A method for regulating the geometric conformation of food-grade biphasic droplets through nanoparticle-mediated droplet engulfment mechanism

Abstract

The invention discloses a method for regulating the geometric conformation of food-grade biphasic droplets by a nanoparticle-mediated droplet engulfment mechanism, which belongs to the field of nanomaterial technology. The present invention breaks the molecular structure of zein by deamidation, and further improves the amphiphilicity of zein by Maillard reaction, thereby constructing nanoparticles with flexible conformation. The nanoparticles are dispersed in a sodium alginate solution, and biphasic droplets are prepared together with a zein solution and medium-chain triglycerides based on a vortex method, which can achieve effective control of the Janus structure or core-shell structure of the biphasic droplets. The method for regulating the geometric conformation of food-grade biphasic droplets provided by the present invention has the advantages of simple preparation of nanoparticles, easy regulation of the geometric conformation of biphasic droplets, and large-scale preparation. At the same time, the material for constructing the biphasic droplets has good biocompatibility.

Description

Technical Field

[0001] The present invention belongs to the technical field of nanomaterials, and in particular relates to a method for regulating the geometric conformation of food-grade two-phase droplets by using a nanoparticle-mediated droplet engulfment mechanism. Background Art

[0002] Traditional single-phase droplets have been extensively studied, possessing a single interface and compartment, but they cannot meet complex application requirements. In contrast, biphasic droplets, composed of two mutually incompatible dispersed phases and possessing multiple interfaces and compartments, are advantageous for meeting complex application requirements. Existing biphasic droplets are typically prepared using microfluidics, which has low efficiency and high cost, making them unsuitable for large-scale production. Collisions between incompatible droplets provide an opportunity for the formation of biphasic droplets and multiple interfaces. Due to the Marangoni effect, droplets with lower interfacial tension tend to spread on the surface of droplets with higher interfacial tension. The interfacial evolution during the collision of incompatible droplets leads to the formation of biphasic droplets with a water-water interface. However, surfactants cannot be stably adsorbed at the interface, but instead undergo a dynamic adsorption-desorption process. In contrast, particulate emulsifiers have excellent interfacial activity and exhibit irreversible adsorption behavior at the interface. Therefore, the conformation of biphasic droplets can be controlled by manipulating the interface of incompatible droplets stabilized by nanoparticles. It is necessary to develop nanoparticles with a specific structure and regulate the geometric conformation of biphasic droplets based on the phagocytosis of droplets.

[0003] As shown in the following patent: Application number: CN202010569775.6, Publication number: CN111701627A, the invention name is "A device and method for rapid generation of core-shell droplets based on surface acoustic wave microfluidics", which discloses a method for preparing core-shell droplets. This method can be used to prepare core-shell droplets, but the size of the droplets will be limited by the microfluidic device, and the preparation of smaller core-shell droplets cannot be achieved. In addition, microfluidic equipment has a high cost, and the production efficiency is also limited by the pores of the microfluidic chip, which is inefficient and not conducive to the large-scale preparation of core-shell droplets.

[0004] As shown in the following patent application: Application Number: CN201711155312.X, Publication Number: CN109806918A, Invention Name: "Preparation Method of Gelatin Methacrylamide Core-Shell Microspheres Based on Microfluidics Technology," a method for preparing core-shell droplets and particles is disclosed. This method uses methylcellulose and gelatin methacrylamide based on microfluidics to prepare core-shell droplets. However, the final particle size of the core-shell droplets is approximately 200 μm. This larger particle size is not conducive to the stability of the core-shell droplets and subsequent applications. In addition, the core-shell droplets are composed of methacrylamide, which is toxic and cannot be used in industries such as food, medicine, and cosmetics.

[0005] The article "Batch-scale preparation of reverse Janus emulsions" describes a technique for scalable preparation of Janus droplets. This method uses sodium carbonate and ethanol as mutually immiscible dispersed phases, constructs batches of Janus droplets via vortexing, and enables the manipulation of the droplet geometry. However, the presence of ethanol in the droplets may hinder their application in food applications, and this method cannot adjust the droplets to a core-shell conformation.

[0006] As mentioned above, there is still a lack of a method to control the geometric conformation of food-grade biphasic droplets and to prepare them on a large scale. Therefore, it is urgent to develop a method to efficiently manipulate the structure of biphasic droplets. Currently, nanoparticles can manipulate the interface structure of droplets, but methods to manipulate the geometric conformation of biphasic droplets by affecting the phagocytosis of droplets have not been reported. Summary of the Invention

[0007] The main purpose of the present invention is to provide a method for regulating the geometric conformation of food-grade biphasic droplets by a nanoparticle-mediated droplet engulfment mechanism and to prepare a food-grade biphasic droplet, so as to overcome the shortcomings of the prior art.

[0008] The principle of the present invention is to effectively control the geometric conformation of food-grade biphasic droplets by regulating the interfacial structure of sodium alginate droplets through nanoparticles. Zein is first deamidated to disrupt its molecular structure. Then, sodium alginate is grafted onto the protein via a Maillard reaction. The conjugate further assembles to form deamidated zein-sodium alginate nanoparticles. Zein-sodium alginate nanoparticles are then constructed via a Maillard reaction. The nanoparticles are dispersed in a sodium alginate solution. The geometric conformation of the biphasic droplets is then controlled by emulsifying the zein, sodium alginate solution, and a PGPR-containing MCT solution. The adsorption of the nanoparticles at the interface between sodium alginate and MCT forms an interfacial film of varying structure. The strength of this interfacial film influences the phagocytosis of the sodium alginate droplets by the zein droplets, further affecting the geometric conformation of the biphasic droplets, ultimately achieving effective control of Janus droplets and core-shell droplets.

[0009] In order to achieve the technical purpose of the present invention, the technical solution of the present invention includes the following steps:

[0010] (1) Disperse zein in deionized water and adjust the pH of the solution to obtain a zein stock solution.

[0011] (2) The zein solution is heated to obtain a deamidated zein solution.

[0012] (3) preparing a sodium alginate solution, adding the sodium alginate solution to the zein solution or the deamidated zein solution obtained in step (1) or (2), heating the zein solution containing sodium alginate, and cooling to obtain a zein-sodium alginate nanoparticle dispersion or a deamidated zein-sodium alginate nanoparticle dispersion.

[0013] (4) Based on zein solution, sodium alginate solution and medium-chain triglycerides (MCT), biphasic droplets were constructed.

[0014] (5) Dispersing the nanoparticles into the sodium alginate solution obtained in step (4).

[0015] (6) Dissolve polyglycerol ricinoleate (PGPR) in MCT.

[0016] (7) Zein solution, sodium alginate solution containing nanoparticles, and MCT solution containing PGPR were added in proportion and biphasic droplets were prepared by vortexing.

[0017] Preferably, the zein concentration in the zein solution in step (1) is 10 wt %, and the solution pH is 12.0;

[0018] Preferably, in step (2), the heating temperature is 80°C and the heating time is 10 h;

[0019] Preferably, in step (3), the concentration of the sodium alginate solution is 2 wt %, and the pH of the solution is 12.0;

[0020] Preferably, in step (3), the concentration of the zein solution or the deamidated zein solution is 2 wt %, and the pH of the solution is 12.0;

[0021] Preferably, in step (3), the mixing ratio of the sodium alginate solution to the zein solution or the deamidated zein solution is 1:1;

[0022] Preferably, in step (3), the sodium alginate solution and the zein solution or the deamidated zein solution are mixed and heated to 80° C. for 3.5 h.

[0023] Preferably, in step (4), the concentration of the zein solution is 10 wt %, and the pH of the solution is 12.0. The concentration of the sodium alginate solution is 1.5 wt %, and the pH of the solution is 12.0.

[0024] Preferably, in step (5), the concentration of the nanoparticles in sodium alginate is 1.5 mg / mL or 3 mg / mL;

[0025] Preferably, in step (6), the concentration of PGPR is 2 wt %;

[0026] Preferably, in step (7), the ratio of zein solution, sodium alginate solution and MCT is 2:1:7;

[0027] Preferably, in step (7), the vortex speed is 3000 rpm and the vortex time is 3 min.

[0028] Compared with the prior art, the present invention has the following beneficial effects:

[0029] (1) The present invention uses zein as one of the raw materials for constructing nanoparticles, which can still maintain an aggregated spherical structure in an alkaline environment. After zein is grafted with sodium alginate, the particle size of the nanoparticles is significantly reduced due to the increase in hydrophilicity, which can play a beneficial role in the oil-water interface. In addition, zein is broken into polypeptides through deamidation treatment, which are then grafted with sodium alginate through the Maillard reaction to form nanoparticles. The particles have a flexible conformation, which is conducive to rearrangement at the oil-water interface to form a complete interface structure, thereby regulating the phagocytosis of zein droplets.

[0030] (2) The present invention is the first to use nanoparticles to regulate the geometric conformation of biphasic droplets. The adsorption and rearrangement behavior of nanoparticles at the interface of sodium alginate droplets will significantly change the interface structure, and exhibit significant strain hardening behavior during the interface compression process, which helps to maintain the interface structure, thereby affecting the phagocytic effect of zein droplets on sodium alginate droplets, and ultimately achieving effective regulation of the geometric conformation of biphasic droplets.

[0031] (3) The zein, sodium alginate, medium-chain triglycerides, and polyglycerol ricinoleate used in the present invention are all food-grade raw materials, and no other substances are introduced during the construction of the nanoparticles. The constructed two-phase droplets have good biocompatibility and do not produce toxic and harmful substances. Therefore, they can be used in the fields of food, medicine, and cosmetics.

[0032] (4) The present invention innovatively proposes the phagocytic behavior between incompatible droplets. Driven by the Marangoni force, the zein droplets will engulf the sodium alginate droplets, which is similar to the phagocytic action of macrophages in the human body. This can better understand the formation process of biphasic droplets and regulate the droplet engulfment process to control the geometric conformation of biphasic droplets.

[0033] (5) The one-step emulsification method used in the present invention can prepare Janus droplets or core-shell droplets on a large scale, which has the beneficial effects of low cost and high efficiency compared with microfluidic technology.

[0034] (6) The two-phase droplets prepared by the present invention have a smaller particle size and exhibit good stability, which is conducive to maintaining stability in various application scenarios.

[0035] (7) The nanoparticles used in the present invention are easy to prepare. The organic combination of deamidation and Maillard reaction enables zein and sodium alginate to form nanoparticles with different structures. The reaction is efficient and safe. At the same time, it gives the nanoparticles better interfacial activity and can effectively regulate the phagocytosis between incompatible droplets. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 2 are transmission electron micrographs of zein-alginate nanoparticles (ZSNPs) (a) in Example 1 and deamidated zein-alginate nanoparticles (DZSNPs) (b) in Example 3.

[0037] Figure 2 These are SDS-PAGE patterns of zein (Zein), deamidated zein (DZein), zein-sodium alginate nanoparticles (ZSNPs), and deamidated zein-sodium alginate nanoparticles (DZSNPs) in Examples 2 and 4.

[0038] Figure 3 is the grafting degree of zein-sodium alginate nanoparticles (ZSNPs) and deamidated zein-sodium alginate nanoparticles (DZSNPs) in Example 1 and Example 3.

[0039] Figure 4 Example 1 and Example 3 are the free amino group contents of the samples before and after the Maillard reaction.

[0040] Figure 5 The absorbance of the samples at 420 nm before and after the Maillard reaction in Example 1 and Example 3 is shown.

[0041] Figure 6 The absorbance of the samples at 304 nm before and after the Maillard reaction in Example 1 and Example 3 is shown.

[0042] Figure 7 These are the intrinsic fluorescence spectra of zein (Zein), deamidated zein (DZein), zein-sodium alginate nanoparticles (ZSNPs), and deamidated zein-sodium alginate nanoparticles (DZSNPs) in Examples 2 and 4.

[0043] Figure 8These are Fourier transform infrared spectra of zein (Zein), deamidated zein (DZein), zein-sodium alginate nanoparticles (ZSNPs), and deamidated zein-sodium alginate nanoparticles (DZSNPs) in Examples 2 and 4.

[0044] Figure 9 These are the Raman spectra of zein (Zein), deamidated zein (DZein), zein-sodium alginate nanoparticles (ZSNPs), and deamidated zein-sodium alginate nanoparticles (DZSNPs) in Examples 2 and 4.

[0045] Figure 10 It is the protein secondary structure content of zein (Zein), deamidated zein (DZein), zein-sodium alginate nanoparticles (ZSNPs) and deamidated zein-sodium alginate nanoparticles (DZSNPs) in Example 2 and Example 4.

[0046] Figure 11 These are the disulfide bond conformations of zein, deamidated zein (DZein), zein-sodium alginate nanoparticles (ZSNPs), and deamidated zein-sodium alginate nanoparticles (DZSNPs) in Examples 2 and 4.

[0047] Figure 12 It is the I850 / I830 of zein, deamidated zein, zein-sodium alginate nanoparticles (ZSNPs), and deamidated zein-sodium alginate nanoparticles (DZSNPs) in Examples 2 and 4.

[0048] Figure 13 Effects of the content and structure of nanoparticles on the morphology of biphasic droplets in Examples 1-4; low zein-alginate nanoparticle addition amount (ZSNPs-L) (a), high zein-alginate nanoparticle addition amount (ZSNPs-H) (b), low deamidated zein-alginate nanoparticle addition amount (DZSNPs-L) (c), high deamidated zein-alginate nanoparticle addition amount (DZSNPs-H) (d).

[0049] Figure 14 The effect of the content and structure of nanoparticles in Examples 1-4 on the curvature of the two-phase droplet interface.

[0050] Figure 15Cryo-scanning electron micrographs of zein droplets (ab), sodium alginate droplets (cd), Janus droplets (ef), and core-shell droplets (gh) in Examples 3-6.

[0051] Figure 16 Effects of nanoparticles on the interfacial structure of sodium alginate droplets; low zein-alginate nanoparticle addition amount (ZSNPs-L) (a), high zein-alginate nanoparticle addition amount (ZSNPs-H) (b), low deamidated zein-alginate nanoparticle addition amount (DZSNPs-L) (c), high deamidated zein-alginate nanoparticle addition amount (DZSNPs-H) (d).

[0052] Figure 17 Lissajous plots of alginate droplets containing nanoparticles at 30% amplitude; low zein-alginate nanoparticle addition amount (ZSNPs-L) (a), high zein-alginate nanoparticle addition amount (ZSNPs-H) (b), low deamidated zein-alginate nanoparticle addition amount (DZSNPs-L) (c), high deamidated zein-alginate nanoparticle addition amount (DZSNPs-H) (d).

[0053] Figure 18 This is the interfacial viscoelastic modulus-amplitude curve of sodium alginate droplets containing nanoparticles.

[0054] Figure 19 The TSI curve of sodium alginate droplets containing nanoparticles.

[0055] Figure 20 The effect of nanoparticles on TSI of biphasic droplets. DETAILED DESCRIPTION

[0056] The following examples are used to illustrate the present invention but are not intended to limit the scope of the present invention.

[0057] Operations not mentioned in the present invention are all routine operations in the art, and materials whose specific sources are not mentioned in the present invention are all conventional materials that can be purchased from the market.

[0058] Example 1

[0059] (1) Prepare a 10 wt% zein solution and adjust its pH to 12.0 to obtain a zein stock solution. Dilute the zein solution to 2 wt%. Prepare a 2 wt% sodium alginate solution and adjust its pH to 12.0. Mix the zein solution and the sodium alginate solution in a 1:1 ratio. Heat the mixed solution at 80°C for 3.5 h to obtain zein-sodium alginate nanoparticles.

[0060] (2) Prepare a 10 wt% zein solution and a 1.5 wt% sodium alginate solution, and adjust the pH of the solution to 12.0. Prepare an MCT solution containing 2 wt% PGPR. Disperse zein-alginate nanoparticles in the sodium alginate solution at a particle concentration of 1.5 mg / mL.

[0061] (3) Zein solution, sodium alginate solution containing nanoparticles, and MCT solution were vortexed in a ratio of 2:1:7 at 3000 rpm for 3 min to prepare biphasic droplets.

[0062] Example 2

[0063] (1) Prepare a 10 wt% zein solution and adjust its pH to 12.0 to obtain a zein stock solution. Dilute the zein solution to 2 wt%. Prepare a 2 wt% sodium alginate solution and adjust its pH to 12.0. Mix the zein solution and the sodium alginate solution in a 1:1 ratio. Heat the mixed solution at 80°C for 3.5 h to obtain zein-sodium alginate nanoparticles.

[0064] (2) Prepare a 10 wt% zein solution and a 1.5 wt% sodium alginate solution, and adjust the pH of the solution to 12.0. Prepare an MCT solution containing 2 wt% PGPR. Disperse zein-alginate nanoparticles in the sodium alginate solution at a particle concentration of 3 mg / mL.

[0065] (3) Zein solution, sodium alginate solution containing nanoparticles, and MCT solution were vortexed in a ratio of 2:1:7 at 3000 rpm for 3 min to prepare biphasic droplets.

[0066] Example 3

[0067] (1) Prepare a 10 wt% zein solution and adjust its pH to 12.0 to obtain a zein stock solution. Heat the 10 wt% zein solution at 80°C for 10 h and cool it to obtain a deamidated zein solution. Dilute the deamidated zein solution to 2 wt%. Prepare a 2 wt% sodium alginate solution and adjust its pH to 12.0. Mix the deamidated zein solution and the sodium alginate solution in a 1:1 ratio. Heat the mixed solution at 80°C for 3.5 h to obtain deamidated zein-sodium alginate nanoparticles.

[0068] (2) Prepare a 10 wt% zein solution and a 1.5 wt% sodium alginate solution, and adjust the pH of the solution to 12.0. Prepare an MCT solution containing 2 wt% PGPR. Disperse deamidated zein-sodium alginate nanoparticles in the sodium alginate solution at a particle concentration of 1.5 mg / mL.

[0069] (3) Zein solution, sodium alginate solution containing nanoparticles, and MCT solution were vortexed in a ratio of 2:1:7 at 3000 rpm for 3 min to prepare biphasic droplets.

[0070] Example 4

[0071] (1) Prepare a 10 wt% zein solution and adjust its pH to 12.0 to obtain a zein stock solution. Dilute the deamidated zein solution to 2 wt%. Prepare a 2 wt% sodium alginate solution and adjust the pH to 12.0. Mix the zein solution or deamidated zein solution with the sodium alginate solution in a 1:1 ratio. Heat the mixed solution at 80°C for 3.5 h to obtain deamidated zein-sodium alginate nanoparticles.

[0072] (2) Prepare a 10 wt% zein solution and a 1.5 wt% sodium alginate solution, and adjust the pH of the solution to 12.0. Prepare an MCT solution containing 2 wt% PGPR. Disperse deamidated zein-alginate nanoparticles in the sodium alginate solution at a particle concentration of 3 mg / mL.

[0073] (3) Zein solution, sodium alginate solution containing nanoparticles, and MCT solution were vortexed in a ratio of 2:1:7 at 3000 rpm for 3 min to prepare biphasic droplets.

[0074] Comparative Example 1

[0075] (1) Prepare a 10 wt% zein solution and adjust its pH to 12.0 to obtain a zein stock solution. Dilute the zein solution to 2 wt%. Prepare a 2 wt% sodium alginate solution and adjust its pH to 12.0. Mix the zein solution and the sodium alginate solution in a 1:1 ratio. Heat the mixed solution at 80°C for 3.5 h to obtain zein-sodium alginate nanoparticles.

[0076] (2) Prepare a 1.5 wt% sodium alginate solution and adjust the pH to 12.0. Prepare an MCT solution containing 2 wt% PGPR. Disperse zein-alginate nanoparticles in the sodium alginate solution at a particle concentration of 1.5 mg / mL.

[0077] (3) The sodium alginate solution containing nanoparticles and the MCT solution were vortexed in a ratio of 3:7 at a speed of 3000 rpm for 3 min to prepare sodium alginate droplets containing nanoparticles.

[0078] Comparative Example 2

[0079] (1) Prepare a 10 wt% zein solution and adjust its pH to 12.0 to obtain a zein stock solution. Dilute the zein solution to 2 wt%. Prepare a 2 wt% sodium alginate solution and adjust its pH to 12.0. Mix the zein solution and the sodium alginate solution in a 1:1 ratio. Heat the mixed solution at 80°C for 3.5 h to obtain zein-sodium alginate nanoparticles.

[0080] (2) Prepare a 1.5 wt% sodium alginate solution and adjust the pH to 12.0. Prepare an MCT solution containing 2 wt% PGPR. Disperse zein-alginate nanoparticles in the sodium alginate solution at a particle concentration of 3 mg / mL.

[0081] (3) The sodium alginate solution containing nanoparticles and the MCT solution were vortexed in a ratio of 3:7 at a speed of 3000 rpm for 3 min to prepare sodium alginate droplets containing nanoparticles.

[0082] Comparative Example 3

[0083] (1) Prepare a 10 wt% zein solution and adjust its pH to 12.0 to obtain a zein stock solution. Heat the 10 wt% zein solution at 80°C for 10 h and cool it to obtain a deamidated zein solution. Dilute the deamidated zein solution to 2 wt%. Prepare a 2 wt% sodium alginate solution and adjust its pH to 12.0. Mix the deamidated zein solution and the sodium alginate solution in a 1:1 ratio. Heat the mixed solution at 80°C for 3.5 h to obtain deamidated zein-sodium alginate nanoparticles.

[0084] (2) Prepare a 1.5 wt% sodium alginate solution and adjust the pH to 12.0. Prepare an MCT solution containing 2 wt% PGPR. Disperse deamidated zein-sodium alginate nanoparticles in the sodium alginate solution at a particle concentration of 1.5 mg / mL.

[0085] (3) The sodium alginate solution containing nanoparticles and the MCT solution were vortexed in a ratio of 3:7 at a speed of 3000 rpm for 3 min to prepare sodium alginate droplets containing nanoparticles.

[0086] Comparative Example 4

[0087] (1) Prepare a 10 wt% zein solution and adjust its pH to 12.0 to obtain a zein stock solution. Heat the 10 wt% zein solution at 80°C for 10 h and cool it to obtain a deamidated zein solution. Dilute the deamidated zein solution to 2 wt%. Prepare a 2 wt% sodium alginate solution and adjust its pH to 12.0. Mix the deamidated zein solution and the sodium alginate solution in a 1:1 ratio. Heat the mixed solution at 80°C for 3.5 h to obtain deamidated zein-sodium alginate nanoparticles.

[0088] (2) Prepare a 1.5 wt% sodium alginate solution and adjust the pH to 12.0. Prepare an MCT solution containing 2 wt% PGPR. Disperse deamidated zein-sodium alginate nanoparticles in the sodium alginate solution at a particle concentration of 3 mg / mL.

[0089] (3) The sodium alginate solution containing nanoparticles and the MCT solution were vortexed in a ratio of 3:7 at a speed of 3000 rpm for 3 min to prepare sodium alginate droplets containing nanoparticles.

[0090] Comparative Example 5

[0091] (1) A 10 wt% zein solution was prepared and its pH was adjusted to 12.0 to obtain a zein stock solution.

[0092] (2) Prepare an MCT solution containing 2 wt% PGPR.

[0093] (3) The zein solution and MCT solution were vortexed in a ratio of 3:7 at a speed of 3000 rpm for 3 min to prepare zein single-phase droplets.

[0094] Comparative Example 6

[0095] (1) Prepare a 1.5 wt% sodium alginate solution and adjust its pH to 12.0 to obtain a sodium alginate stock solution.

[0096] (2) Prepare an MCT solution containing 2 wt% PGPR.

[0097] (3) Sodium alginate solution and MCT solution were vortexed in a ratio of 3:7 at a speed of 3000 rpm for 3 min to prepare sodium alginate single-phase droplets.

[0098] Experimental Example 1 Transmission Electron Microscope Observation

[0099] The sample dispersions obtained in Example 1 and Example 3 were dropped onto a copper mesh. After drying, the morphology of the samples was observed using a transmission electron microscope.

[0100] pass Figure 1 The experimental results show that both samples are spherical nanoparticles, indicating that nanoparticles are successfully constructed based on zein and sodium alginate, and deamidated zein-alginate nanoparticles (DZSNP S ) have a smaller particle size than zein-alginate nanoparticles (ZSNPs).

[0101] Experimental Example 2 SDS-PAGE

[0102] The samples obtained in Example 1 and Example 3 were subjected to electrophoresis. The samples were mixed with the loading buffer and heated in boiling water for 5 minutes. Electrophoresis was performed using a voltage of 80 V in the stacking gel and 120 V in the separating gel, respectively, and stained with Coomassie Brilliant Blue R-250.

[0103] pass Figure 2 The experimental results show that after the Maillard reaction, the protein band moves up significantly, indicating that sodium alginate is grafted onto the protein through the Maillard reaction.

[0104] Experimental Example 3 Determination of free amino content, grafting degree and Maillard reaction products

[0105] The free amino group content of the samples obtained in Examples 2 and 4 was determined using the o-phthalaldehyde (OPA) method. The OPA reagent was mixed with the sample and incubated at 35°C for 2 minutes. The absorbance at 340 nm was measured using a UV-visible spectrophotometer to calculate the free amino acid content and the degree of grafting. After dilution, the sample absorbance was measured at 304 nm and 420 nm.

[0106] pass Figure 3-6 The experimental results show that the content of free amino groups is significantly reduced after the Maillard reaction, and the proteoglycan particles have a higher degree of grafting. The content of Maillard reaction products represented by the absorbance at 304 nm and 420 nm also increases significantly, indicating that the Maillard reaction occurs between the protein and the polysaccharide.

[0107] Experimental Example 4 Fluorescence Spectrum

[0108] The samples obtained in Example 1 and Example 3 were diluted to an appropriate concentration, and fluorescence spectra within the range of 300-500 nm were obtained at an excitation wavelength of 280 nm.

[0109] pass Figure 7The experimental results show that the fluorescence intensity of zein increases after deamidation, which may be caused by the unfolding of the protein structure. The fluorescence intensity decreases significantly after the Maillard reaction, which is caused by the rearrangement of the protein structure after the hydrophilic polysaccharide grafting.

[0110] Experimental Example 5 Fourier Transform Infrared Spectroscopy

[0111] The samples obtained in Example 2 and Example 4 were mixed with potassium bromide and pressed into tablets to obtain 4000-500 cm -1 Fourier transform infrared spectroscopy in the range.

[0112] pass Figure 8 The experimental results show that 3000-3500 cm -1 The characteristic peaks in the range represent C─N and O─H vibrations. After the Maillard reaction, the peaks tend to be flat, which may be caused by the formation of hydrogen bonds. In addition, the infrared spectra of ZSNPs and DZSNPs at 1033 cm -1 A new characteristic peak appeared at , which corresponds to the stretching vibration of the C─O─C bond in sodium alginate, indicating the effective coupling of sodium alginate and zein.

[0113] Experimental Example 6 Raman Spectroscopy

[0114] Raman spectra of the samples obtained in Examples 2 and 4 were measured. Spectral acquisition conditions were: 785 nm, 1800 g / mm, with an acquisition time of 15 s and an accumulation time of 20 times. Changes in disulfide bond conformation, tyrosine (Tyr) microenvironment, and protein secondary structure were further analyzed.

[0115] pass Figure 9 The experimental results show that after the Maillard reaction, the -1 A new characteristic peak appeared at the wavelength, which belongs to sodium alginate COO ─ The vibration of the groups indicates the successful grafting of sodium alginate. The changes in the secondary structure of the protein are further analyzed by spectral information, such as Figure 10 As shown in Figure 5, the Maillard reaction resulted in a decrease in α-helix content and an increase in β-sheet content, indicating that zein underwent structural unfolding and rearrangement. -1 The changes in protein disulfide bond conformation are analyzed at different wavelengths, such as Figure 11 As shown in Figure 2, under high pH conditions, sulfhydryl groups are more likely to be deprotonated, thereby increasing the possibility of disulfide bond formation. GGT and GGG represent intrachain and interchain disulfide bonds, respectively. The GGT content is significantly reduced after the Maillard reaction, indicating that the breakage of intrachain disulfide bonds is beneficial to improving the flexibility of nanoparticles. The changes in the tyrosine microenvironment in proteins are further analyzed by the I850 / I830 value, as shown in Figure 2. Figure 12 As shown in the figure, after grafting with sodium alginate, the I850 / I830 ratio decreases, indicating that tyrosine is buried within the zein. Grafting with sodium alginate causes a structural rearrangement of zein due to changes in hydrophilicity. This results in adjacent tyrosines binding to each other through non-covalent interactions such as hydrogen bonding and π-π stacking.

[0116] Experimental Example 7 Laser Confocal Microscopy

[0117] Laser confocal images of the droplet samples of Examples 1 to 4 were taken. Zein and nanoparticles were labeled with rhodamine B and FITC, respectively, and excited at 561 nm and 488 nm.

[0118] pass Figure 13 The experimental results show that at low particle addition levels, the biphasic droplets still exhibit a Janus structure, but when a high content of DZSNPs is added, the biphasic droplets exhibit core-shell droplets. Figure 14 The experimental results also show that the addition of particles significantly affects the interfacial curvature of the two-phase droplets.

[0119] Experimental Example 8 Cryo-Scanning Electron Microscopy

[0120] Cryo-scanning electron microscope images of droplet samples from Examples 3 and 4 and Comparative Examples 5 and 6 were taken. The samples were placed on a sample stage and frozen in liquid nitrogen. The samples were then transferred to a preparation chamber and fractured. The samples were sublimated and then coated with platinum for morphological observation.

[0121] pass Figure 15 The experimental results show that the single-phase zein droplets ( Figure 15 a and b) and single-phase alginate droplets ( Figure 15 c and d) show obvious differences in their network structures. Zein droplets have a denser interface network, while alginate droplets have a looser and more porous network structure. This is mainly due to the concentration difference between the two phases. After adding particles, the two-phase droplets appear as Janus droplets ( Figure 15 e and f) and core-shell droplets ( Figure 15 g and h), demonstrating that the addition of nanoparticles can regulate the geometric conformation of biphasic droplets.

[0122] Experimental Example 9 Interfacial behavior of nanoparticles at the oil-water interface.

[0123] The interfacial behavior of the nanoparticles at the oil-water interface in Comparative Examples 1 to 4 was photographed using a laser confocal microscope, and the interfacial properties of the nanoparticles were determined by interfacial dilatation rheology.

[0124] pass Figure 16The experimental results show that the nanoparticles are adsorbed on the oil-water interface, and as the amount of nanoparticles added increases, the content of nanoparticles on the interface increases. When the amount of DZSNPs added is high, the nanoparticles form a complete interface layer at the interface, which may be due to the better flexibility of DZSNPs and the rearrangement at the interface. Figure 17 The experimental results show that after adding particles, the Lissajous curve becomes asymmetric. This is because the adsorption of nanoparticles affects the interface structure. The interface undergoes strain hardening during compression. This may be due to the interface blockage caused by nanoparticles, which is conducive to maintaining the interface structure. Figure 18 It can be seen from the experimental results that the addition of nanoparticles affects the interface structure.

[0125] Experimental Example 10 Effect of Nanoparticles on the Stability of Sodium Alginate Droplets

[0126] The stability of the samples of Comparative Examples 1-4 was determined by multiple static light scattering technique. Figure 19 The experimental results show that the addition of nanoparticles enhances the stability of sodium alginate droplets, mainly because the adsorption of nanoparticles at the interface enhances the interfacial film structure.

[0127] Experimental Example 11 Effect of Nanoparticles on the Stability of Biphasic Droplets

[0128] The stability of the samples of Examples 1-4 was determined by multiple static light scattering technique. Figure 20 The experimental results show that TSI is significantly reduced after the addition of nanoparticles, indicating that the addition of nanoparticles enhances the stability of the two-phase droplets.

Claims

1. A method for regulating the geometric conformation of food-grade biphasic droplets using a nanoparticle-mediated droplet engulfment mechanism, characterized by: The specific preparation method comprises the following steps: (1) Dispersing zein in deionized water and adjusting the pH of the solution to obtain a zein solution; (2) heating the zein solution to obtain a deamidated zein (DZein) solution; (3) preparing a sodium alginate solution, adding the sodium alginate solution to the solution of step (2) and heating the solution at a temperature of 60-90° C. for 1-5 h, and cooling the solution to obtain a deamidated zein-sodium alginate nanoparticle (DZSNPs) dispersion; (4) preparing zein solution and sodium alginate solution, and constructing biphasic droplets based on zein solution, sodium alginate solution and medium-chain triglycerides (MCT); (5) dispersing the nanoparticles prepared in step (3) in the sodium alginate solution prepared in step (4); the concentration of the nanoparticles in the sodium alginate solution is 1-5 mg / mL; (6) Dissolve polyglycerol ricinoleate (PGPR) in MCT, with the concentration of PGPR being 1-3 wt%; (7) Zein solution, sodium alginate solution containing nanoparticles, and MCT solution containing PGPR were added in proportion and biphasic droplets were prepared by vortexing; Wherein, in step (3), the concentration of the sodium alginate solution is 1-2 wt %, and the pH of the solution is 11.5-12.5; the concentration of the deamidated zein solution is 1-2 wt %, and the pH of the solution is 11.5-12.5; and the mixing ratio of the sodium alginate solution to the deamidated zein solution is 2:1-1:

2.

2. The method according to claim 1, characterized in that include: The zein concentration in the zein solution in step (1) is 6-10 wt %, and the pH value of the solution is 11.5-12.

5.

3. The method according to claim 1, characterized in that include: In step (2), the heating temperature is 60-90°C and the heating time is 2-10 hours.

4. The method according to claim 1, characterized in that include: In step (4), the concentration of the zein solution is 6-10 wt %, and the pH of the solution is 11.5-12.5; The concentration of the sodium alginate solution is 1-2wt%, and the pH of the solution is 11.5-12.

5.

5. The method according to claim 1, characterized in that include: In step (7), the ratio of zein solution, sodium alginate solution and MCT is 2:1:7 or 1:1:8; the vortex speed is 1000-3000 rpm, and the vortex time is 1-3 min.

6. Food-grade biphasic droplets prepared according to the method according to any one of claims 1 to 5.

7. Use of the biphasic droplets according to claim 6 in the fields of food, medicine or cosmetics.

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