High-strength western blot membrane based on nanofiber network interpenetration and preparation method of high-strength western blot membrane

Abstract

The invention relates to the technical field of western blot membranes, in particular to a high-strength western blot membrane based on nanofiber network interpenetration and a preparation method thereof.The high-strength western blot membrane comprises the following raw materials: a polyvinylidene fluoride membrane matrix, a nanofiber skeleton, a pore-foaming agent, an N, N-dimethylacetamide cosolvent and coagulating bath; the preparation method comprises the following steps: selecting a cellulose nanofibril aqueous dispersion with the concentration of 2wt%, carrying out hydrophobic modification on the cellulose nanofibril aqueous dispersion through octyl triethoxy silane, and taking the modified cellulose nanofibril aqueous dispersion as a nanofiber skeleton; the pore-foaming agent is a compound mixed solution of polyvinylpyrrolidone and PEG-400 (Polyethylene Glycol). The cellulose nanofibrils are subjected to surface hydrophobization modification, and a compound pore-foaming agent system is introduced, so that the mechanical property and the porous structure of the membrane are considered, and the good porosity of the membrane is maintained while the overall strength of the membrane is enhanced.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of protein blotting membrane, in particular to high-strength protein blotting membrane based on nanofiber network interpenetration and a preparation method thereof. BACKGROUND

[0002] As a key consumable in molecular biology, medical detection and related scientific research experiments, white blotting membrane is widely used in protein separation, fixation and detection. The commonly used polyvinylidene fluoride (PVDF) protein blotting membrane has good chemical stability and protein binding capacity, but its mechanical properties are limited, and it is easy to tear or break during operation, which is not conducive to stable use under complex experimental conditions.

[0003] In order to improve the overall performance of the membrane, nanomaterials are introduced into the base film material to improve the toughness and strength of the membrane. However, the introduction of nanomaterials will cause the degradation of the porous structure, resulting in problems such as widening of the pore size distribution, decrease of effective porosity, and limitation of liquid mass transfer. Therefore, we propose a high-strength protein blotting membrane based on nanofiber network interpenetration and a preparation method thereof. SUMMARY

[0004] The purpose of the present application is to provide a high-strength protein blotting membrane based on nanofiber network interpenetration and a preparation method thereof, to solve the problems of pore structure degradation, widening of pore size distribution, decrease of effective porosity, and limitation of liquid mass transfer caused by the introduction of nanofibers in the background art.

[0005] The present application provides a high-strength protein blotting membrane based on nanofiber network interpenetration, comprising the following raw materials: polyvinylidene fluoride membrane base, nanofiber skeleton, pore former, N,N-dimethylacetamide solvent and coagulation bath.

[0006] Among them, the concentration of cellulose nanofiber aqueous dispersion is 2wt%, and the hydrophobic modification is carried out by using octyl triethoxysilane. The modified cellulose nanofiber dispersion (the source of cellulose nanofiber is an aqueous nanofiber dispersion obtained by chemical treatment of natural wood pulp) is used as the nanofiber skeleton.

[0007] The pore former is a complex mixture of polyvinylpyrrolidone and PEG-400, and the mass ratio of polyvinylpyrrolidone to PEG-400 is 3-5:1.

[0008] As a preferred embodiment, the preparation step of the nanofiber skeleton is as follows:

[0009] S1. Dilute the cellulose nanofiber aqueous dispersion to 0.5wt% with deionized water, add acetone dropwise while stirring, centrifuge at 9,000 rpm for 15 minutes, discard the supernatant to obtain cellulose nanofiber gel, wherein the volume ratio of acetone to aqueous dispersion is 3:1.

[0010] S2. Redisperse the cellulose nanofiber gel in pure acetone, centrifuge again, and repeat 2-3 times to completely remove residual moisture.

[0011] S3. Replace the solvent with N,N-dimethylacetamide and repeat step S2 until acetone is gradually replaced with N,N-dimethylacetamide to finally obtain CNF / DMAc predispersant.

[0012] S4. Transfer the CNF / DMAc predispersant to a three-necked flask equipped with a condenser, and add octyltriethoxysilane to the three-necked flask while stirring.

[0013] S5. Add glacial acetic acid dropwise as a catalyst to adjust the pH of the reaction system to 4-5. The amount of glacial acetic acid used is 0.5% of the mass of octyltriethoxysilane.

[0014] S6. Heat the reaction system to 80-90℃ and react under reflux for 4-6 hours. After the reaction is complete, cool to room temperature.

[0015] S7. Pour the reaction mixture into anhydrous ethanol to precipitate the modified cellulose nanofibers. Collect the precipitate by centrifugation and wash it repeatedly with ethanol by centrifugation 3-4 times until the supernatant is colorless and odorless (without silane residue).

[0016] S8. The purified modified cellulose nanofibers were directly dispersed in N,N-dimethylacetamide (DMAc) and treated by an ultrasonic cell disruptor to obtain a modified CNF / DMAc dispersion with a solid content of 2wt% (i.e., the dry weight of the modified cellulose nanofibers was 2wt%), which served as the nanofiber skeleton.

[0017] Preferably, in the 2wt% cellulose nanofiber aqueous dispersion, the amount of cellulose nanofiber added accounts for 0.2-0.5% of the mass of the polyvinylidene fluoride membrane matrix; and the diameter of the cellulose nanofiber is 8-12 nm.

[0018] Preferably, the amount of octyltriethoxysilane (OTES) added is 20-35 wt% of the dry weight of the cellulose nanofibers.

[0019] Preferably, the polyvinylpyrrolidone has a molecular weight of 150,000-160,000 and a solution concentration of 15 wt%.

[0020] Preferably, the PEG-400 has a molecular weight of 400 and a solution concentration of 3 wt%.

[0021] Preferably, the coagulation bath is water at a temperature of 20-25°C.

[0022] On the other hand, the present invention provides a method for preparing a high-strength protein-imprinted membrane based on interpenetrating nanofiber networks, which is used to prepare the above-mentioned high-strength protein-imprinted membrane based on interpenetrating nanofiber networks. The method for preparing the high-strength protein-imprinted membrane based on interpenetrating nanofiber networks is as follows:

[0023] The polyvinylidene fluoride (PVDF) membrane substrate was dissolved in N,N-dimethylacetamide as a co-solvent, heated to 60-70°C with a magnetic stirrer, and stirred at 500 rpm for 1 hour until completely dissolved to obtain a casting solution. The mass fraction of the PVDF membrane substrate in N,N-dimethylacetamide was 15 wt%.

[0024] Add the pore-forming agent and the modified CNF / DMAc dispersion in sequence. Stir with magnetic stirring at 300 rpm for 5 minutes to coarsely mix. Then, sonicate for 10 minutes to disperse the mixture evenly and obtain the casting solution.

[0025] The casting solution was uniformly coated onto a glass plate, and a film was formed by scraping. The film was then immediately immersed in a coagulation bath for 35 minutes to undergo phase inversion, resulting in a high-strength protein-imprinted membrane based on an interpenetrating nanofiber network.

[0026] Preferably, the amount of pore-forming agent added is 5-15 wt% of the mass of the polyvinylidene fluoride membrane substrate.

[0027] Preferably, the ultrasonic processing parameters are: power 200W, frequency 20kHz.

[0028] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0029] 1. In this invention, a high-strength protein-imprinted membrane based on an interpenetrating nanofiber network and its preparation method are described. Polyvinylidene fluoride (PVDF) is selected as the matrix membrane material, combined with cellulose nanofibers modified with octyltriethoxysilane (OTES) as a reinforcing framework. This allows the nanofibers to form a stable interpenetrating network structure with the matrix, significantly improving the membrane's tensile strength, tear strength, and fracture toughness. Simultaneously, a complex pore-forming agent system of polyvinylpyrrolidone (PVP) and PEG-400 is used. By controlling the compounding ratio and addition amount, precise control of the membrane pore size distribution is achieved, forming a hierarchical porous structure combining macropores and micropores. This allows the membrane to maintain good liquid permeability and stable structural uniformity while possessing high porosity.

[0030] 2. In the high-strength protein-imprinted membrane based on the interpenetrating nanofiber network and its preparation method, the combination of PVP and PEG-400 forms a network of interconnected pores with both macropores and micropores, ensuring high porosity and liquid mass transfer efficiency. Compared with using PVP alone, the compound system significantly improves permeability and enhances the uniformity of pore size distribution, solving the problems of strength and toughness of the protein-imprinted membrane while maintaining a uniform pore size distribution and high effective porosity. Detailed Implementation

[0031] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0032] This invention provides a high-strength protein-imprinted membrane based on interpenetrating nanofiber networks, comprising the following raw materials: polyvinylidene fluoride membrane matrix (PVDF), nanofiber framework, pore-forming agent, N,N-dimethylacetamide (DMAc) cosolvent, and coagulation bath;

[0033] Among them, a 2wt% cellulose nanofiber aqueous dispersion was selected and hydrophobically modified by octyltriethoxysilane. The modified cellulose nanofiber dispersion (the source of cellulose nanofibers is an aqueous nanofiber dispersion obtained by chemical treatment of natural wood pulp) was used as the nanofiber skeleton.

[0034] The pore-forming agent is a compound mixture of polyvinylpyrrolidone and PEG-400, with a mass ratio of 3~5:1 between polyvinylpyrrolidone and PEG-400.

[0035] The nanofiber framework preparation steps are as follows:

[0036] S1. Dilute the cellulose nanofiber aqueous dispersion to 0.5 wt% with deionized water, add acetone dropwise while stirring, centrifuge at 9,000 rpm for 15 minutes, discard the supernatant to obtain cellulose nanofiber (CNF) gel, wherein the volume ratio of acetone to aqueous dispersion is 3:1.

[0037] S2. Redisperse the cellulose nanofiber (CNF) gel in pure acetone, centrifuge again, and repeat 2-3 times to completely remove residual moisture;

[0038] S3. Replace the solvent with N,N-dimethylacetamide (DMAc) and repeat step S2 until acetone is gradually replaced with N,N-dimethylacetamide (DMAc) to finally obtain CNF / DMAc predispersant;

[0039] S4. Transfer the CNF / DMAc predispersant to a three-necked flask equipped with a condenser, and add octyltriethoxysilane (OTES) to the three-necked flask while stirring.

[0040] S5. Add glacial acetic acid dropwise as a catalyst to adjust the pH of the reaction system to 4-5. The amount of glacial acetic acid used is 0.5% of the mass of octyltriethoxysilane.

[0041] S6. Heat the reaction system to 80-90℃ and react under reflux for 4-6 hours. After the reaction is complete, cool to room temperature.

[0042] S7. Pour the reaction mixture into anhydrous ethanol to precipitate the modified cellulose nanofibers. Collect the precipitate by centrifugation and wash it repeatedly with ethanol by centrifugation 3-4 times until the supernatant is colorless and odorless (without silane residue).

[0043] S8. The purified modified cellulose nanofibers were directly dispersed in N,N-dimethylacetamide (DMAc) and treated by an ultrasonic cell disruptor to obtain a modified CNF / DMAc dispersion with a solid content of 2wt% (i.e., the dry weight of the modified cellulose nanofibers was 2wt%), which served as the nanofiber skeleton.

[0044] In a 2wt% aqueous dispersion of cellulose nanofibers, the diameter of the cellulose nanofibers is 8-12 nm;

[0045] Polyvinylpyrrolidone has a molecular weight of 150,000-160,000 and a solution concentration of 15 wt%.

[0046] The molecular weight of PEG-400 is 400, and the solution concentration is 3wt%.

[0047] Example 1: A method for preparing a high-strength protein-imprinted membrane based on interpenetrating nanofiber networks, comprising the following steps:

[0048] A 2wt% aqueous dispersion of cellulose nanofibers was selected and hydrophobically modified with octyltriethoxysilane. The modified cellulose nanofiber dispersion (the cellulose nanofibers were obtained from natural wood pulp through chemical treatment) was used as the nanofiber skeleton.

[0049] Polyvinylidene fluoride (PVDF) membrane substrate was dissolved in N,N-dimethylacetamide (DMAc) co-solvent, heated to 60-70°C with a magnetic stirrer, and stirred at 500 rpm for 1 hour until completely dissolved to obtain a casting solution. The mass fraction of the PVDF membrane substrate in N,N-dimethylacetamide (DMAc) solution was 15 wt%.

[0050] Pore-forming agent and modified CNF / DMAc dispersion were added sequentially. After coarse mixing for 5 minutes at 300 rpm using magnetic stirring, the mixture was ultrasonically treated for 10 minutes at 200 W and 20 kHz to achieve uniform dispersion and obtain the casting solution.

[0051] The casting solution was uniformly coated on a glass plate, and a film was formed by scraping. The film was immediately immersed in a water coagulation bath at 25°C and soaked for 35 minutes to carry out phase transformation, thus obtaining a high-strength protein imprinted membrane based on interpenetrating nanofiber networks.

[0052] The amount of pore-forming agent added is 5 wt% of the mass of the polyvinylidene fluoride (PVDF) membrane matrix;

[0053] The amount of octyltriethoxysilane (OTES) added is 20 wt% of the dry weight of cellulose nanofibers (CNF);

[0054] The amount of cellulose nanofibers (CNF) added accounts for 0.2% of the mass of the polyvinylidene fluoride (PVDF) film matrix, and the diameter of the cellulose nanofibers is 8 nm.

[0055] Example 2: The difference between this example and Example 1 is that the amount of octyltriethoxysilane (OTES) added is 25 wt% of the dry weight of cellulose nanofibers (CNF).

[0056] Example 3: The difference between this example and Example 1 is that the amount of octyltriethoxysilane (OTES) added accounts for 30 wt% of the dry weight of cellulose nanofibers (CNF).

[0057] Example 4: The difference between this example and Example 1 is that the amount of octyltriethoxysilane (OTES) added accounts for 35 wt% of the dry weight of cellulose nanofibers (CNF).

[0058] To demonstrate the effect of octyltriethoxysilane on the performance of the finished high-strength protein imprinted membrane, Example 3 was selected as the optimal example, and the following comparative examples were used to compare it with the examples:

[0059] Comparative Example 1: The difference between this example and Example 3 is that the cellulose nanofiber aqueous dispersion was not modified by octyltriethoxysilane (OTES), and the 2wt% cellulose nanofiber aqueous dispersion was directly used as the nanofiber skeleton.

[0060] Experimental Example: The purpose of this experimental example is to test the high-strength protein-blown membranes prepared in Examples 1-4 and Comparative Example 1;

[0061] Experimental objective: The high-strength protein-imprinted membranes prepared in Examples 1-4 and the high-strength protein-imprinted membrane prepared in Comparative Example 1;

[0062] Sample preparation: High-strength protein blot membrane samples were prepared according to the methods of Examples 1-4 and Comparative Example 1;

[0063] Mechanical property testing:

[0064] Tensile strength: Fix the two ends of the sample with clamps and stretch it at a constant rate (e.g., 5 mm / min), and record the maximum stress at fracture;

[0065] Tear strength: A straight-line tear sample is used to determine the force required for tearing to begin;

[0066] Fracture toughness: Record the energy absorption capacity of the sample material and calculate the fracture toughness per unit area (J / m²).

[0067] Table 1 Performance data of the high-strength protein-imprinted membranes prepared in Examples 1-4 and Comparative Example 1

[0068] Example CNF modification OTES addition (wt%) Tensile strength (MPa) Tear strength (N / mm) Fracture toughness (J / m2) Example 1 20 12.5 5.8 450 Example 2 25 13.8 6.2 480 Example 3 30 14.6 6.5 510 Example 4 35 15.2 6.8 540 Comparative Example 1 Unmodified 10.1 4.5 350

[0069] As can be seen from Examples 1-4 and Table 1 above, as the amount of octyltriethoxysilane added as a percentage of the dry weight of cellulose nanofibers increases, the various performance data (tensile strength, tear strength and fracture toughness) of the high-strength protein imprinted membrane are all enhanced.

[0070] As can be seen from Comparative Example 1, compared with Comparative Example 1, the modification effect of octyltriethoxysilane in Examples 1-4 is obvious. The high-strength protein imprinted membrane has significantly improved tensile strength, tear strength and fracture toughness. This shows that the hydrophobic modification of the surface of cellulose nanofibers by octyltriethoxysilane (OTES) effectively enhances the interfacial bonding force between the nanofibers and the polyvinylidene fluoride membrane matrix (PVDF), thereby improving the overall mechanical properties of the membrane.

[0071] As the amount of octyltriethoxysilane (OTES) added increased from 20 wt% to 35 wt%, the tensile strength, tear strength and fracture toughness of the membrane gradually increased, indicating that the higher the degree of modification, the more perfect the interpenetrating network structure of the nanofiber skeleton and the membrane matrix, and the more significant the mechanical reinforcement effect.

[0072] Although Comparative Example 1 also contains cellulose nanofibers (CNF), it has not been modified with octyltriethoxysilane (OTES), and the various properties of the membrane are significantly lower than those of Examples 1-4, indicating that modification is a key step in achieving high-strength protein-imprinted membranes.

[0073] In summary, surface modification of cellulose nanofibers with octyltriethoxysilane results in a more stable interpenetrating network structure between the nanofiber skeleton and the polyvinylidene fluoride (PVDF) membrane matrix, which significantly improves the tensile strength, tear strength, and fracture toughness of the membrane. Moreover, the degree of performance enhancement is positively correlated with the amount of modifier added.

[0074] Furthermore, to demonstrate the influence of cellulose nanofiber diameter on the performance of the finished high-strength protein-imprinted membrane, Example 3 was used as the optimal example, and compared with the following Examples 5-6:

[0075] Example 5: The difference between this example and Example 3 is that the diameter of the cellulose nanofibers is 10 nm.

[0076] Example 6: The difference between this example and Example 3 is that the diameter of the cellulose nanofibers is 12 nm.

[0077] Using the methods described in the above experimental examples, high-strength protein-imprinted membrane samples were prepared according to the methods in Examples 5-6; and the above mechanical tests were performed, resulting in the data in Table 2.

[0078] Table 2 Performance data of the prepared high-strength protein-imprinted membrane

[0079] Example Tensile strength (MPa) Tear strength (N / mm) Fracture toughness (J / m2) Example 3 14.6 6.5 510 Example 5 14.1 6.1 500 Example 6 13.2 5.9 480

[0080] Based on Examples 3, 5, and 6, and in conjunction with Table 2, it can be seen that:

[0081] As the diameter of cellulose nanofibers increases (8nm, 10nm, 12nm), the performance data of high-strength protein-imprinted membranes gradually decrease. Further increasing the fiber diameter leads to a more significant decrease in the tensile strength, tear strength, and fracture toughness of the membrane. Network sparsity has a significant impact on the mechanical properties of the membrane.

[0082] As the diameter of cellulose nanofibers increases, the density of the nanofiber network decreases. Since the key to the interpenetrating nanofiber network lies in the specific surface area of ​​the fibers and the number of network intersections, as the fiber diameter increases, the number of fibers per unit volume decreases, and the tightness of the intersections and the interpenetrating network decreases. Furthermore, thicker fibers lead to a wider distribution of network pores and a relatively smaller number of micropores, thereby affecting the overall toughness and stress dispersion ability of the membrane.

[0083] Therefore, for tensile strength, the larger the diameter of the cellulose nanofibers, the lower the network density, which leads to a slight decrease in the strength of the membrane under tensile conditions.

[0084] Regarding tear strength, the larger the diameter of cellulose nanofibers, the lower the network density, resulting in a sparser interpenetrating network of fibers. This leads to fewer fiber intersections around the crack tip, reducing the resistance to crack propagation and making the membrane more prone to crack propagation.

[0085] Regarding fracture toughness, a decrease in network density (sparse network) leads to significant local stress concentration, which weakens the membrane's ability to absorb and disperse energy under external forces.

[0086] In summary, the diameter of cellulose nanofibers directly affects the density and number of intersections of the interpenetrating network, thus determining the membrane's ability to resist cracks and absorb energy, which is ultimately reflected in tensile strength, tear strength, and fracture toughness. Therefore, when the diameter of cellulose nanofibers is in the range of 8-12 nm, the tensile strength, tear strength, and fracture toughness are optimal when the diameter of cellulose nanofibers is 8 nm.

[0087] Furthermore, to demonstrate the effect of the pore-forming agent on the performance of the finished high-strength protein-imprinted membrane, Example 3 was used as the optimal example, and performance comparison analysis was conducted through Examples 7-9 below:

[0088] Example 7: The difference between this example and Example 3 is that the amount of pore-forming agent added is 10 wt% of the mass of the polyvinylidene fluoride (PVDF) membrane matrix.

[0089] Example 8: The difference between this example and Example 3 is that the amount of pore-forming agent added is 15 wt% of the mass of the polyvinylidene fluoride (PVDF) membrane matrix.

[0090] Example 9: The difference between this example and Example 3 is that the pore-forming agent is a compound mixture of polyvinylpyrrolidone and PEG-400, and the mass ratio of polyvinylpyrrolidone to PEG-400 is 4:1.

[0091] Example 10: The difference between this example and Example 3 is that the pore-forming agent is a compound mixture of polyvinylpyrrolidone and PEG-400, and the mass ratio of polyvinylpyrrolidone to PEG-400 is 5:1.

[0092] Comparative Example 2: The difference between this example and Example 3 is that the pore-forming agent is polyvinylpyrrolidone.

[0093] Among them, the permeability test: high-strength protein imprinted membrane samples were prepared by the preparation methods of Examples 3, 9-10 and Comparative Example 2;

[0094] The high-strength protein blot membrane sample is fixed in the filter device to ensure that the membrane fits tightly and there is no leakage at the edges. A certain height of liquid is added above the membrane (controlled by constant pressure or constant pressure pump). The liquid flowing out from below the membrane is collected, and the volume of liquid passing through the membrane per unit time is measured. The measurement is repeated more than 3 times and the average value is taken.

[0095] Using the methods described in the above experimental examples, high-strength protein-imprinted membrane samples were prepared according to the methods of Examples 7-10 and Comparative Example 2; and the above mechanical tests and permeability tests were performed, resulting in the data in Table 3.

[0096] Table 3 Performance data of high-strength protein-imprinted membranes

[0097] Example Tensile strength (MPa) Tear strength (N / mm) Fracture toughness (J / m2) Permeability (mL / cm2 min) Example 3 14.6 6.5 510 0.50 Example 7 14.2 6.3 515 / Example 8 13.8 6.0 519 / Example 9 14.4 6.3 521 0.45 Example 10 14.2 6.1 530 0.42 Comparative Example 2 12.5 5.8 500 0.30

[0098] Using Example 3 as a comparative example, and referring to Table 3, a comparison of Examples 3 and 7-8 shows that:

[0099] As the amount of pore-forming agent added increased to the mass ratio of polyvinylidene fluoride (PVDF) membrane matrix (increasing to 5wt%, 10wt%, and 15wt% respectively), the performance data (tensile strength and tear strength) of the high-strength protein-imprinted membrane decreased slightly. This was because the increased mass ratio of PVDF membrane matrix led to an increase in the number and porosity of micropores in the membrane. The pores did not participate in the load-bearing under tensile or tearing loads, resulting in a decrease in the effective load-bearing cross-sectional area of ​​the material, which manifested as a slight decrease in tensile strength and tear strength.

[0100] Furthermore, high porosity areas may become local stress concentration points, causing micro-fractures to occur locally during tensile or tearing processes, further reducing the overall strength; increased porosity leads to local sparse networks, resulting in a decrease in the ability to withstand local tensile and tearing loads.

[0101] The fracture toughness increased slightly because micropores and channels can absorb strain energy during crack propagation, thus slowing down the crack propagation rate. When the crack encounters pores or the nanofiber bridging zone around the pores, additional energy is required to overcome the pores and fiber network, thereby improving the fracture toughness. Furthermore, the more pores there are, the more tortuous the crack path becomes, resulting in more energy absorbed per unit area and thus improving the fracture toughness.

[0102] In summary, the addition of pore-forming agents makes the micropore distribution more uniform, which is conducive to the uniform dissipation of energy during crack propagation, rather than the concentration at a few crack tips, thereby enhancing the overall toughness of the membrane.

[0103] By comparing the data in Table 2 with Examples 3, 9-10, and Comparative Example 2, it can be seen that:

[0104] As the mass ratio of polyvinylpyrrolidone to PEG-400 increases (increasing to 3:1, 4:1 and 5:1 respectively), the tensile strength and tear strength decrease slightly. This is because the increased proportion of polyvinylpyrrolidone inhibits micropore formation, resulting in a reduction in micropore formation assistance, which in turn leads to a slight decrease in tensile strength and tear strength.

[0105] As the proportion of polyvinylpyrrolidone (PVP) increases, the energy absorption capacity of the nanofiber skeleton continues to increase, thus gradually improving fracture toughness. This is because the increased proportion of PPVP improves the dispersion of CNF, allowing the energy absorption capacity of the nanofiber skeleton to be fully utilized, greatly enhancing the toughening effect of the nanofiber network. The resulting gain outweighs the loss of microporous toughening effect caused by the reduction of PEG-400, ultimately resulting in an overall improvement in fracture toughness.

[0106] In Comparative Example 2, the lack of polyvinylpyrrolidone resulted in the absence of micropores, concentrated pore size distribution, and a significant decrease in permeability.

[0107] In summary, PEG-400 directly affects the permeability, pore size distribution, and mechanical properties of high-strength protein-imprinted membranes by regulating the number, size, and permeability of membrane micropores. Due to the small molecular weight of PEG-400, it has excellent miscibility with water (coagulation bath). When the cast membrane is immersed in the coagulation bath, PEG-400 molecules diffuse from the polymer-rich phase (PVDF / CNF phase) and dissolve into the water extremely rapidly.

[0108] The combination of polyvinylpyrrolidone (PVP) and PEG-400 has the following advantages: PVP acts as the backbone, responsible for forming a stable macroporous framework with a certain strength; while PEG-400 creates a large number of tiny micropores and channels inside and between the macroporous framework formed by PVP, allowing liquids (such as buffer solutions and antibody solutions) to penetrate the overall thickness of the membrane very efficiently, thus achieving high permeability.

[0109] The foregoing has shown and described the basic principles, main features, and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The embodiments and descriptions in the specification are merely preferred examples and are not intended to limit the invention. Various changes and modifications can be made to the invention without departing from its spirit and scope, and all such changes and modifications fall within the scope of the present invention as claimed. The scope of protection of the present invention is defined by the appended claims and their equivalents.

Claims

1. A high-strength protein-imprinted membrane based on interpenetrating nanofiber networks, characterized in that, The raw materials include: polyvinylidene fluoride membrane matrix, nanofiber skeleton, pore-forming agent, N,N-dimethylacetamide cosolvent and coagulation bath; Among them, a 2wt% cellulose nanofiber aqueous dispersion was selected and hydrophobically modified with octyltriethoxysilane, and the modified cellulose nanofiber dispersion was used as the nanofiber skeleton. The pore-forming agent is a compound mixture of polyvinylpyrrolidone and PEG-400, with a mass ratio of 3~5:1 between polyvinylpyrrolidone and PEG-400.

2. The high-strength protein-imprinted membrane based on interpenetrating nanofiber networks according to claim 1, characterized in that, The nanofiber framework preparation steps are as follows: S1. Dilute the cellulose nanofiber aqueous dispersion to 0.5wt% with deionized water, add acetone dropwise while stirring, centrifuge at 9,000 rpm for 15 minutes, discard the supernatant to obtain cellulose nanofiber gel, wherein the volume ratio of acetone to aqueous dispersion is 3:

1. S2. Redisperse the cellulose nanofiber gel in pure acetone, centrifuge again, and repeat 2-3 times to completely remove residual moisture. S3. Replace the solvent with N,N-dimethylacetamide and repeat step S2 until acetone is gradually replaced with N,N-dimethylacetamide to finally obtain CNF / DMAc predispersant. S4. Transfer the CNF / DMAc predispersant to a three-necked flask equipped with a condenser, and add octyltriethoxysilane to the three-necked flask while stirring. S5. Add glacial acetic acid dropwise as a catalyst to adjust the pH of the reaction system to 4-5. The amount of glacial acetic acid used is 0.5% of the mass of octyltriethoxysilane. S6. Heat the reaction system to 80-90℃ and react under reflux for 4-6 hours. After the reaction is complete, cool to room temperature. S7. Pour the reaction mixture into anhydrous ethanol to precipitate the modified cellulose nanofibers. Collect the precipitate by centrifugation and wash it repeatedly with ethanol by centrifugation 3-4 times until the supernatant is colorless and odorless. S8. The purified modified cellulose nanofibers were directly dispersed in N,N-dimethylacetamide and treated with an ultrasonic cell disruptor to obtain a modified CNF / DMAc dispersion with a solid content of 2wt%, which served as the nanofiber skeleton.

3. The high-strength protein-imprinted membrane based on interpenetrating nanofiber networks according to claim 1, characterized in that, The amount of cellulose nanofibers added accounts for 0.2-0.5% of the mass of the polyvinylidene fluoride membrane matrix; and the diameter of the cellulose nanofibers is 8-12 nm.

4. The method for preparing a high-strength protein-imprinted membrane based on interpenetrating nanofiber networks according to claim 1, characterized in that, The amount of octyltriethoxysilane added is 20-35 wt% of the dry weight of the cellulose nanofibers.

5. The method for preparing a high-strength protein-imprinted membrane based on interpenetrating nanofiber networks according to claim 1, characterized in that, The polyvinylpyrrolidone has a molecular weight of 150,000-160,000 and a solution concentration of 15 wt%.

6. The method for preparing a high-strength protein-imprinted membrane based on interpenetrating nanofiber networks according to claim 1, characterized in that, The PEG-400 has a molecular weight of 400 and a solution concentration of 3wt%.

7. The method for preparing a high-strength protein-imprinted membrane based on interpenetrating nanofiber networks according to claim 1, characterized in that, The coagulation bath is water, and the temperature is 20-25℃.

8. A method for preparing a high-strength protein-imprinted membrane based on interpenetrating nanofiber networks, used to prepare a high-strength protein-imprinted membrane based on interpenetrating nanofiber networks as described in any one of claims 1-7, characterized in that, The method for preparing the high-strength protein-imprinted membrane based on interpenetrating nanofiber networks is as follows: The polyvinylidene fluoride (PVDF) membrane substrate was dissolved in N,N-dimethylacetamide as a co-solvent, heated to 60-70°C with a magnetic stirrer, and stirred at 500 rpm for 1 hour until completely dissolved to obtain a casting solution. The mass fraction of the PVDF membrane substrate in N,N-dimethylacetamide was 15 wt%. Add the pore-forming agent and the modified CNF / DMAc dispersion in sequence. Stir with magnetic stirring at 300 rpm for 5 minutes to coarsely mix. Then, sonicate for 10 minutes to disperse the mixture evenly and obtain the casting solution. The casting solution was uniformly coated onto a glass plate, and a film was formed by scraping. The film was then immediately immersed in a coagulation bath for 35 minutes to undergo phase inversion, resulting in a high-strength protein-imprinted membrane based on an interpenetrating nanofiber network.

9. The method for preparing a high-strength protein-imprinted membrane based on interpenetrating nanofiber networks according to claim 8, characterized in that, The amount of pore-forming agent added is 5-15 wt% of the mass of the polyvinylidene fluoride membrane substrate.

10. The method for preparing a high-strength protein-imprinted membrane based on interpenetrating nanofiber networks according to claim 8, characterized in that, The ultrasonic processing parameters are: power 200W, frequency 20kHz.