Zein and isomaltooligosacharide compound as well as preparation method and application thereof

Abstract

The invention provides a zein and isomaltooligosacharide compound as well as a preparation method and application thereof, and relates to the technical field of biochemistry. The invention provides the zein and isomaltooligosacharide compound, the compound is prepared through Maillard reaction, the combination degree of the compound is high, the particle size distribution range is relatively concentrated, and the particle size is uniform. The preparation method of the compound disclosed by the invention comprises the following steps: dissolving a zein-ethanol solution in an isomaltooligosaccharide solution, then adjusting the pH value to 11.5-12.5, and then heating for 90-150 minutes at the temperature of 70-85 DEG C. The invention also discloses a preparation method of the compound. The compound prepared by the method can effectively improve the functional properties of zein and improve the oxidation resistance and stability of zein, is used for improving the oxidation resistance and stability of active ingredients and promoting the active ingredients to stably exert the functions of the active ingredients, and has a wide market prospect.

Description

Technical Field

[0001] The invention belongs to the technical field of biochemistry, and in particular relates to a complex of zein and isomaltooligosaccharide, and a preparation method and application thereof. Background Art

[0002] Zein is the main storage protein in corn. It is almost insoluble in water and is also insoluble in anhydrous ethanol. Although it is soluble in 60%-90% ethanol aqueous solution and can form nanoparticles through self-assembly, the formed nanoparticles have poor stability and weak antioxidant capacity.

[0003] In the field of covalent binding of proteins and polysaccharides, the existing chemical cross-linking and enzymatic cross-linking methods have many defects. The chemical cross-linking method uses toxic reagents such as glutaraldehyde, which poses a safety hazard in practical applications; the enzymatic cross-linking method requires the use of enzymes that catalyze specific substrate types and react under their optimal catalytic conditions, which not only places harsh demands on the reaction conditions, but also generally requires a large amount of energy consumption throughout the process. There is an urgent need to develop a safe and simple method for zein to form a stable bond and improve its antioxidant capacity, which is of great practical significance for the widespread application of zein in the fields of food, medicine, and chemical industry. Summary of the Invention

[0004] In response to the above technical problems, the first object of the present invention is to provide a complex of zein and isomaltooligosaccharide. The complex has a high degree of binding, a relatively concentrated and uniform particle size distribution, and significantly improves the stability of zein and its antioxidant capacity.

[0005] The second object of the present invention is to provide a method for preparing the complex. The present invention uses a safe and simple construction method to form a complex with stable binding of zein and improve its antioxidant capacity.

[0006] The third object of the present invention is to provide a use of the complex in improving the antioxidant properties and / or stability of active ingredients.

[0007] The fourth object of the present invention is to provide a use of the complex in preparing a product for improving the antioxidant properties and / or stability of active ingredients.

[0008] The fifth object of the present invention is to provide a method for preparing a composite carrier.

[0009] The technical solutions of the present invention are as follows:

[0010] The invention provides a complex of zein and isomaltooligosaccharide, wherein the complex is prepared through Maillard reaction.

[0011] The present invention also provides a method for preparing the complex, which comprises the following steps: dissolving zein in a 60% to 70% ethanol solution, weighing isomaltooligosaccharide in a protein-sugar mass ratio of 1:4 to 4:1 and dissolving it in water, dissolving the zein-ethanol solution in the isomaltooligosaccharide solution, adjusting the pH value to 11.5 to 12.5, and heating at a temperature of 70°C to 85°C for 90 minutes to 150 minutes.

[0012] As an embodiment, the mass fraction of the zein-ethanol solution is 3% to 6%.

[0013] As an embodiment, the mass-to-volume ratio of the isomaltooligosaccharide solution is 10 g / L to 20 g / L.

[0014] As an embodiment, the process further includes distilling off ethanol before adjusting the pH value.

[0015] As an embodiment, the pH value is adjusted and then sealed and heated.

[0016] The present invention also provides a use of the complex in improving the antioxidant property and / or stability of active ingredients.

[0017] The present invention also provides a use of the complex in preparing a product for improving the antioxidant property and / or stability of active ingredients.

[0018] The present invention also provides a method for preparing the composite loaded with active ingredients. After the Maillard reaction is completed, the mixture is cooled, the active ingredient is added at a composite-active ingredient mass ratio of 1:4 to 4:1, stirred for 20 to 40 minutes, centrifuged to collect the supernatant, and freeze-dried.

[0019] As an embodiment, the active ingredients include curcumin, lutein, and resveratrol.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] 1. The composite prepared by the present invention has a high degree of binding, a relatively concentrated particle size distribution range, and a uniform particle size.

[0022] 2. The complex prepared by the present invention has antioxidant properties and stability, can improve the antioxidant properties and stability of active ingredients such as phenols, and promote the active ingredients such as phenols to stably exert their functional effects, and has broad market prospects.

[0023] 3. The complex prepared by the Maillard reaction in the present invention can effectively improve the functional properties of zein, increasing the soluble protein content from 47.04 mg / mL to 85.85 mg / mL. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 The effect of different initial pH on the Maillard reaction of MRPs;

[0025] Figure 2 The apparent color changes of the Maillard reaction of MRPs with different initial pH values ​​(pH increases from left to right);

[0026] Figure 3 The effect of different egg-sugar ratios on the Maillard reaction of MRPs;

[0027] Figure 4 The apparent color changes of the Maillard reaction of MRPs with different egg-sugar ratios;

[0028] Figure 5 The effect of different heating times on the Maillard reaction of MRPs;

[0029] Figure 6 The effect of different heating temperatures on the Maillard reaction of MRPs;

[0030] Figure 7 is the effect of Maillard reaction on pH value of reaction system;

[0031] Figure 8 The standard curve for measuring soluble protein by UV absorption method;

[0032] Figure 9 The effect of Maillard reaction on soluble protein content;

[0033] Figure 10 The fluorescence spectra of Zein and MRPs complexes with different mass ratios are shown in Figure 2.

[0034] Figure 11 is the particle size distribution diagram of Zein and MRPs complex;

[0035] Figure 12 The hydroxyl radical scavenging ability under different Zein:IMO protein-sugar ratios;

[0036] Figure 13 ABTS for different Zein:IMO egg sugar ratios + Free radical scavenging ability;

[0037] Figure 14 is the DPPH scavenging ability under different Zein:IMO;

[0038] Figure 15 is the total reducing force under different Zein:IMO. DETAILED DESCRIPTION

[0039] The invention provides a complex of zein and isomaltooligosaccharide, wherein the complex is prepared through Maillard reaction.

[0040] The present invention also provides a method for preparing the complex, comprising the following steps: dissolving zein in a 60% to 70% ethanol solution, weighing isomaltooligosaccharide at a protein-sugar mass ratio of 1:4 to 4:1 and dissolving it in water, dissolving the zein-ethanol solution in the isomaltooligosaccharide solution, adjusting the pH to 11.5 to 12.5, and heating the mixture at 70°C to 85°C for 90 to 150 minutes.

[0041] In the present invention, zein is first dissolved in a 60% to 70% ethanol solution, isomaltooligosaccharide is dissolved in water, and then the zein-ethanol solution and isomaltooligosaccharide solution are mixed at a mass ratio of 1:4 to 4:1, preferably 1:1, 2:1, 3:1, 4:1, 1:2, 1:3, or 1:4. When the mass of isomaltooligosaccharide is too large, its own steric hindrance effect is large, thereby preventing normal contact between the reaction substrates, reducing the degree of Maillard reaction in the entire system and reducing product concentration. Under the egg-sugar mass ratio of the present invention, sufficient reaction can be achieved and loss is avoided. In the present invention, the mass fraction of the zein-ethanol solution is 3%-6%, preferably 3%, 4%, 5%, or 6%; the mass-to-volume ratio of the isomaltooligosaccharide solution is 10g / L-20g / L, preferably 10g / L, 11g / L, 12g / L, 13g / L, 14g / L, 15g / L, 16g / L, 17g / L, 18g / L, 19g / L, or 20g / L. The zein-ethanol solution is dissolved in the isomaltooligosaccharide aqueous solution using an antisolvent method. After the solutions are uniformly mixed, the ethanol in the mixed solution is evaporated to reduce the effect of the ethanol on the absorbance of subsequent samples when measuring the absorbance. In the present invention, ethanol is evaporated by using a rotary evaporator; the heating temperature for evaporation is the boiling point of ethanol, which is 45-50° C. in the present invention. The present invention can evaporate ethanol at this evaporation heating temperature without affecting the solubility of zein, thereby preventing the solubility of zein in the mixed solution from being affected by excessively high temperature.

[0042] In the present invention, the pH of the steamed mixed solution is adjusted to 11.5-12.5, preferably 11.5, 12.0, or 12.5. In one embodiment, the pH adjuster is an HCl solution or a NaOH solution, and the concentration of the NaOH solution is 8-10 mol / L. The concentration of the HCl solution is not particularly limited in the present invention. By adjusting the initial pH of the reaction system, the present invention facilitates protein stability and promotes the Maillard reaction. After sealing, the mixed solution is heated at a temperature of 70°C to 85°C for 90-150 minutes. In the present invention, the temperature is preferably 70°C, 75°C, 80°C, or 85°C, and the heating time is preferably 90 minutes, 120 minutes, or 150 minutes. The present invention does not specifically limit the heating device; it can be a constant temperature water bath, etc. The degree of carbonylamine condensation in the Maillard reaction is affected by the heating temperature and heating time. The present invention can enhance the degree of the Maillard reaction.

[0043] Based on the high degree of binding of the complex of the present invention and its antioxidant and stability, the present invention also provides a complex for improving the antioxidant and / or stability of active ingredients, and the present invention also provides a complex for preparing a product with improved antioxidant and / or stability of active ingredients.

[0044] The present invention also provides a method for preparing the active ingredient-loaded composite. After the Maillard reaction, the composite is cooled, and the active ingredient is added at a composite-to-active ingredient mass ratio of 1:4 to 4:1. The mixture is stirred for 20 to 40 minutes, and the supernatant is collected by centrifugation and freeze-dried. In the present invention, the stirring speed is 400 to 600 rpm, preferably 400, 500, or 600 rpm, and the stirring time is preferably 20, 30, or 40 minutes. In the present invention, the composite-to-active ingredient mass ratio is preferably 1:1, 2:1, 3:1, 4:1, 1:2, 1:3, or 1:4. At the mass ratios described herein, the binding capacity of the composite and the active ingredient is enhanced, thereby protecting the active ingredient in terms of antioxidant properties or stability. In the present invention, the centrifugal speed is 1000 to 3000 rpm, preferably 1000, 2000, or 3000 rpm. The centrifugation time is 5 to 30 minutes, preferably 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes or 30 minutes. The complex generated after the Maillard reaction is not stable in the system, and the active ingredient will also partially precipitate due to its insolubility in water. The present invention removes the precipitated material through centrifugal separation operation, retaining the complex-active ingredient composite carrier in the liquid system. In the present invention, the active ingredients include curcumin, lutein, and resveratrol. The present invention also includes dispersing the supernatant and then freeze-drying it. In the present invention, the dispersion is to add water to the supernatant in a volume ratio of 1: (2 to 5), preferably 1:2, 1:3, 1:4 or 1:5. In the present invention, the freeze-drying includes spray freeze-drying and vacuum freeze-drying. The freeze-drying temperature is -15°C to -30°C, preferably -15°C, -20°C, -25°C, or -30°C; the freeze-drying time is 24 to 72 hours, preferably 24 hours, 36 hours, 48 ​​hours, 52 hours, 60 hours, 65 hours, or 72 hours. The freeze-drying method of the present invention maintains the original structure and morphology of the complex of zein and isomaltooligosaccharide, avoiding the destruction of the complex structure due to drying, which may affect the activity of the active ingredient.

[0045] In order to make the purpose, technical solutions and advantages of the present invention more clearly understood, the present invention is described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0046] Unless otherwise specified, the materials, reagents, etc. used in the following examples are all commercially available. If specific conditions of use are not specified, they are usually carried out under conventional conditions or under conditions recommended by the company.

[0047] 1. Materials and Reagents

[0048] Reagents level Manufacturer Zein Purity ≥92% Sigma Corporation of America Isomaltooligosaccharide (IMO) Food grade Jiangsu Xinchen Biotechnology Co., Ltd.

[0049] 2. Instruments and Equipment

[0050]

[0051] Example 1

[0052] 0.3 g of zein was dissolved in 10 mL of 70% ethanol solution to prepare a zein-ethanol solution with a mass fraction of 3% (w / v). 0.6 g of oligomaltodextrose was weighed and dissolved in 3 times the volume (30 mL) of distilled water according to a protein-sugar mass ratio of 1:2. After the two were fully dissolved, the zein-ethanol solution was dissolved in the oligomaltodextrose aqueous solution using an antisolvent method, and the ethanol in the system was evaporated from the mixed solution using a rotary evaporator. The heating temperature was 45°C, and the evaporated sample solution was transferred to a flask again. The pH value of the reaction system was adjusted to 11.5 with 9 mol / L sodium hydroxide solution and 1 mol / L hydrochloric acid solution. The sample was sealed and placed in a constant temperature water bath at 75°C for 120 min and then taken out to obtain a complex of zein and oligomaltodextrose (MRPs).

[0053] Example 2

[0054] 0.3 g of zein was dissolved in 10 mL of 60% ethanol solution to prepare a 3% (w / v) zein-ethanol solution. 0.3 g of isomaltooligosaccharide was weighed and dissolved in 3 volumes (30 mL) of distilled water at a protein-sugar ratio of 1:1. After the two were fully dissolved, the zein-ethanol solution was dissolved in the isomaltooligosaccharide aqueous solution using an antisolvent method. The ethanol in the system was evaporated from the mixed solution using a rotary evaporator at a heating temperature of 50°C. The evaporated sample solution was transferred to a flask again, and the pH value of the reaction system was adjusted to 12 with a 10 mol / L sodium hydroxide solution and a 1 mol / L hydrochloric acid solution. The sample was sealed and placed in a constant temperature water bath at 80°C for 90 minutes before being removed to obtain a complex of zein and isomaltooligosaccharide (MRPs).

[0055] Example 3

[0056] 0.6 g of zein was dissolved in 10 mL of 65% ethanol to prepare a 6% (w / v) zein-ethanol solution. 0.3 g of isomaltooligosaccharide was weighed and dissolved in 3 volumes (30 mL) of distilled water at a protein-sugar ratio of 2:1. After both were fully dissolved, the zein-ethanol solution was dissolved in the isomaltooligosaccharide aqueous solution using an antisolvent method. The ethanol in the mixed solution was evaporated using a rotary evaporator at a heating temperature of 50°C. The evaporated sample solution was transferred to a flask again, and the pH of the reaction system was adjusted to 11.5 with a 9 mol / L sodium hydroxide solution and a 1 mol / L hydrochloric acid solution. The sample was sealed and placed in a constant temperature water bath at 85°C for 120 minutes before removal to obtain a complex of zein and isomaltooligosaccharide (MRPs).

[0057] Example 4

[0058] 0.3 g of zein was dissolved in 10 mL of 70% ethanol solution to prepare a 3% (w / v) zein-ethanol solution. 0.6 g of isomaltooligosaccharide was weighed and dissolved in 3 times the volume (30 mL) of distilled water according to a protein-sugar mass ratio of 1:2. After the two were fully dissolved, the zein-ethanol solution was dissolved in the isomaltooligosaccharide aqueous solution using an antisolvent method, and the mixed solution was evaporated using a rotary evaporator to remove the ethanol in the system. The heating temperature was 45°C, and the steamed sample liquid was transferred to a flask again. The pH value of the reaction system was adjusted to 12.0 with a 10 mol / L sodium hydroxide solution and a 1 mol / L hydrochloric acid solution. The sample was placed directly in an 80°C water bath without being sealed and heated for 120 min. After cooling to room temperature, 0.3 g of curcumin was added. The sample was stirred at a speed of 500 r / min with a magnetic stirrer for 30 min, and then centrifuged at a speed of 2000 r / min for 10 min, and the supernatant was collected.

[0059] Example 5

[0060] Same as Example 4, except that curcumin was replaced with 0.6 g lutein.

[0061] Example 6

[0062] Same as Example 4, except that curcumin was replaced with 0.3 g of resveratrol.

[0063] Test Example 1

[0064] 1. Single-factor experiment

[0065] During the preparation of MRPs complex, factors such as initial pH value, mass ratio of zein and isomaltooligosaccharide, heating temperature, and heating time were optimized in sequence.

[0066] 1.1 Effect of different initial pH values ​​on the degree of Maillard reaction

[0067] On the basis of the preparation method of the MRPs complex in Example 1, the pH value of the reaction system was adjusted to 11.5, 12.0, 12.5, 13.0, and 13.5 respectively with appropriate concentrations of hydrochloric acid and sodium hydroxide solution, and the samples cooled to room temperature were respectively diluted to volumetric flasks of the same volume, and the absorbance of the intermediate product and the end product was used to judge the degree of the Maillard reaction. Among them, the method for determining the content of the intermediate product is to take 1 mL of the sample solution cooled after the reaction, dilute it to 80 times the original with distilled water, use distilled water as a blank sample, place the diluted sample in a quartz cuvette, and measure the absorbance A of the sample at an ultraviolet wavelength of 290 nm; the method for determining the content of the end product is to take 1 mL of the sample solution cooled after the reaction, dilute it to 40 times the original with distilled water, use distilled water as a blank sample, and measure the absorbance of the sample at a wavelength of 420 nm to characterize the degree of browning. The effect of the initial pH on the intermediate product and browning degree of the MRPs complex is shown in the following figure. Figure 1 As shown in the results, with increasing pH, the concentration of the intermediate product and the absorbance corresponding to the degree of browning also showed a trend of first increasing and then decreasing, and there was no significant change in the decreasing phase with the increase in initial pH (p>0.05). When the pH was 12.0, the concentration of the intermediate product and the absorbance corresponding to the degree of browning of the sample were the highest (p<0.05).

[0068] The apparent color change of the sample is as follows Figure 2 As shown, when the initial pH is below 12.0, the MRPs complex shows no obvious signs of browning. When the initial pH is 12.0, the sample only exhibits mild browning. When the initial pH is above 12.0, significant browning is observed, and the degree of browning deepens with increasing initial pH. However, there is no significant difference in the absorbance values ​​corresponding to the degree of browning (p>0.05). Simultaneously, the stability of the liquid changes from an initial emulsion to a clear state. This phenomenon may be due to the dual effects of protein denaturation under alkaline conditions and the instability of the Maillard reaction intermediates and brown-black end products at higher pH levels, resulting in a decrease in the sample's absorbance. Therefore, a strongly alkaline environment should not be selected, and an initial pH of 12.0 is used.

[0069] 1.2 Effect of different raw material ratios on the degree of Maillard reaction

[0070] On the basis of the preparation method of the MRPs complex in Example 1, a 3% (w / v) mass fraction of zein-ethanol solution was taken, and the amount was kept unchanged at 10 mL. The corresponding oligosaccharides were weighed in accordance with the protein-sugar mass ratio of 1:1, 2:1, 3:1, 4:1, 1:2, 1:3, and 1:4, and dissolved in 3 volumes (30 mL) of distilled water respectively. The reaction mixture was prepared according to the same anti-solvent method. After the ethanol in the sample was evaporated in turn, the pH value was adjusted to 12 with hydrochloric acid and sodium hydroxide solution of appropriate concentrations. After sealing and marking, the mixture was placed in a constant temperature water bath at 75°C and heated for 120 minutes. After being taken out, the mixture was cooled to room temperature and the samples were respectively diluted to the same volume in volumetric flasks. The degree of Maillard reaction was judged by the absorbance of the intermediate product and the end product. The influence of different raw material ratios on the intermediate product and browning degree of the MRPs complex was shown in the following figure. Figure 3 As shown in the data, when the content of zein remained unchanged, as the mass ratio of protein to sugar continued to increase, the concentration of the intermediate product showed a trend of first increasing and then decreasing. When the mass ratio of MRPs was 1:2, the content of the intermediate product was the highest (p<0.05), indicating that when the mass of oligomaltolsaccharide was too large, its own steric hindrance effect was large, thereby preventing the normal contact between the reaction substrates and reducing the degree of Maillard reaction of the entire system. Therefore, the content of the intermediate product of the reaction was low. Later, as the mass of oligomaltolsaccharide decreased, the degree of contact between the substrates involved in the Maillard reaction also continued to decrease. Therefore, from the overall trend, the concentration of the generated intermediate products continued to decrease, but there was no significant difference between them (p>0.05). Similarly, the overall browning degree of the sample showed a trend of first increasing and then decreasing with the increase of the mass ratio of protein to sugar. The highest degree was achieved when the protein-sugar ratio was 1:2, followed by the mass ratio of 1:1 (p>0.05). When the amount of oligomaltodextrose added was too large or too small, the browning degree was lower than that of the sample with a ratio of 1:2 (p<0.05).

[0071] The apparent color changes of samples under different mass ratios are as follows Figure 4 As shown, overall, when the mass of added IMO is similar to or lower than that of Zein, the color change of the samples is relatively close. However, when the mass of added IMO is larger, although the samples with a larger proportion of Zein have similar absorbance related to the degree of browning, their apparent color is darker than the latter. It is speculated that in addition to the Maillard reaction between Zein and IMO, the remaining excess oligosaccharides undergo caramelization under high temperature and alkaline conditions, resulting in a darker color. In order to reduce the loss of oligosaccharides and allow for more complete binding between the substrates, a mass ratio of Zein to IMO of 1:2 was selected for subsequent experiments.

[0072] 1.3 Effect of different heating times on the degree of Maillard reaction of MRPs complexes

[0073] Based on the preparation method of the MRPs complex in Example 1, the pH value was adjusted to 12 with appropriate concentrations of hydrochloric acid and sodium hydroxide solution. After sealing and labeling, the solution was placed in a constant temperature water bath at 75°C and heated. The same amount of solution was taken out at 30 minutes, 60 minutes, 90 minutes, 120 minutes, and 150 minutes, and diluted to the same multiple. The absorbance of the intermediate product and the end product was used to determine the degree of Maillard reaction. The effect of different heating times on the intermediate product and browning degree of the MRPs complex is shown in the figure below. Figure 5 As shown in the figure, with the extension of heating time, the concentration of the intermediate product shows an overall trend of first increasing and then decreasing, and its concentration reaches the maximum when the reaction time is 120 minutes, indicating that the degree of carbonyl-amine condensation is the highest at this time. After that, with the passage of time, the intermediate product is further transformed into the final product, melanoidin and other substances. When the Maillard reaction is complete, the intermediate product is almost completely converted, so the absorbance of the corresponding mixed system will not change significantly (p>0.05).

[0074] The apparent browning of the sample increased with increasing heating time, but the degree of change was not significant (p>0.05). After a reaction time of more than 1 hour, there was no significant change in absorbance, indicating that the Maillard reaction between the substrates was almost complete. Therefore, based on the concentration of the intermediate product, a reaction time of 120 minutes was selected for subsequent experiments.

[0075] 1.4 Effect of different heating temperatures on the degree of Maillard reaction

[0076] Based on the preparation method of the MRPs complex in Example 1, the pH value was adjusted to 12 with appropriate concentrations of hydrochloric acid and sodium hydroxide solution. After sealing and marking, the constant temperature water bath was set to 70°C, 75°C, 80°C, 85°C, and 90°C. After heating for 120 minutes, the samples were taken out and cooled to room temperature. The samples were then diluted to the same volume in volumetric flasks. The absorbance of the intermediate product and the end product was used to determine the degree of Maillard reaction. The effect of different heating temperatures on the intermediate product and browning degree of the MRPs complex is shown in the figure below. Figure 6 As shown in the images, the concentration of Maillard reaction intermediates and the degree of browning in the mixed system first increase and then decrease with increasing temperature, with the absorbance reaching its highest level at 80°C (p < 0.05). The concentration of intermediates does not change significantly below 80°C (p > 0.05), indicating that the Maillard reaction of MRPs is not easily triggered below 80°C. Above 80°C, the intermediates are easily converted into browning end products, resulting in a transient decrease in absorbance and subsequent stabilization (p > 0.05).

[0077] However, the corresponding browning curve did not show the desired increasing trend. Based on the Maillard reaction principle, when the pH is > 7 and the temperature is high, the intermediate stages of the Maillard reaction and the Amadori rearrangement products undergo Strecker cleavage, further generating products such as methylglyoxal, diacetyl, and 1-hydroxy-2-propanone. These products not only volatilize and absorb moisture at high temperatures but also have tertiary carcinogenic properties, resulting in a corresponding decrease in absorbance. Furthermore, due to the change in the reaction mixture's state before and after, excessive precipitation of zein at high temperatures causes the solution to transition from an emulsion to a solution. This dual effect leads to a sudden decrease in sample absorbance. Therefore, 80°C was selected for subsequent experiments.

[0078] 2. Response Surface Analysis

[0079] On the basis of the above single-factor experimental optimization, the response surface test was used to optimize the three linear change factors (heating time, heating temperature, and reaction system pH) that have a significant impact on the absorbance of the Maillard reaction intermediates and end products. The relevant factors and levels are shown in Table 1.

[0080] Table 1 Experimental design factor levels and coding

[0081]

[0082]

[0083] The optimal conditions of reaction time, reaction temperature and initial pH of the reaction system obtained from the above single-factor experiment were selected as the basis for the level interval of each factor. The absorbance of the intermediate product content and the browning index were used as the dual response values ​​to optimize the Maillard reaction process. The experimental results are shown in Tables 2, 3 and 4.

[0084] Table 2 Response surface test optimization results

[0085]

[0086] The above experimental data were subjected to multiple regression fitting using Design-Expert 12 software, and the polynomial regression equations between reaction time, reaction temperature, initial pH, intermediate product, and browning index were obtained:

[0087] Y1=3.77157-0.003763A+0.01145B-0.32025C;

[0088] Y2=2.89490-0.003513A+0.019025B-0.313C.

[0089] Where: Y1 is the intermediate product, Y2 is the browning index, A is the reaction time, B is the reaction temperature, and C is the initial system pH.

[0090] Table 3 Intermediate product regression model analysis results

[0091]

[0092]

[0093] Note: “*” indicates a significant effect on the results (P < 0.05); “**” indicates an extremely significant effect on the results (P < 0.01).

[0094] Table 4 Analysis results of browning index regression model

[0095]

[0096] Note: “*” indicates a significant effect on the results (P < 0.05); “**” indicates an extremely significant effect on the results (P < 0.01).

[0097] Tables 3 and 4 show that the final fitting results were linear when the absorbance of both the intermediate product and the browning index was used as the response value. No significant interaction was observed between reaction time, reaction temperature, and initial reaction pH. The signal-to-noise ratio (S / N) obtained from fitting the two response values ​​was > 4, further confirming the model's reliability. The influence of the three factors on the absorbance of the intermediate product and the browning index followed the order C > A > B, i.e., initial reaction pH > reaction time > reaction temperature. In the intermediate product determination experiment, the primary term C had an extremely significant effect on the results (P < 0.01), A had a significant effect (P < 0.05), and B had no significant effect (P > 0.05). In the browning index determination experiment, the primary term C had a significant effect (P < 0.05), while A and B had no significant effect (P > 0.05).

[0098] Since there is no obvious interaction between the factors, orthogonal experiments are needed to further optimize the preparation process parameters.

[0099] 3. Orthogonal Experiment Optimization

[0100] On the basis of the above single-factor experimental optimization, a three-factor three-level L9 (34) orthogonal experiment was conducted using the same three factors (heating time, heating temperature, and reaction pH). The absorbance of the intermediate product and the end product were used as the basis to determine the optimal preparation conditions. The factor levels are shown in Table 5.

[0101] Table 5 Orthogonal test factor level table

[0102]

[0103] On the basis of the above single factor experiments, the preparation conditions of zein-isomaltooligosaccharide complex were optimized by orthogonal test, and the results are shown in Tables 6, 7, 8 and 9.

[0104] Table 6 Intermediate product orthogonal test results

[0105]

[0106]

[0107] Table 7 Analysis of variance of intermediate products

[0108] factor sum of squares degrees of freedom mean square F P Reaction time / min 0.043 2 0.022 0.871 0.534 Reaction temperature / ℃ 0.015 2 0.007 0.299 0.770 Reaction pH 0.046 2 0.023 0.934 0.517 error 0.049 2 0.025

[0109] From Table 6 and Table 7, we can see that R A >R C >R B , that is, reaction time > reaction initial pH > reaction temperature, among which the optimal complex preparation conditions are A2B1C1, that is, reaction time 120min, reaction temperature 75℃, and reaction initial pH 11.5. The absorbance of the intermediate product obtained in the experiment reached a maximum value of 0.644A.

[0110] Table 8 Results of orthogonal test of browning index

[0111]

[0112] Table 9 Analysis of variance of browning index

[0113]

[0114]

[0115] From Table 8 and Table 9, we can see that R C >R A >R B , that is, initial reaction pH > reaction time > reaction temperature. Theoretically, a greater degree of browning results in a higher end product content and a higher corresponding sample absorbance. However, in practice, when browning is severe, the corresponding zein aggregates significantly, causing the sample solution to transition from an emulsion to a solution. This change in state results in a sudden drop in sample absorbance. Therefore, considering these considerations, the group with the maximum absorbance in the browning index experiment was ultimately selected as the optimal preparation condition. Analysis of the orthogonal test results revealed that the optimal composite carrier preparation conditions were also A2B1C1: a reaction time of 120 minutes, a reaction temperature of 75°C, and an initial reaction pH of 11.5, resulting in a maximum absorbance of 0.490A for the end product, the browning index.

[0116] Based on the results of the intermediate product and browning index, the optimal preparation conditions for the MRPs complex were reaction time 120 min, reaction temperature 75°C, and initial reaction pH 11.5.

[0117] Experimental Example 2 Changes in pH of the reaction system

[0118] According to the optimal preparation conditions of the single-factor experiment in Experimental Example 1, two groups of samples, a zein solution and a mixed solution of zein and oligomaltodextrose, were prepared. The pH value of the reaction solution system with the optimal heating temperature, heating time, and raw material ratio was measured using a pH meter. The results were repeated three times and the average value was obtained. The corresponding changes in the pH value of the reaction solution are shown in Figure 2. Figure 7 As shown in Figure 2, when the Maillard reaction occurs between proteins and polysaccharides, free -NH2 is consumed and small molecular organic acids are produced, which usually leads to a decrease in the pH value of the reaction system. Figure 7 The initial pH of the MRPs mixed solution was 12.0, but after a period of reaction under optimal conditions, it dropped sharply to 8.28. In contrast, the pH of the control zein sample, which also had a pH of 12.0, only dropped to 11.30 after reaction under optimal reaction conditions without the addition of any reducing sugars. This significant change further confirms that the Maillard reaction between zein and isomaltooligosaccharides occurred, and the reaction was quite intense.

[0119] Experimental Example 3 Changes in the solubility of zein before and after reaction

[0120] Preparation of bovine serum albumin (BSA) solutions of different concentrations: first weigh 0.1 g of bovine serum albumin and dissolve it in 100 mL of distilled water to prepare a BSA solution with a concentration of 1 mg / mL. Select different volumes of the above solution in turn and dilute them to the same volume with distilled water to obtain BSA solutions with concentrations of 0, 0.2, 0.4, 0.6, 0.8, and 1.0 mg / mL. Use distilled water as a blank group for zero adjustment and use a quartz cuvette to measure the absorbance of the corresponding samples at an ultraviolet wavelength of 280 nm. Use the BSA concentration and the corresponding absorbance to plot the graph to obtain the UV-visible spectrophotometer method for determination. A standard curve for soluble protein content was prepared; under the optimal reaction conditions obtained from the single-factor experiment in Experimental Example 1, two groups of zein samples and three corresponding parallel experimental samples were prepared, wherein one group of parallel samples contained only zein, and the other group of parallel samples contained a corresponding mass of isomaltooligosaccharide added according to the optimal protein-sugar ratio obtained from the single-factor experiment. After the reaction was completed, a small amount of sample was taken out and diluted 80 times, and the absorbance of the two groups of samples was measured and compared at an ultraviolet wavelength of 280 nm. The soluble protein content of the two groups of samples was obtained by combining the above standard curve, thereby reflecting the effect of the Maillard reaction on the solubility change of zein.

[0121] By preparing BSA solutions of different concentrations, the absorbance of the solutions at 280 nm was measured, and the standard curve of soluble protein content was drawn. Figure 8 As shown in the experiment, the linear regression equation is Y = 0.602x-0.004, R 2 =0.9994. The soluble protein content in each sample group was calculated using this linear regression equation, thus reflecting the change in the solubility of zein.

[0122] The absorbance of the two groups of experimental samples after dilution of the zein solution and the mixed solution of zein and oligoisomaltolsaccharide was substituted into the above linear regression equation, and the content changes were as follows: Figure 9 As shown. Figure 9 The blank Zein sample had a soluble protein content of 47.04 mg / mL after the reaction. However, after mixing with IMO and undergoing the Maillard reaction, the soluble protein content of the corresponding MRPs increased to 85.85 mg / mL. This suggests that glycation and Maillard reaction can effectively improve the functional properties of zein, namely, enhance its solubility.

[0123] Experimental Example 4: Endogenous Fluorescence Spectroscopy Analysis

[0124] Comparing the differences in intrinsic fluorescence spectra between samples can detect changes in protein structure and further determine whether new chemical bonds have formed between substances, leading to the formation of new substances. The intrinsic fluorescence spectra of Zein nanoparticles and MRPs nanoparticles were analyzed using a microplate reader. While maintaining the Zein concentration constant, different amounts of IMO were added at predetermined protein-sugar ratios of 4:1, 3:1, 2:1, 1:1, 1:2, 1:3, and 1:4. After Maillard reaction under the same conditions, the fluorescence spectra of each sample were measured and compared with a blank Zein sample. Before measurement, each sample was diluted to a protein concentration of 1 mg / mL in 10 mM PBS buffer (pH 7.4) to achieve an appropriate signal intensity. Approximately 50 μL of sample was then added to a 96-well plate. The changes in intrinsic fluorescence intensity of each sample were measured using a microplate reader, and fluorescence intensity curves were plotted for comparison. The corresponding fluorescence measurement parameters were set as follows: excitation wavelength 280 nm, emission wavelength range set between 300 nm and 540 nm, scanning step length of 5 nm, and detection height of 9.5 mm.

[0125] Under the condition of constant zein concentration, the amount of isomaltooligosaccharide (IMO) was changed. The intrinsic fluorescence spectrum of the product after Maillard reaction under different Zein:IMO mass ratios was obtained as shown in the following figure: Figure 10 As shown. Figure 10 It can be seen that when Zein does not undergo Maillard reaction, its fluorescence intensity reaches a peak at 348nm, which is a typical characteristic fluorescence spectrum of tryptophan in protein. When IMO is added, it participates in the Maillard reaction with Zein, and the fluorescence intensity emission peak of the corresponding MRPs complex moves to around 390nm. And as the amount of IMO added continues to increase, the fluorescence intensity shows a regular decrease. This shows that the increase in the Maillard reaction product has continuously enhanced the shielding effect on tryptophan, that is, the fluorescence quenching effect, indicating that a glycosylation reaction has occurred between Zein and IMO, and the degree of occurrence continues to increase with the increase of IMO. At the same time, after Zein undergoes a glycosylation reaction, its corresponding λ max An obvious red shift phenomenon occurred, indicating that the Maillard reaction can change the conformation of Zein and its hydrolyzate to a certain extent, making the structure looser and the solubility enhanced.

[0126] Test Example 5 Particle Size Measurement

[0127] Nanoparticle size and zeta potential analysis were performed on Zein and MRPs composite nanoparticles with varying protein-sugar ratios of 4:1, 3:1, 2:1, 1:1, 1:2, 1:3, and 1:4. The average of three measurements for each sample was used as the final result. Before particle size testing, each sample was diluted with deionized water to a concentration of approximately 1 mg / mL.

[0128] The particle size distribution of Zein and MRPs complexes at different ratios is shown in Figure 2. Figure 11 As shown. Figure 11 Zein particle sizes are distributed within the 50-955 nm and 1110-6440 nm ranges, respectively accounting for 97.95% and 2.05% of the Zein particle size. The Maillard reaction between IMO and Zein at varying addition levels resulted in varying particle size distributions of MRPs due to varying degrees of glycosylation. The particle size distributions of different Zein:IMO combinations are shown below.

[0129] When the Zein:IMO ratio was 4:1, the particle size distribution of the composite was 50–460 nm, with a significantly increased peak height and a leftward shift. When the Zein:IMO ratio was 3:1, the composite was distributed in the 50–220 nm, 250–1480 nm, and 2330–6440 nm ranges, accounting for 50.59%, 44.02%, and 5.39%, respectively. The peak heights were significantly lower than those of the Zein ratio, resulting in a more dispersed distribution range and the emergence of new peaks. When the Zein:IMO ratio was 2:1, the composite was distributed in the 28–51 nm and 58–531 nm ranges, accounting for 3.67% and 96.33%, respectively. The peak heights again increased and shifted to the left. This indicates that when the mass of the IMO is lower than that of the Zein ratio, the particle size distribution is more random. With the exception of the 3:1 Zein:IMO ratio, the particle size distribution range of the composites decreased for all other ratios.

[0130] When the Zein:IMO ratio is 1:1, the particle size distribution range of the composite is 68-396 nm, and the distribution peak reaches the highest among all samples. When the Zein:IMO ratio is 1:2, the particle size distribution range of the composite further expands to 68-615 nm, while the peak height decreases slightly. When the Zein:IMO ratio is 1:3, the particle size distribution range of the composite further expands to 68-825 nm, and the peak height decreases further. When the Zein:IMO ratio is 1:4, the particle size distribution range of the composite again shows a non-normal distribution, and new distribution peaks appear again, including the distribution ranges of 43-122 nm and 142-1280 nm, accounting for 26.57% and 73.43% respectively, and the overall particle size distribution is close to that of Zein. This shows that when the mass of IMO is higher than that of Zein, the particle size distribution pattern and peak change show a regular decreasing trend. With the exception of the Zein:IMO = 1:4 group, which exhibited two distribution peaks, the particle size distribution range of the composites shifted to the left with increasing IMO addition. These experimental results indicate that the introduction of IMO reduced the particle size of MRPs, suggesting that glycosylation can alter the physical and chemical properties of Zein.

[0131] Because Zein possesses a strong hydrophobic effect, the addition of IMO introduces more hydrophilic groups, inhibiting the aggregation of Zein molecules and reducing particle size. Furthermore, the steric hindrance and electrostatic repulsion of the complex formed with IMO are enhanced, further enhancing particle dispersion and reducing the size of the MRPs complex. Comparison of the particle size distributions at different ratios reveals that when the Zein:IMO ratio is 1:1, the Maillard reaction-generated complex has the smallest particle size distribution range and a highly normal distribution, indicating that this ratio of protein to sugar yields the most uniform particle size, with an average particle size concentrated around 122 nm. At Zein:IMO ratios of 1:2 and 1:3, the average particle size dispersion is similar and still follows a normal distribution, with an average particle size concentrated around 142 nm. As the IMO content increases, the particle size distribution range widens further, indicating that at a Zein:IMO ratio of 1:2, the Maillard reaction of Zein can be considered complete.

[0132] Test Example 6 Antioxidant Capacity Determination and Analysis

[0133] 1.1 Hydroxyl radical scavenging ability determination

[0134] First, dilute each sample to a concentration of 1 mg / mL with deionized water, then prepare 9.0 mM FeSO4 solution, 8.8 mM H2O2 solution, and 9.9 mM salicylic acid solution in sequence. Salicylic acid is slightly soluble in water and needs to be prepared with anhydrous ethanol as the solvent. FeSO4 solution needs to be prepared before use because Fe 2+It is easily oxidized to Fe at room temperature. 3+ This will lead to changes in chemical properties, thus affecting the experimental results.

[0135] Accurately pipette 0.5 mL of the above-mentioned FeSO4 solution, salicylic acid-ethanol solution, and sample diluent respectively, add 5.5 mL of deionized water and 0.5 mL of H2O2 solution, and react in the dark at 37°C for 15 minutes. Use deionized water as a reference to adjust to zero, and measure the absorbance of the sample at 510 nm. The result is recorded as sample A; accurately pipette 0.5 mL of the above-mentioned FeSO4 solution, salicylic acid-ethanol solution, and sample diluent respectively, add 6 mL of deionized water to the same volume, and use this as the blank group of the corresponding sample. Also, react in the dark at 37°C for 15 minutes, use deionized water as a reference to adjust to zero, and measure the absorbance of the sample at 510 nm. The result is recorded as A 样0 ; Accurately pipette 0.5 mL of FeSO4 solution and salicylic acid-ethanol solution respectively, add 6.5 mL of deionized water to the same volume, and use this as a blank reference group. After reacting at 37°C in the dark for 15 minutes, adjust the zero value with deionized water as the reference, and measure the absorbance of the sample at 510 nm. The result is recorded as A 参 ; Accurately aspirate 0.5 mL of FeSO4 solution and salicylic acid-ethanol solution respectively, then add 5.5 mL of deionized water and 0.5 mL of H2O2 solution as the control group. After reacting in the dark at 37°C for 15 minutes, use deionized water as a reference to adjust to zero, and measure the absorbance of the sample at 510 nm, recorded as A0; replace the sample diluent with ascorbic acid of the same volume and concentration as a positive control. Each group of samples including ascorbic acid is subjected to three parallel experiments, and the final result is calculated according to the following formula and the average value is taken.

[0136]

[0137] The OH scavenging ability of the Maillard reaction complex prepared at different Zein:IMO mass ratios is as follows Figure 12 As shown in the figure, compared with the blank Zein sample, the ·OH scavenging ability of the glycosylated products decreased to varying degrees and showed strong fluctuations. When the addition amount of IMO was higher than that of Zein, the ·OH scavenging ability of the complex gradually increased.

[0138] 1.2ABTS + Free radical scavenging ability determination

[0139] First, accurately weigh 0.1920 g of ABTS using an analytical balance and dilute to 50 mL with deionized water to prepare a 7 mM ABTS radical solution. Simultaneously, prepare an appropriate amount of 140 mM potassium persulfate solution. Accurately pipette 0.89 mL of this solution into the 50 mL ABTS radical solution and store overnight at 4°C for 12-14 hours before use. Protect from light throughout the process. This is the ABTS stock solution. Dilute the stock solution immediately before use. Accurately pipette 1 mL of the ABTS stock solution and add approximately 100 mL of deionized water to dilute the solution until its absorbance at 734 nm is within the range of 0.70 ± 0.02 A. This provides the ABTS working solution required for the experiment.

[0140] Accurately pipette 0.5mL of sample diluent and 2mL of ABTS working solution in sequence as the sample group, after standing at room temperature in the dark for 6 minutes, use deionized water as a blank to adjust to zero, measure its absorbance at 734nm and record it as sample A; accurately pipette 0.5mL of sample diluent and 2mL of deionized water in sequence as the sample blank group, also after standing at room temperature in the dark for 6 minutes, use deionized water to adjust to zero, and use this as the blank group corresponding to the sample diluent, measure its absorbance at 734nm and record it as sample A 样0 ; Accurately draw 0.5mL of deionized water, add 2mL of ABTS working solution, mix well, and place it at room temperature in the dark for 6 minutes. Adjust the value to zero with deionized water and measure the absorbance A0 at 734nm as the control group. Use the same concentration of ascorbic acid solution to perform three parallel experiments on the positive control. Each group of samples also performs three parallel experiments and takes the average value. The ABTS of each group of samples is calculated by the following formula + Free radical scavenging capacity.

[0141]

[0142] Maillard reaction complex ABTS prepared at different Zein:IMO mass ratios + Changes in free radical scavenging ability Figure 13 As shown in the figure, at the same concentration, the scavenging ability of Zein in the blank group was only 58.90±0.54%, and the ABTS of the complex prepared by adding IMO and undergoing glycosylation reaction was + The scavenging ability changed to varying degrees, and the change was particularly obvious when the Zein:IMO mass ratio was 1:2. + The scavenging capacity can be increased to 93.21±0.71%, and then increases slowly with the increase of IMO mass.

[0143] 1.3 Analysis of DPPH free radical scavenging ability

[0144] Weigh 0.0039 g of DPPH powder using an analytical balance, add 100 mL of anhydrous ethanol to the volume to prepare a 0.1 mM DPPH ethanol solution, and store in the dark at 4°C until use; then measure 30 mL of anhydrous ethanol and add 70 mL of deionized water to prepare a 30% ethanol solution by volume.

[0145] Accurately pipette 1.5mL of sample diluent and DPPH ethanol solution in sequence, shake well, keep it in the dark for 30 minutes, adjust to zero with deionized water, measure the absorbance of the sample group at 517nm and record it as sample A; accurately pipette 1.5mL of sample diluent and 30% ethanol solution in sequence, use this as the blank control group of the corresponding sample diluent, shake well, keep it in the dark for 30 minutes, adjust to zero with deionized water, measure its absorbance at 517nm and record it as A 样0 Accurately pipette 1.5 mL of each DPPH ethanol solution and 30% ethanol solution, shake well, and let stand in the dark for 30 minutes. This is used as a control group. Adjust the absorbance to zero with deionized water and measure the absorbance at 517 nm, recording it as A0. A positive control group, using the same concentration of ascorbic acid, is used. Three parallel experiments are performed on both the sample group and the positive control group. The corresponding DPPH radical scavenging capacity is calculated according to the following formula, and the average value is used as the final experimental result.

[0146]

[0147] The DPPH scavenging ability of Maillard reaction complexes prepared at different Zein:IMO mass ratios is shown in Figure 2. Figure 14 As shown in the figure, the DPPH scavenging ability of the complex decreased to varying degrees compared with the blank group Zein. When the Zein:IMO mass ratio was 1:1 and 1:2, the DPPH scavenging ability increased and was about 79%, and then decreased again.

[0148] 1.4 Determination of total reducing power

[0149] Use an analytical balance to weigh 1.9670 g of sodium dihydrogen phosphate (NaH2PO4·2H2O), 5.4303 g of disodium hydrogen phosphate (Na2HPO4·12H2O), and 1.7 g of sodium chloride (NaCl). Add appropriate amount of deionized water and stir until completely dissolved. Finally, adjust the volume to 400 mL. Use a pH meter to measure the pH of the solution at this time. Sodium hydroxide solution or hydrochloric acid solution can be used to adjust the pH of the solution to 6.6. Finally, a phosphate buffer solution with a concentration of 0.2M and a pH of 6.6 is obtained for use; in addition, 1.0g of potassium ferrocyanide is weighed, deionized water is added to make the volume to 100mL to obtain a potassium ferrocyanide solution with a concentration of 1%, and the solution is stored at 4°C in the dark for use; 10g of trichloroacetic acid (TCA) crystals is weighed, the volume is made up to 100mL with deionized water, and a TCA solution with a concentration of 10% (w / v) is prepared for use; 0.1g of anhydrous ferric chloride solid is weighed, the volume is made up to 100mL with deionized water, and a FeCl3 solution with a concentration of 0.1% is prepared for use.

[0150] Add 1 mL of sample diluent, 2.5 mL of phosphate buffer solution, and 2.5 mL of potassium ferricyanide solution to the test tube in sequence. Add the ascorbic acid positive control group corresponding to the sample volume and concentration in the same manner. After thorough mixing, place the tube in a 50°C constant temperature water bath in the dark and incubate for 20 minutes. Then, add 2.5 mL of TCA solution to terminate the reaction. After thorough mixing, pour the sample solution into a centrifuge tube, level it with deionized water, and centrifuge it at 3000 r / min for 10 minutes. Take 2.5 mL of the supernatant from each sample group and the positive control group, add 2.5 mL of deionized water and 0.5 mL of FeCl3 solution in sequence, mix thoroughly, and let it stand for 10 minutes. Measure the absorbance at 700 nm. Each sample group and positive control group are tested in triplicate. The average absorbance of the three groups reflects the final reducing power of each sample group.

[0151] The total reducing power of the Maillard reaction complexes prepared at different Zein:IMO mass ratios is as follows: Figure 15 As shown, according to Figure 15 The overall trend of change shows that the Maillard reaction has no significant effect on the reducing ability of Zein. It only increases when the Zein:IMO mass ratio is 1:2, and then tends to be flat.

[0152] According to the above antioxidant test results, the glycosylation products obtained by Maillard reaction of MRPs, except for ABTS +Aside from the enhanced free radical scavenging ability, the scavenging abilities of other free radicals showed no significant improvement, and in fact decreased. This suggests that the glycosylation reaction weakens Zein's free radical scavenging ability to some extent. The introduction of IMO into the complex formed by the glycosylation reaction also showed no significant increase in total reducing power. An increase in reducing power was observed only when the Zein:IMO mass ratio exceeded 1:2.

[0153] Based on the results of intrinsic fluorescence spectroscopy, particle size, and antioxidant activity, the final Zein:IMO mass ratio of 1:2 was selected, and the complex prepared via the Maillard reaction effectively protected the active substance. This also indirectly proves that under certain conditions, the Maillard reaction can occur between Zein and IMO, and that they can bind to each other through the Maillard reaction.

[0154] From a molecular binding perspective, polyphenols, as natural antioxidants, not only bind to MRP complexes through non-covalent bonds such as hydrogen bonds and hydrophobic interactions, but also rearrange themselves into quinones through self-oxidation, reacting with nucleophilic groups in the MRP complexes to form covalent bonds (including C-N bonds and C-S bonds) to achieve stable binding, while maintaining their activity. Polyphenols have relatively small molecular weights, typically above 600 Da, making them relatively easy to bind to macromolecules such as proteins. From an encapsulation efficiency perspective, polyphenols bound to MRP complexes differ substantially from proteins in their structural properties, as do active substances that can be bound to and protected by zein, such as curcumin, lutein, and resveratrol. The concentration changes before and after binding can be used to determine the amount of active substance assembled before and after encapsulation. This demonstrates that the MRP complexes of the present invention can bind to curcumin, lutein, and resveratrol to achieve a protective effect.

[0155] The above descriptions are merely embodiments of the present invention and are not intended to limit the patent scope of the present invention. Any equivalent structure or equivalent process transformation made using the contents of the present invention specification, or directly or indirectly applied in other related technical fields, are also included in the patent protection scope of the present invention.

Claims

1. A complex of zein and isomaltooligosaccharide, characterized in that: The complex is prepared through the Maillard reaction.

2. The method for preparing the composite according to claim 1, characterized in that: The method comprises the following steps: dissolving zein in a 60% to 70% ethanol solution, weighing isomaltooligosaccharide in a protein-sugar mass ratio of 1:4 to 4:1 and dissolving the solution in water, dissolving the zein-ethanol solution in the isomaltooligosaccharide solution, adjusting the pH value to 11.5 to 12.5, and heating the solution at 70° C. to 85° C. for 90 to 150 minutes.

3. The preparation method according to claim 2, characterized in that The mass fraction of the zein-ethanol solution is 3% to 6%.

4. The preparation method according to claim 2, characterized in that The mass volume ratio of the isomaltooligosaccharide solution is 10 g / L to 20 g / L.

5. The preparation method according to claim 2, characterized in that The ethanol is also distilled off before the pH adjustment.

6. The preparation method according to claim 2, characterized in that After adjusting the pH, seal and heat.

7. Use of the complex according to claim 1 in improving the antioxidant properties and / or stability of active ingredients.

8. Use of the complex according to claim 1 in the preparation of a product having improved antioxidant properties and / or stability of an active ingredient.

9. The method for preparing a composite loaded with active ingredients according to claim 1, characterized in that: After the Maillard reaction is completed, the mixture is cooled, and the active ingredient is added at a mass ratio of the complex to the active ingredient of 1:4 to 4:

1. The mixture is stirred for 20 to 40 minutes, and the supernatant is collected by centrifugation and freeze-dried.

10. The preparation method according to claim 9, characterized in that The active ingredients include curcumin, lutein and resveratrol.

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