CROSS REFERENCE TO RELATED APPLICATIONS
This application claims priority benefit of U.S. Provisional Application Ser. No. 60/287,172 filed Apr. 27, 2001, the contents of which are incorporated herein by reference.
FIELD OF THE INVENTION
The present invention relates to the field of porous media having a bulk matrix of a first material and a surface coating of a second material. The surface coating comprises a cross-linked terpolymer which has a superior combination of properties, including heat stable biomolecule resistant adsorptive properties, resistance to strong alkaline solutions, and low levels of extractable matter. In some preferred embodiments, the porous media is a porous membrane.
BACKGROUND OF THE INVENTION
Porous media are useful in many separation and adsorption technologies, such as chromatography. One particular type of porous media, porous membranes, are used for a variety of applications. Porous membranes have a first porous surface, a second porous surface, and a continuous porous structure that extends throughout the membrane from the first to the second surface. The continuous porous structure includes the bulk material matrix and the network of pores. The interface separating the bulk matrix from the pore volume (i.e., the surface of the interior pore network) is known as the interstitial surface.
Porous membranes can be classified as “microporous” membranes or “ultrafiltration” membranes on the basis of the size of the pores of the membrane. Generally, the range of pore sizes for microporous membranes is considered to be from approximately 0.05 micron to approximately 10.0 microns, whereas the range of pore sizes for ultrafiltration membranes is considered to be from approximately 0.002 micron to about 0.05 micron. These pore sizes refer to pore diameter for circular or approximately circular pores, or to a characteristic dimension for non-circular pores.
The pore size of a membrane can be denominated by the size of the smallest species (particle or molecule) that cannot pass through the membrane above a specified fractional passage. A common rating is below 10% passage, which corresponds to a 90% cutoff or retention. Other methods are known to those skilled in the art, including image analysis of scanning electron microscopy to obtain pore size distribution characteristics. Microporous membranes are typically used to remove particulates from liquids and gases. An important application of microporous membranes is in sterile filtration of pharmaceutical solutions to remove any bacteria that may be present in the solution. Microporous membranes are also used as sterile gas vents, which allow gas flow but prevent airborne bacteria from passing through the filter. Ultrafiltration membranes are generally used in applications where retention of smaller species is desired. For example, ultrafiltration membranes are used in the biotechnology industry to concentrate proteins, and in diafiltration applications to remove salts and low molecular weight species from protein solutions. Both ultrafiltration and microporous membranes can be fabricated in several forms, including sheets, tubes, and hollow fibers.
Porous membranes are made from a variety of materials, polymers being the most common. Many commercial membranes are made from engineering plastics, such as polyethersulfone, polysulfone, polyvinylidene fluoride, polyethylene, polytetrafluoroethylene, polypropylene and so forth, to take advantage of their robust thermal, mechanical, and chemical-resistance properties. Unfortunately, these materials are hydrophobic and have a high propensity to adsorb biomolecules.
In general, a hydrophilic membrane is preferred for filtration of aqueous solutions. This is because with hydrophobic membranes, contact with a low surface tension prewetting liquid followed by water exchange is required to start permeation. This not only introduces added material cost to the process, but any wetting liquid must be exhaustively flushed, which greatly increases process time and cost.
In addition to permeability and retentive properties, porous membranes have other requirements that are dictated by the nature of the application. For example, they must have sufficient mechanical strength to withstand operating pressures and temperatures. In applications where cleanliness is a major requirement, as in the pharmaceutical or microelectronics wafer-making industry, the amount of material that extracts from the membrane in use must be very small. In applications where the membrane comes in contact with biomolecules, the membrane surface must be resistant to biomolecule adsorption. A biomolecule resistant surface is a surface that adsorbs or binds minimal amounts of biomolecules, such as proteins and nucleic acids; specifically, a surface that adsorbs less than about 30 micrograms of IgG per square centimeter of membrane area as measured by the IgG binding test described herein. It is greatly preferred that a membrane surface be maximally biomolecule resistant, to reduce permeation loss from fouling by surface adsorption or pore blockage, and to prevent product loss by irreversible adsorption or associated biomolecule denaturization.
In many applications, porous membranes comes in contact with highly alkaline solutions in cleaning or sanitation operations. Thus, the membrane must have sufficient chemical resistance to withstand such contact without loss of filtration properties or desirable surface properties.
To impart the aforementioned properties to porous media or membranes, manufacturers typically modify the membrane surface (i.e., the first and second surfaces and the interstitial surface) of the bulk matrix material making up the porous media or membrane to make the surface hydrophilic and biomolecule resistant. This is done by a variety of procedures that coat, attach to, or otherwise cover the surface of the bulk matrix material with a hydrophilic polymer or group. While such modification can solve some problems related to the hydrophobic or high biomolecule binding of the bulk matrix material surface, it also can add other problems. For example, such modifications increase the amount of material able to be extracted from the membrane during use, and the modification material can have low tolerance to exposure to alkaline solutions. In addition, in many applications membranes are heated, either by wet heat as in autoclaving or steam sanitization, or by dry heat, as in a drying step. It has been observed that such heating will reduce hydrophilicity and decrease biomolecule resistance of some modified surfaces.
Some membranes of the prior art are made by modifying the surface of preformed porous membranes with cross-linked hydroxyacrylates, where the crosslinking monomer is a difunctional acrylate (“Case A membranes”). Such membranes have excellent resistance to biomolecule adsorption, excellent heat stability of the biomolecule resistance, and acceptable flow loss (i.e., the reduction in flow or permeability compared to the unmodified membrane). However, while these membranes have acceptable cleanability characteristics (i.e., the ability to clean the membrane by washing such that residual extractable matter (“extractables”) is lowered to an acceptable level, it was found that to lower extractables to below a certain level, about 2 microgram per square centimeter using the TOC test (described herein in the “Methods” section), a very extensive extraction regime was needed. In addition, these membranes were sensitive to strong alkaline solutions.
Much of the prior art describes the use of hydroxyl containing monomers, usually carbonyl ester containing acrylate polymers, to produce membrane surface modifications having hydrophilic character and high resistance to protein binding. However, it is known that polymers from such monomers are not resistant to strong alkaline solutions. For example, a solution of 1.0 normal sodium hydroxide will hydrolyze the carbonyl containing acrylate polymers to acrylic acid containing polymers. Such acrylic acid containing polymers are ionically charged under certain pH conditions, and will attract and bind oppositely charged proteins or biomolecules, thus increasing sorption and membrane fouling. In addition, acrylic acid containing polymers swell in water such that they constrict pore passages, thus reducing membrane permeability and productivity. Moreover, polymers from hydroxyl containing monomers, such as hydroxy acrylates, further react in strong alkaline solutions and degrade into soluble low molecular weight fragments, which dissolve away and expose the underlying porous media or membrane.
Various methods of modifying porous membranes are known in the art. For example, U.S. Pat. No. 4,618,533 discloses and claims a composite porous thermoplastic membrane which comprises a porous membrane substrate having an average pore size between about 0.001 and 10 microns formed of a first polymer, the substrate being directly coated on its entire surface with a cross-linked second polymer formed from a monomer polymerized in situ with a free radical initiator on the substrate, where the composite porous membrane has essentially the same porous configuration as the membrane substrate.
U.S. Pat. No. 4,944,879 discloses a composite porous membrane having desired bulk properties on which is coated a cross-linked polymer having desired surface priorities. The cross-linked surface polymer is produced from a crosslinkable monomer or polymer by energy from an electron beam in the absence of a chemical polymerization initiator.
Similar modified porous media are disclosed in U.S. Pat. Nos. 4,906,374, 4,968,533, and 5,019,260, in which hydroxyl containing polymeric material is derived from monomers having hydroxyl groups and moieties characterized by having one polymerizable unit of unsaturation, such as hydroxy or hydroxy-forming substituted acrylates or methacrylate esters. Polymers from such monomers are known to lack resistance to degradation by strong alkaline solutions.
Chapman et al (J. Am. Chem. Soc. 2000, 122, 8303-8304) describe the use of self assembled monolayers (SAM) to screen functional groups for protein resistance. They report several functional groups to be protein resistant, including N-substituted amide functionalities.
U.S. Pat. Nos. 4,695,592 and 4,678,813 describe a process and product for a hydrophilized porous polyolefin membrane with a crosslinked polymer, which is composed of 50% or more of diacetone acrylamide monomer.
Iwata et al (J. Membrane Sci. 1991 55 119-130) report acrylamide coatings of membranes that have temperature responsive properties (TRP), specifically polyacrylamides, and particularly poly(N-isopropylacrylamide (polyIPAA)). Iwata report the graft polymerization of homopolymers of polyIPAA and copolymers with acrylamide to a first surface of a PVDF membrane. However, they do not cross-link the polymers, as that would impede the polymer TRP.
U.S. Pat. No. 5,929,214 to Peters et al, describes porous monoliths functionalized and/or grafted with TRP polymers, including non-crosslinked copolymers of polyIPAA. These membranes are designed to adsorb biomolecules, and the Peters et al. patent does not teach the production of protein or biomolecule resistant surfaces.
It can be seen that practitioners attempting to develop optimized membranes for sterile filtration and other applications in the pharmaceutical and biotechnology industries have to overcome significant problems. Facing stringent cost, performance and safety requirements, a practitioner has to use materials and develop manufacturing methods that produce membranes with not only optimized flow and retention characteristics, but be economical to produce, meet cleanliness criteria, be stable to the various chemical environments which are commonly encountered, and be very resistant to biomolecule adsorption. Thus, it would be desirable to have a membrane modification that results in a hydrophilic, biomolecule resistant surface that is heat stable, which is resistant to degradation by alkaline solutions, and which has very low levels of material capable of being extracted therefrom. This invention is directed to these, as well as other, important ends.
SUMMARY OF THE INVENTION
This invention is directed to polymeric porous media, preferably porous membranes, that have been modified by forming in situ on the surface thereof a cross-linked polymeric terpolymer coating. In some preferred embodiments, the coated porous media or membranes have substantially the same porous character as the unmodified porous media or membrane, and also have biomolecule resistant sorptive properties, including heat resistant biomolecule resistance, chemical resistance to strong alkaline solutions, and very low levels of extractable matter. In some more preferred embodiments, the modified porous media or membrane is hydrophilic, and does not substantially change pore size as a function of temperature.
Thus, in some preferred embodiments, the present invention provides clean porous membranes comprising a porous substrate and a separately formed heat stable biomolecule resistant surface. In further preferred embodiments, the present invention provides clean porous membranes comprising a porous substrate and a separately formed caustic resistant, heat stable biomolecule resistant surface.
In some preferred embodiments, the porous support and the polymer coating are formed from different materials.
Preferably, the porous substrate is a membrane, more preferably a microporous membrane.
In further preferred embodiments, the invention provides clean, caustic resistant, porous membranes comprising a microporous membrane substrate which is preferably formed from one or more of the group consisting of aromatic sulfone polymers, polytetrafluoroethylene, perfluorinated thermoplastic polymers, polyolefin polymers, ultrahigh molecular weight polyethylene, and polyvinylidene difluoride, and a heat stable biomolecule resistant surface that is a separately formed surface coating which comprises a crosslinked terpolymer, said terpolymer comprising at least two monofunctional monomers selected from the group consisting of acrylamides, methacrylamides, and N-vinyl pyrrolidones, and at least one polyfunctional monomer selected from the group consisting of polyfunctional acrylamides, polyfunctional methacrylamides, and diacroylpiperazines.
In some more preferred embodiments, the invention provides clean, caustic resistant, porous membranes comprising a polyvinylidene difluoride microporous membrane substrate and a heat stable biomolecule resistant surface, wherein said heat stable biomolecule resistant surface is a separately formed surface coating which comprises a crosslinked terpolymer, said crosslinked terpolymer being a copolymer formed from either:
(a) methylene-bis-acrylamide, dimethylacrylamide, and diacetone acrylamide; or
(b) methylene-bis-acrylamide, vinylpyrrolidone, and either of dimethylacrylamide or diacetone acrylamide.
Also provided in accordance with some preferred embodiments of the present invention are methods for the preparation of a clean, caustic resistant porous membrane, said membrane comprising a porous membrane substrate and a heat stable biomolecule resistant surface coating, said method comprising the steps of:
a. providing a porous membrane substrate;
b. optionally washing said porous membrane substrate with a wetting, liquid to wet the surfaces thereof;
c. optionally washing said wet porous membrane substrate with a second wetting liquid to replace said first wetting liquid, leaving said porous membrane substrate wetted with said second liquid;
d. contacting the surfaces of said porous membrane substrate with a reactant solution containing:
(1) at least two monofunctional monomers selected from the group consisting of acrylamides, methacrylamides, and N-vinyl pyrrolidones; and
(2) at least one polyfunctional monomer selected from the group consisting of polyfunctional acrylamides, polyfunctional methacrylamides and diacroylpiperazines;
said solution optionally further comprising one or more polymerization initiators;
e. polymerizing said monomers to form said heat stable biomolecule resistant surface; and
f. washing said membrane.
Preferably, the sizes of the pores of the porous substrate prior to performing steps (a) through (e) are not significantly different from the sizes of said pores after performing steps (a) through (e). In some preferred embodiments, the porous membrane substrate is a microporous membrane.
In some preferred embodiments of the methods and membranes of the invention where the porous substrate is a microporous membrane, the microporous membrane is formed from one or more of the group consisting of aromatic sulfone polymers, polytetrafluoroethylene, perfluorinated thermoplastic polymers, polyolefin polymers, ultrahigh molecular weight polyethylene, and polyvinylidene difluoride, with polyvinylidene difluoride being more preferred.
In some more preferred embodiments of the methods and membranes of the invention, the crosslinked terpolymer comprises at least one monofunctional monomer that is an acrylamide, wherein the acrylamide nitrogen of said acrylamide is substituted with at least one gem dialkyl substituted carbon.
In some particularly preferred embodiments of the methods and membranes of the invention, the crosslinked terpolymer is a copolymer formed from dimethylacrylamide, diacetone acrylamide, and methylene-bis-acrylamide. In other particularly preferred embodiments, the crosslinked terpolymer is a copolymer formed from methylene-bis-acrylamide, N-vinyl pyrrolidone, and either of dimethylacrylamide or diacetone acrylamide.
In some preferred embodiments of the membranes of the invention, the heat stable biomolecule resistant surface of the membranes is a separately formed surface coating comprising a crosslinked terpolymer; the crosslinked terpolymer comprising:
at least one polyfunctional monomer selected from the group consisting of polyfunctional acrylamide monomers, polyfunctional methacrylamide monomers, and diacroylpiperazines; and
at least two different monofunctional monomers selected from the group of N-vinyl pyrrolidone monomers and monomers having the general formula:
alternatively, described as H2C═C(R1)C(═O)N(R2) (R3)
R1 is —H or CH3,
R2 is H or C1-C6, preferably C1-C3 alkyl, either linear or branched,
R3 is H or C1-C6, preferably C1-C3 alkyl, either linear or branched, or C(CH3)2CH2C(═O)CH3, or (P═O)((NCH3)2)2, or C═ON(CH3)2, or CH2—O—R4, where R4 is C1-C5 alkyl, either linear or branched, or (CH2—CH2-0)n—R5, where R5 is —H or —CH3, and n=2 or 3; provided that R2 and R3 are not simultaneously H.
In some preferred embodiments of the methods and membranes of the invention, the crosslinked terpolymer of the membranes of the invention further comprises a supplemental property modifying monomer, which is preferably present in an amount that is less than either of the monofunctional monomers.
In some more preferred embodiments of the methods and membranes of the invention, the supplemental property modifying monomer is selected from the group consisting of positively or negatively charged ion containing monomers, monomers with affinity groups, or monomers with significant hydrophobic character. In further embodiments, the supplemental property modifying monomer is selected from the group consisting of (3-(methacryloylamino)propyl)trimethylammonium chloride, (3-acrylamidopropyl)trimethylammonium chloride, 2-acrylamido-2-methyl-1-propanesulfonic acid and aminopropylmethacrylamide.
In some preferred embodiments of the methods and membranes of the invention, two of the monofunctional monomers of the terpolymer are present in the weight ratio of about 1 to 5, with about 1 to 2 being more preferred.
In further preferred embodiments of the methods and membranes of the invention, the total amount of monofunctional monomers present is from about 0.5% to about 20%, with from about 2% to about 10% being more preferred, and from about 4% to about 8% being even more preferred.
In still further preferred embodiments of the methods and membranes of the invention, the ratio of the total amount of monofunctional comonomers to polyfunctional crosslinker monomer is about 1 to about 10, with about 2 to about 6 being more preferred.
In some preferred embodiments of the membranes of the invention, the heat stable biomolecule resistant surface is hydrophilic.
In some more preferred embodiments of the methods and membranes of the invention, the membranes of the invention have a biomolecule binding of less than about 30 microgram per square centimeter measured by the IgG binding test.
In further preferred embodiments, the membranes of the invention have TOC extractables of less than about 2.0 micrograms of extractable matter per square centimeter of membrane as measured by the NVR Extractables test. More preferably, the membranes of the invention have less than about 1.5, more preferably less than about 1.4, more preferably less than about 1.3, more preferably less than about 1.2, more preferably less than about 1.1, and even more preferably less than about 1.0 micrograms of extractable matter per square centimeter of membrane as measured by the NVR Extractables test.
In further more preferred embodiments of the methods and membranes of the invention, the membranes of the invention have caustic resistance of less than about 1.5, preferably less than about 1.3, more preferably less than about 1.2, and even more preferably less than about 1.0 as measured by the Flow Time Measurement test.
In further preferred embodiments, the present invention also provides methods for removing cells from a solution comprising the steps of providing a solution comprising having undesired cells; and filtering said solution through a membrane of the invention.
In still further preferred embodiments, the present invention also provides methods for sterilizing a solution comprising the steps of providing a non-sterile solution and filtering said solution through a membrane of the invention.
In further embodiments, the present invention provides membranes having a surface coating comprising at least one hydroxymethyldiacetoneacrylamide (HMDAA) monomer of formula:
wherein R1 and R2 are each independently H or CH2OH, preferably wherein R1 and R2 are each CH2OH.
Also provided are methods for preparing a coated polymer membrane comprising the steps of: