Polyamide separation membrane, method for producing the same, and use thereof

Abstract

The application discloses a polyamide separation membrane and a preparation method and application thereof. The polyamide separation membrane takes an organic membrane as a base, is formed by chemical cross-linking of a diamine monomer containing a biphenyl structure with a long molecular size and a trimesoyl chloride through an interfacial polymerization mode, and has a separation membrane with a polyamide network structure. The separation membrane has holes with a proper size on a membrane structure, has a good interception effect in protein separation, and the amide structure on the separation membrane can make the separation membrane have good chemical stability and thermal stability. Meanwhile, the polyamide separation membrane preparation method is simple, low in cost, can realize batch and rapid preparation, meets the requirements of protein separation, and has a good application and promotion prospect.

Description

Technical Field

[0001] This invention relates to the field of protein separation and purification technology, and in particular to a polyamide separation membrane, its preparation method, and its application. Background Technology

[0002] Proteins play a vital role in the composition of organisms and often exist in complex solution systems. This makes protein purification a crucial process in protein production. Proteins are highly sensitive to their environment; changes in pH, temperature, and other factors can easily cause denaturation during separation. Currently, commonly used protein separation and purification methods include precipitation, centrifugation, electrophoresis, ion exchange, and chromatography. However, these methods also have many problems, such as high separation costs, complex equipment, cumbersome processes, and low separation efficiency, hindering their widespread adoption (Membr. Sci. Tech., 2011, 1-6; J. Biol., 2010, 27, 43-46; Polym Bull., 2015, 29-36). Membrane separation technology, on the other hand, is gaining increasing attention due to its advantages such as simple operation, generally no phase change, good economic benefits, high separation coefficient, high efficiency, energy saving, no secondary pollution, mild working environment (can be carried out at room temperature), and ease of scale-up.

[0003] Membrane separation technology primarily relies on the selective permeability of membranes. Under a certain driving force, one or more components in a mixture to be separated permeate through the membrane, thereby achieving the separation of the mixture and realizing the extraction, purification, concentration, fractionation, or enrichment of products (Polym. Int., 2003, 52, 138-145). Commonly used membrane separation technologies include microfiltration (MF), ultrafiltration (UF), nanofiltration (NF), and reverse osmosis (RO). Most membrane separation technologies rely on pressure difference as the driving force to complete the membrane separation process. Applying a certain pressure difference across the membrane allows solutes and solvents smaller than the membrane pore size to pass through, while components larger than the membrane pore size are retained (J. Membr. Sci., 2005, 249, 245-249). Moreover, different preparation methods and materials used in the preparation of separation membranes will result in different effects.

[0004] The main methods for preparing protein separation membranes currently include phase inversion, dissolution, coating (spin coating, spray coating, dip coating), stretching, etching, sintering, calendering, chemical polymerization (interfacial polymerization), in-situ polymerization, plasma polymerization, and chemical grafting. This invention selects interfacial polymerization to prepare the protein separation membrane because it offers advantages such as low temperature, high speed, high product molecular weight, simple equipment, convenient operation, and less stringent requirements on monomer purity and proportions.

[0005] Currently, commonly used separation membrane materials mainly include chitin-based, polyolefin-based, polysulfone-based, fluorinated polymers, cellulose-based, and polyamide-based materials. Among these, cellulose acetate is the most widely studied and applied fiber-based separation membrane prepared by electrospinning. However, due to the easy hydrolysis of the -COOR in its molecular chain under acidic and alkaline conditions, it has poor chemical and thermal stability and is easily degraded. It also suffers from limited selectivity in spinning solvent systems, large fiber size, and wide diameter distribution (ACS Appl. Mater. Inter., 2014, 6, 20958-20967; J. Memb. Sci., 2015, 492, 547-558; J. Eng. Fibers. Fabr., 2014, 9, 146-152). Currently, there is a greater need for more stable separation membrane materials that also possess appropriately sized pores. Summary of the Invention

[0006] The purpose of this invention is to provide a method for preparing a polyamide separation membrane that has both chemical and thermal stability and can effectively separate proteins.

[0007] Another object of the present invention is to provide a polyamide separation membrane prepared by the above-described method for preparing a polyamide separation membrane.

[0008] Another object of the present invention is to provide an application of the above-described polyamide separation membrane in protein separation.

[0009] Therefore, the technical solution of the present invention is as follows:

[0010] A method for preparing a polyamide separation membrane, the specific steps of which are as follows:

[0011] S1. Dissolve the diamine monomer in a solvent to prepare a diamine solution with a concentration of 0.5 mg / mL to 15 mg / mL; wherein the chemical structure of the diamine monomer contains a biphenyl structure, and the biphenyl structure has a substituent at the 2,2' position;

[0012] S2. Add 2 to 10 times the molar amount of the acid-binding agent to the diamine solution, which is the same as the molar amount of the diamine monomer containing the biphenyl structure, and mix well to obtain a diamine solution with the acid-binding agent.

[0013] S3. First, immerse the base film in a solvent to swell for at least 2 hours, then immerse it in a diamine solution containing an acid-binding agent for at least 0.1 hours, and then remove it and remove the excess solution from the surface of the film.

[0014] S4. Dissolve pyromellitic methyl chloride in a hydrocarbon solvent to prepare a 0.1 wt.% to 2 wt.% TMC solution;

[0015] S5. Place the membrane obtained in step S3 in a TMC solution and soak it for at least 0.5 h; then take it out, rinse it with a membrane cleaning solvent, and air dry it naturally to obtain a polyamide separation membrane.

[0016] Based on the above preparation process, it can be seen that the polyamide separation membrane is prepared by crosslinking a diamine monomer containing a biphenyl structure with a relatively long molecular size with tribenzoyl chloride on a base membrane. Among them, the biphenyl structure in the diamine monomer has a large spatial structure, and different substituents on it provide different chemical properties to the polymer, which has a certain degree of designability. Based on this, this application uses a diamine monomer with substituents at the 2,2' position of the biphenyl structure, so as to utilize its large steric hindrance to a certain extent to hinder the rotation of the single bond in the biphenyl structure, ensuring that the product of the polymerization of the diamine monomer and tribenzoyl chloride can better form a planar structure. At the same time, the two can also form a suitable porous structure after polymerization, which is suitable for the application of daily chemical membranes in the separation and purification of proteins.

[0017] Preferably, in step S1, the diamine monomer containing the biphenyl structure is 2,2'-bis(trifluoromethyl)benzidine, 2,2'-dimethylbenzidine, or 2,2'-dihydroxybenzidine.

[0018] Preferably, in step S1, the solvent is at least one selected from methanol, ethanol, isopropanol, N,N-dimethylformamide, acetone, ethyl acetate, dichloromethane, N,N-diisopropylacetamide, N-methylpyrrolidone, and dimethyl sulfoxide. Preferably, 4-dimethylaminopyridine is used as the N-dimethylaminopyridine.

[0019] Preferably, in step S2, the acid-binding agent is at least one of sodium hydroxide, tetrabutylammonium hydroxide, pyridine, N,N-diisopropylethylamine, and N-dimethylaminopyridine.

[0020] Preferably, in step S3, the base membrane is a nylon 6 membrane, polyethersulfone membrane, polysulfone membrane, polyolefin membrane, polyvinylidene fluoride membrane or polypropylene membrane with a pore size range of 0.22μm to 0.8μm.

[0021] Preferably, in step S3, the swelling time of the base film is 2 to 20 hours.

[0022] Preferably, in step S3, the soaking time of the base film is 0.1h to 1h.

[0023] Preferably, in step S4, the hydrocarbon solvent is at least one selected from n-hexane, cyclohexane, n-heptane, toluene, and benzene.

[0024] Preferably, in step S5, the soaking time is 0.5h to 1.5h.

[0025] Preferably, in step S5, the membrane cleaning solvent is at least two of methanol, pure water, n-hexane, ethanol, and isopropanol, and the membrane is rinsed sequentially.

[0026] A polyamide separation membrane prepared by the above-described method for preparing polyamide separation membranes.

[0027] An application using the above-described polyamide separation membrane: the polyamide separation membrane is used for protein separation.

[0028] Compared with existing technologies, this polyamide separation membrane is prepared by crosslinking a diamine monomer with a biphenyl structure and trimesoyl chloride through interfacial polymerization. The membrane structure has pores of suitable size, and the amide structure on the membrane enables it to have good chemical and thermal stability. At the same time, the preparation method of this polyamide separation membrane is simple and low-cost, and it can be prepared in batches and rapidly, meeting the requirements of protein separation and showing good application and promotion prospects. Attached Figure Description

[0029] Figure 1 The infrared spectrum of the polyamide separation membrane prepared in Example 1 of the present invention;

[0030] Figure 2 A scanning electron microscope (SEM) image of the polyamide separation membrane prepared in Example 1 of the present invention;

[0031] Figure 3 DSC image of the polyamide separation membrane prepared in Example 1 of the present invention;

[0032] Figure 4 Thermogravimetric images of the polyamide separation membrane prepared in Example 1 of the present invention;

[0033] Figure 5 The ultraviolet spectrum of the polyamide separation membrane prepared in Example 1 of the present invention is shown in the test results.

[0034] Figure 6 The image shows the XRD pattern of the polyamide separation membrane prepared in Example 1 of this invention. Detailed Implementation

[0035] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, but the following embodiments are by no means intended to limit the present invention.

[0036] Example 1

[0037] A polyamide separation membrane is prepared by the following method:

[0038] S1. Dissolve 300 mg of 2,2'-bis(trifluoromethyl)benzidine in 30 mL of anhydrous methanol to prepare a 10 mg / mL diamine solution.

[0039] S2. Add N,N-diisopropylethylamine, which is 4 times the molar amount of diamine, to the diamine solution as an acid-binding agent, mix and stir evenly to obtain a diamine solution with the acid-binding agent added.

[0040] S3. Take a 0.45μm nylon 6 (Tianjin Jinteng Experimental Equipment Co., Ltd., TJMF0428, Ф25·0.45μ organic system) as the base membrane. First, soak it in anhydrous methanol for 12h to allow it to swell, and then soak it in a diamine solution with an acid-binding agent for 10min. Take out the membrane and remove the excess solution from the surface with filter paper.

[0041] S4. Dissolve 150 mg of trimesoyl chloride in n-hexane to prepare a 0.5 wt.% TMC solution;

[0042] S5. Place the membrane obtained in step S3 into a TMC solution and soak for 1.5 hours. After removing the membrane, rinse it with methanol and n-hexane in sequence and let it air dry to obtain a polyamide separation membrane.

[0043] The chemical reaction process between 2,2'-bis(trifluoromethyl)benzidine and trimesoyl chloride in Example 1 is shown in the following formula:

[0044]

[0045] Example 2

[0046] A polyamide separation membrane is prepared by the following method:

[0047] S1. Dissolve 300 mg of 2,2'-dimethylbenzidine in 30 mL of anhydrous methanol to prepare a 10 mg / mL diamine solution;

[0048] S2. Add 3 times the molar amount of tetrabutylammonium hydroxide to the diamine solution as an acid-binding agent, mix and stir evenly to obtain a diamine solution with added acid-binding agent;

[0049] S3. Take a polyethersulfone (Haining Keluo Membrane Filtration Equipment Co., Ltd.; KL-PES-47mm-0.45μm) with a pore size of 0.45μm as the base membrane. First, soak it in anhydrous methanol for 5 hours to allow it to swell. Then, soak it in a diamine solution with an acid-binding agent for 10 minutes. Take out the membrane and remove the excess solution from the surface with filter paper.

[0050] S4. Dissolve 300 mg of trimesoyl chloride in cyclohexane to prepare a 1 wt.% TMC solution;

[0051] S5. Place the membrane obtained in step S3 into the TMC solution and soak for 30 minutes. After taking out the membrane, rinse it with methanol and n-hexane in sequence and let it air dry to obtain a polyamide separation membrane.

[0052] Example 3

[0053] A polyamide separation membrane is prepared by the following method:

[0054] S1. Dissolve 15 mg of 2,2'-dihydroxybenzidine in 30 mL of anhydrous methanol to prepare a 0.5 mg / mL diamine solution;

[0055] S2. Add pyridine, which is twice the molar amount of diamine, to the diamine solution as an acid-binding agent, mix and stir evenly to obtain a diamine solution with the acid-binding agent added.

[0056] S3. Take a polyethersulfone (Haining Keluo Membrane Filtration Equipment Co., Ltd.; KL-PES-47mm-0.45μm) with a pore size of 0.45μm as the base membrane. First, soak it in anhydrous methanol for 2 hours to allow it to swell. Then, soak it in a diamine solution with an acid-binding agent for 10 minutes. Take out the membrane and remove the excess solution from the surface with filter paper.

[0057] S4. Dissolve 30 mg of trimesoyl chloride in cyclohexane to prepare a 0.1 wt.% TMC solution;

[0058] S5. Place the membrane obtained in step S3 into a TMC solution and soak for 1.5 hours. After removing the membrane, rinse it with methanol and n-hexane in sequence and let it air dry to obtain a polyamide separation membrane.

[0059] Example 4

[0060] A polyamide separation membrane is prepared by the following method:

[0061] S1. Dissolve 300 mg of 2,2'-bis(trifluoromethyl)benzidine in 30 mL of anhydrous ethanol to prepare a 10 mg / mL diamine solution.

[0062] S2. Add 4-dimethylaminopyridine, which is 5 times the molar amount of diamine, to the diamine solution as an acid-binding agent, mix and stir evenly to obtain a diamine solution with the acid-binding agent added.

[0063] S3. Take a polyvinylidene fluoride (Tianjin Jinteng Experimental Equipment Co., Ltd., TJMF0410, Ф13·0.45μm organic system) with a pore size of 0.45μm as the base membrane. First, immerse it in anhydrous methanol for 3 hours to allow it to swell, and then immerse it in a diamine solution with an acid-binding agent for 10 minutes. Take out the membrane and remove the excess solution from the surface.

[0064] S4. Dissolve 150 mg of trimesoyl chloride in cyclohexane to prepare a 0.5 wt.% TMC solution;

[0065] S5. Place the membrane obtained in step S3 into the TMC solution and soak for 30 minutes. After taking out the membrane, rinse it with methanol and n-hexane in sequence and let it air dry to obtain a polyamide separation membrane.

[0066] Example 5

[0067] A polyamide separation membrane is prepared by the following method:

[0068] S1. Dissolve 300 mg of 2,2'-dimethylbenzidine in 30 mL of anhydrous methanol to prepare a 10 mg / mL diamine solution;

[0069] S2. Add N,N-diisopropylethylamine, which is 3 times the molar amount of diamine, to the diamine solution as an acid-binding agent, mix and stir evenly to obtain a diamine solution with the acid-binding agent added.

[0070] S3. Take a polyvinylidene fluoride (Tianjin Jinteng Experimental Equipment Co., Ltd., TJMF0410, Ф13·0.22μ organic system) with a pore size of 0.22μm as the base membrane. First, immerse it in anhydrous methanol for 5 hours to allow it to swell, and then immerse it in a diamine solution with an acid-binding agent for 10 minutes. Take out the membrane and remove the excess solution from the surface with filter paper.

[0071] S4. Dissolve 150 mg of trimesoyl chloride in n-heptane to prepare a 0.5 wt.% TMC solution;

[0072] S5. Place the membrane obtained in step S3 into the TMC solution and soak for 30 minutes. After taking out the membrane, rinse it with methanol and n-hexane in sequence and let it air dry to obtain a polyamide separation membrane.

[0073] Example 6

[0074] A polyamide separation membrane is prepared by the following method:

[0075] S1. Dissolve 300 mg of 2,2'-bis(trifluoromethyl)benzidine in 30 mL of isopropanol to prepare a 10 mg / mL diamine solution.

[0076] S2. Add N,N-diisopropylethylamine, which is 3 times the molar amount of diamine, to the diamine solution as an acid-binding agent, mix and stir evenly to obtain a diamine solution with the acid-binding agent added.

[0077] S3. Take a piece of nylon 6 (Tianjin Jinteng Experimental Equipment Co., Ltd., TJMF0428, Ф25·0.8μm organic system) with a pore size of 0.8μm as the base membrane. First, soak it in anhydrous methanol for 8 hours to allow it to swell. Then, soak it in a diamine solution with an acid-binding agent for 30 minutes. Take out the membrane and remove the excess solution from the surface with filter paper.

[0078] S4. Dissolve 150 mg of trimesoyl chloride in n-hexane to prepare a 0.5 wt.% TMC solution;

[0079] S5. Place the membrane obtained in step S3 into the TMC solution and soak for 30 minutes. After taking out the membrane, rinse it with methanol and n-hexane in sequence and let it air dry to obtain a polyamide separation membrane.

[0080] Example 7

[0081] A polyamide separation membrane is prepared by the following method:

[0082] S1. Dissolve 450 mg of 2,2'-bis(trifluoromethyl)benzidine in 30 mL of anhydrous methanol to prepare a diamine solution with a concentration of 15 mg / mL.

[0083] S2. Add 10 times the molar amount of N,N-diisopropylethylamine to the diamine solution as an acid-binding agent, mix and stir evenly to obtain a diamine solution with the acid-binding agent added.

[0084] S3. Take a polyethersulfone (Haining Keluo Membrane Filtration Equipment Co., Ltd.; KL-PES-47mm-0.45μm) with a pore size of 0.45μm as the base membrane. First, soak it in anhydrous methanol for 20h to allow it to swell, and then soak it in a diamine solution with an acid-binding agent for 1h. Take out the membrane and remove the excess solution from the surface with filter paper.

[0085] S4. Dissolve 600 mg of trimesoyl chloride in n-hexane to prepare a 2 wt.% TMC solution;

[0086] S5. Place the membrane obtained in step S3 into a TMC solution and soak for 1.5 hours. After removing the membrane, rinse it with methanol and n-hexane in sequence and let it air dry to obtain a polyamide separation membrane.

[0087] Example 8

[0088] A polyamide separation membrane is prepared by the following method:

[0089] S1. Dissolve 300 mg of 2,2'-dihydroxybenzidine in 30 mL of anhydrous methanol to prepare a 10 mg / mL diamine solution.

[0090] S2. Add N,N-diisopropylethylamine, which is 4 times the molar amount of diamine, to the diamine solution as an acid-binding agent, mix and stir evenly to obtain a diamine solution with the acid-binding agent added.

[0091] S3. Take a piece of polyvinylidene fluoride (Tianjin Jinteng Experimental Equipment Co., Ltd., TJMF0410, Ф13·0.45μm organic system) with a pore size of 0.45μm as the base membrane. First, immerse it in anhydrous methanol for 2 hours to allow it to swell, and then immerse it in a diamine solution with an acid-binding agent for 10 minutes. Take out the membrane and remove the excess solution from the surface with filter paper.

[0092] S4. Dissolve 150 mg of trimesoyl chloride (TMC) in n-hexane to prepare a 0.5 wt.% TMC solution;

[0093] S5. Place the membrane obtained in step S3 into the TMC solution and soak for 30 minutes. After taking out the membrane, rinse it with methanol and n-hexane in sequence and let it air dry to obtain a polyamide separation membrane.

[0094] Performance testing:

[0095] (I) Infrared structural characterization:

[0096] like Figure 1 The image shows the infrared spectrum of the polyamide separation membrane prepared in Example 1; it can be seen from the image that the wavenumber is around 3331 cm⁻¹. -1 A bending vibration absorption peak of the -NH amino group appears nearby; wavenumber 1662 cm⁻¹ -1 An absorption peak appears near the acyl group; wavenumber 1600 cm⁻¹ -1 Absorption peaks of amide NH appear around the left and right; 1508 cm⁻¹ -1 A vibrational absorption peak of the benzene ring appears nearby; 1415 cm⁻¹ -1 and 1174cm -1 The presence of -CF vibrational absorption peaks on both sides confirms that Example 1 has indeed synthesized a network polyamide polymer with a cross-linked structure. Similarly, the infrared spectra of the polyamide separation membranes prepared in Examples 2-8 also confirm the presence of amide bonds in their chemical structure, indicating that a network polyamide polymer with a cross-linked structure has been synthesized.

[0097] (II) Microstructure characterization:

[0098] like Figure 2 The image shown is a scanning electron microscope (SEM) image of the polyamide separation membrane prepared in Example 1. As can be seen from the image, the polymer exhibits a sheet-like microstructure, indicating that the polyamide polymer has good film-forming properties, meeting the requirements for polymer separation membrane applications. Similarly, the polyamide separation membranes prepared in Examples 2-8 also exhibit a sheet-like structure as verified by SEM images, indicating good film-forming properties.

[0099] like Figure 6 The image shown is an XRD pattern of the polyamide separation membrane prepared in Example 1; it can be seen from the image that the polymer has an amorphous crystalline morphology and 2θ = 21.986°, indicating that the polyamide polymer possesses certain crystallinity and a large interlayer distance. Similarly, the scanning electron microscope (XRD) characterization results of the polyamide separation membranes prepared in Examples 2-8 also show good crystallinity.

[0100] (III) Membrane stability performance test:

[0101] like Figure 3 The image shows the DSC image of the polyamide separation membrane prepared in Example 1. As can be seen from the image, the glass transition onset temperature is 220.9℃, the midpoint is 233.3℃, ​​the inflection point is 234.5℃, and the termination point is 241.9℃, proving that the polyamide separation membrane has excellent thermal stability. Similarly, the DSC test results of the polyamide separation membranes prepared in Examples 2 to 8 confirm that their glass transition temperatures are all >200℃, that is, they all have excellent thermal stability.

[0102] like Figure 4 The image shows the thermogravimetric analysis (TGA) of the polyamide separation membrane prepared in Example 1. As can be seen from the image, a 3% weight loss occurs before approximately 300°C, corresponding to water. The subsequent two slopes represent the weight loss due to the breakage of the two amide groups. Finally, after 800°C, almost only carbon remains, demonstrating that the polyamide separation membrane exhibits good stability before 300°C. Correspondingly, the polyamide separation membranes prepared in Examples 2-8 also showed the same performance characteristics after TGA testing, i.e., good stability before 300°C.

[0103] (iv) Protein retention performance test of the membrane:

[0104] The protein retention performance of the polyamide separation membranes prepared in Examples 1-8 was tested using ultraviolet spectroscopy. Specifically, bovine hemoglobin was used as the protein to be separated. The protein was dissolved in phosphate buffer at pH 7.4 to prepare a protein solution with a concentration of 1 mg / mL.

[0105] like Figure 5 The image shows the UV spectrum of the polyamide separation membrane prepared in Example 1, which is used to test its retention performance. In the UV spectrum, there is a clear absorption peak at a wavelength of 405.5 nm. We use this peak to determine the concentration of bovine hemoglobin before and after separation. The ratio of the peak values ​​shows that the bovine hemoglobin solution after separation is 0.1886 mg / mL. The retention rate of the separation membrane can be calculated to be 96.51% using formula (1).

[0106]

[0107] In the formula, C0 represents the concentration of the protein solution before separation, C p R represents the concentration of the protein solution after separation, and R represents the rejection rate.

[0108] The retention rate test results of the polyamide separation membranes prepared in Examples 1 to 8 are shown in Table 1 below.

[0109] Table 1:

[0110] Experimental Example Retention rate of bovine hemoglobin solution Example 1 96.51％ Example 2 86.68％ Example 3 80.36％ Example 4 82.18％ Example 5 88.95％ Example 6 92.18％ Example 7 85.76％ Example 8 81.14％

[0111] As can be seen from the test results in Table 1 above, the polyamide separation membranes prepared in Examples 1 to 8 have high retention rates for bovine hemoglobin solution, all >80%, and have good protein separation performance. The polyamide separation membrane prepared in Example 1 has the highest retention rate, up to 96.51%.

[0112] In summary, the polyamide separation membranes prepared in Examples 1-8 of this application have both good chemical and thermal stability, and can effectively achieve protein separation.

[0113] The present invention has been described in detail above with reference to specific embodiments and exemplary examples; however, these descriptions should not be construed as limiting the present invention. Those skilled in the art will understand that various equivalent substitutions, modifications, or improvements can be made to the technical solutions and embodiments of the present invention without departing from the spirit and scope of the invention, and all such modifications and improvements fall within the scope of the present invention. The scope of protection of the present invention is defined by the appended claims.

Claims

1. An application of a polyamide separation membrane for protein separation, characterized in that, The preparation steps are as follows: S1. Dissolve the diamine monomer in a solvent to prepare a diamine solution with a concentration of 0.5 mg / mL to 15 mg / mL; wherein the chemical structure of the diamine monomer contains a biphenyl structure, and the biphenyl structure has a substituent at the 2,2' position; The diamine monomer containing the biphenyl structure is 2,2'-bis(trifluoromethyl)benzidine, 2,2'-dimethylbenzidine, or 2,2'-dihydroxybenzidine. S2. Add 2 to 10 times the molar amount of the acid-binding agent to the diamine solution, which is the same as the amount of the diamine monomer containing the biphenyl structure, and mix well to obtain a diamine solution with the acid-binding agent. S3. First, immerse the base film in a solvent to swell for at least 2 hours, then immerse it in a diamine solution containing an acid-binding agent for at least 0.1 hours, and then remove it and remove the excess solution from the surface of the film. S4. Dissolve pyromellitic chloride in a hydrocarbon solvent to prepare a 0.1 wt.%~2 wt.% pyromellitic chloride solution; S5. Place the membrane obtained in step S3 in a pyromellitic methyl chloride solution and soak it for at least 0.5 h; then take it out, rinse it with a membrane cleaning solvent, and air dry it naturally to obtain a polyamide separation membrane.

2. The application of the polyamide separation membrane according to claim 1 for protein separation, characterized in that, In step S1, the solvent is at least one selected from methanol, ethanol, isopropanol, N,N-dimethylformamide, acetone, ethyl acetate, dichloromethane, N,N-diisopropylacetamide, N-methylpyrrolidone, and dimethyl sulfoxide.

3. The application of the polyamide separation membrane according to claim 1 for protein separation, characterized in that, In step S2, the acid-binding agent is at least one of sodium hydroxide, tetrabutylammonium hydroxide, pyridine, N,N-diisopropylethylamine, and N-dimethylaminopyridine.

4. The application of the polyamide separation membrane according to claim 1 for protein separation, characterized in that, The base film in step S3 is a nylon 6 film, a polyethersulfone film, a polysulfone film, a polyolefin film, a polyvinylidene fluoride film, or a polypropylene film.

5. The application of the polyamide separation membrane according to claim 1 for protein separation, characterized in that, In step S4, the hydrocarbon solvent is at least one of n-hexane, cyclohexane, n-heptane, toluene, and benzene.

6. The application of the polyamide separation membrane according to claim 1 for protein separation, characterized in that, In step S5, the membrane cleaning solvent is at least two of methanol, pure water, n-hexane, ethanol, and isopropanol, and the membrane is rinsed sequentially.

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