Membranes formed from anionic monomers suitable for detection, filtration and/or purification of biomolecules and metal ions
阅读说明：本技术 适合检测、过滤和/或纯化生物分子和金属离子的由阴离子单体形成的膜 (Membranes formed from anionic monomers suitable for detection, filtration and/or purification of biomolecules and metal ions ) 是由 B·博格斯 伊丹雄二郎 于 2019-09-04 设计创作，主要内容包括：平均孔径为5nm至5,000nm并且孔隙率为10％以上的膜,所述膜可通过包括将组合物固化的方法获得,所述组合物包含：(i)包含至少一个阴离子基团的交联剂；和(ii)惰性溶剂。所述膜用于检测金属离子以及过滤和/或纯化生物分子和包含金属离子的组合物。(A membrane having an average pore diameter of 5nm to 5,000nm and a porosity of 10% or more, the membrane being obtainable by a process comprising curing a composition comprising: (i) a crosslinker comprising at least one anionic group; and (ii) an inert solvent. The membranes are useful for detecting metal ions and for filtering and/or purifying biomolecules and compositions comprising metal ions.)
1. A membrane having an average pore diameter of 5nm to 5,000nm and a porosity of 10% or more, the membrane being obtainable by a process comprising curing a composition comprising:
(i) a crosslinker comprising at least 1 anionic group; and
(ii) an inert solvent.
2. The membrane of claim 1, wherein component (i) comprises at least 2 anionic groups.
3. The membrane of any one of the preceding claims, wherein component (i) comprises a backbone, at least 2 polymerizable groups, and at least 1 anionic group, wherein the at least 1 anionic group is a pendant group attached to the backbone by a single covalent bond or by a spacer group.
4. A film according to any preceding claim wherein the composition comprises from 10 to 50% by weight of component (i) and from 50 to 90% by weight of component (ii).
5. The film of any of the preceding claims, wherein the composition comprises an organic amine.
6. A membrane according to any preceding claim, wherein component (ii) comprises water and an inert water-miscible organic solvent, wherein the wt% of water is less than the wt% of the inert water-miscible organic solvent.
7. A film according to any preceding claim wherein component (ii) comprises water and a water-miscible organic solvent and component (i) is fully dissolved in the composition.
8. The film of any one of the preceding claims, wherein:
component (ii) comprises a composition comprising one or more inert solvents selected from list (iia) and one or more inert solvents selected from list (iib):
list (iia): isopropanol, methanol, ethanol, acetone, tetramethylurea, hexamethylphosphoramide, hexamethylphosphoric triamide, butanone, cyclohexanone, methyl ethyl ketone, tetrahydrofuran, tetrahydropyran, 2-methyltetrahydrofuran, cyclopentyl methyl ether, propionitrile, acetonitrile, 1, 4-dioxane, 1, 3-dioxolane, ethyl acetate, γ -butyrolactone, ethanolamine, or a mixture comprising two or more thereof; and
list (iib): water, glycerol, ethylene glycol, dimethyl sulfoxide, sulfolane, dimethylimidazolidinone, sulfolane, N-methylpyrrolidone, dimethylformamide, acetonitrile, acetone, 1, 4-dioxane, 1, 3-dioxolane, tetramethylurea, hexamethylphosphoramide, hexamethylphosphoric triamide, pyridine, propionitrile, butanone, cyclohexanone, tetrahydrofuran, tetrahydropyran, 2-methyltetrahydrofuran, ethylene glycol diacetate, cyclopentyl methyl ether, methyl ethyl ketone, ethyl acetate, and γ -butyrolactone, and wherein dimethyl sulfoxide, N-methylpyrrolidone, N-dimethylformamide, dimethylimidazolidinone, N-methylmorpholine, acetone, cyclopentyl methyl ether, methyl ethyl ketone, acetonitrile, tetrahydrofuran, 2-methyltetrahydrofuran, and mixtures comprising two or more thereof.
9. A film according to any preceding claim wherein component (ii) comprises isopropanol and water.
10. The film according to claim 8, wherein component (ii) comprises from 40 to 70% by weight of an inert solvent selected from the list (iia) and from 10 to 40% by weight of an inert solvent selected from the list (iib).
11. The film of any preceding claim, wherein component (ii) comprises 40 to 70 wt% isopropanol, 10 to 40 wt% water and optionally 0.01 to 10 wt% organic amine.
12. A film according to any preceding claim wherein the composition further comprises from 0 to 20% by weight of (iii) a monomer reactive with component (i).
13. A film as claimed in any preceding claim wherein the curing comprises photo-curing.
14. A film according to any preceding claim wherein the composition further comprises a photoinitiator.
15. The film of claim 13 or 14, wherein the photocuring comprises a polymerization-initiated phase separation between the film and the composition.
16. The membrane of any one of the preceding claims, further comprising a porous support.
17. The membrane of any one of the preceding claims, which has good tolerance to pH at pH 1 to pH 10.
18. A method of making the film of any one of claims 1 to 17, the method comprising curing a composition comprising:
(i) a crosslinker comprising at least one anionic group; and
(ii) an inert solvent.
19. A method according to claim 18, wherein the composition is as defined in any one of claims 2 to 12.
20. The method of claim 18 or 19, wherein the curing comprises photo-curing.
21. The method of any one of claims 18 to 20, wherein the curing comprises photo-polymerization induced phase separation between the film and the composition.
22. Use of a membrane according to any one of claims 1 to 17 for the detection, filtration and/or purification of biomolecules.
23. Use of a membrane according to any one of claims 1 to 17 for detecting metal ions or filtering and/or purifying a composition comprising metal ions.
24. A method of purifying and/or separating biomolecules from other biomolecules, the method comprising contacting a biomolecule with the membrane of any one of claims 1 to 17.
25. A method of purifying metal ions and/or separating metal ions from other particles or ionic species, the method comprising contacting metal ions with the membrane of any one of claims 1 to 17.
26. The method of claim 24 or 25, wherein the method comprises membrane size exclusion chromatography or ion exchange chromatography.
Drawings
Fig. 1a and 1b are scanning electron microscope ("SEM") photographs of the top surface and cross-section of the inventive membrane at magnifications of 10,000 and 20,000, respectively. The membrane in fig. 1a and 1b comprises a porous support.
Fig. 2a and 2b are SEM photographs of the top surface and cross section of the membrane of the present invention at magnifications of 2,000 times and 10,000 times, respectively. The membrane in fig. 2a and 2b does not comprise a porous support.
In this specification (including its claims), the verb "to comprise" and its conjugations is used in its non-limiting sense to mean that items following the word are included, but items not specifically mentioned are not excluded. In addition, the indefinite article "a" or "an" does not exclude the possibility that more than one of the element is present, unless the context clearly requires that only one of the elements is present. Thus, the indefinite article "a" or "an" usually means "at least one".
The crosslinking agent preferably comprises at least 2 polymerizable groups, for example at least 2 groups selected from epoxy groups, thiol (-SH), oxetane rings, and in particular ethylenically unsaturated groups. The polymerizable groups in component (i) are typically selected such that they are reactive with each other or with at least one polymerizable group present in another chemically different component (i).
Curing causes the crosslinking agent to crosslink, e.g., form a film, as a crosslinked three-dimensional polymer matrix.
The at least 2 polymerizable groups present in component (i) may be chemically identical or they may also be different.
Preferred ethylenically unsaturated groups are selected from the group consisting of (meth) acrylic groups and vinyl groups (e.g., vinyl ether groups, aromatic vinyl compounds, N-vinyl compounds, and allyl groups).
Examples of suitable (meth) acrylic groups include acrylates (H)2A C ═ CHCO-) group, acrylamide (H)2A C ═ CHCONH-) group, methacrylate (H)2C＝C(CH3) CO-) radical and methacrylamide (H)2C＝C(CH3) CONH-) groups. Acrylic groups are preferred over methacrylic groups because acrylic groups are more reactive.
Preferred ethylenically unsaturated groups are free of ester groups as this may improve the stability and pH tolerance of the resulting film. The ethylenically unsaturated group containing no ester group includes a (meth) acrylamide group and a vinyl ether group (a (meth) acrylamide group is particularly preferable).
As preferred examples of the polymerizable group, there may be mentioned groups of the formula:
the anionic groups in component (i) may help the resulting membrane to distinguish between molecular species (e.g. ionically charged biomolecules) as well as between different metal ions.
Preferred anionic groups include sulfate, phosphate, carboxylate, benzoate, phenolate and urate, especially sulfate.
In a preferred embodiment, component (i) comprises a backbone, at least 2 polymerizable groups and at least 1 anionic group, preferably a pendant anionic group (e.g., an anionic group is attached to the backbone by a single covalent bond or by a spacer group).
In an even more preferred embodiment, component (i) comprises at least 2 anionic groups, for example, 2, 3 or 4 anionic groups. The anionic groups are preferably attached to the remainder of component (i) by a single covalent bond or by a spacer group.
Preferred examples of the component (i) include the following compounds (M1) to (M10):
the amount of component (i) present in the composition is preferably from 1 to 80 wt%, more preferably from 10 to 50 wt%, and most preferably from 15 to 40 wt%, relative to the total weight of the composition.
Preferably, component (i) is completely dissolved in the composition.
In this specification, "inert" means non-polymerizable. Therefore, component (ii) cannot be polymerized with component (i).
Component (ii) preferably consists of one inert solvent or comprises more than one inert solvent, in particular a mixture comprising two or more miscible inert solvents. The inert character of component (ii) helps to form pores in the membrane.
Preferably, component (ii) is non-solvent to the membrane (the membrane is preferably insoluble in component (ii)). Component (ii) has the function of dissolving component (i), component (iv) and optionally component (iii), if present. Component (ii) may also help ensure that the membrane precipitates out of the composition as it is formed, for example by a phase separation process.
The amount of component (ii) present in the composition is preferably from 0 to 99% by weight, more preferably from 50 to 90% by weight and in particular from 60 to 85% by weight, relative to the total weight of the composition.
Curing causes the crosslinking agent to crosslink, e.g., form a film, as a crosslinked three-dimensional polymer matrix.
Preferably, component (ii) comprises water, or a mixture of water and an inert organic solvent (preferably, a water-miscible inert organic solvent) having a water solubility of at least 5 wt.%.
Examples of the inert solvent which can be used as component (ii) or used in component (ii) include alcohol solvents, ether solvents, amide solvents, ketone solvents, sulfoxide solvents, sulfone solvents, nitrile solvents and organophosphorus solvents, with inert aprotic polar solvents being preferred.
Examples of alcoholic solvents (particularly in combination with water) that may be used as or in component (ii) include methanol, ethanol, isopropanol, n-butanol, ethylene glycol, propylene glycol, diethylene glycol, dipropylene glycol and mixtures comprising two or more thereof. Particular preference is given to isopropanol.
In addition, preferred inert organic solvents that can be used as or in component (ii) include dimethyl sulfoxide, dimethyl imidazolidinone, sulfolane, N-methylpyrrolidone, dimethylformamide, acetonitrile, acetone, 1, 4-dioxane, 1, 3-dioxolane, tetramethylurea, hexamethylphosphoramide, hexamethylphosphoric triamide, pyridine, propionitrile, butanone, cyclohexanone, tetrahydrofuran, tetrahydropyran, 2-methyltetrahydrofuran, ethylene glycol diacetate, cyclopentyl methyl ether, methyl ethyl ketone, ethyl acetate, γ -butyrolactone, and mixtures comprising two or more thereof. Preferred are dimethyl sulfoxide, N-methylpyrrolidone, dimethylformamide, dimethylimidazolidinone, sulfolane, acetone, cyclopentyl methyl ether, methyl ethyl ketone, acetonitrile, tetrahydrofuran, 2-methyltetrahydrofuran, and mixtures comprising two or more thereof.
In a preferred embodiment, component (ii) comprises a composition comprising one inert solvent selected from list (iia) and one or more inert solvents selected from list (iib):
list (iia): isopropanol, methanol, ethanol, acetone, tetramethylurea, hexamethylphosphoramide, hexamethylphosphoric triamide, butanone, cyclohexanone, methyl ethyl ketone, tetrahydrofuran, tetrahydropyran, 2-methyltetrahydrofuran, cyclopentyl methyl ether, propionitrile, acetonitrile, 1, 4-dioxane, 1, 3-dioxolane, ethyl acetate, γ -butyrolactone, ethanolamine, or a mixture comprising two or more thereof; and
list (iib): water, glycerol, ethylene glycol, dimethyl sulfoxide, sulfolane, dimethylimidazolidinone, sulfolane, N-methylpyrrolidone, dimethylformamide, acetonitrile, acetone, 1, 4-dioxane, 1, 3-dioxolane, tetramethylurea, hexamethylphosphoramide, hexamethylphosphoric triamide, pyridine, propionitrile, butanone, cyclohexanone, tetrahydrofuran, tetrahydropyran, 2-methyltetrahydrofuran, ethylene glycol diacetate, cyclopentyl methyl ether, methyl ethyl ketone, ethyl acetate, and γ -butyrolactone, and wherein dimethyl sulfoxide, N-methylpyrrolidone, N-dimethylformamide, dimethylimidazolidinone, N-methylmorpholine, acetone, cyclopentyl methyl ether, methyl ethyl ketone, acetonitrile, tetrahydrofuran, 2-methyltetrahydrofuran, and mixtures comprising two or more thereof.
In one embodiment, the composition comprises water and one or more other solvents selected from the list (iia) and/or the list (iib).
Thus, preferred compositions comprise from 1 to 80% by weight of component (i) and from 20 to 99% by weight of component (ii), more preferably from 10 to 50% by weight of component (i) and from 50 to 90% by weight of component (ii), in particular from 15 to 40% by weight of component (i) and from 60 to 85% by weight of component (ii).
Preferably, component (ii) comprises an organic amine (especially C)2-6-alcohol amines), such as methylamine, ethylamine, diethylamine, triethylamine and in particular ethanolamine. The amount of organic amine present in the composition is preferably sufficient to neutralise at least 50%, more preferably at least 75%, and especially all anionic groups present in components (i) and (iii), if present.
Preferably, component (ii) comprises 40 to 70 wt% isopropanol, 10 to 40 wt% water and optionally 0.01 to 10 wt% organic amine.
Optionally, the composition further comprises (iii) a monomer reactive with component (i), for example a monomer comprising one polymerizable group (e.g. an ethylenically unsaturated group) and optionally one or more anionic groups. As described above for component (i), preferred polymerizable groups are ethylenically unsaturated groups, especially (meth) acrylic groups.
Preferably, the number of moles of component (i) exceeds the number of moles of component (iii), if present.
The composition preferably comprises from 0 to 20% by weight of component (iii), based on the total weight of the composition.
In a preferred embodiment, component (i) is present in the composition in an amount of at least 80 wt.%, more preferably at least 90 wt.%, in particular at least 95 wt.%, relative to the total amount of components (i) and (iii). In a most preferred embodiment, the composition comprises 0 wt% of component (iii) (monomer reactive with component (i)).
Preferably, component (iii), if present, is soluble in component (ii).
The composition may be cured by any suitable method, including thermal curing, photo-curing, and combinations thereof. However, it is preferred to cure the composition by photocuring, for example irradiating the composition, thereby polymerising component (i) and any other polymerisable components present in the composition. Typically, component (ii) is inert and does not polymerize, but leaves pores in the resulting film.
Preferably, the composition further comprises (iv) a polymerization initiator, such as a thermal initiator and/or a photoinitiator.
Examples of suitable thermal initiators that may be included in the composition include: 2,2 ' -azobis (2-methylpropanenitrile) (AIBN), 4 ' -azobis (4-cyanopentanoic acid), 2 ' -azobis (2, 4-dimethylpentanenitrile), 2 ' -azobis (2-methylbutyronitrile), 1 ' -azobis (cyclohexane-1-carbonitrile), 2 ' -azobis (4-methoxy-2, 4-dimethylpentanenitrile), dimethyl 2,2 ' -azobis (2-methylpropionate), 2 ' -azobis [ N- (2-propenyl) -2-methylpropionamide, 1- [ (1-cyano-1-methylethyl) azo ] formamide, 2 ' -azobis (N-butyl-2-methylpropionamide), 2,2 '-azobis (N-cyclohexyl-2-methylpropionamide), 2' -azobis (2-methylpropionamidine) dihydrochloride, 2 '-azobis [2- (2-imidazolin-2-yl) propane ] disulfate dihydrate, 2' -azobis [ N- (2-carboxyethyl) -2-methylpropionamidine ] hydrate, 2 '-azobis {2- [1- (2-hydroxyethyl) -2-imidazolin-2-yl ] propane } dihydrochloride, 2' -azobis [2- (2-imidazolin-2-yl) propane ], (N-methyl-2-ethyl-2-methyl-2-propyl-carbamate) hydrochloride, 2,2 ' -azobis (1-imino-1-pyrrolidinyl-2-ethylpropane) dihydrochloride, 2 ' -azobis { 2-methyl-N- [1, 1-bis (hydroxymethyl) -2-hydroxyethyl ] propionamide } and 2,2 ' -azobis [ 2-methyl-N- (2-hydroxyethyl) propionamide ].
Examples of suitable photoinitiators that may be included in the composition include aromatic ketones, acylphosphine compounds, aromatic onium salt compounds, organic peroxides, thio compounds, hexaarylbiimidazole compounds, ketoxime ester compounds, borate compounds, azine (azinium) compounds, metallocene compounds, active ester compounds, compounds having a carbon halogen bond, and alkylamine compounds. Preferred examples of the aromatic ketone, acylphosphine oxide compound and thio compound include compounds having a benzophenone backbone or a thioxanthone backbone, which are described IN "RADIATION CURING IN POLYMER SCIENCE AND TECHNOLOGY", pp.77-117 (1993). More preferable examples thereof include α -thiobenzophenone compounds described in JP1972-6416B (JP-S47-6416B), benzoin ether compounds described in JP1972-3981B (JP-S47-3981B), α -substituted benzoin compounds described in JP1972-22326B (JP-S47-22326B), benzoin derivatives described in JP1972-23664B (JP-S47-23664B), aroylphosphonates described in JP1982-30704A (JP-S57-30704A), dialkoxybenzophenones described in JP1985-26483B (JP-S60-26483B), JP1985-26403B (JP-S60-26403B) and JP1987-81345A (JP-S62-81345A), benzoin ethers described in JP1989-34242B (JP-S34242H 242-34242B), and JP 1982-30242B (JP-S01-34242A), Alpha-aminobenzophenones described in U.S. Pat. No. 4,318,791A and EP0284561A1, p-bis (dimethylaminobenzoyl) benzenes described in JP1990-211452A (JP-H02-211452A), thio-substituted aromatic ketones described in JP1986-194062A (JP-S61-194062A), acylphosphine sulfides described in JP1990-9597B (JP-H02-9597B), acylphosphines described in JP1990-9596B (JP-H02-9596B), thioxanthones described in JP1988-61950B (JP-S63-61950B), coumarins described in JP1984-42864B (JP-S59-42864B). In addition, the photoinitiators described in JP2008-105379A and JP2009-114290A are also preferred. In addition, photoinitiators described in "Ultraviolet Curing System" of Kato Kiyomi (Research Center co., ltd., 1989), pages 65 to 148, may be used.
The polymerization initiator is preferably water-soluble.
The composition preferably comprises from 0.1 to 5 wt%, more preferably from 0.3 to 2 wt%, of the polymerization initiator (iv), based on the total weight of the composition. The polymerization initiator (iv) preferably has a water solubility of at least 1 wt%, more preferably at least 3 wt%, when measured at 25 ℃.
Optionally, the composition may contain one or more additional components in addition to the above components, such as surfactants, polymeric dispersants, polymerization control agents, thickeners or anti-cratering agents, and the like.
Optionally, the membrane of the invention further comprises a support, in particular a porous support. The inclusion of a support may provide greater mechanical strength to the membrane. If desired, the composition may be applied to the support between steps (a) and (b) of the method of producing a film of the second aspect of the present invention described below. In this manner, the porous support may be impregnated with the composition, and the composition may then be polymerized on and/or in the support.
Examples of suitable supports include synthetic textiles and synthetic nonwovens, sponge-like films and films with fine through-holes. The material used to form the optional porous support may be a porous membrane based on: such as polyolefins (polyethylene or polypropylene, etc.), polyacrylonitrile, polyvinyl chloride, polyesters, polyamides or copolymers thereof, or such as polysulfone, polyethersulfone, polyphenylsulfone, polyphenylene sulfide, polyimide, polyetherimide, polyamide, polyamideimide, polyacrylonitrile, polycarbonate, polyacrylate, cellulose acetate, cellulose, polypropylene, poly (4-methyl-1-pentene), polyvinylidene fluoride, polytetrafluoroethylene, polyhexafluoropropylene, polychlorotrifluoroethylene or copolymers thereof. Among them, in the present invention, polyolefin and cellulose are preferable.
As commercially available porous supports, products from Japan Vilene Company, Ltd., Freudenberg Filtration Technologies, Sefar AG or Asahi-Kasei can be used.
When the film comprises a support and curing comprises photocuring, then it is preferred that the support does not shield the wavelength of light used to cure the composition.
The support is preferably a hydrophilic support, for example, a support which has been subjected to corona treatment, ozone treatment, sulfuric acid treatment, silane coupling agent treatment, or two or more of the foregoing treatments.
The membrane of the present invention may optionally comprise more than one support, and the more than one supports may be the same or different from each other.
The average pore diameter of the membrane is preferably 10 to 5,000nm, more preferably 100 to 2,000 nm. Preferably, the pores are deeper than their average diameter.
The average pore size of the membranes of the invention can be measured using a porosimeter, such as PoroluxTMThe porosimeter. For example, a wetting fluid (e.g., Porefil) may be usedTMA wetting liquid, an inert non-toxic fluorocarbon wetting liquid with a contact angle of zero) completely wets the membrane to be tested, and then the wetted membrane is placed in the sample holder of the porosimeter and a pressure of maximum 35mbar is applied. The porosimeter may then provide the bubble point, maximum pore size, mean flow pore size, minimum pore size of the membrane being testedAverage pore size distribution (of uniform material) and gas permeability.
When the membrane does not comprise a support, the porosity of the membrane is preferably from 15 to 99%, preferably from 20 to 99%, in particular from 20 to 85%.
When the membrane does not comprise a support, the porosity of the membrane is preferably 21 to 70%.
The porosity of the membrane can be determined by gas displacement densitometry, for example using a densitometer (particularly AccuPyc available from Micromeritics Instrument Corporation)TMII 1340 gas replacement ratio calculation system).
The porosity of a membrane is the amount of volume accessible to external fluids or gases. Which can be determined as described below. Preferably, the porosity of the membrane of the invention is greater than 20%.
When the film of the present invention includes a support, the thickness of the film including the support in a dry state is preferably 20 μm to 2,000 μm, more preferably 40 μm to 1,000 μm, and particularly preferably 70 μm to 800 μm.
When the film of the present invention does not contain a support, the thickness of the film in a dry state is preferably 20 μm to 2,000 μm, more preferably 100 μm to 2,000 μm, and particularly preferably 150 μm to 2,000 μm.
When the membrane of the present invention includes a support, the thickness of the membrane including the support is preferably 10 μ M to 4,000 μ M, more preferably 20 μ M to 2,000 μ M, and particularly preferably 20 μ M to 1,500 μ M, when measured after being stored in a 0.1M NaCl solution for 12 hours.
When the membrane of the present invention does not contain a support, the thickness of the membrane is preferably 10 μ M to 4,000 μ M, more preferably 50 μ M to 4,000 μ M, particularly 70 μ M to 4,000 μ M, when measured after storage in a 0.1M NaCl solution for 12 hours.
According to a second aspect of the present invention there is provided a method of making a film of the first aspect of the present invention, the method comprising curing a composition as defined in the first aspect of the present invention.
Preferably, the membrane of the first aspect of the invention is obtained by a process comprising the steps of:
(a) mixing components (i), (ii) and preferably (iii) and/or (iv) to form a composition comprising components (i), (ii) and optionally (iii);
(b) curing (e.g. irradiating) the composition resulting from step (a), thereby polymerising component (i) (and component (iii), if present) to form a film; and
(c) optionally washing the membrane resulting from step (b).
Preferably, the process for preparing the film of the invention comprises polymerisation initiated phase separation, more preferably photo polymerisation initiated phase separation, e.g. separation of the film from the composition. In this method, preferably, the polymer is formed as a result of a photopolymerization reaction.
Optionally, step (b) may be performed by one or more additional irradiation and/or heating steps to fully cure the film.
The inclusion of component (ii) in the composition has the advantage of helping the polymerisation in step (b) to proceed uniformly and smoothly.
In a preferred embodiment, component (ii) acts as a solvent for component (i) and helps to form pores in the resulting film.
The method of the second aspect of the invention provides a substantially homogeneous film, which typically has a substantially homogeneous bicontinuous structure. In some embodiments, curing results in component (i) (and component (iii), when present) forming substantially uniform polymer particles that are then fused to form the membranes of the present invention having an average pore size of 5nm to 5,000nm and a porosity of 10% or greater. The interstices between the polymer particles provide a membrane with pores of a desired average size and a desired porosity.
The polymer particles or agglomerates thereof typically have an average diameter in the range of from 0.1nm to 5,000 nm. Preferably, the average particle or agglomerate size of the polymer particles is from 1nm to 2,000nm, most preferably from 10nm to 1,000 nm. The size of the average particles or agglomerates can be determined by cross-sectional analysis using a Scanning Electron Microscope (SEM).
If desired, the composition may be applied to a support, in particular a porous support, between steps (a) and (b) of the method of the second aspect of the invention. Step (b) may be carried out on the composition present on and/or in the support. When it is not necessary for the film to include a support, the film may be peeled off the support. Alternatively, if it is desired that the membrane include a support, the membrane may be left on and/or in the support.
The composition can be applied to or impregnated into the support by various methods, such as curtain coating, extrusion coating, air knife coating, slide coating, roll coating, forward roll coating, reverse roll coating, dip coating, kiss coating, rod coating, and spray coating. The coating of the multiple layers may be performed simultaneously or sequentially. Among the simultaneous multilayer coating, curtain coating, slide coating, slot die coating or extrusion coating is preferable.
The composition may be applied to the support at a temperature that facilitates the desired phase separation of the membrane from the composition. The temperature at which the composition is applied to the support (if present) is preferably less than 80 ℃, more preferably between 10 and 60 ℃, in particular between 15 and 50 ℃.
When the film comprises a support, the surface of the support may be treated, for example, using corona discharge treatment, glow discharge treatment, flame treatment or ultraviolet irradiation treatment, before applying the composition to the surface of the support. In this way, the wettability and adhesion of the support can be improved.
Step (b) optionally further comprises heating the composition.
Thus, in a preferred process, the composition is applied continuously to a moving support, more preferably by a manufacturing unit comprising one or more composition application stations, one or more irradiation sources for curing the composition, a membrane collection station, and means for moving the support from the composition application station to the irradiation sources and the membrane collection station.
The composition application station may be positioned at an upstream location relative to the illumination source, and the illumination source may be positioned at an upstream location relative to the composite film collection station.
Preferably, the curing of the composition of the invention starts within 60 seconds, more preferably within 15 seconds, particularly preferably within 5 seconds, most preferably within 3 seconds from the application of the composition to the support or from the immersion of the support by the composition (when the support is used).
The light irradiation for photocuring is preferably carried out for less than 10 seconds, more preferably for less than 5 seconds, particularly preferably for less than 3 seconds, most preferably for less than 2 seconds. In a continuous process for preparing the film, the film may be continuously irradiated. The speed at which the composition moves through the radiation beam generated by the radiation source determines the curing time and radiation dose.
Preferably, the composition is cured by a process comprising irradiating the composition with Ultraviolet (UV) light. The wavelength of the UV light used depends on the photoinitiator present in the composition, for example, the UV light is UV-A (400nm to 320nm), UV-B (320nm to 280nm) and/or UV-C (280nm to 200 nm).
When the composition is cured using high intensity UV light, a large amount of heat is generated. To prevent overheating, the lamp of the light source and/or the support/membrane are preferably cooled with cooling air. When the composition is irradiated with a high dose of infrared light (IR light) together with UV light, the irradiation of UV light is preferably performed by using an IR reflective quartz plate as a filter.
Examples of the UV light source include mercury arc lamps, carbon arc lamps, low-pressure mercury lamps, medium-pressure mercury lamps, high-pressure mercury lamps, swirl plasma arc lamps, metal halide lamps, xenon lamps, tungsten lamps, halogen lamps, lasers, and ultraviolet-emitting diodes. Medium or high pressure mercury vapor type ultraviolet emitting lamps are particularly preferred. In addition, additives such as metal halides may be present in order to modify the emission spectrum of the lamp. Lamps having a maximum emission at a wavelength of 200nm to 450nm are particularly suitable.
The energy output of the illumination source is preferably from 20W/cm to 1000W/cm, and more preferably from 40W/cm to 500W/cm, but may be higher or lower than the above exposure dose if the desired exposure dose can be achieved. The curing of the film is adjusted by the exposure intensity. The exposure dose is measured in the wavelength range of UV-A by using a high-energy UV irradiator (UV Power Puck (registered trademark) manufactured by EIT-Instrument Markets), and is preferably 40mJ/cm2Above, more preferably 100mJ/cm2To 3,000mJ/cm2And most preferably 150mJ/cm2To 1,500mJ/cm2. The exposure time can be selected fromBy choice, short times are preferred, most preferably less than 2 seconds.
The membranes of the invention are particularly useful for the isolation and purification of biomolecules, such as proteins, peptides, amino acids, antibodies and nucleic acids, in biomedical applications.
The ion exchange capacity of the membrane of the invention is preferably from 0.50meq/g to 8.00meq/g, more preferably from 0.5meq/g to 6.00meq/g, especially from 0.70meq/g to 4.00 meq/g.
The Ion Exchange Capacity (IEC) of the membranes of the present invention can be determined as follows.
The water flux of the membrane of the invention is preferably greater than 100l/m2A/bar/hr, more preferably more than 150l/m2A/bar/hr, in particular more than 500l/m2A/bar/hr and more particularly more than 1000l/m2/bar/hr。
The water flux of the membrane according to the invention can be determined as follows.
The swelling ratio of the membrane of the invention can be determined as follows: the volume of the membrane when dry and wet (wetted with water) was measured and the following calculation was performed:
the swelling ratio of the membrane in water is preferably less than 20%, more preferably less than 10%, in particular less than 5%.
According to a third aspect of the present invention there is provided the use of a membrane of the first aspect of the present invention in the detection, filtration and/or purification of biomolecules.
The membrane of the first aspect of the invention may be used for filtration and/or purification of biomolecules by eluting a solution containing biomolecules, in particular positively charged biomolecules. The positive charge on such biomolecules is attracted by the negative charge on the membrane derived from component (i). Membranes can be used to separate biomolecules by a variety of methods, including the use of membranes for size exclusion chromatography (e.g., when the pores of the membrane are used to separate or purify biomolecules according to their size (i.e., physical exclusion)) and ion exchange chromatography (e.g., when biomolecules are purified or separated according to the strength of their overall ionic interaction with anionic groups in the membrane (i.e., electronic interaction)).
The membrane of the first aspect of the invention may be used for the detection of biomolecules by techniques involving colour detection, particularly when the biomolecules comprise fluorescent or coloured labels.
Thus, another aspect of the invention includes a method of purifying and/or separating a biomolecule from other biomolecules, the method comprising contacting the biomolecule with a membrane of the invention. Preferably, the method for purifying the biomolecule and/or separating the biomolecule from other biomolecules comprises membrane size exclusion chromatography or ion exchange chromatography.
According to a fourth aspect of the present invention there is provided the use of a membrane of the first aspect of the present invention in the detection of metal ions or in the filtration and/or purification of a composition comprising metal ions.
The membrane of the first aspect of the invention may be used to detect metal ions by techniques involving colour detection.
The membrane of the first aspect of the invention may be used for filtration and/or purification of biomolecules by techniques similar to those described above for biomolecules, in particular membrane size exclusion chromatography or ion exchange chromatography. Compositions comprising metal ions (e.g., two or more metal ions and optional contaminants), metal-containing colloids, or other agglomerates of metal ions can be separated and/or purified based on their size (i.e., physical exclusion) or by ion exchange chromatography in which the composition comprising metal ions is purified or different metal ions are separated from each other based on the strength of their overall ionic interaction with the anionic groups in the membranes of the invention.
The membrane can obviously also be used for other purposes.
Preferably, the membranes of the invention are stable for at least 12 hours, more preferably at least 16 hours at pH 1.0 to pH 10.0.
The invention will now be illustrated by the following non-limiting examples. The following abbreviations are used in the examples:
FO-2223-10, which is a polypropylene-based nonwoven porous fabric with a thickness of 100 μm available from the Freudenberg Group. Which acts as a support.
IPA is isopropyl alcohol.
AMPS-Na, which is the sodium salt of 2-acrylamido-2-methylpropanesulfonic acid (from Sigma Aldrich) having the structure shown below.
CN132, which is a cross-linking agent (from Sartomer) having no anionic groups and having the structure shown below.
MBA is a cross-linker (from Sigma-Aldrich) having no anionic groups and having the structure shown below.
M282 is tetraethylene glycol diacrylate from Sigma-Aldrich.
M2 which is a monomer (i) comprising two anionic groups and two bisacrylamido groups as reactive groups. This monomer is obtained from FUJIFILM and has the structure shown in the above description. M2 can be prepared by the method described in paragraphs [0138] - [0148] of WO16024454A1 (compound MB-3).
M6 which is a monomer (i) comprising two anionic groups and two bisacrylamido groups as reactive groups. This monomer is obtained from FUJIFILM and has the structure shown in the above description. M6 can be prepared by the process described in EP2965803 in paragraph [211 ].
The water flux, ion exchange capacity, porosity and thickness of the membranes described in the examples and comparative examples were measured as follows:
2I) water flux of Membrane (L/m/bar/Hr)
The water flux of the membrane was measured using a device in which the weight of water passing through the membrane was measured over time. A column of feed solution (pure water) was brought into contact with the membrane being evaluated and the feed solution was forced through the membrane by applying a constant gas pressure at the top of the water column. By achieving a constant flow of water at a constant applied pressure, the water flux can be determined.
Typically, the membranes to be evaluated are stored in pure water for 12 hours before use. The feed solution (250ml of pure water) was brought into contact with the membrane (membrane contact area 12.19 cm)2). The water column was closed and pressurized with air pressure and the membrane was rinsed with a water column (250 ml). The feed solution was refreshed and a constant gas pressure of 100mbar was applied. Finally, the measurement was performed by monitoring the weight with a scale at a constant flow rate.
II) ion exchange capacity of the Membrane (meq/q) ("IEC")
The membranes were weighed in the dry state before measuring their IEC. The membrane was then stored in a 1.0M KCl solution for 24 hours to completely exchange all possible counterions of the membrane for chloride ions, and then stored in demineralized water for 24 hours. Subsequently, the membrane was equilibrated with 0.1M KBr solution for 24 hours and rinsed with demineralized water for 24 hours. Quantitatively combining the residual KBr solution and the flushing solution of softened water; 1.0 g barium acetate was added and 0.1M AgNO was used3The solution was titrated. The amount of silver ions was measured using an ion selective silver electrode, which resulted in the amount of ions exchanged per unit weight of the membrane.
III) porosity of the film (%)
Apparent density (. rho.) of the filmApparent appearance) And true density determines the porosity of the film being evaluated. The film was weighed and its volume was determined according to its dimensions (length, width and thickness) to measure ρ in airApparent appearance. The true density of the film was determined by densitometric measurement of films having a known weight under a helium atmosphere. Helium gas occupies the pores of the membrane at a known weight and therefore the volume of polymer can be determined. The porosity can thus be determined according to equation (1):
the specific gravity meter used was AccuPyc from Micromeritics Instrument CorporationTMII 1340 gas displacement gravimetric system.
IV) thickness of film (μm)
The thickness of the film was determined by contact mode measurement. Measurements were made at five different locations on the film and the average thickness (in μm) of these five measurements was calculated.
V) pH stability of the film
The pH stability of the membranes was determined by measuring the water flux of the membranes before and after exposing the membranes to aqueous solutions at pH 1,3, 8 and 10 for 16 hours.
The evaluated membranes were stored in pure water (pH 5.5) for 12 to 16 hours, and then their pH stability was measured. The pure water column was closed and pressurized with air pressure, and the membrane was rinsed with a water column (250 ml). Another pure water column was then used as the feed solution to perform the water flux measurements (as described above). The feed solution was brought into contact with the membrane (membrane contact area 12.19 cm)2) And a constant gas pressure of 100mbar is applied. Finally, flux measurements of the membranes were made by monitoring the weight of the filtrate with a scale at a constant flow rate before challenging the membranes with different pH values. The membrane was removed from the apparatus and stored in aqueous hydrochloric acid (when membrane stability was evaluated at pH 1 and pH 3) or aqueous NaOH (when membrane stability was evaluated at pH 8 and pH 10) for 16 hours. The evaluated membrane was removed from the challenge solution and then returned to the device. The feed solution (250ml of pure water column) was contacted with the membrane. The water column was closed and pressurized with air pressure (100mbar) and the membrane was rinsed with a water column (250 ml). The feed solution was refreshed and a constant gas pressure of 100mbar was applied. Finally, measurements after exposure to challenge pH solutions were made by monitoring the weight of the filtrate with a scale at a constant flow rate. Membranes with a water flux difference of less than 10% were considered to have good pH tolerance before and after exposure to all aqueous solutions at pH 1,3, 8 and 10 for 16 hours. Membranes with water flux differences above 10% were considered to have poor pH tolerance before and after 16 hours of exposure to aqueous solutions at pH 1, pH 3, pH 8 and/or pH 10.
Examples 1 to 35
(ai) preparation of the composition
Compositions 1 to 34 were prepared by mixing the ingredients shown in table 1 below in the amounts indicated. In Table 1, component (i) has the structure identified in the description above, component (ii) is as described in Table 1, and component (iv) is IrgacureTM1173. The compositions are each applied to a support (FO-2223-10), as will be described in more detail in this specification.
TABLE 1 compositions
(aii) applying the composition to a support
The compositions described in the above table 1 were each independently applied to the support shown in table 1 at 20 ℃ using a bench coater (manufactured by TQC, model AB3000 automatic film coater). A support was attached to an aluminum plate, and the composition was applied to the support at a speed of about 1cm/sec using a wire rod (stainless steel rod on which 150 μm wire was wound (in the length direction) at 1 turn/3 cm). Excess composition was removed from the coated support using a 12 μm wire rod. A piece of polyethylene was placed on top of the coated support and any air bubbles present in the coating composition were removed by applying a 12 μm wire rod to the polypropylene sheet.
(b) Curing the composition to form a film
The top polypropylene sheet for removing bubbles was removed from the coated support prepared in step (aii), and the composition present on the support was cured by UV irradiation using a Light Hammer LH6 UV exposure machine (manufactured by Fusion UV Systems, inc.). The Light Hammer machine was equipped with an H-type bulb (intensity 100%) and a D-type bulb (intensity 80%). The coated support was passed through a Light Hammer machine at a speed of 10m/min to expose the composition to UV Light from two bulbs. The curing time was 0.8 seconds. The exposure time was two times 0.71 seconds. The first bulb was subjected to the most curing and the second bulb was subjected to the additional curing, thereby improving the mechanical strength of the resulting film. The resulting film was removed from the aluminum plate and stored in a polyolefin bag.
The membrane obtained in step (b) of example 1 had a dry thickness of 118 μm, an ion exchange capacity of 1.17meq/g and a water flux of 793l/m2Bar/hr, average pore diameter 500nm, swelling rate in water 2.3%.
Properties of the resulting film
Example 35
In this example, a membrane containing no support was prepared. In addition, the film is thermally cured rather than light cured.
Preparation of the composition
Composition 35 was prepared by mixing the ingredients shown in table 3 in the amounts indicated.
Mixing composition 35(5.0 cm)3) The placing capacity is 25cm3In a glass vial. The vial was sealed and placed in a vacuum oven at 45 ℃ for 60 minutes. The oven was cooled to room temperature and the vial was removed from the oven. The resulting membrane was taken out from the vial and found to have a dry thickness of 150 μm, an ion exchange capacity of 1.17meq/g, and a water flux of 850l/m2Bar/hr and a swelling ratio in water of 4.8%.
Properties of the resulting film
The film obtained in step (b) above has the properties described in table 2 below:
A)TABLE 2 film Properties
ND means not determined
Measurement of pH tolerance
The pH resistance of the films shown in table 3 below were measured by the above method, and the results (good or bad) are shown in the last column of table 3:
table 3: pH tolerance
Comparative examples 1 to 6
Preparation of the composition
Comparative compositions CEx1 to CEx6 were prepared by mixing the ingredients shown in table 4 below in the indicated amounts. In Table 4, component (i) has the structure identified in the above description, component (ii) consists of the ingredients specified in Table 4, and component (iv) is Irgacure in the amounts indicatedTM1173。
Membranes of comparative examples CEx1 to CEx6 were prepared using steps (aii) and (b) of the process described above for examples 1 to 35, except that the compositions shown in table 4 below were used. The support used in all cases was FO-2223-10.
Table 4: composition used in comparative example
Properties of the comparative film obtained
The films obtained from comparative compositions CEx1 to CEx6 had the properties shown in table 5 below:
table 5: properties of comparative example film