Method for filtering cell culture

Abstract

To provide a more stable filtration method capable of separating a culture containing cells and a product containing an enzyme and an antibody from each other without clogging at a lower tangential flow linear velocity.SOLUTION: This filtration method comprises a filtration step of passing a culture liquid containing a cell, a medium, a product, and an antifoam through a porous film and separating a filtrate containing the product from the culture liquid. A product of a porosity in a thickness from the outermost surface of the filtered liquid side surface of the porous film to 0.12% of a film thickness and an aperture of the filtered liquid side surface is 1800% or higher.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates generally to process filtration systems, and more specifically to a method for filtering cell cultures utilizing hollow fiber membranes having a high porosity and open area on the surface facing the liquid to be filtered. [Background technology]

[0002] Filtration is utilized to separate, purify, modify, and / or concentrate fluid solutions, mixtures, or suspensions. It is often a necessary step in the production, processing, and analysis stages of drugs, diagnostics, and chemicals by the biotechnology, pharmaceutical, and medical industries. Filtration may be used, for example, to remove desired compounds from a solution or to remove by-products, leaving only a more concentrated medium. Such processes may be modified as needed by selecting various filter materials, pore sizes, and / or other filter variables.

[0003] When producing cell cultures, it is often necessary to remove waste products from the growth culture. Advances in biological manufacturing processes now allow for the large-scale production of cell cultures, often using process vessel devices to produce recombinant proteins, virus-like particles (VLPs), gene therapy particles, and vaccines. Cell retention devices are widely available that remove metabolic waste products and add nutrients to renew the culture. Typically, this retention is performed by perfusion filtration of the process vessel culture with hollow fiber membranes using tangential flow filtration. In addition, bioprocessing runs are long term and continuous cultures are utilized. Such runs can last for days, weeks, or months. It is desirable for many typical components, such as filters, to function satisfactorily for such periods without fouling or otherwise requiring maintenance or replacement.

[0004] Hollow fiber membranes are often used for cell culture perfusion and product separation, but such use can result in membrane fouling by cell debris and the like. Fouling can then cause the desired product to be retained within the membrane rather than passing through. Hollow fiber membranes are commonly made of polyethersulfone (PES), polysulfone, cellulose, polyvinylidene fluoride, polypropylene, polyethylene and other materials, and there is a continuing demand for membranes that are resistant to fouling and allow for the filtration of solutions containing high solids. Fluororesins such as polyethylene (PE) and polyvinylidene fluoride (PVDF) are hydrophobic materials that have been commonly used in industrial filtration, and are known to adsorb certain molecules less. Specifically, polyphenols, polysaccharides, and tannins are adsorbed at lower levels. On the other hand, the structure of the membrane varies greatly depending on the manufacturing method, and even if the membrane is made of the same material, the fouling phenomenon on the membrane varies greatly depending on the manufacturing method.

[0005] The following Patent Document 1 describes the relationship between the filtration rate of the permeate in tangential flow filtration and the flow rate of the culture solution. It also describes the use of a porous membrane containing a dense layer. As shown in FIG. 6, tangential flow filtration is also known as cross-flow filtration, and is a method in which a solution sample to be filtered is refluxed in a direction parallel to the membrane surface, generating a certain pressure on the membrane and allowing particles smaller than the membrane pore size to pass through. In contrast, general direct flow filtration is a method in which a liquid is passed from top to bottom on a membrane surface such as a syringe filter or centrifugal filter, in which case, fine particles contained in the sample are captured on the membrane surface or inside and accumulate, causing clogging and generating excessive pressure load on the sample. In contrast, in tangential flow filtration, the sample is constantly refluxed in a direction parallel to the membrane surface, so particles and molecules larger than the membrane pore size remain upstream of the membrane, and the accumulation of fine particles on the membrane is suppressed, resulting in a low load on the sample.

[0006] The following Patent Document 2 describes a cell culture method and device that can remove cell growth inhibitors, perform high-density culture, and return at least one of the serum components, which is an expensive polymeric medium component, to the culture tank, thereby reducing running costs.

[0007] The following Patent Document 3 describes a method for increasing recombinant protein production in a mammalian cell culture expressing the recombinant protein, which describes establishing a mammalian cell culture in a serum-free culture medium in a bioreactor and inducing cell growth arrest by perfusion with a serum-free perfusion medium having an L-asparagine concentration of 5 mM or less.

[0008] The following Patent Document 4 describes a technique for monitoring and measuring the flow rate of the permeate from a filtration membrane in perfusion culture, and switching the filtration membrane when the flow rate of the permeate is at least 10% of the flow rate of the permeate at the start of tangential flow filtration.

[0009] The following Patent Document 5 describes a method for efficiently separating a product using a tangential flow filtration operation method in perfusion culture. [Prior art documents] [Patent documents]

[0010] [Patent Document 1] JP 2018-76291 A [Patent Document 2] JP 2000-32977 A [Patent Document 3] Special Publication No. 2014-520534 [Patent Document 4] JP 2014-024824 A [Patent Document 5] International Publication No. 2015 / 188009 Summary of the Invention [Problem to be solved by the invention]

[0011] Patent Documents 1 to 4 describe a method for recovering a useful substance from a culture solution in continuous culture of cells that produce a useful substance, the method comprising: a bleeding step of discharging the culture solution from a cell culture tank and adding to the culture tank an amount of fresh medium equal to the amount of the discharged culture solution; and a filtration step of filtering the culture solution extracted from the culture tank using a porous membrane that does not substantially have a dense layer, the filtration step being tangential flow filtration, and recovering the useful substance. Furthermore, Patent Document 5 merely describes a technique for efficiently separating the product from impurities.

[0012] However, although Patent Documents 1 to 4 disclose ideas on how to use the membrane, such as the speed of the permeate in the filtration process, there is no description of the membrane that is optimal for tangential flow filtration, and it is not clear what specific structure or material of the membrane should be used to achieve stable tangential flow. When continuing filtration, clogging of the membrane inevitably occurs and it is necessary to deal with it, and in terms of separation, a basic separation function is required in which cells and microorganisms are retained on the concentrated side and the product is permeated to the permeation side without being blocked by the membrane. In addition, when the culture target is animal cells, there is a problem that the linear velocity of the tangential flow cannot be increased because a too high shear rate causes problems in cell growth.

[0013] In view of the above-mentioned state of the prior art, the problem to be solved by the present invention is to provide a more stable filtration method that can separate a culture product from a product at a lower linear tangential flow velocity without clogging. [Means for solving the problem]

[0014] As a result of intensive research and repeated experiments to solve the above-mentioned problems, the inventors unexpectedly discovered that by using a hollow fiber membrane having a high porosity and aperture ratio on the surface facing the liquid to be filtered, the permeability of the culture product is good and furthermore, clogging is less likely to occur even when the linear velocity of the tangential flow is low, which led to the completion of the present invention.

[0015] That is, the present invention is as follows. [1] A filtration method comprising a filtration step of passing a culture solution containing cells, a medium, a product, and an antifoaming agent through a porous membrane to separate a filtrate containing the product from the culture solution, wherein the filtration method is characterized in that the product of the porosity of the porous membrane in a thickness from the outermost surface of the surface facing the liquid to be filtered to a thickness of 0.12% of the membrane thickness and the porosity of the surface facing the liquid to be filtered is 1800%·% or more. [2] The filtration method according to [1] above, wherein the porosity of the porous membrane is 50% or more in a thickness ranging from the outermost surface of the surface facing the liquid to be filtered to a thickness of 0.12% of the membrane thickness. [3] The filtration method according to [1] or [2] above, wherein the porosity of the surface of the porous membrane facing the liquid to be filtered is 30% or more. [4] The filtration method according to any one of [1] to [3] above, wherein the ratio of the porosity in a thickness from the outermost surface of the surface of the porous membrane facing the liquid to 0.12% of the membrane thickness to the porosity of the surface facing the liquid to be filtrated is 1.70 or more. [5] The filtration method according to any one of [1] to [4], wherein the product of the porosity in a thickness range from the outermost surface of the surface of the porous membrane facing the liquid to be filtered to 0.12% of the membrane thickness and the porosity of the surface facing the liquid to be filtered is 2800% % or more. [6] The filtration method according to any one of [1] to [5] above, wherein the product of the porosity at a thickness of 300 nm from the outermost surface of the filtrate side surface of the porous membrane in the film thickness direction and the porosity of the filtrate side surface is 860% % or more. [7] The filtration method according to [6] above, wherein the porosity at a thickness of 300 nm from the outermost surface of the filtrate side surface of the porous membrane in the film thickness direction is 35% or more. [8] The filtration method according to [7] above, wherein the porosity of the filtrate-side surface of the porous membrane is 30% or more. [9] The filtration method according to any one of [1] to [8] above, wherein the cross-sectional pore size of the porous membrane at a thickness of 0.12% of the membrane thickness from the outermost surface on the side of the liquid to be filtered is 1000 nm or less.

[10] The filtration method according to any one of [1] to [9] above, wherein the cross-sectional pore size at a thickness of 300 nm from the outermost surface of the surface of the porous membrane facing the liquid to be filtered in the film thickness direction is 300 nm or less.

[11] The filtration method according to any one of [1] to

[10] above, wherein the porous membrane has a thickness of 100 μm or more and 800 μm or less.

[12] The filtration method according to any one of [1] to

[11] above, wherein the flux of the porous membrane in the filtration step is 1 LMH or more.

[13] The filtration method according to any one of [1] to

[12] above, wherein the porous membrane is a hollow fiber membrane, and the filtration step is perfusion culture filtration.

[14] The filtration method according to any one of [1] to

[13] above, wherein the intramembrane linear velocity of the tangential flow of the hollow fiber membrane during the perfusion culture is 0.3 m / s or less.

[15] The filtration method according to any one of the above [1] to

[14] , wherein the resin constituting the porous membrane is a thermoplastic resin.

[16] The filtration method according to

[15] , wherein the thermoplastic resin contains a fluororesin as a main component.

[17] The filtration method according to

[16] , wherein the fluororesin comprises at least one selected from the group consisting of polyvinylidene fluoride resin (PVDF), chlorotrifluoroethylene resin, tetrafluoroethylene resin, ethylene-tetrafluoroethylene copolymer (ETFE), ethylene-monochlorotrifluoroethylene copolymer (ECTFE), hexafluoropropylene resin, and mixtures of these resins.

[18] The filtration method described in

[15] , wherein the thermoplastic resin contains a polyolefin resin as a main component.

[19] The filtration method described in

[15] , wherein the thermoplastic resin is polyethylene (PE) or polypropylene (PP).

[20] The filtration method according to any one of [1] to

[19] above, wherein the product is selected from the group consisting of proteins including enzymes and antibodies, amino acids, nucleic acids, and organic substances. Effect of the Invention

[0016] The culture solution filtration method of the present invention uses a porous filtration membrane with good pore connectivity from the inside of the membrane, which is the treated liquid side, to the outside of the membrane, which is the filtrate side (i.e., the surface on the filtrate side has high porosity and aperture ratio), so that the filtration method has excellent filtration performance, has good product permeability and is less likely to become clogged with cells, and can separate the culture containing cells from the product containing enzymes and antibodies, even at a lower linear tangential flow velocity, and furthermore does not damage the membrane during filtration operation, so that proteins such as antibodies can be separated efficiently. [Brief description of the drawings]

[0017] [Figure 1] FIG. 1 is a schematic diagram of a three-dimensional network structure. [Diagram 2] FIG. 1 is a diagram showing the configuration of an apparatus for producing a porous hollow fiber membrane. [Figure 3A] FIG. 13 is a diagram for explaining a method for measuring a layer boundary, and is a diagram for explaining a method for determining a line for determining a measurement position for the length of a hole used for boundary measurement. [Figure 3B] FIG. 3B is a diagram for explaining a method for measuring the layer boundary, and a diagram for explaining a method for measuring the length of a hole using the line determined in FIG. 3A. [Figure 4] This is a procedure for obtaining a binary image of only the pores on the outermost surface of the film. [Diagram 5] 1 is an electron microscope photograph of a cross section of a porous hollow fiber membrane near the side of a liquid to be filtered obtained in Example 1. [Figure 6] FIG. 1 is an explanatory diagram of tangential flow filtration. [Figure 7] FIG. 2 is an explanatory diagram of the perfusion culture filtration process in the continuous culture used in the Examples. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0018] A preferred embodiment of the present invention (hereinafter referred to as "embodiment") will be described in detail below. Note that the embodiment shown below is an example of an apparatus and method for embodying the technical idea of ​​the present invention, and the technical idea of ​​the present invention is not limited to the combination of components described below. The technical idea of ​​the present invention can be modified in various ways within the scope of the claims.

[0019] In the following description, the upper or lower limit of a stepwise described numerical range may be replaced with the upper or lower limit of another stepwise described numerical range. In addition, in the following description, the upper or lower limit of a certain numerical range may be replaced with a value described in the examples. Furthermore, with respect to the term "step" in the following description, not only an independent step, but also a step that cannot be clearly distinguished from other steps may be included in this term as long as the function of the "step" is achieved.

[0020] The porous membrane of the present embodiment is a filtration method comprising a filtration step of passing a culture solution containing cells, a medium, a product, and an antifoaming agent through a porous membrane to separate a filtrate containing the product from the culture solution, characterized in that the product of the porosity at a thickness of 0.12% of the membrane thickness from the outermost surface of the porous membrane on the side of the liquid to be filtered and the porosity of the surface on the side of the liquid to be filtered is 1800%·% or more.

[0021] The filtration method of the present embodiment is characterized by including a filtration step in which a culture solution containing cells, medium, product, antifoaming agent, etc. is passed through a porous membrane composed of a resin with a three-dimensional mesh structure to separate a filtrate containing the product from the culture solution and a culture solution containing cells. The product is not particularly limited, but examples thereof include proteins including enzymes and antibodies, amino acids, nucleic acids, organic substances, and the like, and examples thereof include proteins including enzymes and antibodies to be produced by cells, for example, genetically modified cells that produce useful proteins, for example, CHO cells. The method may also include additional processing steps, such as centrifugation, diatomaceous earth filtration, other membrane separation, sieving, or the like, before, after, or both before and after the filtration step. The shape of the porous membrane is not particularly limited, and examples thereof include a flat membrane, a tubular membrane, and a hollow fiber membrane. From the viewpoint of space saving of the filtration device, i.e., because it is possible to increase the membrane area per unit volume of the membrane module, a hollow fiber membrane is preferred.

[0022] The product may be an antibody protein, or as generally defined in biochemistry, a glycoprotein molecule (also called gamma globulin or immunoglobulin) produced by B lymphocytes as a defense mechanism against infection in vertebrates. For example, the antibody protein separated by the porous membrane according to the embodiment is used as a human pharmaceutical and has a structure substantially identical to that of an antibody protein present in the human body to which it is administered.

[0023] The product may be various antibody vaccines, vector viruses, plasmid DNA, or industrial enzymes such as amylase and protease. Various culture objects are selected according to each product. For example, when the product is an enzyme, the culture object may be bacteria such as genetically modified Escherichia coli, and when the product is a protein such as albumin, the culture object may be genetically modified Pichia yeast.

[0024] The structure, material, and manufacturing method of the porous membrane used in the culture solution filtration method of this embodiment will be described in detail below.

[0025] The porous membrane of the present embodiment preferably contains a fluororesin, such as vinylidene fluoride or chlorotrifluoroethylene, as a main component of the polymer component (e.g., thermoplastic resin) constituting the membrane. Here, "containing as a main component" means that the polymer component contains 50 mass% or more in terms of solid content. The polymer component may be only one type or a combination of multiple types.

[0026] The weight average molecular weight (Mw) of the vinylidene fluoride resin is not particularly limited, but is preferably 100,000 to 1,000,000, and more preferably 150,000 to 1,500,000. The vinylidene fluoride resin is not limited to a vinylidene fluoride resin of a single molecular weight, and may be a mixture of vinylidene fluoride resins of different molecular weights. In this embodiment, the weight average molecular weight (Mw) can be measured by gel permeation chromatography (GPC) using a standard resin of known molecular weight as a reference.

[0027] On the other hand, the porous membrane may contain other polymer components. The other polymer components are not particularly limited, but are preferably compatible with the vinylidene fluoride resin, and for example, fluorine-based resins that exhibit high chemical resistance similar to the vinylidene fluoride resins can be suitably used.

[0028] The above-mentioned porous membrane may have, for example, a hollow fiber membrane structure. Here, the hollow fiber membrane means a membrane having a hollow ring shape. By having the porous membrane have a hollow fiber membrane structure, it is possible to increase the membrane area per unit volume of the module compared to a flat membrane. However, the porous membrane of this embodiment is not limited to a porous membrane having a membrane structure of a hollow fiber membrane (a hollow fiber-shaped porous membrane), but may have other membrane structures such as a flat membrane or a tubular membrane. The porous membrane of the present embodiment is preferably a hollow fiber membrane containing a thermoplastic resin, and may be a hollow fiber membrane consisting of only a thermoplastic resin. The thermoplastic resin preferably contains a fluororesin as a main component, and may be consisting of only a fluororesin. The fluororesin preferably contains at least one selected from the group consisting of vinylidene fluoride resin (PVDF), chlorotrifluoroethylene resin, tetrafluoroethylene resin, ethylene-tetrafluoroethylene copolymer (ETFE), ethylene-monochlorotrifluoroethylene copolymer (ECTFE), hexafluoropropylene resin, and mixtures of these resins, and may be at least one selected from the group consisting of vinylidene fluoride resin (PVDF), chlorotrifluoroethylene resin, tetrafluoroethylene resin, ethylene-tetrafluoroethylene copolymer (ETFE), ethylene-monochlorotrifluoroethylene copolymer (ECTFE), hexafluoropropylene resin, and mixtures of these resins.

[0029] In the porous membrane of this embodiment, it is preferable that the ratio of the porosity in a thickness from the outermost surface of the surface facing the liquid to 0.12% of the film thickness (the portion from the outermost surface to a position that is 0.12% of the film thickness in the film thickness direction from the outermost surface) to the open area of ​​the surface facing the liquid to be filtrated is 1.70 or more. When the ratio is 1.70 or more, the interconnectivity between the surface pores and the pores near the surface inside the membrane is good, and the pores near the surface that contribute most to filtration are less likely to be blocked. In addition, since the interconnectivity of the pores near the surface is good, dirt can be easily removed by backwashing or other cleaning or by the crossflow effect, so that high filtration performance can be achieved. The ratio is preferably 1.75 or more, and more preferably 1.70 to 2.50. When the ratio is 2.50 or less, the polymer that forms the surface pores is less likely to deform, and blocking performance can be maintained. As described above, it is desirable that the ratio is 1.70 or more in the thickness from the outermost surface of the surface on the side of the liquid to be filtered to 0.12% of the membrane thickness. This is because the porosity in the vicinity of the surface in contact with the liquid to be filtered is important for achieving high filtration performance. This is because the surface in contact with the liquid to be filtered has the highest concentration of membrane fouling, and pore clogging occurs, which affects the performance of the entire membrane.

[0030] The porous membrane of this embodiment preferably has a porosity of 30% or more on the surface facing the liquid to be filtered. When the porosity is 30% or more, it can have high filtration performance. It is presumed that when the porosity is high, the load of membrane fouling per hole is small and the number of holes that are completely blocked is small, so that high filtration performance can be achieved. Preferably, the porosity is 32% or more, more preferably 35% or more. Also, the porosity may be 60% or less.

[0031] The porous membrane of the present embodiment preferably has a porosity of 50% or more, more preferably 60% or more, in a thickness of 0.12% of the membrane thickness from the surface on the side of the liquid to be filtered. When the porosity is 50% or more, high filtration performance can be exhibited for a wider range of properties of the liquid to be filtered. It is estimated that when the porosity is 50% or more, the load of membrane fouling per hole is small, and the number of holes that are completely blocked is small, as with the porosity, so that high filtration performance can be exhibited. It is preferably 50% or more and 95% or less, more preferably 55% or more and 85% or less, and even more preferably 60% or more and 80% or less. When the porosity is 95% or less, it can have sufficient strength for practical use. The porous membrane of this embodiment preferably has a porosity of 30% or more on the surface facing the liquid to be filtered, and a porosity of 50% or more in a thickness of 0.12% of the membrane thickness from the outermost surface of the surface facing the liquid to be filtered.

[0032] The porous membrane of this embodiment preferably has a cross-sectional pore size of 1000 nm or less in a thickness that is 0.12% of the membrane thickness from the outermost surface on the side of the liquid to be filtered. The cross-sectional pore diameter is preferably 100 nm or more and 900 nm or less, and more preferably 200 nm or more and 800 nm or less. If the cross-sectional pore diameter is 1000 nm or less, sufficient blocking performance can be obtained for practical use. In the porous membrane of this embodiment, the product of the porosity in a thickness from the surface on the liquid to be filtered to 0.12% of the membrane thickness and the porosity of the surface on the liquid to be filtered is preferably 1800%·%. It is preferably 2000%·% or more, and more preferably 2700%·% or more. It is estimated that when the product is 1800%·% or more, the load of membrane fouling per hole is small both on the surface and in the thickness direction, and therefore the number of completely blocked holes is extremely small, resulting in high filtration performance. It may also be 5000%·% or less.

[0033] In the porous membrane of this embodiment, the product of the porosity in a thickness of 300 nm from the filtrate side surface to the membrane thickness and the porosity of the filtrate side surface is preferably 860%·%. It is preferably 875%·% or more. It is presumed that when the product is 860%·% or more, the load of membrane fouling per pore is small both on the surface and in the thickness direction, and therefore the number of completely blocked pores is extremely small, resulting in high filtration performance. It may also be 5000%·% or less.

[0034] The porous membrane of the present embodiment preferably has a porosity of 30% or more, more preferably 35% or more, in a thickness of 300 nm from the filtrate side surface to the membrane thickness. If the porosity is 50% or more, high filtration performance can be achieved for a wider range of filtrate properties. It is estimated that when the porosity is 30% or more, the load of membrane fouling per hole is small, and the number of holes that are completely blocked is small, as with the porosity, so that high filtration performance can be exhibited. It is preferably 35% or more and 85% or less, and more preferably 35% or more and 80% or less. When the porosity is 85% or less, it can have sufficient strength for practical use.

[0035] The porous membrane of the present embodiment preferably has a cross-sectional pore size of 300 nm or less in a thickness of 300 nm from the outermost surface on the filtrate side to the membrane thickness. The cross-sectional pore diameter is preferably 50 nm or more and 250 nm or less. If the cross-sectional pore diameter is 300 nm or less, sufficient blocking performance can be obtained for practical use.

[0036] When the porous membrane of this embodiment is a hollow fiber membrane, the inner diameter is preferably 0.3 mm or more and 5 mm or less. If the inner diameter is 0.3 mm or more, the pressure loss of the liquid flowing through the hollow fiber membrane is not too large, and if the inner diameter is 5 mm or less, sufficient compressive strength and burst strength are easily achieved with a relatively thin membrane thickness. The inner diameter is more preferably 0.4 mm or more and 3 mm or less, and even more preferably 0.5 mm or more and 2 mm or less.

[0037] The membrane thickness is preferably 0.1 mm or more and 1 mm or less. If the membrane thickness is 0.1 mm or more, sufficient compressive strength and burst strength are easily exhibited, and if the membrane thickness is 1 mm or less, sufficient water permeability is easily exhibited. The membrane thickness is preferably 100 μm or more and 800 μm or less, more preferably 0.15 mm or more and 0.8 mm or less, even more preferably 0.16 mm or more and 0.6 mm or less, 0.17 mm or more and 0.5 mm or less, and also preferably 0.1 mm or more and 0.5 mm or less.

[0038] The outer diameter is preferably 0.5 mm or more and 5 mm or less. If the outer diameter is 0.5 mm or more, sufficient tensile strength can be obtained. If the outer diameter is 5 mm or less, the number of porous membranes (preferably porous hollow fiber membranes) packed in a container can be increased. The outer diameter is more preferably 0.6 mm or more and 4 mm or less, and further preferably 0.7 mm or more and 3 mm or less.

[0039] The pure water permeability of the porous membrane of this embodiment is 1000 L / m 2 / hr or more 20000L / m 2 When the pure water permeability is in this range, it is possible to achieve both filtration performance and blocking performance. The pure water permeability is preferably 1200 L / m 2 / hr or more 18000L / m2 / hr or less, and more preferably 3000 L / m 2 / hr or more 12000L / m 2 / hr or less.

[0040] The flux of the porous membrane of this embodiment is preferably 1 LMH or more.

[0041] The porosity of the entire porous membrane of the present embodiment is preferably 50% or more from the viewpoint of water permeability, and is preferably 90% or less from the viewpoint of strength, more preferably 55% or more and 85% or less, and even more preferably 65% ​​or more and 80% or less. The present inventors have found that factors including the porosity of the extreme surface layer, the difference of which cannot be detected by measuring the porosity of the entire membrane, affect the filtration performance. Specifically, they have found that the filtration performance can be improved by controlling the ratio of the porosity in a thickness from the outermost surface of the surface to be filtrated to 0.12% of the membrane thickness to the pore ratio of the surface to be filtrated, and the product of the porosity in a thickness from the outermost surface of the surface to be filtrated to 0.12% of the membrane thickness and the pore ratio of the surface to be filtrated.

[0042] The porous membrane (preferably a porous hollow fiber membrane) preferably has a three-dimensional mesh structure. The three-dimensional mesh structure in the present application refers to a structure as shown in FIG. 1. For example, thermoplastic resin a is bonded to form a mesh, and voids b are formed. In the three-dimensional mesh structure, so-called spherulite structure resin aggregates are hardly seen. It is preferable that the voids b of the three-dimensional mesh structure are surrounded by the thermoplastic resin a, and each part of the voids b is connected to each other. Since most of the thermoplastic resins used form a three-dimensional mesh structure that can contribute to the strength of the porous membrane (preferably a hollow fiber membrane), it is possible to form a support layer with high strength. In addition, chemical resistance is also improved. The reason for the improvement in chemical resistance is not clear, but it is thought that this is because there is a large amount of thermoplastic resin that forms a mesh that can contribute to strength, so even if a part of the mesh is corroded by a chemical, the strength of the layer as a whole is not significantly affected. The porous membrane (preferably a porous hollow fiber membrane) may have a single-layer structure or a multi-layer structure of two or more layers. The layer having the surface facing the liquid to be filtered is referred to as layer (A), and the layer having the surface facing the filtrate is referred to as layer (B). For example, layer (A) is a so-called blocking layer, which has a small surface pore size and functions to prevent foreign matter contained in the liquid to be treated (raw water) from passing through the membrane, and layer (B) is a so-called supporting layer, which has a function of ensuring high mechanical strength and preventing the water permeability from decreasing as much as possible. The functions of layer (A) and layer (B) are not limited to the above. In the porous membrane of this embodiment, only one surface may be the surface on the side of the liquid to be filtered.

[0043] The following describes a two-layer structure in which layer (A) is a blocking layer and layer (B) is a supporting layer. The thickness of layer (A) is preferably 1 / 100 or more and less than 40 / 100 of the membrane thickness. By making layer (A) relatively thick in this way, it can be used even if the raw water contains insoluble matter such as sand and aggregates. This is because the surface pore size does not change even if it is slightly worn. If the thickness is within this range, a balance between desirable blocking performance and high water permeability can be achieved. More preferably, it is 2 / 100 or more and 30 / 100 or less of the membrane thickness. The thickness of layer (A) is preferably 1 μm or more and 100 μm or less, and more preferably 2 μm or more and 80 μm or less.

[0044] The filtration step in the filtration method of this embodiment may be, for example, a so-called internal pressure filtration step in which a culture broth (liquid to be treated) containing cells, a culture medium, useful substances, an antifoaming agent, etc. is supplied to the hollow part (inner surface) of a porous hollow fiber membrane, passed through the membrane thickness (thick wall) part of the porous hollow fiber membrane, and the liquid that seeps out from the outer surface of the porous hollow fiber membrane is taken out as filtrate, or a so-called external pressure filtration step in which the liquid to be treated is supplied from the outer surface of the porous hollow fiber membrane, and the filtrate that seeps out from the inner surface of the porous hollow fiber membrane is taken out via the hollow part. In this specification, the term "inside the porous membrane" refers to the membrane thickness (thick wall) portion where numerous pores are formed.

[0045] A specific production method for a hollow fiber membrane will be described. A method for producing a porous membrane (preferably a porous hollow fiber membrane) of this embodiment preferably includes a step of discharging a molten mixture containing a thermoplastic resin, an organic liquid, and an inorganic fine powder from a spinneret having an annular discharge port to form a hollow fiber-shaped molten mixture, and a step of solidifying the hollow fiber-shaped molten mixture and then extracting and removing the organic liquid and the inorganic fine powder to produce a porous membrane (preferably a porous hollow fiber membrane). The molten mixture may be composed of two components, a thermoplastic resin and a solvent, or may be composed of three components, a thermoplastic resin, an inorganic fine powder, and a solvent.

[0046] The thermoplastic resin used in the manufacturing method of the porous membrane (preferably a porous hollow fiber membrane) of this embodiment is a resin that has elasticity and does not show plasticity at room temperature, but shows plasticity and becomes moldable when appropriately heated. Furthermore, the thermoplastic resin is a resin that returns to its original elastic body when cooled and the temperature is reduced, and does not undergo chemical changes such as molecular structure during this time (see, for example, "Chemical Encyclopedia 6, Condensed Edition, Edited by the Editorial Committee for the Chemical Encyclopedia, Kyoritsu Shuppan, pp. 860 and 867, 1963").

[0047] Examples of thermoplastic resins include those listed in the section on thermoplastics (pages 829-882) of Chemical Products No. 12695 (The Chemical Daily, 1995) and those listed on pages 809-810 of the revised Chemical Handbook, Applied Edition, 3rd Edition (edited by the Chemical Society of Japan, Maruzen, 1980). Specific examples of thermoplastic resins include polyolefins such as polyethylene and polypropylene, fluororesins such as polyvinylidene fluoride, ethylene-vinyl alcohol copolymers, polyamides, polyetherimides, polystyrene, polysulfone, polyvinyl alcohol, polyphenylene ether, polyphenylene sulfide, cellulose acetate, polyacrylonitrile, etc. Among them, crystalline polyolefins, fluororesins such as polyvinylidene fluoride, ethylene-vinyl alcohol copolymers, polyvinyl alcohol, and other crystalline thermoplastic resins can be preferably used in terms of strength development. More preferably, fluororesins such as polyolefins and polyvinylidene fluoride, which are hydrophobic and therefore highly water-resistant, and can be expected to have durability in filtering normal aqueous liquids, can be used. Specifically, the fluororesins are preferably vinylidene fluoride resins (PVDF), chlorotrifluoroethylene resins, tetrafluoroethylene resins, ethylene-tetrafluoroethylene copolymers (ETFE), ethylene-monochlorotrifluoroethylene copolymers (ECTFE), hexafluoropropylene resins, and mixtures of these resins, either alone or in combination of two or more, as the main components, and more preferably comprise only resins consisting of either alone or in combination of two or more. Particularly preferably, the fluororesins can be polyvinylidene fluoride, which has excellent chemical durability such as chemical resistance. Examples of polyvinylidene fluoride include vinylidene fluoride homopolymers and vinylidene fluoride copolymers with a vinylidene fluoride ratio of 50 mol % or more. The vinylidene fluoride copolymer may be a copolymer of vinylidene fluoride and one or more monomers selected from the group consisting of tetrafluoroethylene, hexafluoropropylene, trifluorochloroethylene, and ethylene. As the polyvinylidene fluoride, vinylidene fluoride homopolymer is particularly preferred.

[0048] The concentration of the thermoplastic resin in the melt-kneaded product is desirably 30% by mass to 48% by mass. It is preferably 32% by mass to 45% by mass. If it is 30% by mass or more, mechanical strength is easily ensured, and if it is 48% by mass or less, there is no decrease in water permeability.

[0049] When the porous membrane has a two-layer structure, the concentration of the thermoplastic resin in the molten mixture of layer (B) is preferably 34 to 48% by mass, and more preferably 35 to 45% by mass. The concentration of the thermoplastic resin in the molten mixture of layer (A) is preferably 10% by mass or more and 35% by mass or less, more preferably 12% by mass or more and less than 35% by mass. If it is 10% by mass or more, both the pore size and the mechanical strength of the surface can be achieved, and if it is 35% by mass or less, the water permeability is not reduced.

[0050] The organic liquid used in this embodiment is a latent solvent for the thermoplastic resin used in this embodiment. In this embodiment, the latent solvent refers to a solvent that hardly dissolves the thermoplastic resin at room temperature (25°C) but can dissolve the thermoplastic resin at a temperature higher than room temperature. It is sufficient that the solvent is liquid at the melt-kneading temperature with the thermoplastic resin, and it is not necessarily liquid at room temperature.

[0051] When the thermoplastic resin is polyethylene or polypropylene, examples of organic liquids include phthalate esters such as dibutyl phthalate, diheptyl phthalate, dioctyl phthalate, di(2-ethylhexyl) phthalate, diisodecyl phthalate, ditridecyl phthalate, etc.; sebacate esters such as dibutyl sebacate; adipate esters such as dioctyl adipate; trimellitate esters such as trioctyl trimellitate; phosphate esters such as tributyl phosphate, trioctyl phosphate, etc.; glycerin esters such as propylene glycol dicaprate, propylene glycol dioleate, etc.; paraffins such as liquid paraffin; and mixtures thereof.

[0052] When the thermoplastic resin is polyvinylidene fluoride, examples of the organic liquid include phthalate esters such as dimethyl phthalate, diethyl phthalate, dibutyl phthalate, dicyclohexyl phthalate, diheptyl phthalate, dioctyl phthalate, and di(2-ethylhexyl) phthalate; sebacate esters such as dibutyl sebacate; adipate esters such as dioctyl adipate; benzoate esters such as methyl benzoate and ethyl benzoate; phosphate esters such as triphenyl phosphate, tributyl phosphate, and tricresyl phosphate; ketones such as γ-butyrolactone, ethylene carbonate, propylene carbonate, cyclohexanone, acetophenone, and isophorone; and mixtures thereof.

[0053] When the thermoplastic resin is a polyketone or ethylene vinyl alcohol, examples of the organic liquid include phthalates such as dimethyl phthalate, diethyl phthalate, dibutyl phthalate, dicyclohexyl phthalate, diheptyl phthalate, dioctyl phthalate, and di(2-ethylhexyl) phthalate; sebacates such as dibutyl sebacate; adipates such as dioctyl adipate; trimellitates such as trioctyl trimellitate; benzoates such as methyl benzoate and ethyl benzoate; phosphates such as triphenyl phosphate, tributyl phosphate, and tricresyl phosphate; glycerin esters such as propylene glycol dicaprate and propylene glycol dioleate; polyethers such as polyethylene glycol, paraffins such as liquid paraffin; ketones such as γ-butyrolactone, ethylene carbonate, propylene carbonate, cyclohexanone, acetophenone, and isophorone, glycerin; and mixtures thereof.

[0054] Examples of inorganic fine powder include silica, alumina, titanium oxide, zirconia oxide, calcium carbonate, etc., and particularly, fine silica having an average primary particle diameter of 3 nm to 500 nm is preferred. More preferably, it is 5 nm to 100 nm. Hydrophobic silica fine powder that is less likely to aggregate and has good dispersibility is more preferred, and even more preferably, hydrophobic silica having a MW (methanol wettability) value of 30 volume % or more is preferred. The MW value here is the volume % value of methanol that completely wets the powder. Specifically, silica is placed in pure water, and methanol is added below the liquid surface while stirring, and the volume % of methanol in the aqueous solution is determined when 50 mass % of the silica has settled. The above-mentioned "average primary particle diameter of inorganic fine powder" means a value obtained by analyzing an electron microscope photograph. That is, first, a group of inorganic fine powder is pretreated by the method of ASTM D3849. Then, the particle diameters of 3000 to 5000 particles photographed in a transmission electron microscope photograph are measured, and the average primary particle diameter of the inorganic fine powder is calculated by arithmetically averaging these values.

[0055] The amount of inorganic fine powder added is preferably 5% by mass or more and 50% by mass or less, and more preferably 10% by mass or more and 40% by mass or less, in terms of the mass ratio of the inorganic fine powder in the molten kneaded product. If the ratio of the inorganic fine powder is 5% by mass or more, the effect of kneading the inorganic fine powder can be fully exhibited, and if it is 40% by mass or less, stable spinning can be achieved.

[0056] The melt kneading can be carried out using a normal melt kneading means, for example, an extruder. Although the case where an extruder is used will be described below, the melt kneading means is not limited to an extruder. An example of a production apparatus used to carry out the production method of this embodiment is shown in FIG. 2.

[0057] The porous hollow fiber membrane manufacturing apparatus shown in Fig. 2 includes an extruder 10, a hollow fiber forming nozzle 20, a coagulation bath 30 in which a solution for coagulating the membrane-forming stock solution is stored, and a plurality of rollers 50 for transporting and winding up the porous hollow fiber membrane 40. Reference numeral 60 denotes a suction machine, and 70 denotes a high-temperature container. The space S shown in Fig. 2 is an empty run section through which the membrane-forming stock solution discharged from the hollow fiber forming nozzle 20 passes before reaching the solution in the coagulation bath 30.

[0058] The molten mixture is extruded by the extruder 10 through the hollow fiber molding nozzle 20, which is attached to the tip of the extruder 10 and has one or more annular discharge ports arranged in a concentric circle. When producing a membrane with a multilayer structure, there is a method in which the hollow fiber molding nozzle 20 having two or more annular discharge ports is attached to the tip of the extruder 10, and the molten mixture is supplied from different extruders 10 to each annular discharge port and extruded, or a method in which one layer of the multilayer structure is produced and then the remaining layer is coated. For example, in the former method of production using different extruders, the molten mixtures supplied are merged at the discharge ports and overlapped to obtain a hollow fiber extrudate having a multilayer structure. At this time, by extruding molten mixtures having different compositions from adjacent annular discharge ports, a multilayer membrane in which the pore sizes of adjacent layers are different can be obtained. The different compositions refer to the case where the constituent substances of the molten mixture are different, or the case where the constituent substances are the same but have different composition ratios. Even if the thermoplastic resins are the same type, if they have clearly different molecular weights or molecular weight distributions, they are considered to have different constituent substances. The joining position of the molten mixtures having different compositions may be at the lower end surface of the hollow fiber molding nozzle 20 or may be at a position other than the lower end surface of the hollow fiber molding nozzle 20.

[0059] When extruding the molten mixture from the annular discharge port, it is preferable to discharge the mixture so that the spinneret discharge parameter R (1 / sec) is 10 or more and 1000 or less, since this results in high productivity, stable spinning, and a membrane with high strength. Here, the spinneret discharge parameter R is the value obtained by dividing the discharge linear velocity V (m / sec) by the slit width d (m) of the discharge port. The discharge linear velocity V (m / sec) is the discharge volume (m 3 / sec) to the cross-sectional area of ​​the outlet (m 2 ) When R is 10 or more, there is no problem such as pulsation of the fiber diameter of the hollow extrudate, and the hollow fiber can be spun stably with good productivity. When R is 1000 or less, the breaking elongation, which is one of the important strengths of the obtained porous hollow fiber membrane, can be maintained sufficiently high. The breaking elongation is the elongation rate relative to the original length when the membrane is pulled in the longitudinal direction.

[0060] In the case of a multi-layered porous hollow fiber membrane, the linear extrusion speed V of the laminated molten mixture after the resins join is divided by the slit width d of the extrusion port to obtain the spinneret extrusion parameter R. The range of R is more preferably 50 or more and 1000 or less.

[0061] The hollow fiber-shaped molten kneaded material discharged from the discharge port is solidified by passing through a coolant such as air or water, and then passes through the above-mentioned free-running section S consisting of an air layer due to the intended porous hollow fiber membrane, and then passes through the solidification bath 30 containing water or the like. In other words, the free-running section S is the section from the discharge port of the hollow fiber molding nozzle 20 to the water surface of the solidification bath 30. A container such as a cylinder may be used for the free-running section S from the discharge port as necessary. After passing through the solidification bath 30, the material is wound into a skein or the like as necessary.

[0062] In particular, it is preferable to pass the molten mixture through a high-temperature container such as a cylindrical container for 0.015 seconds or more immediately after the discharge port. If it is 0.015 seconds or more, the solvent vapor accumulated in the high-temperature container suppresses the surface, i.e., the pores, from closing, and furthermore, the resin concentration of the surface layer of the molten mixture is reduced by absorbing the solvent vapor, so that the porosity and the porosity near the surface can be increased. In addition, it was found that the inner surface side of the hollow fiber-shaped molten mixture can also be increased by the heat retention effect of the high-temperature container and the solvent vapor generated from the molten mixture due to the high temperature inside the container. In order to control the pore size of the surface, it is preferable for it to be 1.0 seconds or less. More preferably, it is 0.018 seconds or more and 0.90 seconds or less, and even more preferably, it is 0.018 seconds or more and 0.80 seconds or less.

[0063] Generally, in non-solvent induced phase separation, a cylinder is used for the entire empty runway, because the moisture in the empty runway promotes phase separation. In the present invention, however, a new finding has been discovered that the open area and porosity of the surface can be controlled by solvent vapor.

[0064] The set temperature of the high-temperature container is preferably from (T-60)°C to (T+60)°C relative to the discharge temperature T of the molten kneaded product. More preferably, it is from (T-50)°C to (T+50)°C. If it is (T-60)°C or higher, the effect of the solvent vapor can be fully exerted. Although there is no particular reason, it is preferably (T+60)°C or lower in order to prevent deterioration of the resin kneaded product by excessively increasing the set temperature.

[0065] The time during which the molten mixture passes through the free-running section S is called the free-running time, and the free-running time is preferably 0.20 seconds or more. If the free-running time is 0.20 seconds or more, the polymer molecules are oriented in the free-running section, and the compression resistance strength can be further increased. The free-running time is more preferably 0.20 seconds or more and 5.0 seconds or less. If it is 5.0 seconds or less, stable production is possible. The free-running time is preferably 0.25 seconds or more and 4.0 seconds or less, and more preferably 0.30 seconds or more and 3.5 seconds or less.

[0066] In the free-running section, it is desirable to use a suction machine or the like to apply cooling air perpendicular to the discharge direction at 0.80 m / sec or less. The reason for this is unclear, but it is presumed that at 0.80 m / sec or less, the solvent vapor is moderately retained on the surface of the molten kneaded product, which prevents the solvent vapor from closing the surface, i.e., the pores, and furthermore, the resin concentration in the surface layer of the molten kneaded product decreases as the solvent vapor is absorbed, which increases the open area ratio and porosity near the surface.

[0067] In the hollow fiber after solidification, the polymer-rich phase and the organic liquid-rich phase are finely separated. For example, when inorganic fine powder is added, if the inorganic fine powder is fine silica, the fine silica is unevenly distributed in the organic liquid-rich phase. By extracting and removing the organic liquid and the inorganic fine powder from the hollow fiber, the organic liquid-rich phase becomes pores. Thus, a porous hollow fiber membrane can be obtained.

[0068] The organic liquid and inorganic fine powder can be extracted and removed simultaneously if they can be extracted and removed with the same solvent. Usually, they are extracted and removed separately.

[0069] The organic liquid is extracted and removed using a liquid suitable for extraction that does not dissolve or denature the thermoplastic resin used and is miscible with the organic liquid. Specifically, the liquid can be contacted by a technique such as immersion. The liquid is preferably volatile so that it can be easily removed from the hollow fiber membrane after extraction. Examples of the liquid include alcohols and methylene chloride. If the organic liquid is water-soluble, water can also be used as the extraction liquid.

[0070] The inorganic fine powder is usually extracted and removed using an aqueous liquid. For example, when the inorganic fine powder is silica, the inorganic fine powder is first contacted with an alkaline solution to convert the silica into a silicate, and then contacted with water to extract and remove the silicate.

[0071] It does not matter whether the organic liquid is extracted and removed first or the inorganic fine powder is extracted and removed first. If the organic liquid is immiscible with water, it is preferable to extract and remove the organic liquid first, and then extract and remove the inorganic fine powder. Since the organic liquid and the inorganic fine powder are usually mixed and coexist in the organic liquid-rich partial phase, this is advantageous in that the inorganic fine powder can be extracted and removed smoothly.

[0072] In this manner, the organic liquid and inorganic fine powder are extracted and removed from the solidified porous hollow fiber membrane, thereby obtaining a porous hollow fiber membrane. In addition, the hollow fiber membrane after solidification can be stretched in the longitudinal direction at a stretching ratio of 3 times or less at any of the following stages: (i) before the organic liquid and inorganic fine powder are extracted and removed, (ii) after the organic liquid is extracted and removed but before the inorganic fine powder is extracted and removed, (iii) after the inorganic fine powder is extracted and removed but before the organic liquid is extracted and removed, and (iv) after the organic liquid and inorganic fine powder are extracted and removed. Generally, when a hollow fiber membrane is stretched in the longitudinal direction, the water permeability is improved, but the pressure resistance (burst strength and compressive strength) is reduced, so that the membrane does not have a practical strength after stretching in many cases. However, the porous hollow fiber membrane obtained by the manufacturing method of this embodiment has high mechanical strength. Therefore, stretching at a stretching ratio of 1.1 times or more and 3.0 times or less is possible. The water permeability of the porous hollow fiber membrane is improved by stretching. The stretching ratio referred to here refers to the value obtained by dividing the hollow fiber length after stretching by the hollow fiber length before stretching. For example, when a porous hollow fiber membrane having a hollow fiber length of 10 cm is stretched to a length of 20 cm, the stretching ratio is 2 times according to the following formula. 20cm÷10cm=2

[0073] The stretching is preferably carried out at a space temperature of 0° C. to 160° C. If the temperature is higher than 160° C., the stretching unevenness will be large and the elongation at break and water permeability will decrease, which is undesirable, while if the temperature is lower than 0° C., the possibility of stretching breakage is high and this is not practical. It is more preferable to set the space temperature during the stretching process to 10° C. to 140° C., and even more preferably 20° C. to 100° C.

[0074] In this embodiment, it is preferable to stretch a hollow fiber membrane containing an organic liquid. A hollow fiber membrane containing an organic liquid is less likely to break during stretching than a hollow fiber membrane not containing an organic liquid. Furthermore, a hollow fiber membrane containing an organic liquid can shrink the hollow fiber membrane more after stretching, which increases the degree of freedom in setting the shrinkage rate after stretching.

[0075] It is also preferable to stretch a hollow fiber membrane containing inorganic fine powder. The hollow fiber membrane containing inorganic fine powder is less likely to be flattened during stretching due to the hardness of the hollow fiber membrane caused by the inorganic fine powder contained therein. This also makes it possible to prevent the pore size of the hollow fiber membrane finally obtained from becoming too small or the fiber diameter from becoming too thin. In this embodiment, it is more preferable to stretch a hollow fiber membrane containing both an organic liquid and an inorganic fine powder.

[0076] For the reasons described above, it is preferable to stretch a hollow fiber membrane containing either an organic liquid or an inorganic fine powder rather than stretching the hollow fiber membrane after the extraction is completed, and furthermore, it is more preferable to stretch a hollow fiber membrane containing both an organic liquid and an inorganic fine powder rather than stretching a hollow fiber membrane containing either an organic liquid or an inorganic fine powder.

[0077] The method of extracting a stretched hollow fiber membrane has the advantage that the number of voids on the surface and inside of the hollow fiber membrane is increased by the stretching, so that the extraction solvent can easily penetrate into the inside of the hollow fiber membrane. The method of performing extraction after the step of stretching and then shrinking the hollow fiber membrane has the advantage that, since the hollow fiber membrane has a low tensile modulus and is easily bent, when the extraction is performed in a liquid flow, the hollow fiber membrane is easily shaken by the liquid flow, and the stirring effect is increased, so that efficient extraction can be performed in a short time.

[0078] In this embodiment, the hollow fiber membrane is stretched and then contracted, so that a hollow fiber membrane with a low tensile modulus can be finally obtained. Here, "low tensile modulus" means that the threads are easily stretched with a small force and return to their original shape when the force is removed. If the tensile modulus is low, the hollow fiber membrane does not collapse flat, but is easily bent, and is easily swayed by the water flow during filtration. The bend of the threads is not constant and swayed according to the water flow, so that the layer of contaminants that adhere to and accumulate on the membrane surface is easily peeled off without growing, and the amount of filtrate can be maintained high. Furthermore, when the threads are forcibly swayed by flushing or air scrubbing, the shaking is large and the cleaning recovery effect is high.

[0079] Regarding the degree of shrinkage of the yarn length when shrinking after drawing, it is desirable to set the yarn length shrinkage ratio to the increment in yarn length due to drawing in the range of 0.3 to 0.9. For example, when a 10 cm yarn is drawn to 20 cm and then drawn to 14 cm, the following formula is obtained: Yarn length shrinkage rate = {(maximum yarn length when stretched) - (yarn length after shrinkage)} / [(maximum yarn length when stretched) - (original yarn length)] = (20 - 14) / (20 - 10) = 0.6 The fiber length shrinkage is 0.6. If the fiber length shrinkage is 0.9 or more, the water permeability is likely to be low, and if it is less than 0.3, the tensile modulus is likely to be high, which is not preferable. In the present invention, it is more preferable that the fiber length shrinkage is in the range of 0.50 to 0.85.

[0080] In addition, by employing a process of stretching the hollow fiber membrane to the maximum fiber length when stretched and then shrinking it, the final hollow fiber membrane will not break even when stretched to the maximum fiber length during use. Here, when the draw ratio is X and the yarn length shrinkage rate relative to the yarn length increment due to drawing is Y, the rate Z, which represents the degree of guarantee of breaking elongation, can be defined by the following formula. Z = (Maximum yarn length during stretching - yarn length after shrinkage) / yarn length after shrinkage = (XY-Y) / (X+Y-XY) Z is preferably 0.2 or more and 1.5 or less, and more preferably 0.3 or more and 1.0 or less. If Z is too small, the guarantee of breaking elongation is reduced, and if Z is too large, the possibility of breakage during stretching increases, but the water permeability decreases.

[0081] In addition, since the manufacturing method of this embodiment includes a step of stretching and then shrinking, the tensile breaking elongation is extremely low in breaks at low elongations, and the distribution of tensile breaking elongation can be narrowed.

[0082] The space temperature in the process of stretching and then shrinking is preferably in the range of 0° C. to 160° C. in terms of shrinkage time and physical properties. If it is lower than 0° C., shrinkage takes too long and is not practical, while if it exceeds 160° C., it is not preferable because the breaking elongation and water permeability are reduced.

[0083] In this embodiment, it is also preferable to crimp the hollow fiber membrane during the shrinking step, which makes it possible to obtain a hollow fiber membrane with a high degree of crimping without crushing or damaging the hollow fiber membrane.

[0084] Generally, hollow fiber membranes are straight and unbent, so when they are bundled together to form a filtration module, there is a high possibility that the hollow fibers will not have gaps between them, resulting in a fiber bundle with a low porosity. In contrast, when hollow fiber membranes with a high degree of crimping are used, the individual fibers will bend, leading to wider hollow fiber membrane spacing on average, resulting in a fiber bundle with a high porosity. Furthermore, when a filtration module is made of hollow fiber membranes with a low degree of crimping, the gaps in the fiber bundle are reduced, particularly when used under external pressure, increasing flow resistance, and the filtration pressure is not effectively transmitted to the center of the fiber bundle. Furthermore, the cleaning effect of the inside of the fiber bundle is reduced when filtration deposits are peeled off from the hollow fiber membranes by backwashing or flushing. The degree of crimping A fiber bundle made of high-density hollow fiber membranes has a large porosity, so the gaps between the hollow fiber membranes are maintained even during external pressure filtration, making it less likely that drift will occur.

[0085] In this embodiment, the crimp degree is preferably in the range of 1.5 to 2.5. A crimp degree of 1.5 or more is preferable for the reasons described above, and a crimp degree of less than 2.5 can suppress a decrease in the filtration area per volume.

[0086] Methods for crimping hollow fiber membranes include a method in which the hollow fiber membrane is stretched and then shrunk during the process, and the membrane is taken up while being sandwiched between a pair of gear rolls having periodic projections and recesses or a pair of sponge belts having projections and recesses while being shrunk.

[0087] In this embodiment, the drawing is preferably performed using a take-up machine consisting of a pair of opposing caterpillar belts. In this case, take-up machines are used on the upstream and downstream sides of the drawing, and in each take-up machine, the hollow fiber membrane is sandwiched between the opposing belts, and the two belts are moved in the same direction at the same speed to feed the yarn. In this case, it is preferable to perform drawing by setting the yarn feed speed on the downstream side faster than that on the upstream side. By performing drawing in this manner, it is possible to draw the yarn without slipping due to the drawing tension during drawing, and to prevent the yarn from being crushed flat.

[0088] Here, the endless track belt preferably has an inner surface that contacts the driving roll and is made of a highly elastic belt such as a fiber-reinforced belt, and an outer surface that contacts the hollow fiber membrane and is made of an elastic body. It is more preferable that the compressive elastic modulus in the thickness direction of the elastic body is 0.1 MPa or more and 2 MPa or less, and the thickness of the elastic body is 2 mm or more and 20 mm or less. In particular, it is preferable that the elastic body on the outer surface is made of silicone rubber from the viewpoint of chemical resistance and heat resistance.

[0089] If necessary, the stretched membrane may be heat-treated to increase the compressive strength. The heat treatment is preferably performed at 80°C or higher and 160°C or lower. If the temperature is 160°C or lower, the decrease in breaking elongation and water permeability can be suppressed, and if the temperature is 100°C or higher, the compressive strength can be increased. It is also preferable to perform the heat treatment on the hollow fiber membrane after the extraction is completed, since this reduces the changes in fiber diameter, porosity, pore size, and water permeability.

[0090] When using PVDF (polyvinylidene fluoride) as a thermoplastic resin, it is necessary to select an appropriate solvent for PVDF in order to achieve both a high porosity and high compressive strength. First, there are two ways to increase the porosity: lowering the PVDF concentration, or increasing the temperature of the fluid used to form the hollow section, as mentioned above. When using a method of forming a membrane by lowering the PVDF concentration, the pore size also increases, so it is necessary to select a solvent that can achieve a high porosity and a small pore size. The following parameter P is a relational expression between the three-dimensional solubility parameter of PVDF and the three-dimensional solubility parameter of the solvent, and is used to evaluate the solubility of PVDF and the solvent. The right side represents the solubility range of the Hansen solubility parameters three-dimensionally, and quantitatively represents the distance from the three-dimensional solubility parameters of PVDF (σdp, σpp, σhp) to the three-dimensional solubility parameters of the solvent (σdm, σpm, σhm). P = ((σdm-σdp) 2 +(σpm-σpp) 2 +(σhm-σhp) 2 ) 0.5 [In the formula, σdm and σdp represent the dispersion force terms of the solvent and polyvinylidene fluoride, respectively, σpm and σpp represent the dipole binding force terms of the solvent and polyvinylidene fluoride, respectively, and σhm and σhp represent the hydrogen bond terms of the solvent and polyvinylidene fluoride, respectively.] The above concept is not limited to PVDF.

[0091] In the case of a two-layer porous membrane, the parameter P between the solvent used to prepare the melt-kneaded material B forming the layer (B) and PVDF is preferably greater than 7.88, more preferably from 7.88 to 10.0. If this value is 7.88 or more, the decrease in water permeability can be suppressed.

[0092] In preparing the melt-kneaded material A that forms the layer (A), the parameter P between the solvent used and PVDF is preferably 7.88, more preferably 0 to 7.88, and even more preferably 1.00 to 7.88. When this value is 7.88 or less, a high aperture ratio and a small pore size can be achieved. EXAMPLES

[0093] Hereinafter, the present embodiment will be described more specifically with reference to examples and comparative examples, but the present embodiment is not limited to these examples.

[0094] The measurement method used in this embodiment is as follows. Unless otherwise specified, all of the following measurements were performed at 25° C. Below, the evaluation methods will be explained, followed by a description of the manufacturing methods and evaluation results of the examples and comparative examples. The composition and manufacturing conditions of the membrane, as well as various performance characteristics, are shown in Tables 1 and 2 below.

[0095] (1) Measurement of outer and inner diameters and film thickness (mm) The hollow fiber membrane was cut into thin slices at 15 cm intervals in the vertical direction along the length of the membrane using a razor or similar tool, and the long and short inner diameters and long and short outer diameters of the cross sections were measured using a microscope. The inner and outer diameters were calculated using the following formulas (2) and (3), respectively, and the inner diameter was subtracted from the calculated outer diameter and the result divided by 2 to calculate the membrane thickness. Measurements were taken at 20 points, and the average values ​​were used as the inner diameter, outer diameter, and membrane thickness under those conditions.

number

number

[0096] (2) Pure water permeability (L / m 2 / hr) The hollow fiber membrane was immersed in a 50% by mass aqueous ethanol solution for 30 minutes, and then immersed in water for 30 minutes to wet the hollow fiber membrane. One end of the wet hollow fiber membrane with a length of about 10 cm was sealed, a syringe needle was inserted into the hollow part of the other end, and pure water at 25°C was injected into the hollow part from the syringe needle at a pressure of 0.1 MPa. The amount of pure water permeating to the outer surface was measured, and the pure water permeation flux was determined by the following formula. The effective membrane length here refers to the net membrane length excluding the part where the syringe needle was inserted. In addition, the number of measurements was 10, and the average value was taken as the pure water permeability under each condition.

number

[0097] (3) Breaking strength (MPa), breaking elongation (%) The tensile strength, load and displacement at break were measured under the following conditions. Sample: Wet hollow fiber membrane prepared by method (2) Measuring equipment: Instron type tensile testing machine (Shimadzu Corporation AGS-X) Chuck distance: 5 cm Pulling speed: 20cm / min The breaking strength and breaking elongation were determined according to the following formulas.

number

number

[0098] (4) Porosity of the entire membrane The porosity of the entire membrane is calculated by the following formula: Porosity (whole membrane)% = 100 × (wet membrane weight [g] - dry membrane weight [g]) / water density [g / cm 3 ] / (membrane volume [cm 3 ]) Here, the wet membrane refers to a membrane in a state in which the pores are filled with pure water but the hollow portion does not contain pure water. Specifically, a sample membrane of 10 to 20 cm in length is immersed in ethanol to fill the pores with ethanol, and then the immersion in pure water is repeated 4 to 5 times to fully replace the pores with pure water, and then one end of the hollow fiber is held by hand and shaken well about 5 times, and then the other end is held by hand and shaken well again about 5 times to remove the water in the hollow portion. In addition, a dry membrane can be obtained by measuring the weight of the wet membrane and then drying it in an oven at, for example, 60°C until it reaches a constant weight. The membrane volume is calculated according to the following formula: Membrane volume [cm 3 ]=π×{(Outer diameter [cm] / 2)^2-(Inner diameter [cm] / 2)^2}×Membrane length [cm] If the weight of a single membrane is too small and the weight measurement error becomes large, multiple membranes can be used.

[0099] (5) How to determine the boundaries between layers in the case of a multi-layer structure A Hitachi SU8000 series electron microscope was used to observe the cross section of the membrane at an accelerating voltage of 3 kV. In this example and comparative example, the vicinity of the boundary between the layers was photographed at 1000x magnification. If a boundary line between the layers can be distinguished from the photographed image, the boundary line is regarded as the boundary between the layers. In the porous hollow fiber membranes in this example and comparative example, the boundary line can also be distinguished, so the boundary line is regarded as the boundary between the layers.

[0100] If the boundary between layers cannot be determined by the above method, the boundary can also be determined by the following method. For example, a method for determining the boundary between layer (A) and layer (B) in the case of a two-layered porous hollow fiber membrane is described below. The following method is for the case where layer (A) is the blocking layer and layer (B) is the support layer. The cross section of the hollow fiber membrane was photographed by the above-mentioned electron microscope, and a photograph in which the shape of 20 or more holes could be confirmed was used. In order to observe the entire cross section, multiple images were taken. In this example and comparative example, the measurement was performed at 5000 times magnification. The cross-sectional electron microscope sample was obtained by cutting the membrane sample frozen in ethanol into ring slices. Using the commercially available image analysis software Winroof6.1.3, 100 lines L (i.e. lines connecting points with the same film thickness) at equal distances from the surface FA were drawn at intervals that divided the total film thickness into 101 equal parts, as shown in Figure 3A, and the length Lh of the line L crossing the part corresponding to the void part h in the image was measured, as shown in Figure 3B. The average value of the crossing length Lh was calculated by arithmetic averaging to obtain the cross-sectional pore diameter in each film thickness part. If the magnification of the scanning electron microscope photograph is sufficiently high, the lines at equal distances from the surface FA may be approximated by a straight line. The cross-sectional pore diameter in each film thickness part was normalized using the maximum value of the obtained cross-sectional pore diameter, and the first point from the surface FA where the normalized value was closest to 0.7 was determined to be the boundary layer of the layer.

[0101] (6) Three-dimensional solubility parameter The three-dimensional solubility parameters were taken from the following publication: Hansen, Charles (2007). Hansen Solubility Parameters: A user's handbook, Second Edition. Boca Raton, Fla: CRC Press. (ISBN 978-0-8493-7248-3)

[0102] (7) Inner and outer surface pore diameter and porosity The surface of the filter facing the liquid to be filtered was photographed using the same electron microscope as in (5). The images were taken at a magnification that allowed the shapes of 20 or more pores to be confirmed, and in the present example and comparative example, the images were taken at a magnification of 10,000 times. Using the photographed image, for example, as described in International Publication No. 2001 / 53213, a transparent sheet was placed on top of a copy of the image, and the holes were filled in black with a black pen or the like, and the transparent sheet was then copied onto a blank sheet of paper, so that the holes were clearly distinguished as black and the non-holes as white. Then, binarization was performed using discriminant analysis with the commercially available image analysis software Winroof 6.1.3. The area occupied by the binarized image thus obtained was calculated to determine the open area ratios of the surfaces FA and FB. The pore size was determined by calculating the equivalent circle diameter for each hole present on the surface, adding up the pore areas of each hole in order of largest diameter, and determining the pore size as the hole whose sum reached 50% of the sum of the pore areas of all the holes.

[0103] (8) Permeation pressure (kPa) The membrane permeation pressure was calculated as follows: That is, when the inlet pressure to the hollow fiber membrane module is Pi, the outlet pressure is Po, and the permeation side outlet pressure (back pressure) is Pf, the permeation pressure TMP was calculated by the following formula. TMP = (Pi + Po) / 2 - Pf

[0104] (9) Tangential flow linear velocity (m / s) The tangential flow linear velocity Lv was calculated as follows: When the number of hollow fibers introduced in the hollow fiber membrane module is n (pieces), the inner diameter of the hollow fiber membrane is Di (m), and the flow rate of the raw solution supplied to the hollow fiber membrane module is V (m3 / s), the tangential flow linear velocity was calculated by the following formula. Lv=V / (n×(π×(Di / 2)^2))

[0105] (10)Minimum antibody penetration rate The minimum antibody permeability was determined as follows. That is, after the start of culture, the antibody concentrations in the culture medium and the permeate were measured every day and were designated as Cm and Cf, respectively, and the ratio (Cf / Cm) was designated as the antibody permeability. The minimum antibody permeability during the entire operation period was designated as the minimum antibody permeability. The antibody concentration can be determined by known methods such as ELISA and LC-MS. In the method of this embodiment, the antibody concentration was quantified using an automated immunoassay system of xP workstation manufactured by Gyrolab.

[0106] (11) Porosity, polymer skeleton size, and cross-sectional pore size The porous hollow fiber membrane was cut into a circular shape at a cross section perpendicular to the fiber length direction, and then embedded in epoxy resin. After trimming, the sample cross section was subjected to BIB processing to create a smooth cross section, and conductive treatment was performed to prepare a microscopic specimen. A microscopic specimen was prepared for one cut point for each sample. Using a Hitachi SU7000 electron microscope, scanning electron microscope (SEM) images were taken of the cross-sections of the membranes of the prepared specimens. The image acquisition conditions were as follows: Five fields of view including the outer or inner surface of each specimen were imaged. Image acquisition conditions Acceleration voltage: 1 kV Detector: Backscattered electron detector Imaging magnification: 5,0000x (device display magnification) Image resolution: 2560 x 1920 pixels

[0107] ImageJ was used for image analysis. First, Plugins-Bilateral Filter Fiji (10 times under the conditions of spatial radius = 3, range radius = 50) was performed to perform filtering. The filtered SEM image was subjected to threshold processing (Image-Adjust-Treshold: maximum entropy method (MaxEntropy selected)) to binarize the void portion (portion where voids are embedded by embedding resin) and the polymer skeleton portion. Based on the top of the binarized image, the pixel of the membrane part closest to the top of the image was set as the point of 0 nm membrane thickness. Regions of a predetermined thickness (for example, 100 nm thickness, in some cases 50 nm thickness) were continuously cut in the membrane thickness direction, and the porosity, polymer skeleton size, and cross-sectional pore diameter were calculated from each image by the method shown below. Here, for example, the porosity, polymer skeleton size, and cross-sectional pore diameter of the 0-300 nm region were each taken as the arithmetic mean value of the porosity, polymer skeleton size, and cross-sectional pore diameter of the 0-100 nm, 100-200 nm, and 200-300 nm regions that were continuously cut and calculated as above. In addition, for example, the porosity, polymer skeleton size, and cross-sectional pore diameter of the 0-480 nm region were each taken as the value analyzed in the 0-480 nm region. In addition, since the 0-100 nm region is an image including the pores on the top surface of the film, when calculating the porosity and cross-sectional pore size, it is necessary to define the top surface of the film and calculate the numerical value from the binary image of only the pores. In order to obtain a binary image of only the pores, this time, the border of the pores / embedded resin part on the top surface of the film was manually determined using the pencil tool of Adobe Photoshop (registered trademark) Elements 9, and then the embedded resin part was filled in to obtain a binary image of only the pores. A specific example of the work is shown in Figure 4. A line was drawn with the pencil tool to connect both ends of the pores on the top surface of the film on an image with a brightness of 255 (white) for the polymer skeleton and a brightness of 0 for the pores and embedded resin. Next, the embedded resin part was filled in with a brightness of 0 (black) using a fill tool to obtain a binary image of only the pores in the 0-100 nm region. The positions of both ends of the pores on the top surface of the film were determined at the discretion of the operator. Porosity (%): Analyze-Analyze Particles was applied to the binary image of pores (pores are black with brightness 0), and the arithmetic mean of the 5 fields of view in the %area in the Summary was used as the porosity. The settings for Analyze Particles were as follows: Size(Pixel^2):0-infinity Circularity:0-1.00 Summarize: Checked checkbox Exclude on edges: Check box is not checked Include Holes: Check box is not checked Polymer skeleton size: A local thickness image was obtained by applying ImageJ's Plugins-BoneJ-Thickness to the binary image of the membrane (the part corresponding to the polymer is a black image with a brightness of 0). The local thickness image was then analyzed by Analyze-Histogram to obtain detailed local thickness values. The arithmetic mean was calculated from the local thickness values ​​of the five fields of view obtained, and defined as the polymer skeleton size. Cross-sectional pore diameter: A local thickness image was obtained by applying ImageJ's Plugins-BoneJ-Thickness to the binary image of the pore (the area corresponding to the pore is black with a brightness of 0). The local thickness image was then analyzed by Analyze-Histogram to obtain detailed local thickness values. The arithmetic mean was calculated from the local thickness values ​​of the five fields of view obtained, and defined as the cross-sectional pore diameter.

[0108] Example 1 The thermoplastic resin was vinylidene fluoride homopolymer (KF-W#1000 manufactured by Kureha Corporation), the organic liquid was a mixture of di(2-ethylhexyl) phthalate (DEHP) (manufactured by C.G. Ester Corporation) and dibutyl phthalate (DBP) (manufactured by C.G. Ester Corporation), and the inorganic fine powder was finely powdered silica (manufactured by Nippon Aerosil Co., Ltd., product name: AEROSIL-R972, primary particle diameter approximately 16 nm). A hollow fiber molding nozzle was used to perform melt extrusion of a hollow fiber membrane using an extruder. A molten mixture having a composition of vinylidene fluoride homopolymer: di(2-ethylhexyl) phthalate: dibutyl phthalate: finely divided silica = 40.0: 30.8: 6.20: 23.0 (mass ratio) was prepared. The mixture was extruded from a hollow fiber forming nozzle having an outer diameter of 3.0 mm and an inner diameter of 2.0 mm at a discharge temperature of 240°C using air as a hollow portion forming fluid. The hollow fiber-shaped molten kneaded material extruded at a discharge temperature of 240°C passed through a high-temperature container set at a temperature of 240°C for 0.36 seconds, and after 1.50 seconds of air travel including the high-temperature container section, was introduced into a coagulation bath filled with water at 30°C and taken off at a speed of 5.0 m / min. The air speed in the air travel section was 0.80 m / sec. The obtained hollow fiber-shaped material was immersed in isopropyl alcohol to extract and remove di(2-ethylhexyl) phthalate and dibutyl phthalate, and then dried. Next, it was immersed in a 50% by mass aqueous ethanol solution for 30 minutes, immersed in water for 30 minutes, and then immersed in a 20% by mass aqueous sodium hydroxide solution at 70°C for 1 hour, and further repeatedly washed with water to extract and remove fine silica, thereby obtaining a porous hollow fiber membrane. The obtained porous hollow fiber membrane is a porous membrane with the inner surface (inner diameter side surface) as the surface on the side of the filtrate. Table 1 shows the detailed composition and conditions. FIG. 5 is an electron microscope photograph of a cross section of the obtained porous hollow fiber membrane in the vicinity of the side of the liquid to be filtered.

[0109] The membrane area (converted into the inner surface) of this hollow fiber membrane is 1.9 m 2 The membrane was placed in a module case with an effective length of 20 cm so that the effective length was 20 cm, and both ends were potted with epoxy resin to prepare a hollow fiber filtration membrane mini module with open ends. As shown in Figure 7, using the prepared hollow fiber filtration membrane mini-module, CHO-K1 cells incorporating a gene expressing a monoclonal antibody were introduced into a 15 L (effective volume: 8.5 L) glass bioreactor (Repligen) using a KML-100TFF perfusion system (Repligen), and a continuous culture experiment was performed. A basal medium containing glucose and an antifoaming agent was continuously added until the cell concentration reached 75 ± 10 × 10 6 The permeation flux of the hollow fiber membrane mini module was 2.0 L / m. 2 The flow rate was set to 0.30 m / s / h, and the culture solution was allowed to permeate from the inner surface to the outer surface of the hollow fiber membrane, and the tangential flow linear velocity of the culture solution flowing through the hollow was set to 0.30 m / s. The culture was carried out for about 40 days, during which time the permeation pressure of the hollow fiber membrane was about 12 kPa, and stable operation was achieved. When the antibody transmittance in the permeate was checked, it was found to be 83% or more over a period of 40 days.

[0110] Example 2 The thermoplastic resin was vinylidene fluoride homopolymer (KF-W#1000 manufactured by Kureha Corporation), the organic liquid was a mixture of di(2-ethylhexyl) phthalate (DEHP) (manufactured by C.G. Ester Corporation) and dibutyl phthalate (DBP) (manufactured by C.G. Ester Corporation), and the inorganic fine powder was finely powdered silica (manufactured by Nippon Aerosil Co., Ltd., product name: AEROSIL-R972, primary particle diameter approximately 16 nm). A hollow fiber molding nozzle was used to perform melt extrusion of a hollow fiber membrane using an extruder. A molten mixture having a composition of vinylidene fluoride homopolymer: di(2-ethylhexyl) phthalate: dibutyl phthalate: finely divided silica = 40.0: 30.8: 6.20: 23.0 (mass ratio) was prepared. The mixture was extruded from a hollow fiber forming nozzle having an outer diameter of 2.0 mm and an inner diameter of 0.90 mm at a discharge temperature of 240°C using air as a hollow portion forming fluid. The hollow fiber-shaped molten kneaded material extruded at a discharge temperature of 240°C passed through a high-temperature container set at 240°C for 0.090 seconds, and after 0.60 seconds of air travel including the high-temperature container section, was led into a coagulation bath filled with 30°C water, taken up at a speed of 20 m / min, sandwiched between belts and stretched at a speed of 40 m / min, then contracted at a speed of 30 m / min while being exposed to hot air of 140°C, and wound into a skein. The air speed in the free running section was 0.80 m / sec. The obtained hollow fiber-like product was immersed in isopropyl alcohol to extract and remove di(2-ethylhexyl) phthalate and dibutyl phthalate, and then dried. It was then immersed in a 50% by mass aqueous solution of ethanol for 30 minutes, immersed in water for 30 minutes, and then immersed in a 20% by mass aqueous solution of sodium hydroxide at 70°C for 1 hour, and then repeatedly washed with water to extract and remove the fine silica, thereby obtaining a porous hollow fiber membrane. The air velocity in the free running section was 0.80 m / sec. The obtained hollow fiber-like material was immersed in isopropyl alcohol to extract and remove di(2-ethylhexyl) phthalate and dibutyl phthalate, and then dried. Next, it was immersed in a 50% by mass aqueous ethanol solution for 30 minutes, then in water for 30 minutes, and then in a 20% by mass aqueous sodium hydroxide solution at 70°C for 1 hour, and further repeatedly washed with water to extract and remove the fine silica, thereby obtaining a porous hollow fiber membrane. The obtained porous hollow fiber membrane is a porous membrane with the inner surface (inner diameter side surface) as the surface on the side of the filtrate. Table 1 shows the detailed composition and conditions. The membrane area (converted into the inner surface) of this hollow fiber membrane is 1.9 m 2 The membrane was placed in a module case with an effective length of 20 cm so that the effective length was 20 cm, and both ends were potted with epoxy resin to prepare a hollow fiber filtration membrane mini module with open ends.

[0111] As shown in Figure 7, using the prepared hollow fiber filtration membrane mini-module, CHO-K1 cells incorporating a gene expressing a monoclonal antibody were introduced into a 15 L (effective volume: 8.5 L) glass bioreactor (Repligen) using a KML-100TFF perfusion system (Repligen), and a continuous culture experiment was performed. A basal medium containing glucose and an antifoaming agent was continuously added until the cell concentration reached 75 ± 10 × 106 The permeation flux of the hollow fiber membrane mini module was 2.0 L / m. 2 The flow rate was set to 0.5 / h, and the culture solution was allowed to permeate from the inner surface to the outer surface of the hollow fiber membrane, and the tangential flow linear velocity of the culture solution flowing through the hollow was set to 0.25 m / s. The culture was carried out for about 40 days, during which time the permeation pressure of the hollow fiber membrane was about 11 kPa, and stable operation was achieved. When the antibody transmittance in the permeate was checked, it was found to be 85% or more over a period of 40 days.

[0112] Example 3 Polyethylene (SH800, Asahi Kasei Corporation) was used as the thermoplastic resin, and dibutyl adipate (DBA) (Tokyo Chemical Industry Co., Ltd.) was used as the organic liquid. A porous hollow fiber membrane was obtained in the same manner as in Example 1, except that the composition of the molten mixture was polyethylene:dibutyl adipate:fine silica = 18.5:54.3:27.2 (mass ratio), the discharge temperature was 230°C, the idle time was 3.0 seconds, and the winding speed was 10 m / min. Table 1 shows the detailed composition and conditions.

[0113] The membrane area (converted into the inner surface) of this hollow fiber membrane is 1.9 m 2 The membrane was placed in a module case with an effective length of 20 cm so that the effective length was 20 cm, and both ends were potted with epoxy resin to prepare a hollow fiber filtration membrane mini module with open ends. As shown in Figure 7, using the prepared hollow fiber filtration membrane mini-module, CHO-K1 cells incorporating a gene expressing a monoclonal antibody were introduced into a 15 L (effective volume: 8.5 L) glass bioreactor (Repligen) using a KML-100TFF perfusion system (Repligen), and a continuous culture experiment was performed. A basal medium containing glucose and an antifoaming agent was continuously added until the cell concentration reached 75 ± 10 × 10 6 The flow rate of the hollow fiber membrane mini module was adjusted to maintain the flow rate at 1 L / m. At this time, 3 / 4 of the effective volume was replaced in one day. 2The flow rate was set to 0.5 / h, and the culture solution was allowed to permeate from the inner surface to the outer surface of the hollow fiber membrane, and the tangential flow linear velocity of the culture solution flowing through the hollow was set to 0.25 m / s. The culture was carried out for about 40 days, during which time the permeation pressure of the hollow fiber membrane was about 13 kPa, and stable operation was achieved. When the antibody transmittance in the permeate was checked, it was found to be 84% or more over a period of 40 days.

[0114] Comparative Example 1 The composition of the melt-kneaded product was vinylidene fluoride homopolymer: di(2-ethylhexyl) phthalate: dibutyl phthalate: fine silica powder = 40.0: 45.8: 9.20: 5.0 (mass ratio). A porous hollow fiber membrane was obtained in the same manner as in Example 2, except that the stretching and shrinking steps were not performed. Table 1 shows the detailed composition and conditions. The membrane area (converted into the inner surface) of this hollow fiber membrane is 1.9 m 2 The membrane was placed in a module case with an effective length of 20 cm so that the effective length was 20 cm, and both ends were potted with epoxy resin to prepare a hollow fiber filtration membrane mini module with open ends.

[0115] As shown in Figure 7, using the prepared hollow fiber filtration membrane mini-module, CHO-K1 cells incorporating a gene expressing a monoclonal antibody were introduced into a 15 L (effective volume: 8.5 L) glass bioreactor (Repligen) using a KML-100TFF perfusion system (Repligen), and a continuous culture experiment was performed. A basal medium containing glucose and an antifoaming agent was continuously added until the cell concentration reached 75 ± 10 × 10 6 The flow rate of the hollow fiber membrane mini module was adjusted to maintain the flow rate at 1 L / m. At this time, 3 / 4 of the effective volume was replaced in one day. 2 The flow rate was set to 0.30 m / s / h, and the culture solution was allowed to permeate from the inner surface to the outer surface of the hollow fiber membrane, and the tangential flow linear velocity of the culture solution flowing through the hollow was set to 0.30 m / s. After about 20 days of culture, the permeation pressure of the hollow fiber membrane rose to about 100 kPa, and stable operation was not possible. When the antibody transmittance in the permeate was checked, it was found to have decreased to 50% on the 20th day.

[0116] [Table 1]

[0117] [Table 2] [Explanation of symbols]

[0118] a. Thermoplastic resin b Cavity 10 Extruder 20 Hollow fiber molding nozzle 30 Coagulation bath 40 Porous hollow fiber membrane 50 Lola 60 Suction machine 70 High Temperature Container S empty running club space FA Surface L Line of equal distance from FA h Hole Lh The length L crosses the part of the image that corresponds to h

Claims

1. A filtration method comprising a filtration step of passing a culture solution containing cells, a medium, a product, and an antifoaming agent through a porous membrane to separate a filtrate containing the product from the culture solution, wherein the filtration method is characterized in that the product of the porosity of the porous membrane at a thickness of 0.12% of the membrane thickness from the outermost surface of the surface of the porous membrane facing the liquid to be filtered and the porosity of the surface of the porous membrane facing the liquid to be filtered is 1800%.% or more.

2. 2. The filtration method according to claim 1, wherein the porosity of the porous membrane is 50% or more in a thickness range from the outermost surface of the surface facing the liquid to be filtered to a thickness of 0.12% of the membrane thickness.

3. 3. The filtration method according to claim 1, wherein the porosity of the surface of the porous membrane facing the liquid to be filtered is 30% or more.

4. 3. The filtration method according to claim 1, wherein the ratio of the porosity in a thickness from the outermost surface of the surface of the porous membrane facing the liquid to 0.12% of the membrane thickness to the porosity of the surface facing the liquid to be filtrated is 1.70 or more.

5. 3. The filtration method according to claim 1, wherein the product of the porosity in a thickness range from the outermost surface of the surface of the porous membrane facing the liquid to be filtered to 0.12% of the film thickness and the porosity of the surface facing the liquid to be filtered is 2800%.% or more.

6. 3. The filtration method according to claim 1, wherein the product of the porosity at a thickness of 300 nm from the outermost surface of the filtrate side surface of the porous membrane in the film thickness direction and the porosity of the filtrate side surface is 860%.% or more.

7. 7. The filtration method according to claim 6, wherein the porosity at a thickness of 300 nm from the outermost surface of the filtrate side surface of the porous membrane in the film thickness direction is 35% or more.

8. 8. The filtration method according to claim 7, wherein the porosity of the filtrate-side surface of the porous membrane is 30% or more.

9. 3. The filtration method according to claim 1, wherein the cross-sectional pore size of the porous membrane at a thickness of 0.12% of the membrane thickness from the outermost surface of the surface facing the liquid to be filtered is 1000 nm or less.

10. 3. The filtration method according to claim 1, wherein the cross-sectional pore size at a thickness of 300 nm from the outermost surface of the surface of the porous membrane facing the liquid to be filtered in the film thickness direction is 300 nm or less.

11. The filtration method according to claim 1 or 2, wherein the porous membrane has a thickness of 100 μm or more and 800 μm or less.

12. The filtration method according to claim 1 or 2, wherein the flux of the porous membrane in the filtration step is 1 LMH or more.

13. The filtration method according to claim 1 or 2, wherein the porous membrane is a hollow fiber membrane, and the filtration step is perfusion culture filtration.

14. 3. The filtration method according to claim 1, wherein the hollow fiber membrane has an intramembrane linear velocity of tangential flow during the perfusion culture of 0.3 m / s or less.

15. The filtration method according to claim 1 or 2, wherein the resin constituting the porous membrane is a thermoplastic resin.

16. The filtration method according to claim 15 , wherein the thermoplastic resin contains a fluororesin as a main component.

17. The fluororesin is at least one selected from the group consisting of polyvinylidene fluoride resin (PVDF), chlorotrifluoroethylene resin, tetrafluoroethylene resin, ethylene-tetrafluoroethylene copolymer (ETFE), ethylene-monochlorotrifluoroethylene copolymer (ECTFE), hexafluoropropylene resin, and mixtures of these resins. The filtration method according to claim 16.

18. The filtration method according to claim 15 , wherein the thermoplastic resin contains a polyolefin resin as a main component.

19. The filtration method according to claim 15, wherein the thermoplastic resin is polyethylene (PE) or polypropylene (PP).

20. 3. The method of claim 1, wherein the product is selected from the group consisting of proteins including enzymes and antibodies, amino acids, nucleic acids, and organic substances.

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