Drug-loaded nanoparticle and metal polyphenol network mediated food-borne assembly

Abstract

The invention relates to the technical field of drug-loaded particles, in particular to a drug-loaded nanoparticle and metal polyphenol network mediated food-borne assembly, which is characterized in that natural sulfide is coated in calcium carbonate containing a mesoporous structure to form the drug-loaded nanoparticle; the preparation method comprises the following steps: preparing drug-loaded nanoparticles, and coating the outer layers of the drug-loaded nanoparticles with a nano coating with a metal-polyphenol network structure to obtain a metal polyphenol network mediated food-borne assembly which is used for preparing a drug for treating alcohol injury, and the drug is beneficial to relieving tissue injury and behavior disorder caused by alcohol.

Description

Technical Field

[0001] The present invention relates to the technical field of drug-loaded particles, and in particular to a drug-loaded nanoparticle and an edible assembly mediated by a metal polyphenol network. Background Art

[0002] With the continuous advancement of materials science, the demand for novel functional materials is increasing. Traditional drug carriers have limitations, such as limited drug loading and insufficient stability. Metal-polyphenol networks (MPNs) are a class of materials with three-dimensional network structures formed by the coordination bond interaction between metal ions and polyphenol compounds. This unique coordination chemistry not only endows MPNs with dynamic and reversible interfacial assembly properties, but also exhibits multifunctionality, biocompatibility, and tunable physicochemical properties. In recent years, MPNs have demonstrated broad application prospects in biomedical engineering fields such as drug delivery systems, biosensor interfaces, and biomimetic coating technologies, thanks to their precise and controllable nanostructure construction capabilities and multifunctional integrated properties, becoming a hot material system for interdisciplinary research.

[0003] For example, CN119033957A discloses a metal polyphenol network-coated calcium peroxide nanoparticle, and a preparation method and application thereof, comprising the following steps: (1) mixing calcium peroxide nanoparticles, a drug, and a first solvent, and then subjecting the mixture to a chelation reaction to obtain drug-loaded calcium peroxide nanoparticles; (2) mixing a polymer solution with a polyphenol solution, and then subjecting the mixture to a dehydration condensation reaction to obtain a polymer-modified polyphenol solution; (3) mixing the drug-loaded calcium peroxide nanoparticles obtained in step (1) with a second solvent, the polymer-modified polyphenol solution obtained in step (2), and a metal salt solution, and then subjecting the mixture to a coordination complexation reaction to obtain metal polyphenol network-coated calcium peroxide nanoparticles. However, calcium oxide nanoparticles have a strong alkalinity and are easily reacted with water in a physiological environment or in some common solvents to generate calcium hydroxide, and even further absorb carbon dioxide to convert into substances such as calcium carbonate, which may cause changes in the structure and properties of the nanoparticles, seriously affecting the stability and effect of the drug loading thereof. At the same time, its strong alkalinity may cause certain stimulation and damage to cells and tissues, causing adverse phenomena such as inflammatory reactions, which may not only affect the delivery effect of the drug, but also cause additional damage to the body.

[0004] Alcoholic injury refers to a state of disorder in the body's functions caused by the ingestion of a large amount of alcohol far exceeding the body's metabolic capacity. Alcoholic injury can lead to multi-organ damage, including the nervous, digestive, and cardiovascular systems, especially irreversible damage to the brain and liver. This injury interferes with the normal signal transmission of the nervous system, causing symptoms such as cognitive impairment and motor disorders. At the same time, it also brings a metabolic burden to the liver, which may lead to damaged liver cells and abnormal liver function. After alcohol is ingested into the human body, alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) are activated, and the alcohol metabolism process intensifies intracellular oxidative stress metabolism, increasing intracellular reactive oxygen species (ROS). The production of acetaldehyde exacerbates the damage caused by this oxidative stress. When tissues such as the brain and liver are damaged, inflammatory mediators are released, triggering an inflammatory response and leading to further aggravation of alcohol injury.

[0005] The endogenous signaling molecule H2S has attracted great interest due to its significant antioxidant and anti-inflammatory properties. Studies have shown that H2S can reduce ROS-induced apoptosis, inactivate the MAPK signaling pathway, and reduce oxidative stress-related damage. These findings suggest the potential of H2S as a treatment for alcohol injury. Although H2S donor molecules have been developed, many molecules cannot precisely and controllably release H2S continuously. NaHS and Na2S release unstably and rapidly induce cytotoxicity under physiological conditions. JK1 and JK2 can release H2S in response to acid too quickly. Diallyl disulfide (DADS), an oil-soluble organosulfide derived from garlic, can provide a more controllable and persistent release of H2S in the presence of glutathione (GSH). However, its water solubility is poor (less than 1 μg / mL), its chemical properties are unstable, which largely negatively affects its storage and bioavailability, and limits its practical application. Therefore, it is necessary to develop an effective delivery system to directly preserve and encapsulate hydrophobic drugs during storage to improve their loading capacity and bioavailability. However, in the existing technology, no drug has been successfully prepared by combining it with metal-polyphenol network materials and having a certain alleviating effect on alcohol injury.

[0006] Based on the above situation of the existing technology, there are technical problems to be solved urgently in the existing technology, such as the natural sulfide diallyl disulfide (DADS) has a mitigating effect on alcohol injury, but its poor water solubility and unstable chemical properties make it unable to be directly used as a drug. Therefore, it is necessary to develop an effective delivery system to directly preserve and encapsulate hydrophobic drugs during storage to improve their portability and bioavailability. Summary of the Invention

[0007] To solve the above technical problems, the present invention provides a drug-loaded nanoparticle, which is formed by coating a natural sulfide in calcium carbonate with a mesoporous structure to form the drug-loaded nanoparticle (S@CaCO3). The average particle size of the drug-loaded nanoparticle is 10-300 nm; the loading rate of the natural sulfide in the drug-loaded nanoparticle is greater than or equal to 70%.

[0008] Further, the natural sulfide includes, but is not limited to, one or several of diallyl disulfide (DADS), diallyl trisulfide (DATS), diallyl monosulfide (DAS), and diallyl tetrasulfide.

[0009] The present invention also provides a preparation method of the above drug-loaded nanoparticle, which includes the following steps: Step 1: Dissolve the natural sulfide in a calcium ion solution, and form a mixed solution after ultrasonic mixing; Step 2: React the mixed solution with ammonium bicarbonate by a gas diffusion method to obtain a reaction solution; Step 3: Wash and centrifuge the reaction solution, and the obtained precipitate is the drug-loaded nanoparticle (S@CaCO3).

[0010] Further, the natural sulfide includes a natural H2S donor compound.

[0011] Further, in Step 1, the concentration of calcium ions in the calcium ion solution is 1-4 mg / mL, and the solvent is an organic solvent.

[0012] Further, the calcium source in the calcium ion solution in Step 1 includes, but is not limited to, calcium chloride.

[0013] Further, the organic solvent includes, but is not limited to, anhydrous ethanol.

[0014] Further, in Step 1, the mass ratio of the natural sulfide to the calcium source is 1:(1-50).

[0015] Further, the stirring and mixing time in Step 1 is greater than or equal to 6 h.

[0016] Further, the intensity of the ultrasonic mixing in Step 1 is greater than or equal to 300 W.

[0017] Further, in Step 2, the ammonium bicarbonate (NH4HCO3) is dry ammonium bicarbonate without water after drying.

[0018] Further, the specific reaction principle in Step 2 is as follows: The dried ammonium bicarbonate (NH4HCO3) is placed on one side of the reaction vessel, and the mixed solution prepared in Step 1 is placed on the other side of the reaction vessel. By controlling the reaction temperature, the CO2 and NH3 generated by the decomposition of ammonium bicarbonate diffuse into the mixed solution, triggering a chemical reaction to form calcium carbonate precipitate with a mesoporous structure. Meanwhile, the natural sulfide contained in the mixed solution is loaded in the mesoporous structure. During the loading process, physical and / or chemical interactions occur. Physical interactions include van der Waals forces generated during adsorption; and / or in chemical interactions, sulfide ions (S 2- ) in the natural sulfide undergo ion exchange with calcium ions (Ca 2+ ) on the surface of the calcium carbonate mesoporous structure or form compounds such as calcium sulfide, ultimately achieving the stable loading of natural sulfide in the mesoporous structure.

[0019] Further, the mass ratio of the natural sulfide in Step 1 to the ammonium bicarbonate in Step 2 is 1:(5 - 10).

[0020] Further, the reaction temperature in Step 2 is 25 - 60 °C.

[0021] Further, the washing solvent used in Step 3 is an organic solvent, and the type of the organic solvent is the same as that used for the calcium ion solution in Step 1.

[0022] Further, the number of washing times in Step 3 is at least 3 times to remove unreacted impurities.

[0023] The present invention provides an edible source assembly mediated by a metal polyphenol network (MPN@S@CaCO3@NPs). The edible source assembly mediated by the metal polyphenol network is a nano - coating with a metal - polyphenol network structure wrapped around the outer layer of the above - mentioned drug - loaded nanoparticles (S@CaCO3). The edible source assembly mediated by the metal polyphenol network has a nano - scale spherical structure with an average particle size of 50 - 1000 nm.

[0024] The present invention also provides a preparation method of the above - mentioned edible source assembly mediated by a metal polyphenol network (MPN@S@CaCO3@NPs), including the following steps: Step 1: Disperse the drug - loaded nanoparticles (S@CaCO3) into an organic alcohol solvent to obtain reaction solution A (alcohol dispersion of S@CaCO3). Step 2: Prepare an aqueous polyphenol solution, and add reaction solution A to the aqueous polyphenol solution. After mixing, obtain reaction solution B (polyphenol@S@CaCO3 solution). Step 3: Prepare a metal ion solution, add the metal ion solution to the reaction solution B, mix by ultrasonic treatment and then centrifuge. The obtained precipitate is the metal polyphenol network-mediated edible assembly (MPN@S@CaCO3@NPs).

[0025] Further, the organic alcohol solvent in Step 1 includes but is not limited to ethanol.

[0026] Further, the molar concentration of the natural sulfide in the drug-loaded nanoparticles in Step 1 in the reaction solution A is 0.1 μmol / L - 1 mmol / L.

[0027] Further, the solute polyphenol in the polyphenol aqueous solution in Step 2 is a compound with an ortho-phenol structure.

[0028] Further, the polyphenol includes but is not limited to one or a combination of several of epigallocatechin gallate (EGCG), gallic acid (GA), tannic acid (TA), epicatechin (EC), epicatechin gallate (ECG), and epigallocatechin (EGC).

[0029] Further, the molar concentration of the polyphenol in the polyphenol aqueous solution in Step 2 is 0.1 μmol / L - 5 mmol / L.

[0030] Further, the mass ratio of the drug-loaded nanoparticles (S@CaCO3) in the reaction solution A in Step 2 to the solute polyphenol in the polyphenol aqueous solution is 1:(0.05 - 10).

[0031] Further, the mixing time in Step 2 is greater than or equal to 3 min.

[0032] Further, the stirring speed of the mixing in Step 2 is greater than or equal to 20 rpm.

[0033] Further, the solute metal ions in the metal ion solution in Step 3 are one or several of the trace elements required by the human body.

[0034] Further, the metal ions include but are not limited to one or a combination of several of zinc, iron, magnesium, and copper.

[0035] Further, the molar concentration of the solute metal ions in the metal ion solution in Step 3 is 0.1 μmol / L - 5 mmol / L.

[0036] Further, the mass ratio of the solute metal ions in the metal ion solution in Step 3 to the solute polyphenol in the reaction solution B is 1:(0.1 - 10).

[0037] Further, the mixing time in Step 3 is greater than or equal to 3 min.

[0038] Furthermore, the intensity of the ultrasonic mixing described in step 3 is greater than or equal to 300 W.

[0039] Furthermore, the reaction principle involved in step 3 is that polyphenols contain phenolic hydroxyl groups, and the phenolic hydroxyl groups self-assemble into a metal-polyphenol network structure through coordination chelation with metal ions and adhere to the interface of the drug-loaded nanoparticles (S@CaCO3) to form a nanocoating; Meanwhile, hydrogen bonds are formed between polyphenol molecules and between polyphenols and groups such as hydroxyl groups on the surface of the drug-loaded nanoparticles (S@CaCO3). The existence of hydrogen bonds enables polyphenols to connect with each other and combine with hydroxyl groups, enhancing the stability of the nanocoating; meanwhile, metal ions can also interact with the hydrogen bond network on the surface of polyphenols and the drug-loaded nanoparticles (S@CaCO3), further promoting the self-assembly process.

[0040] The present invention also provides a drug, and the preparation raw materials of the drug include the above-mentioned metal-polyphenol network-mediated edible assembly (MPN@S@CaCO3@NPs).

[0041] Furthermore, the drug is used to relieve drunkenness symptoms.

[0042] The beneficial effects of the present invention are as follows: 1. The present invention provides a drug-loaded nanoparticle (S@CaCO3), in which natural sulfide is coated in calcium carbonate with a mesoporous structure to form the drug-loaded nanoparticle. The average particle size of the drug-loaded nanoparticle is 10 - 300 nm; the loading rate of the natural sulfide in the drug-loaded nanoparticle is greater than or equal to 70%; and a nanocoating with a metal-polyphenol network structure is wrapped on the outer layer of the drug-loaded nanoparticle to obtain a metal-polyphenol network-mediated edible assembly (MPN@S@CaCO3@NPs). The metal-polyphenol network-mediated edible assembly has a nanoscale spherical structure with an average particle size of 50 - 1000 nm; and the present invention creatively develops a metal-polyphenol network-mediated edible assembly using completely edible ingredients for preparing drugs for treating alcohol damage; 2. The polyphenols used in the present invention, such as epigallocatechin gallate (EGCG), as the main polyphenol in green tea, can rapidly self-assemble with metal ions into a metal-polyphenol network structure through coordination and chelation. When an interface exists, it can adhere to the interface to form a nanocoating. Based on this, the present invention uses mesoporous CaCO3 nanoparticles as a template, polyphenols as organic ligands, and metal ions as metal centers to obtain a metal-polyphenol network-mediated edible assembly through coordination reaction. It shows high loading efficiency for natural sulfides such as DADS, and the metal polyphenol coating protects unstable CaCO3 nanoparticles. The resulting metal-polyphenol network-mediated edible assembly is used as a raw material for preparing a drug for alleviating the symptoms of drunkenness. It not only significantly improves the bioavailability of natural sulfides such as DADS, but also exhibits stronger anti-inflammatory and antioxidant activities when used in combination with polyphenols and metal ions. 3. In an alcohol-damaged mouse model, the metal polyphenol network-mediated edible assemblies, such as Zn-EGCG@S@CaCO3@NPs, activated the body's antioxidant system and significantly reduced oxidative stress levels in brain and liver tissues. Simultaneously, they inhibited the decrease in ALDH levels in the liver, thereby alleviating acetaldehyde accumulation caused by intoxication and the suppression of the central nervous system. Behavioral experiments showed that the mice's motor dysfunction was improved. Therefore, the metal polyphenol network-mediated edible assemblies prepared by the present invention will help alleviate alcohol-induced tissue damage and behavioral disorders, and are expected to achieve clinical translation. At the same time, according to the results of the test experiments, it can be clearly seen that after the present invention loads natural sulfide in the mesopores of the drug-loaded nanoparticles and prepares the metal polyphenol network-mediated edible assembly, it can be directly dissolved in PBS buffer solution and can also be used directly as a drug. BRIEF DESCRIPTION OF THE DRAWINGS

[0043] Figure 1 This is an electron micrograph of the drug-loaded nanoparticles prepared in Example 1 of the present invention; Figure 2 This is an electron micrograph of the edible assembly mediated by the metal polyphenol network prepared in Example 2 of the present invention; Figure 3 This is a UV spectrum analysis characterization diagram tested in Example 2 of the present invention; Figure 4 This is a diagram showing the analysis results of a footprint experiment on improving cerebellar ataxia in drunken mice according to Application Example 1 of the present invention; Figure 5 This is a diagram showing the analysis results of the rotarod test for improving cerebellar ataxia in drunken mice according to Application Example 1 of the present invention; Figure 6Analysis result graph of the balance beam experiment on the improvement of cerebellar ataxia in drunk mice in Application Example 1 of the present invention; Figure 7 Analysis result graph of the protection of nerve cells damaged by alcohol in vitro in Application Example 1 of the present invention; Figure 8 Analysis result graph of the oxidative stress caused by alcohol in Application Example 1 of the present invention; Figure 9 Analysis result graph of apoptosis caused by oxidative stress in Application Example 1 of the present invention; Figure 10 Analysis result graph of liver damage caused by alcohol metabolism in Application Example 1 of the present invention; Figure 11 Analysis result graph of the inflammatory response caused by alcohol in Application Example 1 of the present invention; Figure 12 Analysis result graph of the footprint experiment on the improvement of cerebellar ataxia in drunk mice in Application Example 2 of the present invention; Figure 13 Analysis result graph of the rotarod experiment on the improvement of cerebellar ataxia in drunk mice in Application Example 2 of the present invention; Figure 14 Analysis result graph of the balance beam experiment on the improvement of cerebellar ataxia in drunk mice in Application Example 2 of the present invention; Figure 15 Analysis result graph of the protection of nerve cells damaged by ethanol in vitro in Application Example 2 of the present invention; Figure 16 Analysis result graph of the oxidative stress caused by alcohol in Application Example 2 of the present invention. Detailed implementation mode

[0044] Example 1 is a drug-loaded nanoparticle (S@CaCO3) and its preparation method; Examples 2-4 are metal polyphenol network-mediated edible assemblies (MPN@S@CaCO3@NPs) prepared using the drug-loaded nanoparticles (S@CaCO3) prepared in Example 1, and their preparation methods; Application Examples 1-2 are to use the metal polyphenol network-mediated edible assemblies prepared in Examples 2-4 as drugs for relieving drunken symptoms, and conduct relevant experimental verifications on mice.

[0045] Example 1 As Figure 1 (Scale bar: 100nm) shows, this example provides a drug-loaded nanoparticle (S@CaCO3), which is formed by coating the natural sulfide diallyl disulfide (DADS) in calcium carbonate with a mesoporous structure, and the average particle size of the drug-loaded nanoparticle is 200nm; A known amount of DADS was dissolved in absolute ethanol, and the linear regression equation of DADS(@S) was calculated by measuring data using a UV-visible spectrophotometer. Finally, the content of DADS in S@CaCO3 was tested by recording the absorbance at 210 nm, and the loading rate calculated from the average of three average results was 78.7%.

[0046] The preparation method of the drug-loaded nanoparticles (S@CaCO3) includes the following steps: Step 1: 1 g of diallyl disulfide (DADS) was dissolved in a calcium ion solution, i.e., an absolute ethanol solution containing 15 g of calcium chloride. After mixing for 6 h under an ultrasonic intensity of 300 W, a mixed solution was formed; the concentration of calcium ions in the calcium ion solution was 3 mg / mL. Step 2: The mixed solution was reacted with ammonium bicarbonate by the gas diffusion method. Specifically, 5 g of dry ammonium bicarbonate (NH4HCO3) without water after drying was placed on one side of the reaction vessel, and the mixed solution prepared in Step 1 was placed on the other side of the reaction vessel. By controlling the reaction temperature at 35 °C, CO2 and NH3 generated by the decomposition of ammonium bicarbonate diffused into the mixed solution, triggering a chemical reaction to form calcium carbonate precipitate with a mesoporous structure. Meanwhile, the natural sulfide diallyl disulfide contained in the mixed solution was loaded in the mesoporous structure. Physical and chemical interactions occurred during the loading. Among them, the physical interaction was the van der Waals force generated during adsorption; the chemical interaction was that the sulfide ions (S 2- ) in the natural sulfide reacted with the calcium ions (Ca 2+ ) on the surface of the calcium carbonate mesoporous structure to undergo ion exchange or form compounds such as calcium sulfide, ultimately realizing the stable loading of the natural sulfide in the mesoporous structure to obtain a reaction solution. Step 3: The reaction solution was washed 3 times with absolute ethanol and centrifuged to remove unreacted impurities. The finally obtained precipitate was the drug-loaded nanoparticles (S@CaCO3).

[0047] Example 2 As Figure 2 shown (Scale bar: 100 nm), this example provides an edible source assembly mediated by a metal-polyphenol network (Zn-EGCG@S@CaCO3@NPs). The edible source assembly mediated by the metal-polyphenol network is a nano-coating with a metal-polyphenol network structure wrapped on the outer layer of the drug-loaded nanoparticles (S@CaCO3) prepared in Example 1. The edible source assembly mediated by the metal-polyphenol network has a nanoscale spherical structure with an average particle size of 250 nm.

[0048] Preparation method of the metal polyphenol network-mediated edible assembly (Zn-EGCG@S@CaCO3@NPs), comprising the following steps: Step 1: Disperse the drug-loaded nanoparticles (S@CaCO3) prepared in Example 1 into absolute ethanol to obtain reaction solution A (alcohol dispersion of S@CaCO3); wherein the molar concentration of the natural sulfide in the drug-loaded nanoparticles in reaction solution A is 0.1 mmol / L; Step 2: Prepare an aqueous polyphenol solution, add reaction solution A to the aqueous polyphenol solution, and under the condition of a stirring speed of 20 rpm, mix for 3 min to obtain reaction solution B (EGCG@S@CaCO3 solution); in this example, the solute polyphenol of the aqueous polyphenol solution is epigallocatechin gallate (EGCG), and the molar concentration of EGCG in the aqueous polyphenol solution is 0.05 mmol / L; and the mass ratio of the drug-loaded nanoparticles (S@CaCO3) in reaction solution A to EGCG is 1:5; Step 3: Prepare a Zn 2+ solution with a concentration of 0.5 mmol / L of Zn 2+ and add the Zn 2+ solution to reaction solution B, wherein the mass ratio of the solute Zn 2+ in the Zn 2+ solution to the solute polyphenol EGCG in reaction solution B is 1:5. After mixing for 20 min under an ultrasonic intensity of 300 W and then centrifuging, the finally obtained precipitate is the metal polyphenol network-mediated edible assembly (MPN@S@CaCO3@NPs), which is specifically represented as Zn-EGCG@S@CaCO3@NPs in this example.

[0049] Among them, the reaction principle involved in Step 3 is that polyphenols contain phenolic hydroxyl groups, and the phenolic hydroxyl groups self-assemble into a metal-polyphenol network structure through coordination chelation with metal ions and adhere to the interface of the drug-loaded nanoparticles (S@CaCO3) to form a nano-coating; Meanwhile, in this example, hydrogen bonds are also formed between polyphenol molecules and between polyphenols and groups such as hydroxyl groups on the surface of the drug-loaded nanoparticles (S@CaCO3). The existence of hydrogen bonds enables polyphenols to be interconnected and combined with hydroxyl groups, enhancing the stability of the nano-coating; at the same time, metal ions can also interact with the hydrogen bond network on the surface of polyphenols and the drug-loaded nanoparticles (S@CaCO3), further promoting the self-assembly process.

[0050] Among them, the drug-loaded nanoparticles (S@CaCO3), epigallocatechin gallate (EGCG), and metal polyphenol network-mediated edible assemblies (Zn-EGCG@S@CaCO3@NPs) in this example and Example 1 were characterized by UV spectral analysis. The analysis results are as Figure 3 shown. Through the characterization of UV spectral analysis, the interaction between Zn ions, EGCG, and S@CaCO3 was further demonstrated. As Figure 3 shown, EGCG showed two characteristic ultraviolet absorption peaks at 213 nm and 275 nm respectively. S@CaCO3 showed the ultraviolet absorption peak of DADS, indicating the successful loading of DADS by mesoporous CaCO3. In addition, the characteristic ultraviolet absorption peak of EGCG at 275 nm in Zn-EGCG@S@CaCO3@NPs shifted towards the longer wavelength direction, and a new characteristic peak appeared at 570 nm, indicating an ionic crosslinking reaction between EGCG and Zn ions. In addition, Zn-EGCG@S@CaCO3@NPs showed the characteristic UV peak of Zn-EGCG, providing strong support for the successful loading of EGCG and Zn ions in this co-delivery nanosystem.

[0051] Example 3 This example provides a metal polyphenol network-mediated edible assembly (Fe-EGCG @S@CaCO3@NPs). The metal polyphenol network-mediated edible assembly is a nano-coating with a metal-polyphenol network structure wrapped on the outer layer of the drug-loaded nanoparticles (S@CaCO3) prepared in Example 1. The metal polyphenol network-mediated edible assembly has a nanoscale spherical structure with an average particle size of 300 nm.

[0052] The preparation method of the metal polyphenol network-mediated edible assembly (Fe-EGCG@S@CaCO3@NPs) includes the following steps: Step 1: Disperse the drug-loaded nanoparticles (S@CaCO3) prepared in Example 1 into absolute ethanol to obtain reaction solution A (alcohol dispersion of S@CaCO3). The molar concentration of the natural sulfide in the drug-loaded nanoparticles in reaction solution A is 0.1 mmol / L; Step 2: Prepare an aqueous polyphenol solution, add reaction solution A to the aqueous polyphenol solution, and mix for 3 min under the condition of a stirring speed of 20 rpm to obtain reaction solution B (EGCG@S@CaCO3 solution). In this example, the solute polyphenol of the aqueous polyphenol solution is epigallocatechin gallate (EGCG), the molar concentration of EGCG in the aqueous polyphenol solution is 0.05 mmol / L; and the mass ratio of the drug-loaded nanoparticles (S@CaCO3) in reaction solution A to EGCG is 1:5; Step 3. Configure Fe 2+ Fe concentration of 0.5mmol / L 2+ solution, add the Fe 2+ solution, wherein the Fe 2+ Solute Fe in solution 2+ The mass ratio of the solute polyphenol EGCG in the reaction solution B is 1:5, and the mixture is mixed at an ultrasonic intensity of 300 W for 20 minutes and then centrifuged. The final precipitate obtained is the metal polyphenol network-mediated edible assembly (MPN@S@CaCO3@NPs), which is specifically represented by Fe-EGCG@S@CaCO3@NPs in this embodiment.

[0053] The reaction principle involved in step 3 is that polyphenols contain phenolic hydroxyl groups, which self-assemble with metal ions through coordination and chelation to form a metal-polyphenol network structure, which then adheres to the interface of the drug-loaded nanoparticles (S@CaCO3) to form a nanocoating layer. At the same time, in this embodiment, hydrogen bonds are formed between polyphenol molecules and between polyphenols and groups such as hydroxyl groups on the surface of drug-loaded nanoparticles (S@CaCO3). The existence of hydrogen bonds enables polyphenols to connect with each other and combine with hydroxyl groups, thereby enhancing the stability of the nanocoating; at the same time, metal ions can also interact with the hydrogen bond network on the surface of polyphenols and drug-loaded nanoparticles (S@CaCO3), further promoting the self-assembly process.

[0054] Example 4 This embodiment provides a metal polyphenol network-mediated edible assembly (Zn-TA@S@CaCO3@NPs). The metal polyphenol network-mediated edible assembly is a nanocoating having a metal-polyphenol network structure wrapped on the outer layer of the drug-loaded nanoparticles (S@CaCO3) prepared in Example 1. The metal polyphenol network-mediated edible assembly has a nanoscale spherical structure with an average particle size of 550 nm.

[0055] The preparation method of the metal polyphenol network-mediated edible assembly (Zn-TA@S@CaCO3@NPs) comprises the following steps: Step 1: Disperse the drug-loaded nanoparticles (S@CaCO3) prepared in Example 1 in anhydrous ethanol to obtain a reaction solution A (an alcohol dispersion of S@CaCO3); wherein the molar concentration of the natural sulfide in the drug-loaded nanoparticles in the reaction solution A is 0.1 mmol / L; Step 2: Prepare a polyphenol aqueous solution. Add the reaction solution A to the polyphenol aqueous solution, and under the condition of a stirring speed of 20 rpm, mix for 3 min to obtain a reaction solution B (TA@S@CaCO3 solution). In this example, the solute polyphenol in the polyphenol aqueous solution is tannic acid (TA), the molar concentration of TA in the polyphenol aqueous solution is 0.05 mmol / L, and the mass ratio of the drug-loaded nanoparticles (S@CaCO3) in the reaction solution A to TA is 1:5. Step 3: Prepare a Zn 2+ solution with a concentration of 0.5 mmol / L of Zn 2+ and add the Zn 2+ solution to the reaction solution B. Among them, the mass ratio of the solute Zn 2+ in the Zn 2+ solution to the solute polyphenol TA in the reaction solution B is 1:5. Mix under an ultrasonic intensity of 300 W for 20 min and then centrifuge. The finally obtained precipitate is the metal-polyphenol network-mediated edible assembly (MPN@S@CaCO3@NPs), which is specifically represented as Zn-TA@S@CaCO3@NPs in this example.

[0056] Among them, the reaction principle involved in Step 3 is that polyphenols contain phenolic hydroxyl groups, and the phenolic hydroxyl groups self-assemble into a metal-polyphenol network structure through coordination chelation with metal ions and adhere to the interface of the drug-loaded nanoparticles (S@CaCO3) to form a nano-coating. At the same time, in this example, hydrogen bonds are also formed between polyphenol molecules and between polyphenols and groups such as hydroxyl groups on the surface of the drug-loaded nanoparticles (S@CaCO3). The existence of hydrogen bonds enables polyphenols to be connected to each other and combined with hydroxyl groups, enhancing the stability of the nano-coating. At the same time, metal ions can also interact with the hydrogen bond network on the surface of polyphenols and drug-loaded nanoparticles (S@CaCO3), further promoting the self-assembly process.

[0057] Application Example 1 This application example uses the metal-polyphenol network-mediated edible assembly (Zn-EGCG@S@CaCO3@NPs) prepared in Example 2 as the raw material for preparing a drug (abbreviated as Drug A) to relieve hangovers. To illustrate its beneficial effects, the following tests are specifically carried out: (1) Improvement of cerebellar ataxia in mice after drunkenness by the metal-polyphenol network-mediated edible assembly: The improvement effect of drug A on cerebellar ataxia was tested using footprint experiments, balance beam experiments, and rotarod experiments. The mice used in this experiment were 6-8-week-old busulfan-sensitive mice (BALB / c). The metal polyphenol network-mediated edible assembly with a concentration of 0.5 mg / mL was administered to ethanol-intoxicated mice at a dose of 2.5 mg / kg by gavage, denoted as the drug A group. Among them, the solvent used in drug A was phosphate buffer solution (PBS). In all subsequent related application examples, PBS buffer was also used when preparing drugs with the metal polyphenol network-mediated edible assembly. At the same time, two control groups were set up, namely the normal group and the alcohol group. Among them, the normal group was normal mice without ethanol intoxication, and the alcohol group was ethanol-intoxicated mice. After 30 minutes, footprint experiments, balance beam experiments, and rotarod experiments were carried out simultaneously: 1) Footprint experiment: By analyzing the footprints generated when the experimental animals walk, the motor coordination ability of the mice was evaluated by assessing their gait.

[0058] The experimental results are as Figure 4 shown, where (a) is the footprint imprint diagram of the mice, and (b) is the statistical chart of the footprint spacing of the mice. It can be seen from the figure that during walking, the footprints of the hind paws of the mice in the normal group almost overlapped with those of the front paws, indicating their good limb coordination. The footprints of the mice in the alcohol group were of different depths, and the footprints of the front and hind paws did not overlap, and there were phenomena such as trajectory inclination and tail dragging. Compared with the normal group, the distance between their two feet increased significantly. The mice in the drug A group containing the metal polyphenol network-mediated edible assembly prepared in Example 2 could significantly restore the even stride of the ethanol-intoxicated mice again, and to a certain extent restored the motor coordination ability of the mice.

[0059] 2) Rotarod experiment: The performance of the rotating rod was evaluated on the suspension rod of an accelerating rotating rod device (diameter: 3 cm). The rotating rod device accelerated at a constant rate of 1-25 rpm for 300 s. The mice were trained continuously for 3 days and were placed on the rod for 3 trials. The time of each trial was recorded. The trial ended when the mouse fell off the rotating rod or the time reached 300 s. A 180 s rest was allowed between each trial.

[0060] The experimental results are as Figure 5 follows: The movement time of the mice in the alcohol group was reduced by 5 times compared with that of the mice in the normal group in the rotarod test, while the movement time of the mice given the metal polyphenol network-mediated edible assembly was significantly increased compared with that of the mice in the alcohol group, indicating that the metal polyphenol network-mediated edible assembly can improve the motor ability of drunken mice.

[0061] 3) Balance beam experiment: Use a suspended wooden rod with a width of 15 mm, record the time the mouse stays on it, and the maximum recording time is 60 s. The mice are trained continuously for 3 days, and all the trainings are concentrated in a fixed time period every day. After the training, the experiment starts on the 4th day, and the movement state of the mice on the balance beam is scored. A stable balance posture is scored 0 points; being able to hold tightly to the edge of the balance beam is scored 1 point; hugging the balance beam tightly with one limb hanging down from the balance beam is scored 2 points; hugging the balance beam tightly with two limbs hanging down from the balance beam or rotating on the balance beam (>60 s) is scored 3 points; attempting to balance on the balance beam but falling (>40 s) is scored 4 points; attempting to balance on the balance beam but falling (>20 s) is scored 5 points; falling; not attempting to balance on the balance beam (<20 s) is scored 6 points. Repeat three times to end the experiment. A 180 s rest is allowed between each experiment.

[0062] The experimental results are as Figure 6 shown: In the alcohol group, the grasping force and balance ability of the mice on the balance beam were greatly reduced, and they rotated or even fell on the balance beam due to the inability to maintain balance in a short time. While in the Group A mice administered with the drug, the balance ability was restored to a certain extent, indicating that the edible assembly mediated by the metal polyphenol network can relieve the balance ability damage caused by excessive drinking, and the consciousness of the drunken mice can be restored 60 min after administration.

[0063] (2) Protection of the edible assembly mediated by the metal polyphenol network against alcohol-damaged nerve cells in vitro: The MTT method is used to detect the cell viability in vitro. The MTT method is the tetramethyl thiazolyl blue colorimetric method, which is a commonly used method for detecting cell survival and growth. This method is based on the fact that mitochondrial dehydrogenase in living cells can reduce yellow MTT (tetramethyl thiazolyl blue) to insoluble blue-violet formazan particles. By measuring the absorbance of formazan at a specific wavelength, the number and activity of living cells can be indirectly reflected.

[0064] The SH-SY5Y neuroblastoma cells are evenly plated in a 96-well plate at a cell density of 5×10 6 , 100 μL per well. Place it in an incubator containing 5% CO2 at 37 °C. After culturing for 4 h, aspirate the original culture medium, and add fresh culture medium containing 500 mM ethanol (EtOH) to damage the SH-SY5Y neuroblastoma. At the same time, add six concentrations of Drug A containing the edible assembly mediated by the metal polyphenol network at 0, 5, 15, 30, 60, 90, 120, 150, 180 μg / mL. Put the cell plate back into the incubator and continue to culture for 24 h. The next day, aspirate the culture medium, add 100 μL of 1 mg / mL MTT solution, put it back into the incubator and continue to culture for 4 h. Aspirate the MTT, add 150 μL of dimethyl sulfoxide (DMSO) to dissolve the formazan, and use an enzyme-linked immunosorbent assay (ELISA) reader to detect the absorbance value at a wavelength of 490 nm.

[0065] The experimental results are as follows Figure 7 shown: Adding 500 mM ethanol caused about 40% death of neuroblastoma cells. After administering the metal polyphenol network-mediated edible assembly, the survival rate of neuroblastoma cells showed a dose-dependent relationship with the concentration of the metal polyphenol network-mediated edible assembly, and the survival rate of neuroblastoma cells increased by more than 20%, indicating that the metal polyphenol network-mediated edible assembly can improve the survival rate of damaged nerve cells in vitro.

[0066] (3)The metal polyphenol network-mediated edible assembly alleviates oxidative stress caused by alcohol: The intracellular oxidative stress level induced by ethanol was detected by 2',7'-dichlorodihydrofluorescein diacetate probe (DCFH-DA probe). SH-SY5Y neuroblastoma cells were evenly seeded in a 6-well plate at a cell density of 5×10 6 . After being placed in an incubator with 5% CO2 at 37 °C for 4 h, the original medium was aspirated, and fresh medium containing 500 mM ethanol was added. At the same time, drug A containing 120 μg / mL of the metal polyphenol network-mediated edible assembly was added, and the cell plate was returned to the incubator for further culture for 24 h. DCFH-DA was diluted 1:1000 with serum-free Dulbecco's Modified Eagle's Medium (DMEM) to prepare a DCFH-DA working solution. After removing the cell medium and washing with phosphate-buffered saline (PBS), 1 mL of the prepared DCFH-DA working solution was added to each well and incubated in the incubator for 20 min. The DCFH-DA working solution was discarded, and the cells were washed three times with serum-free DMEM, and then imaged with an inverted fluorescence microscope.

[0067] The experimental results are as follows Figure 8 shown: After incubating SH-SY5Y cells with ethanol, bright green fluorescence appeared after the probe was added. This further proved that alcohol can cause oxidative stress. The green fluorescence in the group treated with drug A was significantly weakened, and with the increase of the drug concentration, the fluorescence became weaker, indicating that the metal polyphenol network-mediated edible assembly can effectively reduce cellular oxidative stress.

[0068] (4)The metal polyphenol network-mediated edible assembly alleviates apoptosis caused by oxidative stress: The brain protection effect of metal polyphenol network-mediated edible assemblies was tested using the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining technique. TUNEL staining fluorescesly labels fragmented DNA. When cells are damaged, the DNA breaks and is labeled with green fluorescence. The mice used in this experiment were 6-8-week-old BALB / c mice. The metal polyphenol network-mediated edible assemblies at a concentration of 0.5 mg / mL were administered by gavage at a dose of 2.5 mg / kg, and cardiac perfusion was performed 4 h later. The perfused brain tissues were removed, immersed in 4% paraformaldehyde (PFA) overnight, dehydrated in a gradient with 20%, 30%, and 40% sucrose solutions, and liver slices with a thickness of 20 μm were cut after embedding. Staining steps: The slices were washed in water for 5 min, fixed in 4% PFA for 30 min, washed twice with PBS for 10 min each time, the cells were perforated with a PBS solution containing 0.5% Triton X-100 for 5 min, incubated with the pre-prepared staining solution (5 μL terminal deoxynucleotidyl transferase (TdT) + 45 μL fluorescent labeling solution + 50 μL detection solution) at 37 °C in the dark for 1 h, washed three times with PBS, sealed with glycerol containing 4',6-diamidino-2-phenylindole (DAPI), and imaged with an inverted fluorescence microscope.

[0069] The experimental results are as Figure 9 shown (scale bar: 100 μm): The green fluorescence intensity in the normal group was weak. Compared with the normal group, the green fluorescence intensity in the alcohol group increased significantly, indicating that ethanol caused brain tissue damage. After treatment with the metal polyphenol network-mediated edible assemblies, the green intensity in the A drug group was significantly weaker than that in the alcohol group, indicating that the metal polyphenol network-mediated edible assemblies have a protective effect on nerve apoptosis caused by drunkenness.

[0070] Among them, Figure 9 DAPI: is an anti-fluorescence quenching nuclear fuel. 4',6'-Diamidino-2-phenylindole is a fluorescent dye that can strongly bind to DNA and is commonly used for fluorescence microscopy observation. Since DAPI can penetrate intact cell membranes, it can be used for staining of live cells and fixed cells; TUNEL: is a method for detecting cell apoptosis. When genomic DNA breaks, the exposed 3'-OH can be added with dUTP at the 3'-end by terminal deoxynucleotidyl transferase (TdT) Under the catalysis of [catalyst name], fluorescein-dUTP (fluorescein-labeled dUTP) is added, enabling detection through a fluorescence microscope or flow cytometer. This is the principle of the TUNEL (TdT-mediated dUTP Nick-End Labeling) method for detecting apoptosis; Merge: For overlaying layers.

[0071] (5)Edible assemblies mediated by metal polyphenol networks can alleviate liver damage caused by alcohol metabolism: In this experiment, 6-8-week-old BALB / c mice were used. The edible assemblies mediated by metal polyphenol networks with a concentration of 0.5 mg / mL were administered by gavage at a dose of 2.5 mg / kg. After 4 h, the mice were sacrificed and serum samples from mice in different treatment groups were collected and analyzed using a blood biochemical analyzer (MNCHIP).

[0072] The experimental results are as Figure 10 (scare bar: 100 μm): Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were further used to evaluate liver toxicity in vivo. The results showed that the levels of serum ALT and AST in the PBS-treated group (alcohol group) were significantly higher than those in the normal group, but were significantly lower in the group treated with the edible assemblies mediated by metal polyphenol networks (Drug A group).

[0073] (6)Edible assemblies mediated by metal polyphenol networks alleviate the inflammatory response caused by alcohol The mice used in this experiment were 6 - 8 - week - old BALB / c mice. The metal polyphenol network - mediated edible assembly with a concentration of 0.5 mg / mL was administered by gavage at a dose of 2.5 mg / kg. After 4 h, cardiac perfusion was performed. The perfused brain tissues were removed and homogenized in 1 mL of total RNA extraction reagent (Trizol). The samples were homogenized in 1 mL of Tizol at room temperature. The samples were placed at room temperature (15 - 30 °C) for 5 min to completely separate the nucleic acid - protein complexes. 200 μL of chloroform was added to each milliliter of Tizol, and it was shaken for 15 s and then placed at room temperature for 3 min. Then it was centrifuged in a 4 °C centrifuge for 15 min at a speed of 12,000 rpm. After centrifugation, the sample was divided into three layers. The bottom layer was a yellow organic phase, the upper layer was a colorless aqueous phase and the middle layer. RNA was mainly in the aqueous phase, and the volume of the aqueous phase was about 60% of the volume of Tizol used. Then, it was centrifuged in a 4 °C centrifuge for 10 min at a speed of 12,000 rpm, and the supernatant was discarded. The RNA precipitate was washed with 500 μL of 75% ethanol and centrifuged in a 4 °C centrifuge for 5 min at a speed of 7,500 rpm, and the supernatant was discarded. The RNA precipitate was air - dried and an appropriate amount of RNase - free water (DECP water) was added. It was placed at 55 - 60 °C for 10 min to dissolve the RNA. The obtained RNA was quantified by a ultra - micro spectrophotometer (NanoDrop2000). Then the mRNA was reverse - transcribed into cDNA. The RNA was reverse - transcribed using a reverse transcription kit (TranscriptFirst - Standard cDNAsynethesis Supermix kit). The 20 μL reverse transcription system: 7 μL of RNase - free water, 1 μL of oligo - deoxythymidine (OligoddT), 1 μL of RNA, 10 μL of Rmix, and 1 μL of Emix were added in sequence. It was incubated in a PCR instrument at 42 °C for 5 min and at 85 °C for 5 s.

[0074] Among them, Rmix: is 2×TS Reaction Mix; Emix: is miRNA RT Enzyme Mix; it can complete efficient tailing addition and first - strand cDNA synthesis in one step.

[0075] Among them, the reaction system of RT - qPCR is shown in Table 1: Table 1 RT - qPCR reaction system

[0076] SuperReal PreMix Plus is a premixed reagent for real - time fluorescence quantitative PCR (qPCR) experiments; it is a premixed Mix, and only the template, primers, and ddH2O need to be added to perform Real - Time PCR reactions.

[0077] ROX (Passive Reference Dye) is an inert reference dye.

[0078] Prepare the primers and 2×SuperReal PreMix Plus in a 1.5 mL EP tube according to the reagent ratios in the table. Then prepare the remaining reagents in another 1.5 mL EP tube. Finally, add them sequentially to a 96-well or 384-well PCR plate. Seal the entire plate with a PCR film, mix by shaking, centrifuge at 1000 rpm for 3 min, and then place it in an RT-qPCR instrument for amplification. The program settings are as follows: (1) Pre-denaturation at 95°C for 15 min, 1 cycle, without collecting fluorescence signals.

[0079] (2) PCR reaction: 95°C for 10 s; 60°C for 32 s, 40 cycles, collecting fluorescence signals.

[0080] (3) Record the Ct value, use β-actin as an internal reference, and calculate the relative expression level of the target gene.

[0081] Calculate the mRNA expression level according to the comparative Ct method ( ) The corresponding primer sequences are shown in Table 2.

[0082] Table 2 RT-qPCR primer sequences

[0083] Among them, in the above reverse transcription-polymerase chain reaction (RT-qPCR), Gapdh is glyceraldehyde-3-phosphate dehydrogenase; Forward is the forward of the nucleic acid sequence, and Reverse is the reverse of the nucleic acid sequence. Among them, the sequence of Forward (5’-3) of interleukin-1β (IL-1β) is as shown in SEQ ID NO:1, and the sequence of Reverse (5’-3) is as shown in SEQ ID NO:2; the sequence of Forward (5’-3) of tumor necrosis factor-α (TNF-α) is as shown in SEQ ID NO:3, and the sequence of Reverse (5’-3) is as shown in SEQ ID NO:4; the sequence of Forward (5’-3) of inducible nitric oxide synthase (iNOS) is as shown in SEQ ID NO:5, and the sequence of Reverse (5’-3) is as shown in SEQ ID NO:6; the sequence of Forward (5’-3) of glyceraldehyde-3-phosphate dehydrogenase (Gapdh) is as shown in SEQ ID NO:7, and the sequence of Reverse (5’-3) is as shown in SEQ ID NO:8.

[0084] The experimental results are as Figure 11As shown: Oxidative stress is usually accompanied by the occurrence of inflammation, and severe inflammation can cause irreversible damage to brain tissue and liver tissue. Therefore, we next detected the inflammatory level in the brain. We detected the expression of pro-inflammatory factors interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and inducible nitric oxide synthase (iNOS) in brain tissue and liver tissue by RT-qPCR technology. The results showed that oxidative stress caused by ethanol metabolism did cause neuroinflammation (alcohol group), while the metal-polyphenol network-mediated edible-derived assembly (Group A drug) significantly reduced the inflammatory levels in the brain and liver, which helped to alleviate the brain damage and liver damage caused by alcohol metabolism.

[0085] Application Example 2 This application example uses the metal-polyphenol network-mediated edible-derived assembly (Zn-EGCG@S@CaCO3@NPs) prepared in Examples 3 and 4 as the raw material for preparing a drug to relieve hangovers, abbreviated as Drug B (including the metal-polyphenol network-mediated edible-derived assembly prepared in Example 3) and Drug C (including the metal-polyphenol network-mediated edible-derived assembly prepared in Example 4). To illustrate its beneficial effects, the following tests were specifically conducted: (1)Improvement of metal-polyphenol network-mediated edible-derived assembly on cerebellar ataxia in drunk mice: The footprint experiment, balance beam experiment, and rotarod experiment were used to test the improvement effect of Drug B and Drug C on cerebellar ataxia. The mice used in this experiment were 6-8-week-old busulfan-sensitive mice (BALB / c). The metal-polyphenol network-mediated edible-derived assembly with a concentration of 0.5 mg / mL was administered to ethanol-intoxicated mice at a dose of 2.5 mg / kg by gavage, denoted as Drug B group and Drug C group. At the same time, two control groups were set up, namely the normal group and the alcohol group. Among them, the normal group was normal mice without ethanol intoxication, and the alcohol group was ethanol-intoxicated mice. After 30 min, the footprint experiment, balance beam experiment, and rotarod experiment were carried out simultaneously: 1) Footprint experiment: By analyzing the footprints generated when the experimental animals walk, the motor coordination ability of mice was evaluated by assessing their gait.

[0086] The experimental results are as Figure 12 shown, where (a) is the footprint imprint map of mice, and (b) is the statistical chart of the footprint spacing of mice: During the walking process of normal group mice, the footprints of the hind paws almost overlapped with those of the front paws, indicating their good limb coordination. The footprints of alcohol group mice were of different depths, and the footprints of the front paws and hind paws did not overlap, and there were phenomena such as trajectory inclination and tail dragging. Compared with the normal group, the distance between their two feet increased significantly. Pretreatment with Drug B group and Drug C group significantly made the mouse stride uniform and restored the motor coordination ability of mice to a certain extent.

[0087] 2) Rotarod test: The performance of the rotarod was evaluated on the suspension rod of an accelerating rotarod device (diameter: 3 cm). The rotarod device was accelerated at a constant rate of 1 - 25 rpm for 300 s. Mice were trained continuously for 3 days and placed on the rod for 3 trials. The time of each trial was recorded. The trial ended when the mouse fell off the rotarod or the time reached 300 s. A 180 s rest was allowed between each trial.

[0088] The experimental results are as Figure 13 follows: The exercise time of mice in the alcohol group was reduced by 5 times compared with that of the normal group in the rotarod test. However, the exercise time of mice given the metal polyphenol network-mediated edible assembly was significantly increased compared with that of the mice in the alcohol group, indicating that the metal polyphenol network-mediated edible assembly can improve the exercise ability of drunken mice.

[0089] 3) Balance beam test: A suspended wooden beam with a width of 15 mm was used to record the time the animal stayed on it, with a maximum recording time of 60 s. Mice were trained continuously for 3 days, and all the trainings were concentrated in a fixed time period every day. After the training, the experiment started on the 4th day, and the movement state of the mice on the balance beam was scored. A stable balance posture was scored 0; being able to hold tightly to the edge of the balance beam was scored 1; holding tightly to the balance beam with one limb hanging down from the balance beam was scored 2; holding tightly to the balance beam with two limbs hanging down or rotating on the balance beam (>60 s) was scored 3; attempting to balance on the balance beam but falling (>40 s) was scored 4; attempting to balance on the balance beam but falling (>20 s) was scored 5; falling; not attempting to balance on the balance beam (<20 s) was scored 6. The test was repeated three times to end. A 180 s rest was allowed between each trial.

[0090] The experimental results are as Figure 14 shown: The grasping force and balance ability of mice in the alcohol group on the balance beam were greatly reduced, and they rotated or even fell on the balance beam due to inability to maintain balance in a short time. However, after administration, there was a tendency for the balance ability to recover, indicating that the metal polyphenol network-mediated edible assembly can alleviate the balance ability damage caused by excessive alcohol consumption to a certain extent.

[0091] (2) Protection of metal polyphenol network-mediated edible assembly on nerve cells damaged by ethanol in vitro: The MTT method was used to detect the cell viability in vitro. SH-SY5Y neuroblastoma cells were seeded at 5×10 6Cells were evenly plated in a 96-well plate at a cell density of 100 μL per well. The plate was placed in an incubator with 5% CO2 at 37 °C for 4 h. Then, the original medium was aspirated, and fresh medium containing 500 mM ethanol was added to damage SH-SY5Y neuroblastoma cells. At the same time, 120 μg / mL of Drug B and Drug C were added. The cell plate was returned to the incubator and cultured for another 24 h. The next day, the medium was aspirated, 100 μL of 1 mg / mL MTT solution was added, and the plate was placed back in the incubator for 4 h. The MTT was then aspirated, and 150 μL of DMSO was added to dissolve the formazan. The absorbance at a wavelength of 490 nm was measured using a microplate reader.

[0092] The experimental results are as Figure 15 shown: Adding 500 mM ethanol caused about 50% death of neuroblastoma cells. After administration of Drugs B and C, the survival rate of neuroblastoma cells showed a dose-dependent relationship with the concentration of the metal-polyphenol network-mediated edible-derived assembly. The survival rate of neuroblastoma cells increased by more than 20%, indicating that the metal-polyphenol network-mediated edible-derived assembly can improve the survival rate of damaged nerve cells in vitro.

[0093] (3)The metal-polyphenol network-mediated edible-derived assembly alleviates oxidative stress caused by alcohol: The intracellular oxidative stress level induced by ethanol was detected using a DCFH-DA probe. SH-SY5Y neuroblastoma cells were evenly plated in a 6-well plate at a cell density of 5×10 6 . After culturing in an incubator with 5% CO2 at 37 °C for 4 h, the original medium was aspirated, and fresh medium containing 500 mM ethanol was added. At the same time, 120 μg / mL of Drugs B and C were added. The cell plate was returned to the incubator and cultured for another 24 h. DCFH-DA was diluted 1:1000 with serum-free DMEM to prepare a DCFH-DA working solution. The cell medium was removed, and after washing with PBS, 1 mL of the prepared DCFH-DA working solution was added to each well and incubated in the incubator for 20 min. The DCFH-DA working solution was discarded, and the cells were washed three times with serum-free DMEM. Imaging was performed using an inverted fluorescence microscope.

[0094] The experimental results are as Figure 16 shown: After incubating SH-SY5Y cells with ethanol, bright green fluorescence appeared after the probe was added, further demonstrating that alcohol can cause oxidative stress. In the group treated with the drug, the green fluorescence was significantly weakened, and with the increase in drug concentration, the fluorescence became weaker, indicating that the metal-polyphenol network-mediated edible-derived assembly can effectively reduce cellular oxidative stress.

[0095] As can be seen from the above, the present invention forms the drug-loaded nanoparticles by coating natural sulfides in calcium carbonate with a mesoporous structure, and wraps the outer layer of the drug-loaded nanoparticles with a nano-coating having a metal-polyphenol network structure to obtain a metal-polyphenol network-mediated edible assembly for preparing a drug for treating alcohol injury, and the drug helps to reduce tissue damage and behavioral disorders caused by alcohol.

[0096] It should be understood that the present invention is not limited to the content already described above and shown in the drawings, and various modifications and changes can be made without departing from its scope. The scope of the present invention is only limited by the appended claims.

Claims

1. A drug-loaded nanoparticle, characterized in that, The drug-loaded nanoparticles are formed by coating natural sulfide in calcium carbonate with a mesoporous structure, and the average particle size of the drug-loaded nanoparticles is 10 - 300 nm; the loading rate of the natural sulfide in the drug-loaded nanoparticles is greater than or equal to 70%.

2. The drug-loaded nanoparticles according to claim 1, characterized in that, The natural sulfide is one or more of diallyl disulfide, diallyl trisulfide, diallyl monosulfide, and diallyl tetrasulfide.

3. A method for preparing the drug-loaded nanoparticles according to any one of claims 1-2, characterized in that, It includes the following steps: Step 1: Dissolve the natural sulfide in a calcium ion solution, and form a mixed solution after ultrasonic mixing; Step 2: React the mixed solution with ammonium bicarbonate by gas diffusion method to obtain a reaction solution; Step 3: Wash and centrifuge the reaction solution, and the obtained precipitate is the drug-loaded nanoparticles.

4. The preparation method of the drug-loaded nanoparticles according to claim 3, characterized in that, The calcium source in the calcium ion solution in Step 1 is calcium chloride.

5. A metal-polyphenol network-mediated edible assembly, characterized in that, The metal polyphenol network-mediated edible assembly is to coat a nano-coating with a metal-polyphenol network structure on the outer layer of the drug-loaded nanoparticles according to any one of Claims 1 - 2. The metal polyphenol network-mediated edible assembly has a nano-scale spherical structure with an average particle size of 50 - 1000 nm.

6. A method for preparing the metal-polyphenol network-mediated edible assembly according to claim 5, characterized in that, It includes the following steps: Step 1: Disperse the drug-loaded nanoparticles into an organic alcohol solvent to obtain reaction solution A; Step 2: Prepare an aqueous polyphenol solution, and add reaction solution A to the aqueous polyphenol solution, and obtain reaction solution B after mixing; Step 3: Prepare a metal ion solution, add the metal ion solution to reaction solution B, ultrasonically mix and then centrifuge, and the obtained precipitate is the metal polyphenol network-mediated edible assembly.

7. The preparation method of the metal polyphenol network-mediated edible assembly according to claim 6, wherein, The solute polyphenol in the aqueous polyphenol solution in Step 2 is a compound with an ortho-phenol structure.

8. The preparation method of the metal-polyphenol network-mediated edible assembly according to claim 7, wherein, The polyphenol is one or a combination of several of epigallocatechin gallate, gallic acid, tannic acid, epicatechin, epicatechin gallate, and epigallocatechin.

9. The preparation method of the metal polyphenol network-mediated edible assembly according to claim 6, wherein The solute metal ions in the metal ion solution in Step 3 are one or several of trace elements required by the human body.

10. A drug, characterized in that, The preparation raw materials of the drug include the metal polyphenol network-mediated edible assembly according to Claim 6.

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