Composite drug-loaded nanoparticle based on acylated polysaccharide and berberine as well as preparation method and application of composite drug-loaded nanoparticle

Abstract

The invention relates to the technical field of nano-medicine carriers, in particular to composite drug-loaded nanoparticles based on acylated polysaccharide and berberine as well as a preparation method and application of the composite drug-loaded nanoparticles. The preparation method comprises the following steps: carrying out acylation modification on natural polysaccharide by using a basic catalyst and acid anhydride to obtain acylated natural polysaccharide; wherein the acylation modification mode comprises acetylation modification or propionylation modification; in an organic solvent, the berberine and the acylated natural polysaccharide are subjected to encapsulation treatment, and the composite drug-loaded nanoparticles based on the acylated polysaccharide and the berberine are obtained. According to the preparation method, the natural polysaccharide is subjected to acylation modification, the hydrophobicity of the natural polysaccharide is adjusted, and the encapsulation efficiency of berberine is remarkably improved. On one hand, the composite drug-loaded nanoparticles can significantly improve the bioavailability of berberine and prolong the in-vivo action time of berberine, and on the other hand, the composite drug-loaded nanoparticles can promote the repair and healing of wounds infected by staphylococcus aureus and reduce the drug resistance of staphylococcus aureus.

Description

Technical Field

[0001] This application relates to the field of nanomedicine carrier technology, and in particular to a composite drug-loaded nanoparticle based on acylated polysaccharide and berberine, its preparation method, and its application. Background Technology

[0002] Skin damage or injury is common in daily life. If broken skin is exposed to air, it is extremely easy to cause skin infection. If the skin infection is not treated in time, the wound will worsen, which may lead to amputation or sepsis, or even threaten the patient's life. At present, wound infection is generally treated with a combination of oral and topical antibiotics. However, long-term use of antibiotics can easily lead to antibiotic resistance. Drug-resistant bacteria mainly resist the destruction of antibiotics through three lines of defense: (1) Drug-resistant bacteria produce drug pumps that pump antibiotics out of the bacterial cells to prevent antibiotics from accumulating in the bacterial cells; (2) Drug-resistant bacteria modify antibiotic target molecules to reduce the binding of antibiotics to target molecules, thereby weakening the antibacterial ability of antibiotics; (3) Drug-resistant bacteria synthesize enzymes that can decompose antibiotics, making antibiotics inactive. Based on the above three-line defense mechanism, drug-resistant bacteria have become one of the challenges of current public health, and it has been confirmed that drug-resistant bacteria are the third leading cause of death worldwide. The World Health Organization has clearly stated that antibiotic overuse and misuse are the main factors driving the development of bacterial resistance, which promotes the continuous evolution of drug-resistant bacteria such as MRSA, leading to the failure of anti-infective treatment.

[0003] Berberine is an isoquinoline alkaloid extracted from traditional medicinal plants such as Coptis chinensis and Phellodendron amurense. It possesses a broad spectrum of antibacterial and antifungal activity. Its antibacterial mechanisms include disrupting bacterial cell membrane integrity, inhibiting bacterial metabolic activity, inhibiting bacterial protease activity, inhibiting bacterial protease expression, and preventing DNA replication—a multi-target inhibitory effect. Berberine can not only directly inhibit pathogens but also act as an adjuvant to antibiotics against drug-resistant bacteria. Therefore, berberine is a promising candidate drug for combating current drug-resistant bacteria. Due to its multi-target inhibitory mechanism, it is less likely to induce bacterial resistance. Furthermore, berberine exhibits significant anti-inflammatory and pro-repair effects, downregulating the expression of pro-inflammatory factors and the production of reactive oxygen species (ROS) to reduce the damage to tissues caused by excessive inflammation, thus supporting the transition of wounds from the "inflammatory phase" to the "repair phase." However, berberine has limited antibacterial ability, and its rapid metabolism, wide range of metabolic pathways, and low oral bioavailability limit its application in wound treatment.

[0004] Natural polysaccharides possess excellent biocompatibility and biodegradability. Their degradation products in vivo are small-molecule sugars, which can be metabolized and absorbed by the body. Many natural polysaccharides exhibit antibacterial, antioxidant, anti-inflammatory, and moisturizing activities, and also show some inhibitory effects on certain bacteria. They form a physical barrier on wound surfaces, reducing moisture loss and regulating the release of inflammatory factors, thus laying the foundation for wound healing. Compared to traditional synthetic carriers (such as some chemical polymers), natural polysaccharides have advantages in biocompatibility and wound adaptability; however, their antibacterial and antioxidant capabilities are limited.

[0005] Therefore, combining berberine with natural polysaccharides can have a synergistic effect, promoting wound healing, prolonging the duration of action, and improving efficacy. However, berberine contains five-membered rings such as pyridine and benzene rings, exhibiting strong hydrophobicity, making it difficult to bind with highly hydrophilic natural polysaccharides. This results in extremely low encapsulation efficiency between berberine and natural polysaccharides, affecting the drug loading rate and stability. Summary of the Invention

[0006] This application provides a composite drug-loaded nanoparticle based on acylated polysaccharides and berberine, its preparation method, and its application, in order to solve the technical problem of low encapsulation efficiency between berberine and natural polysaccharides.

[0007] In a first aspect, embodiments of this application provide a method for preparing composite drug-loaded nanoparticles based on acylated polysaccharides and berberine, the preparation method comprising: Acylation modification of natural polysaccharides using an alkaline catalyst and acid anhydride is used to enhance the hydrophobicity of the natural polysaccharides, resulting in acylated natural polysaccharides; wherein the acylation modification includes acetylation modification or propionylation modification. Berberine and the acylated natural polysaccharide were encapsulated in an organic solvent to obtain composite drug-loaded nanoparticles based on acylated polysaccharide and berberine.

[0008] Optionally, when the acylation modification is acetylation modification, the acid anhydride is acetic anhydride, and the mass m1 of the natural polysaccharide, the mass m2 of the acetic anhydride and the mass m3 of the alkaline catalyst satisfy: m1:m2:m3=1:(2 to 5):(2 to 10).

[0009] Optionally, when the acylation modification is acetylation modification, the acylation modification temperature is 50°C to 120°C, and the acylation modification time is 20 min to 1440 min.

[0010] Optionally, when the acylation modification is propionylation modification, the acid anhydride is propionic anhydride, and the mass m1 of the natural polysaccharide, the mass m4 of the propionic anhydride and the mass m3 of the alkaline catalyst satisfy: m1:m4:m3=1:(1 to 5):(2 to 10).

[0011] Optionally, when the acylation modification is propionylation modification, the acylation modification temperature is 60°C to 120°C, and the acylation modification time is 30 min to 1440 min.

[0012] Optionally, the mass m5 of the berberine and the mass m6 of the acylated natural polysaccharide satisfy the following condition: m5:m6 = 1:(2 to 5).

[0013] Optionally, the encapsulation temperature is 30°C to 80°C, and the encapsulation time is 30 min to 1440 min.

[0014] Optionally, the alkaline catalyst may include at least one of pyridine, 4-dimethylaminopyridine, triethylamine, N,N-diisopropylethylamine, and trimethylpyridine; and / or The types of natural polysaccharides include at least one of tobacco polysaccharide, wolfberry polysaccharide, astragalus polysaccharide, and pomegranate peel polysaccharide; and / or The organic solvents include at least one of ethanol, methanol, acetone and ethyl acetate.

[0015] Secondly, embodiments of this application provide a composite drug-loaded nanoparticle based on acylated polysaccharide and berberine, wherein the composite drug-loaded nanoparticle is prepared by the preparation method described in the first aspect; the composite drug-loaded nanoparticle includes a natural polysaccharide matrix and berberine loaded on the natural polysaccharide matrix.

[0016] Thirdly, embodiments of this application provide an application of composite drug-loaded nanoparticles based on acylated polysaccharides and berberine, the application including using the composite drug-loaded nanoparticles described in the second aspect for the treatment of wound infection, diarrhea, enteritis or mucosal ulceration.

[0017] The technical solutions provided in this application have the following advantages compared with the prior art: This application provides a method for preparing composite drug-loaded nanoparticles based on acylated polysaccharides and berberine. The method first involves acylation modification of natural polysaccharides using acid anhydrides under alkaline catalysts. Hydrophobic groups such as acetyl or propionyl groups are introduced onto the molecular chain of the natural polysaccharides, significantly improving their lipophilicity and overall hydrophobicity. This results in the modified acylated polysaccharides possessing hydrophobic properties, thus meeting the carrier requirements of berberine. Subsequently, berberine and the acylated natural polysaccharides are encapsulated in an organic solvent. Due to the hydrophobicity of both, through hydrophobic interactions, berberine can be more fully and stably bound to the molecular structure of the acylated polysaccharides, forming a denser and more stable nanoparticle microstructure, allowing more berberine to be encapsulated within the acylated natural polysaccharides. This preparation method significantly improves the encapsulation efficiency of berberine by acylated natural polysaccharides through both the binding effect of the carrier and the encapsulation effect of nanoparticles, solving the key problem that natural polysaccharides, due to their high hydrophilicity, cannot effectively encapsulate hydrophobic berberine. On the one hand, these composite drug-loaded nanoparticles can significantly improve the bioavailability of berberine and prolong its duration of action in vivo; on the other hand, they can promote the repair and healing of wounds infected with Staphylococcus aureus while reducing the development of drug resistance in this bacterium. Attached Figure Description

[0018] The accompanying drawings, which are incorporated in and form part of this specification, illustrate embodiments consistent with this application and, together with the description, serve to explain the principles of this application.

[0019] To more clearly illustrate the technical solutions in the embodiments of this application or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, for those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0020] Figure 1 A schematic diagram of a method for preparing composite drug-loaded nanoparticles based on acylated polysaccharides and berberine, provided for embodiments of this application; Figure 2 The antibacterial activity of different berberine formulations provided in this application against S. aureus is shown in the graph. Figure 3 The antibacterial activity of different berberine formulations provided in this application against MRSA is shown in the graph. Figure 4 Pharmacokinetic results of different berberine formulations provided in the embodiments of this application; Figure 5 The images provided in this application illustrate the healing process of a circular wound on a mouse at different time points. Figure 5 (a) is an observation of the healing of a circular wound in a mouse at different time points. Figure 5(b) is a bar chart showing the healing rate of circular wounds in mice at different time points; in the figure, *, *, ** represent comparisons between each group and the model group, p<0.05, p<0.01, p<0.001 respectively; #, ##, ### represent comparisons between groups, p<0.05, p<0.01, p<0.001 respectively; Figure 6 Images of Masson trichrome stained sections of a mouse circular wound at different time points, provided in an embodiment of this application. Figure 7 The figures show the VEGF fluorescence staining results and quantitative bar graphs of circular wounds in mice at different time periods provided in the embodiments of this application. In the figures, *, *, and * represent comparisons between each group and the model group, respectively, with p < 0.05, p < 0.01, and p < 0.001; #, ##, and ### represent comparisons between each group (excluding the model group), respectively, with p < 0.05, p < 0.01, and p < 0.001. Detailed Implementation

[0021] To make the objectives, technical solutions, and advantages of the embodiments of this application clearer, the technical solutions of the embodiments of this application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of this application, not all embodiments. Based on the embodiments of this application, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this application.

[0022] The range descriptions used in this application, such as numerical ranges and proportional ranges, include all possible sub-ranges and single numerical values ​​within that range. For example, the range descriptions of "1 to 6" or "1~6" cover all sub-ranges (such as 1 to 3, 2 to 5, etc.) and single numbers (such as 1, 2, 3, 4, 5, 6) between 1 and 6. Unless otherwise specified, the terms "comprising" and others used herein mean "including but not limited to"; relational terms such as "first" and "second" are used only to distinguish different entities or operations and do not imply an actual order or relationship; "and / or" indicates that multiple situations can exist individually or simultaneously; expressions such as "at least one," "multiple," and "at least one" refer to any combination of the corresponding objects, including combinations of single or multiple objects. The proportional relationships involved in this document, such as mass ratios and molar ratios, should be understood as the correspondence between the first and second terms of a proportional formula, according to the order of description. The raw materials, reagents, instruments, and equipment used herein can all be obtained by purchasing from the market or by existing methods.

[0023] It should be noted that berberine is characterized by rapid metabolism in vivo, while natural polysaccharides have insufficient antibacterial activity. Therefore, if natural polysaccharides can be used as carriers for berberine, a synergistic effect between the two can be achieved: berberine acts as an antibacterial agent, while natural polysaccharides promote wound healing. Furthermore, with natural polysaccharides as carriers, the metabolic degradation rate of berberine in vivo can be reduced, prolonging its duration of action and thus improving its efficacy. However, because berberine generally contains five-membered rings such as pyridine and benzene rings, exhibiting strong hydrophobicity, while natural polysaccharides contain numerous polar groups, making them highly hydrophilic, the highly hydrophobic berberine and the highly hydrophilic natural polysaccharide are difficult to bind, resulting in extremely low encapsulation efficiency between berberine and natural polysaccharides. Therefore, it is currently necessary to modify natural polysaccharides to increase their hydrophobicity, improve their encapsulation efficiency of berberine, and thus enhance the drug loading stability of berberine.

[0024] In view of the above-mentioned deficiencies of the prior art, the embodiments of this application provide the following technical solutions: Figure 1 An exemplary schematic diagram of a method for preparing composite drug-loaded nanoparticles based on acylated polysaccharides and berberine provided in this application embodiment is shown. like Figure 1 As shown in the embodiments of this application, a method for preparing composite drug-loaded nanoparticles based on acylated polysaccharides and berberine is provided, the preparation method comprising: S1. Using an alkaline catalyst and an acid anhydride to acylate and modify a natural polysaccharide to enhance its hydrophobicity, thereby obtaining an acylated natural polysaccharide; wherein the acylation modification includes acetylation or propionylation. S2. Berberine and the acylated natural polysaccharide are encapsulated in an organic solvent to obtain composite drug-loaded nanoparticles based on acylated polysaccharide and berberine.

[0025] It should be noted that the natural polysaccharide before acylation modification can be dissolved and dispersed using N,N-dimethylformamide, which is beneficial for the acylation modification between the alkaline catalyst and the acid anhydride and the natural polysaccharide.

[0026] It should be noted that the acylated natural polysaccharide obtained by this acylation modification can be purified by dialysis for 24 hours, and then the dialysis solution can be concentrated and freeze-dried to obtain high-purity acylated natural polysaccharide.

[0027] It should be noted that before this encapsulation process, berberine and acylated natural polysaccharides can be dispersed using an organic solvent, and then encapsulated by stirring and mixing.

[0028] It should be noted that after the encapsulation process is completed, the organic solvent in the composite drug-loaded nanoparticles can be removed by vacuum distillation. Then, the composite drug-loaded nanoparticles can be redispersed using distilled water, phosphate buffer, or physiological saline to obtain high-purity composite drug-loaded nanoparticles.

[0029] It should be noted that this application provides a method for preparing composite drug-loaded nanoparticles based on acylated polysaccharides and berberine. This method first modifies the natural polysaccharide by acylation to control the hydrophobicity of the carrier, thereby significantly improving the encapsulation efficiency of berberine. The core of this method revolves around the compatibility matching between the drug and the carrier. The specific implementation path is as follows: This application addresses the core problem of low encapsulation efficiency caused by the poor compatibility of highly hydrophilic natural polysaccharides with hydrophobic berberine. By using an alkaline catalyst and acid anhydride to acetylate or propionylate natural polysaccharides, hydrophobic acetyl or propionyl groups are introduced into the molecular chain of the natural polysaccharides, directly altering their original physicochemical properties and significantly improving their lipid solubility and overall hydrophobicity. This results in modified acylated polysaccharides with hydrophobic properties, making them suitable as carriers for berberine.

[0030] In an organic solvent encapsulation system, the hydrophobic interaction between acylated polysaccharide carriers and berberine is significantly enhanced through hydrophobic matching. Berberine can be more fully and stably bound to the molecular structure of the acylated polysaccharide, rather than simply being physically mixed. At the same time, the enhanced hydrophobicity of the acylated polysaccharide can form a denser and more stable nanoparticle microstructure in the system, which can encapsulate more berberine inside the nanoparticles. From the perspectives of both the binding ability of the carrier to the drug and the encapsulation effect of the nanoparticles, the encapsulation rate of berberine is greatly improved, solving the key problem that natural polysaccharides cannot effectively encapsulate hydrophobic berberine due to their high hydrophilicity.

[0031] In some alternative embodiments, when the acylation modification is acetylation modification, the acid anhydride is acetic anhydride, and the mass m1 of the natural polysaccharide, the mass m2 of the acetic anhydride and the mass m3 of the basic catalyst satisfy: m1:m2:m3=1:(2 to 5):(2 to 10).

[0032] In these embodiments, when the acylation modification is acetylation modification, the acid anhydride is acetic anhydride, and the mass ratio of natural polysaccharide, acetic anhydride and alkaline catalyst is controlled to be 1:(2 to 5):(2 to 10). This ensures that there is sufficient acetic anhydride and alkaline catalyst in the acylation modification stage. Under the action of sufficient alkaline catalyst, natural polysaccharide and acetic anhydride will react to introduce hydrophobic acetyl groups into the molecular chain of natural polysaccharide, thereby improving the hydrophobicity of natural polysaccharide and meeting the carrier requirements of berberine.

[0033] The mass m2 of the acetic anhydride can be 2, 3, 4 or 5.

[0034] The mass m3 of the alkaline catalyst can be 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0035] In some alternative embodiments, when the acylation modification is acetylation modification, the acylation modification temperature is 50°C to 120°C, and the acylation modification time is 20 min to 1440 min.

[0036] In these embodiments, when the acylation modification is acetylation modification, the acylation modification at a temperature of 50°C to 120°C and a time of 20 min to 1440 min can introduce hydrophobic acetyl groups into the molecular backbone of natural polysaccharides under the action of acetic anhydride and a basic catalyst, thereby improving the hydrophobicity of natural polysaccharides to meet the carrier requirements of berberine.

[0037] The acylation modification temperature can be 50℃, 60℃, 70℃, 80℃, 90℃, 100℃, 110℃ or 120℃.

[0038] The acylation modification time can be 20 min, 30 min, 40 min, 50 min, 100 min, 150 min, 200 min, 250 min, 500 min, 600 min, 700 min, 800 min, 1000 min, 1200 min, 1400 min or 1440 min.

[0039] In some alternative embodiments, when the acylation modification is propionylation modification, the acid anhydride is propionic anhydride, and the mass m1 of the natural polysaccharide, the mass m4 of the propionic anhydride and the mass m3 of the basic catalyst satisfy: m1:m4:m3=1:(1 to 5):(2 to 10).

[0040] In these embodiments, when the acylation modification is propionylation, the acid anhydride is propionic anhydride, and the mass ratio of natural polysaccharide, propionic anhydride, and alkaline catalyst is controlled to be 1:(1 to 5):(2 to 10). This ensures that there is sufficient propionic anhydride and alkaline catalyst in the acylation modification stage. Under the action of sufficient alkaline catalyst, natural polysaccharide and propionic anhydride will react to introduce hydrophobic propionyl groups into the molecular chain of natural polysaccharide, thereby improving the hydrophobicity of natural polysaccharide and meeting the carrier requirements of berberine.

[0041] The mass m4 of the propionic anhydride can be 1, 2, 3, 4 or 5.

[0042] The mass m3 of the alkaline catalyst can be 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0043] In some alternative embodiments, when the acylation modification is propionylation modification, the acylation modification temperature is 60°C to 120°C, and the acylation modification time is 30 min to 1440 min.

[0044] In these embodiments, when the acylation modification is propionylation, the acylation modification at a temperature of 60°C to 120°C and a time of 30 min to 1440 min can introduce the hydrophobic propionyl group into the molecular backbone of the natural polysaccharide under the action of acetic anhydride and a basic catalyst, thereby improving the hydrophobicity of the natural polysaccharide to meet the carrier requirements of berberine.

[0045] The acylation modification temperature can be 60℃, 70℃, 80℃, 90℃, 100℃, 110℃ or 120℃.

[0046] The acylation modification time can be 30 min, 40 min, 50 min, 100 min, 150 min, 200 min, 250 min, 500 min, 600 min, 700 min, 800 min, 1000 min, 1200 min, 1400 min or 1440 min.

[0047] In some alternative embodiments, the mass m5 of the berberine and the mass m6 of the acylated natural polysaccharide satisfy the following: m5:m6 = 1:(2 to 5).

[0048] In these embodiments, a mass ratio of berberine to acylated natural polysaccharide of 1:(2 to 5) can promote a full reaction between berberine and acylated natural polysaccharide during the encapsulation process, so that berberine can be more fully and stably bound to the molecular structure of acylated polysaccharide to form stable composite drug-loaded nanoparticles.

[0049] The mass m6 of the acylated natural polysaccharide can be 2, 3, 4 or 5.

[0050] In some alternative embodiments, the encapsulation process is carried out at a temperature of 30°C to 80°C for a duration of 30 min to 1440 min.

[0051] In these embodiments, encapsulation at temperatures of 30°C to 80°C and for times of 30 min to 1440 min allows berberine to bind more fully and stably to the molecular structure of the acylated polysaccharide, forming stable composite drug-loaded nanoparticles.

[0052] The encapsulation temperature can be 30℃, 35℃, 40℃, 45℃, 50℃, 55℃, 60℃, 65℃, 70℃, 75℃ or 80℃.

[0053] The encapsulation process can be performed for 30 min, 40 min, 50 min, 100 min, 150 min, 200 min, 250 min, 500 min, 600 min, 700 min, 800 min, 1000 min, 1200 min, 1400 min, or 1440 min.

[0054] In some alternative embodiments, the basic catalyst includes at least one selected from pyridine, 4-dimethylaminopyridine, triethylamine, N,N-diisopropylethylamine, and trimethylpyridine; and / or The types of natural polysaccharides include at least one of tobacco polysaccharide, wolfberry polysaccharide, astragalus polysaccharide, and pomegranate peel polysaccharide; and / or The organic solvents include at least one of ethanol, methanol, acetone and ethyl acetate.

[0055] In these embodiments, the use of an alkaline catalyst, including at least one selected from pyridine, 4-dimethylaminopyridine, triethylamine, N,N-diisopropylethylamine, and trimethylpyridine, allows for a sufficient reaction between the natural polysaccharide and propionic anhydride, introducing a propionyl hydrophobic group onto the molecular chain of the natural polysaccharide, thereby improving its hydrophobicity and meeting the carrier requirements of berberine. Furthermore, the use of at least one natural polysaccharide selected from tobacco polysaccharide, wolfberry polysaccharide, astragalus polysaccharide, and pomegranate peel polysaccharide can cover most types of natural polysaccharides, improving the versatility of the preparation method provided in this application. In addition, the use of an organic solvent, including at least one selected from ethanol, methanol, acetone, and ethyl acetate, can sufficiently disperse berberine and the acylated natural polysaccharide, allowing berberine to bind more fully and stably to the molecular structure of the acylated polysaccharide, forming stable composite drug-loaded nanoparticles.

[0056] Based on a general inventive concept, embodiments of this application provide a composite drug-loaded nanoparticle based on acylated polysaccharide and berberine, wherein the composite drug-loaded nanoparticle is prepared by the preparation method described above; the composite drug-loaded nanoparticle comprises a natural polysaccharide matrix and berberine loaded on the natural polysaccharide matrix.

[0057] The composite drug-loaded nanoparticles are prepared based on the above-described preparation method. The specific steps of the preparation method can be referred to the above embodiments. Since the composite drug-loaded nanoparticles adopt some or all of the technical solutions of the above embodiments, they have at least all the beneficial effects brought about by the technical solutions of the above embodiments, which will not be elaborated here.

[0058] Based on a general inventive concept, embodiments of this application provide an application of composite drug-loaded nanoparticles based on acylated polysaccharides and berberine, the application including using the composite drug-loaded nanoparticles for the treatment of wound infection, diarrhea, enteritis or mucosal ulceration.

[0059] This application is based on the above-mentioned composite drug-loaded nanoparticles. The specific composition of the composite drug-loaded nanoparticles can be referred to in the above embodiments. Since this application adopts some or all of the technical solutions of the above embodiments, it has at least all the beneficial effects brought about by the technical solutions of the above embodiments, which will not be elaborated here.

[0060] It should be noted that, specifically, this application refers to the use of composite drug-loaded nanoparticles as a standalone therapeutic agent for the treatment of wound infections, diarrhea, enteritis, or mucosal ulcers, or as a raw material for the preparation of therapeutic agents for wound infections, diarrhea, enteritis, or mucosal ulcers.

[0061] It should be noted that, due to the high encapsulation efficiency of this composite drug-loaded nanoparticle, berberine within the nanoparticles is not easily metabolized and degraded in vivo, significantly improving its bioavailability and enhancing its therapeutic effect. Furthermore, this composite drug-loaded nanoparticle can promote wound healing caused by Staphylococcus aureus infection and reduce the development of Staphylococcus aureus drug resistance, making it a potential alternative to antibiotics.

[0062] The present application is further illustrated below with reference to specific embodiments. Experimental methods in the following embodiments that do not specify specific conditions are generally determined according to national / industry standards; if there is no corresponding national / industry standard, they are performed according to general international standards, conventional conditions, or conditions recommended by the manufacturer.

[0063] Example 1 A method for preparing composite drug-loaded nanoparticles based on acylated polysaccharides and berberine, comprising: S1. First, the natural polysaccharide is dissolved using N,N-dimethylformamide (DMF). Then, the natural polysaccharide is acylated using an alkaline catalyst and acid anhydride to enhance its hydrophobicity. The mixture is then subjected to dialysis for 24 hours, concentration, and freeze-drying to obtain the acylated natural polysaccharide. The acylation modification method includes acetylation modification. The natural polysaccharide used is water-extracted tobacco polysaccharide. S2. Berberine and acylated natural polysaccharide are encapsulated in an organic solvent, and then the organic solvent is removed by rotary evaporation to obtain composite drug-loaded nanoparticles (BB@Ace-TP) based on acetylated polysaccharide and berberine.

[0064] When the acylation modification is acetylation modification, the acid anhydride is acetic anhydride, and the mass m1 of the natural polysaccharide, the mass m2 of the acetic anhydride, and the mass m3 of the basic catalyst satisfy the following: m1:m2:m3=1:2:2.

[0065] When the acylation modification method is acetylation modification, the acylation modification temperature is 120℃ and the acylation modification time is 20min.

[0066] The mass m5 of berberine and the mass m6 of acylated natural polysaccharide satisfy the ratio m5:m6 = 1:2.

[0067] The encapsulation temperature was 60℃ and the encapsulation time was 480 min.

[0068] The type of alkaline catalyst is pyridine; The organic solvent is anhydrous ethanol.

[0069] Example 2 Compared to Example 1, the differences in this example are as follows, while the rest are the same: Acylation modification methods include propionylation modification; the natural polysaccharide used is water-extracted tobacco polysaccharide. The natural polysaccharide can be dissolved in dimethyl sulfoxide (DMSO) before acylation modification.

[0070] When the acylation modification is propionylation, the acid anhydride is propionic anhydride, and the mass m1 of the natural polysaccharide, the mass m4 of the propionic anhydride, and the mass m3 of the alkaline catalyst satisfy the following: m1:m4:m3=1:1:(2 to 10).

[0071] The alkaline catalyst is 4-dimethylaminopyridine.

[0072] When the acylation modification method is propionylation modification, the acylation modification temperature is 60℃ and the acylation modification time is 1440min.

[0073] The mass m5 of berberine and the mass m6 of acylated natural polysaccharide satisfy the ratio m5:m6 = 1:4.

[0074] The encapsulation temperature was 60℃ and the encapsulation time was 720 min.

[0075] The organic solvent is anhydrous ethanol.

[0076] The resulting drug-loaded nanoparticles based on propionyl polysaccharide and berberine are denoted as BB@Prop-TP.

[0077] Example 3 Compared to Example 1, the differences in this example are as follows, while the rest are the same: Acylation modification methods include acetylation modification; the natural polysaccharide used is water-extracted wolfberry polysaccharide. Prior to acylation modification, the natural polysaccharide is dissolved using DMSO.

[0078] When the acylation modification is acetylation modification, the acid anhydride is acetic anhydride, and the mass m1 of the natural polysaccharide, the mass m2 of the acetic anhydride and the mass m3 of the basic catalyst satisfy the following: m1:m2:m3=1:5:10.

[0079] The alkaline catalyst is triethylamine; When the acylation modification method is acetylation modification, the acylation modification temperature is 80℃ and the acylation modification time is 480min.

[0080] The mass m5 of berberine and the mass m6 of acylated natural polysaccharide satisfy the ratio m5:m6 = 1:5.

[0081] The encapsulation temperature was 40℃ and the encapsulation time was 1440 min.

[0082] The organic solvent is methanol.

[0083] The resulting composite drug-loaded nanoparticles based on acetylated polysaccharides and berberine are denoted as BB@Ace-LBP.

[0084] Example 4 Compared to Example 1, the differences in this example are as follows, while the rest are the same: Acylation modification methods include propionylation modification; the natural polysaccharide used is water-extracted Lycium barbarum polysaccharide. Prior to acylation modification, the natural polysaccharide is dissolved in DMF.

[0085] When the acylation modification method is propionylation modification, the acid anhydride is propionic anhydride, and the mass m1 of the natural polysaccharide, the mass m4 of the propionic anhydride and the mass m3 of the alkaline catalyst satisfy the following: m1:m4:m3=1:5:10.

[0086] The alkaline catalyst is N,N-diisopropylethylamine; When the acylation modification method is propionylation modification, the acylation modification temperature is 80℃ and the acylation modification time is 240min.

[0087] The mass m5 of berberine and the mass m6 of acylated natural polysaccharide satisfy the ratio m5:m6 = 1:4.

[0088] The encapsulation temperature was 70℃ and the encapsulation time was 360 min.

[0089] The organic solvent is acetone.

[0090] The resulting composite drug-loaded nanoparticles based on acetylated polysaccharides and berberine are denoted as BB@Prop-LBP.

[0091] Example 5 Compared to Example 1, the differences in this example are as follows, while the rest are the same: Acylation modification methods include acetylation modification; the natural polysaccharide used is water-extracted Astragalus polysaccharide. Prior to acylation modification, the natural polysaccharide is dissolved in acetone.

[0092] When the acylation modification is acetylation modification, the acid anhydride is acetic anhydride, and the mass m1 of the natural polysaccharide, the mass m2 of the acetic anhydride, and the mass m3 of the alkaline catalyst satisfy the following: m1:m2:m3=1:3:7.

[0093] The type of alkaline catalyst is pyridine; When the acylation modification method is acetylation modification, the acylation modification temperature is 70℃ and the acylation modification time is 720min.

[0094] The mass m5 of berberine and the mass m6 of acylated natural polysaccharide satisfy the ratio m5:m6 = 1:3.

[0095] The encapsulation temperature was 80℃ and the encapsulation time was 240 min.

[0096] The organic solvent is anhydrous ethanol.

[0097] The resulting composite drug-loaded nanoparticles based on acetylated polysaccharides and berberine are denoted as BB@Ace-AP.

[0098] Example 6 Compared to Example 1, the differences in this example are as follows, while the rest are the same: Acylation modification methods include propionylation modification; the natural polysaccharide used is water-extracted pomegranate peel polysaccharide. Prior to acylation modification, the natural polysaccharide is dissolved in DMF.

[0099] When the acylation modification method is propionylation modification, the acid anhydride is propionic anhydride, and the mass m1 of the natural polysaccharide, the mass m4 of the propionic anhydride and the mass m3 of the alkaline catalyst satisfy the following: m1:m4:m3=1:3:6.

[0100] The alkaline catalyst is trimethylpyridine.

[0101] When the acylation modification method is propionylation modification, the acylation modification temperature is 65℃ and the acylation modification time is 900min.

[0102] The mass m5 of berberine and the mass m6 of acylated natural polysaccharide satisfy the following ratio: m5:m6 = 1:2.5.

[0103] The encapsulation temperature was 70℃ and the encapsulation time was 240 min.

[0104] The organic solvent is ethanol.

[0105] The resulting drug-loaded nanoparticles based on propionyl polysaccharide and berberine are denoted as BB@Prop--PPP.

[0106] Relevant experimental and effect data: 1. Encapsulation efficiency determination: The composite drug-loaded nanoparticles obtained in each embodiment were collected separately, and a certain mass concentration of the composite drug-loaded nanoparticles was weighed and placed in a 50 kDa ultrafiltration tube, then centrifuged at 3500 r / min for 15 min. The solution outside the ultrafiltration tube was then collected, and the mass content of free berberine in the composite drug-loaded nanoparticles was determined by high-performance liquid chromatography (HPLC). The encapsulation efficiency of the composite drug-loaded nanoparticles was calculated by comparing the peak area of ​​the control sample with the dilution factor. In the HPLC method, a C18 column was used as the stationary phase, and the phosphate buffer solution consisted of a 1:1 volume ratio of 0.05 mol / L potassium dihydrogen phosphate solution and a 0.05 mol / L mass ratio of 0.2% triethylamine solution. The pH of the phosphate buffer was adjusted to 3.0 using phosphoric acid. The mobile phase was then used with a 65:35 volume ratio of phosphate buffer to acetonitrile, and detection was performed at a wavelength of 263 nm. The temperature of the C18 column was 30 °C. The encapsulation efficiency of the composite drug-loaded nanoparticles in each embodiment was measured and is shown in Table 1.

[0107] Table 1 Encapsulation efficiency of composite drug-loaded nanoparticles in various embodiments

[0108] As shown in Table 1, the present application provides a method for preparing composite drug-loaded nanoparticles based on acylated polysaccharides and berberine. This method modifies the hydrophobicity of natural polysaccharides by acylation, thereby significantly improving the encapsulation efficiency of berberine, resulting in an encapsulation efficiency of berberine in the composite drug-loaded nanoparticles of up to 65% or more.

[0109] 2. Physiological activity assay: (1) In vitro antibacterial activity: Use an inoculation loop to pick up a portion of the bacterial strain (e.g., Staphylococcus aureus, methicillin-resistant Staphylococcus aureus) preserved on the slant of a test tube, and inoculate the selected strain into 100 mL of liquid LB medium. Place the tube in a constant temperature shaker and incubate at 120 rpm for 24 h to activate the bacterial strain.

[0110] Experimental groups 1, 2, and 3 were prepared using 50% LB medium at different concentrations. Experimental group 1 consisted of berberine alone (BB), experimental group 2 was a physical mixture of berberine and acetylated tobacco polysaccharide, and experimental group 3 was a composite drug-loaded nanoparticle solution (BB@Ace-TP) based on Example 1. 100 μL of each concentration of the experimental group solution was taken and mixed with 100 μL of bacterial culture at a density of 102. 6 A bacterial suspension of CFU / mL was mixed to obtain a mixed test solution. The initial absorbance of these mixed test solutions was then measured at 600 nm using a microplate reader. These mixed test solutions were incubated at 37°C for 24 h, and the endpoint absorbance was measured again. The initial and endpoint absorbance values ​​can be used to assess bacterial growth in different materials. A mixture of 100 μL of the experimental group solution and 100 μL of 50% LB medium was used as the sample background, and a mixture of 100 μL of 50% LB medium and 100 μL of bacterial suspension was used as the blank control group. The initial and endpoint absorbance values ​​were measured using the same procedure, and the inhibition rate was calculated using the following formula.

[0111] , In the formula, A2 is the endpoint absorbance value of the experimental group, A1 is the initial absorbance value of the experimental group, and A... 02 A represents the endpoint absorbance value of the blank control group. 01 The initial absorbance value of the blank control group; A 2S A represents the endpoint absorbance value of the sample background. 1S The initial absorbance value of the sample background.

[0112] The results showed that the antibacterial activity of different concentrations of the experimental group samples against *S. aureus* was as follows: Figure 3 As shown, the overall antibacterial rate of the drugs in the experimental groups against *S. aureus* increased with increasing concentration, exhibiting a concentration-dependent effect. The order of antibacterial rate from highest to lowest was BB@Ace-TP > Mixture > BB (p<0.05). BB@Ace-TP showed the strongest antibacterial activity, achieving an antibacterial rate of 95.22% ± 1.45% at a concentration of 0.25 mg / mL, while free BB only achieved 69.98% ± 2.77%.

[0113] Similar to the results against S. aureus, the inhibition rates of different concentrations of the experimental groups against MRSA were as follows: Figure 4As shown, the order of antibacterial activity from highest to lowest is BB@Ace-TP > Mixture > BB (p<0.05). The antibacterial activity of BB@Ace-TP provided in this application embodiment is the highest, followed by Mixture. This indicates the synergistic antibacterial efficacy of BB and Ace-TP in the composite drug-loaded nanoparticles provided in this application embodiment. After the composite drug-loaded nanoparticles are prepared, the binding between BB and acylated natural polysaccharides is tighter, enhancing the protective effect of acylated natural polysaccharides on BB, thereby reducing BB degradation. This is an important reason for the improved overall antibacterial activity of the composite drug-loaded nanoparticles. Meanwhile, the experiment revealed that MRSA has low sensitivity to BB. To achieve the same antibacterial rate as *S. aureus*, the concentration of MRSA used in the experimental group samples needs to be 2 to 4 times that used for *S. aureus*.

[0114] (2) Antibacterial effect: To investigate the effects of berberine alone and the antibacterial effects of berberine combined with acetylated polysaccharides in different forms on Staphylococcus aureus, this application used the IC50 calculated from the berberine content in the sample as the evaluation index. The antibacterial activity of each system was determined through antibacterial experiments, and the results are shown in Table 2.

[0115] Table 2. Antibacterial activity data of Staphylococcus aureus as determined by antibacterial test.

[0116] As shown in Table 2, when berberine acts alone, the IC50 value of berberine against Staphylococcus aureus is 0.098 mg / mL, indicating that berberine can exhibit certain antibacterial activity. When berberine and acetylated tobacco polysaccharide are reacted in the form of a physical mixture, the IC50 value of the physical mixture against Staphylococcus aureus decreases to 0.076 mg / mL, which is 22.4% higher than the antibacterial activity of berberine acting alone. This indicates that the addition of acetylated natural polysaccharide can initially enhance the antibacterial ability of berberine. When berberine and acetylated tobacco polysaccharide form the composite drug-loaded nanoparticle solution shown in Example 1, the IC50 value of the composite drug-loaded nanoparticle solution against Staphylococcus aureus further decreases to 0.064 mg / mL, which is 34.7% higher than the antibacterial activity of berberine acting alone, and is significantly higher than the case of berberine and acetylated tobacco polysaccharide acting in the form of a physical mixture.

[0117] Based on the experiment on the antibacterial effect of Staphylococcus aureus, this study investigated the antibacterial effects of berberine alone and in combination with acetylated polysaccharides in different forms against methicillin-resistant Staphylococcus aureus (MRSA). The results are shown in Table 3.

[0118] Table 3. Antibacterial activity data determined by MRSA antibacterial test.

[0119] Table 3 shows that, when berberine acts alone, the IC50 of berberine on MRSA is... 50 The value was 0.538 mg / mL; when berberine reacted with acetylated tobacco polysaccharide in the form of a physical mixture, the IC50 of the physical mixture on MRSA was [value missing]. 50 The value decreased to 0.382 mg / mL, representing a 29.0% increase in antibacterial activity compared to berberine alone; and when berberine reacted with acetylated tobacco polysaccharide to form the composite drug-loaded nanoparticle solution shown in Example 1, the IC50 value of the composite drug-loaded nanoparticle solution against MRSA was... 50 The value was further reduced to 0.237 mg / mL, which increased the antibacterial activity by 56.0% compared to berberine alone, and was significantly higher than that of berberine and acetylated tobacco polysaccharide in the form of a physical mixture.

[0120] As shown in Tables 2 and 3, berberine and acetylated tobacco polysaccharide, in the form of a physical mixture, exhibited good synergistic antibacterial efficacy against Staphylococcus aureus or MRSA. After preparing the composite drug-loaded nanoparticles according to Example 1, the binding between berberine and acetylated natural polysaccharide was more compact, and the protective effect of acetylated natural polysaccharide on berberine was enhanced, reducing the degradation of berberine. This is an important reason for improving the overall antibacterial activity of the composite drug-loaded nanoparticles.

[0121] (3) In vivo pharmacokinetics: Berberine, a physical mixture of berberine and acetylated tobacco polysaccharide, and the BB@Ace-TP solution from Example 1 were administered to rats via oral gavage at a dose of 50 mg / kg (calculated based on the amount of berberine in each group). Rats were kept NPO (no food or water) during the administration period. At different time points after administration, 0.1 mL of blood was collected from the tail vein of each rat and placed in an EDTA-Na tube. 40 μL of plasma was collected by centrifugation, and 30 μL of 19.11 units / mL sulfatase solution was added. The mixture was thoroughly mixed and incubated at 37°C with shaking for 1 h. After incubation, the mixture was cooled to room temperature, and 200 μL of acetonitrile was added to precipitate the protein. Then, 15 μL of hydrocortisone (internal standard) was added, and the mixture was thoroughly mixed. The supernatant was centrifuged, and the supernatant was evaporated to dryness at 40°C. The supernatant was redispersed in the mobile phase, centrifuged again, and injected into an HPLC system to analyze the berberine content in the plasma. In vivo pharmacokinetic parameters were calculated using DAS 2.0 software. The result is Figure 4 As shown, rats treated with berberine, the physical mixture, and BB@Ace-TP solution all reached peak plasma concentrations within 15 minutes after administration. The peak plasma concentrations of the physical mixture and BB@Ace-TP solution were 1.15 times and 6.96 times higher than those of the berberine group, respectively. The area under the curve (AUC) for the physical mixture and BB@Ace-TP solution was also significantly higher.0→4h The concentrations were 1.07 times and 1.69 times higher than those of the berberine group, respectively. Overall, the absorption process of berberine and the physical mixture in vivo was similar, both exhibiting low blood concentrations and poor oral bioavailability. However, the BB@Ace-TP solution in Example 1 was rapidly absorbed in vivo, resulting in a high peak blood concentration. This indicates that the composite drug-loaded nanoparticles in Example 1 significantly improved the oral bioavailability of berberine.

[0122] (4) Pharmacodynamics of in vivo wound healing: 1) Wound healing status: Mice had some back hair removed, and after disinfection with 75% alcohol, skin and subcutaneous mucosa were excised from the sides of the spine of each mouse to create a circular wound approximately 1 cm in diameter, ensuring complete exposure without any mucosal residue. After bleeding from the circular wound stopped, 50 μL of Staphylococcus aureus suspension was applied to the wound to establish a circular wound infection model. The next day, medication was administered. The experimental group received 50 μL of the drug applied to the wound twice daily, morning and evening, and was also given oral gavage. The positive control group (PC) received oral amoxicillin and topical levofloxacin, while the model group received oral and topical saline. The healing of the circular wounds in the mice was observed, and the results are as follows: Figure 5 As shown.

[0123] Depend on Figure 5 The healing rate of circular wounds was observed to be: positive control > BB@Ace-TP > physical mixture > berberine > acetylated tobacco polysaccharide. Among the groups, the BB@Ace-TP group exhibited the fastest wound healing rate, with its healing rate only significantly lower than the positive control group on day 3; the healing rates on other days were not significantly different from the positive control group. The physical mixture showed a slightly higher healing rate than berberine, but significantly lower than BB@Ace-TP. This indicates that, compared to berberine alone or material mixtures, preparing acetylated tobacco polysaccharide and berberine as composite drug-loaded nanoparticles can synergistically enhance the wound healing rate.

[0124] 2) Wound staining condition: Frozen sections of the circular wound tissue were fixed in 4% paraformaldehyde for 10 min, then thoroughly rinsed with distilled water. The sections were then stained with Weigert iron hematoxylin solution for 10 min, bluing with tap water for 5 min, followed by differentiation with hydrochloric acid-alcohol solution for 1 min, and finally rinsed with distilled water to terminate differentiation. The sections were then stained with Ponceau S-acid fuchsin solution for 8 min, rinsed with 2% glacial acetic acid solution for 1 min, differentiated with 1% phosphomolybdic acid solution for 3 min, directly transferred to aniline blue solution for 4 min, and rinsed with 0.2% glacial acetic acid solution for 1 min. Finally, the sections were rapidly dehydrated with graded ethanol, cleared with xylene for 1 min, mounted with neutral resin, and observed under an optical microscope. The Masson trichrome staining results of the circular wound are shown below. Figure 6 As shown.

[0125] Depend on Figure 6 It is known that in circular wounds, muscle fibers and the stratum corneum will be stained red, while cell nuclei will be stained black, and collagen fibers will be stained blue. The maturity of collagen fibers can be represented by the depth of blue staining. Based on the area of ​​the blue-stained region and the arrangement of collagen fibers, the BB@Ace-TP experimental group showed the most significant amount of collagen fiber deposition and the most orderly tissue remodeling effect, indicating that the BB@Ace-TP experimental group can simultaneously promote large-scale early collagen synthesis and later maturation and arrangement, making it the group with the best healing effect. The positive control group showed better collagen deposition and tissue remodeling effects in the later stages, but insufficient collagen density on day 5, indicating that the positive control group had limited ability to remodel deep skin tissue in the early stages of treatment. The experimental group using acetylated tobacco polysaccharide alone showed weaker healing effects on day 5, but significant improvement in collagen deposition and structure on day 10, demonstrating that acetylated tobacco polysaccharide performs better in later collagen fiber deposition and tissue remodeling, thus having a more significant effect in the later stages of wound healing. The experimental group using berberine alone showed some collagen deposition on day 5, but no significant progress on day 10. Compared with acetylated tobacco polysaccharide, berberine exhibited stronger antibacterial and antioxidant activity, which is particularly important in the early stages of wound healing. This also indicates that berberine is more effective in early hemostasis and antibacterial action, with limited impact on later collagen maturation. The model group showed a lack of collagen deposition and failure of tissue remodeling throughout the entire process, validating the effectiveness of the experimental system.

[0126] Therefore, berberine is slightly more effective than acetylated tobacco polysaccharide in the early stages of wound healing, but acetylated tobacco polysaccharide ultimately achieves more complete tissue reconstruction. The physical mixture group showed better results than the groups using acetylated tobacco polysaccharide or berberine alone, but the healing effect was not as good as the BB@Ace-TP group. This indicates that the synergistic effect of the "simple mixture" of berberine and acetylated tobacco polysaccharide is weaker than the precise delivery of the composite drug-loaded nanoparticles in Example 1. Furthermore, the composite drug-loaded nanoparticles in Example 1 combine the complementary effects of berberine and acetylated tobacco polysaccharide, and compared to the groups using either substance alone, the composite drug-loaded nanoparticles BB@Ace-TP showed the most significant healing effect on circular wounds. Although the healing rate of circular wounds in the positive control group was similar to that of the composite drug-loaded nanoparticles BB@Ace-TP, the collagen fiber density in the healed wounds treated in the positive control group was lower, indicating that the positive control group had limited remodeling effect on deep skin tissues.

[0127] 3) Changes in biological factors during wound healing: Vascular endothelial growth factor (VEGF) is one of the most critical pro-angiogenic factors in the body. The key role of VEGF is to stimulate the formation of new blood vessels and increase vascular permeability, drive the formation of dense new capillary networks in granulation tissue, ensure that the wound bed receives sufficient oxygen and nutrients, and induce angiogenesis in the hair follicle microenvironment, which is essential for wound healing.

[0128] The general procedure for determining the fluorescence intensity of wounds during the healing process is as follows: Frozen sections from different sample groups were taken, equilibrated at room temperature, and washed with PBS; 5% BSA was added, and the sections were blocked at room temperature for 30 min, then the blocking solution was discarded; the sections were incubated overnight with primary antibody (1:150 dilution) at 4°C, and washed three times with 1×PBS after incubation; the sections were then incubated with secondary fluorescent antibody (1:100 dilution) at room temperature in the dark for 60 min, and washed three times with 1×PBS; the sections were incubated with DAPI at room temperature for 5 min, washed three times with 1×PBS, and an anti-quenching agent was added. The sections were then mounted and observed under a microscope. The fluorescence intensity was analyzed using ImageJ software, and the results are as follows: Figure 7 As shown.

[0129] Depend on Figure 7It can be seen that the wound fluorescence intensity varied in different drug administration groups. In the quantitative fluorescence chromatogram on the fifth day of wound healing, the order of wound fluorescence intensity was: BB@Ace-TP > positive control group > berberine > physical mixture > acetylated tobacco polysaccharide > model group, which is consistent with the wound healing rate results on the third day. Compared with the wound fluorescence intensity on the fifth day, the wound fluorescence intensity of each drug administration group increased on the tenth day, and the order of wound fluorescence intensity was BB@Ace-TP ≈ positive control group > berberine > physical mixture > acetylated tobacco polysaccharide > model group, corresponding to the final wound healing rate results.

[0130] Therefore, based on the above experimental results, it can be seen that the different VEGF expression in each drug administration group is closely related to the wound healing rate, and BB@Ace-TP can efficiently regulate VEGF expression in wound tissue and promote wound healing.

[0131] In summary, this application provides a method for preparing composite drug-loaded nanoparticles based on acylated polysaccharides and berberine. This method modifies the hydrophobicity of natural polysaccharides through acylation, and then uses the acylated natural polysaccharides as a carrier and berberine as a model drug to prepare composite drug-loaded nanoparticles, achieving a significant improvement in the encapsulation efficiency of berberine. The prepared composite drug-loaded nanoparticles have the following advantages: (1) High encapsulation efficiency, increasing the stability of berberine: The preparation method provided in this application provides acylation modification of natural polysaccharides, which improves the lipophilicity and hydrophobicity of natural polysaccharides. The lipophilic and hydrophobic acylated natural polysaccharides can significantly improve the encapsulation efficiency of berberine. This means that more berberine is encapsulated in the acylated natural polysaccharide carrier. The acylated natural polysaccharide carrier can provide better protection for berberine and reduce the impact of external factors (such as oxidation and hydrolysis) on berberine during storage and transportation, thereby improving the stability of berberine. In addition, the composite drug-loaded nanoparticles can reduce the risk of degradation in vivo and overcome the key problem of low bioavailability of berberine due to rapid degradation in vivo.

[0132] (2) Prolonging the duration of drug action and improving oral bioavailability and efficacy: High encapsulation rate of composite drug-loaded nanoparticles can reduce the degradation of berberine in the body, prolong the duration of action, thereby improving oral bioavailability and efficacy.

[0133] (3) Reduce drug dosage: Due to the high bioavailability of oral drugs, the dosage can be reduced to achieve the same therapeutic effect. This helps to reduce the toxic side effects of drugs, alleviate the physical burden on patients, and also reduce the cost of drug treatment.

[0134] (4) Synergistic effect: The composite drug-loaded nanoparticles based on acylated natural polysaccharides and berberine can also play a synergistic pharmacodynamic role in antibacterial, anti-inflammatory and wound healing promotion, providing a new direction for solving the current problem of antibiotic resistance in wound treatment, effectively reducing bacterial resistance, and providing a new strategy for clinical wound healing treatment.

[0135] The above description is merely a specific embodiment of this application, enabling those skilled in the art to understand or implement this application. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of this application. Therefore, this application is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features claimed in this application.

Claims

1. A method for preparing composite drug-loaded nanoparticles based on acylated polysaccharides and berberine, characterized in that, The preparation method includes: Acylation modification of natural polysaccharides using an alkaline catalyst and acid anhydride is used to enhance the hydrophobicity of the natural polysaccharides, resulting in acylated natural polysaccharides; wherein the acylation modification includes acetylation modification or propionylation modification. Berberine and the acylated natural polysaccharide were encapsulated in an organic solvent to obtain composite drug-loaded nanoparticles based on acylated polysaccharide and berberine.

2. The preparation method according to claim 1, characterized in that, When the acylation modification is acetylation modification, the acid anhydride is acetic anhydride, and the mass m1 of the natural polysaccharide, the mass m2 of the acetic anhydride and the mass m3 of the alkaline catalyst satisfy: m1:m2:m3=1:(2 to 5):(2 to 10).

3. The preparation method according to claim 2, characterized in that, When the acylation modification is acetylation modification, the acylation modification temperature is 50°C to 120°C, and the acylation modification time is 20 min to 1440 min.

4. The preparation method according to claim 1, characterized in that, When the acylation modification is propionylation modification, the acid anhydride is propionic anhydride, and the mass m1 of the natural polysaccharide, the mass m4 of the propionic anhydride and the mass m3 of the alkaline catalyst satisfy: m1:m4:m3=1:(1 to 5):(2 to 10).

5. The preparation method according to claim 4, characterized in that, When the acylation modification is propionylation modification, the acylation modification temperature is 60°C to 120°C, and the acylation modification time is 30 min to 1440 min.

6. The preparation method according to claim 1, characterized in that, The mass m5 of berberine and the mass m6 of acylated natural polysaccharide satisfy the following condition: m5:m6 = 1:(2 to 5).

7. The preparation method according to claim 1, characterized in that, The encapsulation process is carried out at a temperature of 30°C to 80°C for a duration of 30 min to 1440 min.

8. The preparation method according to claim 1, characterized in that, The alkaline catalyst includes at least one of pyridine, 4-dimethylaminopyridine, triethylamine, N,N-diisopropylethylamine, and trimethylpyridine; and / or The types of natural polysaccharides include at least one of tobacco polysaccharide, wolfberry polysaccharide, astragalus polysaccharide, and pomegranate peel polysaccharide; and / or The organic solvent includes at least one of ethanol, methanol, acetone and ethyl acetate.

9. A composite drug-loaded nanoparticle based on acylated polysaccharide and berberine, characterized in that, The composite drug-loaded nanoparticles are prepared by the preparation method according to any one of claims 1 to 8; the composite drug-loaded nanoparticles include a natural polysaccharide matrix and berberine loaded on the natural polysaccharide matrix.

10. An application of composite drug-loaded nanoparticles based on acylated polysaccharides and berberine, characterized in that, The applications include using the composite drug-loaded nanoparticles of claim 9 for the treatment of wound infections, diarrhea, enteritis, or mucosal ulcers.

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