EP3519085A1 - Graphene oxide anti-microbial element - Google Patents

Abstract

Described herein is a crosslinked graphene material based element that provides anti-microbial capabilities. Described is an element that can also comprise a support. Also described is an element where the support can be the article to be protected microbes. Also described are methods for killing microbes or for preventing microbial fouling by applying the aforementioned anti-microbial elements and related devices.

Description

GRAPHENE OXIDE ANTI-MICROBIAL ELEMENT

Inventors: Shijun Zheng, Wanyun Hsieh, Isamu Kitahara, Ozair Siddiqui, John Ericson,

Makoto Kobuke, Peng Wang, Yuji Yamashiro

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Application 62/403,601 , filed October 3, 2016; U.S. Provisional Application 62/420,241 , filed November 10, 2016; and U.S. Provisional Application 62/475,487, filed March 23, 2017; the entire disclosures of which are incorporated by reference.

FIELD

[0002] The present embodiments are related to crosslinked graphene oxide membranes and provide membranes with anti-microbial properties.

BACKGROUND

[0003] The growth of microbes in today's society can present serious issues in applications where the level of microbes must be controlled. In applications such as health industry and in water delivery, treatment, and filtration, the growth of microbes to unhealthy levels can result in widespread sickness. Additionally, the growth of microbes in water filtration and delivery apparatuses can also result in biological fouling, reducing the effective lifespan of the equipment. In Heating, Ventilation, and Air Conditioning (HVAC) systems, microbes multiplying in the moist air ducts can lead to foul odor and health problems if left untreated. Also, for vessels in water, unchecked growth of microbes on the vessel's wetted area can reduce the hydrodynamic efficiency of the hull by disrupting the hull shape and creating drag thereby reducing fuel efficiency.

[0004] While there are means for controlling microbes through photo- catalysts, such means require external energy sources such as a source of ultraviolent light to be effective. As a result, there is a need for a passive means of controlling microbe growth.

SUMMARY

[0005] The present embodiments, a crosslinked GO membrane, may reduce the presence of microbes.

-l- [0006] In some embodiments, an anti-microbial membrane can be described as comprising: (1 ) a support, and (2) a composite coating the support comprising a crosslinked optionally substituted graphene oxide compound, where the graphene was crosslinked by a crosslinker selected form the group consisting of an optionally substituted biphenyl of Formula 1 , an optionally substituted triphenylmethane of Formula 2, an optionally substituted diphenylamine or optionally substituted 9H-carbazole represented by Formula 3A or 3B, and an optionally substituted bishydroxy methyl propanediol compound of Formula 4:

Formula 1 ;

where Ri and R₂ are independently NH₂ or OH; and R₃ and R₄ are independently OH, S0₃H, S0₃Na, or S0₃K;

Formula 2; wherein R₅ is H, CH₃, or C₂H₅; R₆ is H, CH₃, -C0₂H, -C0₂Li, -C0₂Na, -C0₂K, -S0₃H, - S0₃Li, -S0₃Na, or -S0₃K; and n is 0, 1 , 2, 3, 4, or 5;

Formula 3A Formula 3B wherein Ri and R₂ can be independently be NH₂, or OH; R₇ and R₈ are independently H, CH₃, C0₂H, C0₂Li, C0₂Na, C0₂K, S0₃H, S0₃Li, S0₃Na, or S0₃K; k is 0 or 1 ; m is 0, 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; and p is 0, 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10;

Formula 4; wherein R₉ , Ri₀, R-n , and Ri₂, can be independently:

wherein R₁₃ is independently OH, NH₂, C0₂H, C0₂Na, C0₂K, S0₃H, S0₃Na, or S0₃K and r is 0, 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; whereby the membrane kills microbes as determined by having an antibacterial effectiveness of 2.0 or more. In some embodiments, the optionally substituted biphenyl can be selected from:

[0007] In some embodiments, the optionally substituted biphenyl can be selected from:

[0008] The membrane of embodiment 1 , wherein the optionally substituted diphenylamine or optionally substituted 9H-carbazole is:

[0009] The membrane of embodiment 1 , wherein the optionally substituted bishydroxymethyl propanediol compound is:

[0010] For some membranes, the optionally substituted graphene oxide comprises platelets. In some embodiments of membranes, the platelets may be between about 0.05 μηη and about 50 μηη. In some embodiments, the mass ratio of graphene oxide to crosslinker in the composite can be a value ranging from 1 :1000 to 50:1. In some embodiments, the support can be the article to be protected from microbial growth.

[0011] In some embodiments, a method of preventing microbial growth can be described, the method comprising: (1) providing the aforedescribed membrane and (2) exposing the membrane to a working fluid containing microbes, wherein the membrane can kill microbes as a result of exposure to the working fluid as determined by having an antibacterial effectiveness of 2.0 or more. In some embodiments, providing the aforedescribed membrane can comprise coating said membrane on the surface to be protected from microbes. In some embodiments, the mass ratio of graphene oxide to crosslinker in the composite can be a value ranging from 1 :1000 to 50: 1. In some embodiments, the support can comprise the article to be protected from microbes. [0012] These and other embodiments are described in greater detail below.

BRIEF DESCRIPTION OF THE DRAWINGS

[0013] FIG. 1 is a diagram showing the dimensions of a graphene platelet.

[0014] FIG. 2 is a depiction of one possible embodiment of an anti-microbial membrane that may be used in anti-microbial applications.

[0015] FIG. 3 is another possible embodiment of an anti-microbial membrane where the support as part of the object protected; the support being the hull of a boat.

[0016] FIG. 4 is yet another possible embodiment of an anti-microbial membrane where the support as part of the object protected; the support being a reverse osmosis membrane.

[0017] FIG. 5 is a depiction of possible method embodiment(s) for preventing microbial growth and/or microbial fouling. The solid lines indicate a possible embodiment and the dashed lines indicate a more specific possible embodiment of the method for preventing microbial growth.

DETAILED DESCRIPTION

[0018] As referred to herein, killing microbes can be measured by the methods used in JIS Z 2801 :2012 (English Version pub. Sep. 2012) where successful killing of microbes by an object can be defined as that object having an antibacterial activity of 2.0 or higher.

[0019] As used herein, the term "selective permeability" refers to a membrane that is relatively permeable for one material and relatively impermeable for another material. For example, a membrane may be relatively permeable to water vapor and relatively impermeable to oxygen and/or nitrogen. The ratio of permeabilities of the different materials may be useful in describing the selective permeability.

[0020] As used herein the term "rest," "resting," or "rested" refers to the act of leaving a solution stand undisturbed at room temperature and atmospheric pressure for a specific duration of time.

[0021] For convenience, the term "molecular weight" is used with respect to a moiety or part of a molecule to indicate the sum of the atomic masses of the atoms in the moiety or part of a molecule, even though it may not be a complete molecule. [0022] As used herein, the term "phenylene" has the broadest meaning generally understood in the art, and may include a cyclic ring or ring system comprising six carbon atoms where there are at least two ring hydrogen substitutions.

[0023] As used herein, the term the term "biphenyl" has the broadest meaning generally understood in the art, and may refer to the cyclic ring or ring system comprising 12 carbon atoms which includes: where there is at least one hydrogen substitution.

[0024] As used herein, the term the term "triphenylmethane" has the broadest meaning generally understood in the art, and may refer to the cyclic ring or ring system comprising 20 carbon atoms which includes:

[0025] As used herein, the term "diphenylamine" has the broadest meaning generally understood in the art, and may include a heterocyclic ring or ring system

comprising

[0026] As used herein, the term "bishydroxymethyl propanediol" has the broadest meaning generally understo a compound

comprising the following core structure o

[0027] As used herein the term "C_X-C_Y" or "C_X._Y" refers to a carbon chain having from X to Y carbon atoms. For example, C1-12 alkyl or C1-C12 alkyl includes fully saturated hydrocarbon containing 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , or 12 carbon atoms. [0028] Layered GO membranes with lamellar structure can be fabricated from a GO aqueous solution, but may be highly susceptible to be dispersed in environments under high flux or with transient shear forces. To solve this issue, the GO sheets can be cross-linked firmly to withstand the shear forces while keeping the lamellar structure.

Anti-Microbial Element

[0029] In some embodiments, an anti-microbial membrane is described. In some embodiments, the membrane can comprise a composite coating. In some embodiments, the membrane can comprise a support and a composite coating on the support material. In some embodiments, the anti-microbial membrane may be selectively permeable. In other embodiments, the membrane is not selectively permeable. In still other embodiments, the membrane is not permeable. In some embodiments, the membrane can have high water vapor permeability. In some embodiments, the membrane may have low water vapor permeability. In some embodiments, the support may be porous. In other embodiments, the support can be non-porous.

[0030] In some embodiments the composite coating may comprise a graphene material and a crosslinker material. In some embodiments, the graphene material may be arranged amongst a polymer material. In some embodiments, the crosslinker material can also be a polymer. In some embodiments, the graphene material and the crosslinker material are covalently linked to one another. In some embodiments, the crosslinker material can be the same material as the polymer material.

[0031] In some embodiments, the graphene material may be arranged amongst other materials in the composite coating in such a manner as to create an exfoliated nanocomposite, an intercalated nanocomposite, or a phase-separated microcomposite. A phase-separated microcomposite phase may be when, although mixed, the graphene material exists as separate and distinct phases apart from the other materials. An intercalated nanocomposite may be when the other compounds begin to intermingle amongst or between the graphene platelets but the graphene material may not be distributed throughout the polymer. In an exfoliated nanocomposite phase the individual graphene platelets may be distributed within or throughout the other materials. An exfoliated nanocomposite phase may be achieved by chemically exfoliating the graphene material by a modified Hummer's method, a process well known to persons of ordinary skill. In some embodiments, the majority of the graphene material may be staggered to create an exfoliated nanocomposite as a dominant material phase. In some embodiments, the graphene material may be separated by about 10 nm, 50 nm, 100 nm to about 500 nm, to about 1 micron.

[0032] In some embodiments, the graphene material may be in the form of sheets, planes or flakes. In some embodiments, the graphene material may be in the form of platelets. In some embodiments, the graphene may have a platelet size of about 0.05 μηη to about 100 μηη. In some embodiments, the graphene material may have a surface area of between about 100 m²/g to about 5000 m²/g. In some embodiments, the graphene material may have a surface area of about 150 m²/g to about 4000 m²/g. In some embodiments the graphene material may have a surface area of about 200 m²/g to about 1000 m²/g, e.g. , about 400 m²/g to about 500 m²/g.

[0033] In some embodiments, the graphene oxide may be platelets having one or more dimensions in the nanometer to micron range. In some embodiments, as shown in Figure 1 , the platelets may have dimensions in the x, y and/or z dimension. For example, the platelets may have: an average x dimension between about 0.05 um to about 50 um, or any value in a range bounded by, or between, any of these lengths; an average y dimension of 0.05 um to about 50 um, or any value in a range bounded by, or between, any of these lengths. In some embodiments, the graphene oxide comprises GO platelets, the platelets defining an average size of about 0.05 μηη to about 50 μηη.

[0034] In some embodiments, the graphene material may not be modified and may comprise of a non-functionalized graphene base. In some embodiments, the graphene material may comprise a modified graphene. In some embodiments, the modified graphene can comprise an optionally substituted graphene material. In some embodiments, the optionally substituted graphene material may comprise a functionalized graphene. In some embodiments, more than about 90%, about 80%, about 70%, about 60% about 50%, about 40%, about 30%, about 20%, about 10% of the graphene may be functionalized. In other embodiments, the majority of graphene material may be functionalized. In still other embodiments, substantially all the graphene material may be functionalized. In some embodiments, the functionalized graphene may comprise a graphene base and functional compound. A graphene may be "functionalized," becoming functionalized graphene when there is one or more types of functional compounds not naturally occurring on GO are substituted instead of hydroxide in the acetic acid groups of one or more hydroxide locations in the graphene matrix. In some embodiments, the graphene base may be selected from reduced graphene oxide and/or graphene oxide. In some embodiments, the graphene base may be selected from:

reduced Graphene Oxide [RGO],

Graphene oxide [GO],

and/or Graphene.

[0035] In some embodiments, multiple types of functional compounds are used to functionalize the graphene material in addition to comprising at least one epoxide group. In other embodiments, only one type of functional compound can be utilized to functionalize the graphene material. In some embodiments, the functional compounds comprise an epoxide group.

[0036] In some embodiments, the epoxide group may comprise a epoxide- based compound having the functional group:

[0037] In some embodiments, the epoxide groups can be the by-product of oxidation of the graphene to create graphene oxide. In some embodiments, the epoxide groups are formed on the surface of the graphene base by additional chemical reactions. In some embodiments, the epoxide groups are a mix of those formed during oxidation and those formed by additional chemical reactions.

[0038] In some embodiments, the graphene material may be a crosslinked graphene, where the graphene material may be crosslinked with at least one other graphene base by a crosslinker material/bridge. In some embodiments, the graphene material may comprise crosslinked graphene material where at the graphene bases are crosslinked such that at least about 1 %, about 5 %, about 10 %, about 20 %, about 30 %, about 40 % about 50 %, about 60 %, about 70 %, about 80 %, about 90 %, about 95 %, or all of the graphene material may be crosslinked. In some embodiments, the majority of the graphene material may be crosslinked. In some embodiments, some of the graphene material may be crosslinked such that at least 5 % of the graphene material may be crosslinked with other graphene material. The amount of crosslinking may be estimated by the wt% of the crosslinker/precursor as compared with the total amount of polymer present. In some embodiments, one or more of the graphene base(s) that are crosslinked may also be functionalized. In some embodiments the graphene material may comprise both crosslinked graphene and non-crosslinked, functionalized graphene.

[0039] In some embodiments, the adjacent graphene oxide material can be covalently bonded to each other by one or more crosslinks. In some embodiments, the graphene oxide material can be bonded to the support covalently and/or by Van der Waals forces. In some embodiments, the crosslinks can be a product of a crosslinker compound (CLC). In some embodiments, the crosslinker can comprise a crosslinker selected from the group:

H H H N Link N _ N Link O _ and O Link O ; wherein Link can be the body of the crosslinker. In some embodiments, the resulting linkage can be represented as:

H H H GO N Link N GO , GO N Link O GO , and/or

GO O Link O GO^■ wherein GO represents an optionally substituted graphene oxide and Link can be the body of the crosslinker. In some embodiments, the cross-link can be made by a crosslinker to create a covalent linkage that links two or more optionally substituted graphene oxides.

[0040] In some embodiments, the crosslinker compound (CLC) containing nucleophilic groups can comprise an optionally substituted biphenyl, optionally substituted triphenylmethane, optionally substituted diphenylamine, optionally substituted 9H-carbazole, or optionally substituted 2,2-bis(hydroxymethyl)propane-1 ,3- diol. While not wanting to be bound by theory the presence of a nucleophilic group may increase the reactivity of the corresponding position to an epoxide group on the graphene platelet. In some embodiments, the crosslinker can crosslink at least one of the -NH and/or -OH substituents at Ri , R₂, R3 and/or R₄, for example, two adjacent graphene oxides, three adjacent graphene oxides, or four adjacent graphene oxides. In some embodiments, the crosslinker can crosslink at least one of the -NH and/or -OH substituents at Ri and/or R₂, for example, two adjacent graphene oxides. In some embodiments, Ri and R₂ are independently NH₂ or OH. In some embodiments, Ri and R₂ are both NH₂. In some embodiments, Ri and R₂ are both OH.

[0041] In some embodiments, suitable crosslinkers include potassium tetraborate ("KBO"), a benzoic acid derivate (e.g., 3,5-diaminobenzoic acid ("DABA")), and 2,5-dihydroxyterephthalic acid ("DHTA"), which can be used individually or in combination with each other or other crosslinkers.

[0042] In some embodiments, the crosslinker can be an optionally substituted biphenyl represented by Formula 1.

Formula 1 ; wherein R₃ and R₄ can be independently H, OH, NH₂, CH₃, -C0₂H, -C0₂Li, -C0₂Na, - C0₂K, -SO₃H, -SO₃U, -S0₃Na, or -SO₃K. In some embodiments, at least two of Ri , R₂, R₃, and R₄ can be a nucleophilic group. In some embodiments, the site of the nucleophilic group can be the location of the covalent linkage with the epoxide. In some embodiments, Ri and R₂ can be independently a nucleophilic group, for example, NH₂ or OH. In some embodiments, R₃ and R₄ can be independently OH, S0₃H, S0₃Na, or S0₃K. In some embodiments, R₃ and R₄ can be independently OH, S0₃Na, or S0₃K. In some embodiments, the substituted biphenyl can comprise:

[CLC-1.1], [CLC-1.2], and/or

[0043] In some embodiments, the crosslinker can be an optionally substituted triphenylmethane represented by Formula 2:

Formula 2;

wherein R₅ can be H, CH₃, or C₂H₅; R₆ can be H, CH₃, -C0₂H, -C0₂Li, -C0₂Na, - C0₂K, -S0₃H, -S0₃Li, -S0₃Na, or -S0₃K; and n can be 0, 1 , 2, 3, 4, or 5. In some embodiments, R₅ can be H, CH₃, or C₂H₅. In some embodiments, R₅ can be CH₃. In some embodiments, R₆ can be independently H, CH₃, OH, or an organic acid group or a salt thereof, such as -C0₂H, -C0₂Na, -C0₂Li, -C0₂K, -S0₃H, -S0₃Na, -S0₃Li, or -S0₃K. In some embodiments, R₆ can be S0₃Na. In some embodiments, n can be 4. In some embodiments, at least two of Ri, R₂, R₅, and R₆ can be a nucleophilic group. In some embodiments, the site of the nucleophilic group can be the location of the covalent linkage with the epoxide. In some embodiments, Ri and R₂ can be independently a nucleophilic group, for example, NH₂ or OH. In some embodiments, R₅ can be an alkyl group.

[0044] In some embodiments, the optionally substituted triphenylmethane can comprise:

[0045] In some embodiments, the crosslinker is an optionally substituted diphenylamine or optionally substituted 9H-carbazole represented by formula 3A or 3B:

ormu a Formula 3B wherein R₇ and R₈ can be independently H, CH₃, C0₂H, C0₂Li, C0₂Na, C0₂K, S0₃H, S0₃Li, S0₃Na, or S0₃K; k can be 0 or 1 ; m can be 0, 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; and p can be 0, 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10. With respect to any relevant structural representation, such as Formula 3A or 3B, a dashed line represents the presence or absence of a bond. For example, compounds represented by Formulas 3B-1 and 3B-2 as shown below are included.

[0046] In some embodiments, at least two of Ri, R₂, R₆, and R₇ can be a nucleophilic group. In some embodiments, the site of the nucleophilic group can be the location of the covalent linkage with the epoxide. In some embodiments, Ri and R₂ can be independently a nucleophilic group, for example, NH₂ or OH. In some embodiments, R₇ and R₈ can be independently H, CH₃, or an organic acid group or a salt thereof, such as -C0₂H, -C0₂Na, -C0₂Li, -C0₂K, -S0₃H, -S0₃Na, -S0₃Li, or -S0₃K. In some embodiments, R₆ and R₇ can be independently -S0₃K. In some embodiments, R₇ and R₈ can be both -S0₃K. In some embodiments, k can be 0. In some embodiments, k can be 1 . In some embodiments, m can be 0. In some embodiments, m can be 3. In some embodiments, p can be 0. In some embodiments, p can be 3. In some embodiments, m and p can be both 0. In some embodiments, m and p can be both 3.

[0047] In some embodiments, the optionally substituted diphenylamine or optionally substituted 9H-carbazole can be:

[0048] In some embodiments, the optionally substituted bishydroxy methyl propanediol compound can be described by formula 4:

, formula 4; wherein R₉ , Ri₀, R-n, and Ri₂, can be independently:

wherein in an embodiment, each Ri₃ can independently be a nudeophilic group. In some embodiments, the site of the nudeophilic group can be the location of the covalent linkage with the epoxide. In some embodiments, Ri₃ can be independently OH, NH₂, C0₂H, C0₂Na, C0₂K, S0₃H, S0₃Na, or S0₃K. While not wanting to be bound by theory the presence of a nudeophilic group may increase the reactivity of the corresponding position to an epoxide group on the graphene platelet. In some embodiments, r can be 0, 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10.

[0049] In some embodiments, the optionally substituted bishydroxy methyl propanediol compound can comprise:

[0050] In some embodiments, the resulting linkage can be created by a substitution reaction, wherein an epoxide functional group of the functionalized graphene oxide can be opened. While not wanting to be limited by theory, opening the epoxide ring and may result in a carbon becoming covalently bonded to the crosslinker, taking the place of a hydrogen atom in the NH₂ group or hydrogen in an OH group. While not wanting to be limited by theory, the inventors believe that C-N bonding to the epoxide functional groups instead of forming amide bonds will result in higher incidences of crosslinking between vertically stacked graphene oxide (i.e., crosslinks normal to the graphene's surface) because amide bonds may depend on the presence of a carboxyl groups that are predominantly on the edge of the graphene instead of in the body or planar interior of the graphene and may provide in-plane bonding between adjacent graphene materials. In some embodiments, the reaction for the crosslinker and the optionally substituted graphene oxide can form a crosslink vertically between stacked optionally substituted graphene oxides. While not wanting to be limited by theory, it is thought that for antimicrobial applications one of many potential mechanisms for killing microbes can be the presence of active sites on the graphene platelets and that the crosslinker can be chosen such that active sites on the graphene are not completely consumed by the crosslinking process, thus allowing for the generation of reactive species. In some embodiments, the amount or reactivity of the crosslinker can be chosen so as to ensure the existence of graphene active sites. A non-limiting example can be represented as:

-16-

[0051] In some embodiments, the crosslinker material comprises an aqueous solution of about 2 wt% to about 50 wt% crosslinker. In some embodiments, the crosslinker material comprises an aqueous solution of about 2.5 wt% to about 30 wt% crosslinker. In some embodiments, the crosslinker material comprises an aqueous solution of about 5 wt% to about 15 wt% crosslinker.

[0052] In some embodiments, the weight ratio of optionally substituted graphene oxide to substituted biphenyl can be from about 1 : 1000 to about 50: 1 . In some embodiments, the weight ratio of optionally substituted graphene oxide to substituted biphenyl can be from about 1 : 100 to about 15: 1 . In some embodiments, the weight ratio of optionally substituted graphene oxide to substituted biphenyl can be about 1 : 15 to about 1 : 1 . In some embodiments, the weight ratio of optionally substituted graphene oxide to substituted biphenyl can be about 1 1 to about 1 : 1 .

[0053] In some embodiments, the crosslinker can crosslink a first interior carbon atom on a face of a first optionally substituted graphene oxide platelet to a second interior carbon atom on a face of a second optionally substituted graphene oxide platelet. An interior carbon atom on a face of an optionally substituted graphene oxide platelet is a carbon atom that is not on an outer border of the optionally substituted graphene oxide platelet. For example, for the graphene oxide platelet depicted below, the interior carbon atoms are shown in bold. It should be noted that the structure below is depicted only to illustrate the principle of an interior carbon atom, and does not limit the structure of graphene oxide.

[0054] In some embodiments, the support can be a part of the membrane. Non limiting examples of such supports include reverse osmosis membranes, tapes, or anything that can be used as a substrate, either flexible or non-flexible. In some embodiments, the support material may be polymeric. In some embodiments, the support material can comprise hollow fibers. In other embodiments, the support can be the article to be protected from microbial growth. In some embodiments, the article to be protected can be any item where biological growth is undesirable. Examples include but are not limited to ship hull's, treatment basins, pipes, desalination filters, air filters, HVAC system components, hospital equipment and furnishings, counter-tops, lavatory furnishings, and the like.

[0055] In some embodiments, where the support may comprise a porous material. In some embodiments, the support can comprise a non-porous material. In some embodiments, the material may be polymeric. In some embodiments, the polymer may be polyamide, polyvinylidene fluoride, polyethylene terephthalate, polysulfone, polyether sulfone, and/or mixtures thereof. In some embodiments, the porous support can comprise a polyamide (e.g. Nylon). In some embodiments, the porous material may be a polysulfone based ultrafiltration membrane. In some embodiments, the porous material can be polyvinylidene fluoride. In some embodiments, the porous material may comprise hollow fibers. The hollow fibers may be cast or extruded. The hollow fibers may be made, for example, as described in United States Patent Nos., 4,900,626 and 6,805,730 and United States Patent Publication No. 2015/0165,389, which are all incorporated by reference in their entireties. [0056] In some embodiments, the gas permeability of the membrane may be less than 0.100 cc/m²-day, 0.010 cc/m²-day, and/or 0.005 cc/m²-day. A suitable method for determining gas permeability is disclosed in United States Patent Publication US2014/0272.350, ASTM D3985, ASTM F1307, ASTM 1249, ASTM F2622, and/or ASTM F1927, which are incorporated by reference in their entireties for their disclosure of determining gas (oxygen) permeability %, e.g., oxygen transfer rate (OTR).

[0057] In some embodiments, the moisture permeability of the membrane may be greater than 10.0 gm/m²-day, 5.0 gm/m²-day, 3.0 gm/m²-day, 2.5 gm/m²-day, 2.25 gm/m²-day and/or 2.0 gm/m²-day. In some embodiments, the moisture permeability may be a measure of water vapor permeability/transfer rate at the above described levels. Suitable methods for determining moisture (water vapor) permeability are disclosed in Caria, P. F., Ca test of Al₂0₃ gas diffusion barriers grown by atomic deposition on polymers, Applied Physics Letters 89, 031915-1 to 031915-3 (2006), ASTM D7709, ASTM F1249, ASTM398 and/or ASTME96, which are incorporated by reference in their entireties for disclosure of determining moisture permeability %, e.g., water vapor transfer rate (WVTR).

[0058] In some embodiments, the selective permeability of the membrane may be reflected in a ratio of permeabilities of water vapor and at least one selected gas, e.g., oxygen and/or nitrogen, permeabilities. In some embodiments, the membrane may exhibit a water-vapor permeability to gas permeability ratio, e.g., WVTR/OTR, of greater than 50, greater than 100, greater than 200, and/or greater than 400. In some embodiments, the selective permeability may be a measure of water vapor: gas permeability/transfer rate ratios at the above described levels. Suitable methods for determining water vapor permeability and/or gas permeability have been disclosed herein.

[0059] In some embodiments, the membrane can have anti-microbial properties, or kill microbes in a working fluid. In some embodiments, the microbes killed can comprise escherichia coli (ATCC® 8739, American Type Culture Collection (ATCC), Manassas, VA USA). In some embodiments, the membrane can have an antibacterial effectiveness of 2.0 or more. The antibacterial effectiveness can be determined by standard JIS Z 2801 (2010). In some embodiments, the working fluid can be either liquid, gas, or a combination thereof (e.g., saturated air). Non-limiting examples of a liquid working fluid can be the brine/salt water or fresh water in a desalination plant, water in a waste treatment plant, ocean water for a ship, air in a HVAC system, or air in an enclosed space.

[0060] In some embodiments, the anti-microbial membrane may be disposed between an object to be protected and a fluid reservoir. In some embodiments, the fluid reservoir can contain microbes. In some embodiments, the membrane can kill microbes on the membrane.

[0061] In some embodiments, solvents may also be present in the antimicrobial element. Used in manufacture of material layers, solvents include, but are not limited to, water, a lower alkanol such as but not limited to ethanol, methanol, isopropyl alcohol, xylenes, cyclohexanone, acetone, toluene and methyl ethyl ketone, and mixtures thereof. Some embodiments can use water as a solvent.

[0062] In an embodiment, as seen in Figure 2, the anti-microbial membrane, 100, may comprise at least a substrate element, 120, and the aforementioned composite coating, 110. The coating is exposed to the working fluid, 130. In some embodiments, as shown in Figures 3 and 4, the substrate, 120, can comprise the article to be protected from microbes. In Figure 3 the article to be protected is a reverse osmosis membrane and the membrane is on the surface of the membrane. In Figure 4 the article to be protected is the hull of a ship and the membrane is a coating on the hull. As a result of the membrane, the growth of microbes on said membrane and on the protected article can be precluded by killing of the microbes by the membrane.

[0063] In some embodiments, a material may be included in the antimicrobial membrane 100 to increase or improve the interaction membrane 100 has with the working fluid 130. In some embodiments, the added material or spacer material may improve the flux or movement of the working fluid over or through membrane 100. In some embodiments, the added material creates space or volume within the anti-microbial membrane 100. In some embodiments, the added material creates or increases the roughness or irregularity of the surface of the anti-microbial membrane 100. In some embodiments, the added material is silica, such as silica nanoparticles, or another suitable material that creates the desired fluid flux or surface texture. In some embodiments utilizing silica particles or nanoparticles, the size of the particles can be between 1 nm and 500 nm, between 40 nm and 300 nm, or between 70 nm and 250 nm. In some embodiments, the particle size is 5 nm, 7 nm, 10 nm, 20 nm, 60 nm, 80 nm, 100 nm, 120 nm, 140 nm, 160 nm, 180 nm, 200 nm, or 220 nm. In addition, other nanoparticles having similar size and behavioral characteristics include nanoparticles of Fe₃0₄, ΤΊΟ2, Zr0₂, or AI2O3. In some embodiments, the spacer material has a weight percentage of about 1 % to about 10% relative to the total weight of composite coating 110. In some embodiments, the spacer material has a weight percentage of about 6% or about 6.6% relative to the total weight of composite coating 110.

[0064] In some embodiments, composite coating 110 has a thickness ranging from about 10 nm to about 10 μιη. Composite coating 110 can have a thickness of 50 nm, 100 nm, 1 10 nm, 150 nm, 180 nm, 200 nm, 220 nm, 300 nm, 400 nm, 500 nm, 600 nm, 1 μιη, 1 .4 μιη, 5 μιη, or any value close to or between these values. In some embodiments, the thickness is less than about 20 μιη, less than about 15 μιη, less than 10, or less than about 5 μιη. In some embodiments, composite coating 110 is not thick enough to be self-supported. In other words, in some embodiments, composite coating 110 must be applied to or adhered to a support structure or surface, such as substrate element 120.

[0065] In some embodiments, membrane 100 is prepared by applying composite coating 110 to substrate element 120 and then exposing the resulting membrane to an elevated temperature for a period time. In some embodiments, this process cures membrane 100. In some embodiments, after being applied to substrate element 120 composite coating 110 is allowed to air dry for a period of time before being exposed to an elevated temperature. In some embodiments, the elevated temperature ranges from about 30 °C to about 300 °C, from about 60 °C to about 200 °C, or from about 70 °C to about 150 °C. In some embodiments, the elevated temperature is about 70 °C, about 85 °C, about 90 °C, about 130 °C, about 140 °C, or any value close to or between these values. In some embodiments, the period of exposure is from about 1 minute to about 180 minutes, from about 2 minutes to about 150 minutes, from about 3 minutes to about 120 minutes. In some embodiments, the period of exposure is about 3 minutes, about 6 minutes, about 8 minutes, about 20 minutes, about 30 minutes, about 60 minutes, about 90 minutes, about 120 minutes, or any value close to or between these values. Method for Protecting an Article from Microbes

[0066] In some embodiments, a method for killing microbes on a surface can be described, as shown in Figure 5. In some embodiments, the method can comprise providing any of aforedescribed antimicrobial membranes. In some embodiments, providing any of the aforedescribed membranes can comprise coating the surface to be protected with any of the said membranes. In some embodiments, the membrane can comprise a composite coating. In some embodiments, the membrane can comprise a support and a composite coating on the support material. In some embodiments, the composite coating can comprise graphene oxide and a crosslinker. In some embodiments, the crosslinker can comprise potassium tetraborate, 3,5-diaminobenzoic acid, 2,5-dihydroxyterephthalic acid, an optionally substituted biphenyl, optionally substituted triphenylmethane, optionally substituted diphenylamine, optionally substituted 9H-carbazole, or optionally substituted 2,2-bis(hydroxymethyl)propane-1 ,3- diol as described elsewhere in this application. In some embodiments, the substituted biphenyl can be described by Formula 1 :

Formula 1.

[0067] In some embodiments, the substituted biphenyl can comprise:

[CLC-1.1], [CLC-1.2], and/or

[0068] In some embodiments, the crosslinker can be an optionally substituted triphenylmethane represented by Formula 2:

Formula 2.

[0069] In some embodiments, the optionally substituted triphenylmethane can comprise:

[0070] In some embodiments, the crosslinker is an optionally substituted diphenylamine or optionally substituted 9H-carbazole represented by Formula 3A or 3B:

[0071] In some embodiments, the optionally substituted diphenylamine can comprise:

[0072] In some embodiments, the optionally substituted bishydroxy methyl propanediol compound can be described by Formula 4:

, Formula 4; wherein R₉ , Ri₀, Rn , and Ri₂, can be independently:

[0073] In some embodiments, the optionally substituted bishydroxy methyl propanediol compound can comprise:

[0074] In some embodiments, the mass ratio of graphene oxide to crosslinker can be from 1 : 1000 to 50:1 . In some embodiments, the mass ratio of graphene material to crosslinker can range from about 1 : 100 to about 15:1. In some embodiments, the mass ratio of graphene material to crosslinker can range from about 1 :15 to about 1 :1.

In some embodiments, the support can be porous. In other embodiments, the support can be non-porous. In some embodiments, the support can be part of the coating. In other embodiments, the support can be separate from the coating. In some embodiments, where the support is separate from the coating, the support can comprise the article to be protected from microbes. Examples include but are not limited to ship hull's, treatment basins, pipes, desalination filters, air filters, HVAC system components, hospital equipment and furnishings, counter-tops, lavatory furnishings, and the like.

[0075] In some embodiments, the method further comprises exposing the membrane to a working fluid. In some embodiments, the working fluid can contain microbes, whereby the membrane kills microbes as a result of exposure to the working fluid. In some embodiments, the microbes controlled can comprise escherichia coli (ATCC® 8739, ATCC). In some embodiments, the membrane can have an antibacterial effectiveness of 2.0 or more. The antibacterial effectiveness can be determined by standard JIS Z 2801 (2012). In some embodiments, the working fluid can comprise air. In some embodiments, the working fluid can comprise water. In some embodiments, the working fluid can comprise a mixture of air and water vapor. In some embodiments, the mixture of air and water vapor can have a relative humidity ranging from about 100 % to about 0 %. In some embodiments, the relative humidity can range from 0 %, 20 %, 50 %, 60 %, 78 %, 80 %, to 90 %, to 93 % to 100%, or any combination thereof.

EXAMPLES

[0076] It has been discovered that embodiments of the anti-microbial membrane elements described herein have improved resistance to microbes. These benefits are further shown by the following examples, which are intended to be illustrative of the embodiments of the disclosure, but are not intended to limit the scope or underlying principles in any way.

Example 1.1.1 : Preparation of Graphene Oxide

[0077] GO Preparation: GO was prepared from graphite using modified Hummers method. Graphite flake (2.0g, Aldrich, 100 mesh) was oxidized in a mixture of NaN0₃ (2.0g), KMn0₄ (10g) and concentrated 98% H₂S0₄ (96 mL) at 50 °C for 15 hours. The resulting pasty mixture was then poured into ice (400g) followed by the addition of 30% hydrogen peroxide (20 mL). The resulting solution was stirred for 2 hours to reduce the manganese dioxide, filtered through filter paper, and washed with deionized (Dl) water. The solid was collected and dispersed in Dl water by stirring, centrifuged at 6300 rpm for 40 minutes, and demayted the aqueous layer. The remaining solid was dispersed in Dl water, and washing process repeated 4 times. The purified GO was then dispersed in Dl water under sonication (20 W) for 2.5 hours for a GO dispersion (0.4% wt).

Example 1.1.2: Synthesis of Crosslinker Compound #1 .1 (CLC-1.1)

[0078] '-Diamino-ttr-biphenyll^'-diol (CLC-1.1) Preparation: To a mixture of 3-nitrophenol (6.95g, 0.05mol) (Aldrich) in ethanol (50 ml) (Aldrich), was added 12M NaOH aqueous solution (25ml) (Aldrich), and zinc powder (13g, 0.2 mol) (Aldrich) under a nitrogen atmosphere (Airgas, San Marcos, CA, USA). The resulting mixture was stirred for 10 hours, then filtrated. The filtrate was acidified by acetic acid (Aldrich) to pH of 4, and a precipitate formed. The solid was collected by filtration, washing with water to neutral, and drying under vacuum using a vacuum oven (Thermo Scientific Precision 6500, Thermo Fisher Scientific Waltham, MA USA) at 60 °C at 2 torr to afford desired product (3.8g, 70% yield), or CLC-1.1. The compound was characterized by LCMS: calc'd for C₁₂H₁₃N202 (M+H): 217.1 ; found: 217. H NMR (DMSO): δ 8.9 (bs, 4H), 6.77 (bs, 2H), 6.15 (bs, 4H).

Example 1.1.3: Synthesis of Crosslinker Compound #1.2 (CLC-1.2)

[0079] Potassium 4,4'-dihydroxy-[1,1^,-biphenyl]-2,2^,-disulfonate (CLC-2) Preparation: A mixture of 4,4'-diaminobiphenyl-2,2'-disulfonic acid (9.5 g, 27.6 mmol, Aldrich) and sodium hydroxide (2.08 g, 52 mmol, Aldrich) were dissolved in 65 mL Dl water, 59 g ice was charged to the solution. Then sodium nitrite (3.52 g, 51 mmol, Aldrich) was added, followed by a solution of sulfuric acid (1 1.5mL H₂S0₄ (98%, Aldrich) in 59 mL Dl water) was added drop wise at 0 °C. The solution turned red and then a yellow precipitate formed. The mixture was stirred at 0 °C for one hour. The suspension was then filtered; the solid of diazonium salt washed with 120 mL of icy-water. The salt was then suspended in 40 mL water, and heated to 100 °C for one hour. All of solid was dissolved; the solution was then cooled to room temperature, and neutralized by KOH solution (2.3 g in 5 mL water, Aldrich) to pH of 7. The resulted solution was added to ethanol/isopropanol (300 mL/100 mL) (Aldrich) for extraction and a yellow-orange precipitate formed. After filtration and drying in a vacuum, a yellow solid, CLC-1.2, was obtained (6.7g, in 60% yield). Confirmed by H NMR (D₂0): δ 7.39 (dd, 2H), 7.18 (dd, 2H), 6.95 (dd, 2H).

Example 1.1.4: Preparation of Crosslinker Compound #1 .3 (CLC-1.3)

CLC-1.3

[0080] Sodium '-diamino-tl '-biphenylJ^^'-disulfonate (CLC-1.3) Preparation: To a solution of 4,4'-diaminobiphenyl-2,2'-disulfonic acid (4.82 g, 14 mmol, Aldrich) in 30 mL methanol (Aldrich), NaOH aqueous solution (1.12 g, 28 mmol in 10 mL water) (Aldrich) was added. The whole mixture was stirred for 30 min and then the solvents were removed using a vacuum oven (Thermo Scientific Precision 6500, Thermo Fisher) at 60 °C at 2 torr to give a solid (5.44 g, 100% yield), or CLC-1.3.

Example 1.1.5: Preparation of Crosslinker Compound #2.1 (CLC-2.1)

CLC-2.1 [0081] Sodium 4-(4-(1,1-bis(4-hydroxyphenyl)ethyl)phenoxy)butane-1- sulfonate (CLC-2.1) Preparation: To a stirring quantity of tert-butanol (90 mL) (Aldrich) at room temperature, 4,4',4"-(ethane-1 ,1 ,1-triyl)triphenol (5 g, 16 mmol) (Aldrich) was added followed by sodium tert-butoxide (1.57g, 16 mmol) (Aldrich). The mixture was then stirred at 1 10 °C for 15 minutes. Subsequently 1 ,4-butanesultone (1.67mL, 16 mmol) was added to the mixture and the reaction was let go overnight. After about 16 hours, the reaction was then cooled. To the solution hexanes (200 mL) (Aldrich) were added and the solution was stirred for 30 minutes. The collected precipitate was then washed again for 30 minutes in fresh hexanes solution (about 500 mL) (Aldrich). Then the collected precipitate was put in isopropanol (400 mL) (Aldrich) and then stirred for 2 hours. The final step added hexanes (400 mL) (Aldrich) to the solution and after 5 minutes of stirring, the precipitate was collected. The product was dried at 60 °C at 2 torr in a vacuum oven (Thermo Scientific Precision 6500, Thermo Fisher Scientific Waltham, MA USA) overnight to give a white powder (6.25 g, 82.4% yield), or CLC-2.1. H-NMR (D₂0): δ 1.63 (4H, m), 1 .77(31-1, s), 2.72 (2H, t), 3.65 (2H, t), 6.51 (6H, t), 6.76 (6H, t).

Example 1.1.5: Preparation of Crosslinker Compound #2.1 (CLC-3.1)

CLC-3.1

[0082] N1, N3-bis(4-nitrophenyl)benzene-1,3-diamine (CLC-3.1) Preparation: A mixture of 4-fluoro-1 -nitrobenzene (10.6 mL, 100 mmol) (Aldrich), meta- phenylenediamine (5.4g, 50 mmol) (Aldrich) and potassium carbonate (16.6g, 120 mmol) (Aldrich) in anhydrous dimethyl sulfoxide (DMSO) (80 mL) (Aldrich) was heated to 105 °C for 20 hours. The resulting mixture was poured into water (250 mL) slowly and then extracted with dichloromethane (500 mL) (Aldrich). The organic was washed with brine, dried over Na₂S0₄, then loaded on silica gel (Aldrich) for flash column chromatography using eluents of dichloromethane/hexanes (1 :10 to 3:2) (Aldrich). The desired fractions were collected and concentrated to give orange solid (4.8 g, in 27% yield), CLC-3.1. Confirmed by LCMS: calc'd for Ci₈Hi₄N₄0₄: 350.1 ; Found: 350. Example 1.1.3: Synthesis of Crosslinker Compound #2.2 (CLC-3.1).

ONa

CLC-4.1

[0083] Sodium 4-(2-(3-(2-hydroxyethoxy)-2,2-bis((2-hydroxyethoxy)- methyl)propoxy)ethoxy)butane-1-sulfonate (CLC-3.1 ) Preparation: While stirring a solution of te/ -butanol (100ml_) (Aldrich) at room temperature, pentaerythritol ethoxylate (7g, 22.4 mmol) (Aldrich 416150, M_n=270 avg, ¾ EO/OH) was added followed by sodium te/ -butoxide (2.15g, 22.4 mmol) (Aldrich). Continuing stirring, the mixture was then heated to 1 10 °C for 70 minutes. Subsequently, 1 ,4-butanesultone (2.29ml_, 22.4 mmol) (Aldrich) was added to the mixture and the reaction let go overnight. After 17 hours of reaction, the excess solution was decanted. The precipitates were then washed with hexanes (Aldrich). The precipitates were then dissolved in methanol (125 mL) (Aldrich) and dried in vacuo in a 50 °C bath giving a viscous, transparent wax, or CLC-3.1 (8.77 g, yield 73%). H-NMR (D₂0): δ 1.7-1.8(41-1, m), 2.90 (2H, t), 3.3 (8H, s), 3.4-3.7 (21 H, m).

Example 1.1.4: Synthesis of Crosslinker Compound #4 (CLC-4.2).

IC-1 [0084] Dimethyl 4,4'-((2,2-bis((4-(methoxycarbonyl)phenoxy)methyl)- propane-1 ,3-diyl)bis(oxy))dibenzoate (IC-1): Into Λ/,Λ/'-dimethylformamide (100 mL) (Aldrich) at room temperature, pentaerythritol tetrabromide (6 g, 15.5 mmol, Aldrich) was added with stirring followed by methyl 4-hydroxybenzoate (9.42 g, 61.9 mmol, Aldrich), and then potassium carbonate (27.80 g, 201.5 mmol, Aldrich). The resulting mixture was heated to 150 °C overnight. After about 22 hours, the reaction was cooled down to room temperature and the reaction mixture was poured into Dl water (1000 mL) and extracted with ethyl acetate (800 mL, Aldrich). The organic layer was separated and concentrated under vacuum on a rotary evaporator. The resulting residue was purified by column chromatography eluting with a gradient of hexanes and ethyl acetate to give a white powder as the desired product IC-1 (7.19 g, 69% yield). H-NMR (TCE ???): δ 3.8 (s, 12H), 4.4 (s, 8H), 6.9 (d, 8H), and 7.9 (d, 8H).

IC-1

[0085] Synthesis of Intermediate Compound #2 (IC-2): Into 50 mL of anhydrous tetrahydrofuran cooled in an ice bath at 0 °C, IC-1 (6.5 g, 9.7 mmol) was added. LiAIH₄ (1 M in diethyl ether, 58 mL, 58.2 mmol, Aldrich) cooled to 0 °C was added dropwise. Thenthe reaction solution was allowed to warm to room temperature and stirred for 4 hours. The reaction mixture was poured into chilled water (1000 mL) and neutralized with HCI (1 M, Aldrich). Then, the solution was extracted with ethyl acetate (800 mL, Aldrich), layers were separated, and the organic layer was concentrated. The crude product was purified by column chromatography eluting with a gradient of ethyl acetate and methanol to give a white powder as the desired product IC-2 (3.44 g, 63.5% yield). H-NMR (DMSO-d₆): δ 4.25 (s, 8H), 4.39 (s, 8H), 5.04 (s, 4H), 6.91 (d, 8H), 7.21 (d, 8H).

[0086] Synthesis of Crosslinker #3.2 (CLC-4.2): To tert-butanol (60 ml_, Aldrich) at room temperature, IC-2 was added with stirring followed by sodium tert- butoxide (566 mg, 5.89 mmol, Aldrich). The mixture was heated at 1 10 °C for 40 minutes. Then 1 ,4-butane sultone (0.60 ml_, 5.89 mmol, Aldrich), additional te/ -butanol (75 ml_, Aldrich) and N,N'-dimethylformamide (20 ml_, Aldrich) were added to the reaction mixture. After stirring for 24 hours, the product was precipitated out by adding hexanes (400 ml_, Aldrich). The resulting mixture was stirred for 15 minutes and then filtered. The collected solid was then added into hexanes (100 ml_, Aldrich). After stirring for another 15 minutes, the precipitate was collected by filteration again. The solid corrected was then added into a mixture of hexanes (100 ml_, Aldrich) and isopropanol (30 ml_, Aldrich). After stirring for 15 minutes, the precipitate was filtered and dried at 60 °C in a vacuum oven at 2 torr (Thermo Scientific Precision 6500, Thermo Fisher Scientific Waltham, MA USA) for 4 hours to give a white powder as the desired product CLC-4.2 (3.25 g, 76.8% yield). H-NMR (DMSO-d₆): δ 1.56 (m, 4H), 2.50 (m, 2H), 4.25-4.38 (m, 16H), 5.06 (s, 2H), 6.93 (d, 8H), and 7.20 (d, 8H).

Example 1.2.1 : Preparation of Anti-Microbial Element #1 (AM-1)

[0087] Crosslinked GO membrane (AM-1) Preparation: 4 mg/mL of a graphene oxide (GO) aqueous dispersion prepared as described in Example 1.1 was diluted to 0.1 wt% by de-ionized water. Second, a 0.1 wt% CLC-1.1 aqueous solution was created by dissolving appropriate amounts of CLC-1.1 in Dl water. Then, a coating mixture was created by mixing a mixture consisting of 0.1 wt% CLC-1.1 aqueous solution and a mixture consisting of 0.1 wt% graphene oxide aqueous dispersion at a weight ratio of 3: 1. The resulting solution was then was stirred at room temperature for 10 minutes. The resulting solution was cast onto a Reverse Osmosis (RO) membrane (ESPA Membrane, Hydranautics) by dropping the solution on membrane surface using a die caster (Taku-Die 200, Die-Gate Co., Ltd., Tokyo, Japan), set to coat 0.6 g of mixture per 90 cm². After drying in air, the membrane was put in an oven (DX400, Yamato Scientific Co., Ltd. Tokyo, Japan) at 85 °C for 30 minutes in order to remove water and crosslink the membrane, resulting in a membrane that was 1 .4 μηη thick with 1 :3 mass ratio GO/CLC-1.1 membrane, or AM-1.

Example 1.2.2: Preparation of Additional Anti-Microbial Elements (AM-2 thru AM-3)

[0088] For Example 1 .2.2, additional anti-microbial elements were constructed. The methods used were similar to the one in Example 1.2.1 with the exception that parameters were varied for the specific elements as identified in Table 1.

Table 1 : Anti-microbial Crosslinked GO Elements.

Comparative Example 1 .2.3: Preparation of Stock Substrate (CM-1 )

[0089] Comparative Element/Hydranautics Membrane (CM-1) Preparation: In another example (CM-1), a membrane was cut from stock reverse osmosis (RO) membrane from Hydranautics (ESPA Membrane, Hydranautics).

Example 2.1 : Measurement of Anti-Microbial Properties.

[0090] To test the membrane anti-microbial, example AM-1 was measured using a procedure that conformed to Japanese Industrial Standard (JIS) Z 2801 :2012 (English Version pub. Sep. 2012) for testing anti-microbial product efficacy, which is incorporated herein in its entirety. The organisms used in the verification of antimicrobial capabilities were escherichia coli. (ATCC® 8739, ATCC).

[0091] For the test, a broth was prepared by suspending 8 g of the nutrient powder (Difco™ Nutrient Broth, Becton, Dickinson and Company, Franklin Lakes, NJ USA) in 1 L of filtered, sterile water, mixing thoroughly and then heating with frequent agitation. To dissolve the powder the mixture was boiled for 1 minute and then autoclaved at 121 °C for 15 minutes. The night before testing, the escherichia coli. were added to 2-3 mL of the prepared broth and grown overnight.

[0092] On the day of the test, the resulting culture was diluted in fresh media and then let grow to a density of 10⁸ CFU/mL (or approximately diluting 1 mL of culture into 9 mL of fresh nutrient broth). The resulting solution was then left to re-grow for 2 hours. The re-growth was then diluted by 50 times in sterile saline (NaCI 8.5 g (Aldrich) in 1 L of distilled water) to achieve an expected density of about 2 x 10⁶ CFU/mL. 50 μί of the dilute provides the inoculation number.

[0093] The samples were then cut into 1 inch by 2 inch squares and placed in a petri dish with the GO-coated side up. Then 50 μί of the dilute was taken and the test specimens were inoculated. A transparent cover film (0.75 in. x 1.5 in., 3M, St. Paul, MN USA) was then used to help spread the bacterial inoculums, define the spread size, and reduce evaporation. Then, the petri dish was covered with a transparent lid, and left to grow.

[0094] When the desired measurement points of 2 hours and 24 hours were achieved, the test specimens and cover film were transferred with sterile forceps into 50 mL conical tubes with 20 mL of saline and the bacteria for each sample was washed off each sample by mixing them for at least 30 seconds in a vortex mixer (120V, VWR Arlington Heights, IL USA). The bacteria cells in each solution were then individually transferred using a pump (MXPPUMP01 , EMD Millipore, Billerica, MA USA) combined with a filter (Millflex-100, 100 mL, 0.45 μπι, white gridded, MXHAWG124, EMD Millipore) into individual cassettes prefilled with tryptic soy agar (MXSMCTS48, EMD Millipore).

[0095] Then, the cassettes were then invented and then placed in an incubator at 37 °C for 18 hours. After 18 hours, the number of colonies on the cassettes was counted. If there were no colonies a zero was recorded. For untreated pieces, after 24 hours the number of colonies was not less than 1 x 10³ colonies. The tests were run three times for each sample type to assure validity and repeatability of the data. Similar experiments were run for samples AM-2 thru AM-23, with the results shown in Table 2. Even assuming that the TNTC samples had counts equivalent to the approximate maximum count value of around 4000 colonies, the antibacterial activity can be estimated at 3.8. Thus, the antibacterial activity is at least 3.8, which supports an antibacterial activity of 2.0 or higher. As a result, it was determined that the crosslinked GO coatings disclosed herein are an effective biocide that could help prevent microbe buildup on surfaces.

Table 2: Antimicrobial Effect of Graphene Coatings.

Inoculation 1^st ₃rd

Material _t Activif y Comment

Control Count Count Coun

20 AM-5 GO+CLC-4.2/ 2.09 x 10^B 0 0 N/A 3.8 All Microbes

Glass Killed

21 CM-1 RO 1.31 x 10^J TNTC TNTC N A — Negligible

Membrane Microbes Killed

(Washed)

22 AM-4 GO+CLC-4.1/ 1.31 x 10^s TNTC TNTC N/A 0.0 Negligible

Glass Microbes Killed

23 AM-5 GO+CLC-4.2/ 1.31 x 10^B 0 0 N/A 3.8 All Microbes

Glass Killed

Note the term TNTC denotes too numerous to count.

Example 3.1 : Preparation and Testing of additional Anti-Microbial Elements

[0096] A number of antimicrobial elements were prepared in a manner similar to AM-1 using different crosslinkers in varying ratios and deposited in varying thicknesses. In some samples, additional crosslinkers or other materials were used to test the effect of those materials on the performance. The details of each antimicrobial element and the results obtained for each are shown in Table 3 below.

Table 3: Preparation and testing of antimicrobial coatings.

⁵ The silica nanoparticles in this sample have a size of 80 nm. The same is true for samples 7, 8, and 9.

^† DABA: 3,5-diaminobenzoic acid; KBO: potassium tetraborate

EMBODIMENTS

[0097] The authors of the present disclosure contemplate a number of specific embodiments including at least the following:

Embodiment 1. An anti-microbial membrane comprising:

a support; and

a composite coating the support comprising a crosslinked optionally substituted graphene oxide compound, where the graphene was crosslinked by a crosslinker selected form the group consisting of a benzoic acid derivative, an optionally substituted biphenyl of Formula 1 , an optionally substituted triphenylmethane of Formula 2, an optionally substituted diphenylamine or an optionally substituted 9H- carbazole represented by Formula 3A or 3B, and an optionally substituted bishydroxymethyl propanedi 4:

Formula 1 ;

where Ri and R₂ are independently NH₂ or OH; and R₃ and R₄ are independently OH, S0₃H, S0₃Na, or S0₃K;

Formula 2; wherein R₅ is H, CH₃, or C₂H₅; R₆ is H, CH₃, -C0₂H, -C0₂Li, -C0₂Na, - C0₂K, -S0₃H, -S0₃Li, -S0₃Na, or -S0₃K; and n is 0, 1 , 2, 3, 4, or 5;

Formula 3A Formula 3B wherein R₇ and R₈ are independently H, CH₃, C0₂H, C0₂Li, C0₂Na, C0₂K, SO₃H, SO₃L1, S0₃Na, or S0₃K; k is 0 or 1 ; m is 0, 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; and p is O, 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10;

Formula 4; wherein R₉ , Ri₀, Rn , and Ri₂, can be independently: wherein R13 is independently OH, NH₂, C0₂H, C0₂Na, C0₂K, SO3H , S0₃Na, or SO3K and r is 0, 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; whereby the membrane kills microbes as determined by having an antibacterial effectiveness of 2.0 or more.

Embodiment 2. The membrane of embodiment 1 , wherein the optionally substituted biphenyl is:

Embodiment 3. The membrane of embodiment 1 , wherein the optionally substituted triphenylmethane is:

Embodiment 4. The membrane of embodiment 1 , wherein the optionally substituted diphenylamine or optionally substituted 9H-carbazole is:

Embodiment 5. The membrane of embodiment 1 , wherein the optionally substituted bishydroxymethyl propanediol compound is:

Embodiment 6. The membrane of embodiment 1 , wherein the benzoic acid derivative is 3,5-diaminobenzoic acid.

Embodiment 7. The membrane of embodiment 1 or 6, wherein the composite further comprises at least one of potassium tetraborate and 2,5- dihydroxyterephthalic acid. Embodiment s. The membrane of embodiment 1 , 2, 3, 4, 5, 6, or 7, wherein the optionally substituted graphene oxide comprises platelets.

Embodiment 9. The membrane of embodiment 8, wherein the platelets are between about 0.05 μηη and about 50 μηη.

Embodiment 10. The membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, or 9, wherein the mass ratio of graphene oxide to crosslinker in the composite is a value ranging from 1 : 1000 to 50: 1.

Embodiment 11. The membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, or 9, wherein the mass ratio of graphene oxide to crosslinker in the composite is a value ranging from 1 :4 to 12: 1.

Embodiment 12. The membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, or 9, wherein the mass ratio of graphene oxide to crosslinker in the composite is a value ranging from 1 :4 to 1 : 1.

Embodiment 13. The membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, or 9, wherein the mass ratio of graphene oxide to crosslinker in the composite is a value ranging from 4: 1 to 1 1 : 1.

Embodiment 14. The membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, or 13, wherein the composite further comprises a spacer material.

Embodiment 15. The membrane of embodiment 14, wherein the spacer material comprises silica nanoparticles.

Embodiment 16. The membrane of embodiment 15, wherein the silica nanoparticles have a size of about 3 nm to about 20 nm.

Embodiment 17. The membrane of embodiment 15, wherein the silica nanoparticles have a size of about 50 nm to about 250 nm.

Embodiment 18. The membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, or 17, wherein the composite coating on the support has a thickness of about 0.9 μηη to about 3 μηη.

Embodiment 19. The membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, or 17, wherein the composite coating on the support has a thickness of about 10 nm to about 500 nm. Embodiment 20. The membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, or 17, wherein the composite coating on the support has a thickness of about 100 nm to about 300 nm.

Embodiment 21. The membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, or 20, wherein the membrane is prepared by applying the composite to the support and exposing the resulting membrane to a temperature of about 70 °C to about 200 °C for a period of about 2 minutes to about 60 minutes.

Embodiment 22. The membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, or 20, wherein the membrane is prepared by applying the composite to the support and exposing the resulting membrane to a temperature of about 80 °C to about 150 °C for a period of about 3 minutes to about 30 minutes.

Embodiment 23. The membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , or 22, wherein the support is the article to be protected from microbial growth.

Embodiment 24. A method of killing microbes, the method comprising:

providing the membrane of embodiment 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, or 23; and

exposing the membrane to a working fluid containing microbes;

wherein the membrane kills microbes as a result of exposure to the working fluid as determined by having an antibacterial effectiveness of 2.0 or more.

[0098] Unless otherwise indicated, all numbers expressing quantities of ingredients, properties such as molecular weight, reaction conditions, and so forth used in the specification and claims are to be understood as being modified in all instances by the term "about." Accordingly, unless indicated to the contrary, the numerical parameters set forth in the specification and attached claims are approximations that may vary depending upon the desired properties sought to be obtained. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. [0099] The terms "a," "an," "the" and similar referents used in the context of describing the invention (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. All methods described herein may be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., "such as") provided herein is intended merely to better illuminate the invention and does not pose a limitation on the scope of any claim. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the invention.

[00100] Groupings of alternative elements or embodiments disclosed herein are not to be construed as limitations. Each group member may be referred to and claimed individually or in any combination with other members of the group or other elements found herein. It is anticipated that one or more members of a group may be included in, or deleted from, a group for reasons of convenience and/or patentability.

[00101] Certain embodiments are described herein, including the best mode known to the inventors for carrying out the invention. Of course, variations on these described embodiments will become apparent to those of ordinary skill in the art upon reading the foregoing description. The inventor expects skilled artisans to employ such variations as appropriate, and the inventors intend for the invention to be practiced otherwise than specifically described herein. Accordingly, the claims include all modifications and equivalents of the subject matter recited in the claims as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is contemplated unless otherwise indicated herein or otherwise clearly contradicted by context.

[00102] In closing, it is to be understood that the embodiments disclosed herein are illustrative of the principles of the claims. Other modifications that may be employed are within the scope of the claims. Thus, by way of example, but not of limitation, alternative embodiments may be utilized in accordance with the teachings herein. Accordingly, the claims are not limited to embodiments precisely as shown and described.

Claims

Claims:

1. An anti-microbial membrane comprising:

a support; and

a composite coating the support comprising a crosslinked optionally substituted graphene oxide compound, where the graphene was crosslinked by a crosslinker selected form the group consisting of a benzoic acid derivative, an optionally substituted biphenyl of Formula 1 , an optionally substituted triphenylmethane of Formula 2, an optionally substituted diphenylamine or an optionally substituted 9H- carbazole represented by Formula 3A or 3B, and an optionally substituted bishydroxymethyl propanedi 4:

Formula 1 ;

where Ri and R₂ are independently NH₂ or OH; and R₃ and R₄ are independently OH, S0₃H, S0₃Na, or S0₃

Formula 2;

wherein R₅ is H, CH₃, or C₂H₅; R₆ is H, CH₃, -C0₂H, -C0₂Li, -C0₂Na, - C0₂K, - -S0₃Li, -S0₃Na, or -S0₃K; and n is 0, 1 , 2, 3, 4, or 5;

wherein R₇ and R₈ are independently H, CH₃, C0₂H, C0₂Li, C0₂Na, C0₂K, S0₃H, S0₃Li, S0₃Na, or S0₃K; k is 0 or 1 ; m is 0, 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; and p is O, 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10;

Formula 4;

wherein R₉ , Ri₀, Rn , and Ri₂, can be independently: wherein R₁₃ is independently OH, NH₂, C0₂H, C0₂Na, C0₂K, S0₃H, S0₃Na, or S0₃K and r is 0, 1 ,

2,

3, 4, 5, 6, 7, 8, 9, or 10; whereby the membrane kills microbes as determined by having an antibacterial effectiveness of 2.0 or more.

The membrane of claim 1 , wherein the optionally substituted biphenyl is:

The membrane of claim 1 , wherein the optionally substituted triphenylmethane is:

4. The membrane of claim 1 , wherein the optionally substituted diphenylamine or optionally substituted -carbazole is:

5. The membrane of claim 1 , wherein the optionally substituted bishydroxymethyl propanediol compound is:

6. The membrane of claim 1 , wherein the benzoic acid derivative is 3,5- diaminobenzoic acid.

7. The membrane of claim 1 or 6, wherein the composite further comprises at least one of potassium tetraborate and 2,5-dihydroxyterephthalic acid.

8. The membrane of claim 1 , 2, 3, 4, 5, 6, or 7, wherein the optionally substituted graphene oxide comprises platelets.

9. The membrane of claim 8, wherein the platelets are between about 0.05 pm and about 50 pm.

10. The membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, or 9, wherein the mass ratio of graphene oxide to crosslinker in the composite is a value ranging from 1 : 1000 to 50:1.

1 1. The membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, or 9, wherein the mass ratio of graphene oxide to crosslinker in the composite is a value ranging from 1 :4 to 12: 1.

12. The membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, or 9, wherein the mass ratio of graphene oxide to crosslinker in the composite is a value ranging from 1 :4 to 1 : 1.

13. The membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, or 9, wherein the mass ratio of graphene oxide to crosslinker in the composite is a value ranging from 4:1 to 1 1 : 1.

14. The membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, or 13, wherein the composite further comprises a spacer material.

15. The membrane of claim 14, wherein the spacer material comprises silica nanoparticles.

16. The membrane of claim15, wherein the silica nanoparticles have a size of about 3 nm to about 20 nm.

17. The membrane of claim 15, wherein the silica nanoparticles have a size of about 50 nm to about 250 nm.

18. The membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, or 17, wherein the composite coating on the support has a thickness of about 0.9 pm to about 3 pm.

19. The membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, or 17, wherein the composite coating on the support has a thickness of about 10 nm to about 500 nm.

20. The membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, or 17, wherein the composite coating on the support has a thickness of about 100 nm to about 300 nm.

21. The membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, or 20, wherein the membrane is prepared by applying the composite to the support and exposing the resulting membrane to a temperature of about 70 °C to about 200 °C for a period of about 2 minutes to about 60 minutes.

22. The membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, or 20, wherein the membrane is prepared by applying the composite to the support and exposing the resulting membrane to a temperature of about 80 °C to about 150 °C for a period of about 3 minutes to about 30 minutes.

23. The membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , or 22, wherein the support is the article to be protected from microbial growth.

24. A method of killing microbes, the method comprising:

providing the membrane of claim 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, or 23; and

exposing the membrane to a working fluid containing microbes;

wherein the membrane kills microbes as a result of exposure to the working fluid as determined by having an antibacterial effectiveness of 2.0 or more.