Antioxidant nanomaterial based on self-assembly of traditional chinese medicine and copper ions and preparation method thereof

Abstract

The application belongs to the field of nanomedicine and traditional Chinese medicine, and specifically discloses an antioxidant nano material based on self-assembly of traditional Chinese medicine and copper ions and a preparation method thereof. The nano material is BPCu nanoparticles formed by self-assembly of berberine and procyanidin B2 under the induction of copper ions, and is further surface-modified by polyvinylpyrrolidone to obtain stable and dispersed PVP@BPCu nano structures. The material has small particle size, uniform distribution, good water solubility and storage stability, has excellent free radical scavenging capacity and peroxidase-like activity, is suitable for auxiliary treatment of oxidative stress related diseases, and the preparation method is simple to operate and does not need organic solvents, and has good industrialization and disease treatment potential.

Description

TECHNICAL FIELD

[0001] The application belongs to the field of nanomedicine and traditional Chinese medicine, and particularly relates to an antioxidant nanomaterial based on self-assembly of traditional Chinese medicine and copper ions and a preparation method thereof. BACKGROUND

[0002] Oxidative stress refers to a pathological state caused by imbalance between the production and clearance of reactive oxygen species (ROS) in the body, which has been proven to play a key role in the development of various diseases, especially in cardiovascular and cerebrovascular diseases, neurodegenerative diseases, tumors and aging-related diseases. Excessive accumulation of free radicals can cause cell damage, inflammatory response and dysfunction. Therefore, developing efficient and biocompatible antioxidants to scavenge excess free radicals in the body has become an important direction in current drug development and material science research.

[0003] Natural products have been widely used in antioxidant research in recent years due to their safe biological sources, structural diversity and good pharmacological activities. Among them, berberine (BBR), as a representative alkaloid in traditional Chinese medicine Coptis, has multiple biological activities such as antioxidant, anti-inflammatory and hypolipidemic activities. However, berberine has poor stability in body fluids, low water solubility and limited bioavailability, which significantly limits its clinical application potential.

[0004] To overcome the limitations of direct application of natural products, self-assembly nanotechnology has shown great advantages in the functional enhancement and delivery of natural products. In particular, the coordination between metal ions and natural small molecules (such as polyphenols, alkaloids, etc.) can construct stable and controllable nanostructures through non-covalent interactions (such as coordination bonds, π-π stacking, hydrogen bonds). This kind of metal-induced self-assembly technology can enhance the stability of natural products to some extent. However, the existing self-assembly nanomaterials still have problems of poor stability, low utilization rate and poor water solubility. SUMMARY

[0005] To solve the above technical problems, the application provides an antioxidant nanomaterial based on self-assembly of traditional Chinese medicine and copper ions and a preparation method thereof. The antioxidant material of the application takes berberine and procyanidin B2 as the core active ingredients of traditional Chinese medicine, cooperates with copper ions to form a nanostructure by self-assembly, and is surface-modified by PVP, which significantly improves its water solubility and stability, thereby overcoming the limitations of poor solubility and low stability of traditional natural antioxidants, and constructing a functional nanomaterial with high efficiency, stability and excellent free radical scavenging capacity.

[0006] The technical scheme protected by the application is: an antioxidant nanomaterial based on self-assembly of traditional Chinese medicine and copper ions, wherein the nanomaterial is formed by self-assembly of berberine, procyanidin B2 and divalent copper ions in an aqueous phase to form BPCu nanoparticles, and the nanoparticles are surface-modified by polyvinylpyrrolidone (PVP) to form PVP@BPCu nanoparticles.

[0007] Further, the average particle size of the PVP@BPCu nanoparticles is 111.37±3.71 nm, the polymer dispersity index (PDI) is 0.207±0.021, and the Zeta potential is -23.47±0.96 mV.

[0008] Further, the berberine drug loading of the PVP@BPCu nanoparticles is 30%-36%, and the copper element content is 8%-9%.

[0009] Further, the berberine drug loading of the PVP@BPCu nanoparticles is 34.20%, and the copper element content is 8.87%.

[0010] Another scheme to be protected by the application is a preparation method of an antioxidant nanomaterial based on self-assembly of traditional Chinese medicine and copper ions, characterized by the following steps:

[0011] Step S1: prepare aqueous solutions of berberine, procyanidin B2, CuCl2·2H2O, NaOH and polyvinylpyrrolidone respectively for standby use;

[0012] Step S2: under the condition of an aqueous phase, sequentially add the berberine solution, the procyanidin B2 solution, the Cu 2+ solution into a reaction system, and stir at room temperature for 25-35 minutes to uniformly mix them;

[0013] Step S3: then add the NaOH solution to adjust the pH to 7-8, and continue to stir for 3 hours to promote complexation and self-assembly to form BPCu nanoparticles;

[0014] Step S4: then add the PVP solution, and ultrasonically treat for 10-15 minutes to realize surface modification of the nanoparticles to form PVP@BPCu nanomaterial;

[0015] Step S5: purify the obtained material by an ultrafiltration centrifugation method, resuspend and store after washing to remove impurities.

[0016] Further, the molecular weight of the polyvinylpyrrolidone used in step S1 is 45,000-58,000.

[0017] Furthermore, in step S1, the concentration of NaOH solution is 5 mg / mL, the concentration of CuCl2·2H2O solution is 20 mg / mL, the concentration of proanthocyanidin B2 aqueous solution is 5 mg / mL, the concentration of berberine solution is 1 mg / mL, and the concentration of polyvinylpyrrolidone aqueous solution is 40 mg / mL.

[0018] Furthermore, in step S2, the mass ratio of berberine, proanthocyanidin B2, and CuCl2·2H2O is 1:0.9-1.1:0.5-0.75.

[0019] Furthermore, in step S2, the mass ratio of berberine, proanthocyanidin B2, and CuCl2·2H2O is 1:1:0.67.

[0020] Furthermore, in step S5, the nanoparticles are purified using a 100kDa ultrafiltration centrifuge tube, centrifuged at 3000 rpm for 10 minutes, and washed three times.

[0021] Compared with the prior art, the present invention has the following advantages:

[0022] 1. This invention utilizes copper ion-induced self-assembly to form a stable nanoparticle structure from berberine and proanthocyanidin B2. By employing a ternary synergistic strategy of "traditional Chinese medicine active molecules + metal ions + surface-modified polymers", the solubility and bioavailability of natural products can be significantly improved.

[0023] 2. The present invention uses PVP surface modification to effectively improve the dispersibility and storage stability of nanomaterials, avoid nanoparticle aggregation and precipitation. After continuous storage in aqueous solution, PBS buffer, DMEM medium and RPMI-1640 medium for 7 days, the particle size and distribution remain stable, which shows good biological stability.

[0024] 3. The antioxidant nanomaterials prepared in this invention exhibit highly efficient scavenging ability against various free radicals, including DPPH• and ABTS•. + Hydroxyl radical (•OH) and superoxide anion (•O2) - Meanwhile, the antioxidant nanomaterials of this invention possess catalase-mimicking activity, which can effectively decompose hydrogen peroxide into water and oxygen, enhancing the ability to decompose hydrogen peroxide and further improving the antioxidant effect.

[0025] 4. No organic solvents are used in the entire preparation process of this invention. The preparation method is simple and mild. This material has potential antioxidant application prospects, especially in the treatment of diseases caused by oxidative stress. Attached Figure Description

[0026] The present invention will now be described in further detail with reference to the accompanying drawings.

[0027] Figure 1 The particle size distribution diagrams (measured by dynamic light scattering DLS) of the antioxidant nanomaterials BPCu and PVP@BPCu of this invention reflect the average particle size and uniformity of the distribution.

[0028] Figure 2 The zeta potential measurements of the nanomaterials BPCu and PVP@BPCu show that PVP has been successfully modified on the surface of BPCu nanoparticles.

[0029] Figure 3 The particle size distribution of the nanomaterial PVP@BPCu in different solutions shows that it has good dispersibility.

[0030] Figure 4 The particle size stability of the nanomaterial PVP@BPCu in different solutions indicates that it has good stability.

[0031] Figure 5 The nanomaterials are PVP@BPCu, berberine, proanthocyanidin B2, and Cu. 2+ The ultraviolet-visible absorption spectrum.

[0032] Figure 6 The UV absorption spectra of the nanomaterial PVP@BPCu at different concentrations are shown.

[0033] Figure 7 The nanomaterial PVP@BPCu and its components (BBR, PAC B2 and Cu) are mentioned. 2+ In terms of aqueous solution appearance, BBR solution is bright yellow, anthocyanin B2 (PAC B2) solution is nearly colorless, Cu²⁺ solution is light blue, and PVP@BPCu is a dark brown transparent solution.

[0034] Figure 8 The figure shows the experimental results of the DPPH free radical scavenging ability of the nanomaterial PVP@BPCu.

[0035] Figure 9 The figure shows the experimental results of the ABTS free radical scavenging ability of the nanomaterial PVP@BPCu.

[0036] Figure 10 The figure shows the experimental results of the •OH free radical scavenging ability of the nanomaterial PVP@BPCu.

[0037] Figure 11 To remove •O2 from the nanomaterial PVP@BPCu - Figure showing the experimental results of free radical scavenging ability.

[0038] Figure 12The figure shows the experimental results of the catalase activity of the nanomaterial PVP@BPCu, demonstrating its decomposition effect on H2O2. Detailed Implementation

[0039] To make the objectives, features, and advantages of the present invention readily apparent, the specific embodiments of the present invention will be described in detail below with reference to the accompanying drawings.

[0040] This invention relates to an antioxidant nanomaterial based on the self-assembly of traditional Chinese medicine and copper ions. Berberine, proanthocyanidin B2, and divalent copper ions self-assemble in an aqueous phase to form BPCu nanoparticles, which are then surface-modified with polyvinylpyrrolidone (PVP) to form PVP@BPCu nanoparticles. The average particle size of the PVP@BPCu nanoparticles is 111.37±3.71 nm, the polymer dispersibility index (PDI) is 0.207±0.021, and the zeta potential is -23.47±0.96 mV. The berberine loading of the PVP@BPCu nanoparticles is 34.20%, and the copper content is 8.87%.

[0041] This invention is significantly innovative in terms of component combination and structural design. The nanoparticles of this invention employ a ternary synergistic strategy of "active molecules of traditional Chinese medicine + metal ions + surface-modified polymers," resulting in traditional Chinese medicine-based organometallic nanomaterials with excellent antioxidant activity and stability, possessing significant theoretical and practical application value.

[0042] In existing technologies, common organometallic nanoparticles are typically assembled from metal ions and functional monomers (such as polyphenolic compounds, organic molecules containing carboxyl or amino groups) through coordination interactions. Most of these functional monomers have simple structures and single functions; their main role is to provide coordination sites to bind with metal ions, thus constructing basic coordination networks or nanostructure frameworks. In contrast, berberine, used in this invention, is a natural isoquinoline alkaloid with unique structural characteristics and functions. In self-assembly systems, it not only acts as a structural regulation unit but also simultaneously endows the material with functional activity, possessing a dual role of structural regulation and functional activity. Its advantages are reflected in the following two aspects:

[0043] In terms of structural regulation: Berberine molecules possess a rigid large π-conjugated aromatic structure, and also contain quaternary ammonium positive charge centers (N... + This structural feature endows berberine with excellent π-π stacking ability, enabling it to undergo intermolecular stacking interactions with other aromatic molecules (proanthocyanidins B2) in the system, promoting orderly aggregation. On the other hand, its cationic structure can participate in electrostatic interactions with anions or negatively charged groups in the system, thereby providing spatial stability and structural directionality during the overall self-assembly process, unlike conventional functional monomers which only provide coordination sites.

[0044] In terms of functional activity, berberine not only participates in the formation of self-assembled structures, but also possesses clearly defined natural pharmacological activities, including but not limited to antioxidant and anti-inflammatory biological effects. Therefore, introducing berberine into metal-organic self-assembly systems can endow nanomaterials with endogenous drug functionality while forming them, achieving integrated "structure + function" design. This significantly expands the applications of such nanoparticles, especially suitable for therapeutic, synergistic delivery, or targeted functional development in the biomedical field.

[0045] Selected copper ions (Cu) 2+ As a common transition metal ion, copper not only forms stable coordination networks with various natural small molecules, but also possesses certain catalase-like mimicry activity. Procyanidin B2 (PACB2), a widely sourced polyphenol, exhibits excellent antioxidant properties and can form coordination complexes with copper ions, enhancing its structural stability and free radical scavenging ability.

[0046] To further improve the stability and water solubility of nanomaterials, this invention uses polyvinylpyrrolidone (PVP) for surface modification, which has good biocompatibility and stability. Modification on the surface of nanoparticles can significantly improve their dispersibility and storage stability, solving the problems of poor stability, low utilization rate and poor water solubility of natural antioxidants.

[0047] The preparation method of antioxidant nanomaterials based on the self-assembly of traditional Chinese medicine and copper ions of the present invention is carried out according to the following steps:

[0048] Step S1: Prepare aqueous solutions of berberine, proanthocyanidin B2, CuCl2·2H2O and polyvinylpyrrolidone for later use;

[0049] Step S2: Under aqueous phase conditions, berberine solution, proanthocyanidin B2 solution, and Cu... 2+ The solution was added to the reaction system in sequence. The mass ratio of berberine, proanthocyanidin B2 and CuCl2·2H2O was selected as 1:1:0.67. The mixture was stirred at room temperature for 25-35 minutes to ensure uniform mixing.

[0050] Step S3: Then add NaOH solution to adjust the pH to 7-8, and continue stirring for 3 hours to promote complexation and self-assembly to form BPCu nanoparticles;

[0051] Step S4: Next, add PVP solution and sonicate for 10-15 minutes to achieve surface modification of nanoparticles and form PVP@BPCu nanomaterials.

[0052] Step S5: Purify the obtained material using ultrafiltration centrifugation, wash to remove impurities, and then resuspend and store it.

[0053] No organic solvents were used in the entire preparation process of this invention, and the preparation method is simple and mild. The prepared nanoparticle materials have potential antioxidant applications, especially showing good application prospects in the treatment of diseases caused by oxidative stress.

[0054] The antioxidant nanomaterials and their preparation methods of the present invention have been described in detail above. The following section describes the preparation of PVP@BPCu antioxidant nanomaterials through specific experiments, and the prepared antioxidant materials are then characterized and their performance evaluated.

[0055] Step S1, Preparation of raw material solutions: Berberine (purity ≥99%) and copper chloride dihydrate (purity ≥99.99% metalsbasis) were purchased from Shanghai Aladdin Biochemical Technology Co., Ltd., proanthocyanidin B2 (purity ≥99%) and sodium hydroxide (purity 97%) were purchased from Shanghai Maclean Biochemical Technology Co., Ltd., and polyvinylpyrrolidone K-30 (molecular weight 45000-58000) was purchased from Beijing Coolplay Technology Co., Ltd. All chemical reagents were used in their received condition without further purification.

[0056] Weigh 13.2 mg of berberine (BBR), add 13.2 mL of deionized water, and dissolve it under water bath heating and ultrasonic assistance to prepare a 1 mg / mL BBR aqueous solution; weigh 11 mg of procyanidin B2 (PACB2), dissolve it in 2.2 mL of deionized water to prepare a 5 mg / mL PACB2 aqueous solution; weigh 49 mg of copper chloride dihydrate (CuCl2·2H2O), dissolve it in 2.45 mL of water to prepare a 20 mg / mL CuCl2·2H2O solution; separately weigh 24.3 mg of sodium hydroxide (NaOH), dissolve it in 4.86 mL of water to prepare a 5 mg / mL NaOH solution.

[0057] Step S2, Mixing of raw material solutions: Add 10 mL of deionized water, 5 mL of 1 mg / mL BBR aqueous solution, 1 mL of 5 mg / mL PACB2 aqueous solution, and 167.5 μL of 20 mg / mL CuCl2·2H2O solution sequentially to a 40 mL sample vial. Stir magnetically at 300 rpm for 30 min at room temperature in a dark environment to ensure thorough mixing of all components.

[0058] Step S3: Adjusting the pH of the reaction system: Add 500 μL of 5 mg / mL NaOH solution to the above reaction system to adjust the pH to 7–8. Continue stirring at 300 rpm for 3 hours at room temperature and in the dark to promote the complexation and self-assembly of metal ions and ligands, forming BPCu nanoparticles.

[0059] Step S4, Surface PVP Modification: Weigh 106.4 mg of polyvinylpyrrolidone (PVPK30, average molecular weight 45,000–58,000), dissolve it in 2.66 mL of deionized water to prepare a 40 mg / mL PVP solution. Under ultrasonic conditions, add 500 μL of the 40 mg / mL PVP aqueous solution to the above system; continue ultrasonic treatment at room temperature for 10 minutes to promote the adsorption of PVP on the surface of the nanoparticles to form a stable coating layer, thereby obtaining PVP@BPCu nanoparticles.

[0060] Step S5, Purification of Nanoparticles: The obtained nanoparticles were purified using 100 kDa ultrafiltration centrifuge tubes (brand: Millipore, model: UFC910096, capacity: 15 mL). The nanomaterials were recovered by centrifugation at 3000 rpm for 10 min, followed by washing three times with deionized water (3000 rpm, 10 min each time) to remove free reactants and unreacted components.

[0061] The resulting PVP@BPCu nanoparticles were dispersed in deionized water, filtered through a 0.45 μm filter membrane to remove large particulate impurities, and then stored at 4 °C in the dark for later use.

[0062] The following series of characterization and testing were conducted to assess the physicochemical properties and antioxidant performance of the PVP@BPCu antioxidant nanomaterials prepared by the above method.

[0063] 1. Particle size, PDI and Zeta potential testing

[0064] The average particle size, particle size distribution index (PDI), and zeta potential of BPCu and PVP@BPCu nanoparticles were measured using dynamic light scattering (DLS) technology. Figure 1 and Figure 2 The results showed that the average particle size of BPCu nanoparticles was 70.86±1.08 nm, the PDI was 0.228±0.009, and the Zeta potential was -16.57±0.57 mV; the average particle size of PVP@BPCu nanoparticles was 111.37±3.71 nm, the PDI was 0.207±0.021, and the Zeta potential was -23.47±0.96 mV. These results indicate that PVP modification increased the nanoparticle size and made the potential more negative, demonstrating that PVP was successfully modified onto the BPCu surface, which helps to further improve stability.

[0065] 2. Stability assessment

[0066] Based on the confirmed good dispersibility of PVP@BPCu in aqueous solution, its stability in different physiologically relevant media was further investigated. PVP@BPCu nanoparticles were dispersed in water, PBS, DMEM, and RPMI-1640 media, respectively, and their particle size distribution was analyzed. The results are as follows: Figure 3 As shown, it maintains good dispersibility and homogeneity in various solutions. Furthermore, to assess its long-term stability, particle size changes were continuously monitored for 7 days in the aforementioned solutions. Figure 4 The results showed that the particle size of PVP@BPCu remained relatively stable in aqueous solution, PBS, DMEM and RPMI-1640 medium, indicating that the nanosystem has good storage stability in different biological media and meets the basic requirements for subsequent biological applications.

[0067] 3. Ultraviolet-Vis absorption spectroscopy (UV-Vis) analysis

[0068] UV-Vis spectrophotometer for PVP@BPCu and its components (BBR, PACB2, Cu) 2+ The characteristic absorption peaks of ( ) are analyzed. For example Figure 5 and Figure 6 As shown, PVP@BPCu exhibits a significant absorption peak in the 250-300 nm range, and the peak intensity increases with increasing concentration, indicating that the functional components have been successfully assembled into the nanosystem. Figure 7 The macroscopic appearance of each component and the final assembled product in aqueous solution is shown: BBR solution is bright yellow, PACB2 solution is nearly colorless, and Cu... 2+ The solution was light blue, while the aqueous solution of PVP@BPCu nanoparticles was dark brown and transparent, clearly different from the precursor components, further confirming the successful realization of the self-assembly process and metal coordination reaction.

[0069] 4. DPPH • Cleanup Test

[0070] DPPH is a commonly used organic free radical, widely used to evaluate antioxidant activity. DPPH free radicals possess unpaired electrons and exhibit a stable purple color in ethanol. Antioxidants can capture these unpaired electrons, scavenging the free radical and causing the color to weaken or even disappear.

[0071] In the experiment, DPPH was dissolved in ethanol to prepare a working solution of 30 μg / mL. 100 μL of different concentrations of PVP@BPCu nanoparticle solutions (0, 12.5, 25, 50, 100, 150, and 200 μg / mL) were added to 1 mL of the DPPH working solution, mixed, and incubated at 37 °C in the dark for 30 min. The absorbance of the reaction system at 519 nm was then measured using a UV-Vis spectrophotometer to evaluate the scavenging ability of PVP@BPCu on DPPH free radicals.

[0072] like Figure 8 As shown, the addition of PVP@BPCu significantly reduced the intensity of the absorption peak at 519 nm and noticeably weakened the purple color of the reaction system, indicating its effective scavenging of DPPH free radicals. This scavenging effect was concentration-dependent; when the concentration of PVP@BPCu increased from 12.5 μg / mL to 200 μg / mL, the DPPH scavenging rate increased from 4.79% to 72.67%, demonstrating that PVP@BPCu possesses excellent free radical scavenging capabilities.

[0073] 5. ABTS• + Cleanup test

[0074] The effect of PVP@BPCu nanoparticles on ABTS• was evaluated using the Total Antioxidant Assay Kit (ABTS method). + Free radical scavenging ability. This method uses 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonicacid) (ABTS) as a substrate, and generates green ABTS• under the action of a suitable oxidant. + Free radicals, which have a characteristic absorption peak at 734 nm. The presence of antioxidants can effectively inhibit ABTS• + The formation of [a substance] leads to a decrease in absorbance. By measuring the change in absorbance at 734 nm, the total antioxidant capacity of the sample can be indirectly reflected.

[0075] Trolox (a vitamin E analog) was used as the standard reference antioxidant in the experiment, and the antioxidant level of the sample was expressed as Trolox-Equivalent Antioxidant Capacity (TEAC). For example, when the free radical scavenging rate of a sample is consistent with the scavenging rate corresponding to 0.6 mM Trolox, the TEAC value of the sample is considered to be 0.6 mM.

[0076] The experimental procedure is as follows: Prepare the ABTS working solution according to the kit instructions. Mix different concentrations of PVP@BPCu nanoparticle solutions with the ABTS working solution, react at room temperature for a certain time, and then measure the absorbance at 734 nm using a microplate reader. The results are as follows. Figure 9 As shown, with the increase of PVP@BPCu concentration, the absorbance at 734 nm gradually decreased, while the TEAC value increased significantly, indicating that PVP@BPCu nanoparticles have good free radical scavenging ability and strong total antioxidant activity.

[0077] 6. Hydroxyl radical (•OH) scavenging test

[0078] •OH can be converted from Fe by the Fenton reaction. 2+ It reacts with H2O2 to produce, and the reaction equation is as follows: Fe 2+ +H₂O₂→Fe 3+ +OH - •OH. To evaluate the scavenging ability of PVP@BPCu nanoparticles for •OH, 3,3′,5,5′-tetramethylbenzidine (TMB) was used as a colorimetric probe. •OH generated in the Fenton reaction oxidizes colorless TMB to form a blue oxidized product (oxTMB), which has a characteristic absorption peak at 650 nm. In the experiment, a reaction system containing FeSO4 (1 mM), H2O2 (5 mM), and TMB (1 mM) was prepared in PBS buffer at pH 7.4, and different concentrations of PVP@BPCu nanoparticles (25, 50, 100, 150, 200, and 250 μg / mL) were added. After mixing and reaction, the absorbance at 650 nm was measured to evaluate the •OH scavenging efficiency.

[0079] like Figure 10 As shown, the addition of PVP@BPCu significantly suppressed the absorption peak intensity of oxTMB at 650 nm, and the blue color of the solution was significantly reduced, indicating its good scavenging ability for •OH. Further analysis revealed that this scavenging effect was concentration-dependent: when the concentration of PVP@BPCu increased from 25 μg / mL to 250 μg / mL, the •OH scavenging rate significantly increased from 6.02% to 77.68%, and the blue color almost disappeared.

[0080] 7. •O2 - Cleanup test

[0081] The classic nitroblue tetrazolium (NBT) colorimetric method was used, and the total SOD activity assay kit (NBT method) was employed to evaluate the effect of PVP@BPCu nanoparticles on superoxide anion radicals (•O2). -The system exhibits the ability to scavenge xanthine. Using xanthine as a substrate and xanthine oxidase (XO) as a catalyst, the reaction continuously generates •O2. - This free radical can reduce NBT to blue, insoluble formazan, which has a characteristic absorption peak at 560 nm. Superoxide dismutase (SOD) can catalyze the reduction of O2. - The O2 content of the sample is disproportionated into H2O2 and O2, thereby inhibiting the formation of formazan. Therefore, the change in absorbance at 560 nm can indirectly reflect the O2 content of the sample. - Cleaning efficiency.

[0082] In the specific experiment, different concentrations of PVP@BPCu nanoparticles (25, 50, 100, 150, 200, and 250 μg / mL) were added to the NBT / enzyme working solution and incubated at 37°C in the dark for 30 min. After the reaction, the absorbance at 560 nm was measured using a microplate reader, and •O2 was calculated accordingly. - The inhibition rate was used to characterize its SOD-like antioxidant activity.

[0083] Experimental results are as follows Figure 11 As shown, PVP@BPCu nanoparticles affect •O2 - The scavenging efficiency of •O2 showed a concentration-dependent effect, increasing with concentration from 25 μg / mL to 250 μg / mL. - The inhibition rate increased from 12.36% to 62.46%, indicating that the material has a significant superoxide anion scavenging ability.

[0084] 8. Measurement of catalase activity (H2O2 scavenging)

[0085] The catalase-like catalytic activity of PVP@BPCu nanoparticles was evaluated using a commercially available catalase assay kit. Under sufficient hydrogen peroxide concentration, the catalytically active material mimics the natural CAT activity, catalyzing the decomposition of H₂O₂ to produce water and oxygen. Residual H₂O₂ in the reaction system undergoes an oxidation reaction with a chromogenic substrate under the catalysis of peroxidase, generating a red oxidation product, N-(4-antipyryl)-3-chloro-5-sulfonate-p-benzoquinonemonoimine, which exhibits a characteristic absorption peak at 520 nm. The CAT-like catalytic ability of the sample can be indirectly assessed by detecting the absorbance change at this wavelength.

[0086] In the experiment, different concentrations of PVP@BPCu nanoparticles (50, 75, 100, 150, 200 and 250 μg / mL) were added to H2O2 solution for reaction. After the reaction was terminated, the diluted reaction solution was mixed with the colorimetric working solution and incubated at 25°C for 15 minutes. Then, the absorbance at 520 nm was measured using an ELISA reader.

[0087] like Figure 12 As shown, the concentration of PVP@BPCu nanoparticles is positively correlated with their CAT-like catalytic activity, exhibiting excellent hydrogen peroxide scavenging ability. Their CAT activity is mainly attributed to the Cu content in the material. + Its coordination structure can effectively catalyze the decomposition of H2O2 into O2 and H2O, demonstrating its potential application value in ROS removal.

[0088] 9. Analysis of drug loading and copper content

[0089] The berberine (BBR) loading in PVP@BPCu nanoparticles was determined by UV-Vis spectrophotometry, and the result was 34.20% ± 0.016. The copper content was analyzed by inductively coupled plasma optical emission spectrometry (ICP-OES), and the copper content in the PVP@BPCu nanoparticles was found to be 8.87 wt%.

[0090] These characterization results fully demonstrate that the PVP@BPCu nanomaterials prepared in this invention have stable structure, controllable size, good antioxidant capacity and enzyme-mimicking activity, and are suitable for subsequent biomedical research.

[0091] The embodiments of the present invention have been described in detail above with reference to the accompanying drawings. However, the present invention is not limited to the above embodiments. Within the scope of knowledge possessed by those skilled in the art, various changes can be made without departing from the spirit of the present invention.

Claims

1. Antioxidant nanomaterials based on the self-assembly of traditional Chinese medicine and copper ions, characterized in that: The nanomaterials are formed by the self-assembly of berberine, proanthocyanidin B2 and divalent copper ions in an aqueous phase to form BPCu nanoparticles, and then surface-modified by polyvinylpyrrolidone (PVP) to form PVP@BPCu nanoparticles.

2. The antioxidant nanomaterial based on the self-assembly of traditional Chinese medicine and copper ions according to claim 1, characterized in that: The average particle size of the PVP@BPCu nanoparticles is 111.37±3.71 nm, the polymer dispersibility index (PDI) is 0.207±0.021, and the zeta potential is -23.47±0.96 mV.

3. The antioxidant nanomaterial based on the self-assembly of traditional Chinese medicine and copper ions according to claim 1, characterized in that: The PVP@BPCu nanoparticles have a berberine loading of 30%-36% and a copper content of 8%-9%.

4. The antioxidant nanomaterial based on the self-assembly of traditional Chinese medicine and copper ions according to claim 3, characterized in that: The PVP@BPCu nanoparticles have a berberine loading of 34.20% and a copper content of 8.87%.

5. The method for preparing the antioxidant nanomaterials based on the self-assembly of traditional Chinese medicine and copper ions as described in claim 1, characterized in that: Please follow these steps: Step S1: Prepare aqueous solutions of berberine, proanthocyanidin B2, CuCl2·2H2O, NaOH and polyvinylpyrrolidone for later use; Step S2: Under aqueous phase conditions, berberine solution, proanthocyanidin B2 solution, and CuCl2·2H2O solution are added sequentially to the reaction system and stirred at room temperature for 25-35 minutes to ensure uniform mixing. Step S3: Then add NaOH solution to adjust the pH to 7-8, and continue stirring for 3 hours to promote complexation and self-assembly to form BPCu nanoparticles; Step S4: Next, add PVP solution and sonicate for 10-15 minutes to achieve surface modification of nanoparticles and form PVP@BPCu nanomaterials. Step S5: Purify the obtained material using ultrafiltration centrifugation, wash to remove impurities, and then resuspend and store it.

6. The method for preparing antioxidant nanomaterials based on the self-assembly of traditional Chinese medicine and copper ions according to claim 5, characterized in that: The molecular weight of the polyvinylpyrrolidone used in step S1 is 45,000-58,000.

7. The method for preparing antioxidant nanomaterials based on the self-assembly of traditional Chinese medicine and copper ions according to claim 6, characterized in that: In step S1, the concentration of NaOH solution is 5 mg / mL, the concentration of CuCl2·2H2O solution is 20 mg / mL, the concentration of proanthocyanidin B2 aqueous solution is 5 mg / mL, the concentration of berberine solution is 1 mg / mL, and the concentration of polyvinylpyrrolidone aqueous solution is 40 mg / mL.

8. The method for preparing antioxidant nanomaterials based on the self-assembly of traditional Chinese medicine and copper ions according to claim 7, characterized in that: The mass ratio of berberine, proanthocyanidin B2, and CuCl2·2H2O selected in step S2 is 1:0.9-1.1:0.5-0.

75.

9. The method for preparing antioxidant nanomaterials based on the self-assembly of traditional Chinese medicine and copper ions according to claim 8, characterized in that: In step S2, the mass ratio of berberine, proanthocyanidin B2, and CuCl2·2H2O is 1:1:0.

67.

10. The method for preparing antioxidant nanomaterials based on the self-assembly of traditional Chinese medicine and copper ions according to claim 5, characterized in that: In step S5, the nanoparticles are purified using a 100kDa ultrafiltration centrifuge tube, centrifuged at 3000rpm for 10 minutes, and washed three times.

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