Nano-micelle for embedding curcumin as well as preparation method and application of nano-micelle

Abstract

The invention belongs to the technical field of preparation of drug carriers, and particularly provides a nano-micelle for embedding curcumin as well as a preparation method and application of the nano-micelle. The preparation method of the nano-micelle for embedding curcumin comprises the following steps: dissolving shellac, PGPR, beta-sitosterol succinate and curcumin in liquid coconut oil, and adding ethanol to form an organic phase; and after mixing with a xanthan gum aqueous solution, carrying out pre-emulsification, fine emulsification, gradient cooling and cold storage curing to obtain the nano-micelle. The encapsulation efficiency of the nano-micelle is greater than or equal to 93%, the drug loading capacity reaches 1.58%-4.15%, and the nano-micelle has efficient pH response targeting property; wherein the beta-sitosterol succinate enhances the drug-loading core density, the shellac constructs a gastric juice barrier, and the PGPR maintains the micelle dispersibility, so that the oral bioavailability of the curcumin can be improved by 46-55 times compared with that of free curcumin, and the curcumin is suitable for efficient delivery of food functional components or drugs.

Description

Technical Field

[0001] This invention belongs to the field of drug carrier preparation technology, specifically providing a nanomicelle for encapsulating curcumin, its preparation method and its application. Background Technology

[0002] Curcumin, an extract from ginger plants, possesses well-defined biological activities such as antioxidant, liver-protective, and anti-inflammatory properties, and has been widely studied for its use as a functional food ingredient and active pharmaceutical ingredient. However, curcumin faces key limitations, including extremely poor water solubility, low oral bioavailability, and easy degradation in physiological environments. Its degradation rate can reach 90% within 30 minutes in a neutral environment at pH 7.2, and the peak blood concentration of free curcumin after oral administration is less than 1 / 30th that of its nanocarrier form, severely restricting its industrial application.

[0003] To address the aforementioned issues, nanomicelles have become a research hotspot due to their advantages of strong drug-carrying capacity in a hydrophobic core and improved pharmacokinetic properties in a hydrophilic shell. Among existing technologies, patent application CN109820815A discloses a pH-responsive curcumin micelle hydrogel, which constructs a carrier by combining a surfactant with gellan gum. However, this method suffers from limitations in solubilizing hydrophobic drugs and a drug loading capacity of only 60.9-65.8 mg / g. Other technologies employ polyethylene glycol (PEG) to modify curcumin to prepare prodrug nanomicelles. For example, patent application CN112336234A discloses a method for preparing PEG-modified curcumin prodrug nanomicelles. While this improves stability, it requires a chemical synthesis step and carries the risk of nanoparticle aggregation. Furthermore, single carrier materials such as pure shellac (CN108743156A) exhibit poor stability and are prone to brittle fracture.

[0004] This invention addresses the shortcomings of existing technologies, such as low drug loading efficiency, complex preparation processes, and insufficient intestinal targeting. It employs a composite carrier of shellac, PGPR, and β-sitosterol succinate, leveraging the synergistic effect of the three components—shellac providing pH responsiveness, PGPR maintaining micelle flexibility, and β-sitosterol succinate enhancing drug loading core density—to construct a highly efficient targeted drug delivery system with a simple process. Summary of the Invention

[0005] To address the core bottlenecks of curcumin's poor water solubility, low oral bioavailability, and easy degradation in the physiological environment, as well as the shortcomings of existing nanocarrier technologies such as low drug loading efficiency, complex preparation processes, insufficient intestinal targeting, and easy aggregation and brittleness of micelles, this invention aims to provide a curcumin nanomicelle with high encapsulation efficiency, high intestinal targeting, simple preparation process, and good biocompatibility, along with its corresponding preparation method and applications.

[0006] To achieve the above objectives, the present invention adopts the following technical solution: A nanomicelle for encapsulating curcumin is prepared mainly from shellac, polyglycerol ricinoleate (PGPR), and β-sitosterol succinate.

[0007] Preferably, in the nanomicelles described above, the shellac, polyglycerol ricinoleate, and β-sitosterol succinate are in a mass ratio of 2~8:2~5:1~3.

[0008] A nanomicelle for encapsulating curcumin uses shellac, polyglycerol ricinoleate (PGPR), and β-sitosterol succinate as the core composite carrier to load curcumin as the active ingredient.

[0009] Preferably, in the nanomicelles described above, the shellac, PGPR, and β-sitosterol succinate are in a mass ratio of 2~8:2~5:1~3, and the total carrier mass is 5~16 parts; the mass ratio of curcumin to the total carrier is 1:2.5~8.

[0010] The key performance indicators of the nanomicelles used to encapsulate curcumin, as described above, are: average particle size 56~80nm, PDI≤0.20, Zeta potential -25~-30mV, encapsulation efficiency ≥93%, drug loading 1.58%~4.15%, release rate in simulated intestinal fluid ≥95%, and encapsulation retention rate ≥84% after 28 days of storage at 40℃.

[0011] A method for preparing nanomicelles for encapsulating curcumin, comprising the following steps: S1. Take shellac, PGPR, β-sitosterol succinate, and curcumin and dissolve them together in liquid coconut oil at 55~70℃, and add food-grade ethanol to obtain a homogeneous and transparent organic phase mixture. S2. Add xanthan gum aqueous solution to the mixture in step S1, and adjust the pH to acidic using citric acid aqueous solution; S3. Homogenize the material obtained in step S2: After preheating, pre-emulsify first, then finely emulsify; S4. Gradient cooling: First, cool to the target temperature at a set rate and hold at that temperature, then continue cooling to the refrigeration temperature to solidify and obtain nanomicelles.

[0012] In the preparation method described above, preferably, in step S1, the mass ratio of shellac, PGPR, and β-sitosterol succinate is 2~8:2~5:1~3.

[0013] In the preparation method described above, preferably, in step S1, the temperature of the liquid coconut oil is 55~70℃, the amount of liquid coconut oil used is 5~10 times the total mass of shellac, PGPR, and β-sitosterol succinate; the amount of ethanol added is 10%~30% of the volume of liquid coconut oil, and the ethanol is 95% food-grade ethanol by volume.

[0014] In the preparation method described above, preferably, in step S2, the mass concentration of xanthan gum aqueous solution is 0.5~2 g / L, the amount of xanthan gum aqueous solution used is 0.5~1.5 times the volume of liquid coconut oil; the mass volume concentration of citric acid aqueous solution is 0.4~1%, and the pH of the system is adjusted to 3~4.

[0015] In the preparation method described above, preferably, in step S3, the mixing system is preheated to 65~75℃; the pre-emulsification speed is 8000~10000 rpm and the time is 1.5~3 min; the fine emulsification speed is 13000~18000 rpm and the time is 5~10 min, and the system temperature does not exceed 75℃ during the emulsification process.

[0016] In the preparation method described above, preferably, in step S4, the gradient cooling specifically involves: first cooling to 35~40℃ at a cooling rate and holding at that temperature, then cooling to 0~8℃ at a cooling rate and then refrigerating for solidification.

[0017] Furthermore, first lower the temperature by 5℃ / min to 35~40℃ and hold for 3~5 minutes, then lower the temperature by 2℃ / min to 0~8℃ and refrigerate for 3~4 hours to solidify.

[0018] The nanomicelles and the above preparation method can be used to prepare food functional preparations (such as nutritional supplements) containing curcumin or other functional agents or for use in the preparation of drugs. In specific applications, curcumin or other functional agents or drugs can be directly added to the organic phase liquid coconut oil along with the composite carrier simultaneously without additional pretreatment, making it suitable for industrial mass production.

[0019] The beneficial effects of this invention are as follows: The nano micelles provided by this invention have strong stability. The added PGPR maintains the flexibility of the micelles and reduces the risk of aggregation and brittleness during storage. The encapsulation retention rate is ≥91% after 28 days of storage at 4°C, which is better than the existing pure shellac carrier (retention rate ≤85%).

[0020] The nanomicelles provided by this invention have excellent drug loading and targeting performance. The added β-sitosterol succinate enhances the hydrophobic core density, improving the encapsulation efficiency and drug loading (15%~20% higher than that of a single shellac carrier). The shellac imparts pH-responsive properties, resulting in a release rate of ≤4% in gastric juice (0-2h) and ≥95% in intestinal juice (2-10h), achieving precise intestinal targeting.

[0021] The nanomicelles provided by this invention have high bioavailability. Oral administration experiments in mice showed that the AUC0-12h of curcumin in the nanomicelle group was 46-55 times that of the free curcumin group, and the peak blood concentration was increased by 28-33 times. It is suitable for the efficient delivery of functional ingredients in food or drugs.

[0022] The nanomicelle process provided by this invention is both safe and efficient: there are no chemical synthesis steps involved, all raw materials used (shellac, PGPR, xanthan gum, etc.) are food-grade, the preparation cycle is short (≤8h), and it is suitable for large-scale production, thus solving the problems of complex and poor biocompatibility of existing prodrug nanomicelle processes. Detailed Implementation

[0023] This invention uses shellac, polyglycerol ricinoleate (PGPR), and β-sitosterol succinate as building blocks to construct a nanomicelle system with targeted delivery characteristics, high structural stability, and drug loading efficiency through the synergistic mechanism of functional groups. This effectively overcomes the application bottlenecks of drugs such as curcumin, which have poor water solubility and low bioavailability.

[0024] This invention elucidates the molecular mechanism of action. Shellac, as a pH-sensitive polymer, undergoes protonation in the acidic environment of the stomach (pH 1.5-3.5) to form a dense cross-linked shell, inhibiting the early release of drugs such as curcumin. In the alkaline environment of the intestine (pH 6.8-7.4), it deprotonates and dissociates, triggering controlled drug release. PGPR molecules, with their unique hydrophilic-lipophilic amphiphilic structure, embed themselves in the micelle interface layer through hydrophobic interactions, regulating micelle membrane fluidity and effectively inhibiting interparticle aggregation. β-sitosterol succinate, relying on a steroidal framework, forms a stable hydrophobic core, enhancing the loading efficiency of drugs such as curcumin through π-π stacking and van der Waals forces, while simultaneously strengthening the thermodynamic stability of the micelle system.

[0025] The mass ratios of the components exhibit a significant synergistic effect: an excessively high proportion of shellac leads to a decrease in shell mechanical strength, affecting drug release kinetics; exceeding the critical value of PGPR reduces drug loading efficiency and interferes with pH response performance; and excessive use of β-sitosterol succinate causes deterioration in nanoparticle dispersibility. Response surface methodology optimization experiments verified that when the mass ratio of the three components is controlled within the range of 2–8:2–5:1–3, an optimal balance between targeted delivery efficiency, structural stability, and drug loading is achieved, meeting the core performance parameter requirements for preclinical applications.

[0026] The following embodiments are used to further illustrate the present invention, but should not be construed as limiting the present invention. Any modifications or substitutions made to the present invention without departing from its spirit and essence are within the scope of the present invention.

[0027] Unless otherwise specified, the technical means used in the embodiments are conventional means well known to those skilled in the art, and unless otherwise specified, the raw materials used in the embodiments may be commercially available products.

[0028] Example 1: Preparation of BPS-1 nanomicelles 1. Raw material ratio: Shellac:PGPR:β-sitosterol succinate = 8:5:3, with a total carrier mass of 16g. Specifically, the following ingredients are used: 2g curcumin (purity ≥98%, available from Shanghai Yuanye Biotechnology Co., Ltd.), 8g shellac (bleached and refined shellac, available from Anning Daike Biotechnology Co., Ltd.), 5g polyglycerol ricinoleate (PGPR) (available from Danisco (China) Co., Ltd.), 3g β-sitosterol succinate (β-sitosterol purity ≥98%, available from Shanghai Yuanye Biotechnology Co., Ltd.), 80g liquid coconut oil (melting point 24-27℃, available from Hainan Coconut Island Group Co., Ltd.), 16g 95% (v / v) food-grade ethanol (available from Sinopharm Chemical Reagent Co., Ltd.), 100mg xanthan gum (available from Shandong Fufeng Group Co., Ltd.), 60mg citric acid (available from Anhui Fengyuan Group Co., Ltd.), and 80mL purified water (available from Hangzhou Wahaha Group Co., Ltd.).

[0029] 2. Preparation steps S1. Organic phase mixing: Add 8g shellac, 5g PGPR, 3g β-sitosterol succinate, and 2g curcumin to 80g of liquid coconut oil preheated to 60℃, then add 16g of 95% (v / v) food-grade ethanol. Place the mixture on a magnetic stirrer and stir at 1000rpm for 10min until all solid components are completely dissolved, forming a uniform and transparent organic phase mixture. S2. Aqueous phase adjustment: Prepare a 1.25 g / L xanthan gum aqueous solution using 80 mL of purified water. Stir the solution on a magnetic stirrer at 700 rpm for 12 min until the xanthan gum is completely dissolved. Add the entire xanthan gum aqueous solution to the organic phase mixture prepared in step S1 and continue stirring at 1000 rpm for 6 min to allow the system to mix initially. Then, prepare 10 mL of 0.6% (w / v, g / mL) citric acid aqueous solution and slowly add it dropwise to the above mixture using a pipette while stirring. Measure the pH of the system with a pH meter after each 1 mL drop is added until the pH of the system stabilizes at 3. S3. Homogenization: Transfer the mixture obtained in step S2 to the dispersion cup of a high-speed homogenizer. Place the dispersion cup in a 65°C constant temperature water bath for preheating. When the system temperature reaches 70°C, perform pre-emulsification at 8500 rpm for 2 minutes. During this period, monitor the system temperature in real time with a thermometer to ensure that the temperature does not exceed 75°C. After the pre-emulsification is completed, immediately increase the speed to 15000 rpm for fine emulsification for 8 minutes. Keep the water bath temperature stable during the process, and finally form a milky white homogeneous emulsion. S4. Gradient cooling molding: The finely emulsified emulsion is rapidly transferred to a constant temperature cooling bath. The initial temperature of the cooling bath is set to 70℃, and the temperature is reduced to 38℃ at a cooling rate of 5℃ / min. The temperature is maintained at this temperature for 5 minutes. Then, the cooling rate is adjusted to 2℃ / min, and the temperature is further reduced to 4℃. The mixture is then refrigerated and solidified at this temperature for 3 hours. The system status is observed every hour during this period to ensure that there is no stratification or precipitation. The final product is a milky white, non-stratified curcumin nanomicelles, denoted as BPS-1. If long-term storage is required, the product can be transferred to a suitable container, 8g of mannitol (food grade, Shandong Lukang Pharmaceutical Co., Ltd.) can be added as a protective agent, and the container can be pre-frozen in an ultra-low temperature freezer at -80℃ for 4 hours. Then, the container can be transferred to a vacuum freeze dryer and freeze-dried for 24 hours under conditions of vacuum degree ≤10Pa and cold trap temperature ≤-50℃ to obtain BPS-1 curcumin nanomicelle freeze-dried powder.

[0030] 3. Performance test results: Particle size and Zeta potential: Experimental method: Take 0.5 mL of BPS-1 nanomicelle product, dilute it with pure water to 10 mL (ensure the concentration is within the detection range of the dynamic light scattering instrument), place it in an ultrasonic instrument for ultrasonic dispersion for 1 min, and then inject the diluted solution into the instrument's detection cell; The measurement parameters are set as follows: temperature 25℃, scattering angle 90°, equilibration time 2 min, and three parallel measurements are performed, and the average value ± standard deviation is taken; Results: The average particle size of BPS-1 nanomicelles is 65±2 nm, the PDI (polydispersity index) is 0.18±0.01, and the Zeta potential is -30±2 mV.

[0031] Encapsulation efficiency and drug loading: Experimental methods: ① Encapsulation efficiency determination: Take 1 mL of the finished product and put it into a dialysis bag with a molecular weight cutoff of 10 kDa. Dialyze it in 50 mL of purified water for 24 h (replace the dialysate every 6 h and combine all dialysate). The concentration of curcumin in the dialysate was determined by high performance liquid chromatography (HPLC), and the total amount of free curcumin was calculated. Chromatographic conditions: C18 column (4.6 × 250 mm, 5 μm); mobile phase: acetonitrile-0.1% phosphoric acid aqueous solution (60:40, v / v); flow rate: 1.0 mL / min; detection wavelength: 425 nm; column temperature: 30 ℃; injection volume: 20 μL. Calculation formula: Encapsulation efficiency (%) = [(Total amount of curcumin added - Free curcumin amount) / Total amount of curcumin added] × 100% (Note: Free curcumin amount is the total mass of curcumin measured in the dialysis fluid) ② Drug loading determination: Take 1 mL of the finished product and accurately weigh its total mass; the mass of encapsulated curcumin is calculated according to "Total amount of curcumin added - Free curcumin amount" (the free curcumin amount is the same as the encapsulation efficiency determination result). Calculation formula: Drug loading (%) = [(Total amount of curcumin added - Free curcumin amount) / Total mass of finished product] × 100%; Results: The encapsulation efficiency of BPS-1 nanomicelles is 95.1±0.6%, and the drug loading is 2.28±0.08%.

[0032] Storage stability: Experimental method: 10 mL of BPS-1 nanomicelles were aliquoted into 3 groups of sterile centrifuge tubes (3 replicates per group), labeled as 4℃ group, 25℃ group, and 40℃ group, respectively. The 4℃ group was placed in a constant temperature refrigerator (RH 50%, humidity controlled by saturated magnesium nitrate solution), the 25℃ group was placed in a constant temperature and humidity chamber (RH 60%, controlled by saturated magnesium chloride solution), and the 40℃ group was placed in a high temperature and high humidity chamber (RH 75%, controlled by saturated sodium chloride solution). Samples were taken at 0, 7, 14, and 28 days, with 0.5 mL of sample taken each time. Particle size and PDI were measured according to the "Particle Size and Zeta Potential Determination Method", and encapsulation efficiency was measured according to the "Encapsulation Efficiency Determination Method". The encapsulation efficiency retention rate was calculated (retention rate % = encapsulation efficiency at each time point / encapsulation efficiency on day 0 × 100%). The presence of stratification, precipitation, and color changes in the samples were recorded daily. The results are shown in Table 1.

[0033] Table 1. Storage stability results of BPS-1 at different temperatures. Simulated gastric-intestinal fluid stepwise release experiment: Experimental method: ① Sample preparation: Accurately weigh 0.5g of BPS-1 nanomicelles, place them in a centrifuge tube, add 5mL of sterile physiological saline (0.9% NaCl solution, matching the osmotic pressure of the physiological environment), and sonicate for 2min to prepare a homogeneous micelle suspension; ② Dialysis bag pretreatment: Boil dialysis bags with a molecular weight cutoff of 10kDa in physiological saline for 10min to remove surface impurities and activate the membrane pore size; ③ Gastric fluid stage release: Transfer all of the above micelle suspension into the pretreated dialysis bag, seal the bag opening with a rope (ensuring no leakage), and place the dialysis bag in a solution containing 5% saline. 0 mL of pH 1.2 simulated gastric juice (containing 0.1 mol / L HCl and 0.5% pepsin, preheated to constant temperature in a 37℃ water bath) was placed in a stoppered conical flask and incubated at 100 rpm in a 37℃ water bath shaker. At 0.5, 1, 1.5, and 2 hours of incubation, 1 mL of the dialysate was precisely pipetted (simultaneously, 1 mL of fresh pH 1.2 simulated gastric juice preheated to 37℃ was added via pipette to maintain a constant volume of the dialysate). The dialysate was filtered through a 0.22 μm organic phase filter membrane, and the concentration of free curcumin was determined using the above-described high-performance liquid chromatography method. Calculate the total amount of free curcumin in the dialysis fluid at each time point; ④ Intestinal fluid stage release: After 2 hours of incubation in the gastric fluid stage, remove the dialysis bag and quickly rinse the outer surface of the bag with sterile saline (to remove residual simulated gastric fluid). Immediately transfer the dialysis bag into a stoppered conical flask containing 50 mL of simulated intestinal fluid (pH=6.8, containing 0.1 mol / L PBS and 1% trypsin, preheated to 37°C) and continue incubation at 37°C and 100 rpm; every 1 hour, aspirate 1 mL of dialysis fluid and replenish with an equal amount of fresh simulated intestinal fluid, and determine the concentration and total amount of free curcumin; ⑤ Release rate calculation: according to the formula, cumulative release rate = (at each time point) / (at each time point) The total release rate was calculated as (total free curcumin) / (total mass of curcumin embedded in micelles) × 100%, where the total mass of curcumin embedded in micelles was estimated from "0.5g finished product mass × drug loading". Results: The cumulative release rate in the gastric juice stage (0-2h) was 3.5±0.5% (total free curcumin at 2h was 0.00017±0.00003mg); the cumulative release rate in the intestinal juice stage (2-10h) was 96.2±1.2% (total free curcumin at 10h was 0.0047±0.00028mg), showing an overall highly efficient pH-responsive targeting characteristic with extremely low leakage in gastric juice and near-complete release in intestinal juice.

[0034] Antioxidant capacity (DPPH method): Experimental Methods: ① Reagent Preparation: Accurately weigh 12.1 mg of DPPH powder with a purity ≥98%, dissolve it in anhydrous ethanol, and bring the volume to 50 mL to prepare a 0.1 mmol / L DPPH stock solution. Store the solution under light-protected conditions (shelf life 3 days). Take the BPS-1 nanomicelle product, dilute it with 0.1 mol / L, pH 7.4 phosphate buffered saline (PBS), determine the curcumin concentration by high performance liquid chromatography (HPLC), and adjust it to the target concentration of 50 μmol / L. ② Reaction System Construction: Transfer 1 mL of the diluted sample and add 1 mL of DPPH stock solution. Simultaneously set up a blank group (1 mL PBS + 1 mL DPPH stock solution) and a control group (1 mL anhydrous ethanol + 1 mL DPPH stock solution). Vortex mix for 10 s, then incubate at 37℃ in a light-protected water bath for 30 min. ③ Absorbance measurement: Using a UV-Vis spectrophotometer, the absorbance of the sample group and the control group was measured at a wavelength of 517 nm with the blank solution as the zero point, and recorded as A1 and A0 respectively. ④ Scavenging rate calculation: The scavenging rate % was calculated according to the formula: Scavenging rate % = [1 - (A - A0) / A1] * %, and each sample was measured in triplicate, and the arithmetic mean was taken.

[0035] Results: The DPPH radical scavenging rate of the BPS-1 nanomicelle system was determined to be 91.5±1.4%. Compared with the free curcumin group at the same concentration of 50 μmol / L (scavenging rate 67.3±1.8%), the difference was statistically significant (p<0.05).

[0036] Oral pharmacokinetics in mice: Experimental methods: ① Animal grouping: 20 SPF-grade Balb / c mice weighing 20±2g (purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd.) were randomly divided into 2 groups of 10 mice each, namely the BPS-1 nanomicelle group and the free curcumin group. ② Drug administration preparation: BPS-1 nanomicelle group: The known drug loading of BPS-1 nanomicelles is 2.28%. A curcumin dosage of 50 mg / kg means that each kg of mouse needs to ingest 50 mg of curcumin active ingredient. Taking a 20g (0.02kg) mouse as an example, a single mouse needs to ingest 1 mg of curcumin; the required mass of the finished micelle product is 43.86 mg. In actual operation, take the finished BPS-1 nanomicelle product and dilute it with sterile physiological saline to an appropriate concentration to ensure that the curcumin intake calculated based on the mouse's body weight is 50 mg / kg; Free curcumin group: Take curcumin powder and suspend it in 0.5% CMC-Na solution; ③ Blood sample collection: Both groups of mice were administered the drug orally via gavage. At 0.25, 0.5, 1, 2, 4, 8 and 12 h after administration, 0.5 mL of blood was collected by orbital blood sampling. The blood was placed in a centrifuge tube containing heparin sodium and centrifuged at 3000 rpm for 10 min. The supernatant plasma was separated and stored at -80℃ for later testing. ④ Plasma sample processing: Take 0.2 mL of frozen plasma, add 0.6 mL of ethyl acetate, vortex mix for 3 min, then centrifuge at 12000 rpm for 5 min, collect the upper organic phase, and dry it with nitrogen in a nitrogen blower; add 0.2 mL of mobile phase (acetonitrile-0.1% phosphoric acid water = 60:40, v / v) to the dried residue, vortex to dissolve, filter through a 0.22 μm organic phase filter membrane, and collect the filtrate; ⑤ Detection and parameter calculation: The concentration of curcumin in the filtrate was determined by HPLC-MS / MS, and the pharmacokinetic parameters were calculated by pharmacokinetic software based on the blood drug concentration data at each time point. Results: The peak plasma concentration (Cmax) of mice in the BPS-1 nanomicelle group was 285±22 ng / mL, the area under the curve (AUC0-12h) was 1250±35 ng・h / mL, and the time to peak concentration (Tmax) was 0.5±0.1h. The Cmax of mice in the free curcumin group was 9.2±1.5 ng / mL, and the AUC0-12h was 24.1±1.8 ng・h / mL. The AUC of the BPS-1 nanomicelle group was 52±4 times that of the free curcumin group.

[0037] Example 2: Preparation of BPS-2 nanomicelles 1. Raw material ratio: Shellac:PGPR:β-sitosterol succinate = 2:2:1 by mass, with a total carrier mass of 5g, including 2g curcumin, 2g shellac, 2g PGPR, 1g β-sitosterol succinate, 50g liquid coconut oil, 10g 95% (v / v) food grade ethanol, 50mg xanthan gum, 40mg citric acid, and 25mL purified water.

[0038] 2. Preparation steps: S1. Organic phase mixing: Add 2g shellac, 2g PGPR, 1g β-sitosterol succinate, and 2g curcumin to 50g of liquid coconut oil preheated to 55℃, add 10g of 95% (v / v) food-grade ethanol, place on a magnetic stirrer and stir at 1000rpm for 15min until all solid components are completely dissolved to form a uniform and transparent organic phase mixture; S2, Aqueous Phase Adjustment: Prepare a 0.5 g / L xanthan gum aqueous solution using 25 mL of purified water, and stir at 600 rpm for 10 min until the xanthan gum is completely dissolved; add this aqueous solution to the organic phase mixture from step S1, and continue stirring for 5 min; then prepare 10 mL of a 0.4% (w / v, g / mL) citric acid aqueous solution, and slowly add it dropwise to the mixture while stirring (300 rpm), measuring the pH after each drop of 1 mL is added, until the pH of the system stabilizes at 3; S3. Homogenization: Transfer the above mixture to a high-speed homogenizer dispersion cup, preheat to 65°C, pre-emulsify at 8000 rpm for 1.5 min, then increase the speed to 13000 rpm for 5 min to form a milky white homogeneous emulsion. S4. Gradient cooling molding: The emulsion is transferred to a constant temperature cooling bath with an initial temperature of 65℃. The temperature is then reduced to 35℃ at a rate of 5℃ / min and held for 5 min. The temperature is then reduced to 0℃ at a rate of 2℃ / min and refrigerated for 2 h to obtain the BPS-2 nano micelles.

[0039] 3. Performance test results: Particle size and Zeta potential: The experimental method was the same as in Example 1 (0.5 mL of the finished product was diluted to 10 mL, ultrasonically dispersed, and measured using a dynamic light scattering instrument, in triplicate); Results: The average particle size of BPS-2 nanomicelles was 78±3 nm, PDI was 0.19±0.02, and Zeta potential was -26±2 mV; Encapsulation efficiency and drug loading: The experimental method was the same as in Example 1 (dialysis for 24 hours, HPLC was used to measure the concentration of the dialysate, and the encapsulation efficiency and drug loading were calculated). Results: The encapsulation efficiency was 94.3±0.8%, and the drug loading was 4.15±0.10%; stability results are shown in Table 2.

[0040] Table 2. Storage stability results of BPS-2 at different temperatures. Temperature group Encapsulation rate (%) Encapsulation retention rate (%) Particle size (nm) PDI System status 4℃ 91.5±0.7 97.7 76±3 0.19±0.02 No layering, no sedimentation, and the color remains milky white. 25℃ 89.6±0.8 95.0 78±4 0.21±0.02 There was no stratification, but a small amount of flocculent precipitate appeared at the bottom, and the color remained milky white. 40℃ 84.9±0.9 90.0 82±4 0.24±0.03 There was no obvious layering, but a small amount of sediment appeared at the bottom, and the color turned milky yellow. Simulated gastric-intestinal fluid stepwise release experiment: The experimental method was the same as in Example 1 (sample volume was 0.5g, physiological saline was 5mL, dialysis fluid volume was 50mL, 1mL sample was taken at each time point and fluid was replenished, and the concentration of free curcumin was measured by HPLC). Results: The cumulative release rate in the gastric fluid stage (0-2h) was 3.8±0.6% (total free curcumin at 2h was 0.00037±0.00006mg), and the cumulative release rate in the intestinal fluid stage (2-10h) was 95.8±1.3% (total free curcumin at 10h was 0.0093±0.0005mg). Antioxidant capacity: DPPH scavenging rate 94.1±1.1%; Pharmacokinetics of mice after oral administration: The experimental method was the same as in Example 1 (20 mice were divided into groups, and the drug was administered at a dose of 50 mg / kg, and the blood drug concentration was measured at each time point); Results: The AUC0-12h of mice in the BPS-2 nanomicelle group was 1170±30 ng・h / mL, Cmax was 268±18 ng / mL, and Tmax was 0.5±0.1h; the AUC0-12h of the free curcumin group was 24.1±1.8 ng・h / mL, and the AUC of the BPS-2 group was 48±3 times that of the free group.

[0041] Storage stability: After reconstitution of the lyophilized powder at -20℃ for 3 months, the encapsulation rate was 93.6±0.7% (retention rate 99.3%).

[0042] Example 3: Preparation of BPS-3 nanomicelles 1. Raw material ratio: Shellac:PGPR:β-sitosterol succinate = 3:4:2 by mass, with a total carrier mass of 9g. Specifically, weigh out 2g curcumin, 3g shellac, 4g PGPR, 2g β-sitosterol succinate, 90g liquid coconut oil, 27g 95% (v / v) food grade ethanol, 180mg xanthan gum, 90mg citric acid, and 135mL purified water.

[0043] 2. Preparation steps: S1. Organic phase mixing: Add 3g shellac, 4g PGPR, 2g β-sitosterol succinate, and 2g curcumin to 90g of liquid coconut oil at 70℃, add 27g of 95% ethanol, and stir at 1000rpm for 20min until completely dissolved to form a transparent organic phase; S2. Aqueous phase adjustment: Prepare a 2 g / L xanthan gum aqueous solution with 135 mL of purified water, add it to the organic phase and stir for 8 min; prepare a 9 mL 1% (w / v, in g / mL) citric acid aqueous solution and add it dropwise to adjust the pH to 3; S3. Homogenization: The mixed system is preheated to 75°C, pre-emulsified at 10,000 rpm for 3 min, and finely emulsified at 18,000 rpm for 10 min to obtain an emulsion; S4. Gradient cooling molding: 75℃→40℃ (5℃ / min, hold for 5min)→4℃ (2℃ / min), refrigerate for 4h to obtain BPS-3 nano micelles; 3. Performance test results: Particle size and Zeta potential: Same as in Example 1; Results: Particle size 58±2nm, PDI 0.17±0.01, Zeta potential -28±1mV; Encapsulation efficiency and drug loading: same as in Example 1; Results: encapsulation efficiency 96.5±0.6%, drug loading 1.58±0.07%; storage stability results are shown in Table 3.

[0044] Table 3. Storage stability results of BPS-3 at different temperatures. Temperature group Encapsulation rate (%) Encapsulation retention rate (%) Particle size (nm) PDI System status 4℃ 94.6±0.5 98.0 57±2 0.17±0.01 No layering, no sedimentation, and a uniform milky white color. 25℃ 92.7±0.6 96.1 59±3 0.19±0.02 No stratification, no precipitation, slight opacity, and the color remains milky white. 40℃ 89.8±0.7 93.1 62±3 0.22±0.02 There was no stratification; the system was moderately turbid, with a small amount of flocculent precipitate at the bottom, and the color changed to light milky yellow. Simulated gastric-intestinal fluid stepwise release experiment: same as in Example 1; Results: The release rate in the gastric fluid stage (0-2h) was 3.2±0.4% (0.00012±0.00002mg of free curcumin at 2h), and the release rate in the intestinal fluid stage (2-10h) was 97.1±1.0% (0.0034±0.00024mg of total free curcumin at 10h), which was the highest release efficiency in the intestinal fluid stage among all groups; Antioxidant capacity: Scavenging rate was 82.5 ± 1.2%; Oral pharmacokinetics in mice: Same as in Example 1; Results: AUC0-12h was 1375±40 ng・h / mL, Cmax was 302±25 ng / mL, which was 55±4 times that of the free group (optimal effect group).

[0045] Example 4: Preparation of BPS-4 nanomicelles 1. Raw material ratio: Shellac:PGPR:β-sitosterol succinate = 5:3:2 by mass, with a total carrier mass of 10g. Weigh out 2g curcumin, 5g shellac, 3g PGPR, 2g β-sitosterol succinate, 60g liquid coconut oil, 6g 95% (v / v) food grade ethanol, 120mg xanthan gum, 60mg citric acid, and 60mL purified water.

[0046] 2. Preparation steps: S1. Organic phase mixture: Add 5g shellac, 3g PGPR, 2g β-sitosterol succinate, and 2g curcumin to 60g coconut oil at 65℃, add 6g ethanol, and stir at 1000rpm for 17min until dissolved. S2. Aqueous phase adjustment: Prepare a 2 g / L xanthan gum aqueous solution (120 mg xanthan gum, stirred at 750 rpm for 13 min) in 60 mL of water, add it to the organic phase and stir for 7 min; adjust the pH to 3 with 0.6% (w / v) citric acid aqueous solution. S3. Homogenization treatment: Pre-emulsify at 9000 rpm for 2.5 min at 68℃, then fine emulsify at 16000 rpm for 7 min; S4. Gradient cooling molding: 68℃→37℃ (5℃ / min, hold for 5min)→5℃ (2℃ / min), refrigerate for 3.5h to obtain BPS-4 nanomicelles.

[0047] Performance test results: Particle size and Zeta potential: Same as in Example 1; Results: Particle size 72±2nm, PDI 0.18±0.01, Zeta potential -27±2mV; Encapsulation efficiency and drug loading: same as in Example 1; Results: encapsulation efficiency 95.8±0.5%, drug loading 2.85±0.09%; storage stability results are shown in Table 4.

[0048] Table 4. Storage stability results of BPS-4 at different temperatures. Temperature group Encapsulation rate (%) Encapsulation retention rate (%) Particle size (nm) PDI System status 4℃ 94.9±0.6 99.1 71±2 0.18±0.01 No layering, no sedimentation, and the color remains milky white. 25℃ 92.9±0.7 97.0 73±3 0.20±0.02 No stratification, no sedimentation, slight turbidity, and no color change. 40℃ 88.6±0.8 92.5 76±3 0.23±0.02 There was no stratification; a small amount of sediment appeared at the bottom; the system was moderately turbid; and the color changed to milky white with a yellowish tinge. Simulated gastric-intestinal fluid stepwise release experiment: same as in Example 1; results: gastric fluid release rate 3.6±0.5% (2h free curcumin 0.00025±0.00003mg), intestinal fluid release rate 96.5±1.1% (10h total free curcumin 0.0065±0.00038mg); Antioxidant capacity: DPPH scavenging rate 92.8±1.3%; Oral pharmacokinetics in mice: Same as in Example 1; Results: AUC 0-12h 1225±32 ng・h / mL, which was 50±3 times that of the free group.

[0049] Example 5: Preparation of BPS-5 nanomicelles 1. Raw material ratio: Shellac:PGPR:β-sitosterol succinate = 4:4:1 by mass, with a total carrier mass of 9g. Weigh out 2g curcumin, 4g shellac, 4g PGPR, 1g β-sitosterol succinate, 70g liquid coconut oil, 14g 95% (v / v) food grade ethanol, 140mg xanthan gum, 70mg citric acid, and 70mL purified water.

[0050] 2. Preparation steps: S1. Organic phase mixture: Add 4g shellac, 4g PGPR, 1g β-sitosterol succinate, and 2g curcumin to 70g coconut oil at 62℃, add 14g ethanol, stir at 1000rpm for 16min until dissolved, and form a uniform and transparent organic phase mixture. S2. Aqueous phase adjustment: Prepare a 2 g / L xanthan gum aqueous solution in 70 mL of water (weigh 140 mg of xanthan gum and stir at 720 rpm for 14 min until completely dissolved). Add all of this aqueous solution to the organic phase mixture and continue stirring at 1000 rpm for 6 min. Prepare a 0.7% (w / v) citric acid aqueous solution (70 mg of citric acid dissolved in 10 mL of purified water) and slowly add it dropwise to the mixture while stirring (300 rpm). Measure the pH after each drop of 1 mL is added until the pH of the system stabilizes at 3. S3. Homogenization: Transfer the mixed system to the dispersion cup of a high-speed homogenizer, preheat to 72°C, pre-emulsify at 8800 rpm for 2.2 min, then increase the speed to 14000 rpm for 6 min to form a milky white homogeneous emulsion. S4. Gradient cooling molding: The emulsion is transferred to a constant temperature cooling bath with an initial temperature of 72°C. The temperature is then reduced to 36°C at a rate of 5°C / min and held for 5 min. The temperature is then reduced to 3°C at a rate of 2°C / min and refrigerated for 3 h to obtain the BPS-5 nano micelles.

[0051] Performance test results: Particle size and Zeta potential: Same as in Example 1 (0.5 mL of the finished product was diluted to 10 mL, ultrasonically dispersed, and then measured); Results: Average particle size 80 ± 3 nm, PDI 0.20 ± 0.02, Zeta potential -25 ± 1 mV; Encapsulation efficiency and drug loading: same as in Example 1; Results: encapsulation efficiency 94.6±0.7%, drug loading 2.42±0.08%; storage stability results are shown in Table 5.

[0052] Table 5. Storage stability results of BPS-5 at different temperatures. Temperature group Encapsulation rate (%) Encapsulation retention rate (%) Particle size (nm) PDI System status 4℃ 93.8±0.7 99.2 79±3 0.20±0.02 No layering, no sedimentation, and the color remains milky white. 25℃ 91.7±0.8 96.9 81±3 0.22±0.02 No stratification, no sedimentation, slight opacity, and no color change. 40℃ 86.5±0.9 91.4 84±4 0.25±0.03 There was no obvious layering, but a small amount of sediment appeared at the bottom, and the color turned milky yellow. Simulated gastric-intestinal fluid stepwise release experiment: same as in Example 1; Results: cumulative release rate in the gastric fluid stage (0-2h) was 3.9±0.6% (total free curcumin in 2h was 0.00021±0.00004mg), and cumulative release rate in the intestinal fluid stage (2-10h) was 96.0±1.2% (total free curcumin in 10h was 0.0052±0.0004mg). Antioxidant capacity: DPPH scavenging rate 93.2±1.3%; Oral pharmacokinetics in mice: Same as in Example 1; Results: The AUC0-12h of the BPS-5 nanomicelle group was 1125±28 ng・h / mL, Cmax was 252±16 ng / mL, and Tmax was 0.5±0.1h; the AUC0-12h of the free curcumin group was 24.1±1.8 ng・h / mL, and the AUC of the BPS-5 group was 46±3 times that of the free group.

[0053] Comparative Example 1: Preparation of shellac-deficient nanomicelles (denoted as BPS-01) 1. Raw material ratio (only shellac is missing, the rest is the same as in Example 1) Curcumin 2g, PGPR 5g, β-sitosterol succinate 3g, liquid coconut oil 80g, 95% (v / v) food grade ethanol 16g, xanthan gum 100mg, citric acid 60mg, purified water 80mL (all food grade or meeting corresponding standards); 2. Preparation steps (same as in Example 1, no adjustments) S1. Organic phase mixing: Add PGPR, β-sitosterol succinate, and curcumin to 60℃ liquid coconut oil, add ethanol, and stir at 1000rpm for 10min until completely dissolved; S2. Aqueous phase adjustment: Prepare a 1.25 g / L xanthan gum aqueous solution (100 mg xanthan gum) with 80 mL of purified water. Add all of this solution to the organic phase, stir at 1000 rpm for 6 min, and add 0.6% (w / v) citric acid aqueous solution to adjust the pH to 3. S3. Homogenization: Preheat the mixed system to 70°C, pre-emulsify at 8500 rpm for 2 min, and then finely emulsify at 15000 rpm for 8 min. S4. Gradient cooling: 70℃→38℃ (5℃ / min, hold for 5min)→4℃ (2℃ / min), refrigerate for 3 hours to obtain the finished product.

[0054] 3. Performance test results (The experimental method is the same as in Example 1, and the test was performed in parallel for 3 times. The results are taken as the average value ± error) Basic performance: Particle size 128±3 nm, PDI 0.32±0.02, Zeta potential -18±2 mV, encapsulation efficiency 62.8±1.2%, drug loading 0.58±0.05%, simulated gastric juice release rate 45.3±2.1%, simulated intestinal juice release rate 52.1±1.8%, oral AUC0-12h in mice 312±25 ng・h / mL; stability results are shown in Table 6.

[0055] Table 6. Storage stability results of BPS-01 at different temperatures. Temperature group Encapsulation rate (%) Encapsulation retention rate (%) Particle size (nm) PDI System status 4℃ 58.5±1.2 93.2 135±4 0.35±0.02 Slightly cloudy, with no obvious sediment. 25℃ 52.3±1.5 83.3 145±5 0.40±0.02 Moderately turbid, with trace amounts of sediment at the bottom. 40℃ 42.1±1.8 67.0 160±6 0.45±0.03 Severely turbid with a small amount of sediment at the bottom. Antioxidant capacity (DPPH method): The DPPH free radical scavenging rate was 47.9±1.6%, which was significantly lower than that of Example 1 (91.5±1.4%) and free curcumin (67.3±1.8%) (p<0.05).

[0056] Comparative Example 2: Preparation of nanomicelles lacking PGPR (denoted as BPS-02) 1. Raw material ratio (only PGPR is missing, the rest is the same as in Example 1) Curcumin 2g, shellac 8g, β-sitosterol succinate 3g, liquid coconut oil 80g, 95% (v / v) food grade ethanol 16g, xanthan gum 100mg, citric acid 60mg, purified water 80mL.

[0057] 2. Preparation steps (same as in Example 1, no adjustments) S1. Organic phase mixing: Add shellac, β-sitosterol succinate, and curcumin to 60°C liquid coconut oil, add ethanol, and stir at 1000 rpm for 10 min (some shellac is not completely dissolved, so stirring needs to be extended to 15 min). S2-S4: Same as Example 1.

[0058] 3. Performance test results (The experimental method is the same as in Example 1, and the test was performed in parallel for 3 times. The results are taken as the average value ± error) Basic performance: Particle size was 156±4 nm, PDI was 0.41±0.03, Zeta potential was -15±2 mV, encapsulation efficiency was 58.3±1.5%, drug loading was 0.52±0.04%, simulated gastric juice release rate was 38.7±2.3%, simulated intestinal juice release rate was 48.5±1.6%, and AUC0-12h in mice was 278±22 ng・h / mL; stability results are shown in Table 7.

[0059] Table 7. Storage stability results of BPS-02 at different temperatures. Temperature group Encapsulation rate (%) Encapsulation retention rate (%) Particle size (nm) PDI System status 4℃ 53.2±1.3 91.3 168±5 0.48±0.02 Slightly flocculent suspension, without stratification 25℃ 47.8±1.6 82.0 180±6 0.52±0.03 Obvious flocculent matter, with a small amount of sediment at the bottom. 40℃ 36.2±2.0 62.1 195±8 0.58±0.03 The upper layer is clear, while the lower layer contains a large amount of sediment. Antioxidant capacity (DPPH method): The DPPH free radical scavenging rate was 43.3±1.8%, which was significantly lower than that of Example 1 (91.5±1.4%) and free curcumin (67.3±1.8%) (p<0.05).

[0060] Comparative Example 3: Preparation of nanomicelles lacking β-sitosterol succinate (denoted as BPS-03) 1. Raw material ratio (only β-sitosterol succinate is missing, the rest are the same as in Example 1) Curcumin 2g, shellac 8g, PGPR 5g, liquid coconut oil 80g, 95% (v / v) food grade ethanol 16g, xanthan gum 100mg, citric acid 60mg, purified water 80mL.

[0061] 2. Preparation steps (same as in Example 1, no adjustments) S1. Organic phase mixing: Add shellac, PGPR, and curcumin to 60℃ liquid coconut oil, add ethanol, and stir at 1000rpm for 10min until completely dissolved; S2-S4: Same as Example 1.

[0062] 3. Performance test results (The experimental method is the same as in Example 1, and the test was performed in parallel for 3 times. The results are taken as the average value ± error) Basic performance: Particle size 92±2nm, PDI 0.25±0.02, Zeta potential -22±1mV, encapsulation efficiency 76.5±1.1%, drug loading 0.71±0.06%, simulated gastric juice release rate 18.5±1.4%, simulated intestinal juice release rate 68.2±1.9%, oral AUC0-12h in mice 845±30ng・h / mL; stability test results are shown in Table 8.

[0063] Table 8. Storage stability results of BPS-03 at different temperatures. Temperature group Encapsulation rate (%) Encapsulation retention rate (%) Particle size (nm) PDI System status 4℃ 71.3±1.1 93.2 97±2 0.28±0.02 Slight devitrification, no precipitation 25℃ 66.8±1.2 87.3 100±3 0.31±0.02 Moderate devitrification, no obvious precipitation 40℃ 59.6±1.4 77.9 108±3 0.34±0.02 Noticeably cloudy with trace amounts of sediment at the bottom. Antioxidant capacity (DPPH method): The DPPH free radical scavenging rate was 65.8±1.5%, which was higher than that of Comparative Examples 1 and 2, but significantly lower than that of Example 1 (91.5±1.4%), and close to that of free curcumin (67.3±1.8%) (p<0.05).

[0064] The comparison results between the above examples and comparative examples show that the nanomicelles in Examples 1-5 all performed excellently: encapsulation efficiency ≥94.3%, gastric juice release ≤3.9%, intestinal juice release ≥95.8% (strong targeting), encapsulation retention rate ≥93.5% after 28 days at 4℃, DPPH clearance rate ≥82.5%, and mouse AUC 46-55 times that of free curcumin. Among them, BPS-3 (encapsulation efficiency 96.5%, AUC 1375±40ng・h / mL) showed the best effect.

[0065] The performance of the comparative examples dropped sharply due to the lack of a single component: the lack of shellac resulted in a gastric juice release of 45.3% and an AUC of only 25% of that in Example 1; the lack of PGPR resulted in a particle size of 156±4nm and a PDI of 0.41±0.03, making it easy to separate into layers; the lack of β-sitosterol succinate resulted in a decrease in encapsulation efficiency and a weakening of intestinal juice release.

[0066] In summary, the present invention achieves high encapsulation, targeted release, and high bioavailability of micelles through the synergistic effect of shellac barrier function, PGPR emulsification function, and β-sitosterol succinate hydrophobic anchoring function.

Claims

1. A nanomicelle for encapsulating curcumin, characterized in that, It is mainly prepared from shellac, polyglycerol ricinoleate, and β-sitosterol succinate.

2. The nanomicelles according to claim 1, characterized in that, The shellac, polyglycerol ricinoleate, and β-sitosterol succinate are present in a mass ratio of 2~8:2~5:1~3.

3. A method for preparing nanomicelles for encapsulating curcumin, characterized in that, It includes the following steps: S1. Take shellac, PGPR, β-sitosterol succinate, and curcumin and dissolve them together in liquid coconut oil at 55~70℃, and add food-grade ethanol to obtain a homogeneous and transparent organic phase mixture. S2. Add xanthan gum aqueous solution to the mixture in step S1, and adjust the pH to acidic using citric acid aqueous solution; S3. Homogenize the material obtained in step S2: After preheating, pre-emulsify first, then finely emulsify; S4. Gradient cooling: First, cool to the target temperature at a set rate and hold at that temperature, then continue cooling to the refrigeration temperature to solidify and obtain nanomicelles.

4. The preparation method according to claim 3, characterized in that, In step S1, the mass ratio of shellac, PGPR, and β-sitosterol succinate is 2~8:2~5:1~3.

5. The preparation method according to claim 3, characterized in that, In step S1, the temperature of the liquid coconut oil is 55~70℃, and the amount of liquid coconut oil used is 5~10 times the total mass of shellac, PGPR, and β-sitosterol succinate; the amount of ethanol added is 10%~30% of the volume of liquid coconut oil, and the ethanol is 95% food-grade ethanol by volume.

6. The preparation method according to claim 3, characterized in that, In step S2, the mass concentration of the xanthan gum aqueous solution is 0.5~2 g / L, and the amount of xanthan gum aqueous solution used is 0.5~1.5 times the volume of liquid coconut oil; The mass-volume concentration of citric acid aqueous solution is 0.4-1%, and the pH of the system is adjusted to 3-4.

7. The preparation method according to claim 3, characterized in that, In step S3, the mixing system is preheated to 65~75℃; the pre-emulsification speed is 8000~10000rpm and the time is 1.5~3min; the fine emulsification speed is 13000~18000rpm and the time is 5~10min, and the system temperature does not exceed 75℃ during the emulsification process.

8. The preparation method according to claim 3, characterized in that, In step S4, the gradient cooling process specifically involves: first cooling to 35~40℃ and holding at that temperature, then cooling to 0~8℃ and refrigerating for solidification.

9. The preparation method according to claim 8, characterized in that, First, lower the temperature to 35~40℃ at 5℃ / min and hold for 3~5 minutes, then lower the temperature to 0~8℃ at 2℃ / min and refrigerate for 3~4 hours to solidify.

10. The use of the nanomicelles according to claim 1 or the preparation method according to any one of claims 3-8 in the preparation of functional food formulations or pharmaceuticals.

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