A cellulose porous membrane for virus removal and a method for manufacturing the same

Abstract

This invention provides a cellulose porous membrane for virus removal and its preparation method. The porous membrane comprises a main body, one side of which is a first outer surface and the other side is a second outer surface. The average pore size of the first outer surface is larger than that of the second outer surface. The porosity of the porous membrane is 15%-45%. In the wetted porous membrane body, the region that captures colloidal gold with a diameter of 20 nm is designated as D20. D20 is located in a region of 5%-100% of the thickness of the main body, and the thickness of the D20 region is 15-60 μm. The membrane also captures colloidal gold with a diameter of 3 nm. The 0nm colloidal gold is located at D30, and the overlapping region of D30 and D20 is L1. L1 is located on the side of the D20 region near the first outer surface, and the thickness ratio of L1 to D20 is not less than 0.15. This porous membrane is integrally formed, and the preparation method is green. At the same time, it has a relatively thick pore region, and the pore size changes very little with thickness in the pore region. This makes the porous membrane have a strong retention effect on small viruses with a particle size of 20nm and above, and there is no risk of virus leakage. It also has high protein yield and high throughput.

Description

Technical Field

[0001] This invention relates to the field of membrane materials technology, and more specifically to a cellulose porous membrane for virus removal and its preparation method. Background Technology

[0002] Membrane separation technology is a modern, highly efficient separation technology. Compared with traditional distillation and rectification techniques, it has advantages such as high separation efficiency, low energy consumption, and small footprint. The core of membrane separation technology is the separation membrane. Polymer porous membranes are a type of separation membrane made from organic polymers according to specific processes. Based on the type of polymer, polymer porous membranes can be further subdivided into cellulose-based polymer porous membranes, polyamide-based polymer porous membranes, sulfone-based polymer porous membranes, polytetrafluoroethylene-based polymer porous membranes, etc. Furthermore, they can also be classified according to pore size into microporous membranes, ultraporous membranes, nanoporous membranes, and reverse osmosis membranes.

[0003] In recent years, in addition to plasma fractionation preparations derived from human blood, there is also a need to improve the safety of biological drugs against viruses. Therefore, pharmaceutical manufacturers have been researching the introduction of virus removal / inactivation processes into the manufacturing process. Among them, the virus removal method using a virus-removing membrane for filtration is an effective method that can reduce the virus without denaturing useful proteins.

[0004] For example, Chinese patent CN1759924B (applied by EMD Millipore) discloses a multilayer composite ultraporous membrane (attached). Figure 14 and 15 The composite ultraporous membrane comprises at least one first porous membrane layer having a first surface and an equivalent second surface, and at least one second porous membrane layer having an equivalent first surface and a second surface. The first and second layers are superimposed and have a porosity transition region from the equivalent first surface of the second layer to the equivalent second surface of the first layer. At least one of the layers is an asymmetric ultraporous membrane. The membrane structure formed by this composite has a strong retention effect on parvoviruses, meeting the needs of practical applications. However, the following problems exist: First, since the porous membrane is a composite membrane, there is a risk of delamination / layer separation during pleating. In addition, the membrane-forming material used to prepare the composite membrane is mainly polyethersulfone. The sulfone groups of polyethersulfone have benzene rings on both sides, which makes its hydrophilicity poor. Ultimately, the membrane has a certain adsorption effect on proteins, resulting in a general protein yield and reduced economic benefits.

[0005] Meanwhile, US Patent US20200238221A1 (Sartorius Corporation application) also discloses a porous monolayer polymer membrane, wherein at least one major surface of the polymer membrane has a surface porosity of at least 40%, and the total porosity of the polymer membrane is 0.8 to 1.4 times that of the at least 40% surface porosity; and the polymer membrane has an asymmetry factor of 1.5 to 10; this polymer membrane is a monolayer porous membrane with good flux and long service life, mainly used for filtering viruses, proteins or macromolecules; however, the average pore size of this polymer membrane is relatively large, and it can only retain large particles with a particle size of several hundred nanometers, and cannot retain small viruses with a particle size of about 20 nm (currently, the typical parvovirus is mouse parvovirus, whose particle size is about 20 nm).

[0006] To further improve protein yield, Chinese patent CN201580007740.0 (application by Asahi Kasei Corporation) also discloses a virus-removing membrane comprising cellulose, used to remove viruses from a protein-containing solution. This virus-removing membrane has a first surface for supplying the protein-containing solution and a second surface for discharging the permeate through the membrane. The average pore size of the membrane is 13nm-21nm, thus easily and efficiently trapping viruses. Simultaneously, the membrane is made of cellulose material, which has strong hydrophilicity, resulting in a significant improvement in protein yield. However, this porous membrane also has certain drawbacks. Firstly, the virus-removing membrane is prepared using the cuprammonium method, which involves adding the film-forming material to a cuprammonium solution for various treatments. The method not only pollutes the environment but is also extremely dangerous, posing a significant threat to the lives of researchers. Secondly, to ensure retention efficiency, the pore size of the virus-removing membrane's separation layer is extremely small (even excessively small), achieving good retention rates for 15nm colloidal gold (the portion capturing 15nm diameter colloidal gold is located between 60% and 90% of the membrane thickness from the first side). This easily leads to a significant increase in protein retention (especially in large-size protein fluids), resulting in a protein yield that still cannot meet practical application requirements (proteins are very expensive, and maximizing protein yield is desirable). In summary, the existence of these problems also limits the development of virus-removing membranes to some extent. Summary of the Invention

[0007] To address the shortcomings of existing technologies, the present invention aims to provide a cellulose porous membrane for virus removal and its preparation method. This cellulose porous membrane is integrally formed without the need for composites, and the preparation process is relatively simple, green, and environmentally friendly. At the same time, the prepared cellulose porous membrane has a strong virus retention effect and can achieve a high protein yield, meeting the needs of practical applications.

[0008] To achieve the above objectives, the present invention provides the following technical solution: a cellulose porous membrane for virus removal, comprising a main body having non-directional tortuous pathways within the main body, one side surface of the main body being a first outer surface, and the other side surface of the main body being a second outer surface; the average pore size of the first outer surface is greater than the average pore size of the second outer surface; the porosity of the porous membrane is 15%-45%;

[0009] In a porous membrane substrate in a wet state, the region that captures colloidal gold with a diameter of 20 nm is designated as D20. D20 is located in the region of 5%-100% of the thickness of the substrate, and the thickness of the D20 region is 15-60 μm.

[0010] The region that captures colloidal gold with a diameter of 30nm is designated as D30. The overlapping region of D30 and D20 is designated as L1. L1 is located on the side of the D20 region closer to the first outer surface. The ratio of the thickness of L1 to the thickness of the D20 region is not less than 0.15.

[0011] The porous membrane of this invention is made of cellulose materials (such as cellulose acetate, regenerated cellulose, etc.). Compared with other organic polymer membrane materials such as PES (polyethersulfone), the porous membrane made of cellulose materials has strong hydrophilicity, good biocompatibility and non-toxicity, and low adsorption capacity for various proteins (it is generally believed that cellulose porous membranes are the least likely organic membrane materials to adsorb proteins). Therefore, it is particularly suitable for use as a virus removal membrane.

[0012] In the membrane structure of the porous cellulose membrane provided by this invention, it can be clearly seen that the pore sizes on the two outer surfaces of the porous membrane are different, with a certain gap; the pore size on one outer surface is larger, while the pore size on the other outer surface is smaller; the outer surface with the larger pore size is referred to as the first outer surface (which is both the macropore surface and the liquid inlet surface) in this invention, while the outer surface with the smaller pore size is referred to as the second outer surface (which is both the micropore surface and the liquid outlet surface); the main body has non-directional tortuous pathways, which refer to randomly oriented groove structures and / or a discretely distributed porous structure, with each non-directional tortuous pathway interconnected, and the fibers forming the porous membrane structure are continuous. It can be understood that "continuous" means that essentially all the fibers are interconnected as a whole, as if formed in one piece, without the use of additional adhesives or the like to connect them together. Unless torn by external force, the network of fibers cannot be separated from each other. At the same time, the continuous network of fibers is also interconnected with the first and second outer surfaces. Such a membrane structure ensures the filtration accuracy of the porous membrane and is beneficial to improving the retention of small viruses by the porous membrane.

[0013] When the membrane porosity is too high, it leads to low tensile strength, poor mechanical properties, and low industrial practical value, failing to meet market demands. Conversely, when the membrane porosity is too low, it affects the membrane flow rate, resulting in slower filtration speed, longer filtration time, and higher time costs. Furthermore, it results in low dirt-holding capacity and a short service life, requiring frequent membrane replacements and significantly increasing economic costs. Therefore, porous membranes require a suitable porosity, which is closely related to the porous membrane material. The porous membrane of this invention is made of cellulose-based materials, with a porosity of 15%-45% (preferably 20-40%). This porosity, combined with the membrane material, results in a membrane that not only has good tensile strength but also a fast filtration speed, high flow rate, and high dirt-holding capacity, capable of retaining a large number of impurity particles, long service life, and low economic cost.

[0014] As is well known, the typical parvovirus is mouse parvovirus, with a particle size of approximately 20 nm. This invention reflects the retention capacity of a porous membrane for various parvoviruses by measuring the location and retention status of 20 nm colloidal gold within the membrane. After the porous membrane retains colloidal gold of a certain particle size, the distribution of the colloidal gold within the membrane can be determined according to Chinese Patent CN105980038B – Test Method for Virus-Removing Membranes: A section is cut from the virus-removing porous membrane after filtering the colloidal gold solution. The brightness distribution of multiple sites in the cross-section stained with colloidal gold is measured using an optical microscope. Since colloidal gold absorbs light, the brightness shift depends on the amount of colloidal gold captured. It should be noted that background noise can be removed by analyzing the brightness distribution as needed. Then, a displacement graph with film thickness on the horizontal axis and brightness on the vertical axis is generated; thereby obtaining the region where colloidal particles of a certain size are trapped in the film thickness direction; (in this invention, the first outer surface is 0% of the film thickness, and the second outer surface is 100% of the film thickness); in addition, gold element determination can be performed on the film cross-section by EDS to understand the distribution of colloidal gold of corresponding particle size in the film cross-section; of course, those skilled in the art can also obtain the distribution of colloidal gold in the film cross-section by other measurement methods, and the above measurement methods are for reference only.

[0015] After testing, it was found that the region D20, where 20nm colloidal gold is trapped, is located within 5%-100% of the body thickness from the first outer surface (i.e., D20 can be 40%-100% or 20%-100% of the body thickness), and the thickness of the D20 region is 15-60μm. The trapping effect of the porous membrane on 20nm colloidal gold shows that, on the one hand, the relatively thick D20 region, combined with the tortuous pathways, ensures a high trapping rate for various small viruses (particle size of 20nm and above), without the risk of virus leakage. On the other hand, compared to patent CN201580007740.0 (applied by Asahi Kasei Corporation), the pores in the small-pore region (virus trapping region) of this invention are relatively large, and the 20nm colloidal gold is trapped at the second outer surface and its vicinity, making further trapping of 15μm impossible. The use of colloidal gold in nm particles prevents the porous membrane from retaining proteins, especially large ones, while further reducing protein adsorption (because smaller pores result in stronger adsorption). This significantly improves protein yield (proteins in viral removal solutions are very expensive, and maximizing protein yield is crucial). Space that would otherwise trap proteins can now be used to trap viruses, further increasing the membrane's load capacity and extending its lifespan. Furthermore, it enhances the membrane's pressure resistance, allowing for higher operating pressures. Currently, Asahi Kasei's highest operating pressure is no more than 15 psi; however, this invention can operate at 30 psi during filtration. Higher operating pressures result in higher flux and shorter operating times. During integrity testing, the operating pressure can reach 50 psi. This increased pressure resistance also simplifies membrane manufacturing and reduces membrane damage.

[0016] To better understand the structure of the porous membrane, in addition to retention experiments using 20nm colloidal gold, retention tests were also conducted using colloidal gold with particle sizes of 30nm. By observing the retention locations of colloidal gold with different particle sizes in the porous membrane, a better understanding of the membrane structure can be obtained. As is well known, colloidal gold with different particle sizes is retained at different locations in porous membranes, and the greater the difference in particle size, the more different the retention locations (for example, the retention locations D100 of 100nm colloidal gold and D20 of 20nm colloidal gold have almost no overlap, while D20 and D30 overlap to a certain thickness). The overlapping region (referred to as L1 in this invention) has been studied and found to be a region where the ratio of L1 to D20 is larger. This indicates that the pore size of the small pore region (the region near the second outer surface, used to trap various small viruses) in the porous membrane changes less with thickness. Conversely, the smaller the ratio of L1 to D20, the greater the pore size of the small pore region (the region near the second outer surface) in the porous membrane changes with thickness. In other words, the membrane pore size changes significantly with thickness, and even the thickness of L1, where D20 and D30 overlap, is very small, or even non-overlapping.

[0017] In this invention, the ratio of the thickness of L1 to the thickness of the D20 region is not less than 0.15, which is relatively high. This indicates that the pore size of the small pore region (the region near the second outer surface) in the porous membrane changes little with thickness, or even almost does not change with thickness. This, combined with the location where D20 is trapped, demonstrates the high efficiency of the porous membrane in trapping viruses, even mouse parvovirus with a particle size of 20 nm, without any risk of virus leakage, and with a long service life. At the same time, the pore size of the small pore region is generally large, and almost no protein is trapped, ensuring that the porous membrane has a high yield. It also has high loading capacity and pressure resistance.

[0018] The porous membrane of this invention has a single-layer asymmetric membrane structure, meaning it is integrally formed without undergoing any "composite" or similar processes. The entire membrane is made from a single cellulose-based material, resulting in a uniform material composition throughout without any material variation. The only change in the membrane's structure is its overall structure. In contrast, composite membranes have multiple layers, and the pore size changes abruptly as one layer transitions to another. Therefore, this porous membrane is less prone to delamination or similar phenomena.

[0019] It should be noted that although there is a certain amount of colloidal gold in some regions along the film thickness direction, it is very low. Therefore, these regions are not considered to be areas where colloidal gold is trapped. These regions only have some residual colloidal gold (rather than trapped). Therefore, in the virus-removing membrane, it is preferable to form a continuous area along the film thickness direction that captures colloidal gold with a diameter of 20 nm, which is the region where colloidal gold of the corresponding particle size is truly trapped.

[0020] Here, in the film thickness direction, a first distance a is measured from the first outer surface of the virus-free porous membrane to the portion of the colloidal gold capture site closest to the first outer surface;

[0021] In addition, in the membrane thickness direction, a second distance b is measured from the first outer surface of the virus-free porous membrane to the portion of the colloidal gold capture site closest to the second porous surface; then, the first distance a divided by the membrane thickness c of the virus-free membrane is calculated at multiple sites, and the value A (expressed as a percentage of a / c) is calculated, and the average value of the value A at multiple sites is used as the first reach.

[0022] Furthermore, the second distance b divided by the membrane thickness c of the virus-free membrane is calculated at multiple sites, and the value B is expressed as a percentage (expressed as a percentage of b / c). The average value of the values ​​B at multiple sites is used as the second reachability. The area where colloidal gold with a diameter of 20 nm is retained is A20-B20 (i.e., the D20 region described in this invention, so the thickness of the D20 region = (B20-A20) * membrane thickness c); the area where colloidal gold with a diameter of 30 nm is retained is A30-B30 (i.e., the D30 region described in this invention).

[0023] As a further improvement of the present invention, D30 is located in the region of 5%-90% of the thickness of the main body; the thickness of L1 is 5-50 μm, and L1 is located in the region of 5%-90% of the thickness of the main body; the ratio of the thickness of L1 to the thickness of the D20 region is not less than 0.3.

[0024] Retention experiments using 30nm colloidal gold revealed that D30 is located within a region of 5%-90% of the main body thickness (i.e., D30 can be a region of 5%-70% or 20-90% of the main body thickness). Understanding the location of D30 within the porous membrane further demonstrates that this porous membrane has a high retention rate for various small viruses, and the virus removal area is mainly near the second outer surface, where the pore size is relatively small. Simultaneously, the combined effect of D20 within the membrane ensures efficient retention of 20nm viruses, while also exhibiting good flux and a long lifespan, with the membrane pores not easily becoming clogged prematurely (i.e., flux decay is too rapid).

[0025] Furthermore, it can be seen that the thickness of L1, the overlapping region of D20 and D30, is 5-50 μm, which is relatively thick. This further indicates that in the main cross-section of the porous membrane, the small pore region near the second outer surface can fully and completely capture viruses of various diameters without any risk of leakage. Moreover, L1 is located in the region of 5%-90% of the main body thickness from the first outer surface (i.e., L1 can be in the region of 70%-90% of the main body thickness or 40-80% of the main body thickness). This further indicates that in the structure of the membrane body, the region near and including the second outer surface is a small pore region with a suitable thickness to ensure the efficient capture of viruses by the porous membrane. In the large pore region near and including the first outer surface, a pre-filtration function is performed to intercept large particulate impurities in the fluid, giving the porous membrane a large dirt holding capacity and a fast flow rate. At the same time, it protects the small pore region, ensuring that the membrane as a whole has high mechanical strength and good pressure resistance.

[0026] As a further improvement of the present invention, D20 is located in a region of 15%-100% of the thickness of the main body, and the thickness of the D20 region is 20-50 μm; the ratio of the thickness of L1 to the thickness of the D20 region is 0.4-0.9.

[0027] Research has shown that when the 20nm colloidal gold is more effectively retained within the membrane and has an appropriate thickness, the membrane is more likely to achieve high retention efficiency and high throughput. Simultaneously, a larger overlap region L1 with a greater ratio to D20 can further improve protein yield and enhance the membrane's pressure resistance. Furthermore, due to the thicker D20 layer, minor defects will not significantly impact the membrane's retention efficiency, allowing for sustained high retention efficiency (when the retention area is small, even a tiny defect can be magnified hundreds or thousands of times, leading to a risk of virus leakage during actual use).

[0028] As a further improvement of the present invention, the capture peak of colloidal gold with a diameter of 20 nm is located in the region of 75-92% of the body thickness from the first outer surface, and the distance between the peak location and the second outer surface is 3-12 μm.

[0029] Tests revealed that the area where 20nm colloidal gold is trapped is mainly located near the second porous surface. The capture peak is the area where the most colloidal gold of the corresponding particle size is trapped. If the capture peak is too close to the second outer surface (or even on the second outer surface), there is a risk of virus leakage, meaning the LRV of the porous membrane for various viruses is too low and cannot meet the actual needs. If the capture peak is too far from the second outer surface, it indicates that the 20nm colloidal gold is trapped in the membrane too early, leaving many areas unused and the overall pore size of the membrane too small. This would significantly affect the flux of the porous membrane, resulting in low flux and loading capacity. In this invention, the capture peak for trapping 20nm colloidal gold is located in the region from the first outer surface at 75%-92% of the main body thickness, and the distance between this peak and the second outer surface is 3-12μm. This structure ensures that the porous membrane can efficiently trap various small viruses while maintaining high flux and loading capacity.

[0030] After cutting a section from the virus-free porous membrane after filtering a 20nm colloidal gold solution, the section was measured using an optical microscope. The darkest part of the section was found to be the capture peak. Alternatively, after testing with an EDS spectrometer, the location of the peak was found to be the capture peak.

[0031] As a further improvement of the present invention, the porous membrane is used to capture 20 nm colloidal gold, and the porous membrane containing the captured 20 nm colloidal gold is top-washed. After top-washing, in the cross section of the virus-free membrane, the distance between the peak portion of the captured 20 nm colloidal gold and the second outer surface is 1-8 μm, and the ratio of this distance to the D20 thickness is 3%-20%.

[0032] To achieve higher protein yields, whether in laboratory validation or actual production processes in biopharmaceutical companies, a top-wash operation (typically at 30 psi) is generally required after virus filtration using a membrane filter (or porous membrane). This is because some protein remains trapped in the porous membrane during virus filtration, and since proteins are very expensive, a top-wash operation is necessary to remove them from the membrane to further improve protein yield. Taking laboratory validation as an example (although the actual production process may differ, the mechanism is the same and can be characterized through laboratory validation), the top-wash operation typically occurs during continuous filtration. When the flow rate of the membrane filter (or porous membrane) decreases to 75% or only a small amount of feed remains, the external pressure is removed and the process is paused for a period of time (e.g., 5 to 15 minutes) to release the protein from the pore structure. Subsequently, a liquid such as a buffer solution is added to elute the released protein from the porous membrane, thereby improving protein yield.

[0033] However, while top washing can indeed improve protein yield, during the removal of external pressure, viruses with stronger motility are more likely to be released from the porous structure and move to the effluent surface of the porous membrane. During top washing, these released and moved viruses are easily eluted, resulting in a virus concentration in the top wash solution that is often much higher than that in the filtrate obtained during retention filtration. Therefore, the top wash solution usually needs to be processed separately. However, if the virus concentration in the top wash solution is too high (LRV < 4), various purification processes are required, which are cumbersome, time-consuming, and labor-intensive. Furthermore, it is even considered that the integrity of the porous membrane is compromised.

[0034] After testing, the distance between the peak of the 20nm colloidal gold capture site and the second outer surface in the cross-section of the virus-removing membrane after top washing was 1-8μm. This distance ensures a very low virus concentration in the top washing solution, with an LRV value above 4, while also maintaining high throughput and a short top washing time. Furthermore, the ratio of the distance between the peak and the second outer surface to the D20 thickness was 3%-20%, further demonstrating that after top washing, the porous membrane can still fully and completely retain the virus without prematurely retaining it, thus ensuring the throughput and load capacity of the porous membrane.

[0035] As a further improvement of the present invention, before and after top washing, the displacement of the peak position of the captured 20nm colloidal gold in the porous membrane is 0.5-4μm; and this displacement accounts for 2-12% of the D20 region.

[0036] Before and after top washing, the peak capture site of 20nm colloidal gold may shift downward in the direction of the virus membrane thickness (based on the flow direction of the feed solution, the first outer surface of the virus membrane is considered the upper end, and the second outer surface of the virus membrane is considered the lower end; downward shift means moving towards the second outer surface). Generally, it is considered that the smaller the distance (change in displacement) the peak capture site of 20nm colloidal gold moves downward before and after top washing, the better, so that the LRV of top washing decreases less, and the treatment cost of top washing solution will naturally decrease.

[0037] However, the inventors of this application have discovered that a smaller displacement variation is not always better. While a smaller displacement variation often means a higher wash LRV compared to the initial LRV, it also often indicates that the pore size in the micropore region near the second outer surface is too small, or that the average pore size in the micropore region changes rapidly along the membrane thickness direction (other conditions that can affect the downward movement of 20nm colloidal gold can also produce this result). Regardless of the reason, this can hinder the downward movement of 20nm colloidal gold, which often also affects the downward movement of the feed solution and protein, thus affecting throughput or protein yield. Therefore, a smaller displacement variation is not always better. Studies have shown that a displacement variation of 0.5-4μm, and this displacement variation accounting for 2-12% of the D20 region, not only ensures high protein yield and wash LRV, but also ensures high throughput of the virus removal membrane.

[0038] As a further improvement of the present invention, the region for capturing colloidal gold with a diameter of 40 nm is D40, and D40 is located in the region of 5%-80% of the thickness of the main body; the region where D40 and D30 overlap is L2; ​​the ratio of the thickness of L1 to the thickness of L2 is 1.2-2.5:1; the ratio of the thickness of the main body region for capturing colloidal gold with a diameter of 20-40 nm to the thickness of the main body region for capturing colloidal gold with a diameter of 20-30 nm is 1.1-1.8.

[0039] Research has shown that parvoviruses are generally various viruses with a particle size of 20-40 nm; that is, the virus membrane mainly removes viruses with a particle size of 20-40 nm. Therefore, a 40 nm colloidal gold interception test was also conducted. The test revealed that D40 is located in the region from the first outer surface at 5%-80% of the body thickness (i.e., D40 can be located at 5-60% or 20%-80% of the body thickness). However, the interception position of D40 cannot be very close to or even reach the second outer surface (i.e., reach 100%), because in this case, the porous membrane cannot efficiently capture 20 nm mouse parvoviruses.

[0040] By observing the overlapping regions of colloidal gold particles of different sizes within the membrane where they are trapped, we can better understand the membrane pore structure (the variation of membrane pore size with thickness). We set the overlapping region of D40 and D30 as L2. Although both have a particle size difference of 10nm, the thickness of L2 is less than that of L1. This structural setting ensures more sufficient trapping of smaller viruses, ensuring efficient trapping of 20nm viruses. Furthermore, we found that the ratio of L1 thickness to L2 thickness is 1.2-2.5:1, meaning that the thickness of L2 cannot be too small. This ensures that viruses with a particle size of 30-40nm are trapped at a reasonable position on the membrane, neither trapping them too early, resulting in low membrane flux, nor trapping them too late (20-40nm viruses are basically trapped at the same position on the membrane with almost no distinction), which would cause the porous membrane to rapidly decrease flux, resulting in low load and short service life during virus trapping and filtration.

[0041] The region capturing colloidal gold with a diameter of 20nm is A20-B20; the region capturing colloidal gold with a diameter of 30nm is A30-B30; the region capturing colloidal gold with a diameter of 40nm is A40-B40; therefore, the region capturing colloidal gold with a diameter of 20-30nm is A30-B20 from the first outer surface, which is the thickness of the main body, and the thickness of this region is (B20-A30)*c. Note that if the thicknesses of the two films are different, c is the average of the thicknesses of the two films; therefore, the region capturing colloidal gold with a diameter of 20-40nm is A40-B20 from the first outer surface, which is the thickness of this region, and the thickness of this region is (B20-A40)*c.

[0042] Testing revealed that the thickness ratio of the main region capturing colloidal gold with a diameter of 20-40 nm to the main region capturing colloidal gold with a diameter of 20-30 nm in this invention is 1.1-1.8. Further investigation revealed that the portion retaining 20-30 nm colloidal gold in the pore region of the membrane is relatively thick. The thickness of this portion significantly affects the virus retention efficiency and the overall membrane flux. If the thickness of this portion is too small, the overall membrane retention efficiency for various small viruses will be too low to meet the needs of practical applications; if the thickness of this portion is too large, it will greatly reduce the membrane flux, resulting in a lower membrane filtration rate and reduced economic efficiency per unit time; it will also reduce protein yield to some extent. The goal is to achieve high virus retention efficiency while maximizing protein yield and membrane flux.

[0043] As a further improvement of the present invention, the first outer surface includes a plurality of strip-shaped continuous first fibers, and adjacent continuous first fibers surround each other to form circular holes; the average diameter of the first fibers is 60-600 nm; and the area ratio of the holes on the first outer surface is 5-35%.

[0044] In the membrane structure of the porous membrane provided by this invention, it can be clearly seen that there are several first fibers on the first outer surface (macropore surface) of the membrane. The first fibers are elongated structures, and the pores on the first outer surface are formed by the first fibers surrounding each other. The average diameter of the first fibers is 60-600 μm. This thickness of the first fibers ensures the stability of the pores. Under high pressure of 30 psi or even 50 psi, the pores are not easy to collapse, ensuring that the porous membrane can filter fluids for a long time. At the same time, it performs a certain pre-filtration effect on the fluid, intercepting large particulate impurities. It also ensures the mechanical strength of the membrane, meets the needs of practical applications, and is suitable for various processing.

[0045] Furthermore, a certain number of pores exist on the first outer surface of the membrane. As is well known, the number and shape of membrane pores have a significant impact on the filtration accuracy (retention efficiency) and membrane flux of the membrane. In this invention, the pores on the first outer surface are circular, with some pores being circular and others being elliptical. This shape of pore facilitates fluid filtration. In addition, the pore area ratio on the first outer surface is closely related to the thickness of the first fiber. When the diameter of the first fiber remains basically constant, an excessively large pore area ratio can lead to deformation or even collapse of the membrane pores, making it impossible to achieve efficient retention for a long time. An excessively low pore area ratio will result in a low membrane flux. In this invention, the pore area ratio on the first outer surface is 5-35% (the ratio of the sum of the areas of the first pores to the membrane area). This pore area ratio, combined with a suitable thickness of the first fiber, is beneficial for the porous membrane to have a high flow rate, facilitating the rapid passage of fluid through the porous membrane, shortening the filtration time, and also possessing high tensile strength and compressive strength, meeting the needs of practical applications.

[0046] The average diameter of the first fiber can be measured by characterizing the membrane structure using a scanning electron microscope, followed by measurement and calculation using computer software (such as Matlab, NIS-Elements, etc.) or manually; a certain area is selected, for example, 1 μm. 2 (1μm x 1μm) or 25μm 2 (5μm x 5μm), the specific area size depends on the actual situation. Then, the diameter of the first fiber on this area is measured by the corresponding computer software or manually, and then the average value is calculated. The average diameter of the first fiber on the first outer surface is represented by the average diameter of the first fiber in this area. The porosity is obtained by summing the areas of all pores in this area and then dividing by the corresponding area. Of course, those skilled in the art can also obtain the above parameters by other measurement methods. The above measurement methods are for reference only.

[0047] Note: Cellulose membranes are generally used under humid conditions. After drying, the membrane pores are prone to shrinkage and the thickness is also prone to change. Therefore, the relevant parameters in this invention are all measured when the membrane is humid, or the membrane is freeze-dried and then photographed under SEM to obtain the corresponding parameters.

[0048] As a further improvement of the present invention, the thickness of the porous body is 25-70 μm, the average pore diameter of the first outer surface is 150-800 nm, the ratio of the average pore diameter of the first outer surface to that of the second outer surface is 6-30, and the ratio of the average pore diameter of the first outer surface to the average diameter of the first fiber is 2-8.

[0049] The porous membrane prepared by this invention has a relatively thin portion with a thickness of 25-70 μm. This relatively small thickness, combined with the synergistic effect of an average pore size of 150-800 nm on the first outer surface, facilitates a higher flux. Furthermore, the combined effect of this membrane thickness, the average pore size on the first outer surface, and the porosity of the first outer surface allows the porous membrane to effectively pre-filter large particles, reducing their impact on the small pore areas and enabling these areas to better trap viruses. Moreover, the combination of the relatively thin overall membrane thickness and the thicker small pore areas ensures that the porous membrane, while maintaining a high flux, also exhibits high initial LRV and top-wash LRV.

[0050] Furthermore, the ratio of the average pore size of the first outer surface to that of the second outer surface is 6-30, indicating that the overall pore size variation of the membrane will not be too large, which is more conducive to ensuring the mechanical strength of the membrane. At the same time, by setting the ratio of the average pore size of the first outer surface to the average diameter of the first fiber to 2-8, that is, the first fiber is relatively thick, the stability of the first pore can be ensured, and it is not easy to shrink or collapse. Even under high pressure (30psi, or even 50psi), the flux of the porous membrane remains stable, which can retain viruses quickly and efficiently for a long time.

[0051] The thickness of the membrane can be obtained by characterizing the morphology of the membrane structure using a scanning electron microscope, and then by measuring it using computer software (such as Matlab, NIS-Elements, etc.) or manually. Of course, those skilled in the art can also obtain the above parameters by other measurement methods (such as freeze-drying first, and then measuring with measuring tools). The above measurement methods are for reference only.

[0052] The average pore size of the membrane surface can be measured by characterizing the membrane structure using a scanning electron microscope and then selecting a certain area, such as 1 μm. 2 (1μm x 1μm) or 25μm 2(5μm by 5μm), the specific area size depends on the actual situation. Then, use appropriate computer software or manual measurement to determine the diameter of all holes on this area, and then calculate to obtain the average pore diameter of the surface. Of course, those skilled in the art can also obtain the above parameters through other measurement methods. The above measurement methods are for reference only.

[0053] As a further improvement of the present invention, the thickness of the main body region for capturing colloidal gold with a diameter greater than 40 nm is 3-25 μm, and it is located in the region from the first outer surface within 0-40% of the main body thickness.

[0054] In the host region for capturing colloidal gold with a diameter greater than 40 nm, the pore size change rate near the first outer surface is greater than that near the second outer surface.

[0055] In the field of virus removal, substances with a particle size greater than 40 nm are often considered large particulate impurities. Under normal circumstances, the macroporous region of the membrane (the region near the first outer surface) will remove these large particulate impurities. However, research has found that in these thin porous membranes, the macroporous region is relatively thin (the thickness of the macroporous region accounts for a small proportion of the overall membrane thickness). In this invention, the thickness of the main region for capturing colloidal gold with a diameter greater than 40 nm is 3-25 μm, and it is located in the region of 0-40% of the main thickness from the first outer surface. This effectively removes various large particulate impurities while making the microporous region thicker and accounting for a higher proportion, thereby improving the virus removal capacity of the virus removal membrane. As a result, the thin virus removal membrane has high throughput and high protein yield, while still having high initial LRV and top wash LRV.

[0056] Furthermore, we found that in the main region where colloidal gold with a diameter greater than 40 nm is captured, the pore size change gradient near the first outer surface is greater than that near the second outer surface. Such a membrane structure is beneficial to improving the overall porosity of the membrane. Although the pore region is relatively thick, the membrane still has a high flux due to the combined effect of the overall thin membrane thickness and the change in pore size with thickness.

[0057] In this invention, the average value of the corresponding colloidal gold capture area is used to represent the entire area of ​​colloidal gold capture; that is, C40 is used to represent the entire area of ​​40nm colloidal gold captured in the main structure of the film, and C40 = (A40 + B40) / 2; the main area of ​​colloidal gold with a capture diameter greater than 40nm is 0% - C40, and the thickness is C40*c.

[0058] As a further improvement of the present invention, the thickness of the porous membrane is 60-120 μm, the average pore size of the first outer surface is 400-2000 nm, and the ratio of the average pore size of the first outer surface to the second outer surface is 20-100.

[0059] In the porous membranes prepared by this invention, some membranes have a relatively large thickness, ranging from 60 to 120 μm. Due to this large thickness, the membrane flux is likely to be low. In order to ensure that the porous membrane still has a high flux, the average pore size of the first outer surface in this invention is relatively large, ranging from 400 to 2000 nm. That is, the pore size on the macropore surface (liquid inlet surface) is relatively large, which is beneficial to improving the flux of the porous membrane.

[0060] Meanwhile, the combined effect of such membrane thickness, average pore size of the first outer surface, and pore area ratio of the first outer surface enables the porous membrane to perform excellent pre-filtration of large particles, greatly increasing its dirt-holding capacity, making it particularly suitable for filtering fluids containing a large number of large particulate impurities. In addition, the ratio of the average pore size of the first outer surface to that of the second outer surface is 20-100, indicating that there is a certain variation in the overall pore size of the membrane. That is, although the pore size in the macropore region (the region near the first outer surface) is larger, the pore size in the micropore region is still smaller, ensuring that the porous membrane has a high initial LRV and top wash LRV. Furthermore, the combined effect of the relatively thick overall membrane thickness, the appropriate thickness of the micropore region, and the appropriate thickness of the macropore region enables the porous membrane to have high flux and high mechanical strength.

[0061] As a further improvement of the present invention, the thickness of the main region for capturing colloidal gold with a diameter greater than 40 nm is 35-100 μm, and it is located in the region of 0%-75% of the main thickness from the first outer surface.

[0062] Research has revealed that in these thicker porous membranes, the macropore region is relatively thick (the macropore region accounts for a large proportion of the overall membrane thickness). In this invention, the thickness of the main region capturing colloidal gold with a diameter greater than 40 nm is 35-100 μm, and it is located in the region from the first outer surface within 0-75% of the main thickness. This pore structure gives the porous membrane a high porosity, resulting in a large membrane flux and fast filtration speed despite the large membrane thickness. Simultaneously, the porous membrane also has a large dirt-holding capacity, long service life, and slow flux decay. Although the macroporous region is relatively thick, the large pore size and the fact that the porous membrane is a cellulose membrane (highly hydrophilic) mean that almost no protein is adsorbed in the macroporous region, resulting in a still high protein yield. However, in the Chinese patent CN201580007740.0 (application by Asahi Kasei Corporation), the thickness of the macroporous region (the area with an average pore size greater than 40 nm in the membrane cross-section is called the pre-filtration layer) is very small, which easily leads to a lower virus membrane contaminant holding capacity. Large particulate impurities can easily clog the internal flow channels of the membrane, resulting in a short service life and low viral load.

[0063] As a further improvement of the present invention, the thickness of the main region for capturing colloidal gold with a diameter of 60-80 nm is 0.5-4 μm greater than the thickness of the main region for capturing colloidal gold with a diameter of 80-100 nm.

[0064] The thickness of the host region for capturing colloidal gold with a diameter of 40-60 nm is 1-7 μm greater than that for capturing colloidal gold with a diameter of 60-80 nm.

[0065] The thickness of the host region for capturing colloidal gold with a diameter of 20-40 nm is 2-12 μm greater than that for capturing colloidal gold with a diameter of 80-100 nm.

[0066] Research revealed that larger colloidal gold particles are more likely to be trapped closer to the first outer surface, and the thickness of this trapped area is relatively small. Furthermore, a suitable thickness difference exists, indicating that in the macroporous region of this invention, the membrane pore size changes at a relatively decelerated rate; that is, the closer to the second outer surface in the macroporous region, the smaller the change in membrane pore size with thickness. This membrane structure is beneficial for the porous membrane to have a large overall flux and fast filtration speed. Simultaneously, the porous membrane also has a large dirt-holding capacity, long service life, and slow flux decay. The membrane as a whole also possesses high mechanical properties (high tensile strength and elastic modulus), with the macroporous region providing sufficient support for the microporous region. In addition, the microporous region has a relatively high thickness (although not large in proportion to the overall membrane thickness), ensuring sufficient capture of various small viruses without the risk of leakage.

[0067] As a further improvement of the present invention, the thickness of the main region for capturing colloidal gold with a diameter of 80-100nm is 2-16μm; the thickness of the main region for capturing colloidal gold with a diameter of 60-80nm is 3-20μm.

[0068] The thickness of the host region for capturing colloidal gold with a diameter of 40-60 nm is 4.5-23 μm.

[0069] In these thicker porous membranes, we found that the macropore region was also quite thick. To better understand the membrane structure, especially the variation of membrane pore size with thickness in the macropore region, we conducted retention experiments on colloidal gold with different particle sizes (40nm, 60nm, 80nm, 100nm, etc.). After the experiments, we found that the membrane regions retaining colloidal gold of corresponding particle sizes all had suitable thicknesses. This thickness ensures that impurities of the corresponding particle size in the fluid can be sufficiently retained, being retained at appropriate locations within the membrane. This prevents premature retention, which would result in low membrane flux, and also prevents premature retention, which would affect the efficient retention of small viruses in the fluid in the subsequent micropore region. This ensures a long membrane lifespan, with both high initial LRV and high LRV after top washing.

[0070] The region capturing colloidal gold with a diameter of 100nm is A100-B100; the region capturing colloidal gold with a diameter of 80nm is A80-B80; the region capturing colloidal gold with a diameter of 60nm is A60-B60; the region capturing colloidal gold with a diameter of 40nm is A40-B40; then the region capturing colloidal gold with a particle size of 80-100nm is A100-B80 from the first outer surface at the thickness of the main body, and the thickness of this region is (B80-A100)*c.

[0071] The portion capturing colloidal gold particles with a diameter of 60-80 nm is located at A80-B60 of the body thickness from the first outer surface, with a thickness of (B60-A80)*c. Similarly, the portion capturing colloidal gold particles with a diameter of 40-60 nm is located at A60-B40 of the body thickness from the first outer surface, with a thickness of (B40-A60)*c. As a further improvement of the invention, when the diameter of the captured colloidal gold is 20-40 nm, the capturing gradient K1 along the thickness direction of the body is 0.5-3.5 nm / μm; when the diameter of the captured colloidal gold is 40-100 nm, the capturing gradient K2 along the thickness direction of the body is 1-6 nm / μm.

[0072] Gradient capture = Change in diameter of colloidal gold captured / Thickness of the main region of colloidal gold with the corresponding diameter captured.

[0073] When the diameter of the captured colloidal gold is 20-40 nm, the region capturing 40 nm diameter colloidal gold is designated A40-B40; the region capturing 20 nm diameter colloidal gold is designated A20-B20. When calculating the average gradient of the captured colloidal gold diameter, this invention uses the average value of the corresponding colloidal gold capture region to represent the entire region of colloidal gold capture; that is, C40 represents the entire region of 40 nm colloidal gold captured in the main film structure, where C40 = (A40 + B40) / 2. C20 represents the entire region in the main structure where 20nm colloidal gold is captured, and C20 = (A20 + B20) / 2; then the change in the diameter of the colloidal gold is 40nm - 20nm = 20nm; the thickness of the main body for capturing the corresponding colloidal gold diameter is: C20 - 40 = (C20 - C40) * c; when the diameter of the captured colloidal gold is 20-40nm, the average gradient of the captured colloidal gold diameter along the thickness direction is K1 = 20nm / C20 - 40μm;

[0074] In this invention, the main structure of the porous membrane is divided into two regions. The region with relatively large pore sizes mainly serves a pre-filtration function, while the region with relatively small pore sizes mainly serves to trap small viruses. The porous membrane of this invention is integrally formed, and the pore size gradually changes with the overall thickness of the membrane (within a certain region, the pore size may not change significantly with membrane thickness), without abrupt changes. K2 refers to the macropore region of the thicker porous membrane, where the pore size changes rapidly with thickness, but not too rapidly, because excessively rapid changes would result in excessively large pores in the porous membrane, which would then... This results in low overall mechanical strength of the membrane, making it susceptible to pressure damage. Furthermore, if the pore size in the macropore region changes too slowly with thickness, it can lead to low retention efficiency or low flux, failing to meet the needs of practical applications. In contrast, the pore size in the micropore region changes very little or not at all with thickness, and the dense second outer surface (micropore surface) allows for efficient retention of various parvoviruses (especially mouse parvovirus with a particle size of approximately 20 nm). Parvovirus leakage is less likely, and the LRV after top washing is greater than 4, achieving both high protein yield and high retention efficiency.

[0075] As a further improvement of the present invention, the ratio of K2 to K1 is 1.1-2.5.

[0076] Wherein K1 represents the change of membrane pore size with thickness in the micropore region, and K2 represents the change of membrane pore size with thickness in the macropore region. The study found that in thicker porous membranes, K2 and K1 need to have an appropriate ratio, which is 1.1-2.5. Such a ratio can ensure that the membrane has a suitable average pore size, which can achieve high retention efficiency for various small viruses (especially mouse parvovirus with a particle size of about 20 nm), with high initial LRV and high LRV after top washing, and no risk of virus leakage. At the same time, the membrane has high overall flux, fast filtration speed, high mechanical properties and good elasticity, and large dirt holding capacity.

[0077] As a further improvement of the present invention, the pressure resistance of the cellulose porous membrane is not less than 30 psi; the flux of the cellulose porous membrane is greater than 40 L*h. -1 *m -2 @30psi; the cellulose porous membrane has an LRV greater than 4 for 20nm colloidal gold; the protein yield of the cellulose porous membrane is not less than 98%.

[0078] The porous membrane of this invention does not collapse or shrink under a pressure of 30 psi, thus ensuring efficient retention of small viruses in various protein fluids. In addition, the porous membrane of this invention has a wet tensile strength of 2-10 MPa and an elongation at break of 5-40%, indicating that the porous membrane of this invention has high tensile strength and elongation at break, good mechanical properties, high industrial practical value, and can fully meet market demand.

[0079] Permeation flux, also known as permeation rate or simply flux, refers to the amount of substance that passes through a unit area of ​​a porous membrane per unit time under a certain operating pressure during the separation process. The magnitude of the flux reflects the speed of filtration; the higher the flux, the faster the membrane's filtration speed. In this invention, the flux of the porous membrane is greater than 40 L*h. -1 *m -2 @30psi indicates a relatively high flux, suggesting that the porous membrane has a fast filtration speed. While ensuring retention efficiency, the fluid can pass through the porous membrane quickly, resulting in lower time costs and higher economic benefits.

[0080] The viruses intercepted by this invention are mainly various viruses with a particle size of 20 nm and above (such as mouse parvovirus, whose particle size is about 20 nm). Therefore, 20 nm colloidal gold was intercepted. After interception tests, it was found that the LRV of the porous membrane of this invention for 20 nm colloidal gold was greater than 4, which shows that the porous membrane has a very high rejection rate for various viruses and bacteria, and plays a sufficient role in retaining viral impurities, meeting the needs of practical applications. In addition, the protein yield of the porous membrane is not less than 98%, which shows that the effective substance protein in the fluid is not easily adsorbed on the membrane. On the one hand, it will not block the membrane pores, ensuring that the porous membrane still has a high service life. On the other hand, it ensures that the content of effective substance protein in the fluid changes very little, and the protein is basically not lost, thus ensuring economic benefits. The test method for viral impurities can refer to patents CN105980037B-Virus Removal Membrane, CN101816898B-Ultraporous Membrane and Preparation Method Thereof, CN1759924B-Ultraporous Membrane and Preparation Method Thereof, etc.

[0081] On the other hand, the present invention also provides a method for preparing a cellulose porous membrane for virus removal, comprising the following steps:

[0082] S1: Prepare a casting solution and cast it onto a carrier to form a liquid film; the casting solution comprises the following components by weight: 10-30 parts of film-forming polymer; 30-60 parts of good solvent; 20-40 parts of pore-forming agent;

[0083] The mass of the good solvent in the casting solution is 1.1-2.5 times the mass of the pore-forming agent;

[0084] The film-forming polymer is a cellulose polymer; the pore-forming agent is a non-good solvent substance with a surface tension of less than 30 mN / m, and the substance is miscible with the coagulation bath;

[0085] S2: Pretreatment: The liquid film is placed in a high-concentration organic solvent vapor environment for pretreatment for 2-10 seconds to obtain a raw film; wherein the high-concentration organic solvent vapor refers to the ratio of the actual organic solvent vapor pressure in the environment to the saturated organic solvent vapor pressure at the same temperature being not less than 80%; the temperature of the organic solvent vapor is 2-8°C higher than the temperature of the casting solution, and the flow velocity of the organic solvent vapor is 0.3m / s-0.9m / s;

[0086] S3: Immerse the film-forming material in a coagulation bath for phase separation and solidification. The temperature of the coagulation bath is 5-15°C lower than the temperature of the casting solution, and the duration is 30-80 seconds to obtain the film. The coagulation bath is water.

[0087] As a further improvement of the present invention, the casting solution temperature is 15-40°C; the cellulose polymer is at least one of nitrocellulose and cellulose acetate; the pore-forming agent is any one of ethanol, 1-propanol, isopropanol, n-butanol, 1-pentanol, and diethyl ether; the good solvent is at least one of acetone, dioxane, dimethylacetamide, N-methylpyrrolidone, acetic acid, propionic acid, butyric acid, and valeric acid; and the organic solvent in the organic solvent vapor is at least one of acetone, dioxane, dimethylacetamide, N-methylpyrrolidone, acetic acid, propionic acid, butyric acid, and valeric acid.

[0088] As a further improvement of the present invention, the film is placed in an aqueous sodium hydroxide solution for hydrolysis at a temperature of 30-60°C for a time of 40-60 min; after hydrolysis, the film is washed to obtain a solid film; wherein 0.1-1 wt% of an anti-shrinkage agent is added to the sodium hydroxide solution, and the anti-shrinkage agent is a mixture of glycerol and butyl orthosilicate in a mass ratio of 3:1.

[0089] As a further improvement of the present invention, the hydrolyzed solid membrane is placed in an alkaline environment and crosslinked with a water-soluble crosslinking agent. After the crosslinking is completed, the membrane is washed to obtain the finished product membrane. The crosslinking agent is at least one of halogenated epoxide, dihalogenated alkane and dihalogenated alcohol. The crosslinking time is 20-400 min and the temperature is 30℃-60℃.

[0090] In preparing the cellulose porous membrane of the present invention, a casting solution is first prepared, which includes a film-forming polymer, a good solvent, and a pore-forming agent; wherein the film-forming polymer is a cellulose polymer, preferably at least one of nitrocellulose and cellulose acetate (including cellulose diacetate and cellulose triacetate); these cellulose polymers are readily soluble in their respective good solvents and have good film-forming and fiber-forming properties; the resulting membrane has strong hydrophilicity and does not easily adsorb effective substances (mainly proteins) in the fluid; the presence of the good solvent is used to fully dissolve the corresponding cellulose polymer, and during phase separation, the good solvent is dissolved in the coagulation bath, thereby causing the cellulose polymer to precipitate and form a porous membrane with ideal pore size and porosity; currently commonly used good solvents are at least one of acetone, dioxane, dimethylacetamide, N-methylpyrrolidone, acetic acid, propionic acid, butyric acid, and valeric acid;

[0091] In addition, a pore-forming agent is added to the casting solution. This pore-forming agent is a low-surface-tension non-solvent substance (a non-good solvent substance with a surface tension below 30 mN / m, and miscible with the coagulation bath; commonly used are any one of ethanol, 1-propanol, isopropanol, n-butanol, 1-pentanol, and diethyl ether). The addition of these low-surface-tension non-good solvent substances allows for rapid miscibility with the coagulation bath. With the assistance of these low-surface-tension substances, after proper pretreatment, the coagulation bath can quickly enter the liquid membrane near the carrier side, causing rapid phase separation in the region near the carrier side, forming corresponding small pore areas—areas used to capture small viruses. These small pore areas are relatively thick, thus achieving the requirement of efficient virus retention. Meanwhile, the liquid membrane away from the carrier side (i.e., the air side) and its vicinity form corresponding macropore areas. These macropore areas are used for pre-filtration of the fluid, improving the lifespan of the porous membrane. The diameter and thickness are closely related to the formulation of the casting solution and the pretreatment process. Furthermore, the study found that the content of the pore-forming agent in the casting solution needs to be strictly controlled. Too much pore-forming agent can negatively impact the porosity of the porous membrane, making it impossible to obtain a high-porosity membrane. Too little pore-forming agent prevents the coagulation bath from quickly penetrating the region near the carrier side of the liquid membrane to achieve rapid phase separation, thus resulting in a thicker pore area. In this invention, the mass of the organic solvent in the casting solution is generally controlled to be 1.1-2.5 times the mass of the pore-forming agent. This allows the pore-forming agent to be uniformly dispersed in the casting solution. Combined with suitable pretreatment conditions and phase separation and solidification in the coagulation bath, an ideal membrane size and pore structure are obtained (the pore area is thicker, thus exhibiting high retention efficiency for 20nm viruses). Simultaneously, this porous membrane has high porosity, indicating high flux and high protein yield.

[0092] After the casting solution is prepared, it is cast onto a carrier to form a liquid film. The liquid film is then pretreated. Through proper pretreatment and appropriate phase separation, relatively large pores appear on the air side of the liquid film (which subsequently forms the first outer surface). The specific pore size depends on the casting solution formulation and pretreatment conditions. The presence of these pores facilitates the coagulation bath's penetration into the liquid film near the carrier, resulting in more complete phase separation. This leads to smaller pore size variations along the film thickness in the pore region and a relatively larger thickness in the pore region, thus improving virus retention efficiency. Furthermore, the final porous membrane has higher porosity, resulting in higher flux.

[0093] The pretreatment of this invention is carried out in a high-concentration organic vapor environment, where high-concentration organic solvent vapor refers to an environment where the ratio of the actual organic solvent vapor pressure to the saturated organic solvent vapor pressure at the same temperature is not less than 80%. By controlling the concentration and wind speed of the organic solvent vapor in the environment, the solid content of the surface attachments on the air side of the liquid film is greatly reduced (acting as a dilution effect), while the internal solid content remains basically unchanged. Under the synergistic effect of the casting solution and the subsequent coagulation bath for further phase separation, it is beneficial for the appearance of ideal pore size and ideal number of pores on the first outer surface of the porous membrane, and the pores are evenly distributed on the first outer surface. At the same time, the temperature of the organic solvent vapor is relatively high, which is beneficial for the presence of first fibers of reasonable thickness on the first outer surface, ensuring that the pores do not easily collapse when the fluid passes through the large pores, and the membrane as a whole has high mechanical strength. The organic solvent in the organic solvent vapor is at least one of acetone, dioxane, dimethylacetamide, N-methylpyrrolidone, acetic acid, propionic acid, butyric acid, and valeric acid. This organic solvent can be the same as or different from the good solvent.

[0094] After pretreatment, the film is placed in a coagulation bath for phase separation and solidification. It should be noted that, due to the relatively low content of pore-forming agent, in order to ensure the formation of an ideal pore structure in some areas of the porous membrane, the temperature of the coagulation bath is relatively low, which is 5-15℃ lower than the temperature of the casting solution. This setting helps to accelerate the phase separation rate and ultimately obtain a porous membrane with an ideal membrane structure.

[0095] Preferably, the membrane is hydrolyzed in a 0.01-0.1 mol / L sodium hydroxide aqueous solution at a temperature of 30-60℃ for 40-60 min. After hydrolysis, the membrane is washed to obtain a solid membrane. This solid membrane exhibits better hydrophilicity and lower non-specific protein adsorption, thereby further improving protein yield and economic efficiency. Furthermore, research has revealed that the membrane pores may shrink during hydrolysis, leading to excessively low porosity and flux. Therefore, 0.1-1 wt% of an anti-shrinkage agent, a mixture of glycerol and butyl orthosilicate in a 3:1 mass ratio, is added to the sodium hydroxide aqueous solution. The combined action of these two substances ensures that the membrane pores do not shrink during hydrolysis, resulting in a high-porosity solid membrane that meets the requirements of practical applications.

[0096] Preferably, the hydrolyzed solid membrane is placed in an alkaline environment (pH 9-12) and crosslinked with a water-soluble crosslinking agent (concentration of 5%-15%), wherein the crosslinking agent is at least one of halogenated epoxides, dihaloalkanes, and dihaloalcohols; the crosslinking time is 20-400 min, and the temperature is 30℃-60℃; during the crosslinking modification process, the hydroxyl groups in the solid membrane react with epoxy and halogen functional groups, thereby increasing the mechanical strength of the membrane structure, making it less prone to swelling, and also improving the alkali resistance of the membrane and extending its service life.

[0097] The beneficial effects of this invention are as follows: A cellulose porous membrane for virus removal comprises a main body with non-directional tortuous pathways within it. One side surface of the main body is a first outer surface, and the other side surface is a second outer surface. The average pore size of the first outer surface is larger than that of the second outer surface. During virus removal filtration, the first outer surface serves as the inlet surface, and the second outer surface serves as the outlet surface. The porosity of the porous membrane is 15%-45%, resulting in a large flux, fast filtration speed, high loading capacity, and long service life, meeting the needs of practical applications. In the wetted state of the porous membrane, the region that captures colloidal gold with a diameter of 20 nm is designated as D20. D20 is the region extending from the first outer surface to within 5%-100% of the thickness of the main body, and the D20 region... The membrane has a thickness of 15-60 μm; the region that captures colloidal gold with a diameter of 30 nm is designated as D30, and the overlapping region of D30 and D20 is designated as L1. L1 is located on the side of the D20 region closer to the first outer surface, and the thickness ratio of L1 to the thickness of the D20 region is not less than 0.15. The membrane body has a relatively thick pore region, and the pore size of the pore region changes little with the thickness, thereby ensuring that the porous membrane has a strong retention effect on small viruses with a particle size of 20 nm and above. At the same time, the pore size of the pore region is not too small, thus achieving a high protein yield and higher pressure resistance of the porous membrane, making it particularly suitable for the field of virus removal. In addition, this invention also provides a method for preparing this porous membrane, which is convenient, fast, effective, simple to operate, environmentally friendly, and suitable for large-scale promotion. Attached Figure Description

[0098] Figure 1 The image shows a scanning electron microscope (SEM) image of the first outer surface of the porous membrane prepared in Example 1, with a magnification of 50K×.

[0099] Figure 2 The image shows a scanning electron microscope (SEM) image of the second outer surface of the porous membrane prepared in Example 1, with a magnification of 50K×.

[0100] Figure 3 The image shown is a scanning electron microscope (SEM) image of the longitudinal section of the porous membrane prepared in Example 1, with a magnification of 2K×.

[0101] Figure 4 The image shows a scanning electron microscope (SEM) image of the longitudinal section of the porous membrane prepared in Example 1 near the second outer surface, with a magnification of 20K×.

[0102] Figure 5 The image shows a scanning electron microscope (SEM) image of the first outer surface of the porous membrane prepared in Example 8, with a magnification of 5K×.

[0103] Figure 6The image shows a scanning electron microscope (SEM) image of the second outer surface of the porous membrane prepared in Example 8, with a magnification of 20K×.

[0104] Figure 7 The image shows a scanning electron microscope (SEM) image of the longitudinal section of the porous membrane prepared in Example 8 near the first outer surface, with a magnification of 20K×.

[0105] Figure 8 The image shows a scanning electron microscope (SEM) image of the longitudinal section of the porous membrane prepared in Example 8 near the second outer surface, with a magnification of 20K×.

[0106] Figure 9 The image shows a scanning electron microscope (SEM) image of the longitudinal section of the porous membrane prepared in Example 9, near the first outer surface, with a magnification of 10K×.

[0107] Figure 10 The image shows a scanning electron microscope (SEM) image of the larger pore area in the longitudinal section of the porous membrane prepared in Example 9, with a magnification of 10K×.

[0108] Figure 11 The image shows a scanning electron microscope (SEM) image of the longitudinal section of the porous membrane prepared in Example 9, near the second outer surface, with a magnification of 10K×.

[0109] Figure 12 This is a schematic diagram of the porous membrane flux testing device of the present invention;

[0110] Figure 13 This is a schematic diagram of the testing apparatus for testing the retention efficiency of colloidal gold in porous membranes according to the present invention;

[0111] Figure 14 Scanning electron microscope (SEM) image of the cross-section of a multilayer composite ultraporous membrane prepared for patent CN1759924B;

[0112] Figure 15 A schematic diagram of the composite device used to prepare a multilayer composite ultraporous membrane for patent CN1759924B. Detailed Implementation

[0113] To more clearly illustrate the overall concept of this application, a detailed description is provided below using examples. Unless otherwise specified, in the following examples, the raw materials and equipment used to prepare the porous membrane are commercially available; wherein, a Hitachi S-5500 scanning electron microscope is used to characterize the structural morphology of the virus-removing membrane.

[0114] Example 1: A method for preparing a cellulose porous membrane for virus removal, comprising the following steps:

[0115] S1: Prepare a casting solution and cast it onto a carrier to form a liquid film; the casting solution comprises the following components by weight: 14 parts film-forming polymer; 40 parts good solvent; 24 parts pore-forming agent; the casting solution temperature is 25℃;

[0116] The film-forming polymer is cellulose diacetate; the good solvent is acetone; and the pore-forming agent is isopropanol.

[0117] S2: Pretreatment: The liquid film is placed in a high-concentration acetone vapor environment for pretreatment for 3 seconds to obtain a green film; the acetone vapor temperature is 27℃ and the flow rate is 0.4m / s; high-concentration acetone vapor refers to the ratio of the actual acetone vapor pressure in the environment to the saturated acetone vapor pressure at the same temperature being 0.82; S3: The green film is immersed in a coagulation bath for phase separation and solidification; the coagulation bath temperature is 12℃ and the duration is 55 seconds to obtain a solid film; the coagulation bath is water;

[0118] S4: The film is placed in a 0.05 mol / L sodium hydroxide aqueous solution for hydrolysis at a temperature of 50°C for 50 min. After hydrolysis, the film is washed to obtain a solid film. The sodium hydroxide aqueous solution contains 0.5 wt% anti-shrinkage agent, which is a mixture of glycerol and butyl orthosilicate in a mass ratio of 3:1.

[0119] The main difference between Examples 2-6 and Example 1 lies in the formulation of the casting solution and the process parameters for each step in preparing the porous membrane, as detailed in the table below:

[0120] Examples 2-6 Casting solution formulations

[0121]

[0122] Examples 2-6 Pretreatment Process

[0123]

[0124] Examples 2-6 Phase Separation Curing and Hydrolysis Process

[0125]

[0126] Compared to Examples 2, 4, 5 and 6, Example 3 did not undergo the corresponding hydrolysis process, which would result in a certain decrease in the protein yield of the porous membrane.

[0127] Example 7: A method for preparing a cellulose porous membrane for virus removal, comprising the following steps:

[0128] S1: Prepare a casting solution and cast it onto a carrier to form a liquid film; the casting solution comprises the following components by weight: 15 parts film-forming polymer; 50 parts good solvent; 24 parts pore-forming agent; the casting solution temperature is 25℃;

[0129] The film-forming polymer is cellulose diacetate; the good solvent is dioxane; and the pore-forming agent is isopropanol.

[0130] S2: Pretreatment: The liquid film is placed in a high-concentration acetone vapor environment for pretreatment for 4 seconds to obtain a raw film; wherein the acetone vapor temperature is 30℃ and the flow rate is 0.6m / s.

[0131] S3: The film is immersed in a coagulation bath for phase separation and solidification. The coagulation bath temperature is 15℃ and the duration is 65s to obtain the film. The coagulation bath is water.

[0132] S4: The film is placed in a 0.05 mol / L sodium hydroxide aqueous solution for hydrolysis at a temperature of 50°C for 50 min; after hydrolysis, it is washed to obtain a solid film; 0.6 wt% of an anti-shrinkage agent is added to the sodium hydroxide aqueous solution, which is a mixture of glycerol and butyl orthosilicate in a mass ratio of 3:1;

[0133] S5: The hydrolyzed solid membrane is placed in an alkaline environment of pH=10 and crosslinked with a water-soluble crosslinking agent. After crosslinking, it is washed to obtain the finished membrane. The crosslinking agent is epichlorohydrin, the concentration of crosslinking agent in the aqueous solution is 10%, the crosslinking time is 100 min, and the temperature is 45℃.

[0134] In Example 7, by cross-linking the hydrolyzed solid membrane, the tensile strength and compressive strength of the finished membrane were further improved, enhancing its industrial applicability. At the same time, it exhibited better alkali resistance, further increasing its service life when filtering fluids and enabling it to efficiently trap various small viruses for extended periods.

[0135] Example 8: A method for preparing a cellulose porous membrane for virus removal, comprising the following steps:

[0136] S1: Prepare a casting solution and cast it onto a carrier to form a liquid film; the casting solution comprises the following components by weight: 26 parts of film-forming polymer; 54 parts of good solvent; 35 parts of pore-forming agent; the casting solution temperature is 30℃;

[0137] The film-forming polymer is cellulose diacetate; the good solvent is dioxane; and the pore-forming agent is 1-propanol.

[0138] S2: Pretreatment: The liquid film is placed in a high-concentration organic solvent (butyric acid) vapor environment for pretreatment for 8 seconds; a raw film is obtained; wherein the high-concentration organic solvent vapor refers to the ratio of the actual organic solvent vapor pressure in the environment to the saturated organic solvent vapor pressure at the same temperature being 0.95; the organic solvent vapor temperature is 36℃, and the flow velocity of the organic solvent vapor is 0.7m / s;

[0139] S3: The film is immersed in a coagulation bath for phase separation and solidification. The temperature of the coagulation bath is 18°C ​​lower than the temperature of the casting solution, and the duration is 46s to obtain the film. The coagulation bath is water.

[0140] S4: The film is placed in a 0.05 mol / L sodium hydroxide aqueous solution for hydrolysis at a temperature of 50°C for 50 min; after hydrolysis, it is washed to obtain a solid film; wherein the sodium hydroxide solution contains 0.5 wt% anti-shrinkage agent, which is a mixture of glycerol and butyl orthosilicate in a mass ratio of 3:1.

[0141] Example 9: A method for preparing a cellulose porous membrane for virus removal, comprising the following steps:

[0142] S1: Prepare a casting solution and cast it onto a carrier to form a liquid film; the casting solution comprises the following components by weight: 28 parts of film-forming polymer; 57 parts of good solvent; 38 parts of pore-forming agent; the casting solution temperature is 30℃;

[0143] The film-forming polymer is cellulose triacetate; the good solvent is dimethylacetamide; and the pore-forming agent is n-butanol.

[0144] S2: Pretreatment: The liquid film is placed in a high-concentration organic solvent (dimethylacetamide) vapor environment for pretreatment for 9 seconds; a raw film is obtained; wherein the high-concentration organic solvent vapor refers to the ratio of the actual organic solvent vapor pressure in the environment to the saturated organic solvent vapor pressure at the same temperature being 96%; the organic solvent vapor temperature is 37℃, and the flow velocity of the organic solvent vapor is 0.8m / s;

[0145] S3: The film is immersed in a coagulation bath for phase separation and solidification. The temperature of the coagulation bath is 18°C ​​lower than that of the casting solution, and the duration is 48s to obtain the film. The coagulation bath is water.

[0146] Example 10: A method for preparing a cellulose porous membrane for virus removal, comprising the following steps:

[0147] S1: Prepare a casting solution and cast it onto a carrier to form a liquid film; the casting solution comprises the following components by weight: 25 parts film-forming polymer; 52 parts good solvent; 32 parts pore-forming agent; the casting solution temperature is 35℃;

[0148] The film-forming polymer is a mixture of cellulose diacetate and cellulose triacetate in a mass ratio of 2:3; the good solvent is N-methylpyrrolidone; and the pore-forming agent is ethanol.

[0149] S2: Pretreatment: The liquid film is placed in a high-concentration organic solvent vapor environment for pretreatment for 7 seconds; a raw film is obtained; wherein the high-concentration organic solvent vapor (dimethylacetamide) refers to the ratio of the actual organic solvent vapor pressure in the environment to the saturated organic solvent vapor pressure at the same temperature being 94%; the organic solvent vapor temperature is 41℃, and the flow velocity of the organic solvent vapor is 0.6m / s;

[0150] S3: The film is immersed in a coagulation bath for phase separation and solidification. The temperature of the coagulation bath is 20°C lower than the temperature of the casting solution, and the duration is 42s to obtain the film. The coagulation bath is water.

[0151] S4: The membrane is hydrolyzed in a 0.05 mol / L sodium hydroxide aqueous solution at 50°C for 50 min; after hydrolysis, it is washed to obtain a solid membrane; wherein the sodium hydroxide solution contains 0.4 wt% of an anti-shrinkage agent, which is a mixture of glycerol and butyl orthosilicate in a mass ratio of 3:1. Example 11: A method for preparing a cellulose porous membrane for virus removal, comprising the following steps:

[0152] S1: Prepare a casting solution and cast it onto a carrier to form a liquid film; the casting solution comprises the following components by weight: 24 parts film-forming polymer; 50 parts good solvent; 30 parts pore-forming agent; the casting solution temperature is 35℃;

[0153] The film-forming polymer is a mixture of cellulose diacetate and cellulose triacetate in a mass ratio of 1:2; the good solvent is butyric acid; and the pore-forming agent is isopropanol.

[0154] S2: Pretreatment: The liquid film is placed in a high-concentration organic solvent vapor environment for pretreatment for 6 seconds; a raw film is obtained; wherein the high-concentration organic solvent (propionic acid) vapor refers to the ratio of the actual organic solvent vapor pressure in the environment to the saturated organic solvent vapor pressure at the same temperature being 92%; the temperature of the organic solvent vapor is 40℃, and the flow velocity of the organic solvent vapor is 0.5m / s;

[0155] S3: The film is immersed in a coagulation bath for phase separation and solidification. The temperature of the coagulation bath is 20°C lower than the temperature of the casting solution, and the duration is 39s to obtain the film. The coagulation bath is water.

[0156] S4: The film is placed in a 0.05 mol / L sodium hydroxide aqueous solution for hydrolysis at a temperature of 50°C for 45 min; after hydrolysis, it is washed to obtain a solid film; wherein the sodium hydroxide solution contains 0.3 wt% anti-shrinkage agent, which is a mixture of glycerol and butyl orthosilicate in a mass ratio of 3:1.

[0157] Example 12: A method for preparing a cellulose porous membrane for virus removal, comprising the following steps:

[0158] S1: Prepare a casting solution and cast it onto a carrier to form a liquid film; the casting solution comprises the following components by weight: 30 parts of film-forming polymer; 60 parts of good solvent; 40 parts of pore-forming agent; the casting solution temperature is 28℃;

[0159] The film-forming polymer is nitrocellulose; the good solvent is acetone; and the pore-forming agent is 1-pentanol.

[0160] S2: Pretreatment: The liquid film is placed in a high-concentration organic solvent vapor environment for pretreatment for 2-10 seconds; a raw film is obtained; wherein the high-concentration organic solvent (acetone) vapor refers to the ratio of the actual organic solvent vapor pressure in the environment to the saturated organic solvent vapor pressure at the same temperature being 98%; the organic solvent vapor temperature is 36℃, and the flow velocity of the organic solvent vapor is 0.9m / s;

[0161] S3: The film is immersed in a coagulation bath for phase separation and solidification. The temperature of the coagulation bath is 16°C lower than the temperature of the casting solution, and the duration is 52s to obtain the film. The coagulation bath is water.

[0162] S4: The film is placed in a 0.08 mol / L sodium hydroxide aqueous solution for hydrolysis at a temperature of 50°C for 58 min; after hydrolysis, it is washed to obtain a solid film; wherein the sodium hydroxide solution contains 0.9 wt% anti-shrinkage agent, which is a mixture of glycerol and butyl orthosilicate in a mass ratio of 3:1.

[0163] S5: The hydrolyzed solid membrane is placed in an alkaline environment with pH=9 and crosslinked with a water-soluble crosslinking agent. After crosslinking, it is washed to obtain the finished membrane. The crosslinking agent is epichlorohydrin, the concentration of crosslinking agent in the aqueous solution is 12%, the crosslinking time is 240 min, and the temperature is 50℃.

[0164] Comparative Example 1 replaced the pore-forming agent isopropanol in Example 1 with polyvinylpyrrolidone, and kept the other conditions unchanged. After testing, it was found that there was a dense surface layer in the membrane with a very small pore size of 20-30 nm. However, the thickness of the dense surface layer was very small, about 1 μm, and it had no loading capacity at all, so it could not form a virus-removing membrane that could be used in practice.

[0165] 1. Structural Characterization

[0166] The morphology of the nanoscale polymer porous membranes obtained in each embodiment was characterized using scanning electron microscopy, and the required data were then obtained; the specific results are shown in the table below:

[0167] Table 1:

[0168]

[0169] Table 2

[0170]

[0171] Table 3

[0172]

[0173] The virus-free membranes prepared in each embodiment were used as the test objects. Two samples were cut from each sample, with the distance between the two samples not exceeding 1 cm, and were named Sample 1 and Sample 2.

[0174] Peak capture region of colloidal gold at 20nm before top washing

[0175] Sample 1 was subjected to a 20nm colloidal gold capture test, and the brightness of the cross section of the virus-removing membrane containing the 20nm colloidal gold was measured. The maximum peak of the spectrum is the peak capture location of the 20nm colloidal gold. At the same time, the distance between the maximum peak of the spectrum on the cross section of the virus-removing membrane and the second outer surface was recorded.

[0176] Peak capture region of 20nm colloidal gold after top washing

[0177] Using sample 1, a 20 nm colloidal gold capture test was performed. Filtration was stopped when the flow rate of colloidal gold decreased by 75% or when only a small amount of colloidal gold remained. The pressure was then slowly released to completely relieve the pressure inside the container holding the colloidal gold, and the remaining colloidal gold was poured out. Approximately 30 mL of buffer solution was added, and the container was allowed to stand. After a 30-minute pressure interruption, pressure was applied for top-wash filtration. After top-washing, the brightness of the cross-section of the virus-removing membrane containing the captured 20 nm colloidal gold was measured. The maximum peak of the spectrum was the peak capture location of the 20 nm colloidal gold. Simultaneously, the distance between the maximum peak of the spectrum on the cross-section of the virus-removing membrane and the second outer surface was recorded.

[0178] Table 4

[0179]

[0180] Table 5

[0181]

[0182] Note: As shown in Tables 1-5, the porous membranes prepared in Examples 1-10 of this invention all have ideal membrane structures. These porous membranes are integrally formed without composite processes, making the preparation process simple. Among them, the porous membranes prepared in Examples 1-5 are relatively thin. In the main structure of these membranes, the thickness of the macropore region is small, while the thickness of the micropore region is large and accounts for a large proportion of the overall membrane thickness. The overall pore size of the membrane changes relatively little.

[0183] The porous membranes prepared in Examples 6-10 are relatively thick. In these membrane structures, the macropore region is thicker, and the pore size change rate near the first outer surface is greater than that near the second outer surface (the membrane pore size changes at a slower rate in the macropore region). At the same time, the thickness of the micropore region is relatively small compared to the overall membrane thickness, and the overall membrane pore size change is larger. Both of these membrane structures can ensure efficient virus retention, high flux and load capacity, long service life, and low protein adsorption, making them particularly suitable for use in the field of virus removal.

[0184] Performance characteristics

[0185] Membrane flux is calculated as follows:

[0186] The formula for calculating membrane flux (J) is: J = V / (T × A) where:

[0187] J -- Membrane flux unit: L*h -1 *m -2

[0188] V -- Sampling volume (L); T -- Sampling time (h); A -- Effective membrane area (m2)

[0189] The operating conditions used for determining the porous membrane separation performance in this invention are as follows: the feed solution is deionized water, the operating pressure is 30 psi, the operating temperature is 25°C, and the solution pH is 7; the flux testing device is... Figure 12 ;

[0190] Example 1 120 6.0 5.0 Example 2 90 6.2 5.1 Example 3 150 4.8 4.0 Example 4 125 5.6 4.7 Example 5 115 5.9 4.9 Example 6 180 3.9 3.2 Example 7 60 7.1 5.9 Example 8 100 6.4 5.1 Example 9 140 5.1 4.1 Example 10 120 5.8 4.6 Example 11 80 6.6 5.3 Example 12 130 5.0 4.2

[0191] After testing, it was found that the porous membranes prepared in Examples 1-10 (except Example 6) had an LRV of no less than 4 for 20nm colloidal gold, and the LRV remained high after top washing, thus indicating that the porous membrane of the present invention has sufficient retention for viruses of 20nm and above; moreover, the porous membrane has good flux and fast filtration speed; therefore, the porous membrane is particularly suitable for use in the field of virus removal; while although the LRV of the single layer of Example 6 is less than 4, due to its large flux, it can be used by stacking two layers in actual use, and its LRV can reach more than 7, and the flux is still large, which can meet the needs of practical applications.

[0192] Furthermore, according to the test method used in paragraph 114 of CN201010154974.7-Ultraporous membrane and its preparation method: protein yield test of the sample was performed, and it was found that the protein yield of the porous membrane of the present invention is not less than 98%, and can even reach 99%, with basically no protein retention or adsorption, thus ensuring economic benefits.

[0193] In addition, the pressure resistance of the samples in Examples 1-10 was tested, and the pressure resistance was greater than 30 psi. Under the pressure of 30 psi, they were able to filter the corresponding fluid stably and quickly. At the same time, during the integrity test, they were subjected to a pressure of 50 psi, and the membrane pores remained intact. Furthermore, the tensile strength of Examples 7 and 10 was tested, and their wet tensile strength was significantly increased, which is due to the crosslinking treatment of the membrane.

[0194] The above description is merely a preferred embodiment of the present invention. The scope of protection of the present invention is not limited to the above embodiments. All technical solutions falling within the scope of the present invention's concept are within the scope of protection of the present invention. It should be noted that for those skilled in the art, any improvements and modifications made without departing from the principles of the present invention should also be considered within the scope of protection of the present invention.

Claims

1. A porous cellulose membrane for virus removal, characterized in that: The membrane comprises a main body having non-directional tortuous pathways within it. One side surface of the main body is a first outer surface, and the other side surface of the main body is a second outer surface. The average pore size of the first outer surface is larger than the average pore size of the second outer surface. The porosity of the porous membrane is 15%-45%. In a porous membrane substrate in a wet state, the region that captures colloidal gold with a diameter of 20 nm is designated as D20. D20 is located in the region of 5%-100% of the thickness of the substrate, and the thickness of the D20 region is 15-60 μm. The region that captures colloidal gold with a diameter of 30nm is designated as D30. The overlapping region of D30 and D20 is designated as L1. L1 is located on the side of the D20 region closer to the first outer surface. The ratio of the thickness of L1 to the thickness of the D20 region is not less than 0.

15.

2. The cellulose porous membrane for virus removal according to claim 1, characterized in that: The D30 is located in the region of 5%-90% of the thickness of the main body; The thickness of L1 is 5-50 μm, and L1 is located in the region of 5%-90% of the thickness of the main body. The ratio of the thickness of L1 to the thickness of the D20 region is not less than 0.

3.

3. The cellulose porous membrane for virus removal according to claim 1, characterized in that: The D20 is located in the region of 15%-100% of the thickness of the main body, and the thickness of the D20 region is 20-50 μm; the ratio of the thickness of L1 to the thickness of the D20 region is 0.4-0.

9.

4. The cellulose porous membrane for virus removal according to claim 1, characterized in that: The capture peak of colloidal gold with a diameter of 20 nm is located in the region of 75-92% of the body thickness from the first outer surface, and the distance between the peak and the second outer surface is 3-12 μm.

5. The cellulose porous membrane for virus removal according to claim 1, characterized in that: The porous membrane is used to capture 20 nm colloidal gold, and the porous membrane with captured 20 nm colloidal gold is top-washed. After top-washing, the distance between the peak part of the porous membrane with captured 20 nm colloidal gold and the second outer surface is 1-8 μm, and the ratio of this distance to the D20 thickness is 3%-20%.

6. The cellulose porous membrane for virus removal according to claim 4, characterized in that: Before and after top washing, the peak position of the captured 20nm colloidal gold changed displacement within the porous membrane by 0.5-4μm; and this displacement accounted for 2-12% of the D20 region.

7. The cellulose porous membrane for virus removal according to claim 1, characterized in that: The region that captures colloidal gold with a diameter of 40 nm is designated as D40, which is located in the area of ​​5%-80% of the body thickness. The overlapping area between D40 and D30 is L2; ​​the ratio of the thickness of L1 to the thickness of L2 is 1.2-2.5:1; The ratio of the thickness of the main region capturing colloidal gold with a diameter of 20-40 nm to the thickness of the main region capturing colloidal gold with a diameter of 20-30 nm is 1.1-1.

8.

8. The cellulose porous membrane for virus removal according to claim 1, characterized in that: The first outer surface includes a plurality of strip-shaped continuous first fibers, and adjacent continuous first fibers surround each other to form circular holes; the average diameter of the first fibers is 60-600 nm. The area ratio of the holes on the first outer surface is 5-35%.

9. A cellulose porous membrane for virus removal according to any one of claims 1-8, characterized in that: The thickness of the porous membrane body is 25-70 μm, the average pore size of the first outer surface is 150-800 nm, and the ratio of the average pore size of the first outer surface to that of the second outer surface is 6-30. The ratio of the average pore size of the first outer surface to the average diameter of the first fiber is 2-8.

10. A cellulose porous membrane for virus removal according to claim 8, characterized in that: The thickness of the host region for capturing colloidal gold with a diameter greater than 40 nm is 3-25 μm, and it is located in the region of 0-40% of the host thickness from the first outer surface. In the host region for capturing colloidal gold with a diameter greater than 40 nm, the pore size change rate near the first outer surface is greater than that near the second outer surface.

11. A cellulose porous membrane for virus removal according to any one of claims 1-8, characterized in that: The thickness of the porous membrane is 60-120 μm, the average pore size of the first outer surface is 400-2000 nm, and the ratio of the average pore size of the first outer surface to that of the second outer surface is 20-100.

12. A cellulose porous membrane for virus removal according to claim 11, characterized in that: The thickness of the host region for capturing colloidal gold with a diameter greater than 40 nm is 35-100 μm, and it is located in the region of 0%-75% of the host thickness from the first outer surface.

13. A cellulose porous membrane for virus removal according to claim 11, characterized in that: The thickness of the host region for capturing colloidal gold with a diameter of 60-80 nm is 0.5-4 μm greater than that for capturing colloidal gold with a diameter of 80-100 nm. The thickness of the host region for capturing colloidal gold with a diameter of 40-60 nm is 1-7 μm greater than that for capturing colloidal gold with a diameter of 60-80 nm. The thickness of the host region for capturing colloidal gold with a diameter of 20-40 nm is 2-12 μm greater than that for capturing colloidal gold with a diameter of 80-100 nm.

14. A cellulose porous membrane for virus removal according to claim 11, characterized in that: The thickness of the main region for capturing colloidal gold with a diameter of 80-100nm is 2-16μm; The thickness of the main region for capturing colloidal gold with a diameter of 60-80 nm is 3-20 μm; The thickness of the host region for capturing colloidal gold with a diameter of 40-60 nm is 4.5-23 μm.

15. A cellulose porous membrane for virus removal according to claim 11, characterized in that: When the diameter of the captured colloidal gold is 20-40 nm, the capture gradient K1 along the thickness direction of the host is 0.5-3.5 nm / μm; When the diameter of the captured colloidal gold is 40-100 nm, the capture gradient K2 along the thickness direction of the host is 1-6 nm / μm; Capture gradient = Capture colloidal gold diameter change value / Capture thickness of the main region of colloidal gold with the corresponding diameter.

16. A cellulose porous membrane for virus removal according to claim 15, characterized in that: The K2:K1 = 1.1-2.

5.

17. A cellulose porous membrane for virus removal according to claim 11, characterized in that: The pressure resistance of the porous membrane is not less than 30 psi; The flux of the porous membrane is greater than 40 L*h -1 *m -2 @30psi; The porous membrane has an LRV greater than 4 for 20nm colloidal gold; The protein yield of the porous membrane is not less than 98%.

18. A method for preparing a cellulose porous membrane for virus removal according to any one of claims 1-17, characterized in that: Includes the following steps: S1: Prepare a casting solution and cast it onto a carrier to form a liquid film; the casting solution comprises the following components by weight: 10-30 parts of film-forming polymer; 30-60 parts of good solvent; 20-40 parts of pore-forming agent; The mass of the good solvent in the casting solution is 1.1-2.5 times the mass of the pore-forming agent; The film-forming polymer is a cellulose polymer; The pore-forming agent refers to a non-good solvent substance with a surface tension of less than 30 mN / m, and the substance is miscible with the coagulation bath; S2: Pretreatment: The liquid film is placed in a high-concentration organic solvent vapor environment for pretreatment for 2-10 seconds; thus, a raw film is obtained. High-concentration organic solvent vapor refers to a ratio of the actual organic solvent vapor pressure in the environment to the saturated organic solvent vapor pressure at the same temperature of not less than 80%; the temperature of the organic solvent vapor is 2-8℃ higher than the temperature of the casting solution, and the flow velocity of the organic solvent vapor is 0.3m / s-0.9m / s; S3: Immerse the film-forming material in a coagulation bath for phase separation and solidification. The temperature of the coagulation bath is 5-15°C lower than the temperature of the casting solution, and the duration is 30-80 seconds to obtain the film. The coagulation bath is water.

19. The method for preparing a cellulose porous membrane for virus removal according to claim 18, characterized in that: The temperature of the casting solution is 15-40℃; The cellulose polymer is at least one of nitrocellulose and cellulose acetate; The pore-forming agent is any one of ethanol, 1-propanol, isopropanol, n-butanol, 1-pentanol, and diethyl ether; The good solvent is at least one selected from acetone, dioxane, dimethylacetamide, N-methylpyrrolidone, acetic acid, propionic acid, butyric acid, and valeric acid; The organic solvent in the organic solvent vapor is at least one of acetone, dioxane, dimethylacetamide, N-methylpyrrolidone, acetic acid, propionic acid, butyric acid, and valeric acid.

20. The method for preparing a cellulose porous membrane for virus removal according to claim 18, characterized in that: The film is placed in an aqueous sodium hydroxide solution for hydrolysis at a temperature of 30-60℃ for 40-60 minutes. After hydrolysis, the film is washed to obtain a solid film. The aqueous sodium hydroxide solution contains 0.1-1 wt% of an anti-shrinkage agent, which is a mixture of glycerol and butyl orthosilicate in a mass ratio of 3:

1.

21. The method for preparing a cellulose porous membrane for virus removal according to claim 20, characterized in that: The hydrolyzed solid membrane is placed in an alkaline environment and crosslinked with a water-soluble crosslinking agent. After crosslinking is completed, it is cleaned to obtain the finished membrane. The crosslinking agent is at least one of haloepoxides, dihaloalkanes and dihaloalcohols; the crosslinking time is 20-400 min and the temperature is 30℃-60℃.

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