A mid-internal phase Pickering emulsion gel, its preparation method and application

Abstract

This invention belongs to the field of hydrogel preparation. It discloses a mid-internal phase Pickering emulsion gel, its preparation method, and its applications. The gel comprises glycyrrhizic acid, a cationic compound, an oil phase, and water. The glycyrrhizic acid in the emulsion gel is 0.7-1.5 wt% by weight (based on available glycyrrhizic acid); the cationic compound is 0.1-0.5 wt% by weight, and is selected from at least one of lysozyme, polylysine, zinc chloride, calcium chloride, or magnesium chloride; the oil phase has a volume fraction of 30%-70%. This invention achieves efficient gelation under mid-internal phase conditions through electrostatic bridging of low-concentration glycyrrhizic acid and trace amounts of cations, overcoming the dependence of traditional techniques on high-concentration emulsifiers, composite systems, or high oil phases.

Description

Technical Field

[0001] This invention relates to the field of hydrogel preparation technology, specifically to a mid-internal phase Pickering emulsion gel, its preparation method, and its application. Background Technology

[0002] Pickering emulsion gels, as soft solid materials stabilized by solid particles, combine the dispersibility of emulsions with the mechanical properties of gels, and have significant application prospects in food, medicine, and other fields. Based on the internal phase volume fraction, they can be divided into two categories: high internal phase (φ≥74%) and medium-low internal phase (φ<74%). However, existing technologies have the following key limitations:

[0003] 1. High concentration dependence and high oil phase limitation

[0004] Traditional methods require high concentrations of emulsifiers or complex systems to achieve gelation. For example, the paper "The self-assembly of glycyrrhizic acid into nanofibrils" published in the Journal of Colloid and Interface Science points out that glycyrrhizic acid (GA) needs a concentration of ≥4 wt% or a complex polysaccharide (total >1 wt%) to stabilize high internal phase emulsions (oil phase >74%). This relies on molecular self-assembly to form a nanofiber network (critical aggregation concentration CAC = 0.22 mM, critical gel concentration CGC = 2 mM), but it is only suitable for extreme conditions with an oil phase >74%, and the introduction of a high oil phase will lead to a high fat risk in the product, limiting low-fat applications.

[0005] 2. System complexity and cumbersome processes

[0006] To reduce the oil phase ratio, existing technologies require the introduction of composite particles or external energy input. For example, using whey protein isolate + γ-oryzanol composite particles requires pH adjustment and heating at 85°C for 1 hour to stabilize a 15% low oil phase emulsion, which relies on protein denaturation and complex processes; or using ultrasonic disruption to assist composite particles in stabilizing a 20% oil phase emulsion, which relies on mechanical force rather than intermolecular forces, resulting in low production efficiency and high costs.

[0007] 3. Technical gaps in mesophase gelation

[0008] Current research focuses on systems with high internal phase (>74%) or low oil phase (<30%), while systems with medium internal phase (30%-70%) are difficult to gel with low-concentration emulsifiers due to their moderate oil droplet spacing and weak interfacial interactions. Traditional low-concentration systems (such as <2wt% glycyrrhizic acid) can only form fluid emulsions and cannot simultaneously meet the dual requirements of "medium internal phase" and "low-concentration stability".

[0009] The main problems currently existing are:

[0010] (1) The contradiction between cost and stability caused by high concentration of glycyrrhizic acid. The paper "Herbal small molecule-based low / medium internalphase supramolecular gel emulsion for diabetic wound healing" published in the Journal of Colloid and Interface Science relies on ≥4wt% glycyrrhizic acid to stabilize high internal phase emulsion (oil phase >74%). Although it can form a gel structure, the high concentration leads to a surge in raw material costs, and the emulsion is prone to separation when the oil phase is <60%.

[0011] (2) Interfacial competition and functional limitations caused by composite emulsion systems. CN114557440A discloses a “Pickering emulsion gel stabilized by zein-glycyrrhizic acid-chitosan and its preparation method”, which requires the addition of >1wt% chitosan and zein to co-stabilize the emulsion. This design leads to: decreased interfacial adsorption efficiency (zein and chitosan compete for the oil-water interface); loss of stimuli responsiveness: the chitosan cross-linked and solidified network cannot achieve dynamic structural recombination (such as self-healing, shear thinning). Summary of the Invention

[0012] Therefore, the purpose of this invention is to provide a Pickering emulsion gel based on electrostatic bridging of low-concentration glycyrrhizic acid and cations, and its preparation method. This invention achieves efficient gelation under meso-internal phase conditions by electrostatic bridging of low-concentration glycyrrhizic acid (≤1.5 wt%) and trace amounts of cations (0.1-0.5 wt%), overcoming the dependence of traditional techniques on high-concentration emulsifiers, composite systems, or high oil phases. This provides a novel approach for the low-cost and green preparation of meso-internal phase Pickering emulsion gels.

[0013] The technical solution provided by this invention is as follows:

[0014] A mid-internal phase Pickering emulsion gel comprises glycyrrhizic acid, a cationic compound, an oil phase, and water;

[0015] The glycyrrhizic acid, calculated as available glycyrrhizic acid, constitutes 0.7-1.5 wt% of the emulsion gel.

[0016] The cationic compound has a weight percentage of 0.1-0.5 wt% in the emulsion gel, and the cationic compound is selected from at least one of lysozyme, polylysine, zinc chloride, calcium chloride, and magnesium chloride.

[0017] The volume fraction of the oil phase is 30%-70%.

[0018] The oil phase is selected from at least one of soybean oil, corn oil, sunflower oil, and olive oil.

[0019] The oil phase also contains paeonol, and the weight percentage of paeonol is 0.5-5 wt% based on the total weight of the oil phase.

[0020] Preferably, the glycyrrhizic acid in the emulsion gel is 0.7-1.0 wt% based on available glycyrrhizic acid.

[0021] Preferably, the cationic compound has a weight percentage of 0.1-0.2 wt%.

[0022] Preferably, the volume fraction of the oil phase is 40-50%.

[0023] In a second aspect, the present invention provides a method for obtaining an internal phase Pickering emulsion gel as described above, comprising the following steps:

[0024] 1) Preparation of aqueous solution: Dissolve glycyrrhizic acid in water, heat to 70-90℃ and maintain for 20-40 minutes to form a transparent aqueous solution;

[0025] 2) Pickering emulsion preparation: The oil phase is added to the aqueous solution in step 1), and emulsified at high speed of 20,000-25,000 rpm for 1-3 minutes at room temperature to obtain the Pickering emulsion;

[0026] 3) Emulsion gel construction: Add an aqueous solution of a cationic compound to the Pickering emulsion of step 2), and let it stand at room temperature for 5-30 minutes to induce gelation to obtain the middle internal phase Pickering emulsion gel.

[0027] In step 3), the concentration of the cationic compound aqueous solution is 40-60 mg / mL, and the volume added is such that the weight percentage of the cationic compound in the emulsion gel is 0.1-0.5 wt%.

[0028] Thirdly, the present invention provides the application of the above-described mid-internal phase Pickering emulsion gel in drug delivery systems, wound dressings, tissue engineering scaffolds, topical skin preparations, or cosmetics.

[0029] Compared with the prior art, the present invention has the following beneficial effects:

[0030] (1) By using a method of electrostatic bridging of low concentration glycyrrhizic acid (≤1.5wt%) and trace amounts of cations (0.1-0.5wt%), the problem of traditional Pickering emulsions relying on high concentrations of emulsifiers (>4wt% glycyrrhizic acid) or complex polysaccharides (total amount >1wt%) to prepare emulsion gels (oil phase >74%) has been solved. The technical effect of achieving gelation of the middle internal phase (oil phase 40-50%) with extremely low concentration (glycyrrhizic acid dosage: 0.7-1.5wt%) has been achieved, and the storage modulus (G') has been increased to 1000 Pa (about 100 times higher than the cation-free system), breaking through the industry's cognitive barrier of linear relationship between concentration and performance.

[0031] (2) By using the method of constructing an ordered three-dimensional network based on electrostatic bridging of cationic and glycyrrhizic acid, the key contradiction of traditional Pickering emulsions relying on high concentrations of emulsifiers or physical blending is difficult to balance mechanical strength and dynamic functions (such as self-healing). Breakthrough results have been achieved in achieving high mechanical strength (G'≥100 Pa) and simultaneously endowing self-healing (recovery rate≥90%) at an ultra-low concentration of 1.0 wt%.

[0032] (3) By adopting the sequential process of “preparing Pickering emulsion first and then introducing cations to construct Pickering emulsion gel through electrostatic bridging”, the problems of cation-glycyrrhizic acid cross-linking blocking the equipment and electrostatic failure caused by high temperature in the traditional one-step mixing method were solved, and a construction strategy of constructing Pickering emulsion gel by simple two-step method was obtained. Attached Figure Description

[0033] Other features, objects, and advantages of the present invention will become more apparent from the following detailed description of non-limiting embodiments with reference to the accompanying drawings:

[0034] Figure 1 A comparison chart of Pickering emulsion (PE1) and Pickering emulsion gel AD;

[0035] Figure 2 A comparison chart of Pickering emulsion (PE1) and Pickering emulsion gel E;

[0036] Figure 3 A comparison diagram of Pickering emulsion (PE1) and Pickering emulsion gel F;

[0037] Figure 4 A comparison diagram of Pickering emulsion (PE1) and Pickering emulsion gel G;

[0038] Figure 5A comparison diagram of Pickering emulsion (PE1) and Pickering emulsion gel H;

[0039] Figure 6 A comparison diagram of Pickering emulsion (PE1) and Pickering emulsion gel I;

[0040] Figure 7 A comparison diagram of Pickering emulsion (PE1) and Pickering emulsion gel J;

[0041] Figure 8 A comparison diagram of Pickering emulsion (PE1) and Pickering emulsion gel K;

[0042] Figure 9 The results of free radical scavenging by Pickering emulsion (PE1), Pickering emulsion gel B, and Pickering emulsion gel L are shown; (a) full wavelength scan (PBS as negative control, H2O2 as positive control); (b) DPPH scavenging rate (%) of different sample amounts.

[0043] Figure 10 The cell viability (%) of Pickering emulsion (PE1) and Pickering emulsion gels B and L was used; among them, the PBS group was the negative control group and the H2O2 group was the positive control group.

[0044] Figure 11 The effects of Pickering emulsion (PE1) and Pickering emulsion gels B and L on cell morphology were investigated; the PBS group served as the negative control group, and the H2O2 group served as the positive control group. The scale bar was 100 μm.

[0045] Figure 12 The effect of Pickering emulsion (PE1) and Pickering emulsion gels B and L on intracellular ROS scavenging was evaluated; the PBS group was the negative control group and the H2O2 group was the positive control group. The scale bar was 100 μm.

[0046] Figure 13 The effects of Pickering emulsion (PE1) and Pickering emulsion gels B and L on the repair of diabetic wounds were studied; the PBS group served as the negative control group; (a) a schematic diagram of the rat diabetic model and wound healing process; (b) representative photographs of the rat diabetic wound healing process after different sample treatments. n = 3), representing days 0, 3, 7, and 14 respectively; (c) Quantitative analysis of wound area in each group of diabetic patients ( n= 3); (d) H&E staining of the wound area on day 7 and day 14 ( n = 3, scale bar is 100 μm) and Masson staining of the wound area on day 14 ( n = 3, scale bar is 100 μm).

[0047] Figure 14 Rheological data for Pickering emulsion (PE1) and Pickering emulsion gel (AD) are given; where G' is the storage modulus (represented by solid pattern) and G" is the loss modulus (represented by hollow pattern). Detailed Implementation

[0048] The present invention will be described in detail below with reference to embodiments. These embodiments will help those skilled in the art to further understand the present invention, but do not limit the invention in any way. It should be noted that those skilled in the art can make several adjustments and improvements without departing from the concept of the present invention. These all fall within the protection scope of the present invention.

[0049] Example 1

[0050] 1. Preparation of glycyrrhizic acid aqueous solution

[0051] Accurately weigh 0.10 g of glycyrrhizic acid (brand: Aladdin, CAS No.: 53956-04-0, product number: G109353-100g) and add it to 10 mL of distilled water to obtain a 0.7 wt% glycyrrhizic acid aqueous solution (the glycyrrhizic acid content is the mass of glycyrrhizic acid contained in the aqueous phase. Aladdin's glycyrrhizic acid is ≥70% (HPLC), and an actual weighing of 1 g is equivalent to 0.7 g of glycyrrhizic acid). Heat at 80℃ for 30 min to dissolve the solution, resulting in a clear glycyrrhizic acid aqueous solution.

[0052] 2. Pickering emulsion preparation

[0053] Add 5 mL of glycyrrhizic acid aqueous solution to a 20 mL sample bottle, slowly add 5 mL of soybean oil (COFCO Group), and disperse at high speed of 24000 rpm for 2 min to obtain a glycyrrhizic acid-stable O / W type internal phase (φ = 50%) Pickering emulsion (named PE).

[0054] 3. Preparation of Pickering emulsion gels with different concentrations of lysozyme

[0055] Add different amounts of lysozyme aqueous solution (stock solution concentration 50 mg / mL) to Pickering emulsion (PE1) according to Table 1, and let stand at room temperature (25℃±2℃) for 10 minutes to obtain Pickering emulsion gel AD. Figure 1Comparison graphs of Pickering emulsion (named PE1) and Pickering emulsion gel AD. The emulsion gel was tested using a rheometer (TA, DHR-2, USA); the rheological properties of the medium-low internal phase Pickering gel emulsion with different lysozyme contents are shown below. Figure 14 As shown, Figure 14 In the table, G' represents the storage modulus (represented by solid patterns), and G" represents the loss modulus (represented by hollow patterns). The higher the lysozyme content, the stronger the resulting gel emulsion; the results are shown in Table 1.

[0056] Table 1. Effect of different lysozyme dosages on Pickering emulsion gel.

[0057]

[0058] Example 2

[0059] 1. Preparation of glycyrrhizic acid aqueous solution

[0060] Same as Example 1.

[0061] 2. Pickering emulsion preparation

[0062] Same as Example 1.

[0063] 3. Preparation of Pickering emulsion gel using polylysine

[0064] Add 100 μL of 50 mg / mL polylysine aqueous solution to Pickering emulsion (PE1) and let it stand at room temperature (25℃±2℃) for 10 minutes to obtain Pickering emulsion gel E.

[0065] The results are as follows Figure 2 As shown, 100 μL of 50 mg / mL polylysine can induce the Pickering emulsion to form Pickering emulsion gel E.

[0066] Example 3

[0067] 1. Preparation of glycyrrhizic acid aqueous solution

[0068] Same as Example 1.

[0069] 2. Pickering emulsion preparation

[0070] Same as Example 1.

[0071] 3. Preparation of Pickering emulsion gel using zinc chloride

[0072] Add 100 μL of 50 mg / mL zinc chloride aqueous solution to Pickering emulsion (PE1) and let it stand at room temperature (25℃±2℃) for 10 minutes to obtain Pickering emulsion gel F.

[0073] The results are as follows Figure 3 As shown, 100 μL of 50 mg / mL zinc chloride aqueous solution can induce Pickering emulsion to form Pickering emulsion gel F.

[0074] Example 4

[0075] 1. Preparation of glycyrrhizic acid aqueous solution

[0076] Same as Example 1.

[0077] 2. Pickering emulsion preparation

[0078] Same as Example 1.

[0079] 3. Add 100 μL of 50 mg / mL calcium chloride aqueous solution to Pickering emulsion (PE1) and let it stand at room temperature (25℃±2℃) for 10 minutes to obtain Pickering emulsion gel G.

[0080] The results are as follows Figure 4 As shown, 100 μL of 50 mg / mL calcium chloride aqueous solution can induce Pickering emulsion to form Pickering emulsion gel G.

[0081] Example 5

[0082] 1. Preparation of glycyrrhizic acid aqueous solution

[0083] Same as Example 1.

[0084] 2. Pickering emulsion preparation

[0085] Same as Example 1.

[0086] 3. Preparation of Pickering emulsion gel using magnesium chloride

[0087] Add 100 μL of 50 mg / mL magnesium chloride aqueous solution to Pickering emulsion (PE1) and let it stand at room temperature (25℃±2℃) for 10 minutes to obtain Pickering emulsion gel H.

[0088] The results are as follows Figure 5 As shown, 100 μL of 50 mg / mL magnesium chloride aqueous solution can induce Pickering emulsion to form Pickering emulsion gel H.

[0089] Comparative Example 1

[0090] The difference between this comparative example and Example 1 is that the concentration of the glycyrrhizic acid aqueous solution was adjusted to 10.0 wt%, that is, 1.00 g of glycyrrhizic acid was accurately weighed, 10 mL of distilled water was added, and the solution was heated at 80°C for 30 min to obtain a 10.0 wt% glycyrrhizic acid aqueous solution; the remaining steps were the same as in Example 1.

[0091] The results are as follows Figure 6 As shown, due to the high concentration of glycyrrhizic acid, it rapidly gelled after being heated to 80°C and cooled to room temperature. This caused some of the aqueous solution to gel directly during the emulsion preparation process, resulting in phase separation between the gel and the emulsion gel, and making it impossible to obtain a uniform Pickering emulsion gel I.

[0092] Comparative Example 2

[0093] The difference between this comparative example and Example 1 is that lysozyme was replaced with chitosan (100 μL 50 mg / mL).

[0094] The results are as follows Figure 7 Chitosan has poor dispersibility in Pickering emulsions, leading to the local formation of Pickering emulsion gels.

[0095] Comparative Example 3

[0096] The difference between this comparative example and Example 3 is that zinc chloride is replaced with sodium chloride.

[0097] The results are as follows Figure 8 A 100 μL 50 mg / mL sodium chloride aqueous solution could not induce the Pickering emulsion to form Pickering emulsion gel K.

[0098] Example 6

[0099] Preparation of Pickering emulsion gel loaded with paeonol:

[0100] 1. Preparation of glycyrrhizic acid aqueous solution

[0101] Same as Example 1.

[0102] 2. Preparation of Paeonol Solution

[0103] Dissolve 0.10 g of paeonol in 5 mL of soybean oil to obtain a 2.0 wt% paeonol oil solution (the paeonol content is the mass of paeonol contained in the oil phase).

[0104] 3. Preparation of Pickering emulsion loaded with paeonol

[0105] Add 5 mL of glycyrrhizic acid aqueous solution to a 20 mL sample bottle, slowly add 5 mL of 2.0 wt% paeonol oil solution, and disperse at 24000 rpm for 2 min to obtain a glycyrrhizic acid-stable O / W type internal phase (φ = 50%) Pickering emulsion (named PE2).

[0106] 4. Preparation of Pickering emulsion gel loaded with paeonol

[0107] Pickering emulsion (PE2) was mixed with 100 μL of 50 mg / mL lysozyme aqueous solution and allowed to stand for 10 minutes to obtain Pickering emulsion gel L loaded with paeonol.

[0108] Performance Test Example 1: DPPH Free Radical Scavenging Ability Test

[0109] 1. Preparation of DPPH solution

[0110] 1.2 mg DPPH (1,1-diphenyl-2-trinitrophenylhydrazine) was dissolved in 30 mL of anhydrous ethanol to prepare a 0.1 mM DPPH solution.

[0111] 2. Testing Methods

[0112] Experimental groups: PBS group (negative control group), hydrogen peroxide (H2O2) group (positive control group), Pickering emulsion (PE1) group, Pickering emulsion gel B group, and Pickering emulsion gel L group.

[0113] 10 mg, 50 mg, and 100 mg of each group of samples were placed in 3 mL of DPPH solution and stirred in the dark for 30 min. The absorbance of the DPPH solution at 534 nm was then measured by UV-Vis spectrophotometry. The antioxidant capacity of the samples was evaluated by calculating the scavenging rate, using the following formula:

[0114]

[0115] in, A 0 represents the absorbance of a DPPH-containing solution only;

[0116] A 1 represents the absorbance of the sample group;

[0117] 3. Test Results

[0118] Figure 9Results of free radical scavenging by Pickering emulsion (PE1), Pickering emulsion gel B, and Pickering emulsion gel L; (a) full-wavelength scan (with PBS as negative control and H2O2 as positive control); (b) DPPH scavenging efficiency (%) of sample amounts at different concentrations. Figure 9 In the figure, the curves corresponding to PBS and H2O2 partially overlap.

[0119] like Figure 9 As shown in (a), the characteristic peak of DPPH at 517 nm was detected by UV-Vis full-band scanning. Compared with PBS, glycyrrhizic acid, being a natural antioxidant, can neutralize DPPH free radicals and has a certain free radical scavenging effect. Therefore, Pickering emulsion (PE1) and Pickering emulsion gel B containing glycyrrhizic acid have a certain scavenging effect. Paeonol contains many phenolic hydroxyl groups, and Pickering emulsion gel L containing paeonol has the strongest antioxidant effect. Furthermore, through… Figure 9 Figure (b) shows the DPPH scavenging effect of different sample concentrations. When only 10 mg of sample was added, the DPPH scavenging rates of Pickering emulsion (PE1), Pickering emulsion gel B, and Pickering emulsion gel L were 19.7%±0.5%, 21.1%±0.3%, and 33.7%±0.6%, respectively. When 50 mg of sample was added, the DPPH scavenging rates of Pickering emulsion (PE1), Pickering emulsion gel B, and Pickering emulsion gel L reached 26.6%±0.6%, 31.6%±0.5%, and 46.2%±0.8%, respectively. When 100 mg of sample was added, the scavenging effect further increased, with the DPPH scavenging rates of Pickering emulsion (PE1), Pickering emulsion gel B, and Pickering emulsion gel L reaching 64.7%±0.4%, 65.7%±0.2%, and 74.0%±0.8%, respectively. The data above shows that the addition of paeonol significantly improves the free radical scavenging effect compared to glycyrrhizic acid.

[0120] Performance Test Example 2: Cytotoxicity Test

[0121] Cell line: Rat skin fibroblast RS1 (Suzhou Haixing Biotechnology)

[0122] Experimental groups: PBS group (negative control group), hydrogen peroxide (H2O2) group (positive control group), Pickering emulsion (PE1) group, Pickering emulsion gel B group, and Pickering emulsion gel I group.

[0123] Sample extract preparation: 100 uL of each group of samples were soaked in 37℃ and 900 uL of DMEM medium for 24 hours to obtain the corresponding extracts. The extracts were then filtered through a 0.22 μm filter membrane for sterilization and set aside for later use.

[0124] Blank group: containing only DMEM medium.

[0125] 1. Quantitative detection of cell viability using the CCK-8 assay

[0126] RS 1 cells were loaded at 5 × 10⁻⁶ 4 Cells were seeded at a density of 10 cells / well in 96-well plates and cultured at 37°C with 5% CO2 for 24 hours (until cells adhered). The supernatant was discarded, and 100 μL of different extraction media (with an equal volume of DMEM added to the blank control group) was added to each well. Cells were cultured for another 24 hours. Then, 10 μL of CCK-8 reagent was added to each well, and after incubation for 2 hours, the absorbance (OD value) at 450 nm was measured using a microplate reader, and cell viability was calculated. Results are shown below. Figure 10 .

[0127]

[0128] in: As =Absorbance of experimental wells (including cells, culture medium, CCK-8 solution and analyte);

[0129] Ac =Absorbance of control wells (containing cells, culture medium, and CCK-8 solution, but excluding the analyte);

[0130] Ab =Absorbance of blank wells (including culture medium and CCK-8 solution, but excluding cells and analyte).

[0131] Result: From Figure 10 As can be seen, the cell viability of Pickering emulsion (PE1) and Pickering emulsion gels B and L is within the concentration range of 1~500 mg / mL, and the cell viability of RS 1 cells is greater than 95%, which proves that the above materials have low toxicity. Figure 10 In this study, the PBS group served as the negative control group, while the H2O2 group served as the positive control group.

[0132] 2. Fluorescent staining method (qualitative observation of cell morphology)

[0133] Cell line: Rat skin fibroblasts (RS1) (Suzhou Haixing Biotechnology)

[0134] RS 1 cells at 5 × 10 5Cells were seeded per well in a 24-well plate and cultured for 24 hours. The culture medium was discarded, and 500 μL of each group's extract (sample concentration of 100 mg / mL) was added to each well. The cells were then cultured for another 24 hours. Afterward, the cells were stained with the Calcein / PI cell viability and cytotoxicity assay kit and incubated at 37°C for 30 minutes. The cells were then washed three times with PBS. Live cells (green fluorescence) and dead cells (red fluorescence) were observed and photographed under an inverted fluorescence microscope.

[0135] from Figure 11 It can be seen that after the Pickering emulsion (PE1) with 100 mg / mL, Pickering emulsion gel B and Pickering emulsion gel L were treated with RS 1, the cells showed better morphology. Among them, the PBS group was the negative control group and the H2O2 group was the positive control group.

[0136] Performance Test Example 3: Validation of Intracellular ROS Scavenging Capacity

[0137] Cell line: Rat skin fibroblasts (RS1) (Suzhou Haixing Biotechnology)

[0138] Experimental groups: PBS group (negative control group), hydrogen peroxide (H2O2) group (positive control group), Pickering emulsion (PE1) group, Pickering emulsion gel B group, and Pickering emulsion gel L group.

[0139] Sample extract preparation: 100 uL of each group of samples were soaked in 37℃ and 900 uL of DMEM medium for 24 hours to obtain the corresponding extracts. The extracts were then filtered through a 0.22 μm filter membrane for sterilization and set aside for later use.

[0140] RS1 cells were used at 5 × 10 4 Seeds were inoculated per well in confocal culture dishes and incubated for 12 hours (adherence). The culture medium was discarded, and samples from different groups were added and incubated for 6 hours. The DCFH-DA probe (CAS No.: 4091-99-0, Solarbio: D6470-25mg, final concentration 10 μM) was added, and the sample was incubated at 37°C for 30 minutes. The sample was washed three times with PBS. The green fluorescence intensity (ROS level) was observed and photographed using a confocal laser scanning microscope (Nikon A1).

[0141] Result: From Figure 12It can be seen that, compared with the hydrogen peroxide (H2O2) group, both 100 mg / mL Pickering emulsion (PE1) and Pickering emulsion gel B still exhibited significant green fluorescence after ROS removal, indicating that their ROS removal effects were generally limited. However, 100 mg / mL Pickering emulsion gel L showed no obvious green fluorescence in cells after ROS removal, demonstrating its superior ROS removal effect.

[0142] Performance Test Example 4: Verification of Diabetic Wound Healing Ability

[0143] Pancreatic injury was induced in healthy rats (Vitallix) using streptozotocin (CAS No.: 18883-66-4, Aladdin: S766968-1g, STZ). One week later, blood glucose levels were measured using a glucometer; a blood glucose concentration greater than 16.7 mol / L was considered a successful establishment of a diabetic rat model. Then, circular holes of the same size (8 mm) were made on the backs of the diabetic rats, and PBS, Pickering emulsion PE1, Pickering emulsion gel B, and Pickering emulsion gel L were placed on them. Wound healing was observed after 3, 7, and 14 days. After the wounds had largely healed, tissue samples from different wound sites were taken for immunological analysis.

[0144] Figure 13 The effects of Pickering emulsion (PE1) and Pickering emulsion gels B and L on the repair of diabetic wounds were studied; the PBS group served as the negative control group; (a) a schematic diagram of the rat diabetic model and wound healing process; (b) representative photographs of the rat diabetic wound healing process after different sample treatments. n = 3), representing days 0, 3, 7, and 14 respectively; (c) Quantitative analysis of wound area in each group of diabetic patients ( n = 3); (d) H&E staining of the wound area on day 7 and day 14 ( n = 3, scale bar is 100 μm) and Masson staining of the wound area on day 14 ( n = 3, scale bar is 100 μm); from Figure 13 As can be seen, there are significant differences in the wound repair effects of PBS, Pickering emulsion PE1, Pickering emulsion gel B, and Pickering emulsion gel L on diabetic rats. After 14 days, the wound areas were 11.81, 10.28, 9.80, and 6.44%, respectively. Therefore, it can be seen that Pickering emulsion gel L has a better wound repair effect on diabetic rats.

[0145] The specific embodiments of the present invention have been described above. It should be understood that the present invention is not limited to the specific embodiments described above, and those skilled in the art can make various modifications or variations within the scope of the claims, which do not affect the essence of the present invention.

Claims

1. A mid-internal phase Pickering emulsion gel, characterized in that, Includes glycyrrhizic acid, lysozyme and / or polylysine, oil phase and water; The glycyrrhizic acid, calculated as available glycyrrhizic acid, constitutes 0.7-1.5 wt% of the emulsion gel. The lysozyme and / or polylysine constitute 0.1-0.5 wt% of the emulsion gel. The volume fraction of the oil phase is 30%-70%; The mid-internal phase Pickering emulsion gel was prepared by a method comprising the following steps: 1) Preparation of aqueous solution: Dissolve glycyrrhizic acid in water, heat to 70-90℃ and maintain for 20-40 minutes to form a transparent aqueous solution; 2) Pickering emulsion preparation: The oil phase is added to the aqueous solution in step 1), and emulsified at high speed of 20,000-25,000 rpm for 1-3 minutes at room temperature to obtain the Pickering emulsion; 3) Emulsion gel construction: Add lysozyme and / or polylysine aqueous solution to the Pickering emulsion in step 2), and let it stand at room temperature for 5-30 minutes to induce gelation to obtain the mid-internal phase Pickering emulsion gel.

2. The Pickering emulsion gel according to claim 1, characterized in that, The oil phase is selected from at least one of soybean oil, corn oil, sunflower oil, and olive oil.

3. The Pickering emulsion gel according to claim 1, characterized in that, The oil phase also contains paeonol, and the weight percentage of paeonol is 0.5-5 wt% based on the total weight of the oil phase.

4. The Pickering emulsion gel according to claim 1, characterized in that, The glycyrrhizic acid in the emulsion gel is 0.7-1.0 wt% based on available glycyrrhizic acid.

5. The Pickering emulsion gel according to claim 1, characterized in that, The lysozyme and / or polylysine constitute 0.1-0.2 wt% of the emulsion gel.

6. The Pickering emulsion gel according to claim 1, characterized in that, The volume fraction of the oil phase is 40-50%.

7. The Pickering emulsion gel according to claim 1, characterized in that, In step 3), the concentration of the lysozyme and / or polylysine aqueous solution is 40-60 mg / mL, and the added volume is such that the weight percentage of lysozyme and / or polylysine in the emulsion gel is 0.1-0.5 wt%.

8. The use of the mid-internal phase Pickering emulsion gel as described in any one of claims 1-7 in the preparation of drug delivery systems, wound dressings, tissue engineering scaffolds, topical skin preparations or cosmetics.

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