore, in step (2), the concentration of the quercetin solution is 0.008-0.012 g / mL.

[0034] Furthermore, in step (2), the concentration of the quercetin solution is 0.01 g / mL.

[0035] Furthermore, in step (2), the pH value is adjusted to 8.5-9.5.

[0036] Furthermore, in step (2), the pH value is adjusted to 9.

[0037] Furthermore, in step (3), the volume ratio of the buckwheat protein hydrogel to the quercetin solution is 1:0.8-1.2.

[0038] Furthermore, in step (3), the volume ratio of the buckwheat protein hydrogel to the quercetin solution is 1:1.

[0039] Furthermore, in step (3), heating is performed in a water bath at 90-100° C. for 50-70 min.

[0040] Furthermore, in step (3), heating is performed in a water bath at 90° C. for 60 min.

[0041] Furthermore, in step (3), ultrasonic emulsification is performed at an ultrasonic power of 450 W for 10-30 min.

[0042] Furthermore, in step (3), ultrasonic emulsification is performed at an ultrasonic power of 450 W for 10 minutes.

[0043] Further, in step (3), freeze drying is performed at -80 to -70°C and 0 to 6 Pa for 40 to 50 hours.

[0044] The invention also provides the tartary buckwheat protein complex prepared by the method.

[0045] The present invention also provides application of the tartary buckwheat protein complex in preparing Pickering emulsion.

[0046] Furthermore, the preparation method of Pickering emulsion comprises the following steps: dissolving the above-mentioned tartary buckwheat protein complex in distilled water to obtain a water phase, adding an oil phase, and ultrasonically emulsifying to obtain a Pickering emulsion.

[0047] Furthermore, the mass volume ratio of the tartary buckwheat protein complex and distilled water is 0.2 g:20 mL.

[0048] Furthermore, the volume ratio of the water phase to the oil phase is 1:1.

[0049] Furthermore, the oil phase is soybean oil.

[0050] Furthermore, ultrasonic emulsification was performed at an ultrasonic power of 450 W for 10-30 min.

[0051] Furthermore, ultrasonic emulsification was performed at an ultrasonic power of 450 W for 10 min.

[0052] Furthermore, the Pickering emulsion is a Pickering emulsion loaded with β-carotene.

[0053] Furthermore, the preparation method of the Pickering emulsion loaded with β-carotene comprises the following steps: dissolving β-carotene in an oil phase, performing ultrasonic emulsification for the first time, then adding a tartary buckwheat protein complex solution, performing ultrasonic emulsification for the second time, and obtaining the Pickering emulsion loaded with β-carotene.

[0054] Furthermore, the oil phase is soybean oil.

[0055] Furthermore, the mass volume ratio of β-carotene to the oil phase was 1 mg:1 mL.

[0056] Furthermore, the first ultrasonic emulsification was performed at an ultrasonic power of 450 W for 10 min.

[0057] Furthermore, the volume ratio of the oil phase to the tartary buckwheat protein complex solution is 1:1.

[0058] Furthermore, the concentration of the tartary buckwheat protein complex solution was 10 mg / mL.

[0059] Furthermore, a second ultrasonic emulsification was performed at an ultrasonic power of 450 W for 10-30 min.

[0060] Furthermore, a second ultrasonic emulsification was performed at an ultrasonic power of 450 W for 10 min.

[0061] The present invention has the following beneficial effects:

[0062] 1. Buckwheat protein is a well-balanced plant protein with comprehensive physiological and biochemical properties and high nutritional value. Buckwheat protein contains hydrophilic and hydrophobic amino acids, which have good emulsification and can be used to stabilize Pickering emulsions. Buckwheat protein combines with polyphenols through non-covalent and covalent interactions, which can change the structure of the protein, improve the functional properties of buckwheat protein, and make the formed complex have better stability, emulsification and antioxidant properties, which is conducive to the preparation of a more stable and safe Pickering emulsion.

[0063] 2. Quercetin (Q) belongs to the class of polyphenolic compounds. It is composed of two heterocyclic rings, ring A and ring B, connected by an oxygen-containing heterocyclic ring. There are multiple hydrophilic hydroxyl groups and hydrophobic ring structures in its molecule. Quercetin is found in large quantities in plant foods, such as Sophora japonica, onion, sea buckthorn, etc. It is also found in large quantities in bitter buckwheat. Due to its low water solubility, difficulty in absorption and high metabolic rate, the bioavailability of quercetin is relatively low (<20%). Quercetin has antioxidant, anti-tumor and anti-hypertensive effects. Current studies have shown that quercetin can be used to prevent and treat diseases such as tumors, and that high doses of quercetin can inhibit acrylamide-induced toxicity. The phenolic hydroxyl groups in the structure of quercetin give it unique chemical properties, which can interact with biological macromolecules such as proteins, and can improve its solubility, stability and bioavailability.

[0064] 3. The present invention provides a non-toxic, harmless and pollution-free natural emulsifier and stabilizer, and at the same time prepares a Pickering emulsion that can load functional substances and nutrients, providing a reference for the screening research of natural emulsifiers, the basic research of buckwheat protein and the development of buckwheat products.

[0065] 4. The complex prepared under alkaline and high quercetin concentration and high temperature conditions has better stability, enhanced surface hydrophobicity, emulsification and emulsion stability, and is more suitable for the preparation of buckwheat protein complex Pickering emulsion; the emulsion prepared under high temperature and high concentration conditions is less affected by centrifugation, salt ions and freeze-thaw, and has better stability; buckwheat protein Pickering emulsion can effectively slow down the photodegradation of β-carotene and improve the stability of β-carotene. Among them, the buckwheat protein complex stabilized Pickering emulsion prepared under the conditions of quercetin concentration of 0.01 g / mL, pretreatment temperature above 90°C, and pH 9 has the most significant effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0066] Figure 1 is the particle size of the buckwheat protein complex;

[0067] Figure 2 is the zeta potential of the tartary buckwheat protein complex;

[0068] Figure 3 The emulsifying activity of the tartary buckwheat protein complex;

[0069] Figure 4 For the emulsification stability of buckwheat protein complex;

[0070] Figure 5 is the surface hydrophobicity of the buckwheat protein complex;

[0071] Figure 6 is the creaming index of the Pickering emulsion made from the tartary buckwheat protein complex;

[0072] Figure 7 The thermal stability of Pickering emulsions prepared from tartary buckwheat protein complex;

[0073] Figure 8 Salt ion stability of Pickering emulsion prepared from buckwheat protein complex;

[0074] Fig. 9 β-carotene retention rate of Pickering emulsion prepared from buckwheat protein complex. DETAILED DESCRIPTION

[0075] The principles and features of the present invention are described below in conjunction with the accompanying drawings. The examples are only used to explain the present invention and are not used to limit the scope of the present invention. If specific conditions are not specified in the embodiments, they are carried out according to normal conditions or conditions recommended by the manufacturer. If the manufacturer is not specified for the reagents or instruments used, they are all conventional products that can be purchased commercially.

[0076] In the following examples, the buckwheat protein hydrogel was prepared by the following method:

[0077] S1: Dissolve the tartary buckwheat flour with a PBS buffer solution having a concentration of 0.01 mol / L and a pH value of 7, place it on a magnetic stirrer and stir for 2 hours, centrifuge it at 4000 r / min for 30 minutes to obtain a first supernatant, add ammonium sulfate to make the concentration of ammonium sulfate in the first supernatant 40wt%, stir it for 60 minutes, centrifuge it at 10000 r / min for 15 minutes to obtain a second supernatant, continue to add ammonium sulfate, the concentration of ammonium sulfate in the second supernatant is 80wt%, continue to stir for 60 minutes, centrifuge it at 10000 r / min for 15 minutes to obtain a precipitate; wherein, the mass volume ratio of tartary buckwheat flour and PBS buffer solution is 1g:10mL; redissolve the obtained precipitate with water, dialyze it at 4°C with a 3500Da dialysis bag for 48 hours (change the water every 4 hours), adjust the pH value to 9 with a 1 mol / L NaOH solution, and then freeze-dry it at -76°C and 5Pa for 48 hours to obtain tartary buckwheat protein;

[0078] S2: dissolving the tartary buckwheat protein obtained in step S1 in a PBS buffer solution with a concentration of 0.01 mol / L and a pH value of 7 to obtain a tartary buckwheat protein solution with a concentration of 6 wt%, stirring the solution at room temperature for 2 hours, then placing the solution in a refrigerator and fully hydrating the solution at 4°C for 12 hours, adjusting the pH value to 8 with a 1 mol / L NaOH solution to obtain a hydrated tartary buckwheat protein solution; placing the obtained hydrated tartary buckwheat protein solution in a refrigerated centrifuge, centrifuging the solution at 10,000 r / min for 15 minutes, heating the supernatant in a water bath at 80°C for 10 minutes, then treating the solution in an ice-water bath and diluting the solution to obtain a tartary buckwheat protein hydrogel.

[0079] Embodiment 1:

[0080] A tartary buckwheat protein complex, the preparation method of which comprises the following steps:

[0081] (1) diluting the buckwheat protein hydrogel with distilled water, adjusting the pH value to 9 with 0.1 mol / L NaOH to prepare a protein hydrogel (concentration 0.02 g / mL);

[0082] (2) mixing quercetin and ethanol solution (60 vt%), adjusting the pH value to 9 with 0.1 mol / L NaOH to prepare a quercetin solution (concentration 0.01 g / mL);

[0083] (3) The protein hydrogel prepared in step (1) and the quercetin solution prepared in step (2) were mixed in a volume ratio of 1:1, heated in a water bath at 90° C. for 60 min, ultrasonically emulsified for 10 min using a cell disruptor at an ultrasonic power of 450 W, and freeze-dried at -76° C. and 5 Pa for 45 h to obtain a buckwheat protein complex.

[0084] A Pickering emulsion, the preparation method of which comprises the following steps:

[0085] The above-mentioned buckwheat protein complex was dissolved in distilled water at a mass volume ratio of 0.2 g:20 mL to obtain an aqueous phase, soybean oil was added at a volume ratio of 1:1, and ultrasonic emulsification was performed using a cell disruptor at an ultrasonic power of 450 W for 10 minutes to obtain a Pickering emulsion.

[0086] A β-carotene loaded Pickering emulsion, the preparation method of which comprises the following steps:

[0087] β-carotene was dissolved in soybean oil at a mass volume ratio of 1 mg:1 mL, and ultrasonic emulsification was performed for the first time at an ultrasonic power of 450 W for 10 min using a cell disruptor. Then, buckwheat protein complex solution (concentration of 10 mg / mL) was added at a volume ratio of 1:1, and ultrasonic emulsification was performed for the second time at an ultrasonic power of 450 W for 10 min using a cell disruptor to obtain a Pickering emulsion loaded with β-carotene.

[0088] Embodiment 2:

[0089] A tartary buckwheat protein complex, the preparation method of which comprises the following steps:

[0090] (1) The buckwheat protein hydrogel was diluted with distilled water, and the pH value was adjusted to 8.5 with 0.1 mol / L NaOH to prepare a protein hydrogel (concentration 0.015 g / mL);

[0091] (2) mixing quercetin and ethanol solution (50 vt%), adjusting the pH value to 8.5 with 0.1 mol / L NaOH to prepare a quercetin solution (concentration 0.008 g / mL);

[0092] (3) The protein hydrogel prepared in step (1) and the quercetin solution prepared in step (2) were mixed in a volume ratio of 1:0.8, heated in a water bath at 95° C. for 70 min, ultrasonically emulsified for 20 min using a cell disruptor at an ultrasonic power of 450 W, and freeze-dried at -80° C. and 0 Pa for 40 h to obtain a buckwheat protein complex.

[0093] A Pickering emulsion, the preparation method of which comprises the following steps:

[0094] The above-mentioned buckwheat protein complex was dissolved in distilled water at a mass volume ratio of 0.2 g:10 mL to obtain an aqueous phase, soybean oil was added at a volume ratio of 1:1, and ultrasonic emulsification was performed for 20 minutes using a cell disruptor at an ultrasonic power of 450 W to obtain a Pickering emulsion.

[0095] A beta-carotene-loaded Pickering emulsion, the preparation method of which comprises the following steps: dissolving beta-carotene in soybean oil at a mass volume ratio of 1 mg:1 mL, using a cell disruptor to perform ultrasonic emulsification for 10 minutes at an ultrasonic power of 450 W for the first time, then adding a tartary buckwheat protein complex solution (with a concentration of 10 mg / mL) at a volume ratio of 1:1, and using a cell disruptor to perform ultrasonic emulsification for 20 minutes at an ultrasonic power of 450 W for the second time to obtain the beta-carotene-loaded Pickering emulsion.

[0096] Embodiment 3:

[0097] A tartary buckwheat protein complex, the preparation method of which comprises the following steps:

[0098] (1) diluting the buckwheat protein hydrogel with distilled water, adjusting the pH value to 9.5 with 0.1 mol / L NaOH to prepare a protein hydrogel (concentration 0.025 g / mL);

[0099] (2) mixing quercetin and ethanol solution (70 vt%), adjusting the pH value to 9.5 with 0.1 mol / L NaOH to prepare a quercetin solution (concentration 0.012 g / mL);

[0100] (3) The protein hydrogel prepared in step (1) and the quercetin solution prepared in step (2) were mixed in a volume ratio of 1:1.2, heated in a water bath at 100° C. for 50 min, ultrasonically emulsified for 30 min using a cell disruptor at an ultrasonic power of 450 W, and freeze-dried at -70° C. and 6 Pa for 50 h to obtain a buckwheat protein complex.

[0101] A Pickering emulsion, the preparation method of which comprises the following steps:

[0102] The above-mentioned buckwheat protein complex was dissolved in distilled water at a mass volume ratio of 0.2 g:20 mL to obtain an aqueous phase, soybean oil was added at a volume ratio of 1:1, and ultrasonic emulsification was performed for 30 minutes using a cell disruptor at an ultrasonic power of 450 W to obtain a Pickering emulsion.

[0103] A beta-carotene-loaded Pickering emulsion, the preparation method of which comprises the following steps: dissolving beta-carotene in soybean oil at a mass volume ratio of 1 mg:1 mL, using a cell disruptor to perform ultrasonic emulsification for 10 minutes at an ultrasonic power of 450 W for the first time, then adding a tartary buckwheat protein complex solution (with a concentration of 10 mg / mL) at a volume ratio of 1:1, and using a cell disruptor to perform ultrasonic emulsification for 30 minutes at an ultrasonic power of 450 W for the second time to obtain the beta-carotene-loaded Pickering emulsion.

[0104] Embodiment 4-7:

[0105] A tartary buckwheat protein complex, the preparation method of which comprises the following steps:

[0106] In step (2), the concentrations of the quercetin solution were 0.008 g / mL, 0.009 g / mL, 0.011 g / mL, and 0.012 g / mL, respectively, and the rest was the same as in Example 1.

[0107] A Pickering emulsion, the preparation method of which comprises the following steps:

[0108] Same as Example 1.

[0109] A β-carotene loaded Pickering emulsion, the preparation method of which comprises the following steps:

[0110] Same as Example 1.

[0111] Embodiment 8-9:

[0112] A tartary buckwheat protein complex, the preparation method of which comprises the following steps:

[0113] In step (3), heating is performed in a water bath at 95° C. and 100° C. respectively, and the rest is the same as in Example 1.

[0114] A Pickering emulsion, the preparation method of which comprises the following steps:

[0115] Same as Example 1.

[0116] A β-carotene loaded Pickering emulsion, the preparation method of which comprises the following steps:

[0117] Same as Example 1.

[0118] Embodiment 10-11:

[0119] A tartary buckwheat protein complex, the preparation method of which comprises the following steps:

[0120] In step (3), ultrasonic emulsification was performed for 20 min and 30 min, and the rest was the same as in Example 1.

[0121] A Pickering emulsion, the preparation method of which comprises the following steps:

[0122] Same as Example 1.

[0123] A β-carotene loaded Pickering emulsion, the preparation method of which comprises the following steps:

[0124] Same as Example 1.

[0125] Comparative Example 1:

[0126] A tartary buckwheat protein complex, the preparation method of which comprises the following steps:

[0127] In step (1), distilled water is replaced by acetic acid, and the pH value is not adjusted. The rest is the same as in Example 1.

[0128] A Pickering emulsion, the preparation method of which comprises the following steps:

[0129] Same as Example 1.

[0130] A β-carotene loaded Pickering emulsion, the preparation method of which comprises the following steps:

[0131] Same as Example 1.

[0132] Comparative Example 2-3:

[0133] A tartary buckwheat protein complex, the preparation method of which comprises the following steps:

[0134] In step (3), heating in a water bath is performed at 60° C. and 70° C. respectively, and the rest is the same as in Example 1.

[0135] A Pickering emulsion, the preparation method of which comprises the following steps:

[0136] Same as Example 1.

[0137] A β-carotene loaded Pickering emulsion, the preparation method of which comprises the following steps:

[0138] Same as Example 1.

[0139] Comparative Example 4:

[0140] A tartary buckwheat protein complex, the preparation method of which comprises the following steps:

[0141] In step (3), ultrasonic emulsification was performed for 5 minutes, and the rest was the same as in Example 1.

[0142] A Pickering emulsion, the preparation method of which comprises the following steps:

[0143] Same as Example 1.

[0144] A β-carotene loaded Pickering emulsion, the preparation method of which comprises the following steps:

[0145] Same as Example 1.

[0146] Test example

[0147] 1. Performance test of buckwheat protein complex

[0148] The buckwheat protein complexes prepared in Examples 1, 4-11 and Comparative Examples 1-4 were tested as follows:

[0149] 1. Particle size and zeta potential analysis

[0150] The specific method for particle size and ζ potential analysis is as follows: dilute the protein concentration of the complex to 0.5 mg / mL with distilled water, set the test temperature to 25°C, use a quartz cuvette, test each sample three times, and use a nanoparticle size analyzer (ZEN3600) to measure the complex solution. The results are shown in Figure 1-2 .

[0151] Depend on Figure 1-2 It can be seen that the composite prepared by the present invention reaches the nanometer level, and the absolute value of the potential reaches about -40mv, indicating that the composite has a smaller particle size and better stability.

[0152] 2. Emulsification activity and emulsification stability

[0153] The buckwheat protein complex solution and soybean oil were mixed in a centrifuge tube at a ratio of 4:1, and ultrasonic emulsification was performed for 5 minutes using a cell disruptor at a power of 450W to prepare an emulsion. 50 μL of the emulsion was quickly drawn from the bottom of the centrifuge tube, and the solution was diluted to 50 mL with 0.1% (w / w) sodium dodecyl sulfate (SDS). The absorbance A0 was measured at a wavelength of 500 nm using an ultraviolet spectrophotometer. After the complex sample was allowed to stand for 10 minutes, 50 μL of the emulsion was drawn from the bottom of the centrifuge tube using the same method, and the absorbance A was measured after dilution. 10 The emulsifying activity index (EAI) and emulsifying stability (ESI) of the tartary buckwheat protein complex were calculated according to the formula:

[0154]

[0155] Wherein, N represents the dilution multiple of the complex; Φ represents the volume ratio of the oil phase in the emulsion; C represents the initial concentration of buckwheat protein (mg / mL); A0, A 10 represent the initial absorbance and the absorbance after 10 min, respectively; t 10 -t0 is the time difference between two measurements, which is 10 minutes.

[0156] Results Figure 3-4 As can be seen from the figure, quercetin can significantly improve the emulsifying activity and emulsifying stability of buckwheat protein, and the complex prepared by the present invention has good emulsifying activity and emulsifying stability.

[0157] 3. Surface hydrophobicity

[0158] The changes in the surface hydrophobicity of the sample were measured using a fluorescent probe (ANS). 10, 20, 30, 40, and 50 μL of protein solution (1 mg / mL) were added to a mixed solution of phosphate buffered saline and ANS solution, and the mixture was stirred thoroughly by vortex vibration. Then, a series of samples without ANS solution were used as blank controls to measure their fluorescence intensity. A linear equation was constructed using the protein concentration (horizontal axis) and the sample fluorescence intensity value (vertical axis) without the blank control (protein solution without ANS). The slope is the surface hydrophobicity. The results are shown in Figure 5 .

[0159] Depend on Figure 5 It can be seen that quercetin can significantly improve the surface hydrophobicity of buckwheat protein. The prepared composite has good surface hydrophobicity and can be used to stabilize Pickering emulsion.

[0160] 2. Pickering emulsion performance test

[0161] The Pickering emulsions prepared in Examples 1, 4-11 and Comparative Examples 1-4 were tested as follows:

[0162] 1. Pickering emulsion creaming index

[0163] Take the freshly prepared Pickering emulsion and place it in a centrifuge tube and store it at room temperature for 30 days. Observe and measure the emulsion on the 1st, 3rd, 7th, 15th and 30th day respectively, and record the stratification of the emulsion. Calculate the emulsion index (Cl) according to the formula. The results are shown in Figure 6 .

[0164] CI (%) = (H1 / H2) × 100%

[0165] Among them, H1 refers to the height of the lower clear night, and H2 refers to the total height of the emulsion.

[0166] Depend on Figure 6 It can be seen that the buckwheat protein-quercetin nanocomposite prepared by the present invention and its stable Pickering emulsion have a very small creaming index, indicating good emulsion stability.

[0167] 2. Thermal stability

[0168] The freshly prepared emulsion was placed in a centrifuge tube, heated in a water bath at 100°C for 20 min, cooled to room temperature, and the state of the emulsion was observed. Figure 7 .

[0169] Depend on Figure 7 It can be seen that the Pickering emulsion prepared by the present invention is less affected by temperature and has good thermal stability.

[0170] 3. Salt ion stability

[0171] Take 4 mL of fresh emulsion in a centrifuge tube, add 4 mL of 300 mmol / L NaCl solution, mix well, store at room temperature, and observe the emulsion state after 1 hour. The results are shown in Figure 8 .

[0172] Depend on Figure 8 It can be seen that the Pickering emulsion prepared by the present invention is less affected by salt ions and has good salt ion stability.

[0173] 3. Performance test of Pickering emulsion loaded with β-carotene

[0174] The β-carotene retention rate of the Pickering emulsions loaded with β-carotene obtained in Examples 1, 8-11 and Comparative Example 4 was determined, and the β-carotene content was determined by the standard curve method. The specific determination method is:

[0175] (1) Preparation of β-carotene standard curve

[0176] Accurately weigh 2 mg of β-carotene and dilute to 20 mL with n-hexane to obtain a 0.1 mg / mL β-carotene standard solution. Take five 10 mL standard volumetric flasks, add 1, 2, 3, 4, and 5 mL of the β-carotene standard solution, respectively, and dilute to volume with n-hexane. Then use an ultraviolet spectrophotometer to measure the absorbance A at 450 nm, and make a standard curve with β-carotene concentration and absorbance;

[0177] (2) The β-carotene-loaded Pickering emulsion was placed in a 10 mL centrifuge tube and stored at room temperature without protection from light. The β-carotene content in the Pickering emulsion was measured at 1, 3, 5, and 7 days. Take 0.5 mL of the emulsion and place it in a 10 mL centrifuge tube. Then inject 3 mL of n-hexane solution and 2 mL of anhydrous ethanol. Vortex and oscillate to completely break the emulsion. Let it stand at room temperature for 10 min. Then take the n-hexane solution on the upper layer and aspirate it and collect it in a 25 mL volumetric flask. Then add 3 mL of n-hexane solution and repeat the operation for a total of 3 times. Put all the n-hexane collected in the 3 times in a 25 mL volumetric flask and make up to volume with n-hexane solution. Then take the solution after making up to volume and dilute it 5 times with n-hexane. Measure the absorbance at 450 nm. Calculate its content according to the standard curve of β-carotene. Use the following formula to calculate the retention rate of β-carotene. The results are shown in Table 2. Fig. 9 :

[0178]

[0179] Where: C t represents the concentration of β-carotene in the Pickering emulsion after storage at room temperature for t days; C0 represents the concentration of β-carotene in the original Pickering emulsion.

[0180] Depend on Fig. 9 It can be seen that during the storage process without protection from light, β-carotene is gradually degraded, and the Pickering emulsion can effectively slow down the oxidation and photodecomposition of β-carotene. The Pickering emulsion stabilized by the buckwheat protein-quercetin nanocomposite prepared within the scope of rights protected by this patent is particularly effective. In the absence of protection from light, the retention rate of β-carotene can reach more than 85% after 7 days, indicating that the Pickering emulsion has good antioxidant and anti-photolysis capabilities, and can better protect β-carotene.

[0181] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for preparing a tartary buckwheat protein complex, It is characterized in that The following steps are involved: (1) diluting the buckwheat protein hydrogel with distilled water and adjusting the pH value to prepare the protein hydrogel; (2) mixing quercetin and an ethanol solution and adjusting the pH value to prepare a quercetin solution; (3) mixing the protein hydrogel prepared in step (1) and the quercetin solution prepared in step (2), heating in a water bath, ultrasonic emulsification, and freeze drying to prepare a buckwheat protein complex.

2. The method for preparing the tartary buckwheat protein complex according to claim 1, It is characterized in that In step (1), the pH value is adjusted to 8.5-9.

5.

3. The method for preparing the tartary buckwheat protein complex according to claim 1, It is characterized in that In step (2), the concentration of the quercetin solution is 0.008-0.012 g / mL.

4. The method for preparing the tartary buckwheat protein complex according to claim 1, It is characterized in that In step (2), the pH value is adjusted to 8.5-9.

5.

5. The method for preparing the tartary buckwheat protein complex according to claim 1, It is characterized in that In step (3), the mixture is heated in a water bath at 90-100° C. for 50-70 min.

6. The tartary buckwheat protein complex obtained by the method for preparing the tartary buckwheat protein complex according to any one of claims 1 to 5.

7. Use of the tartary buckwheat protein complex according to claim 6 in preparing Pickering emulsion.

8. Use of the tartary buckwheat protein complex according to claim 7 in preparing Pickering emulsion, It is characterized in that The preparation method of Pickering emulsion comprises the following steps: dissolving the tartary buckwheat protein complex according to claim 6 in distilled water to obtain a water phase, adding an oil phase, and ultrasonically emulsifying to obtain the Pickering emulsion.

9. Use of the tartary buckwheat protein complex according to claim 7 in preparing Pickering emulsion, It is characterized in that The Pickering emulsion is a Pickering emulsion loaded with beta-carotene.

10. Use of the tartary buckwheat protein complex according to claim 9 in preparing Pickering emulsion, It is characterized in that The preparation method of the Pickering emulsion loaded with beta-carotene comprises the following steps: dissolving beta-carotene in an oil phase, performing ultrasonic emulsification for the first time, then adding a tartary buckwheat protein complex solution, performing ultrasonic emulsification for the second time, and preparing the Pickering emulsion loaded with beta-carotene.

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