Ultrasonic modified walnut protein-chitosan composite emulsion as well as preparation method and application thereof

Abstract

The invention provides an ultrasonic modified walnut protein-chitosan composite emulsion as well as a preparation method and application thereof. The composite emulsion comprises an ultrasonic modified walnut protein-chitosan composite solution and an oil phase, the preparation method of the ultrasonic modified walnut protein-chitosan composite solution comprises the following steps: carrying out gradient ultrasonic treatment on a walnut protein solution, and freeze-drying to obtain ultrasonic modified walnut protein powder; preparing an ultrasonic modified walnut protein powder solution; and mixing and stirring the ultrasonic modified walnut protein powder solution and a chitosan solution. According to the ultrasonic modified walnut protein-chitosan composite emulsion disclosed by the invention, the walnut protein modified by different gradients of ultrasonic waves is adopted as a raw material and is compounded with the chitosan for use, so that the walnut protein-chitosan composite emulsion not only shows excellent performance in the aspects of appearance and texture, but also remarkably improves the thermal stability of the prepared high-internal-phase emulsion.

Description

Technical Field

[0001] The invention belongs to the technical field of food processing, and in particular relates to an ultrasonically modified walnut protein-chitosan composite emulsion and a preparation method and application thereof. Background Art

[0002] In recent years, with the increasing emphasis on healthy eating and functional foods, the application of plant proteins and their composite materials in food processing and nutritional supplementation has attracted increasing attention. Walnuts are nutritious nuts rich in protein, fatty acids, and antioxidants, and therefore have broad application prospects in the food and health care products industries. Walnut protein (WP) is rich in amino acids and can serve as an important source of protein for the human body. Chitosan (CS), a natural polysaccharide, has good biocompatibility, is non-toxic, and has excellent emulsifying properties, making it an effective food additive.

[0003] Emulsion is an important food form, especially in the fields of dairy products, seasonings, sauces, etc. High internal phase emulsions are increasingly favored by consumers because of their characteristics such as not easy to stratify and excellent taste. The invention patent with publication number CN105310066A discloses a plant protein / soy polysaccharide nanoemulsion for embedding natural fat-soluble pigments and a preparation method. After the natural fat-soluble pigment is dissolved in oil, it is added to a mixed solution of plant acid-soluble protein and soybean polysaccharide, and the mixture is obtained by homogenizing emulsification and heating reaction. The invention patent with publication number CN107897915A discloses a lotus seed protein-pectin composite emulsion loaded with curcumin, its preparation method and application, using lotus seed protein and pectin complex as emulsifier and stabilizer, the emulsion system is an oil-in-water emulsion system, and the embedding rate of curcumin can reach 85-90%. The prior art does not disclose a technical solution for preparing high internal phase emulsions using walnut protein. Furthermore, conventional emulsion preparation methods for walnut protein-based high internal phase emulsions also face problems such as poor emulsion stability, especially poor thermal stability, which limits their further application in the food processing field. Therefore, it is particularly important to develop a method for preparing walnut protein-based high internal phase emulsions and improve the stability and functionality of the emulsions. Summary of the Invention

[0004] In view of this, the present invention aims to propose an ultrasonically modified walnut protein-chitosan composite emulsion, a preparation method and application thereof, and to prepare a stable high internal phase emulsion based on an ultrasonically modified walnut protein-chitosan complex by gradient ultrasonic treatment of walnut protein.

[0005] To achieve the above object, the technical solution of the present invention is achieved as follows:

[0006] In a first aspect, the present invention provides an ultrasonically modified walnut protein-chitosan composite emulsion, wherein the composite emulsion comprises an ultrasonically modified walnut protein-chitosan composite solution and an oil phase. The preparation method of the ultrasonically modified walnut protein-chitosan composite solution comprises the steps of subjecting the walnut protein solution to gradient ultrasonic treatment and then freeze-drying the solution to obtain ultrasonically modified walnut protein powder, preparing the ultrasonically modified walnut protein powder solution, and mixing and stirring the ultrasonically modified walnut protein powder solution with the chitosan solution.

[0007] Furthermore, the oil phase is vegetable oil; preferably, the oil phase is selected from one of soybean oil, corn oil, walnut oil, peanut oil, rapeseed oil, and olive oil; further preferably, the oil phase is walnut oil.

[0008] In a second aspect, the present invention provides a method for preparing the ultrasonically modified walnut protein-chitosan composite emulsion as described in the first aspect, the preparation method comprising the following steps:

[0009] (1) subjecting the walnut protein solution to gradient ultrasonic treatment and then freeze-drying to obtain ultrasonically modified walnut protein powder;

[0010] (2) preparing an ultrasonically modified walnut protein powder solution;

[0011] (3) mixing the ultrasonically modified walnut protein powder solution and the chitosan solution to obtain an ultrasonically modified walnut protein-chitosan composite solution;

[0012] (4) The ultrasonically modified walnut protein-chitosan composite solution is mixed with the oil phase, and sheared and homogenized to obtain an ultrasonically modified walnut protein-chitosan composite emulsion.

[0013] Furthermore, the walnut protein in the walnut protein solution is prepared by an alkali dissolution and acid precipitation method; preferably, the preparation method of the walnut protein comprises the following steps: mixing walnut meal with deionized water, adjusting the pH to 9.0, heating and stirring while maintaining the pH at 9.0, and then centrifuging to obtain the supernatant; adjusting the pH of the supernatant to 4.5, refrigerating and standing, and collecting the precipitate by centrifugation; washing the precipitate, mixing it with water, adjusting the pH to 7.0, and freeze-drying to obtain walnut protein powder.

[0014] Furthermore, the step (1) of subjecting the walnut protein solution to gradient ultrasonic treatment comprises the following steps:

[0015] (11) The pH of the walnut protein solution was adjusted to 9 and ultrasonicated for 5-10 min;

[0016] (12) adjusting the pH of the walnut protein solution to 10 and ultrasonically treating for 5-10 min;

[0017] (13) Adjust the pH of the walnut protein solution to 11 and ultrasonicate for 5–10 min.

[0018] Furthermore, the concentration of the walnut protein solution is 3% to 6% (w / v); preferably 3% to 5%; and more preferably 4%.

[0019] Furthermore, step (1) also includes a step of adjusting the pH of the walnut protein solution after gradient ultrasonic treatment to neutral before freeze-drying.

[0020] Furthermore, the power of the gradient ultrasonic treatment is 300-600 W / L, and the total time of the gradient ultrasonic treatment is 20-30 min; preferably, the power of the gradient ultrasonic treatment is 350-450 W / L, and the total time of the gradient ultrasonic treatment is 22-28 min; further preferably, the power of the gradient ultrasonic treatment is 400 W / L, and the total time of the gradient ultrasonic treatment is 25 min.

[0021] Furthermore, the concentration of the ultrasonically modified walnut protein powder solution is 20 to 60 mg / mL; preferably 30 to 50 mg / mL; and more preferably 40 mg / mL.

[0022] Furthermore, the concentration of the chitosan solution is 10-30 mg / mL; preferably 15-25 mg / mL; and more preferably 20 mg / mL.

[0023] Furthermore, in step (3), the ultrasonically modified walnut protein powder solution and the chitosan solution are mixed at a mass ratio of ultrasonically modified walnut protein powder to chitosan of 8-12:2-5; preferably 9-11:3-5; and further preferably 10:4.

[0024] Furthermore, in step (3), the stirring rate is 200-800 rpm, and the stirring time is 1-3 h; preferably, the stirring rate is 400-600 rpm, and the stirring time is 1.5-2.5 h; further preferably, the stirring rate is 500 rpm, and the stirring time is 2 h.

[0025] Furthermore, the mass ratio of the ultrasonically modified walnut protein-chitosan composite solution to the oil phase in step (4) is 1:2-5; preferably 1:2.5-3.5; and more preferably 1:3.

[0026] Furthermore, in step (4), the speed of shear homogenization is 12000-18000 rpm, and the time of shear homogenization is 1-5 min; preferably, the speed of shear homogenization is 13000-16000 rpm, and the time of shear homogenization is 1.5-3 min; further preferably, the speed of shear homogenization is 15000 rpm, and the time of shear homogenization is 2 min.

[0027] In a third aspect, the present invention provides an application of gradient ultrasonic treatment of a walnut protein solution in preparing an ultrasonically modified walnut protein-chitosan composite emulsion, wherein the gradient ultrasonic treatment of the walnut protein solution comprises the following steps:

[0028] (11) The pH of the walnut protein solution was adjusted to 9 and ultrasonicated for 5-10 min;

[0029] (12) adjusting the pH of the walnut protein solution to 10 and ultrasonically treating for 5-10 min;

[0030] (13) Adjust the pH of the walnut protein solution to 11 and ultrasonicate for 5–10 min;

[0031] Preferably, the application includes improving the thermal stability of ultrasonically modified walnut protein-chitosan composite emulsion.

[0032] In a fourth aspect, the present invention provides the use of the ultrasonically modified walnut protein-chitosan composite emulsion as described in the first aspect or the preparation method as described in the second aspect in the food field.

[0033] Compared with the prior art, the ultrasonically modified walnut protein-chitosan composite emulsion and its preparation method and application described in the present invention have the following advantages:

[0034] (1) The ultrasonically modified walnut protein-chitosan composite emulsion of the present invention uses walnut protein modified by different gradient ultrasonic waves as a raw material and combines it with chitosan, which not only shows excellent performance in appearance and texture, but also significantly improves the thermal stability of the prepared high internal phase emulsion.

[0035] (2) The present invention can improve the dispersibility of walnut protein and reduce the degree of particle aggregation through ultrasonic treatment, thereby significantly improving the stability of the prepared emulsion. Further, by optimizing the gradient to adjust the pH node and controlling the ultrasonic treatment time, a composite system with good emulsification performance and stability is formed as a new emulsifier, and a walnut protein-based high internal phase emulsion is successfully prepared. This not only improves the quality of the emulsion, but also greatly improves the thermal stability of the emulsion, providing a reference for the research and development of walnut protein-based high internal phase emulsions. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] The accompanying drawings, which constitute part of the present invention, are provided to provide a further understanding of the present invention. The exemplary embodiments of the present invention and their descriptions are provided to explain the present invention and do not constitute an undue limitation of the present invention. In the accompanying drawings:

[0037] Figure 1 Schematic diagram of the appearance and micromorphology of UWP-CS based high internal phase emulsion;

[0038] Figure 2Schematic diagram of shear slope and initial viscosity results of UWP-CS based high internal phase emulsion;

[0039] Figure 3 Schematic diagram of the droplet size results of UWP-CS based high internal phase emulsion;

[0040] Figure 4 Schematic diagram of the storage modulus results of UWP-CS based high internal phase emulsion;

[0041] Figure 5 Schematic diagram of the loss modulus results of UWP-CS based high internal phase emulsion;

[0042] Figure 6 Schematic diagram of the appearance change of UWP-CS based high internal phase emulsion during heat treatment;

[0043] Figure 7 Schematic diagram of the shear slope and initial viscosity of the UWP-CS based high internal phase emulsion after heat treatment;

[0044] Figure 8 Schematic diagram of the microscopic morphology of the UWP-CS based high internal phase emulsion after heat treatment;

[0045] Figure 9 Schematic diagram of the droplet size results of UWP-CS based high internal phase emulsion after heat treatment. DETAILED DESCRIPTION

[0046] It should be noted that, in the absence of conflict, the embodiments of the present invention and the features in the embodiments may be combined with each other.

[0047] The present invention will be described in detail below with reference to the accompanying drawings and in conjunction with embodiments.

[0048] Example 1

[0049] The preparation method of the ultrasonically modified walnut protein-chitosan composite emulsion of this embodiment comprises the following steps:

[0050] (1) Preparation of walnut protein

[0051] In the present embodiment, walnut protein is extracted based on the principle of alkali dissolution and acid precipitation, and the specific method is as follows: 30g cold-pressed walnut meal is weighed, passed through a 40-mesh sieve, and deionized water is added to the walnut meal at a solid-liquid ratio of 1:15 (w / v). The walnut meal is evenly dispersed in deionized water by magnetic stirring for 30min, and the pH is adjusted to 9.0 with 0.5M NaOH. Then, the assisted extraction is performed by heating magnetic stirring for 2h at 50°C, and the pH is readjusted every 30min to maintain it at 9.0. The centrifuge speed is set to 8000rpm for 15min, and the supernatant is retained after centrifugation, and the supernatant pH is adjusted to 4.5. The supernatant is allowed to stand for 2h in a 4°C refrigerator and the supernatant is discarded. Set the centrifuge speed to 9000 rpm, centrifuge for 15 minutes, collect the precipitate, and wash the precipitate three times with deionized water. Finally, dissolve the precipitate in deionized water and stir evenly. Adjust its pH to 7.0 with 0.5 M NaOH. Place the protein solution in a petri dish and refrigerate it in a -20 ° C refrigerator overnight, then put it into a freeze dryer and freeze-dry for 36 hours. Finally, walnut protein powder (purity is 86.51 ± 0.52%) is obtained, collected and placed in a 4 ° C refrigerator for use.

[0052] (2) Preparation of ultrasonically modified walnut protein

[0053] The walnut protein prepared in step (1) is ultrasonically treated using an ultrasonic cell disruptor: a walnut protein solution with a concentration of 4% (w / v) is prepared and ultrasonically modified using an ultrasonic cell disruptor. The ultrasonic power is set to 400 W / L, and the protein solution is subjected to gradient ultrasonic treatment, with the first stage: adjusting the pH to 9.0 and ultrasonic treatment for 5 minutes; the second stage: adjusting the pH to 10 and ultrasonic treatment for 10 minutes; and the third stage: adjusting the pH to 11 and ultrasonic treatment for 10 minutes. After the ultrasonic treatment, the pH of the walnut protein solution is adjusted to neutral, and the solution is freeze-dried in a freeze dryer to obtain ultrasonically modified walnut protein powder.

[0054] (3) Preparation of ultrasonically modified walnut protein-chitosan complex

[0055] The ultrasonically modified walnut protein powder prepared in step (2) was prepared into a 40 mg / mL stock solution, and a 20 mg / mL chitosan stock solution was prepared. The protein solution and chitosan solution were compounded at a concentration ratio of 10:4 and stirred at 500 rpm for 2 hours to obtain a composite solution (abbreviated as UWP / CS). The composite solution was then placed in a 4°C refrigerator for later use.

[0056] (4) Preparation of UWP-CS-based high internal phase emulsion

[0057] In this example, the oil phase ratio of the high internal phase emulsion (HIPE) was fixed at 75 wt %. The specific preparation method was as follows: 15 mL of walnut oil and 5 mL of the UWP / CS solution prepared in step (3) were added to a clean centrifuge tube. After thorough mixing, the oil-water mixture was shear-homogenized at 15,000 rpm for 2 minutes using a high-speed homogenizer to form uniform HIPEs, which were recorded as sample ①.

[0058] Example 2

[0059] The difference from Example 1 is that in step (2), the protein solution is subjected to a gradient ultrasonic treatment: the first stage: adjusting the pH to 9.0 and ultrasonic treatment for 10 minutes; the second stage: adjusting the pH to 10 and ultrasonic treatment for 5 minutes; the third stage: adjusting the pH to 11 and ultrasonic treatment for 10 minutes. The other steps are the same as in Example 1, and sample ② is obtained.

[0060] Example 3

[0061] The difference from Example 1 is that in step (2), the protein solution is subjected to gradient ultrasonic treatment: the first stage: adjusting the pH to 9.0 and ultrasonic treatment for 10 minutes; the second stage: adjusting the pH to 10 and ultrasonic treatment for 10 minutes; the third stage: adjusting the pH to 11 and ultrasonic treatment for 5 minutes. The other steps are the same as in Example 1, and sample ③ is obtained.

[0062] Example 4

[0063] The difference from Example 1 is that in step (2), the concentration of the walnut protein solution is 4% (w / v) and the ultrasonic power is 350 W / L; in step (3), the ultrasonically modified walnut protein powder is prepared into a 30 mg / mL stock solution, and a 15 mg / mL chitosan stock solution is prepared at the same time. The protein solution and the chitosan solution are compounded at a concentration ratio of 9:3 and stirred at 400 rpm for 1.5 hours to obtain a composite solution; in step (4), 12.5 mL of walnut oil and 5 mL of the UWP / CS solution obtained in step (3) are added to a clean centrifuge tube, and after thorough mixing, the oil-water mixture is sheared and homogenized at a speed of 13,000 rpm for 1.5 minutes using a high-speed homogenizer to form uniform HIPEs.

[0064] The other steps are the same as those in Example 1.

[0065] Example 5

[0066] The difference from Example 1 is that in step (2), the concentration of the walnut protein solution is 5% (w / v) and the ultrasonic power is 450 W / L; in step (3), the ultrasonically modified walnut protein powder is prepared into a 50 mg / mL stock solution, and a 25 mg / mL chitosan stock solution is prepared at the same time. The protein solution and the chitosan solution are compounded at a concentration ratio of 11:5 and stirred at 600 rpm for 2.5 hours to obtain a composite solution; in step (4), 19.5 mL of walnut oil and 5 mL of the UWP / CS solution obtained in step (3) are added to a clean centrifuge tube, and after thorough mixing, the oil-water mixture is sheared and homogenized at a speed of 16,000 rpm for 3 minutes using a high-speed homogenizer to form uniform HIPEs.

[0067] The other steps are the same as those in Example 1.

[0068] Comparative Example 1

[0069] The difference from Example 1 is that in step (2), the protein solution is subjected to gradient ultrasonic treatment: the first stage: adjusting the pH to 9.0 and ultrasonic treatment for 15 minutes; the second stage: adjusting the pH to 11 and ultrasonic treatment for 10 minutes. The other steps are the same as in Example 1, and sample ④ is obtained.

[0070] Comparative Example 2

[0071] The difference from Example 1 is that in step (2), the protein solution is subjected to gradient ultrasonic treatment: the first stage: adjusting the pH to 9.0 and ultrasonic treatment for 5 minutes; the second stage: adjusting the pH to 11 and ultrasonic treatment for 20 minutes. The other steps are the same as in Example 1, and sample ⑤ is obtained.

[0072] Comparative Example 3

[0073] The difference from Example 1 is that in step (2), the protein solution is ultrasonically treated: the pH is adjusted to 9.0, and ultrasonic treatment is performed for 25 minutes to obtain sample ⑥.

[0074] Comparative Example 4

[0075] The difference from Example 1 is that in step (2), the protein solution is ultrasonically treated: the pH is adjusted to 11.0, and ultrasonic treatment is performed for 25 minutes to obtain sample ⑦.

[0076] Performance testing methods

[0077] Test Example 1 Appearance and Micromorphology

[0078] Use a rubber-tipped dropper to transfer the prepared emulsion into a black-capped bottle and observe its appearance.

[0079] At the same time, after the prepared HIPEs were appropriately diluted, 10 μL of sample was dropped on a glass slide, covered with a cover glass, and the morphology of the emulsion droplets was observed under a microscope. The results were as follows: Figure 1 shown.

[0080] Test Example 2 Droplet Size Analysis

[0081] The droplet size of HIPEs was measured using a Biotek laser particle size analyzer. The refractive indices of the oil phase and water phase were 1.465 and 1.330, respectively. HIPEs were diluted 100-fold with ultrapure water and added to the sample cell. The average diameter of the droplets in the HIPEs was recorded. The results are shown in Figure 2. Figure 3 shown.

[0082] Test Example 3 Rheological Analysis

[0083] The dynamic viscoelastic properties of high internal phase emulsions were measured using a rotational rheometer. The HIPEs sample was placed on a flat plate with a diameter of 25 mm, and the gap between the two parallel plates was adjusted to 1.0 mm. In the amplitude sweep experiment, oscillatory shear experiments were performed at 25°C, in the range of 0.1 to 100 rad / s, with a strain of 1%. The storage modulus (G') and loss modulus (G") related to elasticity and viscosity were measured, respectively. In the shear ramp experiment, the shear stress of each sample was measured at 25°C, in the range of 0.1 to 100 rad / s. -1 Variation of viscosity with shear rate within the range of 0.1s -1 The viscosity value obtained under the above conditions is taken as the initial viscosity of the sample. The results are as follows Figure 2 、 Figure 4 、 Figure 5 shown.

[0084] Test Example 4 Determination of Thermal Stability of HIPEs

[0085] To evaluate the thermal stability of HIPEs, freshly prepared samples were heat-treated in a constant temperature water bath at 90°C for 10 min, 20 min, and 30 min. The appearance and micromorphology, droplet size, and viscosity of the samples at the initial and end points of heat treatment were measured to observe the effect of ultrasonic gradient treatment of walnut protein / chitosan composites on the thermal stability of the prepared high internal phase emulsions. The results are shown in Figure 2. Figure 6-9 As shown in Table 1.

[0086] Performance test results

[0087] like Figure 1 As shown, all samples did not flow when inverted after being initially transferred into the vial, and had good viscoelasticity. Figure 3It can be seen that compared with samples ④⑤ prepared by two-stage gradient ultrasonic treatment and samples ⑥⑦ prepared by one-stage ultrasonic treatment, HIPEs samples ①②③ prepared by three-stage gradient ultrasonic treatment and then compounded with chitosan as the aqueous phase have smaller droplet sizes, proving that three-stage gradient ultrasonic treatment can reduce the droplet size of HIPEs. In Example 1, the walnut protein treated with ultrasonic gradient (parameters are pH 9 treatment for 5 min, pH 10 treatment for 10 min, and pH 11 treatment for 10 min) and compounded with chitosan as the aqueous phase have the smallest droplet size (17.79±1.76 μm).

[0088] By measuring the changes in emulsion viscosity at different shear rates, we can understand its flow characteristics, which is helpful to study the flow behavior of emulsion in practical applications. Figure 2 As shown in the figure, the apparent viscosity of the seven HIPEs increases with the increase of shear rate (0.1-100s -1 ) and decreased, indicating that WP / CS-based HIPEs all have shear-thinning properties and can be classified as pseudoplastic fluids. Compared with samples ④⑤ prepared by two-stage gradient ultrasonic treatment and samples ⑥⑦ prepared by one-stage ultrasonic treatment, HIPEs samples ①②③ prepared by three-stage gradient ultrasonic treatment and then compounded with walnut protein and chitosan as the aqueous phase have higher initial viscosities, proving that three-stage gradient ultrasonic treatment can increase the initial viscosity of HIPEs. In Example 1, the HIPEs prepared by compounding walnut protein treated with ultrasonic gradient (parameters: pH 9 treatment for 5 minutes, pH 10 treatment for 10 minutes, pH 11 treatment for 10 minutes) and chitosan as the aqueous phase have the highest initial viscosity (854.26±12.33 Pa·s). This is because the gradient ultrasonic effect reduces the size of UWP particles, causing interactions between emulsion droplets. At the same time, the increase in droplet number and specific surface area improves the interactions between droplets and further enhances the adhesion between emulsions.

[0089] The ability of an emulsion to resist deformation can be determined by analyzing the difference between the storage modulus (G') and loss modulus (G") of the emulsion. Figure 4 、 5As shown in the figure, in the entire angular frequency range (0.1-100rad / s), the G' values ​​of all HIPEs are significantly greater than the G" value. This indicates that UWP / CS can be effectively adsorbed on the oil-water interface, so that the emulsion forms an elastic gel-like network structure, showing a rheological behavior similar to that of a solid. In addition, the G' value is independent of the angular frequency, indicating that all samples can resist deformation. Compared with samples ④⑤ prepared by two-stage gradient ultrasonic treatment and samples ⑥⑦ prepared by one-stage ultrasonic treatment, HIPEs samples ①②③ prepared by three-stage gradient ultrasonic treatment and then compounded with walnut protein and chitosan as the aqueous phase have higher G' values, which proves that three-stage gradient ultrasonic treatment can improve the G' value of HIPEs. In Example 1, the walnut protein treated with ultrasonic gradient (parameters are pH 9 for 5 min, pH 10 for 10 min, pH 7 for 15 min, pH 8 for 20 min, pH 9 for 30 min, pH 10 for 40 min, pH 10 for 50 min, pH 10 for 60 min, pH 10 for 70 min, pH 10 for 80 min, pH 10 for 90 min, pH 10 for 15 min, pH 10 for 20 min, pH 10 for 30 min, pH 10 for 50 min, pH 10 for 60 min, pH 10 for 70 min, pH 10 for 80 min, pH 10 for 15 min, pH 10 for 15 min, pH 10 for 15 min, pH 10 for 20 min, pH 10 for 30 min, pH 10 for 15 min, pH 10 for 20 min, pH 10 for 30 min, pH 10 for 15 min, pH 10 for 15 min, pH 10 for 15 min, pH 1 HIPEs prepared by combining UWP (treated with CS for 10 min) with chitosan as the aqueous phase exhibited the highest G' values. This is because the HIPEs produced by ultrasonic gradient treatment of UWP and CS have smaller droplet sizes and larger specific surface areas, which facilitates the formation of a more compact structure between droplets, thereby increasing the interaction between proteins at the emulsified interface.

[0090] like Figure 6 As shown in the figure, after the seven groups of HIPEs were placed in a 90°C constant-temperature water bath for 10 minutes, no significant changes in the appearance of the samples were observed, demonstrating the excellent stability of UWP / CS-based HIPEs after short-term high-temperature heat treatment. However, after 20 minutes of heat treatment at 90°C, samples ④ and ⑥ showed increased fluidity in the vial and lost their inversion stability. However, no oil precipitation was observed, and the emulsions retained their milky white appearance. Further increasing the heat treatment time to 30 minutes, the fluidity of sample ⑦ began to increase, and the inversion stability in the vial was lost. However, samples ①, ②, ③, and ⑤ still maintained good stability.

[0091] like Figure 7 As shown in the figure, by analyzing the change of HIPEs viscosity with shear rate after heat treatment for 30 minutes, it can be found that compared with samples ④⑤ prepared by two-stage gradient ultrasonic treatment and samples ⑥⑦ prepared by one-stage ultrasonic treatment, HIPEs samples ①②③ prepared by walnut protein and chitosan composited as the aqueous phase prepared by three-stage gradient ultrasonic treatment still have higher viscosity after heat treatment, which proves that three-stage gradient ultrasonic treatment can increase the viscosity of HIPEs after heat treatment, and sample ① has the highest viscosity (812.31±18.98Pa·s).

[0092] like Figure 8 、 9As shown, further characterization of the emulsion droplet size revealed that the HIPEs (sample ①) prepared by combining walnut protein treated with ultrasonic gradient in Example 1 (parameters were pH 9 treatment for 5 min, pH 10 treatment for 10 min, and pH 11 treatment for 10 min) with chitosan as the aqueous phase still had a small droplet size (29.71±1.53 μm) after 30 min of heat treatment. As shown in Table 1, compared with samples ④⑤ prepared by two-stage gradient ultrasonic treatment and samples ⑥⑦ prepared by one-stage ultrasonic treatment, HIPEs samples ①②③ prepared by combining walnut protein with chitosan as the aqueous phase prepared by three-stage gradient ultrasonic treatment had lower particle size change and change rate after heat treatment, indicating that the three-stage ultrasonic gradient treatment of walnut protein can significantly improve the thermal stability of HIPEs prepared by combining walnut protein with chitosan.

[0093] Table 1 Changes in droplet size of UWP / CS-based high internal phase emulsions before and after heat treatment

[0094]

[0095]

[0096] In summary, the preparation method of the ultrasonically modified walnut protein-chitosan-based high internal phase emulsion in the present invention significantly improves the thermal stability of the prepared high internal phase emulsion by using walnut protein modified by gradient ultrasonication as a raw material and combining it with chitosan. The emulsion thus prepared exhibits excellent performance in appearance and texture.

[0097] This technology has broad application prospects in food, health products, cosmetics, and other fields. It can meet the market demand for high-performance emulsions and provide new ideas and methods for the development of related products. This innovative preparation process provides strong support for improving product quality, extending shelf life, and meeting consumer expectations for healthy products, with significant social and economic value.

[0098] The embodiments described above are only part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, other embodiments obtained by those skilled in the art without creative work shall fall within the scope of protection of the present invention.

Claims

1. An ultrasonically modified walnut protein-chitosan composite emulsion, characterized in that: The composite emulsion includes an ultrasonically modified walnut protein-chitosan composite solution and an oil phase. The preparation method of the ultrasonically modified walnut protein-chitosan composite solution includes the steps of subjecting the walnut protein solution to gradient ultrasonic treatment and then freeze-drying the solution to obtain ultrasonically modified walnut protein powder, preparing the ultrasonically modified walnut protein powder solution, and mixing and stirring the ultrasonically modified walnut protein powder solution with the chitosan solution.

2. The method for preparing the ultrasonically modified walnut protein-chitosan composite emulsion according to claim 1, wherein: The preparation method comprises the following steps: (1) subjecting the walnut protein solution to gradient ultrasonic treatment and then freeze-drying to obtain ultrasonically modified walnut protein powder; (2) preparing an ultrasonically modified walnut protein powder solution; (3) mixing the ultrasonically modified walnut protein powder solution and the chitosan solution to obtain an ultrasonically modified walnut protein-chitosan composite solution; (4) The ultrasonically modified walnut protein-chitosan composite solution is mixed with the oil phase, and sheared and homogenized to obtain an ultrasonically modified walnut protein-chitosan composite emulsion.

3. The preparation method according to claim 2, characterized in that The step (1) of subjecting the walnut protein solution to gradient ultrasonic treatment comprises the following steps: (11) The pH of the walnut protein solution was adjusted to 9 and ultrasonicated for 5-10 min; (12) adjusting the pH of the walnut protein solution to 10 and ultrasonically treating for 5-10 min; (13) Adjust the pH of the walnut protein solution to 11 and ultrasonicate for 5–10 min.

4. The preparation method according to claim 3, wherein: The power of the gradient ultrasonic treatment is 300-600 W / L, and the total time of the gradient ultrasonic treatment is 20-30 minutes.

5. The preparation method according to claim 2, wherein: In step (3), the ultrasonically modified walnut protein powder solution and the chitosan solution are mixed at a mass ratio of ultrasonically modified walnut protein powder to chitosan of 8-12:2-5.

6. The preparation method according to claim 2, wherein: In step (3), the stirring rate is 200-800 rpm, and the stirring time is 1-3 h.

7. The preparation method according to claim 2, characterized in that: The mass ratio of the ultrasonically modified walnut protein-chitosan composite solution to the oil phase in step (4) is 1:2-5.

8. The preparation method according to claim 2, wherein: The rotation speed of the shear homogenization in step (4) is 12000-18000 rpm, and the shear homogenization time is 1-5 min.

9. Application of gradient ultrasonic treatment of a walnut protein solution in the preparation of an ultrasonically modified walnut protein-chitosan composite emulsion, the gradient ultrasonic treatment of the walnut protein solution comprising the following steps: (11) The pH of the walnut protein solution was adjusted to 9 and ultrasonicated for 5-10 min; (12) adjusting the pH of the walnut protein solution to 10 and ultrasonically treating for 5-10 min; (13) Adjust the pH of the walnut protein solution to 11 and ultrasonicate for 5–10 min; Preferably, the application includes improving the thermal stability of ultrasonically modified walnut protein-chitosan composite emulsion.

10. Use of the ultrasonically modified walnut protein-chitosan composite emulsion according to claim 1 or the preparation method according to any one of claims 2 to 8 in the food field.

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