Method for improving stability of black corn anthocyanin-protein compound

Abstract

A method for improving stability of a black corn anthocyanin-protein compound comprises the following steps: adding a stabilizer which is glucose or maltodextrin into a black corn anthocyanin-protein compound solution; when the stabilizer is glucose, the concentration of the glucose in the solution is 1-5%; when the stabilizer is maltodextrin, the concentration of the maltodextrin in the solution is 1-3%. According to the invention, maltodextrin or glucose is adopted as a stabilizer, so that the stability of the black corn anthocyanin-protein compound is improved. The maltodextrin is combined with the protein part or the anthocyanin part in the compound, so that the structure of the compound is more compact and stable, dissociation and denaturation of the compound in the external environment are reduced, the combination state between the anthocyanin and the protein is kept, the stability of the anthocyanin is indirectly protected, meanwhile, the stability of the protein structure is facilitated, and the stability of the protein structure is improved. Therefore, the protection effect of the protein on the anthocyanin can be better exerted.

Description

Technical Field

[0001] The invention relates to the field of anthocyanin-protein complexes, in particular to a method for improving the stability of black corn anthocyanin-protein complexes. Background Art

[0002] Black corn, also known as black glutinous corn, is a general term for corn with black and purple kernels. It is also called purple corn and belongs to the genus Zea of ​​the Poaceae family. It is rich in anthocyanins, polysaccharides, amino acids and various minerals, and has extremely high nutritional value. The content of protein, fat and selenium in black corn kernels is 1.23 times, 1.3 times and 3 to 8.5 times that of ordinary corn, respectively. The content of anthocyanin is 4.3 times that of blueberries. Compared with ordinary corn, it has higher nutritional value and better health care effects, and has multiple functions such as anti-oxidation, anti-cancer, and memory regulation. At present, there are related products such as black corn vinegar, black corn wine, and black corn noodles on the market, but the small number of product types limits the further upgrading of the black corn industry, and it is urgent to develop new black corn products.

[0003] Anthocyanin is a natural colored flavonoid phenolic compound formed by the combination of anthocyanidins and various monosaccharides in plants to form glycosides. It is a general term for natural pigments in plants, which can make the roots, stems, flowers and fruits of many plants appear in different colors such as orange, pink, red, purple and blue. Studies have shown that anthocyanins have a variety of unique physiological activities, such as preventing heart disease, lowering blood sugar, anti-aging, anti-cancer and anti-inflammatory effects. Due to their natural origin, non-toxic safety and excellent biological functions, anthocyanins have attracted widespread attention from researchers and are often used in the fields of medicine, food, cosmetics, etc. However, anthocyanins have low stability to environmental conditions (such as temperature, pH, light, water, oxygen, metal ions, etc.) during processing and storage, which makes their application in food face huge challenges.

[0004] Proteins have good surface properties, self-assembly properties and gelation characteristics. They can form a combination through non-covalent and covalent interactions without changing the structure of anthocyanins. Anthocyanins are fixed in the protein structure cavity, which can reduce the loss of pyranose cations and thus have a positive effect on the stability of anthocyanins. After anthocyanins are complexed with proteins, the functional properties of proteins will be improved, such as surface hydrophobicity, emulsification, and foaming properties. The interaction between the two can affect each other's morphology, antioxidant capacity, thermal stability, emulsification properties, and bioavailability in the food system.

[0005] However, the stability of anthocyanin-protein complexes is not high. On the one hand, anthocyanin is a natural water-soluble pigment, which is susceptible to environmental stress in aqueous solution, causing degradation or structural changes in anthocyanin, thereby affecting its binding stability with protein; on the other hand, the binding mode between anthocyanin and protein is mainly through non-covalent binding to reduce the loss of anthocyanin, among which hydrogen bonds and hydrophobic effects are the main forces of anthocyanin-protein complexes. This binding mode is also easily affected by multiple factors, resulting in the instability of the complex.

[0006] In order to improve the stability of anthocyanin-protein complexes, extend the shelf life of anthocyanin-protein complexes and maintain their activity, the existing technology generally uses technical means such as low temperature, avoiding vibration and light, and there is no way to improve the stability of black corn anthocyanin-protein complexes by adding stabilizers. Summary of the invention

[0007] In order to solve the deficiencies in the prior art, the present invention provides a method for improving the stability of black corn anthocyanin-protein complex.

[0008] In order to achieve the above-mentioned purpose, the specific scheme adopted by the present invention is: a method for improving the stability of black corn anthocyanin-protein complex, adding a stabilizer to the black corn anthocyanin-protein complex solution, the stabilizer is glucose or maltodextrin; when the stabilizer is glucose, the concentration of glucose in the solution is 1-5%; when the stabilizer is maltodextrin, the concentration of maltodextrin in the solution is 1-3%.

[0009] As a further optimization of the above technical solution, the concentration of the black corn anthocyanin-protein complex solution is 2-8 mg / mL.

[0010] As a further optimization of the above technical solution, the glucose is anhydrous glucose.

[0011] As a further optimization of the above technical solution, the glucose equivalent of maltodextrin is 10-20.

[0012] As a further optimization of the above technical solution, when the stabilizer is maltodextrin, the concentration of maltodextrin in the solution is 2%.

[0013] As a further optimization of the above technical solution, when the stabilizer is maltodextrin, the concentration of maltodextrin in the solution is 1%.

[0014] As a further optimization of the above technical solution, when the stabilizer is maltodextrin, the concentration of maltodextrin in the solution is 3%.

[0015] As a further optimization of the above technical solution, when the stabilizer is glucose, the concentration of glucose in the solution is 5%.

[0016] As a further optimization of the above technical solution, when the stabilizer is glucose, the concentration of glucose in the solution is 1%.

[0017] As a further optimization of the above technical solution, the preparation method of the black corn anthocyanin-protein complex is to defatt the black corn flour with petroleum ether, then add ethanol aqueous solution to the defatted black corn flour, stir, filter, take the supernatant and add NaCl solution, centrifuge, dialyze the precipitate, and freeze-dry to obtain the black corn anthocyanin-protein complex.

[0018] Compared with the prior art, the present invention has the following beneficial effects:

[0019] The invention adopts maltodextrin or glucose as a stabilizer to improve the stability of the black corn anthocyanin-protein complex.

[0020] Maltodextrin can interact with anthocyanin-protein complexes to further stabilize the overall structure of the complex. By binding to the protein or anthocyanin part of the complex, the structure of the complex becomes tighter and more stable, thereby reducing its dissociation and denaturation in the external environment, maintaining the binding state between anthocyanin and protein, thereby indirectly protecting the stability of anthocyanin, and also contributing to the stability of protein structure, thereby better exerting the protective effect of protein on anthocyanin.

[0021] The principle by which glucose improves the stability of black corn anthocyanin-protein complex is that the glucose molecule has a large number of hydroxyl groups, which can compete with water molecules for the interaction sites with the black corn anthocyanin-protein complex in the solution, reducing the adverse effects of water molecules on the complex; glucose can act as an antioxidant, thereby protecting the anthocyanin-protein complex from the attack of free radicals; glucose molecules have a certain spatial structure, which can fill in the area around the anthocyanin-protein complex in the solution, preventing the complex molecules from contacting other substances that may cause its decomposition.

[0022] The present invention extracts black corn anthocyanin-protein complexes through ethanol aqueous solution, combines green technology with the use of green solvents, is environmentally friendly, and the stability of the black corn anthocyanin-protein complex obtained by the method of the present invention can be significantly improved when glucose or maltodextrin is added as a stabilizer. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 The figures are the physical pictures of the samples of Examples 1-3 and Comparative Examples 1-3;

[0024] Figure 2 It is a line graph of the absorbance values ​​of the samples of Examples 1-3 and Comparative Examples 1-3;

[0025] Figure 3 It is a physical picture of the samples of Example 6, Comparative Examples 1, and 4-7;

[0026] Figure 4 It is a line graph of the absorbance values ​​of the samples of Example 6, Comparative Examples 1, and 4-7;

[0027] Figure 5 The figures are the physical pictures of the samples of Comparative Example 1 and Comparative Examples 8-12;

[0028] Figure 6 It is a bar graph of the absorbance values ​​of the samples of Comparative Example 1 and Comparative Examples 8-12;

[0029] Figure 7 The figures are the physical pictures of the samples of comparative example 1 and comparative example 13;

[0030] Figure 8 It is a line graph of the absorbance values ​​of the samples of Comparative Example 1 and Comparative Example 13;

[0031] Fig. 9 It is a bar graph showing the effect of temperature on the preservation rate of anthocyanins in black corn anthocyanin-protein complex;

[0032] Fig.10 It is a bar graph showing the effect of pH on the anthocyanin preservation rate of black corn anthocyanin-protein complex;

[0033] Fig.11 The SDS-PAGE profile of black corn anthocyanin-protein complex;

[0034] Fig.12 is the UV spectrum of black corn protein and black corn anthocyanin-protein complex;

[0035] Fig.13 This is the infrared spectrum of black corn protein and black corn anthocyanin-protein complex. DETAILED DESCRIPTION

[0036] The technical solution of the present invention is further elaborated in detail below in conjunction with specific embodiments. The parts of the present invention that are not described and disclosed in detail in the following embodiments should be understood as the prior art known or should be known to those skilled in the art.

[0037] The invention calculates the corresponding masses of stabilizers with different mass fractions according to the volume of the black corn anthocyanin-protein complex solution, and then mixes the black corn anthocyanin-protein complex solution with the stabilizer.

[0038] In the following embodiments of the present invention, when the stabilizer selected is glucose, the glucose is anhydrous glucose. Anhydrous glucose does not contain water of crystallization and has higher purity. It can ensure the stability of product quality in products that require precise control of the moisture content in the formula. The production process of monohydrate glucose is relatively simple, and the cost is usually lower than that of anhydrous ordinary sugar. When the stabilizer selected is maltodextrin, the glucose equivalent of maltodextrin is 10-20, the glucose equivalent (DE value) is higher, the solubility is better, it can be quickly dissolved, and it provides a certain consistency and taste. Maltodextrin with a high DE value has a lower cost and is easier to obtain.

[0039] Example 1

[0040] A method for improving the stability of black corn anthocyanin-protein complex, the method comprising the following steps:

[0041] 1) Select mature, fresh and dried black corn kernels, use a universal grinder to grind the black corn kernels into powder, and then sieve the black corn powder through 80 mesh for later use.

[0042] 2) According to the ratio of 1:10, weigh 10g of black corn flour, add 100ml of petroleum ether, stir magnetically at room temperature for 8h, dry the defatted corn flour on filter paper, and place it in a cool and ventilated place to dry the petroleum ether.

[0043] 3) Add 100 mL of 80% ethanol solution to the corn flour and extract with magnetic stirring at 50°C for 4 h.

[0044] 4) Filtration: Filter the sample that has been magnetically stirred at 50°C for 4 hours, take the supernatant and remove the residue precipitate.

[0045] A 1% NaCl solution was added to the solution extracted with 80% ethanol at a volume ratio of 1:1 and the mixture was allowed to stand at 4°C for 12 h.

[0046] Centrifugation: Set the centrifuge speed to 6000 r / min, the centrifugation time to 15 min, and the temperature to 4 °C to centrifuge the sample after it has been placed for 12 hours.

[0047] Dialysis: Collect the precipitate from the centrifuged sample, transfer it into a dialysis bag, and place it in ice water for magnetic stirring.

[0048] Freeze drying: freeze drying the dialyzed sample to obtain alcohol-soluble protein, namely, black corn anthocyanin-protein complex.

[0049] 5) Dissolve the black corn anthocyanin-protein complex in a phosphate buffer with a pH value of 7.4 to obtain a black corn anthocyanin-protein complex solution with a concentration of 4 mg / mL. Take 15 ml of the black corn anthocyanin-protein complex solution, add 0.15 g of maltodextrin, and stir evenly. The glucose equivalent of maltodextrin is 15, and the concentration of maltodextrin in the solution is 1%.

[0050] Example 2

[0051] A method for improving the stability of black corn anthocyanin-protein complex, the method comprising the following steps:

[0052] The preparation method of the black corn anthocyanin-protein complex, namely steps 1) to 4), is the same as that of Example 1 and will not be described in detail herein.

[0053] 5) Dissolve the black corn anthocyanin-protein complex in a phosphate buffer with a pH value of 7.4 to obtain a black corn anthocyanin-protein complex solution with a concentration of 4 mg / mL; take 15 ml of the black corn anthocyanin-protein complex solution, add 0.3 g of maltodextrin, and stir evenly; the glucose equivalent of maltodextrin is 15, and the concentration of maltodextrin in the solution is 2%.

[0054] Example 3

[0055] A method for improving the stability of black corn anthocyanin-protein complex, the method comprising the following steps:

[0056] The preparation method of the black corn anthocyanin-protein complex, namely steps 1) to 4), is the same as that of Example 1 and will not be described in detail herein.

[0057] 5) Dissolve the black corn anthocyanin-protein complex in a phosphate buffer with a pH value of 7.4 to obtain a black corn anthocyanin-protein complex solution with a concentration of 4 mg / mL; take 15 ml of the black corn anthocyanin-protein complex solution, add 0.45 g of maltodextrin, and stir evenly; the glucose equivalent of maltodextrin is 15, and the concentration of maltodextrin in the solution is 3%.

[0058] Example 4

[0059] A method for improving the stability of black corn anthocyanin-protein complex, the method comprising the following steps:

[0060] The preparation method of the black corn anthocyanin-protein complex, namely steps 1) to 4), is the same as that of Example 1 and will not be described in detail herein.

[0061] 5) Dissolve the black corn anthocyanin-protein complex in a phosphate buffer with a pH value of 7.4 to obtain a black corn anthocyanin-protein complex solution with a concentration of 4 mg / mL. Take 15 ml of the black corn anthocyanin-protein complex solution, add 0.15 g of anhydrous glucose, and stir evenly. The concentration of glucose in the solution is 1%.

[0062] Example 5

[0063] A method for improving the stability of black corn anthocyanin-protein complex, the method comprising the following steps:

[0064] The preparation method of the black corn anthocyanin-protein complex, namely steps 1) to 4), is the same as that of Example 1 and will not be described in detail herein.

[0065] 5) Dissolve the black corn anthocyanin-protein complex in a phosphate buffer with a pH value of 7.4 to obtain a black corn anthocyanin-protein complex solution with a concentration of 4 mg / mL. Take 15 ml of the black corn anthocyanin-protein complex solution, add 0.45 g of anhydrous glucose, and stir evenly. The concentration of glucose in the solution is 3%.

[0066] Example 6

[0067] A method for improving the stability of black corn anthocyanin-protein complex, the method comprising the following steps:

[0068] The preparation method of the black corn anthocyanin-protein complex, namely steps 1) to 4), is the same as that of Example 1 and will not be described in detail herein.

[0069] 5) Dissolve the black corn anthocyanin-protein complex in a phosphate buffer with a pH value of 7.4 to obtain a black corn anthocyanin-protein complex solution with a concentration of 4 mg / mL. Take 15 ml of the black corn anthocyanin-protein complex solution, add 0.75 g of anhydrous glucose, and stir evenly. The concentration of glucose in the solution is 5%.

[0070] Example 7

[0071] A method for improving the stability of black corn anthocyanin-protein complex, the method comprising the following steps:

[0072] The preparation method of the black corn anthocyanin-protein complex, namely steps 1) to 4), is the same as that of Example 1 and will not be described in detail herein.

[0073] 5) Dissolve the black corn anthocyanin-protein complex in a phosphate buffer with a pH value of 7.4 to obtain a black corn anthocyanin-protein complex solution with a concentration of 2 mg / mL; take 15 ml of the black corn anthocyanin-protein complex solution, add 0.3 g of maltodextrin, and stir evenly; the glucose equivalent of maltodextrin is 15, and the concentration of maltodextrin in the solution is 2%.

[0074] Example 8

[0075] A method for improving the stability of black corn anthocyanin-protein complex, the method comprising the following steps:

[0076] The preparation method of the black corn anthocyanin-protein complex, namely steps 1) to 4), is the same as that of Example 1 and will not be described in detail herein.

[0077] 5) Dissolve the black corn anthocyanin-protein complex in a phosphate buffer with a pH value of 7.4 to obtain a black corn anthocyanin-protein complex solution with a concentration of 8 mg / mL. Take 15 ml of the black corn anthocyanin-protein complex solution and add 0.75 g of glucose, that is, the concentration of glucose in the solution is 5%.

[0078] Comparative Example 1 (i.e., control group)

[0079] Comparative Example 1 is a dilution solution, and the preparation method of the dilution solution is: using a phosphate buffer solution with a pH value of 7.4, mixing the black corn anthocyanin-protein complex with the phosphate buffer solution, and controlling the concentration of the black corn anthocyanin-protein complex solution to be 4 mg / mL.

[0080] Comparative Examples 2-3

[0081] The overall steps of Comparative Example 2-3 are the same as those of Example 1-3, except that the amount of maltodextrin added is adjusted so that the concentrations of maltodextrin in Comparative Examples 2 and 3 are 4% and 5%, respectively.

[0082] Comparative Examples 4-7

[0083] The overall steps of Comparative Examples 3-6 are the same as those of Example 4-6, except that the amount of glucose added is adjusted so that the concentrations of glucose in Comparative Examples 4-7 are 10%, 15%, 20%, and 25%, respectively.

[0084] Comparative Examples 8-12

[0085] The overall steps of Comparative Examples 8-12 are the same as those of Example 1, except that maltodextrin is replaced with ascorbic acid, and the amount of ascorbic acid added is adjusted so that the concentrations of ascorbic acid in Comparative Examples 8-12 are 2%, 4%, 6%, 8%, and 10%, respectively.

[0086] Comparative Example 13

[0087] The overall steps of Comparative Example 13 are the same as those of Example 1, except that maltodextrin is replaced with a compound stabilizer having a concentration of 2% maltodextrin and 5% glucose.

[0088] Results Analysis

[0089] <Effect of maltodextrin on the stability of black corn anthocyanin-protein complex>

[0090] The test results of Examples 1-3 and Comparative Examples 1-3 were analyzed. Figure 1 , 2 It can be seen that after adding maltodextrin to the black corn anthocyanin-protein complex solution, when the maltodextrin concentration of Examples 1-3 is 1%, 2%, and 3%, the absorbance values ​​are all higher than that of Comparative Example 1 (diluted solution), and the absorbance values ​​all show a downward trend, which has a protective effect on the black corn anthocyanin-protein complex; when the maltodextrin concentration of Comparative Examples 2-3 is 4% and 5%, the absorbance values ​​are all lower than that of Comparative Example 1.

[0091] As time goes by, the absorbance values ​​of anthocyanins in each experimental group show a downward trend. However, when low concentrations (1%, 2%, and 3%) of maltodextrin are added, the absorbance value of the solution decreases less and is always higher than that of Comparative Example 1, indicating that low concentrations of maltodextrin can improve the stability of the black corn anthocyanin-protein complex; and when the concentration of maltodextrin is 2%, the effect of maintaining the stability of the black corn anthocyanin-protein complex is best.

[0092] The reason for this analysis is that maltodextrin can interact with anthocyanin-protein complexes, further stabilizing the overall structure of the complex. This interaction may involve hydrogen bonds, van der Waals forces, etc. By binding to the protein part or anthocyanin part of the complex, the structure of the complex is made tighter and more stable, thereby reducing its dissociation and denaturation in the external environment. Maltodextrin focuses on maintaining the overall structural stability of the complex. By binding to the complex, the dissociation of the complex in the external environment is reduced, and the binding state between anthocyanins and proteins is maintained, thereby indirectly protecting the stability of anthocyanins. It also helps to stabilize the protein structure, thereby better exerting the protective effect of protein on anthocyanins.

[0093] As the concentration of maltodextrin increases, the space it occupies in the solution also increases. Maltodextrin molecules are a kind of high molecular carbohydrates, and the molecular chains can stretch in the solution. When the concentration increases, they can form a tighter wrapping layer around the anthocyanin-protein complex, thereby enhancing the steric hindrance effect; and maltodextrin can adjust the properties of the solution. High concentrations of maltodextrin can better control parameters such as the viscosity and osmotic pressure of the solution. Higher viscosity can slow down the movement speed of the complex molecules, allowing the complex to exist in a relatively stable environment. The increase in maltodextrin concentration helps to maintain a stable osmotic pressure environment, preventing the complex from losing water or absorbing water due to changes in osmotic pressure, thereby avoiding structural changes; functional groups such as hydroxyl groups in maltodextrin molecules can interact with anthocyanin-protein complexes through hydrogen bonds, thereby further stabilizing the structure of the complex. When the concentration of maltodextrin is further increased, due to the certain viscosity of maltodextrin, the higher the concentration, the less likely it is to completely dissolve in water. High concentrations of maltodextrin solutions will adsorb some anthocyanin-protein complexes, resulting in a decrease in the stability of black corn anthocyanin-protein complexes.

[0094] <Effect of Glucose on the Stability of Black Corn Anthocyanin-Protein Complex>

[0095] The test results of Example 6, Comparative Example 1, and Comparative Examples 4-7 were analyzed. Figure 3 , 4 It can be seen that after glucose was added to the black corn anthocyanin-protein complex solution, except for Example 6 when the glucose concentration was 5%, the absorbance values ​​of other groups were lower than that of Comparative Example 1, and the absorbance values ​​all showed a downward trend.

[0096] When the glucose concentration in Example 6 is 5%, the absorbance value is higher than that in Comparative Example 1, indicating that glucose forms a non-covalent complex with the anthocyanin-protein complex to improve the stability of the anthocyanin-protein complex, and low concentration of glucose can maintain the stability of the black corn anthocyanin-protein complex, but when it reaches a certain concentration (such as the glucose concentration exceeds 5%), this stabilizing effect will show a weakening trend.

[0097] Therefore, a certain concentration of glucose can be added during the processing to improve the stability of the black corn anthocyanin-protein complex, and a glucose concentration of 5% is preferred.

[0098] <Effect of ascorbic acid on the stability of black corn anthocyanin-protein complex>

[0099] Analyze the test results of Comparative Example 1 and Comparative Examples 8-12. Figure 5 , 6It can be seen that within the experimental range, after adding different concentrations of ascorbic acid, the absorbance of the black corn anthocyanin-protein complex solution showed a significant downward trend, indicating that the stability of the solution decreased, and with the increase of ascorbic acid concentration, the absorbance of the anthocyanin-protein complex decreased, and the solution was transparent, and yellow flocs appeared. This shows that ascorbic acid has a destructive effect on the stability of the black corn anthocyanin-protein complex, and the reason is that ascorbic acid itself is oxidized, and the produced hydrogen peroxide causes the structure of the anthocyanin-protein complex to change, resulting in a decrease in the stability of the anthocyanin-protein complex.

[0100] <Effect of compound stabilizer on the stability of black corn anthocyanin-protein complex>

[0101] Analyze the test results of Comparative Example 1 and Comparative Example 13. Figure 7 , 8 It can be seen that as time goes by, the absorbance values ​​of the comparative example 1 without adding the compound stabilizer and the solution with the compound stabilizer both show a decreasing trend, indicating that the stability of the solution decreases. The absorbance values ​​of the solution with the compound stabilizer are all less than that of the comparative example 1, indicating that the compound stabilizer reduces the stability of the black corn anthocyanin-protein complex. The poor compounding effect of glucose and maltodextrin may be due to the change of the structure caused by the combination of anthocyanins and proteins in the black corn anthocyanin-protein complex, resulting in a decrease in the absorbance of the solution. Therefore, it is best to use only a single stabilizer in the processing and production of the black corn anthocyanin-protein complex.

[0102] Further analysis of the reasons shows that, on the one hand, the protective mechanism of glucose on anthocyanins is different from that of maltodextrin. Glucose and anthocyanins are mainly combined through glycosidic bonds to form anthocyanin-glucoside, which plays a certain protective role on anthocyanins. This combination is relatively stable and can change the chemical structure and properties of anthocyanins to a certain extent, thereby enhancing their stability. Due to its relatively small molecular structure, glucose has relatively limited changes in the spatial structure of anthocyanins after combining with anthocyanins, but the anthocyanin-glucoside formed can reduce the contact area between anthocyanins and the external environment to a certain extent, thereby reducing the degree to which they are affected by external factors. The formation of anthocyanin-glucoside may also change the electron cloud distribution of anthocyanins, making it more difficult for them to undergo chemical reactions such as oxidation.

[0103] Maltodextrin is a polysaccharide composed of multiple glucose units connected by α-1,4 glycosidic bonds. It is mainly bonded to anthocyanins through non-covalent bonds such as hydrogen bonds and hydrophobic interactions. Polar groups such as hydroxyl groups in maltodextrin molecules can form hydrogen bonds with corresponding groups in anthocyanin molecules, wrapping or adsorbing anthocyanins around their molecular structure, thereby protecting anthocyanins. Maltodextrin has a larger molecular structure and spatial network, which can provide a more effective physical barrier for anthocyanins, greatly reducing the direct contact between anthocyanins and external environmental factors such as oxygen, light, and moisture. Maltodextrin reduces the water activity of the environment in which anthocyanins are located and reduces the impact of moisture on anthocyanins, thereby improving their stability during storage and processing.

[0104] As an embedding agent, maltodextrin forms non-covalent complexes with anthocyanins mainly through hydrogen bonds, hydrophobic interactions, etc., which improves the stability of anthocyanins. When the concentration of maltodextrin is too high, it over-embeds anthocyanins, resulting in a decrease in the absorbance of the solution. Maltodextrin acts directly on anthocyanin molecules, reduces the water activity of the environment in which anthocyanins are located, and reduces the impact of water on anthocyanins, thereby improving their stability during storage and processing.

[0105] On the other hand, some studies have found that the combination of glucose and maltodextrin can improve the stability of anthocyanins, as follows:

[0106] 1. Reduce degradation reactions: Maltodextrin has good film-forming properties and emulsification stability, and can form a protective film around anthocyanins, reducing their contact with the external environment, thereby reducing the probability of degradation reactions. Glucose can react with anthocyanins to produce products with antioxidant properties, further preventing the oxidative degradation of anthocyanins.

[0107] 2. Reduce water activity: Water activity is one of the important factors affecting the stability of anthocyanins. High water activity will accelerate the degradation of anthocyanins. Maltodextrin has strong water absorption and can reduce the water activity of the system, thereby slowing down the degradation of anthocyanins. When glucose and maltodextrin are compounded, the effect of reducing water activity may be more significant, further improving the stability of anthocyanins.

[0108] 3. Change the structure of anthocyanins: Glucose and maltodextrin may form hydrogen bonds or other chemical bonds with anthocyanins, increasing the steric hindrance of anthocyanin molecules and thus improving their stability.

[0109] However, the combination of glucose and maltodextrin in the present invention cannot improve the stability of the anthocyanin-protein complex, so it can be inferred that the mechanism by which maltodextrin protects anthocyanins is different from the mechanism by which maltodextrin protects anthocyanin-protein complex.

[0110] In addition, the present invention also studied the effects of temperature and pH value on the anthocyanin retention rate of the black corn anthocyanin-protein complex.

[0111] <Effect of Temperature on Anthocyanin Retention Rate of Black Corn Anthocyanin-Protein Complex>

[0112] As Fig. 9 shown, when the temperature is 35°C, with the passage of time, the anthocyanin retention rate of the black corn anthocyanin-protein complex shows a downward trend. The change in anthocyanin retention rate is not obvious between 1 - 2 hours. As the time extends, the change in anthocyanin retention rate becomes more obvious. When the time is 1 hour, the highest anthocyanin retention rate is 98.23%, and when the time is 4 hours, the lowest anthocyanin retention rate is 66.67%.

[0113] <Effect of pH on Anthocyanin Retention Rate of Black Corn Anthocyanin-Protein Complex>

[0114] As Fig.10 shown, when the pH is 4, with the passage of time, the anthocyanin retention rate of the black corn anthocyanin-protein complex shows a downward trend, and the change in anthocyanin retention rate becomes more obvious. When the time is 1 hour, the highest anthocyanin retention rate is 68.27%, and when the time is 4 hours, the lowest anthocyanin retention rate is 1.61%.

[0115] It should be noted that the black corn anthocyanin-protein complex used in the present invention is an anthocyanin-protein complex extracted from black corn through an ethanol aqueous solution. The use of green technology and green solvents in the extraction process is environmentally friendly. Moreover, for the black corn anthocyanin-protein complex obtained by this method, its stability can be significantly improved when adding glucose or maltodextrin as a stabilizer.

[0116] The following detections were carried out on both the black corn anthocyanin-protein complex prepared in the present invention and the black corn anthocyanin-protein complex extracted by the ethanol-ice water method:

[0117] Among them, the process of extracting the black corn anthocyanin-protein complex by the ethanol-ice water method is the same as that in Example 1, except that in step 4), an equal volume of pre-cooled ice water is added to the solution extracted with 80% ethanol above, and it is placed at 4°C for 24 hours.

[0118] The detection results are as follows:

[0119] 1) Determination of protein content

[0120] The BCA method for determining protein content has the characteristics of simple operation, high sensitivity, small coefficient of variation, accurate and reliable results, and little influence by interfering substances. The protein content can intuitively reflect the difference in extraction effect between ethanol-NaCl extraction and ethanol-ice water extraction. The protein content of the complex extracted by alcohol is higher, among which the protein content in the complex extracted by ethanol-NaCl is 24.81±0.45mg / g black corn, and the protein content in the complex extracted by ethanol-ice water is 28.45±0.34mg / g black corn. Table 1 Protein extraction content in black corn

[0121] 2) Determination of anthocyanin content

[0122] The pH differential method has sufficiently high accuracy, is relatively simple to operate, and is more conducive to wide application. The anthocyanin content can also intuitively reflect the difference in extraction effect between ethanol-NaCl extraction and ethanol-ice water extraction. The anthocyanin content in the complex extracted by ethanol-NaCl measured by the pH differential method was 0.121±0.085mg / mg complex, while the anthocyanin content in the complex extracted by ethanol-ice water was 0.061±0.479mg / g complex. This shows that the anthocyanin can be better retained in the complex extracted by the ethanol-NaCl method. Table 2 Anthocyanin extraction content in black corn

[0123] 3) Molecular weight detection

[0124] SDS-PAGE electrophoresis is used for qualitative identification, purity and impurity control, and quantitative determination of proteins. The difference in molecular weight between the complex extracted by ethanol-NaCl and that extracted by ethanol-ice water can be detected by SDS-PAGE electrophoresis. Fig.11 The figure shows the SDS-PAGE pattern of the anthocyanin-protein complex of black corn. The molecular weights of the anthocyanin-protein complex extracted by ethanol-NaCl and ethanol-ice water were determined to be 100 kDa, 40 kDa, 25 kDa and 20 kDa respectively by SDS-PAGE electrophoresis.

[0125] 4) UV spectrum of black corn anthocyanin-protein complex

[0126] Ultraviolet-visible spectroscopy (UV-Vis) is a method that uses the continuous spectrum of electromagnetic waves in the ultraviolet-visible region as a light source to illuminate the sample and study the relative intensity of light absorption by the molecules of the substance. The mass concentration of the black corn anthocyanin-protein complex sample was adjusted to 0.1 mg / mL using 80% ethanol solution, and the spectrum was scanned at 190-1000 nm with a wavelength interval of 1 nm. Fig.12 As shown in the figure, the black corn protein can be detected by UV spectroscopy to have two main characteristic absorption peaks, a peptide bond characteristic absorption peak at about 200nm, and an absorption peak of tryptophan residues and tyrosine residues in the protein at a wavelength of 280nm. In addition to these two absorption peaks, the black corn anthocyanin-protein complex also has an absorption peak of anthocyanin acylation structure at 300-330nm. The ultraviolet absorption of the black corn anthocyanin-protein complex is weakened and slightly blue-shifted, indicating that anthocyanins interact with proteins.

[0127] 4) Infrared spectrum of black corn anthocyanin-protein complex

[0128] Infrared spectroscopy is the ability of molecules to selectively absorb infrared rays of certain wavelengths, which causes transitions in the vibrational and rotational energy levels of the molecules. By detecting the absorption of infrared rays, the infrared absorption spectrum of the substance can be obtained, which is also called molecular vibration spectrum or vibration-rotation spectrum. Fig.13 The infrared spectrum of the black corn anthocyanin-protein complex is shown below. The amide A is at 3400-3440 cm -1 There is an absorption peak at 3431 cm, which is related to the NH stretching vibration. The amide A of black corn protein is at 3431 cm -1 The amide A of the black corn anthocyanin-protein complex appears at 3417 cm -1 Black corn protein appears near 1647cm -1 The absorption at 1619 cm-1 is the C=O stretching vibration of amide I, while the absorption peak intensity of amide I in the black corn anthocyanin-protein complex is significantly weakened and red-shifted to 1619 cm-1. -1 Black corn protein and black corn anthocyanin-protein complex at 1513 cm -1 、1562cm -1 The absorption at 1331 cm is the characteristic absorption peak of the amide II band, reflecting the CN stretching vibration and NH bending vibration of the protein. -1 、1338cm -1 The weak absorption peak at 1478 cm is caused by NH deformation vibration. -1 、1464cm -1 、1415cm -1The CH bending vibration occurs at 3417 cm -1 The peak at 2975 cm-1 may be the hydroxyl stretching vibration absorption peak, and the composite -1 The nearby absorption peak may be the CH stretching vibration absorption peak of the methoxy group, 2890 cm -1 The peak at the center may be the methylene peak. There are characteristic groups such as hydroxyl, methoxy, and oxygen-containing heterocycles in the complex, which are also characteristic groups of anthocyanins. The peak shape of the amide band in the black corn anthocyanin-protein complex has changed significantly, which may be due to the binding of anthocyanins to proteins, resulting in changes in the protein structure.

[0129] The above description of the disclosed embodiments enables one skilled in the art to implement or use the present invention. Various modifications to these embodiments will be apparent to one skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to the embodiments shown herein, but rather to the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for improving the stability of black corn anthocyanin-protein complex, characterized in that: adding a stabilizer to the black corn anthocyanin-protein complex solution, wherein the stabilizer is glucose or maltodextrin; When the stabilizer is glucose, the concentration of glucose in the solution is 1-5%; When the stabilizer is maltodextrin, the concentration of maltodextrin in the solution is 1-3%.

2. A method for improving the stability of black corn anthocyanin-protein complex according to claim 1, characterized in that: The concentration of the black corn anthocyanin-protein complex solution is 2-8 mg / mL.

3. The method for improving the stability of black corn anthocyanin-protein complex according to claim 1, characterized in that: Glucose is anhydrous glucose.

4. The method for improving the stability of black corn anthocyanin-protein complex according to claim 1, characterized in that: The glucose equivalent of maltodextrin is 10-20.

5. The method for improving the stability of black corn anthocyanin-protein complex according to claim 1, characterized in that: When the stabilizer is maltodextrin, the concentration of maltodextrin in the solution is 2%.

6. The method for improving the stability of black corn anthocyanin-protein complex according to claim 1, characterized in that: When the stabilizer is maltodextrin, the concentration of maltodextrin in the solution is 1%.

7. The method for improving the stability of black corn anthocyanin-protein complex according to claim 1, characterized in that: When the stabilizer is maltodextrin, the concentration of maltodextrin in the solution is 3%.

8. The method for improving the stability of black corn anthocyanin-protein complex according to claim 1, characterized in that: When the stabilizer is glucose, the concentration of glucose in the solution is 5%.

9. The method for improving the stability of black corn anthocyanin-protein complex according to claim 1, characterized in that: When the stabilizer is glucose, the concentration of glucose in the solution is 1%.

10. The method for improving the stability of black corn anthocyanin-protein complex according to claim 1, characterized in that: The preparation method of the black corn anthocyanin-protein complex comprises the following steps: defatting black corn flour with petroleum ether, adding ethanol aqueous solution to the defatted black corn flour, stirring, filtering, taking the supernatant and adding NaCl solution, centrifuging, dialyzing the precipitate, and freeze-drying to obtain the black corn anthocyanin-protein complex.

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