Application of angelica keiskei chalcone in preparation of intervention medicine for metabolism-related fatty liver diseases

Abstract

The invention relates to the technical field of pharmaceutical preparations, and discloses an application of angelica keiskei chalcone in preparation of an intervention drug for metabolism-related fatty liver diseases, and the intervention drug comprises the following raw materials: an angelica keiskei chalcone extract, a methacrylic acid-methyl methacrylate copolymer, L-arginine, nicotinamide and L-tartaric acid. The preparation process adopts a fluid phase change coprecipitation method. In the liquid preparation stage, L-arginine is utilized to realize preliminary solubilization of chalcone. Nicotinamide and chalcone construct a charge transfer complex to inhibit chemical degradation of active components in an alkaline environment. In the coprecipitation stage, L-tartaric acid constructs a spatial multidentate cross-linked network in a polymer matrix, locks the amorphous state of drug molecules, and cuts off a recrystallization channel. In the post-treatment stage, pre-cooled acid washing liquid is adopted for replacing and washing waste salt, and powder is prevented from absorbing moisture and deliquescing. The in-vitro dissolution rate, the chemical stability and the physical stability of long-term storage of the chalcone are effectively improved.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical formulation technology, specifically to the application of Ashitaba chalcone in the preparation of drugs for the intervention of metabolism-related fatty liver disease. Background Technology

[0002] Metabolic fatty liver disease seriously endangers human health, and chalcone from Ashitaba has a definite intervention effect on this disease. However, chalcone compounds are highly hydrophobic substances with extremely low solubility in water, resulting in significant deficiencies in clinical bioavailability under oral administration conditions. Therefore, improving the solubility and in vitro dissolution rate of chalcone is a prerequisite for developing relevant intervention drugs.

[0003] In pharmaceutical research and development, alkaline excipients are often added to disrupt the crystal structure of poorly soluble drugs, enabling preliminary solubility in fluid media. Building upon this, preparing drugs into amorphous solid dispersions is a common process to improve in vitro drug release rates. This involves dispersing and encapsulating drug monomers within a polymer matrix, utilizing the amorphous state to eliminate the original lattice dissolution barrier. Coprecipitation is a conventional technique for the large-scale preparation of solid dispersion particles, primarily through fluid interaction inducing a phase transition in the polymer and trapping drug molecules. In the industrial post-processing stage, the filter cake surface is typically washed with purified water at room temperature to remove waste salt byproducts from acid-base neutralization reactions. The filter cake is then vacuum-dried to obtain the final drug powder.

[0004] However, because chalcone molecules contain an unsaturated ketone skeleton, their active sites are easily attacked when exposed to alkaline liquid environments, leading to reverse aldol condensation and isomerization degradation. Therefore, conventional alkaline solubilization causes significant structural damage to the active ingredient during the solution preparation stage. Furthermore, the amorphous microstructure is thermodynamically metastable, and conventional polymer carriers only provide macroscopic physical encapsulation. Under high-temperature and high-humidity storage conditions, drug monomers easily undergo translation and spatial rearrangement within the polymer's free pores. The spontaneous aggregation and recrystallization structure completely negates the drug release advantage of the solid dispersion. Moreover, conventional vacuum drying processes cannot remove these trapped waste salt crystals. Due to their strong affinity for water, these waste salts become capillary nuclei inducing moisture absorption in the powder, ultimately causing rapid deliquescence when the product is exposed to air. Therefore, this invention provides a multifunctional antibacterial and antifungal soybean protein composite adhesive and its preparation method to address the shortcomings of existing technologies. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides the application of Ashitaba chalcone in the preparation of intervention drugs for metabolism-related fatty liver disease. It solves the problems of poor water solubility leading to low bioavailability in the preparation of intervention drugs for metabolism-related fatty liver disease using existing Ashitaba chalcone. Conventional alkaline solubilization techniques cause chalcone to undergo reverse aldol condensation and isomerization degradation. Under humid and hot storage conditions, the internal active ingredients are prone to physical recrystallization, and the by-product salts generated during the preparation process can cause the powder to absorb moisture and deliquesce, reducing the long-term stability of the product.

[0006] To achieve the above objectives, the present invention provides the following technical solution: In a first aspect, the present invention provides an intervention drug for metabolism-related fatty liver disease containing Ashitaba chalcone, employing the following technical solution: The interventional drug is made from the following raw materials in parts by weight: 10 to 15 parts of Ashitaba chalcone extract; 30 to 50 parts of methacrylate-methyl methacrylate copolymer; 4 to 8 parts of L-arginine; 3 to 6 parts of nicotinamide; and 8.1 to 15.5 parts of L-tartaric acid.

[0007] By employing the above technical solutions, this invention establishes multiple chemical and physical stabilization mechanisms at the formulation level, thereby improving drug solubility and inhibiting its degradation and recrystallization. The reaction mechanism and process are as follows: L-arginine in the system provides a weakly alkaline environment, which disrupts the original crystal structure of chalcone molecules, thereby achieving the initial solubilization of the target drug in the fluid medium.

[0008] The added nicotinamide contains a pyridine ring structure, which spatially approaches the chalcone molecule in the Ashitaba chalcone extract, forming a charge-transfer complex. This complex forms a spatial shield around the easily hydrolyzed site of chalcone, blocking the attack path of the nucleophile and inhibiting the reverse aldol condensation reaction of chalcone under alkaline conditions.

[0009] The added L-tartaric acid molecule contains free carboxyl and hydroxyl groups, which provide hydrogen bond donor and acceptor sites. The L-tartaric acid molecule directionally binds to free nicotinamide amide groups and carboxyl residues on the methacrylic acid copolymer segments in the coprecipitation system, constructing a spatial multidentate crosslinking network. This network physically restricts the translational and rotational trajectories of the chalcone monomer, locking the thermodynamic degrees of freedom of the target molecule and truncates the evolutionary path from the amorphous to the crystalline state.

[0010] The methacrylic acid-methyl methacrylate copolymer undergoes a phase transition during acid-base environment switching, forming a dense polymer skeleton that encapsulates the above components and blocks the direct influence of the external humid and hot environment.

[0011] Preferably, the mass ratio of methacrylic acid to methyl methacrylate in the methacrylic acid-methyl methacrylate copolymer is 1:1.

[0012] By adopting the above technical solution, the copolymer in a specific ratio has anti-alkali hydrolysis properties. The polymer backbone will not undergo destructive degradation at the structural level in weakly alkaline liquids, and will undergo rapid hydrophobic collapse when in contact with acidic environments, thus ensuring the molding rate of the drug-loaded skeleton.

[0013] Preferably, the Ashitaba chalcone extract is prepared by the following steps: Ashitaba herb powder is extracted twice by heating and reflux with a 75% (v / v) ethanol aqueous solution to obtain an extract filtrate; the filtrates are combined and concentrated under reduced pressure to obtain an aqueous extract; the aqueous extract is dynamically adsorbed onto an adsorption resin chromatography column and then eluted sequentially with purified water and an ethanol aqueous solution; the ethanol eluent is collected, concentrated under reduced pressure, vacuum dried, pulverized, and sieved to obtain the final product.

[0014] By adopting the above technical solution and using a combination of ethanol and resin chromatography steps, macromolecular impurities and water-soluble inactive components in the medicinal materials are removed, the purity of chalcone monomers is improved, and a uniform raw material basis is provided for the subsequent preparation of solid dispersions.

[0015] Preferably, the chalcone molecules in the Ashitaba chalcone extract are dispersed in an amorphous state within a mesoporous framework formed by the hydrophobic collapse of the methacrylic acid-methyl methacrylate copolymer.

[0016] By employing the above technical solution, the amorphous state eliminates the original lattice dissolution barrier of the active ingredient. After the aqueous medium penetrates the porous framework, chalcone molecules can directly diffuse and dissolve outward, improving the in vitro dissolution rate of the drug.

[0017] Secondly, the present invention provides a preparation process for a drug for intervening in metabolism-related fatty liver disease containing Ashitaba chalcone, using the following technical solution: The intervention drug is an amorphous solid dispersion of Ashitaba chalcone, and the preparation process of the intervention drug includes the following steps: S1. Mix purified water and anhydrous ethanol, add L-arginine and nicotinamide and stir to dissolve to form a weakly alkaline base solution; S2. Add methacrylic acid-methyl methacrylate copolymer to the weakly alkaline base solution, stir to dissolve and form a colloidal solution, then add Ashitaba chalcone extract and stir in the dark to form a composite feed solution. S3. Add purified water to the crystallization reactor, dissolve it with L-tartaric acid to form an acidic bottom solution, cool it down and start the high-shear homogenizer; S4. The composite feed liquid is pumped into the crystallization reactor and injected into the acidic bottom liquid through the submerged microporous distribution ring, and a co-precipitated microparticle suspension is precipitated in the fluid jet confluence. S5. The obtained suspension is subjected to solid-liquid separation to retain the filter cake. The filter cake is washed by replacing it with an acidic washing solution prepared with purified water and L-tartaric acid and pre-cooled. The washed filter cake is then vacuum dried, pulverized and sieved to obtain the intervention drug containing the amorphous solid dispersion of Ashitaba chalcone.

[0018] By employing the above technical solution, this process combines small molecule protection with polymer phase transition. The specific process is as follows: In the feed solution preparation stage, L-arginine and nicotinamide are used to maintain the chemical structural integrity of chalcone. In the fluid jetting and mixing stage, the feed solution is injected into a low-pH acidic substrate, causing an instantaneous phase transition. The anionic copolymer undergoes hydrophobic collapse, precipitating co-precipitated particles that retain the drug. In the washing stage, an acidic washing solution containing L-tartaric acid is used for displacement, preventing the backflow of water-soluble components and ultimately solidifying the internal structure.

[0019] Preferably, in step S1, the mass ratio of purified water to anhydrous ethanol is 100 to 150 to 40 to 60, the system temperature is controlled at 20 to 25 degrees Celsius, and the pH value of the weakly alkaline base solution is 9.0 to 9.5; in step S2, the stirring time for adding Ashitaba chalcone extract in the dark is 30 to 45 minutes.

[0020] By employing the above technical solution, the introduction of a specific proportion of anhydrous ethanol reduces the overall dielectric constant of the mixed solvent. This environment weakens the nucleophilic attack capability of water molecules and reduces the kinetic rate of hydrolysis side reactions. Simultaneously, it improves the spread of hydrophobic polymers in the feed solution, allowing polymer segments to achieve physical capture of drug molecules with a larger surface area during subsequent phase transitions, thereby increasing the encapsulation efficiency of the final product.

[0021] Preferably, in step S3, the pH value of the acidic bottom solution is 2.5 to 3.0, the temperature of the bottom solution is reduced to 5 to 10 degrees Celsius, and the rotation speed of the high-shear homogenizer is set to 1500 to 2500 revolutions per minute; in step S4, the submerged microporous distribution ring is installed directly below the high-shear impeller, the pore diameter is 0.3 to 0.5 mm, and the feed flow rate of the composite feed liquid is 10 to 20 parts per minute.

[0022] By employing the above technical solution, the microporous distribution ring disperses the feed liquid into fine droplets, and the high-shear environment and low-temperature conditions accelerate the mass transfer process at the fluid mixing section. These hydrodynamic conditions ensure that the polymer completes protonation and precipitation instantaneously at the phase intersection, preventing drug monomers from escaping into the aqueous medium during the phase transition delay period, thereby improving the overall process yield.

[0023] Preferably, in step S5, the pH value of the pre-cooled acidic washing solution is 2.5 to 3.0, and the pre-cooling temperature is 0 to 5 degrees Celsius; the pre-cooled acidic washing solution keeps the polymer molecular chains in a contracted glassy state, forming a hydrophobic barrier on the surface of the co-precipitated particles, and removes free tartaric acid arginine salts by piston flow scouring.

[0024] By employing the above technical solution, the low pH and low temperature conditions forcibly maintain the protonated state of the free carboxyl groups on the methacrylic acid segments, causing the polymer network to contract and preventing mass exchange between the washing liquid and the particles. The fluid, in a piston-flow manner, washes away the waste salt crystals adhering to the macroscopic gaps in the compacted filter cake. Removing the highly hygroscopic waste salt eliminates the capillary cores that allow environmental moisture to penetrate, preventing subsequent deliquescence and agglomeration of the powder during storage.

[0025] Preferably, in step S5, the obtained suspension is subjected to solid-liquid separation, with the separation factor set to 1000 to 2000 x g, and the acidic washing liquid is sprayed in while the centrifuge is rotating; the vacuum drying conditions are a temperature of 40 to 50 degrees Celsius, a vacuum degree of -0.08 to -0.10 MPa, and a drying time of 10 to 16 hours.

[0026] By adopting the above technical solution, the spraying method ensures that the washing liquid evenly covers the surface of the filter cake, and the gentle drying temperature and vacuum conditions can remove residual moisture without damaging the multi-tooth cross-linked network structure, thus maintaining the physicochemical stability of the powder.

[0027] Thirdly, the present invention provides the application of Ashitaba chalcone in the preparation of an intervention drug for metabolism-related fatty liver disease, wherein the drug is made from the raw materials of the aforementioned intervention drug for metabolism-related fatty liver disease containing Ashitaba chalcone, or the drug is made using the aforementioned preparation process.

[0028] By adopting the above technical solution, Ashitaba chalcone, a solid dispersion with high solubility, high chemical stability and high hygroscopic resistance, is applied to the disease field, solving the pharmaceutical bottleneck of this active ingredient in clinical application and meeting the actual needs of drugs for intervention of metabolism-related fatty liver disease for bioavailability and formulation stability.

[0029] This invention provides the application of ashwagandha chalcone in the preparation of drugs for the intervention of metabolism-related fatty liver disease. It has the following beneficial effects: 1. This invention introduces L-arginine to disrupt the chalcone lattice for initial solubilization. Simultaneously, the added nicotinamide utilizes its pyridine ring structure to construct a charge-transfer complex with the chalcone molecule. This structure creates a steric hindrance outside the easily hydrolyzed sites of chalcone, blocking the attack path of nucleophiles and preventing the reverse aldol condensation and isomerization reactions of the active ingredient in a weakly alkaline fluid environment, thus maintaining the structural integrity of the active ingredient in the feed solution.

[0030] 2. The L-tartaric acid added in this invention utilizes the free carboxyl and hydroxyl groups within the molecule to directionally bind with carboxyl residues on the nicotinamide and methacrylic acid copolymer segments, constructing a multidentate cross-linked network within the matrix. This network physically restricts the movement trajectory of chalcone monomers, locks the amorphous state of drug molecules, and cuts off the recombination channels that degenerate from the amorphous state to the crystalline state, ensuring that the powder still possesses a high cumulative dissolution rate after experiencing a humid and hot environment.

[0031] 3. In the preparation method of this invention, an acidic washing solution pre-cooled to 0 to 5 degrees Celsius and with a pH value of 2.5 to 3.0 is used to replace and wash the filter cake. This low temperature and acidic condition maintains the protonated state of the carboxyl groups on the polymer chains, causing the polymer network to contract and preventing washing water from penetrating into the particles. The fluid, in a piston-flow manner, removes highly hygroscopic waste salts adhering to the interparticle gaps, eliminating factors that induce product deliquescence and ensuring the final drug powder's flowability and dryness. Attached Figure Description

[0032] Figure 1 This is a flowchart of the preparation method of the present invention; Figure 2 The graph shows a comparison of the chemical stability of the feed liquid and the encapsulation efficiency of the product in this invention. (a) is a trend graph of the retention rate of chalcone monomer over time, and (b) is a comparison graph of the encapsulation efficiency of the solid dispersion powder. Figure 3 The graph shows the test results of the yield and degradation rate of the effective components in the preparation of the solid dispersion of the present invention. Figure 4 The above are comparison diagrams of in vitro dissolution curves of the solid dispersion of the present invention under accelerated aging conditions. (a) is the in vitro dissolution characteristic curve of the initial unaged sample, and (b) is the in vitro dissolution characteristic curve of the sample after three months of accelerated aging. Figure 5 This is a graph showing the evolution trend of the equilibrium moisture absorption and weight gain of the solid dispersion powder of the present invention. Detailed Implementation

[0033] The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0034] The main raw materials and reagents used in the following examples and comparative examples have the following sources and specifications. Reagents not specifically mentioned are all commercially available analytical grade or higher grade products.

[0035] Ashitaba chalcone extract is a conventional commercially available plant extract. Its total effective component of chalcones is greater than or equal to 80% as determined by high performance liquid chromatography. Its characteristic active components are angelicol (CAS No. 62949-76-2) and 4-hydroxydrycin (CAS No. 55912-03-3).

[0036] The CAS number for L-arginine is 74-79-3.

[0037] The CAS number for M-nicotinamide is 98-92-0.

[0038] The CAS number for NL-tartaric acid is 87-69-4.

[0039] Methacrylic acid-methyl methacrylate copolymer is a commercially available anionic polymer compound with CAS number 25086-15-1. Its macromolecular structure is formed by the random copolymerization of repeating units of methacrylic acid and methyl methacrylate through a free radical copolymerization reaction. The main chain is composed of carbon-carbon single bonds. The molar ratio of repeating units of methacrylic acid to repeating units of methyl methacrylate in the macromolecular chain is 1:1. The weight-average molecular weight is in the range of 100,000 to 135,000 Daltons, and the molecular weight distribution index is between 1.5 and 2.5. It is soluble in aqueous solutions with a pH greater than 6.0 and precipitates in aqueous solutions with a pH less than 5.0.

[0040] Preparation example: This preparation example provides a method for preparing high-purity Ashitaba chalcone extract, including the following steps: 1000 parts by weight of dried Ashitaba medicinal material was chopped and pulverized through a 40-mesh sieve to obtain coarse powder. The coarse powder was put into an extraction tank, and 8000 parts by weight of a 75% (v / v) ethanol aqueous solution was added. The mixture was extracted under reflux for 2 hours. Filter the extract, add 6000 parts by weight of 75% ethanol aqueous solution to the filtered residue, and heat under reflux for 1.5 hours. The filtrates obtained from the two extractions and filtrations were combined and concentrated under reduced pressure at a vacuum of -0.08 MPa and a temperature of 55 degrees Celsius. Ethanol was recovered until the concentrate had no alcohol odor, yielding 2000 parts by weight of aqueous extract. The aqueous extract was loaded at a flow rate of 10 parts by weight per minute into a pre-treated chromatography column packed with 3000 parts by weight of D101 macroporous adsorption resin for dynamic adsorption. After adsorption, elution was first performed using 10,000 parts by weight of purified water at a flow rate of 20 parts by weight per minute. The water eluent was discarded. Then, desorption and elution were performed using 8,000 parts by weight of an 80% ethanol aqueous solution at a flow rate of 15 parts by weight per minute. The ethanol eluent containing chalcone was collected. The collected ethanol eluent was concentrated under reduced pressure at 50°C to a thick paste, and then transferred to a vacuum drying oven. It was then vacuum dried for 24 hours at 60°C and a vacuum degree of -0.09 MPa. The dried block material was then crushed and passed through an 80-mesh sieve to obtain high-purity Ashitaba chalcone extract.

[0041] See attached document Figure 1 Example 1: This embodiment provides an amorphous solid dispersion of Ashitaba chalcone, including the following steps: S1. Pour 100 parts purified water and 40 parts anhydrous ethanol into a mixing vessel and start mechanical stirring. Under the temperature control of 20 degrees Celsius, add 4 parts L-arginine and 3 parts nicotinamide in sequence, and stir for 10 minutes until completely dissolved to form a weakly alkaline base solution with a pH of 9.0.

[0042] S2. Add 30 parts of methacrylic acid-methyl methacrylate copolymer at a mass ratio of 1:1 to the weakly alkaline base solution, and maintain the stirring speed at 200 rpm until the polymer is completely dissolved to form a transparent colloidal solution. Add 10 parts of Ashitaba chalcone extract prepared in Preparation Example 1 to the system, and stir continuously for 30 minutes in the dark to form a homogeneous dark-colored composite feed solution.

[0043] S3. In a crystallization reactor equipped with a rotor-stator high-shear homogenizer, add 300 parts of purified water. Add 8 parts of L-tartaric acid and start regular stirring to completely dissolve it. The pH of the bottom solution is measured to be 2.5. Start the reactor jacket cooling circulation to lower the temperature of the bottom solution to 5 degrees Celsius, start the high-shear homogenizer, and set the speed to 1500 rpm.

[0044] S4. A constant-flow horizontal pump is used to pump the prepared composite feed solution into the crystallization reactor. The feed solution is injected into the bottom liquid through a submerged microporous distribution ring installed directly below the high-shear impeller. The orifice diameter is 0.3 mm, and the feed flow rate is controlled at 10 parts per minute. During the fluid jetting and confluence process, a phase change occurs in the system, precipitating a co-precipitated microparticle suspension.

[0045] S5. Prepare an acidic washing solution with a pH of 2.5 by mixing 200 parts purified water with 0.1 parts L-tartaric acid, and pre-cool to 0 degrees Celsius. Pump the resulting suspension into a centrifuge and perform solid-liquid separation at a separation factor of 1000 x g, retaining the compacted filter cake. While the centrifuge is rotating, spray the pre-cooled acidic washing solution into the filter cake layer for displacement washing. Transfer the washed filter cake to a vacuum dryer and dry it for 10 hours at a temperature of 40 degrees Celsius and a vacuum of -0.08 MPa. Crush the dried lumps and pass them through an 80-mesh sieve to obtain the amorphous solid dispersion powder of Ashitaba chalcone.

[0046] Example 2: This embodiment provides an amorphous solid dispersion of Ashitaba chalcone, including the following steps: S1. Pour 125 parts purified water and 50 parts anhydrous ethanol into a mixing vessel and start mechanical stirring. At a temperature controlled at 22 degrees Celsius, add 6 parts L-arginine and 4.5 parts nicotinamide in sequence, and stir for 12 minutes until completely dissolved to form a weakly alkaline base solution with a pH of 9.2.

[0047] S2. Add 40 parts of methacrylic acid-methyl methacrylate copolymer at a mass ratio of 1:1 to the weakly alkaline base solution, and maintain the stirring speed at 250 rpm until the polymer is completely dissolved to form a transparent colloidal solution. Add 12.5 parts of Ashitaba chalcone extract prepared in Preparation Example 1 to the system, and stir continuously for 35 minutes in the dark to form a homogeneous dark-colored composite feed solution.

[0048] S3. In a crystallization reactor equipped with a rotor-stator high-shear homogenizer, add 400 parts of purified water. Add 11.5 parts of L-tartaric acid and start regular stirring to completely dissolve it. Measure the pH of the bottom solution to be 2.8. Start the reactor jacket cooling circulation to lower the temperature of the bottom solution to 8 degrees Celsius, start the high-shear homogenizer, and set the speed to 2000 rpm.

[0049] S4. A constant-flow horizontal pump is used to pump the prepared composite feed solution into the crystallization reactor. The feed solution is injected into the bottom liquid through a submerged microporous distribution ring installed directly below the high-shear impeller. The orifice diameter is 0.3 mm, and the feed flow rate is controlled at 15 parts per minute. During the fluid jetting and confluence process, a phase change occurs in the system, precipitating a co-precipitated microparticle suspension.

[0050] S5. Prepare an acidic washing solution with a pH of 2.8 by mixing 250 parts purified water with 0.3 parts L-tartaric acid, and pre-cool to 3 degrees Celsius. Pump the resulting suspension into a centrifuge and perform solid-liquid separation at a separation factor of 1500 x g, retaining the compacted filter cake. While the centrifuge is rotating, spray the pre-cooled acidic washing solution into the filter cake layer for displacement washing. Transfer the washed filter cake to a vacuum dryer and dry it for 12 hours at a temperature of 45 degrees Celsius and a vacuum of -0.09 MPa. Crush the dried lumps and pass them through a 90-mesh sieve to obtain the amorphous solid dispersion powder of Ashitaba chalcone.

[0051] Example 3: This embodiment provides an amorphous solid dispersion of Ashitaba chalcone, including the following steps: S1. Pour 150 parts purified water and 60 parts anhydrous ethanol into a mixing vessel and start mechanical stirring. At a temperature controlled at 25 degrees Celsius, add 8 parts L-arginine and 6 parts nicotinamide in sequence, and stir for 15 minutes until completely dissolved to form a weakly alkaline base solution with a pH of 9.5.

[0052] S2. Add 50 parts of methacrylic acid-methyl methacrylate copolymer at a mass ratio of 1:1 to the weakly alkaline base solution, and maintain the stirring speed at 300 rpm until the polymer is completely dissolved to form a transparent colloidal solution. Add 15 parts of Ashitaba chalcone extract prepared in Preparation Example 1 to the system, and stir continuously for 45 minutes in the dark to form a homogeneous dark-colored composite feed solution.

[0053] S3. In a crystallization reactor equipped with a rotor-stator high-shear homogenizer, add 500 parts of purified water. Add 15 parts of L-tartaric acid and start regular stirring to completely dissolve it. Measure the pH of the bottom solution to be 3.0. Start the reactor jacket cooling circulation to lower the temperature of the bottom solution to 10 degrees Celsius, start the high-shear homogenizer, and set the speed to 2500 rpm.

[0054] S4. A constant-flow horizontal pump is used to pump the prepared composite feed solution into the crystallization reactor. The feed solution is injected into the bottom liquid through a submerged microporous distribution ring installed directly below the high-shear impeller. The orifice diameter is 0.5 mm, and the feed flow rate is controlled at 20 parts per minute. During the fluid jetting and confluence process, a phase change occurs in the system, precipitating a suspension of co-precipitated microparticles.

[0055] S5. Prepare an acidic washing solution with a pH of 3.0 by mixing 300 parts purified water with 0.5 parts L-tartaric acid, and pre-cool to 5 degrees Celsius. Pump the resulting suspension into a centrifuge and perform solid-liquid separation at a separation factor of 2000 x g, retaining the compacted filter cake. While the centrifuge is rotating, spray the pre-cooled acidic washing solution into the filter cake layer for displacement washing. Transfer the washed filter cake to a vacuum dryer and dry it for 16 hours at a temperature of 50 degrees Celsius and a vacuum of -0.10 MPa. Crush the dried lumps and pass them through a 100-mesh sieve to obtain the amorphous solid dispersion powder of Ashitaba chalcone.

[0056] Comparative Example 1: The difference from Example 2 is that nicotinamide is not added in step S1, but all other steps are the same.

[0057] Comparative Example 2: Compared with Example 2, the difference is that in step S2, the methacrylic acid-methyl methacrylate copolymer is replaced by an equal mass of the conventional enteric material hydroxypropyl methylcellulose acetate succinate, while the rest are the same.

[0058] Comparative Example 3: Compared with Example 2, the difference is that in the base liquid of step S3 and the washing liquid of step S5, L-tartaric acid and other chemical equivalents are replaced with inorganic acid salts, while the rest are the same.

[0059] Comparative Example 4: Compared with Example 2, the difference is that in step S5, the acidic washing solution pre-cooled to 3 degrees Celsius is replaced with purified water at room temperature for washing, and the rest are the same.

[0060] Comparative Example 5: Compared with Example 2, the difference is that in step S1, 50 parts of anhydrous ethanol are replaced with 50 parts of purified water, and the rest are the same.

[0061] Test Example 1: Experimental objective: By comparing the example scheme with a comparative scheme lacking specific excipients or with a changed solvent system, this experiment aims to verify the actual effectiveness of the L-arginine and nicotinamide combination system in maintaining the chemical structure integrity of chalcone during the feed solution preparation stage.

[0062] Experimental subjects: Example 1, Example 2, Example 3, Comparative Example 1 and Comparative Example 5.

[0063] Experimental steps: 1. Fresh liquid samples were extracted from Examples 1 to 3, Comparative Example 1 and Comparative Example 5 at the moment the composite feed liquid was prepared, and they were dispensed into light-proof glass containers and placed in a constant temperature environment of 25 degrees Celsius.

[0064] 2. At the three test points of 0 hours, 4 hours and 8 hours of standing time, equal amounts of sample were accurately drawn from the above container, diluted with the mobile phase and injected into the high performance liquid chromatograph. The chromatographic peak area of ​​the chalcone core component was recorded, and the chalcone monomer retention rate at different time points was calculated according to the external standard method.

[0065] 3. Accurately weigh the dry solid dispersion powder collected after the preparation process of each group, add sufficient methanol solvent to it and sonicate it to destroy the polymer skeleton, and then centrifuge it at 8000 rpm for 10 minutes.

[0066] 4. Collect the supernatant obtained by centrifugation, and use high performance liquid chromatography to determine the absolute content of chalcone in it. Calculate the final product encapsulation rate by combining the initial input amount of materials and the total mass of the product.

[0067] The experimental data are shown in Table 1: Table 1: Test data on chemical stability of feed liquid and product encapsulation efficiency for each scheme

[0068] in conclusion: According to the appendix Figure 2 As shown in Table 1, the sample solutions maintained extremely high chemical stability throughout the 8-hour solution preparation and fluid transport cycle, with chalcone monomer retention rates consistently above 94%. In contrast, the results of Comparative Example 1 revealed a significant collapse in monomer retention rate after the removal of the nicotinamide component, dropping to approximately 68% by the end of the test. This discrepancy objectively reflects that while the weakly alkaline environment provided by L-arginine can disrupt the crystal lattice and achieve solubilization, it also makes the exposed chalcone skeleton a susceptible target for hydrolysis. In the examples, the added nicotinamide, with its pyridine ring structure, spontaneously binds to the aromatic conjugated system of chalcone, constructing a charge-transfer complex that effectively blocks the attack pathway of the nucleophile on the active site, thereby inhibiting reverse aldol condensation and isomerization degradation.

[0069] Comparing the data distributions of the examples and Comparative Example 5, the change in the dielectric constant of the aqueous system also had a substantial impact on the encapsulation efficiency and the stability of the base liquid. Comparative Example 5 used purified water as the sole solvent; the highly polar environment accelerated the kinetic rate of the hydrolysis side reactions, leading to a rapid decline in retention over time, and the final product encapsulation efficiency failed to exceed 80%. In the examples, the introduction of a specific proportion of anhydrous ethanol reduced the overall dielectric constant of the mixed solvent, weakening the nucleophilic attack capability of water molecules, while simultaneously improving the spread of the hydrophobic polymer in the feed liquid. This state allows the polymer segments to achieve physical capture of drug molecules with a larger surface area when they subsequently undergo a transient phase transition upon contact with the acidic receiving liquid.

[0070] Examples 1 to 3 all achieved a high retention rate of over 90%. The polymer backbone did not undergo structural degradation in the weakly alkaline composite feed solution and completed the expected hydrophobic collapse instantly upon injection into the low-pH substrate. The formed mesoscopic polymer framework tightly encapsulates the chalcone molecules, which are protected by spatial shielding, thereby successfully converting and fixing the chemical stability established in the feed solution stage into the final solid dispersion particles.

[0071] Test Example 2: Experimental objective: To investigate the practical effects of specific small molecule excipients, hydrolysis-resistant polymers, and mixed solvent systems in preventing chemical degradation of active ingredients and ensuring the macroscopic phase transition retention efficiency of the system by comparing the example schemes with comparative schemes that replace or omit specific process components.

[0072] Experimental subjects: Example 2, Comparative Example 1, Comparative Example 2 and Comparative Example 5.

[0073] Experimental steps: 1. Collect the final solid products of Examples 2, 1, 2, and 5 after vacuum drying, pulverization, and sieving. Accurately weigh the total mass of the powder actually obtained using an analytical balance. Calculate the overall preparation yield of each process based on the theoretical total dry mass of the raw materials added during the solution preparation stage for each group.

[0074] 2. Accurately weigh appropriate amounts of sample from each of the above-mentioned solid dispersion powders, place them in volumetric flasks, add methanol, and sonicate at room temperature for 20 minutes to completely depolymerize the polymer backbone encapsulating the drug and release the internal components.

[0075] 3. The ultrasonically treated suspension was filtered through an organic microporous membrane with a pore size of 0.22 micrometers to remove insoluble excipient residues, and the clear filtrate was collected as the test solution.

[0076] 4. Inject each group of test solutions into the high-performance liquid chromatograph, set the detection wavelength to the characteristic absorption peak of chalcone, and record the complete chromatogram. Using the area normalization method, with the peak area of ​​the main component of chalcone as a reference, calculate the total area ratio of all relevant degradation impurity peaks in the chromatogram to obtain the total degradation rate of impurities.

[0077] The experimental data are shown in Table 2: Table 2: Test data on the yield of solid dispersions and the degradation rate of active ingredients

[0078] in conclusion: According to the appendix Figure 3 As shown in Table 2, Example 2 demonstrated excellent material retention and chemical protection throughout the entire preparation process, achieving an actual process yield of over 92%, with the total impurity degradation rate strictly controlled at an extremely low level. In contrast, comparing the test results of Comparative Example 2, when the polymer backbone in the system was replaced with the conventional enteric material hydroxypropyl methylcellulose acetate succinate, the process yield plummeted to only 42.57%. This significant loss of macroscopic materials stemmed from the fact that conventional enteric polymers are unable to withstand the weakly alkaline feed solution environment. The ester bonds on the polymer chains underwent irreversible saponification and hydrolysis, transforming into fully water-soluble substances. The polymer, having lost its phase change precipitation capability, could not undergo hydrophobic collapse when injected into the acidic substrate, causing the drug monomers that should have been encapsulated to be directly exposed and dissolved or lost with the washing liquid. This, in turn, verifies the absolute necessity of methacrylic acid-methyl methacrylate copolymer in resisting alkaline hydrolysis and ensuring the formation of the co-precipitated backbone.

[0079] Comparative Example 1 exhibited chemical degradation, with the total degradation rate of impurities increasing to 15.12%. L-arginine, in providing proton exchange to achieve chalcone lattice disruption and solubilization, inevitably stimulated the reactivity of the unsaturated ketone structure in the substrate molecule. The examples introduced nicotinamide to construct a space charge-transfer complex, forming a shielding layer around the easily hydrolyzed sites, thereby effectively inhibiting the reverse aldol condensation reaction pathway. In the absence of this supramolecular protection mechanism, the exposed chalcone monomer in Comparative Example 1 suffered continuous chemical degradation in an alkaline liquid environment, demonstrating the crucial role of small molecule auxiliary components in maintaining the structural stability of sensitive drugs.

[0080] Comparative Example 5 used purified water to prepare the feed solution, eliminating the effect of anhydrous ethanol on the dielectric constant of the system. This resulted in a higher impurity degradation rate compared to the example, while the product yield decreased to 78.54%. The highly polar aqueous environment not only reduced the thermodynamic stability of the substrate and accelerated the collision frequency of the hydrolysis reaction, but also affected the extended morphology of the anionic copolymer segments at the microscopic level. The polymer prematurely underwent local coiling before reaching the optimal solvation state, weakening the specific surface area for capturing drug molecules upon injection into the substrate solution, causing some drug to escape at the coprecipitation interface.

[0081] Test Example 3: Experimental objective: By comparing the examples with a comparative example using inorganic acids as an alternative, this study aims to verify the effectiveness of small organic molecules in blocking the translational and rotational degrees of freedom of active ingredients and inhibiting the physical recrystallization evolution of the system under humid and hot storage conditions.

[0082] Experimental subjects: Example 2 and Comparative Example 3.

[0083] Experimental steps: 1. Take fresh powder samples from Example 2 and Comparative Example 3 that have just been prepared, and divide them into two equal parts. One part is used as the initial test sample and is properly sealed and protected from light. The other part is laid flat in a large-diameter petri dish and placed in a constant temperature and humidity test chamber with a set temperature of 40 degrees Celsius and a relative humidity of 75 percent for accelerated aging treatment for three months.

[0084] 2. Following the paddle method operation standard in the dissolution test method specified in the pharmacopoeia, 900 ml of degassed phosphate buffer with a pH of 6.8 was used as the simulated intestinal dissolution medium. The water bath temperature of the dissolution apparatus was kept constant at 37 degrees Celsius, and the paddle speed was set to 75 revolutions per minute.

[0085] 3. Accurately weigh the initial and aged samples of the examples and comparative examples containing 50 mg of Ashitaba chalcone active ingredient, and quickly put them into the corresponding dissolution cups that are in operation.

[0086] 4. At the set time points of 5 minutes, 10 minutes, 20 minutes, 30 minutes, 45 minutes and 60 minutes when the instrument is running, use a syringe equipped with a 0.45 micrometer filter membrane to automatically draw 5 ml of dissolution medium and immediately add an equal volume of blank phosphate buffer at the same temperature to the dissolution vessel.

[0087] 5. The sampled and filtered test solutions at each time point were directly injected into the high performance liquid chromatograph to determine the concentration of chalcone monomer. The cumulative dissolution percentage was calculated by volume conversion, and the release evolution trajectory from time to dissolution was plotted.

[0088] The experimental data are shown in Table 3: Table 3: Data on Cumulative Dissolution Rate of Solid Dispersions Before and After Accelerated Aging

[0089] in conclusion: According to the appendix Figure 4 As shown in Table 3, both Example 2 and Comparative Example 3 exhibited highly consistent rapid release characteristics in their initial, unaged state. Within the first 30 minutes after introduction into the simulated intestinal fluid dissolution medium, the cumulative release rate of the drug monomers in both groups successfully climbed to over 90%. This confirms that during the rigid phase transition coprecipitation stage using different types of acidic activators, the polymer copolymers can uniformly disperse the drug substrate in an amorphous state and fix it within the matrix network. The high-energy-distribution amorphous microstructure eliminates the original lattice energy dissolution barrier, enabling the free diffusion and dissolution of active molecules into the aqueous medium through the permeation of water molecules.

[0090] The dissolution trajectory of Comparative Example 3 exhibited an irreversible, precipitous decline, with its cumulative release rate barely reaching 54.89% at the end of the 60-minute test. Using hydrochloric acid as a single inorganic neutralizing agent in the substrate, while macroscopically inducing the transient protonation and precipitation of the polymer backbone, the tiny chloride and hydrogen ions could not provide any spatial intermolecular binding forces within the polymer matrix. The chalcone monomers, in a thermodynamically metastable state, broke through the activation energy barrier after continuously absorbing energy from the external humid and hot environment. In situ translation and rotational rearrangement occurred within the free pores of the microscopic polymer network, leading to spontaneous molecular polymerization and the formation of a stable crystal structure with low-energy characteristics. The regeneration of lattice energy directly blocked the rapid dissociation path of aging products in the dissolution medium.

[0091] Example 2 maintained a dissolution performance highly consistent with the initial state even after the accelerated aging cycle, with the final drug release rate remaining stably in the excellent range of 95%. The introduced L-tartaric acid, with its two free carboxyl groups and two adjacent hydroxyl structures, acts as a supramolecular-scale multidentate bridging element in the coprecipitation microregion. At the moment of fluid intersection and interception, this organic acid molecule utilizes its abundant hydrogen bond donor and acceptor sites to directionally bind to the free nicotinamide amide groups and carboxyl residues on the methacrylic acid copolymer segments in the coprecipitation system, constructing an extremely dense three-dimensional cross-linked network.

[0092] Test Example 4: Experimental objective: By comparing the effects of specific low-temperature acidic washing solution and conventional room-temperature purified water washing in the post-treatment stage of solid dispersions in the examples and comparative examples, this study aims to verify the feasibility and necessity of displacement washing process in removing hygroscopic reactive waste salts, preventing mass transfer and penetration of the washing solution into the polymer framework, and maintaining the long-term physical stability of the powder.

[0093] Experimental subjects: Example 2 and Comparative Example 4.

[0094] Experimental steps: 1. Take a clean and dry flat glass weighing bottle in advance and accurately weigh the empty bottle on an analytical balance. Take the solid dispersion powder of Example 2 and Comparative Example 4 that has just been vacuum dried and sieved, spread it evenly on the bottom of the weighing bottle, so that the powder layer thickness is maintained at about 2 mm, and accurately weigh the initial mass of each sample.

[0095] 2. Smoothly transfer the open weighing bottle into a sealed, temperature-controlled glass desiccator containing a saturated potassium chloride solution at the bottom. Set the internal temperature of the desiccator to 25 degrees Celsius and maintain a constant relative humidity of 80%. Start the timer.

[0096] 3. At the set time points of 24 hours, 72 hours, 120 hours and 168 hours after placement, quickly cover the weighing bottle and remove it. Weigh the current total mass on the analytical balance. After weighing, immediately leave the bottle open and return it to the original high humidity environment for continued exposure.

[0097] 4. Calculate the absolute moisture absorption of the powder by subtracting the mass of the empty bottle from the mass measured at each time point, and then calculate the percentage increase in weight based on the initial powder mass. At the end of the 168-hour test, pour out the powder, touch the powder surface with a stainless steel spatula, and record its changes in macroscopic physical state and flowability.

[0098] The experimental data are shown in Table 4: Table 4: Test Data on Equilibrium Moisture Absorption Weight Gain and Appearance Changes of Powders

[0099] in conclusion: According to the appendix Figure 5 According to the data in Table 4, the powder of Example 2 demonstrated moisture resistance during a 168-hour high-humidity exposure test. Its moisture absorption weight gain rate plateaued after reaching 1.3-1.4%, and the powder maintained a good loose and dispersed state macroscopically. In contrast, the test results of Comparative Example 4 showed that the method of washing the compacted filter cake with purified water at room temperature resulted in moisture accumulation in the later moisture absorption test. The weight gain rate of this group of samples showed a steep upward trend with exposure time, eventually exceeding 14%, indicating significant deliquescence and agglomeration after the powder surface absorbed a large amount of moisture.

[0100] The hygroscopic collapse observed in Comparative Example 4 stems from the disruption of the microscopic water-repellent barrier in the co-precipitation system caused by room-temperature neutral water washing. During the phase transition and convergence of the bottom liquid, a large amount of arginine tartrate, a byproduct, was generated within the system. This waste salt possesses an extremely strong affinity for water, and if it remains within the porous framework, it will become the capillary core for the permeation of environmental moisture. The pH of the room-temperature purified water is close to the dissociation critical point of the methacrylic acid copolymer, causing localized hydration and swelling of the polymer segments when the washing liquid passes through the filter cake. The expanded polymer network not only leads to the reverse permeation and loss of a large number of water-soluble nicotinamide molecules, but also causes the dissolved waste salt carried by the washing water to be deeply trapped inside the mesopores. In the subsequent vacuum drying stage, the moisture is forcibly removed, but the waste salt crystals remain deep within the particles, becoming a ticking time bomb for deliquescence in subsequent high-humidity storage environments.

[0101] The embodiment employs a cold, acidic solution near its critical freezing point for displacement washing, effectively blocking the aforementioned negative mass transfer pathways from both hydrodynamic and polymer thermodynamic perspectives. The low pH environment forcibly maintains the fully protonated state of the free carboxyl groups on the methacrylic acid chains. Combined with the low temperature of 3 degrees Celsius, the polymer molecular chains exhibit a highly contracted glassy state. An extremely dense, hydrophobic skin forms on the surface of the precipitated particles, preventing any substantial mass exchange between the washing liquid and the particles. The fluid can only rapidly flush away free waste salts adhering to the macroscopic gaps in the compacted filter cake in a plug flow manner. This displacement washing mechanism, which isolates surface rinsing from internal shielding, not only removes external impurities that cause deliquescence but also completely locks in small-molecule excipients and active pharmaceutical ingredients encapsulated deep within the microscopic network, ensuring the phase stability of the final powder under complex temperature and humidity alternating environments.

[0102] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. The application of Ashitaba chalcone extract in the preparation of drugs for the intervention of metabolism-related fatty liver disease, characterized in that, The intervention drug is made from raw materials comprising the following parts by weight: Ashitaba chalcone extract: 10-15 parts; 30-50 parts of methacrylic acid-methyl methacrylate copolymer; L-arginine: 4-8 parts; Nicotinamide: 3-6 parts; L-tartaric acid: 8.1-15.5 parts.

2. The application of the Ashitaba chalcone extract according to claim 1 in the preparation of an intervention drug for metabolism-related fatty liver disease, characterized in that, The mass ratio of methacrylic acid to methyl methacrylate in the methacrylic acid-methyl methacrylate copolymer is 1:

1.

3. The application of the Ashitaba chalcone extract according to claim 1 in the preparation of an intervention drug for metabolism-related fatty liver disease, characterized in that, The Ashitaba chalcone extract was prepared by the following steps: The crude powder of Ashitaba was extracted twice by heating and reflux with a 75% ethanol aqueous solution. The filtrates were combined and concentrated under reduced pressure to obtain an aqueous extract. The aqueous extract was dynamically adsorbed onto an adsorption resin chromatography column and then eluted sequentially with purified water and an aqueous ethanol solution. The ethanol eluent was collected, concentrated under reduced pressure, dried under vacuum, pulverized, and sieved to obtain the final product.

4. The application of the Ashitaba chalcone extract according to claim 1 in the preparation of an intervention drug for metabolism-related fatty liver disease, characterized in that, The chalcone molecules in the Ashitaba chalcone extract are dispersed in an amorphous state within a mesoporous framework formed by the hydrophobic collapse of the methacrylic acid-methyl methacrylate copolymer.

5. The use of the Ashitaba chalcone extract according to any one of claims 1-4 in the preparation of an intervention drug for metabolism-related fatty liver disease, characterized in that, The intervention drug is an amorphous solid dispersion of Ashitaba chalcone, and the preparation process of the intervention drug includes the following steps: S1. Mix purified water and anhydrous ethanol, add L-arginine and nicotinamide and stir to dissolve to form a weakly alkaline base solution; S2. Add methacrylic acid-methyl methacrylate copolymer to the weakly alkaline base solution, stir to dissolve and form a colloidal solution, then add Ashitaba chalcone extract and stir in the dark to form a composite feed solution. S3. Add purified water to the crystallization reactor, dissolve it with L-tartaric acid to form an acidic bottom solution, cool it down and start the high-shear homogenizer; S4. The composite feed liquid is pumped into the crystallization reactor and injected into the acidic bottom liquid through the submerged microporous distribution ring, and a co-precipitated microparticle suspension is precipitated in the fluid jet confluence. S5. The obtained suspension is subjected to solid-liquid separation to retain the filter cake. The filter cake is washed by replacing it with an acidic washing solution prepared with purified water and L-tartaric acid and pre-cooled. The washed filter cake is then vacuum dried, crushed and sieved to obtain the amorphous solid dispersion of Ashitaba chalcone.

6. The application of the Ashitaba chalcone extract according to claim 5 in the preparation of an intervention drug for metabolism-related fatty liver disease, characterized in that, In step S1, the mass ratio of purified water to anhydrous ethanol is 100-150:40-60, the system temperature is controlled at 20-25℃, and the pH value of the weakly alkaline base solution is 9.0-9.

5. In step S2, the addition of Ashitaba chalcone extract and stirring time in the dark is 30-45 minutes.

7. The process for preparing a drug to intervene in metabolism-related fatty liver disease using chalcone of Ashitaba according to claim 5, characterized in that, In step S3, the pH value of the acidic substrate is 2.5-3.0, the temperature of the substrate is reduced to 5-10℃, and the speed of the high shear homogenizer is set to 1500-2500 rpm. In step S4, the submerged microporous distribution ring is installed directly below the high-shear impeller, with a pore diameter of 0.3-0.5 mm, and the feed flow rate of the composite feed liquid is 10-20 parts per minute.

8. The application of the Ashitaba chalcone extract according to claim 5 in the preparation of an intervention drug for metabolism-related fatty liver disease, characterized in that, In step S5, the pH value of the pre-cooled acidic washing solution is 2.5-3.0, and the pre-cooling temperature is 0-5℃.

9. The application of the Ashitaba chalcone extract according to claim 5 in the preparation of an intervention drug for metabolism-related fatty liver disease, characterized in that, In step S5, the obtained suspension is subjected to solid-liquid separation, with the separation factor set to 1000-2000xg. Acidic washing liquid is sprayed into the centrifuge while it is rotating. The conditions for vacuum drying are a temperature of 40-50℃, a vacuum degree of -0.08 to -0.10 MPa, and a drying time of 10-16 hours.

10. The use of the Ashitaba chalcone extract according to any one of claims 1-9 in the preparation of an intervention drug for metabolism-related fatty liver disease.

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