A uniform, ultrafine polymer nanomicelle, its preparation method, and its application.

Abstract

This invention discloses a uniform, ultrafine polymer nanomicelle, its preparation method, and its applications. The polymer nanomicelle is first formed by modifying a material based on poly(lactic-co-liquid) with ligands containing double bonds to create a polymer containing unsaturated bonds. The polymer then encapsulates berberine (BBR) via a self-emulsification process, followed by UV cross-linking and pH adjustment. The polymer nanomicelle uses materials obtained from FDA-approved excipient PLGA through simple modification, and subsequently, drug-loaded nanomicelles with a particle size of less than 10 nm are prepared via self-emulsification. Due to their small particle size, the nanomicelles can be transported through the intercellular spaces of epithelial cells, effectively improving drug bioavailability. Combined use with the intestinal permeabilizer SNAC can further enhance the transintestinal absorption capacity of the nanoparticles. The polymer nanomicelles exhibit high uniformity, good stability, good safety and biocompatibility, and no toxic side effects, making them a promising candidate for treating type 2 diabetes.

Description

Technical Field

[0001] This invention relates to a uniform, ultrafine polymer nanomicelle, its preparation method, and its application, belonging to the fields of polymer materials science and pharmaceutical formulation. Background Technology

[0002] Diabetes mellitus is a chronic disease characterized by hyperglycemia. It is a metabolic endocrine disorder, with type 2 diabetes accounting for the majority of diabetes patients in my country. Patients with long-term type 2 diabetes often suffer from serious complications (Nutr Metab (Lond), 2017, 14:60). In addition to insulin, berberine hydrochloride (BBR) has been shown to improve lipid metabolism disorders in patients with type 2 diabetes by inhibiting mitochondrial function, stimulating glycolysis, and activating the AMPK pathway, thereby indirectly lowering blood glucose levels (Chinese Journal of Traditional Chinese Medicine, 2014, 39(08):1374-1378). According to previous research (CN115531309A), BBR can reduce blood glucose levels in STZ-induced type 2 diabetic mice, improve insulin tolerance and glucose tolerance, and at the same time weaken glucose production and gluconeogenesis gene expression in hepatocytes induced by glucagon.

[0003] Due to the mucus barrier, berberine has very low direct oral bioavailability. Therefore, many studies have used nanoparticles to encapsulate and deliver the drug. Polymer materials are widely used in the preparation of nanoparticle carriers due to their good biocompatibility. However, common polymer carrier materials are simple in design and cannot overcome the complex physiological barriers of the gastrointestinal tract, limiting their effectiveness in improving drug bioavailability. Many studies have improved the internalization efficiency of hypoglycemic drugs through complex polymer carrier designs or by modifying carriers with ligands of multiple functions. However, complex ligand designs and compound structures increase costs and safety risks, hindering product updates and industrialization. Furthermore, there is still considerable room for improvement in the oral bioavailability of these drugs.

[0004] Significant progress and innovations have been made in carrier design for the use of nanoparticles in oral drug delivery. However, the impact of nanoparticle size on absorption has not been extensively studied and applied. Some literature has explored the efficiency of gold nanoparticles with sizes of 5, 10, and 20 nm in penetrating the epithelial cell barrier, finding that 5 nm gold nanoparticles can induce nanoscale gaps between endothelial cells, enabling nanoparticles to cross the endothelial cell barrier via the paracellular pathway. The ultrafine particle size of ultrafine nanoparticles inherently provides a certain transmembrane capacity, allowing them to exert transmembrane efficacy without complex carrier design and modified ligands. However, conventional ultrafine nanoparticle preparation processes are complex, and polymer ultrafine nanomicelles are rare. Common ultrafine nanoparticles are mostly prepared from materials such as MOFs, but MOFs and similar materials have poor biocompatibility, pose safety risks, and require stringent preparation conditions, resulting in poor stability, which is detrimental to the development of oral drugs. Summary of the Invention

[0005] Objectives of this invention: The first objective is to provide a uniform, ultrafine polymer nanomicelle; the second objective is to provide a method for preparing the uniform, ultrafine polymer nanomicelle; and the third objective is to provide the application of the uniform, ultrafine polymer nanomicelle in the preparation of drugs for treating type 2 diabetes. The uniform, ultrafine polymer nanomicelle of this invention is a biodegradable polymer material, and it leverages its ultrasmall particle size to exert transmembrane activity, ultimately achieving effective internalization of hypoglycemic drugs.

[0006] Technical solution: The present invention provides a uniform and ultrafine polymer nanomicelle, wherein the polymer nanomicelle comprises a material mainly composed of poly(lactic acid) glycolide (PLGA) modified with ligands containing double bonds to form a polymer containing unsaturated bonds, the polymer then encapsulates berberine (BBR) in a self-emulsifying manner, crosslinks it under ultraviolet light, and adjusts the pH to form the final product.

[0007] Furthermore, the ligand containing a double bond is a hydroxyl-containing double bond ligand or an acyl halide-containing double bond ligand.

[0008] Furthermore, the hydroxyl-containing double bond ligand is hydroxyethyl acrylate (HEA), allyl alcohol, or 3-buten-2-ol.

[0009] Furthermore, the double-bonded ligands containing acyl halides are methacryloyl chloride, acryloyl chloride, or acryloyl bromide.

[0010] Furthermore, the material based on poly(lactic acid lactide) (PLGA) is PLGA, polylactic acid-glycolic acid copolymer-polyethylene glycol (PLGA-PEG), or polyethylene glycol dimer-lactic acid-glycolic acid copolymer (PLGA-PEG-PLGA).

[0011] Furthermore, in PLGA-PEG or PLGA-PEG-PLGA, the molecular weight of PLGA is much larger than that of polyethylene glycol (PEG), and the mass ratio of lactide (LA) to glycolide (GA) in PLGA is 25:25-75.

[0012] Furthermore, the self-emulsifying method for encapsulating berberine involves first dissolving the polymer in an organic solvent, adding linear high-grade fatty acids and an emulsifier, mixing thoroughly, and then adding berberine. Using microfluidic technology, an oil-in-water biphase microcapsule is prepared with the organic solvent phase as the inner phase and water as the outer phase.

[0013] The method for preparing uniform ultrafine polymer nanomicelles according to the present invention includes the following steps:

[0014] (1) Preparation of polymers containing unsaturated bonds:

[0015] In an anhydrous, oxygen-free, and nitrogen-protected environment, the carboxyl-terminated PLGA-based material was dissolved in an organic solvent, a catalyst was added, and the mixture was stirred at room temperature to activate the carboxyl groups. A hydroxyl-containing double bond ligand was added, and the reaction was carried out at room temperature. The mixture was dialyzed, precipitated with ice-cold ether, and the lower precipitate was collected by centrifugation, washed, and dried to obtain a polymer containing unsaturated bonds.

[0016] Alternatively, under nitrogen protection, the hydroxyl-terminated PLGA-based material is dissolved in an organic solvent, and a double-bonded ligand containing an acyl halide is slowly added dropwise in an ice bath. Triethylamine is added as a binding acid agent, the reaction is carried out at room temperature, dialyzed, precipitated with ice-cold ether, and the lower precipitate is collected by centrifugation, washed, and dried to obtain a polymer containing unsaturated bonds.

[0017] (2) Preparation of uniform ultrafine polymer nanomicelles:

[0018] A polymer containing unsaturated bonds was dissolved in an organic solvent to obtain a polymer solution containing unsaturated bonds. A linear high fatty acid and a drop of emulsifier were added and mixed evenly. Berberine methanol solution was then added to obtain an organic phase. Using microfluidic technology, a homogeneous oil-in-water microcapsule was prepared with the organic phase as the inner phase and water as the outer phase. The microcapsules were collected in NaOH solution, and the microcapsules spontaneously emulsified and dispersed to form a polymer nanomicelle solution.

[0019] (3) Add a crosslinking agent to the polymer nanomicelle solution and crosslink the unsaturated bonds under a UV lamp to obtain a uniform and ultrafine polymer nanomicelle solution.

[0020] Further, in step (1), the organic solvent is N,N-dimethylformamide (DMF), dichloromethane (DCM), or tetrahydrofuran (THF).

[0021] Further, in step (1), the carboxyl-terminated PLGA-based material is a carboxyl-terminated poly(lactic acid-glycolic acid) copolymer (PLGA-COOH) or a polyethylene glycol dimer lactic acid-glycolic acid copolymer (PEG-PLGA-COOH).

[0022] Further, in step (1), the hydroxyl-terminated PLGA-based material is a hydroxyl-terminated polyethylene glycol dimer lactic acid-glycolic acid copolymer (OH-PLGA-PEG-PLGA-OH).

[0023] Furthermore, in PLGA-based materials, the mass ratio of PLGA to PEG is 5-10:1, and the mass ratio of LA to GA is 50:50.

[0024] Further, in step (1), the catalyst is 4-dimethylaminopyridine (DMAP) and dicyclohexylcarbodiimide (DCC), or carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS).

[0025] Further, in step (1), the hydroxyl-containing double bond ligand is hydroxyethyl acrylate (HEA).

[0026] Further, in step (1), the acyl halide-containing double bond ligand is methacryloyl chloride.

[0027] Furthermore, in step (1), the molar ratio of the carboxyl-terminated PLGA-based material, the catalyst, and the hydroxyl-containing double-bonded ligand is 1:1.5:2-5.

[0028] Furthermore, in step (1), the molar ratio of the hydroxyl-terminated PLGA-based material, the acyl halide-containing double bond ligand, and triethylamine is 1:20-30:30-40.

[0029] Furthermore, in step (2), the device used in the microfluidic technology is an oil-in-water (O / W) two-phase microfluidic device, with an inner phase pore size of 200-400 μm, an outer phase inner diameter of 600-800 μm, and an inner phase to outer phase flow rate ratio of 5-10:1.

[0030] Further, in step (2), the organic solvent is DCM or chloroform, and the concentration of the polymer solution containing unsaturated bonds is 20-30 mg / mL.

[0031] Furthermore, in step (2), the linear higher fatty acid is a linear fatty acid with a C value of 10-14.

[0032] Furthermore, straight-chain fatty acids with a C value of 10-14 are either decanoic acid or lauric acid.

[0033] Further, in step (2), the emulsifier is Span 80.

[0034] Furthermore, in step (2), the concentration of the NaOH solution is 0.1-0.25 mol / L.

[0035] Furthermore, in step (2), the straight-chain higher fatty acids react with OH- - The molar ratio is 1:20-50.

[0036] Furthermore, in step (2), the concentration of the berberine methanol solution is 20-50 mg / mL.

[0037] Furthermore, in step (2), the volume ratio of the polymer solution containing unsaturated bonds to the higher fatty acids in the straight chain is 1:0.5-2.

[0038] Furthermore, in step (3), the crosslinking agent is photoinitiator 2959 (I2959).

[0039] Furthermore, in step (3), the power of the ultraviolet lamp is 13000 uw / cm. 2 above.

[0040] Furthermore, in step (3), the crosslinking time is 10-20 min.

[0041] Furthermore, in step (3), evaporation is carried out at a temperature below 40°C, and the pH is adjusted to neutral.

[0042] The application of the uniform, ultrafine polymer nanomicelles described in this invention in the preparation of drugs for treating type 2 diabetes.

[0043] Furthermore, during the self-emulsification process, berberine hydrochloride is added to combine with the polymer containing unsaturated bonds to form a drug-loaded polymer micelle solution, which is then cross-linked under ultraviolet light, adjusted to neutral pH, and used in combination with a penetration enhancer.

[0044] Furthermore, the penetration enhancer is sodium 8-(2-hydroxybenzamido)octanoate (SNAC for short).

[0045] The preparation process is preferred.

[0046] (1) Preparation of polymer materials containing unsaturated bonds:

[0047] Under an anhydrous, oxygen-free, and nitrogen-protected environment, carboxyl-terminated PLGA-COOH was first dissolved in DCM, then the catalysts DMAP and DCC were added, and the mixture was stirred at room temperature for 30 min to activate the carboxyl groups. After activation, HEA (PLGA to HEA molar ratio of 1:2-5) was added, and the reaction was carried out at room temperature for 12 h. After the reaction, the reaction solution was placed in a 3.5 kDa dialysis bag and dialyzed for 3 h. After dialysis, the solution was precipitated with ice-cold diethyl ether (4-6 times the volume of the reaction solution), and the lower precipitate was collected by centrifugation. The precipitate was then vacuum dried to obtain the polymer material PLGA-HEA (abbreviated as PH) containing unsaturated bonds.

[0048] Alternatively, when hydroxyl-terminated PLGA-PEG-PLGA (abbreviated as OH-PPP-OH, PLGA to PEG mass ratio of 8-10:1) reacts with a double-bonded ligand containing an acyl halide, the hydroxyl-terminated OH-PPP-OH is first dissolved in tetrahydrofuran under nitrogen protection. Methacrylamide halide (OH-PPP-OH to HEA molar ratio of 1:20-30) is slowly added dropwise in an ice bath, along with a small amount of triethylamine (OH-PPP-OH to triethylamine molar ratio of 1:30-40) as an acid-binding agent. After the addition is complete, the mixture is transferred to room temperature and reacted for 12 hours. After the reaction, the reaction solution is dialyzed in a 3.5 kDa dialysis bag for 3 hours. After dialysis, the precipitate is collected by precipitating with ice-cold diethyl ether and centrifuging. The precipitate is washed, dried, and the resulting polymer material containing unsaturated bonds (abbreviated as PPP) is obtained.

[0049] (2) Preparation of uniform ultrafine polymer nanomicelles:

[0050] A berberine (BBR) solution with a concentration of 35 mg / mL was prepared in advance using a methanol solution, and a two-phase (oil-in-water) microfluidic device was constructed.

[0051] First, a polymer material containing unsaturated bonds was dissolved in DCM to prepare a 40 mg / mL polymer solution. Then, a certain amount of lauric acid (LA, with a volume ratio of LA to DCM of 1:1-3) was added as an excipient, along with the emulsifier Span 80. After mixing thoroughly, an appropriate amount of BBR solution was added to prepare nanomicelles with a drug loading of 20 wt%. Using the polymer solution as the inner phase and water as the outer phase, uniform microcapsules were prepared using a microfluidic device at an inner-phase to outer-phase flow rate ratio of 8-10:1. The pore size of the inner phase of the microfluidic device was 200-300 μm, and the inner diameter of the glass tube was 800 μm. The microcapsules were collected and placed in a 0.1-0.2 M NaOH solution (with a volume ratio of DCM to NaOH of 1:20-40). The microcapsules spontaneously emulsified and dispersed due to the slow reaction of linear higher fatty acids with OH-, ultimately forming uniform ultrafine polymer nanomicelles.

[0052] Berberine has been shown to indirectly control blood sugar in patients with type 2 diabetes. However, its direct oral bioavailability is low. Existing studies often employ complex polymer carrier designs to achieve effective internalization of hypoglycemic drugs, but these designs are complex and their oral bioavailability still has room for improvement. Other studies have used MOFs and other materials to prepare ultrasmall nanoparticles, which can leverage their ultrasmall particle size to achieve transmembrane activity and improve drug bioavailability. However, MOFs and other materials have poor stability and biocompatibility. Based on these studies, we propose the following: can commonly used polymer materials be prepared into ultrasmall nanoparticles to simplify material design and improve oral drug bioavailability? Based on this proposal, this invention discloses a method for preparing uniform ultrafine polymer nanomicelles and their applications. The polymer material is obtained by simple modification of FDA-approved excipient PLGA, followed by self-emulsification to prepare drug-loaded nanomicelles with a particle size of less than 10 nm. The nanomicelles exhibit high particle size uniformity, good stability, and great application potential.

[0053] The uniform ultrafine polymer nanomicelles of the present invention are first prepared by esterification condensation of PLGA-COOH (carboxyl-terminated) and OH-PPP-OH (two hydroxyl-terminated) to obtain two polymer materials PH and PPP containing unsaturated bonds. Then, drug-loaded microcapsules are prepared using a microfluidic device, and the microcapsules are finally prepared into uniform ultrafine polymer nanomicelles by self-emulsification.

[0054] This invention optimizes and explores the conditions for preparing uniform ultrafine polymer nanomicelles via self-emulsification. It investigates the effects of polymer type and concentration, excipient type and volume, and NaOH concentration and volume ratio on the particle size, uniformity, and dispersion of the nanomicelles. The method for preparing uniform ultrafine polymer nanomicelles via self-emulsification is improved, and the optimal preparation scheme is selected. The nanomicelles have a particle size below 10 nm, which allows them to be transported through the intercellular spaces of epithelial cells, effectively improving their bioavailability. The application of UV crosslinking technology and the protective effect of SNAC enhance the stability of the nanomicelles in the stomach and achieve long-term drug circulation. The uniform ultrafine polymer nanomicelles are simple in material composition, have high uniformity, and the excipients are already clinically approved, demonstrating great application potential, particularly in the treatment of diabetes.

[0055] Beneficial effects: Compared with the prior art, the present invention has the following advantages:

[0056] (1) The present invention uses PLGA as an FDA-certified excipient, which has excellent safety. Furthermore, the modified ligands in the present invention are simple, the materials have good biocompatibility, and no toxic side effects.

[0057] (2) The uniform ultrafine polymer nanomicelles of this invention have a particle size of less than 10 nm. Due to their small particle size, they can be transported through the intercellular spaces of epithelial cells, effectively improving the bioavailability of the nanomicelles. When used in combination with the intestinal permeabilizer SNAC, the transintestinal absorption capacity of the nanoparticles can be further enhanced. The transmembrane capacity of the ultrafine nanomicelles is due to their own physical properties, requiring no excessive ligand modification, and is easy to prepare and has high safety.

[0058] (3) This invention utilizes double bonds on polymer materials containing unsaturated bonds to achieve ultraviolet light crosslinking and curing, thereby increasing the stability of the micelles. In the plasma pH environment, the crosslinked structure slowly cleaves and releases hypoglycemic drugs, which is less likely to cause hypoglycemia and achieves a long-lasting circulatory effect. Attached Figure Description

[0059] Figure 1 The hydrogen NMR spectrum of PLGA-HEA in Example 1;

[0060] Figure 2 The hydrogen NMR spectrum of PLGA-PEG-PLGA modified with methacryloyl chloride at both ends in Example 2;

[0061] Figure 3 The graph shows the relationship between NaOH concentration and hydrated particle size of nanoparticles in Example 3, and the relationship between PPP concentration and hydrated particle size of nanoparticles in the internal phase solution.

[0062] Figure 4 The particle size distribution of s-PPP@B NPs, PH@B NPs and b-PPP@B NPs in Example 4 is shown.

[0063] Figure 5 This is a diagram showing the simulated in vivo release results of photocrosslinked drug-loaded nanomicelles in Example 4;

[0064] Figure 6 This is a schematic diagram showing the cell viability of the excipients, polymer materials, and nanomicelles in Example 4, respectively, in the cytotoxicity assays of Caco-2 and HT-29 cells.

[0065] Figure 7 The uptake of nanomicelles after treatment with different nanomicelles and inhibitors in Example 4 is shown.

[0066] Figure 8 The polymer micelles P in Example 4 app Horizontal map;

[0067] Figure 9 The TEER value of the polymer micelles in Example 4;

[0068] Figure 10 This is a graph showing the in vivo blood glucose level of the drug-loaded polymer micelles in Example 4. Detailed Implementation

[0069] The technical solution of the present invention will be further described below with reference to the accompanying drawings.

[0070] Example 1

[0071] The synthesis route and process of PLGA-HEA are as follows:

[0072]

[0073] Under an anhydrous, oxygen-free, and nitrogen-protected environment in a glove box, 600 mg of carboxyl-terminated PLGA-COOH (M: 10000, LA:GA = 50:50) was accurately weighed and dissolved in 3 mL of anhydrous DCM. Simultaneously, 24.7 mg of DCC and 7.3 mg of DMAP were added as catalysts, and the mixture was stirred at room temperature for 30 min to activate the carboxyl groups. After activation, 34.8 mg of HEA (PLGA to HEA molar ratio of 1:5) was added, and the reaction was carried out at room temperature for 12 h. After the reaction, the reaction solution was placed in a 3.5 kDa dialysis bag and dialyzed against DCM solution to remove unreacted HEA and catalyst. After dialysis, the solution was precipitated with refluxed ether (4-6 times the volume of the reaction solution), and the lower precipitate was collected by centrifugation. The precipitate was then vacuum-dried to obtain the PLGA material. 10k -HEA (abbreviated as PH).

[0074] The PLGA-HEA prepared in this embodiment was subjected to proton nuclear magnetic resonance analysis. 10k -The hydrogen NMR characterization of HEA is attached. Figure 1 As shown, 1 The ¹H NMR (400MHz, Chloroform-d) peaks at δ 7.26–7.32 (s, 2H) correspond to double bond peaks, indicating that the unsaturated bonds were successfully modified onto the polymer, and the original LA, GA, and methyl peaks of the polymer were retained. Based on peak intensities, PLGA… 10k -HEA yield is approximately 69.72%.

[0075] Example 2

[0076] The synthesis of PPP, the synthesis route and process are as follows:

[0077]

[0078] Under nitrogen protection, 660 mg of hydroxyl-terminated OH-PLGA was dissolved in 3 mL of tetrahydrofuran. 10k -PEG 2k -PLGA 10k -OH(M: 22000, M) PLGA :M PEG=10:1). 94 μL of methacryloyl chloride (OH-PPP-OH to methacryloyl chloride molar ratio 1:20) was slowly added dropwise under ice bath conditions. After the addition was complete, the ice bath was removed, and 91 μL of triethylamine (OH-PPP-OH to triethylamine molar ratio 1:30) was added as an acid-binding agent. The mixture was sealed and transferred to room temperature for 12 hours of reaction. After the reaction, the reaction solution was first filtered through filter paper to remove the triethylamine salt, and then placed in a 3.5 kDa dialysis bag for dialyzing in tetrahydrofuran solution to remove unreacted raw materials and triethylamine. After dialysis, the solution was precipitated with ice-cold diethyl ether (4-6 times the volume of the reaction solution) and centrifuged to collect the lower precipitate. The precipitate was then vacuum dried to obtain a polymer material containing unsaturated bonds (PPP).

[0079] The PPP prepared in this embodiment was subjected to hydrogen nuclear magnetic resonance analysis. The hydrogen nuclear magnetic resonance characterization of PPP is shown in the appendix. Figure 2 As shown, 1 The ¹H NMR (400MHz, Sulfoxide-d) peaks at δ 5.72–5.81 (s, 2H) correspond to double bond peaks, indicating that the unsaturated bonds were successfully modified onto the polymer, and the original LA, GA, Peg, and methyl peaks of the polymer were retained. Based on the peak intensities, the PPP yield was calculated to be approximately 88.26%.

[0080] Example 3

[0081] The study and optimization of a method for preparing uniform ultrafine polymer nanomicelles are as follows:

[0082] (1) Preparation of internal phase solution

[0083] In preparing the polymer solution, the polymer material PPP obtained in Example 2 was first dissolved in DCM to prepare a 40 mg / mL polymer solution. After the polymer was completely dissolved, it was then added to the solution according to the volume ratio V... 脂肪酸 :V 聚合物溶液 Straight-chain medium-to-high fatty acids (using decanoic acid, lauric acid, and tetradecanoic acid, respectively) were added in ratios of 1:1, 1:2, and 1:3 as excipients, with 1 drop of Span 80 added as an emulsifier. The mixture was then used as the inner phase of a microfluidic device, with water as the outer phase (mobile phase). The pore size of the inner phase was 200-240 μm, the flow rate of the outer phase was 100 μL / min, and the flow rate of the inner phase was 10 μL / min (the flow rate ratio of the inner to outer phase was 10:1). Microcapsules were prepared and placed in a 0.2 mol / L NaOH solution (volume ratio V... 内相 :V NaOH Nanomicelles were prepared by ratio 1:20.

[0084] The formation of nanomicelles depends on the slow reaction of linear higher fatty acids with OH- ions. Therefore, the volume ratio of DCM to linear higher fatty acids should not be too large. After several attempts, it was found that V 脂肪酸 :V聚合物溶液 The optimal ratio of 1:1 resulted in the best nanomicelle state. Among the selected straight-chain higher fatty acids, decanoic acid produced nanomicelles with a particle size around 100 nm, making it impossible to prepare uniform ultrafine polymer nanomicelles; tetradecanoic acid had a high melting point and dissolved slowly at room temperature, making nanomicelle preparation difficult. Considering the later application conditions and preparation difficulty, V was ultimately selected. 脂肪酸 :V 聚合物溶液 Polymer solutions were prepared under a ratio of 1:3. Decanoic acid was used as the preparation material for the control group of nanomicelles, and lauric acid (LA, dodecanoic acid) was used as the preparation material for the experimental group of nanomicelles.

[0085] Based on the above conditions (selecting lauric acid, V...), 月桂酸 :V 聚合物溶液 =1:3) Continue to investigate the effects of NaOH concentration and polymer concentration in the internal phase, with other conditions as described above:

[0086] Microcapsules containing PPP spontaneously emulsified and dispersed in NaOH solution, eventually forming uniform ultrafine polymer nanomicelles. We investigated the relationship between NaOH concentration (0.05 mol / L, 0.10 mol / L, 0.15 mol / L, 0.20 mol / L, and 0.25 mol / L) and the concentration of PPP in the internal phase (1 mg / mL, 2 mg / mL, 4 mg / mL, 8 mg / mL, 12 mg / mL, and 24 mg / mL) and the hydrated particle size of the nanoparticles, as a guide for optimizing the preparation conditions of the nanomicelles. The results are as follows: Figure 3 As shown. Figure 3 This is a graph showing the relationship between NaOH concentration and hydrated particle size of nanoparticles in Example 3, and the relationship between PPP concentration and hydrated particle size of nanoparticles in the internal phase solution. In Figure (A), the graph shows the relationship between NaOH concentration and hydrated particle size of nanoparticles, and in Figure (B), the graph shows the relationship between PPP concentration and hydrated particle size of nanoparticles in the internal phase solution. Figure 3 It is evident that uniform ultrafine polymer nanomicelles cannot be prepared when the NaOH solution concentration is below 0.15 mol / L or the PPP concentration in the polymer solution is above 12 mg / mL. Therefore, the final experimental scheme selected was a NaOH solution concentration of 0.2 mol / L and a PPP concentration of 10 mg / mL in the inner phase solution.

[0087] Example 4

[0088] The preparation process is the same as in Example 3.

[0089] (1) Preparation of drug-loaded nanomicelles in the experimental group

[0090] A berberine (BBR) solution with a concentration of 35 mg / mL was prepared in advance by dissolving it in hot methanol.

[0091] First, dissolve the PPP prepared in Example 2 in DCM to prepare a PPP solution with a concentration of 40 mg / mL. Take 100 μL of the PPP solution, and then, according to the density of lauric acid (LA), weigh out and add 1 / 3 V PPP溶液 LA powder was mixed with 2 drops of Span 80 as an emulsifier. Then, 30 μL of BBR solution was added and mixed to form the internal phase, with water as the external phase. Drug-loaded microcapsules were prepared using a microfluidic device. The microcapsules were collected and placed in 2.5 mL of 0.2 mol / L NaOH solution, and allowed to stand at 40 °C for 12 h to prepare homogeneous ultrafine polymer nanomicelles.

[0092] After the nanomicelles were prepared, 5 μL of photoinitiator I2929 was added, and the mixture was subjected to ultraviolet light (13000 uw / cm²) under stirring. 2 Irradiation for 15 min induced photocrosslinking of the material. The prepared nanomicelle solution was denoted as s-PPP@B NPs, and particle size analysis was performed. The results are as follows: Figure 4 As shown in (A).

[0093] After crosslinking, the organic solvent was removed by heating and evaporation. Finally, the pH was adjusted to 7 with a 0.2 mol / L hydrochloric acid solution, and 15 mg of SNAC was added to obtain the s-PPP@B+SNAC micelle solution.

[0094] (2) Preparation of drug-loaded nanomicelles in the control group

[0095] The preparation process is the same as step (1), except that the pH prepared in Example 1 is dissolved in DCM to prepare a pH solution with a concentration of 40 mg / mL. The nanomicelle solution prepared in this example is denoted as PH@B NPs and particle size analysis is performed. The results are as follows. Figure 4 As shown in (B).

[0096] The preparation process is the same as step (1), except that the PPP prepared in Example 2 is dissolved in DCM to prepare a PPP solution with a concentration of 40 mg / mL. Then 1 / 3 V PPP溶液 A solution of n-decanoic acid (n-OA) was prepared. The nanomicelle solution prepared in this example was denoted as b-PPP@B NPs, and particle size analysis was performed. The results are as follows: Figure 4 As shown in (C).

[0097] Similarly, the pH of the nanomicelle solution in the above experiment was adjusted to 7, and 15 mg of SNAC was added to obtain PH@B+SNAC and b-PPP@B+SNAC micelle solutions, respectively.

[0098] Alternatively, BBR was directly dissolved in water to prepare a BBR solution with a concentration of 0.3 mg / mL. 1 mL of the prepared BBR solution was then added to 15 mg of SNAC to prepare a BBR+SNAC solution for later use.

[0099] Figure 4 The image shows the particle size distribution of s-PPP@B, PH@B, and b-PPP@B nanomicelles in Example 4, where (A) represents s-PPP@B NPs, (B) represents PH@B NPs, and (C) represents b-PPP@B NPs. Figure 4 It can be seen that the hydrated particle size is below 10nm, and the percentage is above 30%.

[0100] (3) Determination of encapsulation efficiency and drug loading of nanoparticles

[0101] Encapsulation efficiency (LE) and drug loading (LC) are commonly used to represent the drug loading capacity of nanomicelles. This invention employs an indirect method, using high-performance liquid chromatography (HPLC) to determine the content of free berberine hydrochloride.

[0102] The HPLC conditions were as follows: Dionex Ultimate 3000 high-performance liquid chromatography, with a C18 alkylsilane-bonded silica column (250 × 4.6 mm, 5 μm), acetonitrile as mobile phase A, and 0.1% potassium dihydrogen phosphate solution (pH 3.03) as mobile phase B, with an acetonitrile (A) : potassium dihydrogen phosphate solution (B) ratio of 30:70, v / v; the flow rate was fixed at 1 mL / min; the UV detector was set to λ = 345 nm, the column temperature was 25.0 °C, and the injection volume was 20 μL.

[0103] By measuring the mass of free berberine hydrochloride, the encapsulation efficiency and actual drug loading were further calculated. Drug loading is the percentage of drug mass encapsulated in the nanomicelles to the total mass (carrier and encapsulated drug mass). Mass encapsulation efficiency refers to the percentage of drug mass encapsulated in the nanoparticles to the total drug mass. The calculation formulas are as follows:

[0104]

[0105] Where, m 总 For the mass (mg) of BBR added, m 游 The mass (mg) of free BBR; m 药 m represents the mass (mg) of BBR encapsulated within nanomicelles. 载体 The value represents the mass (mg) of the polymer in the final prepared nanomicelles.

[0106] Following the preparation processes in steps (1) and (2) of this embodiment, PPP and pH solutions with a concentration of 40 mg / mL of DCM were prepared, and BBR solutions with a concentration of 35 mg / mL of methanol were added. Nanomicelles were prepared with BBR drug loadings of 10 wt%, 20 wt%, and 30 wt%, and were designated as PH@B-10, PH@B-20, PH@B-30, s-PPP@B-10, s-PPP@B-20, s-PPP@B-30, b-PPP@B-20, and b-PPP@B-30.

[0107] The nanoparticles in the polymer micelle solution prepared in this embodiment were characterized and analyzed, and the results are shown in Table 1.

[0108] Table 1 Characterization of drug-loaded polymer micelles

[0109]

[0110] As shown in Table 1, when the theoretical drug loading (i.e., BBR mass / final nanoparticle mass) is 10wt%, 20wt%, and 30wt%, the encapsulation efficiency of the polymer micelles for hypoglycemic drugs shows a slow decreasing trend, with the final encapsulation efficiency around 80%. Furthermore, the encapsulation ability exhibits the order PH@B > s-PPP@B > b-PPP@B. This may be because PH is a completely hydrophobic material, while PPP is an amphiphilic material. During the self-emulsification preparation of nanomicelles, the hydrophobic BBR is more easily captured by PH, thus resulting in a better encapsulation efficiency for the PH group than for the PPP group.

[0111] (4) Simulated in vivo release experiment of drug-loaded nanoparticles

[0112] According to this embodiment, 1 mL each of photocrosslinked PH@B-30, s-PPP@B-30, and b-PPP@B-30 (abbreviated as s-PPP@B, PH@B, and b-PPP@B, respectively) micelle solutions with a BBR loading of 30 wt% were prepared and placed in a dialysis bag (3.5 kD). The sample was placed in a 50 mL centrifuge tube and 3 mL of 100 mmol / L PBS buffer solution with a pH of 1.2 was added to simulate the pH environment of the stomach. After 2 hours, the dialysis bag was removed and the residual solution on the outside of the bag was wiped dry. The solution in the 50 mL centrifuge tube was replaced with 3 mL of 100 mmol / L PBS buffer solution with a pH of 6.8 to simulate the pH environment of the intestine. The above operation was repeated at the 6th hour, and the solution was replaced with 3 mL of 100 mmol / L PBS buffer solution with a pH of 7.4 to simulate the pH environment of the blood. Centrifuge tubes containing PH@B-30, s-PPP@B-30, and b-PPP@B-30 samples were placed in a shaker at 37°C and 80 rpm. 100 μL of each sample was collected and transferred to an ep tube at 1, 2, 3, 4, 6, 8, 10, 12, 24, and 36 hours. Each sample was measured in triplicate. (After each sample collection, 300 μL of the corresponding pH buffer solution was added to ensure the buffer solution volume remained constant.)

[0113] After sampling, the sample was lyophilized and reconstituted with 50 μL of methanol. After filtration through a 0.22 μm filter, the berberine hydrochloride content was determined by high-performance liquid chromatography (HPLC). The HPLC conditions were the same as in step (3) of this embodiment. The cumulative release percentage of BBR at different times was calculated using the following formula:

[0114]

[0115] Where: V—volume of 100 mmol / L PBS buffer solution, mL;

[0116] C—The concentration of BBR in the PBS buffer solution, in mg / mL;

[0117] W — the mass of the final prepared micelle solution, in mg;

[0118] LC – Drug loading capacity of drug-loaded nanoparticles, %.

[0119] Meanwhile, the organic solvent was removed by heating and evaporation after cross-linking. Finally, the pH was adjusted to 7 with a 0.2 mol / L hydrochloric acid solution, but 15 mg of SNAC was not added as a control.

[0120] Simulated in vivo release results of drug-loaded nanomicelles are shown in Figure 5 . Figure 5 This is a diagram showing the simulated in vivo release results of drug-loaded nanomicelles, by... Figure 5As shown, the PH material group exhibited better drug release at pH 1.2 than the PPP group, possibly due to the material's stronger hydrophobicity, making it less prone to rupture and drug release. The PPP material, being a triblock material, showed relatively weak stability, but still achieved approximately 40% drug release in the first 6 hours, subsequently releasing BBR gradually in a buffer solution at pH 7.4, providing evidence for the drug's hypoglycemic effect in vivo.

[0121] (5) Cytotoxicity assay of excipients and polymers (MTT)

[0122] According to literature reports, the excipients lauric acid, n-decanoic acid, and SNAC used in this invention can cause certain cell damage upon direct contact with cells, and therefore are often used orally as food additives. To guide the selection of excipient ratios in subsequent experiments, solutions of lauric acid (LA), n-decanoic acid (n-OA), and SNAC at concentrations of 5, 10, 15, 20, and 25 mg / mL, and PPP and pH solutions at concentrations of 4, 8, 12, 16, and 20 mg / mL were prepared. 50 μL of each solution was used for MTT assays of human clonal colon adenocarcinoma (Caco-2) and human colon adenocarcinoma (HT-29). Based on the differences in cell types and drug administration methods, the cells were divided into four groups: Group A was given Caco-2 cells with different concentrations of LA, n-OA, and SNAC, denoted as LA(Caco-2), n-OA(Caco-2), and SNAC(Caco-2); Group B was given HT-29 cells with different concentrations of LA, n-OA, and SNAC, denoted as LA(HT-29), n-OA(HT-29), and SNAC(HT-29); Group C was given Caco-2 cells with different concentrations of PPP and pH solutions, denoted as PPP(Caco-2) and pH(Caco-2); and Group D was given HT-29 cells with different concentrations of PPP and pH solutions, denoted as PPP(HT-29) and pH(HT-29). The results are shown in the appendix. Figure 6 .

[0123] Following the methods in steps (1) and (2) of this embodiment (without adding BBR solution during the preparation process, and the other steps being the same), empty micelle solutions of s-PPP NPs, PH NPs, and b-PPP NPs were prepared. After adjusting the pH, SNAC was added to finally prepare four groups of nanoparticles: s-PPP NPs, s-PPP NPs+SNAC, PH NPs+SNAC, and b-PPP NPs+SNAC. 50 μL of each of the above nanomicelle solutions were used for MTT assays of human clonal colon adenocarcinoma (Caco-2) and human colon adenocarcinoma (HT-29). Based on the cell type, the cells were divided into two groups: Group E received Caco-2 cells with different concentrations of the above nanomicelles, denoted as s-PPP NPs(Caco-2), s-PPP NPs+SNAC(Caco-2), PH NPs+SNAC(Caco-2), and b-PPP NPs+SNAC(Caco-2); Group F received HT-29 cells with different concentrations of the above nanomicelles, denoted as s-PPP NPs(HT-29), s-PPP NPs+SNAC(HT-29), PH NPs+SNAC(HT-29), and b-PPP NPs+SNAC(HT-29). Results are shown in the appendix. Figure 6 .

[0124] Figure 6 This diagram illustrates the cell viability of the excipients, polymer materials, and nanomicelles in Example 4 during cytotoxicity assays of Caco-2 and HT-29 cells, respectively. (A, B) represent the cell viability of the excipients in the cytotoxicity assays of Caco-2 and HT-29 cells, while (C, D) represent the cell viability of the polymer materials in the cytotoxicity assays of Caco-2 and HT-29 cells. Figure 6 (E, F) are schematic diagrams illustrating cell viability in Caco-2 and HT-29 cytotoxicity assays. Figure 6 According to AD, the polymer material has low cytotoxicity. Based on the toxicity of the excipients, the preparation process of the nanomicelles was optimized, with lauric acid concentration of 5 mg / mL and SNAC concentration of 20 mg / mL in the nanomicelles. Figure 6 As can be seen from EF, compared with the cell viability in the blank cell culture medium, the cell survival rate of the nanomicelle group was over 70%, and the cytotoxicity was relatively low.

[0125] (6) Study on the cellular uptake capacity of polymer micelles

[0126] Caco-2 cells were fed at a rate of 1×10 5Cells were seeded at a density of 100 cells / well in 12-well plates and cultured in a Thermo Forma 371 cell culture incubator for 3 days until the cells filled the wells. 100 mg of PH and PPP prepared in Examples 1 and 2 were dissolved in 3 mL of dichloromethane and reacted with 1,2-ethylenedithiol and cyanogen dye Cy5 at a molar ratio of 1:1.2:1.5 for 6 h to prepare CY5-labeled materials PH-Cy5 and PPP-Cy5.

[0127] Following steps (1) and (2) of this embodiment, PH-Cy5 NPs, PH-Cy5 NPs+SNAC, s-PPP-Cy5 NPs, s-PPP-Cy5 NPs+SNAC, b-PPP-Cy5 NPs, and b-PPP-Cy5 NPs+SNAC micelle solutions were prepared by replacing PH and PPP with PH-Cy5 and PPP-Cy5 respectively. 100 μL of each solution was used to treat an in vitro Caco-2 cell monolayer model. Simultaneously, the inhibitory effects of inhibitors such as polyethylene glycol 2000, L-arginine, M-β-cyclodextrin, and lovastatin on s-PPP-Cy5 NPs+SNAC micelles were investigated.

[0128] Cells were co-cultured with micelles and inhibitors for 3 hours, and cell uptake was analyzed by flow cytometry. The PBS group was used as a control during the analysis to investigate cell uptake. The results are as follows: Figure 7 As shown, Figure 7 The uptake of nanomicelles after treatment with different nanomicelles and inhibitors is shown. Figure 7 It is known that PLGA, due to its lack of hydrophobicity and weak transmucosal ability, exhibits extremely poor uptake capacity in nanomicelles, making it unsuitable for subsequent experiments. SNAC can effectively enhance the permeability of nanomicelles, achieving a near 100% cellular uptake rate when used in combination with ultrafine nanomicelles. The uptake of ultrafine nanomicelles is less closely related to the PAT1 pathway and cell membrane cavitation protein-mediated endocytosis, suggesting that its cellular internalization is the result of a combination of micropinocytosis and passive cellular transport.

[0129] (7) Study on transport of polymer micelles to epithelial cells

[0130] BBR, BBR+SNAC solutions, s-PPP@B, s-PPP@B+SNAC, and b-PPP@B+SNAC micelle solutions were prepared according to steps (1) and (2) of this embodiment. 100 μL of each solution was used to treat in vitro Caco-2 and HT-29 cell monolayer models to evaluate the transepithelial transport activity of the polymer micelles. Cells were sputtered at a rate of 1 × 10⁻⁶. 5Cells were seeded at a density of 9:1 (Caco-2:HT-29 = 9:1) and cultured in chambers for 21 days until the transepithelial electrical resistance (TEER) value exceeded 600 cm² and HT-29 cells secreted a mucus layer. Cell monolayers were washed three times with 200 μL of 100 mM sterile PBS solution at pH 7.4, then preheated Hank's balanced salt solution (HBSS) at pH 7.4 was added until the cell monolayer was submerged, and the cells were incubated at 37°C in a 5% CO₂ incubator for 30 minutes. In the presence of HT-29 cells, the HBSS medium at the top was replaced with an HBSS solution containing BBR, BBR+SNAC, s-PPP@B, s-PPP@B+SNAC, and b-PPP@B+SNAC nanomicelles, with a final BBR concentration of 15 mg / mL. At predetermined time intervals, 200 μL of sample was collected from the substrate side, and the same volume of fresh HBSS solution was added to the substrate side. The concentration of berberine hydrochloride in the sample was determined by liquid chromatography. The apparent permeability coefficients (P0.05) of berberine hydrochloride and insulin were compared. app Calculated using the following formula:

[0131]

[0132] in, It represents the gradient relationship between the cumulative amount of insulin or berberine hydrochloride delivered and time. A is the membrane area (cm²) secreted by Caco-2 cells. 2 C0 is the initial concentration of berberine hydrochloride in the donor chamber.

[0133] As attached Figure 8 As shown, Figure 8 For polymer micelles P app Horizontal map, by Figure 8 It is known that when culturing HT-29 cells (secreting mucus), free BBR has poor permeability in the cell monolayer, with a Papp value of approximately 3 × 10⁻⁷ cm / s. The use of SNAC can effectively improve permeability, possibly because SNAC opens tight junctions, allowing small BBR molecules to directly cross the cell monolayer. Ultrafine nanomicelles, due to their small particle size, can cross the mucus layer on their own and cross monolayers via the paracellular pathway and endocytosis. The combined use of ultrafine nanomicelles and SNAC further enhances the drug's transmembrane ability.

[0134] (8) Polymer micelles' ability to cross intestinal epithelial cells (TEER)

[0135] Take the Caco-2 and HT-29 cell monolayers from the polymer micelle epithelial cell transport study in step (7), and then culture them in apical culture medium with 200 μL of the BBR, BBR+SNAC, s-PPP@B, s-PPP@B+SNAC, and b-PPP@B+SNAC micelle solutions prepared in step (7) of this embodiment. After incubation for 2 hours, remove the cell culture and replace it with fresh HBSS solution. TEER values ​​were measured by the Millicell-resistance system (Millipore, USA) at predetermined time intervals, and the results are as follows: Figure 9 As stated above. Figure 9 The TEER value of polymer micelles; derived from Figure 9 It is known that SNAC, as a permeation enhancer, can disrupt the integrity of tight junctions in cells, providing conditions for the paracellular transport of small molecule drugs and ultrafine nanomicelles. The perturbation effect of SNAC gradually recovers after 100 minutes, preventing antigens from crossing epithelial cells and entering the body, thus preventing harm to the organism.

[0136] (9) In vivo blood glucose lowering experiment

[0137] Four-week-old C57 mice were purchased and cultured for four weeks on a high-fat diet and a 10% sucrose solution. Then, streptozotocin (STZ) solution was injected at a concentration of 55 mg / kg for one week to induce the mice to transform into type 2 diabetic mice.

[0138] Hyperglycemic model mice were randomly divided into 5 groups of 3 mice each. The mice were fasted the night before the experiment but allowed free access to water. In vivo experiments were conducted to evaluate the therapeutic effects on hyperglycemia using BBR, BBR+SNAC solutions, and drug-loaded nanomicelles of s-PPP@B, s-PPP@B+SNAC, and b-PPP@B+SNAC (BBR concentration was 15 mg / mL for all mice, and berberine was administered at a dose of 15 mg / kg). Blood samples (~3 μL) were collected via the tail vein, and hourly blood glucose levels were continuously monitored and recorded using a glucometer.

[0139] The blood glucose change curve is attached. Figure 10 As shown, Figure 10 This is a graph showing the in vivo blood glucose level of drug-loaded nanomicelles. Figure 10 It can be seen that, except for the direct oral administration of BBR, all groups showed a certain hypoglycemic effect. The nanomicelles could maintain normal blood glucose for about 6 hours. Among them, s-PPP@B+SNAC had the best hypoglycemic effect. Moreover, after mice had a short period of food intake, the blood glucose level rose briefly and then dropped rapidly to normal levels, which may be related to the dual promoting effect of SNAC and ultrafine nanomicelles. The ultrafine nanomicelles also showed a long-lasting hypoglycemic effect, which may be because ultraviolet light crosslinking enhanced the stability of the nanomicelles, enabling long-lasting hypoglycemic effects in vivo.

Claims

1. A uniform, sub-micron polymeric nanomicelle, characterized in that, The polymer nanomicelles comprise unsaturated bond-containing polymers formed by modifying a polyglycolide-based material with a double bond-containing ligand, and the polymers self-emulsify to encapsulate berberine, are crosslinked by UV light, and are formed after pH adjustment; the preparation method of the uniform superfine polymer nanomicelles comprises the following steps: (1) Preparation of unsaturated bond-containing polymers: In anhydrous and oxygen-free and nitrogen-protected environment, a carboxyl-terminated polyglycolide-based material is dissolved in an organic solvent, a catalyst is added, and the carboxyl group is activated by stirring at room temperature, a hydroxyl-containing double bond ligand is added, and reaction is carried out at room temperature, dialysis is performed, and the lower layer precipitate is obtained by precipitation and centrifugation with ice ether, washed, and dried to obtain the unsaturated bond-containing polymer; Alternatively, in a nitrogen-protected environment, a hydroxyl-terminated polyglycolide-based material is dissolved in an organic solvent, a double bond ligand containing acid halide is slowly added dropwise in an ice bath environment, triethylamine is added as an acid binding agent, and reaction is carried out at room temperature, dialysis is performed, and the lower layer precipitate is obtained by precipitation and centrifugation with ice ether, washed, and dried to obtain the unsaturated bond-containing polymer; (2) Preparation of polymer nanomicelles: The unsaturated bond-containing polymer solution is obtained by dissolving the unsaturated bond-containing polymer in an organic solvent, lauric acid and a drop of Span 80 are mixed uniformly, and then berberine methanol solution is added to obtain an organic phase, and the uniform microcapsules are prepared by using the microfluidic technology with the organic phase as the inner phase and water as the outer phase, and the microcapsules are collected in NaOH solution, and the microcapsules are spontaneously emulsified and dispersed to form a polymer nanomicelle solution; (3) Preparation of uniform superfine polymer nanomicelles: The crosslinking agent is added to the polymer nanomicelle solution, and the unsaturated bond is crosslinked by UV light to obtain a uniform superfine polymer nanomicelle solution; In the step (1), the carboxyl-terminated PLGA-based material is PLGA-COOH or PEG-PLGA-COOH; the hydroxyl-containing double bond ligand is hydroxyethyl acrylate; the hydroxyl-terminated PLGA-based material is OH-PLGA-PEG-PLGA-OH; and the acyl halide-containing double bond ligand is methacryloyl chloride. In the step (2), the concentration of the unsaturated bond-containing polymer solution is 20-30 mg / mL, the concentration of the NaOH solution is 0.1-0.25 mol / L, the molar ratio of lauric acid to OH - is 1:20-50, the concentration of the berberine methanol solution is 20-50 mg / mL, and the volume ratio of the unsaturated bond-containing polymer solution to lauric acid is 1:0.5-2.

2. The uniform, sub-micron polymeric micelles according to claim 1, wherein, The double bond-containing ligand is a hydroxyl-containing double bond ligand or a double bond ligand containing acid halide, the hydroxyl-containing double bond ligand is hydroxyethyl acrylate, and the double bond ligand containing acid halide is methacryloyl chloride.

3. The uniform, sub-micron polymeric micelles of claim 1, wherein, The polyglycolide-based material is PLGA, PLGA-PEG or PLGA-PEG-PLGA.

4. The uniform, sub-micron polymeric micelles of claim 1, wherein, The particle size of the uniform superfine polymer nanomicelles is 10 nm or less, the molecular weight of PLGA in PLGA-PEG or PLGA-PEG-PLGA is much larger than that of PEG, and the mass ratio of LA to GA in PLGA is 25:25-75.

5. The method of producing uniform, sub-micron polymeric nanomicelles according to any one of claims 1 to 4, characterized in that, The method comprises the following steps: (1) Preparation of unsaturated bond-containing polymers: In anhydrous and oxygen-free and nitrogen-protected environment, a carboxyl-terminated polyglycolide-based material is dissolved in an organic solvent, a catalyst is added, and the carboxyl group is activated by stirring at room temperature, a hydroxyl-containing double bond ligand is added, and reaction is carried out at room temperature, dialysis is performed, and the lower layer precipitate is obtained by precipitation and centrifugation with ice ether, washed, and dried to obtain the unsaturated bond-containing polymer; Alternatively, in a nitrogen-protected environment, a hydroxyl-terminated polyglycolide-based material is dissolved in an organic solvent, a double bond ligand containing acid halide is slowly added dropwise in an ice bath environment, triethylamine is added as an acid binding agent, and reaction is carried out at room temperature, dialysis is performed, and the lower layer precipitate is obtained by precipitation and centrifugation with ice ether, washed, and dried to obtain the unsaturated bond-containing polymer; (2) Preparation of polymer nanomicelles: The polymer containing unsaturated bond is dissolved in an organic solvent to obtain a polymer solution containing unsaturated bond, lauric acid and one drop of Span 80 are mixed uniformly, and then the berberine methanol solution is added to obtain an organic phase, the uniform microcapsules of oil-in-water are prepared by using the microfluidic technology with the organic phase as the inner phase and water as the outer phase, the microcapsules are collected in the NaOH solution, the microcapsules are spontaneously emulsified and dispersed to form a polymer nanomicelle solution; (3) Preparation of uniform ultrafine polymer nanomicelles: The crosslinking agent is added to the polymer nanomicelle solution, and the unsaturated bond is ultraviolet crosslinked under the ultraviolet lamp to obtain a uniform ultrafine polymer nanomicelle solution; In the step (1), the carboxyl-terminated PLGA-based material is PLGA-COOH or PEG-PLGA-COOH; the hydroxyl-containing double bond ligand is hydroxyethyl acrylate; the hydroxyl-terminated PLGA-based material is OH-PLGA-PEG-PLGA-OH; and the acyl halide-containing double bond ligand is methacryloyl chloride. In the step (2), the concentration of the unsaturated bond-containing polymer solution is 20-30 mg / mL, the concentration of the NaOH solution is 0.1-0.25 mol / L, the molar ratio of lauric acid to OH - is 1:20-50, the concentration of the berberine methanol solution is 20-50 mg / mL, and the volume ratio of the unsaturated bond-containing polymer solution to lauric acid is 1:0.5-2.

6. The process for the preparation of uniform, ultra-fine polymeric nanomicelles according to claim 5, characterized in that, In step (1), the organic solvent is DMF, DCM or tetrahydrofuran; the catalyst is DMAP and DCC, or EDC and NHS; the molar ratio of the carboxyl-terminated PLGA-based material, the catalyst and the hydroxyl-containing double bond ligand is 1:1.5:2-5; the molar ratio of the hydroxyl-terminated PLGA-based material, the acyl halide-containing double bond ligand and triethylamine is 1:20-30:30-40.

7. The method of claim 5, wherein the polymer nanomicelles are homogeneous. In step (2), the device used in the microfluidic technology is an oil-in-water two-phase microfluidic device, and the microfluidic device parameters are as follows: the inner phase pore diameter is 200-400 μm, the outer phase inner diameter is 600-800 μm, and the inner phase and outer phase flow rate ratio is 5-10:1; the organic solvent is DCM or chloroform.

8. The method for preparing uniform, ultrafine polymer nanomicelles according to claim 5, characterized in that, In step (3), the crosslinking agent is I2959, and the power of the UV lamp is 13000 uw / cm 2 The crosslinking time is 10-20 min, and the pH is adjusted to neutral after evaporation below 40℃.

9. Use of the uniform, sub-micron polymeric nanomicelles according to any one of claims 1 to 4 for the manufacture of a medicament for the treatment of type 2 diabetes, characterized in that, The berberine hydrochloride is added to the polymer solution containing unsaturated bond to form a mixed solution, a drug-loaded polymer micelle solution is prepared through a self-emulsification process, ultraviolet crosslinking is performed, the pH is adjusted to neutral, and the permeation enhancer 8-(2-hydroxybenzamide) sodium octanoate is mixed and used.

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