Transdermal liraglutide nanoparticle preparation as well as preparation method and application thereof

Abstract

The invention discloses transdermal liraglutide nanoparticles as well as a preparation method and application thereof, the liraglutide nanoparticles are of a core-shell structure, the core comprises a complex of liraglutide and a phenolic hydroxyl polyphenol compound, and the core-shell is quaternized chitosan. A plurality of phenolic hydroxyl groups of the phenolic hydroxyl polyphenol compound can serve as hydrogen bond acting sites to be combined with liraglutide to form the nano-particles, the technical blank of liraglutide nano-particles is filled, and the preparation method is also suitable for preparing other polypeptide molecular nano-particles. And then the nanoparticles are combined with electropositive quaternized chitosan (HTCC) to form the liraglutide nanoparticles with the core-shell structure. The HTCC promotes the medicine to permeate into the skin by changing the secondary structure of keratin, high-concentration positive charges carried on the surface of the HTCC can form strong electrostatic adsorption with a negative-charge phospholipid bimolecular layer of a skin cuticle, and compared with chitosan, the HTCC has higher skin permeation capacity and is beneficial to being applied to a transdermal drug delivery preparation.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedical materials, and particularly relates to a transdermal liraglutide nanoparticle preparation, a preparation method thereof, and an application thereof. Background Art

[0002] Obesity is a global health problem. A series of complications brought about by obesity, such as diabetes, "three highs" and other diseases, have seriously affected the normal life of people and their families. As an adjuvant drug, weight loss drugs play a crucial role in the treatment of obesity. Lira (Lira) is a drug that can be used to treat obesity in China. It is a glucagon-like peptide-1 (GLP-1) receptor agonist and belongs to a long-acting human GLP-1 analogue, which is used for the treatment of type 2 diabetes and obesity. Lira regulates glucose metabolism, increases insulin secretion, reduces blood sugar levels, and can also reduce body weight and improve cardiovascular function by mimicking the action of natural GLP-1. Its current administration method is injection, which causes Lira to be easily degraded by dipeptidyl peptidase 4 in the body, and its plasma half-life is less than 2 minutes. It must be continuously infused intravenously to produce therapeutic effects, which greatly increases the pain of patients during administration.

[0003] In order to achieve better administration effects, a new administration method, "transdermal administration", has been developed. Compared with injection administration, it has advantages such as avoiding the first-pass effect, maintaining a stable blood drug concentration, improving patient compliance, and being convenient for self-administration, and is more adaptable to the needs of the weight loss drug market to treat obese patients. Although transdermal administration is a non-invasive administration method with high patient compliance, due to the barrier effect of the skin stratum corneum, especially for large molecular weight proteins and peptide drugs, it is difficult to effectively penetrate into the skin, which limits the application of such drugs. Therefore, various transdermal administration technologies have been developed in recent years, but several current mainstream transdermal administration technologies have their own insurmountable disadvantages: The first is chemical enhancers: Although chemical enhancers can effectively promote drug absorption, they may cause irritation or allergic reactions to the skin. The second is microneedles: The production and operation costs are relatively high and difficult to promote. And due to the size limitation of microneedles, the amount of drug that each microneedle can carry is limited, which may be insufficient for the administration effects of high-dose or macromolecular drugs, and frequent repeated administrations are required to maintain the drug effect. The third is electroporation: Electroporation can increase skin permeability, but high voltage may cause discomfort such as skin stinging and burning. The fourth is iontophoresis: Iontophoresis is suitable for drugs with positive or negative charges, but has poor effects on uncharged molecules and may cause slight skin discomfort or even damage. Although the above transdermal administration technologies have a certain effect on enhancing penetration, there are still problems such as insufficient drug penetration depth, low efficiency, and possible skin irritation. Therefore, it is necessary to develop a new preparation of Lira that can be transdermally administered. Summary of the Invention

[0004] To overcome the problems existing in the above-mentioned prior art, one of the objectives of the present invention is to provide a transdermal liraglutide nanoparticle preparation. Another objective of the present invention is to provide a polypeptide drug molecule. A third objective of the present invention is to provide a preparation method of the above-mentioned polypeptide drug molecule. A fourth objective of the present invention is to provide the application of the above-mentioned polypeptide drug molecule. To achieve the above objectives, the technical solutions adopted by the present invention are as follows:

[0005] In the first aspect of the present invention, a transdermal liraglutide nanoparticle is provided. The liraglutide nanoparticle has a core-shell structure, wherein the core includes a complex of liraglutide and a phenolic hydroxyl polyphenol compound, and the shell is quaternized chitosan.

[0006] In the second aspect of the present invention, a polypeptide drug molecule nanoparticle is provided. The polypeptide drug molecule nanoparticle has a core-shell structure, wherein the core includes a complex of a polypeptide drug molecule and a phenolic hydroxyl polyphenol compound, and the shell is quaternized chitosan; the polypeptide drug molecule nanoparticle includes liraglutide or other polypeptide drug molecules.

[0007] Preferably, the particle size of the polypeptide drug molecule nanoparticle is 10 - 50 nm.

[0008] Preferably, the complex of the polypeptide drug molecule and the phenolic hydroxyl polyphenol compound is a spherical-like nanoparticle with a particle size of 5 - 25 nm.

[0009] Preferably, the phenolic hydroxyl polyphenol compound is selected from at least one of tannic acid, gallic acid, and ellagic acid.

[0010] Preferably, the quaternized chitosan is 2-hydroxypropyltrimethylammonium chloride chitosan (HTCC).

[0011] In the third aspect of the present invention, a preparation method of the polypeptide drug molecule nanoparticle described in the second aspect is provided, including the following steps:

[0012] S1. Mix a polypeptide drug molecule solution, a phenolic hydroxyl polyphenol compound solution, and an aluminum ion solution by using the rapid nano-composite method to obtain a solution containing a complex of the polypeptide drug molecule and the phenolic hydroxyl polyphenol compound;

[0013] S2. Mix the solution containing the complex of the polypeptide drug molecule and the phenolic hydroxyl polyphenol compound obtained above with a quaternized chitosan solution by using the rapid nano-composite method to obtain the polypeptide drug molecule nanoparticle.

[0014] The rapid nano-hybrid technology adopted in the present invention, also known as flash nano-compounding (FNC), is different from the usual thermodynamically controlled methods. It is a method for preparing nanoparticles through kinetically controlled enhanced self-assembly. By utilizing the dynamic mixing generated by turbulence, the formation of nanoparticles can be completed within a time scale of milliseconds, and it is commonly used for the delivery of hydrophobic drugs. FNC is a dynamically controlled mixing process evolved from the rapid nanoprecipitation method (FNP). Its preparation system adopts technologies such as multi-channel drainage, porous shunting, and vortex confluence. Under specific hydrodynamic conditions, the active substance and the carrier material will undergo continuous instantaneous assembly to form drug-loaded nanoparticles.

[0015] Preferably, the solvent of the polypeptide drug molecule solution is water, and the concentration is 0.1 - 1 mg / mL.

[0016] More preferably, the concentration of the polypeptide drug molecule solution is 0.2 - 0.5 mg / mL. Even more preferably, it is 0.25 - 0.35 mg / mL.

[0017] Preferably, the solvent of the phenolic hydroxyl polyphenol compound solution is an zwitterionic buffer, and the concentration is 0.1 - 1 mg / mL.

[0018] More preferably, the zwitterionic buffer is HEPES buffer with a pH of 4 - 6.

[0019] More preferably, the concentration of the phenolic hydroxyl polyphenol compound solution is 0.15 - 0.6 mg / mL. Even more preferably, it is 0.25 - 0.35 mg / mL.

[0020] Preferably, the solvent of the aluminum ion solution is water, and the concentration is 0.01 - 0.2 mg / mL.

[0021] More preferably, the concentration of the aluminum ion solution is 0.01 - 0.1 mg / mL.

[0022] Preferably, the volume ratio of the polypeptide drug molecule solution, the phenolic hydroxyl polyphenol compound solution, and the aluminum ion solution is 1:(1 - 3):(1 - 3).

[0023] Preferably, the solvent of the quaternized chitosan solution is water, and the concentration is 0.1 - 1 mg / mL.

[0024] More preferably, the concentration of the quaternized chitosan solution is 0.25 - 0.35 mg / mL.

[0025] Preferably, the volume ratio of the solution containing the complex of the polypeptide drug molecule and the phenolic hydroxyl polyphenol compound to the quaternized chitosan solution is 1:(0.5 - 1.5).

[0026] Preferably, the rapid nano-composite method is carried out in a multi-channel vortex mixer.

[0027] More preferably, the polypeptide drug molecule solution, the phenolic hydroxyl polyphenol compound solution, and the aluminum ion solution respectively reach the vortex mixing area through different channels in a multi-channel vortex mixer for mixing, wherein the flow velocity ratio of the polypeptide drug molecule solution, the phenolic hydroxyl polyphenol compound solution, and the aluminum ion solution in the channel is 1:(1 - 3):(1 - 3).

[0028] Further preferably, the flow velocity of the polypeptide drug molecule solution in the channel is 0.1 - 0.4 mL / s. Even more preferably, it is 0.25 - 0.35 mL / s.

[0029] More preferably, the solution containing the complex of the polypeptide drug molecule and the phenolic hydroxyl polyphenol compound and the quaternized chitosan solution respectively reach the vortex mixing area through different channels in a multi-channel vortex mixer for mixing, wherein the flow velocity ratio of the solution containing the complex of the polypeptide drug molecule and the phenolic hydroxyl polyphenol compound and the quaternized chitosan solution in the channel is 1:(0.5 - 1.5).

[0030] Further preferably, the flow velocity of the quaternized chitosan solution in the channel is 0.2 - 0.4 mL / s.

[0031] The fourth aspect of the present invention provides the use of the polypeptide drug molecule nanoparticles described in the second aspect in the preparation of a drug, and the dosage form of the drug includes a skin-administered preparation.

[0032] Preferably, the drug is a weight loss drug.

[0033] Preferably, the skin-administered preparation is selected from one of external solution, lotion, liniment, ointment, plaster, paste, and patch.

[0034] The beneficial effects of the present invention are:

[0035] (1) The present invention provides a liraglutide nanoparticle with a core-shell structure and a polypeptide drug molecule nanoparticle. Multiple phenolic hydroxyl groups of the phenolic hydroxyl polyphenol compound can serve as hydrogen bond action sites to bind with the polypeptide drug molecule to form nanoparticles, and then bind with positively charged quaternized chitosan (HTCC) to form a core-shell structure nanoparticle. HTCC promotes drug penetration through the skin by changing the secondary structure of keratin, and the high concentration of positive charges on its surface can form strong electrostatic adsorption with the negatively charged phospholipid bilayer of the skin stratum corneum, and has stronger skin penetration ability than chitosan, which is helpful for application in transdermal drug delivery preparations.

[0036] (2) The present invention provides a method for preparing the above-mentioned polypeptide drug molecular nanoparticles. In the preparation method, trivalent aluminum ions (Al3+) are introduced as cross-linking agents and stabilizers, and the rapid nanocomposite (FNC) technology is combined to achieve the efficient complexation of polypeptide drug molecules and polyphenolic compounds. Compared with the method that only relies on hydrogen bond complexation, Al3+ significantly improves the stability and particle size uniformity of the particles through ionic bond action, while reducing unnecessary aggregation in the system. The average particle size of the prepared polypeptide drug molecular nanoparticles is less than 25 nm, which is particularly suitable for the high particle size requirements in transdermal delivery systems. Description of the Drawings

[0037] Figure 1 Schematic diagram of the synthesis of liraglutide nanoparticles; where a is the amino acid sequence of Lira, and the structural formulas of TA and HTCC; b is the schematic diagram of the rapid nanocomposite technology process;

[0038] Figure 2 Effect of TA concentration on the particle size, polydispersity index (PDI) and zeta potential of Lira NP1;

[0039] Figure 3 Effect of channel flow rate on the particle size and PDI of Lira NP1;

[0040] Figure 4 Particle size distribution diagram of Lira NP1;

[0041] Figure 5 Particle size distribution diagram of Lira NP1 without adding aluminum ions;

[0042] Figure 6 Transmission electron microscope image of Lira NP1, scale bar 200 nm;

[0043] Figure 7 Effect of HTCC on the particle size, PDI and zeta potential of Lira NP1;

[0044] Figure 8 Particle size distribution diagram of Lira NP2;

[0045] Figure 9 Transmission electron microscope image of Lira NP2, scale bar 200 nm;

[0046] Figure 10 Potential comparison between optimized Lira NP1 and Lira NP2;

[0047] Figure 11 Evaluation of the stability of Lira NP2 in water, expressed by the changes in particle size and PDI within 30 days;

[0048] Figure 12The change in the viability of Caco-2 cells with the concentration of Lira;

[0049] Figure 13 The in vitro release profile of Lira NP2 in double-distilled water (pH = 7.4);

[0050] Figure 14 Confocal images of FITC-Lira NPs permeating mouse skin at 4 and 12 hours after administration (scale bar 250 μm);

[0051] Figure 15 The fluorescence optical density of FITC-Lira NPs in the stratum corneum of mouse skin at 4 and 12 hours after administration respectively;

[0052] Figure 16 The fluorescence optical density of FITC-Lira NPs in the epidermis and dermis of mouse skin at 4 and 12 hours after administration respectively.

[0053] Figure 17 The change in blood glucose levels of obese mice after administration in the injection group (10 mg / kg) and the transdermal administration group (10 mg / kg);

[0054] Figure 18 The change in body weight levels of obese mice after administration in the injection group and the transdermal administration group. Detailed implementation mode

[0055] The content of the present invention will be further described in detail through specific examples below. The raw materials used in the following examples can be obtained from conventional commercial channels or prepared and separated by simple synthesis, unless otherwise specified; the processes adopted, unless otherwise specified, are conventional processes in the art.

[0056] Example 1

[0057] This example provides a liraglutide nanoparticle, which is completed by two-step synthesis. First, multiple phenolic hydroxyl groups in TA can serve as hydrogen bond interaction sites to bind with Lira, preparing drug-loaded nanoparticles (Lira NP1), and introducing Al 3+ , to improve the stability of the particles. Subsequently, through charge interaction, the obtained Lira NP1 is combined with positively charged quaternized chitosan (HTCC) to synthesize drug-loaded nanoparticles (Lira NP2). And to ensure the rapid and uniform mixing of each component, a fast nanoparticle mixing system (FNC-R12.5, Guangzhou Guorui Scientific Instruments Co., Ltd.) provides key kinetic control for this synthesis process. The synthesis schematic diagram is as Figure 1 shown, and the specific step scheme is as follows:

[0058] 1. Prepare the Lira solution: Dissolve Lira in deionized water (pH = 7.5) at a concentration of 0.3 mg / mL; prepare the AlCl₃·6H₂O solution: Dissolve AlCl₃·6H₂O in deionized water, and the concentration of Al 3+ is 0.05 mg / mL; prepare the tannic acid solution: Dissolve TA in 50 mM HEPES buffer (pH = 5.0) at concentrations of 0.15, 0.30, 0.45, and 0.60 mg / mL.

[0059] Place the Lira solution in the first channel, the TA solution in the second channel, and the AlCl₃ solution in the third channel. The volume ratio and flow rate ratio of the three channels are 1:2:2; adjust the flow rate of the first channel to 0.10, 0.20, 0.30, 0.40 mL / s to fully mix the three solutions in the vortex mixing zone to obtain the nano-composite core of Lira and TA (Lira NP1).

[0060] 2. Dissolve HTCC in double-distilled water at concentrations of 0.20, 0.30, 0.40, and 0.60 mg / mL, place the Lira NP1 solution obtained in the first step in the first and second channels, and the HTCC aqueous solution in the third and fourth channels. The flow rates of the four channels are the same. Adjust the channel flow rate to 0.3 mL / s to obtain liraglutide nanoparticles (Lira NP2).

[0061] Comparative Example 1

[0062] This comparative example is different from Example 1 in that no aluminum ions were added in the preparation of Lira NP1. The specific steps are as follows:

[0063] Prepare the Lira solution: Dissolve Lira in deionized water (pH = 7.5) at a concentration of 0.3 mg / mL; prepare the tannic acid solution: Dissolve TA in 50 mM HEPES buffer (pH = 5.0) at a concentration of 0.30 mg / mL.

[0064] Place the Lira solution in the first channel and the TA solution in the second channel. The volume ratio and flow rate ratio of the two channels are 1:2; adjust the flow rate of the first channel to 0.30 mL / s to fully mix the two solutions in the vortex mixing zone to obtain the nano-composite core of Lira and TA (Lira NP1).

[0065] Optimization of process conditions

[0066] During the preparation process of FNC, in the early stage, it is a stage to establish a stable flow. The outflow solution may contain preliminary unknown products, so the first 1 mL of the product is discarded.

[0067] 1. Optimization of the preparation process conditions of Lira NP1

[0068] When the TA concentration was increased from 0.15 to 0.3 mg / mL, Lira fully combined with TA to prepare nanoparticles with small and uniform particle sizes. The surface potential of Lira NP1 decreased from -17 mV to -23 mV, the particle size decreased from 23 nm to 19 nm, and the PDI increased from 0.19 to 0.21( Figure 2 ). However, when the TA concentration was increased to 0.6 mg / mL, Al3+ could not effectively inhibit the complexation of TA and Lira to form larger nanoparticles, resulting in an increase in the particle size of the synthesized Lira NP1 (20 to 60 nm) and an increase in PDI to 0.4; further increasing the TA concentration caused the particle size to continue to increase to 30 - 100 nm, and the PDI remained at a high level (about 0.40), and further increasing the TA concentration did not change the surface potential of Lira NP1 (-23 mV to -25 mV). Therefore, when the TA concentration was selected as 0.3 mg / mL, Lira NP1 had the smallest particle size and uniform distribution, and this condition was determined as the optimal condition for preparing Lira NP1.

[0069] In addition, the channel flow rate has a significant impact on the mixing uniformity of each component in the FNC mixing chamber, thereby affecting the properties of the nanoparticles. As Figure 3 shown, when the channel flow rate increased from 0.1 mL / min to 0.4 mL / min, the PDI did not change significantly (0.19 to 0.23). It is worth noting that when the flow rate reached 0.3 mL / min, the particle size decreased from 34 nm to 20 nm; after the flow rate continued to increase, the particles increased to 40 nm. Therefore, 0.3 mL / min was finally selected as the preferred condition for preparing Lira NP1. The particle size distribution diagram of Lira NP1 prepared under the optimized conditions showed an obvious and uniform single peak( Figure 4 ). Without Al 3+ , the complexes formed by TA and Lira were prone to aggregation and were non-uniform, resulting in a multi-peak result in dynamic light scattering (DLS)( Figure 5 ). The size of Lira NP1 observed under TEM was consistent with the dynamic light scattering (DLS) data, showing a uniform spherical shape with an average diameter of 12 nm( Figure 6 ).

[0070] The above results indicate that the uniform distribution of Al 3+ is crucial for the formation of stable and uniformly sized nanoparticles during the fast nano-composite (FNC) process. Research shows that by increasing the mixing rate, Al 3+ can be better distributed, controlling the growth of nanoparticles, and thus obtaining smaller and more uniform particles.

[0071] 2. Optimization of the Preparation Process Conditions of Lira NP1

[0072] Similarly, using the FNC technology, the prepared HTCC was wrapped around the periphery of the particles. From Figure 7 It can be seen that when the concentration of HTCC increased from 0.2 mg / mL to 0.3 mg / mL, the particle size of Lira NP2 increased from 32 nm to 41 nm, and the surface potential increased from 24 mV to 30 mV. When the concentration of HTCC continued to increase from 0.3 mg / mL to 0.6 mg / mL, the changes in particle size and surface potential were not significant. The particle size remained between 40 and 42 nm, and the surface potential remained between 29 and 32 mV. In addition, when the concentration of HTCC increased from 0.2 mg / mL to 0.6 mg / mL, the PDI also did not show significant changes and remained between 0.22 and 0.24. We speculate that when the concentration of HTCC is 0.3 mg / mL, the HTCC loaded on the surface of Lira NP1 has reached saturation. Therefore, the concentration of HTCC was determined to be 0.3 mg / mL as the optimal condition for the preparation of LiraNP2, and the particle size distribution diagram showed an obvious and uniform single peak ( Figure 8 ). Lira NP2 observed under TEM presented a uniform spherical structure ( Figure 9 ), with an average diameter of 22 nm. The particle size was significantly increased compared to that of Lira NP1. From the surface charge reversal ( Figure 10 ) and the significant increase in particle size, both can confirm the successful coating of HTCC. The calculation results showed that the encapsulation efficiency (EE) of Lira was 47.3%, and the drug loading rate (LC) was 28.4%. In addition, the prepared Lira NP2 maintained a constant particle size even after being stored at 4 °C for 30 days, indicating its good stability ( Figure 11 ).

[0073] Experimental Analysis

[0074] 1. To study the biocompatibility of Lira NP2, different concentrations of Lira NP2 were co-incubated with Caco-2 cells for 24 hours, and the cell viability was detected by the CCK-8 method. From Figure 12 It can be seen that within the measured drug concentration range (15.6 - 250 μg / mL), the cell viability exceeded 50%, indicating that Lira NP2 has low cytotoxicity and good biocompatibility. And the pH environment of simulated body fluid was used in a solution with pH 7.4 to study the in vitro drug release behavior of Lira NP2. As Figure 13 shown, at 37 °C, Lira was rapidly released within the first 8 hours, and the release amount reached 36%. Subsequently, the release rate gradually slowed down, and the cumulative release amount reached the maximum value of 57% on the 4th day.

[0075] 2. Transdermal Drug Delivery Experiment Test

[0076] (1) Skin penetration of Lira NPs in obese mice.

[0077] Methods: Liraglutide loaded with FITC (fluorescein isothiocyanate, as a fluorescent dye) was prepared into liraglutide-loaded nanoparticles (FITC-Lira NPs), and was topically administered (at a dose of 10 mg / kg) to the back skin of obese mice through a glycerogelatin matrix. At 4 hours and 10 hours after administration, the mice were sacrificed by overdose anesthesia, and back skin tissue samples were obtained and made into frozen sections. To better observe the penetration of FITC-Lira NPs in the skin, KLK7 staining was performed to distinguish the stratum corneum from other skin layers. Incubate with KLK7 antibody (1:100, diluted with 5% goat serum) at 4 °C for 24 hours. After washing with PBS solution, immediately incubate with Alexa Fluor-647 secondary antibody at 37 °C for 1 hour. After washing with PBST, counterstain the cell nuclei with DAPI (1:300, diluted with PBS), cover the sections for 15 minutes to stain the cell nuclei in the tissue, and finally mount the sections with an anti-fluorescence quenching mounting medium. Finally, to observe the distribution of FITC-Lira NPs in the skin of obese mice, the skin tissue sections were imaged and analyzed at 100-fold magnification using a laser confocal microscope. ImageJ software was used to quantitatively analyze the average fluorescence intensity of FITC-Lira NPs in different layers (stratum corneum, epidermis, and dermis) of mouse skin. Through fluorescence microscope images, regions of each skin layer were selected respectively, and the fluorescence signals of each region were measured and their average fluorescence intensity was calculated to evaluate the distribution of FITC-Lira NPs in different skin layers.

[0078] Results: As Figure 14 shown, Lira NP2 encapsulated with HTCC successfully penetrated the stratum corneum and reached the epidermis and dermis 4 hours after administration, while Lira NP1 mainly concentrated near the stratum corneum and failed to penetrate deeper skin layers. Figure 15 、 16 The quantitative analysis in

[0079] (2) Hypoglycemic effect of a single dose of Lira NP2 on an obese mouse model

[0080] Method: By monitoring the blood glucose changes in obese mice after Lira NP2 administration, the hypoglycemic effect of the nanoparticles loaded with liraglutide was evaluated. Specifically, first, the model mice were randomly divided into a control group, an injection administration group, and a transdermal administration group (5 mice in each group), and single injection administration and transdermal administration were carried out respectively. During the whole experiment, the experimental mice could freely access food. At 1, 4, 8, 24, 48, 72, and 96 hours after administration, blood was collected from the tail vein, and the blood glucose level was measured by a blood glucose meter (590, Yuwell, China).

[0081] Results: As Figure 17 shown, the blood glucose value in the injection administration group decreased significantly to 70% of the normal blood glucose level 1 hour after administration, and continued to decrease to 62% within 8 hours, reaching the lowest value. In the transdermal administration group, the blood glucose value began to decline 8 hours after administration, and the decline rate was relatively slow. The blood glucose value reached the lowest value of 71% at 48 hours. It is worth noting that compared with direct injection of Lira (the hypoglycemic effect lasts for 48 to 72 hours), the transdermal administration group showed a more persistent hypoglycemic effect (the hypoglycemic effect lasts until 96 hours or even longer). The above results indicate that it takes several hours for the drug to penetrate the skin and then play a role in the mouse body, and the encapsulation of Lira NPs has a sustained-release effect, thus prolonging the treatment time of the drug.

[0082] (3) Therapeutic effect of Lira NP2 on obesity in obese mouse model

[0083] Method: By monitoring the body weight changes of obese mice after Lira NP2 administration, the effect of the nanoparticles loaded with liraglutide in the treatment of obesity was evaluated. First, the model mice were randomly divided into a control group, an injection administration group, and a transdermal administration group (5 mice in each group), and administration was carried out every four days as a cycle, with a total of 3 administrations. During the whole experiment, the experimental mice could freely access food. The body weight changes of each obese mouse were monitored daily and compared with the body weights of normal mice of the same age without modeling.

[0084] Results: As Figure 18Results showed that the body weights of the control group mice without drug treatment remained basically stable throughout the experiment. By the 12th day, their body weights were still maintained at over 95% of the initial body weight, indicating that under free-feeding conditions, the body weights of the model obese mice showed no significant changes. In contrast, the body weights of the mice in the injection group and the transdermal administration group both showed a significant downward trend. By the 12th day, the body weight of the injection group was 87% of the initial body weight, and that of the transdermal administration group was 78% of the initial body weight, which indicated that the transdermal administration group was superior to the injection group in the treatment effect of obesity. Further analysis of the body weight change trends of each administration group found that on the first day after drug administration (on the 0th, 4th, and 8th days), the body weight of the injection group decreased significantly, but there was an obvious body weight rebound within the 2nd to 4th days after drug administration. While the transdermal administration group showed a more persistent drug effect after administration, the body weight of the obese mice continued to decrease and was maintained for at least 4 days, and this result was consistent with that in the hypoglycemic experiment of the obese mouse model. This might be due to the limitation of the administration area. When a certain amount of drug was applied to the skin, the absorption of the drug reached saturation, and increasing the dose could not significantly increase the total amount of drug entering the body, resulting in the high-dose group not showing a more significant weight loss effect.

[0085] In summary, the present invention provides a liraglutide nanoparticle with a core-shell structure. Multiple phenolic hydroxyl groups of the phenolic hydroxyl polyphenolic compound can serve as hydrogen bond action sites to bind with liraglutide to form nanoparticles, filling the technical gap of liraglutide nanoparticles. Then, it combines with positively charged quaternized chitosan (HTCC) to form a liraglutide nanoparticle with a core-shell structure. HTCC has a high concentration of positive charges on its surface, which can form strong electrostatic adsorption with the negatively charged phospholipid bilayer of the skin stratum corneum and has a stronger skin penetration ability compared with chitosan, contributing to its application in transdermal drug delivery preparations.

[0086] The above are the preferred embodiments of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and refinements can still be made, and these improvements and refinements are also regarded as the protection scope of the present invention.

Claims

1. A transdermal liraglutide nanoparticle preparation, characterized in that: The liraglutide nanoparticles are of a core-shell structure, wherein the core comprises a complex of liraglutide and a phenolic hydroxyl polyphenol compound, and the shell is quaternized chitosan.

2. A polypeptide drug molecule nanoparticle, characterized in that: The polypeptide drug molecule nanoparticles are of a core-shell structure, wherein the core comprises a complex of a polypeptide drug molecule and a phenolic hydroxyl polyphenol compound, and the shell is quaternized chitosan; the polypeptide drug molecule nanoparticles comprise liraglutide or other polypeptide drug molecules.

3. The polypeptide drug molecule nanoparticle according to claim 2, characterized in that: The particle size of the polypeptide drug molecule nanoparticles is 10-50nm.

4. The polypeptide drug molecule nanoparticle according to claim 2, characterized in that: The phenolic hydroxyl polyphenol compound is selected from at least one of tannic acid, gallic acid and ellagic acid.

5. The method for preparing the polypeptide drug molecule according to any one of claims 2 to 4, characterized in that: The following steps are involved: S1. Using a rapid nanocomposite method, a polypeptide drug molecule solution, a phenolic hydroxyl polyphenol compound solution and an aluminum ion solution are mixed to obtain a solution containing a complex of polypeptide drug molecules and phenolic hydroxyl polyphenol compounds; S2. The solution containing the complex of the polypeptide drug molecule and the phenolic hydroxyl polyphenol compound is mixed with the quaternized chitosan solution by a rapid nanocomposite method to obtain the polypeptide drug molecule nanoparticles.

6. The method for preparing the polypeptide drug molecule according to claim 5, characterized in that: The solvent of the polypeptide drug molecule solution is water, and the concentration is 0.1-1 mg / mL; and / or, the solvent of the phenolic hydroxyl polyphenol compound solution is a zwitterionic buffer with a concentration of 0.1-1 mg / mL; And / or, the solvent of the aluminum ion solution is water, and the concentration is 0.01-0.2 mg / mL; And / or, the volume ratio of the polypeptide drug molecule solution, the phenolic hydroxyl polyphenol compound solution and the aluminum ion solution is 1:(1-3):(1-3).

7. The method for preparing the polypeptide drug molecule nanoparticles according to claim 5, characterized in that: The solvent of the quaternized chitosan solution is water, and the concentration is 0.1-1 mg / mL; And / or, the volume ratio of the solution containing the complex of polypeptide drug molecules and phenolic hydroxyl polyphenol compounds to the quaternized chitosan solution is 1:(0.5-1.5).

8. The method for preparing the polypeptide drug molecule nanoparticles according to claim 5, characterized in that: The rapid nanocomposite process was carried out in a multi-channel vortex mixer.

9. The method for preparing the polypeptide drug molecule nanoparticles according to claim 8, characterized in that: The polypeptide drug molecule solution, the phenolic hydroxyl polyphenol compound solution and the aluminum ion solution are respectively mixed in the vortex mixing area through different channels in the multi-channel vortex mixer, wherein the flow rate ratio of the polypeptide drug molecule solution, the phenolic hydroxyl polyphenol compound solution and the aluminum ion solution in the channel is 1:(1-3):(1-3).

10. Use of the polypeptide drug molecule nanoparticles according to any one of claims 2 to 4 in the preparation of drugs, characterized in that: The dosage form of the drug includes a skin administration agent.

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