WO2024138043A2

WO2024138043A2 - Zeolitic materials for the inhibition and disruption of biofilms, deactivation of viruses, and inhibition of fungal growth

Info

Publication number: WO2024138043A2
Application number: PCT/US2023/085523
Authority: WO
Inventors: Prabir K. Dutta
Original assignee: Zeovation Inc.
Priority date: 2022-12-21
Filing date: 2023-12-21
Publication date: 2024-06-27
Also published as: WO2024138043A3

Abstract

Described herein are wound dressings comprising a substrate material having a patient contacting surface, and a population of zeolite nanoparticles and a quaternary ammonium compound disposed on the patient contacting surface. The zeolite nanoparticles can comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion¬ exchangeable sites within the zeolite nanoparticles. Also provided are methods of inhibiting or disrupting biofilms, methods of killing, inhibiting, and/or affecting a virus, and methods of killing, inhibiting, and/or affecting a fungus by contacting the biofilm, virus, or fungus with a composition comprising a population of zeolite nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier. The zeolite nanoparticles can comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles.

Description

Zeolitic Materials for the Inhibition and Disruption of Biofilms, Deactivation of Viruses, and Inhibition of Fungal Growth

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims benefit of priority of U.S. Provisional Application No. 63/434,339, filed December 21, 2022, which is incorporated herein by reference.

BACKGROUND

Silver can exhibit antimicrobial activity towards Gram positive and Gram negative bacteria, as well as antifungal and antiviral activity. Specifically, silver can exhibit activity against methicillin-sensitive and methicillin-resistant Staphylococcus aureus (S. aureus', MSSA and MRSA, respectively), Enterococcus faecalis (E. faecalis), Pseudomonas aeruginosa (P. aeruginosa), Escherichia coli (E. coli), Enterobacter cloacae, Proteus vulgaris, Acinetobacter baumannii, Vibrio cholera and Samonella typhi, Candida albicans. Silver can also exhibit antiviral activity against retroviridae Human Immunodeficiency Virus- 1 (HIV-1) by interaction with gpl20 surface glycoprotein and disrupting binding to CD4 receptors on host cells. Since Ag⁺ forms complexes with ligands containing S, N and O, it can interact with thiols, carboxylic acids, phosphates, amines present in these organisms and compete with the native binding metals in enzymes. On a macroscopic level, this is evident in electron microscopy studies of bacteria exposed to Ag⁺, where morphological changes including membrane separation from cell wall can be observed. At the biological level, inhibition of both phosphate uptake and exchange, as well as causing the efflux of succinate, glutamine and proline of E.coli bacteria can be observed. Silver ions also disrupt the proton gradients across membranes, cause cell death by massive proton leakage through the cell membrane. Ag⁺ may also induce the formation of reactive oxygen species, which further contribute antimicrobial effects.

Copper ions can also exhibit antimicrobial activity towards E. coli, S. aureus, Streptococcus group D, and Pseudomonas spp, Salmonella enterica, Campylobacter jejuni, E. faecalis, P. aeruginosa and Klebsiella pneumoniae. Copper can reduce the infectivity of bronchitis virus, poliovirus ribonucleic acid (RNA), viruses phi X174, T7, phi 6, Junin, and HSV and Human Immunodeficiency Virus Type 1 (HIV-1). Copper ions are believed to exert antimicrobial effects by binding to thiol and oxygen containing groups, displacing essential metals from protein active sites, as well as altering the conformational structure and damage of DNA, RNA and proteins, interfering with oxidative phosphorylation and osmotic balance, and/or creating ROS species and increase permeability barrier of the plasma membrane and leakage of amino acids and potassium ions. Copper chelates such as copper- 8-quinolinolate and its derivates have can also exhibit fungicidal activity against Aspergillus spp.

A challenge associated with the use of transition metals in biological environment, especially with silver, is the rapid precipitation upon interaction with biological media. Accordingly, improved compositions and methods that exploit the antimicrobial activity of these metals are needed.

SUMMARY

As described herein, antimicrobially active transition metals can be incorporated into nanoscopic zeolites, thereby protecting them from becoming inactive because of the environment as compared to free silver ions and silver nanoparticles and releasing the ions upon interaction with pathogens. As such, compositions can exhibit improved antimicrobial activity. For example, without wishing to be bound by theory, when used in wound healing applications (e.g., when provided on a wound dressing), the zeolite nanoparticles can protect the antimicrobial transition metal ions (e.g., from precipitation via interaction with proteins) within the wound and provide a lasting source of antimicrobial transition metal ions within the wound. Further, the zeolite nanoparticles because of their nanosize can facilitate deeper penetration of the antimicrobially active transition metals into the wound, improving antimicrobial effects.

Further, as illustrated herein, biofilms, viruses, and fungi often contain a protective layer/membrane through which antimicrobially active transition metals must pass to engage with the cell machinery. As described herein, by combining positively charged “quats” (quaternary ammonium compounds) with negatively charged zeolite with the antimicrobial transition metals, improved antimicrobial activity can be achieved. Without wishing to be bound by theory, the quat can interact with the protective layer/membrane, facilitating a pathway for the associated transition metal in the zeolite to penetrate and bind to functional groups, thereby disrupting pathogen activity. The combination of a quat with antimicrobial transition metal ions (e.g., antimicrobial metal nanoparticles, such as silver nanoparticles (AgNPs) or a platform for the delivery of antimicrobial transition metal ions (e.g., zeolite nanoparticles comprising an antimicrobially effective amount of antimicrobial metal ions, for example, retained at ion-exchangeable sites within the zeolite nanoparticles)) can thus provide for improved antimicrobial effects, including the ability to inhibit, prevent, and/or disrupt a biofilm.

By way of example, Figures 1A-1C shows a possible mechanism by which a nanozeolite containing antimicrobially active transition metal ions in combination with a quat can disrupt and penetrate through a lipid protective layer of microorganisms. The synergy of a quat and antimicrobial transition metal ions can lead to enhanced potency, meaning that these combinations can exhibit one or more of the following: increased potency (e.g., less active agent(s) are required to achieve antimicrobial effect), reduced kill times, increased disinfection, reduce cytotoxicity towards eukaryotic cells (e.g. human cells), or a combination thereof. More importantly, increased residual activity of the product can be realized if a nanozeolite host entraps the antimicrobially active transition metal ions, since the encapsulation enhances the active lifetime of the transition metal ions. In some embodiments, this association (positively charged quat + negatively charged zeolite nanoparticles comprising an antimicrobially effective amount of antimicrobial metal ions, for example, retained at ion-exchangeable sites within the zeolite nanoparticles) can disrupt already formed biofilms, envelope viruses, and fungi due to the penetration of the transition metal thorough the protective layer, which is disrupted by the quat. Moreover, this combination can achieve a longer acting effect since antimicrobial activity depends on the release of the stored transition metal ions (which can occur over an extended period of time).

Accordingly, provided herein are methods for inhibiting, preventing, or disrupting a biofilm that comprises contacting the biofilm (or a surface on which a biofilm may form) with a composition that comprises (1) a population of zeolite nanoparticles comprising an antimicrobially effective amount of antimicrobial metal ions; and (2) a quaternary ammonium compound (also referred to as a “quat”). The antimicrobial metal ions can comprise nanoparticles formed from an antimicrobial metal disposed on and/or within the zeolite nanoparticles. In certain embodiments, the antimicrobial metal ions can comprise antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles. The population of zeolite nanoparticles and the quaternary ammonium compound can be dissolved or dispersed in a carrier, such as an aqueous carrier (e.g., water or a water/alcohol solution), and this stable colloidal solution can be uniformly dispersed on soft and hard surfaces (e.g., bandages). Separate solutions of zeolite colloidal suspension and quat can also be dispersed on soft and hard surfaces (e.g., bandages).

Accordingly, provided herein are methods for inhibiting, preventing, or disrupting a biofilm that comprises contacting the biofilm (or a surface on which a biofilm may form) with a composition that comprises (1) a population of zeolite nanoparticles comprising an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles; and (2) a quaternary ammonium compound (also referred to as a “quat”). The population of zeolite nanoparticles and the quaternary ammonium compound can be dissolved or dispersed in a carrier, such as an aqueous carrier (e.g., water or a water/alcohol solution), as well as dispersed on soft and hard surfaces (e.g., bandages).

Accordingly, also provided herein are wound dressings that comprise a substrate material having a patient contacting surface, and a population of zeolite nanoparticles and a quaternary ammonium compound disposed on the patient contacting surface. The zeolite nanoparticles can comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles.

DESCRIPTION OF DRAWINGS

Figures 1 A-1C illustrate a possible mechanism by which the zeolite quat association can disrupt and penetrate through a lipid protective layer of organisms.

Figure 2 illustrates the mechanisms by which a wound becomes non-healing due to the formation of a protective layer, known as extracellular polymeric substances (EPS) that surrounds the bacteria and prevents drugs from reaching the bacteria.

Figure 3 is a plot showing biofilm inhibition by nanozeolites including both silver ions and zinc ions retained at ion-exchangeable sites within the zeolite nanoparticles (“AM- 30”).

Figure 4 is a plot comparing the antimicrobial activity of free silver ions and silver ions protected within zeolite nanoparticles at the same concentration of silver (2.5 ppm Ag) to kill planktonic cells of Pseudomonas aeruginosa upon a two-hour exposure.

Figure 5 shows an example of a Pseudomonas aeruginosa biofilm grown on a cellulose membrane.

Figure 6 is a plot showing the relative anti-biofilm activity of a variety of extracellular membrane-based wound dressings against a Pseudomonas aeruginosa biofilm grown on a cellulose membrane.

Figure 7 illustrates the structure of envelope and non-enveloped viruses.

Figure 8 is a plot comparing the antiviral activity of a quat, nanozeolites including antimicrobial transition metal ions retained at ion-exchangeable sites within the zeolite nanoparticles, and combinations of a quat and nanozeolites including antimicrobial transition metal ions retained at ion-exchangeable sites within the zeolite nanoparticles.

Figure 9 illustrates the structure of fungi.

Figure 10 is a photograph showing fungal growth on a control textile. Figure 11 is a photograph showing fungal growth on a textile containing a quat.

Figure 12 is a photograph showing fungal growth on a textile containing a quat in combination with nanozeolites including both silver ions and zinc ions retained at ionexchangeable sites within the zeolite nanoparticles (AM 30, 100 pg/cm²). DETAILED DESCRIPTION

Provided herein are wound dressings that comprise a substrate material having a patient contacting surface, and a population of zeolite nanoparticles and quaternary ammonium compound disposed on the patient contacting surface. The zeolite nanoparticles can comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion- exchangeable sites within the zeolite nanoparticles. In some embodiments, the population of zeolite nanoparticles and the quaternary ammonium compound are Coulombically associated with one another.

The substrate material can comprise any suitable substrate used in conventional wound dressings. For example, in some embodiments, the substrate material can comprise a dry or impregnated gauze, a film, a gel, a foam, a hydrocolloid, a woven or non-woven fabric, a hydrogel, a paste, granules, beads, or a combination thereof. In some embodiments, the substrate material can comprise collagen, cellulose, carboxymethylcellulose, oxidized regenerated cellulose (ORC), an alginate, extracellular matrix, a polyester, a polyurethane, or combinations thereof. In certain embodiments, the substrate material can comprise extracellular matrix, (or a protein component thereof).

In some embodiments, the quaternary ammonium compound can be deposited on the patient contacting surface at a concentration of from 10-500 pg/cm².

In some embodiments, the zeolite nanoparticles can be deposited on the patient contacting surface at a concentration of from 10-1200 pg/cm².

In some embodiments, the zeolite nanoparticles can be deposited on the patient contacting surface such that the antimicrobial metal ions are present on the patient contacting surface at a concentration of from 10-500 pg/cm². In some embodiments, the quaternary ammonium compound can be defined by the Formula I or Formula II below

Formula I Formula II wherein

R¹ is selected from C1-20 alkyl, C2-20 alkenyl, C2-20 alkynyl, Ci-2ohaloalkyl, C3-10 cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 4-20 membered heterocycloalkyl, C3.10cycloalkyl-C1.10 alkylene, 4-10 membered heterocycloalkyl-Ci-10 alkylene, 6-10 membered aryl-Ci-10 alkylene, and 5-10 membered heteroaryl-Ci-10 alkylene, each optionally substituted with 1, 2, 3, or 4 independently selected R^x groups;

R' is, individually for each occurrence, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-4 haloalkyl, C3-10 cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 4-10 membered heterocycloalkyl, C3 10 cycloalkyl-C 1-4 alkylene, 4-10 membered heterocycloalkyl-Ci-4 alkylene, 6-10 membered aryl-Ci-4 alkylene, and 5-10 membered heteroaryl-Ci-4 alkylene, each optionally substituted with 1, 2, 3, or 4 independently selected R^x groups; and each R^x, when present, is independently selected from OH, NO2, CN, halo, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, CM haloalkyl, C1-6 alkoxy, C 1-6 haloalkoxy, cyano-Ci-3 alkyl, HO- C1-3 alkyl, amino, C 1-6 alkylamino, di(C 1-6 alky l)amino, thio, C 1-6 alkylthio, C 1-6 alkylsulfinyl, C1-6 alkylsulfonyl, carbamyl, C 1-6 alkylcarbamyl, di(Ci-6 alkyl)carbamyl, carboxy, C1-6 alkylcarbonyl, C 1-6 alkoxycarbonyl, C 1-6 alkylcarbonylamino, C 1-6 alkylsulfonylamino, aminosulfonyl, C 1-6 alkylaminosulfonyl, di(C 1-6 alky l)aminosulfonyl, aminosulfonylamino, C1-6 alkylaminosulfonylamino, di(Ci-6 alkyl)aminosulfonylamino, aminocarbonylamino, Ci-6 alkylaminocarbonylamino, and di(Ci-6 alkyljaminocarbonylamino.

In some embodiments, the antimicrobial metal ions can comprise silver, zinc, copper, or any combination thereof.

In some embodiments, the antimicrobial metal ions can be present in an amount of from 1% up to 50% of the ion exchange capacity of the zeolite nanoparticles, such as from 1% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 1% up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 1% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 25% of the ion exchange capacity of the zeolite nanoparticles, or from 10% up to 20% of the ion exchange capacity of the zeolite nanoparticles.

In some embodiments, the antimicrobial metal ions can comprise silver and zinc

In some embodiments, the antimicrobial metal ions can comprise silver and copper.

In certain embodiments, the zeolite nanoparticles can be deposited on the patient contacting surface such that silver ions are present on the patient contacting surface at a concentration of from 1-100 pg/cm², such as from 5-50 pg/cm², from 10-40 pg/cm², from 20- 40 pg/cm², or from 25-35 pg/cm².

In certain embodiments, the zeolite nanoparticles can be deposited on the patient contacting surface such that zinc ions are present on the patient contacting surface at a concentration of from 0.25-25 pg/cm², such as from 1.25-12.5 pg/cm², from 2.5-10 pg/cm², from 5-10 pg/cm², or from 6.25-8.75 pg/cm².

In certain embodiments, the antimicrobial metal ions can comprise silver and zinc, and the silver and zinc can be present on the patient contacting surface at a weight ratio of silverzinc of from 1: 1 to 10: 1, such as from 2:1 to 8:1, or from 3: 1 to 5:1.

In certain embodiments, the antimicrobial metal ions comprise silver and zinc, and the silver and zinc can be present on the patient contacting surface at a weight ratio of silver:zinc of about 4: 1.

Also provided herein are methods of inhibiting or disrupting a biofilm. These methods can comprise contacting the biofilm with a composition comprising a population of zeolite nanoparticles and a quaternary ammonium compound dissolved or dispersed within or on the surface of a carrier. The zeolite nanoparticles can comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles. In some embodiments, these methods can comprise disrupting an existing biofilm.

Also provided herein are methods of preventing or inhibiting the formation of a biofilm on a surface. These methods can comprise contacting the surface with a composition comprising a population of zeolite nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier. The zeolite nanoparticles can comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles. In some embodiments, the surface can comprise a fomite surface. In some embodiments, the surface can comprise a surface of a medical device or a wound dressing.

In some of these embodiments, the population of zeolite nanoparticles and the quaternary ammonium compound are Coulombically associated with one another.

In some of these embodiments, the quaternary ammonium compound can be deposited on the surface at a concentration of from 10-500 pg/cm².

In some of these embodiments, the zeolite nanoparticles can be deposited on the surface at a concentration of from 10-1200 pg/cm².

In some of these embodiments, the zeolite nanoparticles can be deposited on the surface such that the antimicrobial metal ions are present on the surface of the carrier at a concentration of from 10-500 pg/cm².

In some of these embodiments, the zeolite nanoparticles can be deposited on the surface such that silver ions are present on the surface at a concentration of from 1-100 pg/cm², such as from 5-50 pg/cm², from 10-40 pg/cm², from 20-40 pg/cm², or from 25-35 pg/cm².

In some of these embodiments, the zeolite nanoparticles can be deposited on the surface such that zinc ions are present on the surface at a concentration of from 0.25-25 pg/cm², such as from 1.25-12.5 pg/cm², from 2.5-10 pg/cm², from 5-10 pg/cm², or from 6.25-8.75 pg/cm².

In some of these embodiments, the antimicrobial metal ions comprise silver and zinc, and the silver and zinc can be present on the surface at a weight ratio of silver: zinc of from 1:1 to 10:1, such as from 2:1 to 8:1, or from 3:1 to 5:1.

In some of these embodiments, the antimicrobial metal ions comprise silver and zinc, and the silver and zinc are present on the surface at a weight ratio of silver:zinc of about 4: 1.

Also provided herein are method of killing, inhibiting, and/or affecting a virus. These methods can comprise contacting the virus with a composition comprising a population of zeolite nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier. The zeolite nanoparticles can comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles.

In some embodiments, the virus can comprise an enveloped virus. In certain embodiments, the enveloped virus can comprise a DNA virus, such as a virus in the family Herpesviridae, Poxviridae, Hepadnaviridae, or Asfarviridae. In certain embodiments, the enveloped virus can comprise an RNA virus, such as a flavivirus, an alphavirus, a togavirus, a coronavirus, a hepatitis virus, an orthomyxovirus, a paramyxovirus, a rhabdovirus, a bunyavirus, or a filovirus.

Also provided herein are methods of killing, inhibiting, and/or affecting a fungus. These methods can comprise contacting the fungus with a composition comprising a population of zeolite nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier. The zeolite nanoparticles can comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles.

In some of these embodiments, the quaternary ammonium compound can be defined by the Formula I or Formula II below

R'

R¹— N-R'

R¹— NH₃ '

Formula I Formula II wherein

R¹ is selected from C1-20 alkyl, C2-20 alkenyl, C2-20 alkynyl, Ci-2ohaloalkyl, C3-10 cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 4-20 membered heterocycloalkyl, Cs-iocycloalkyl-Ci-io alkylene, 4-10 membered heterocycloalkyl-Ci-10 alkylene, 6-10 membered aryl-Ci-10 alkylene, and 5-10 membered heteroaryl-Ci-10 alkylene, each optionally substituted with 1, 2, 3, or 4 independently selected R^x groups;

R' is, individually for each occurrence, Ci-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-4 haloalkyl, C3-10 cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 4-10 membered heterocycloalkyl, C3-iocycloalkyl-C 1-4 alkylene, 4-10 membered heterocycloalkyl-Ci-4 alkylene, 6-10 membered aryl-Ci-4 alkylene, and 5-10 membered heteroaryl-Ci-4 alkylene, each optionally substituted with 1, 2, 3, or 4 independently selected R^x groups; and each R^x, when present, is independently selected from OH, NO2, CN, halo, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-4 haloalkyl, C1-6 alkoxy, C 1-6 haloalkoxy, cyano-Ci-3 alkyl, HO- C1-3 alkyl, amino, C 1-6 alkylamino, di(C 1-6 alky l)amino, thio, C 1-6 alkylthio, C 1-6 alkylsulfinyl, Ci .6 alkylsulfonyl, carbamyl, C1-6 alkylcarbamyl, di(Ci-6 alkyl)carbamyl, carboxy, C1-6 alkylcarbonyl, C 1-6 alkoxycarbonyl, C 1-6 alkylcarbonylamino, C 1-6 alkylsulfonylamino, aminosulfonyl, C 1-6 alkylaminosulfonyl, di(C 1-6 alky l)aminosulfonyl, aminosulfonylamino, Ci-6 alkylaminosulfonylamino, di(Ci-6 alkyl)aminosulfonylamino, aminocarbonylamino, Ci-6 alkylaminocarbonylamino, and di(Ci-6 alkyl)aminocarbonylamino.

In some of these embodiments, the quaternary ammonium compound can be present in the composition in an amount of from 10 ppm to 500 ppm.

In some of these embodiments, the antimicrobial metal ions can comprise silver, zinc, copper, or any combination thereof.

In some of these embodiments, the antimicrobial metal ions can be present in an amount of from 1 % up to 50% of the ion exchange capacity of the zeolite nanoparticles, such as from 1% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 1% up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 1% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 25% of the ion exchange capacity of the zeolite nanoparticles, or from 10% up to 20% of the ion exchange capacity of the zeolite nanoparticles.

In some of these embodiments, the antimicrobial metal ions can comprise silver and zinc.

In some of these embodiments, the antimicrobial metal ions can comprise silver and copper.

In some of these embodiments, the zeolite nanoparticles are present in the composition in an amount of from 10 ppm to 10,000 ppm.

Also provided herein are methods of inhibiting or disrupting a biofilm. These methods can comprise contacting the biofilm with a composition comprising an antimicrobially effective amount of antimicrobial metal ions from a metal salt or metal nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier. In some embodiments, the method can comprise disrupting an existing biofilm.

Also provided herein are methods of preventing or inhibiting the formation of a biofilm on a surface, the method comprising contacting the surface with a composition comprising an antimicrobially effective amount of antimicrobial metal ions from a metal salt or metal nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier. In some embodiments, the surface can comprise a fomite surface. In some embodiments, the surface can comprise a surface of a medical device or a wound dressing.

In some embodiments, the quaternary ammonium compound can be deposited on the surface at a concentration of from 10-600 qg/cm².

In some embodiments, the antimicrobial metal ions can be deposited on the surface at a concentration of from 10-1200 qg/cnr.

In some embodiments, the antimicrobial metal ions can comprise silver ions, and the silver ions can be deposited on the surface at a concentration of from 1-100 pg/cnr, such as from 5-50 qg/cm², from 10-40 pg/cm², from 20-40 pg/cm², or from 25-35 pg/cm².

In some embodiments, the antimicrobial metal ions can comprise zinc ions, and the zinc ions can be deposited on the surface at a concentration of from 0.25-25 pg/cm², such as from 1.25-12.5 pg/cm², from 2.5-10 pg/cm², from 5-10 pg/cm², or from 6.25-8.75 pg/cm².

In some embodiments, the antimicrobial metal ions can comprise silver and zinc, and the silver and zinc can be present on the surface at a weight ratio of silverzinc of from 1:1 to 10:1, such as from 2: 1 to 8: 1 , or from 3:1 to 5 : 1.

In some embodiments, the antimicrobial metal ions can comprise silver and zinc, and the silver and zinc can be present on the surface at a weight ratio of silverzinc of about 4:1.

Also provided herein are methods of killing, inhibiting, and/or affecting a virus. These methods can comprise contacting the virus with a composition comprising an antimicrobially effective amount of antimicrobial metal ions from a metal salt or metal nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier.

Also provided herein are methods of killing, inhibiting, and/or affecting a fungus, the method comprising contacting the fungus with a composition comprising an antimicrobially effective amount of antimicrobial metal ions from a metal salt or metal nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier.

In some of these embodiments, the quaternary ammonium compound can be defined by the Formula I or Formula II below R'

R¹— N-R' 1— NH₃ '

Formula I Formula II wherein

R¹ is selected from Ci-20 alkyl, C2-20 alkenyl, C2-20 alkynyl, Ci-2ohaloalkyl, C3-10 cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 4-20 membered heterocycloalkyl, Cs-iocycloalkyl-Ci-io alkylene, 4-10 membered heterocycloalkyl-Ci-10 alkylene, 6-10 membered aryl-Ci-10 alkylene, and 5-10 membered heteroaryl-Ci-10 alkylene, each optionally substituted with 1, 2, 3, or 4 independently selected R^x groups;

R' is, individually for each occurrence, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-4 haloalkyl, C3-10 cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 4-10 membered heterocycloalkyl, C3-iocycloalkyl-Ci-4 alkylene, 4-10 membered heterocycloalkyl-Ci-4 alkylene, 6-10 membered aryl-Ci-4 alkylene, and 5-10 membered heteroaryl-Ci-4 alkylene, each optionally substituted with 1, 2, 3, or 4 independently selected R^x groups; and each R^x, when present, is independently selected from OH, NO2, CN, halo, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, CM haloalkyl, C1-6 alkoxy, C 1-6 haloalkoxy, cyano-Ci-3 alkyl, HO- C1-3 alkyl, amino, C 1-6 alkylamino, di(Ci-6 alkyl)amino, thio, C 1-6 alkylthio, C 1-6 alkylsulfinyl, C1-6 alkylsulfonyl, carbamyl, C1-6 alkylcarbamyl, di(Ci-6 alkyl)carbamyl, carboxy, CM alkylcarbonyl, C 1-6 alkoxycarbonyl, C 1-6 alkylcarbonylamino, C 1-6 alkylsulfonylamino, aminosulfonyl, C 1-6 alkylaminosulfonyl, di(C 1-6 alky l)aminosulfonyl, aminosulfonylamino, C1-6 alkylaminosulfonylamino, di(C 1-6 alky l)aminosulfonylamino, aminocarbonylamino, C1-6 alkylaminocarbonylamino, and di(Ci-6 alkyl)aminocarbonylamino.

In some of these embodiments, the antimicrobial metal ions are present in the composition in an amount of from 10 ppm to 10,000 ppm. In some of these embodiments, the antimicrobial metal ions can comprise antimicrobial metal nanoparticles.

In other of these embodiments, the antimicrobial metal ions can comprise antimicrobial metal nanoparticles disposed on or within zeolite nanoparticles. In some embodiments, the antimicrobial metal ions can be retained at ion-exchangeable sites within zeolite nanoparticles. In some embodiments, the antimicrobial metal ions can be present in an amount of from 1% up to 50% of the ion exchange capacity of the zeolite nanoparticles, such as from 1 % up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 1 % up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 1% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 25% of the ion exchange capacity of the zeolite nanoparticles, or from 10% up to 20% of the ion exchange capacity of the zeolite nanoparticles. In some of these embodiments, the zeolite nanoparticles are present in the composition in an amount of from 10 ppm to 10,000 ppm.

Zeolite Nanoparticles

As described above, the compositions and methods described herein can comprise zeolite nanoparticles. The zeolite nanoparticles are generally aluminosilicate having a three- dimensionally grown skeleton structure and is generally shown by xMz/nO' AhCh- ySiCh- zFhO, wherein M represents an ion-exchangeable metal ion; n corresponds to the valence of the metal; x is a coefficient of the metal oxide; y is a coefficient of silica; and z is the number of water of crystallization. The zeolite nanoparticles can have varying frameworks and differing Si/ Al ratios. In some embodiments, the zeolite nanoparticles can comprise zeolite having a faujasite structure. For example, the zeolite nanoparticles can be zeolite X or Y.

The zeolite nanoparticles can have an average particle size of less than 250 nm (e.g., less than 200 nm, less than 150 nm, less than 100 nm, less than 90 nm, less than 80 nm, less than 70 nm, less than 60 nm, less than 50 nm, less than 40 nm, less than 30 nm, or less than 20 nm). In some embodiments, the zeolite nanoparticles can have an average particle size of at least 10 nm (e.g., at least 20 nm, at least 30 nm, at least 40 nm, at least 50 nm, at least 60 nm, at least 70 nm, at least 80 nm, at least 90 nm, at least 100 nm, at least 150 nm, or at least 200 nm).

The zeolite nanoparticles can have an average particle size ranging from any of the minimum values described above to any of the maximum values described above. For example, in some embodiments, the zeolite nanoparticles can have an average particle size of from 10 to 250 nm (e.g., from 10 to 200 nm, from 10 to 150 nm, from 10 to 100 nm, from 20 to 80 nm, or from 20 to 60 nm). In certain embodiments, the zeolite nanoparticles can have an average particle size of from 20 nm to 40 nm, such as from 25 nm to 35 nm. In certain embodiments, the zeolite nanoparticles can have an average particle size of 31 nm ± 2 nm.

The term “average particle size,” as used herein, generally refers to the statistical mean particle size (diameter) of the particles in a population of particles. The diameter of an essentially spherical particle may refer to the physical or hydrodynamic diameter. The diameter of a non-spherical particle may refer preferentially to the hydrodynamic diameter. As used herein, the diameter of a non-spherical particle may refer to the largest linear distance between two points on the surface of the particle. Mean particle size can be measured using methods known in the art, such as dynamic light scattering or electron microscopy.

In some embodiments, the population of zeolite nanoparticles can comprise a population of zeolite nanoparticle having a monodisperse particle size distribution. The term “monodisperse,” as used herein, describes a population of nanoparticles where all of the nanoparticles are the same or nearly the same size. As used herein, a monodisperse particle size distribution refers to particle distributions in which 80% of the distribution (e.g., 85% of the distribution, 90% of the distribution, or 95% of the distribution) lies within 20% of the median particle size (e.g., within 15% of the median particle size, within 10% of the median particle size, or within 5% of the median particle size).

The zeolite nanoparticles can possess a very regular pore structure of molecular dimensions. In some cases, the zeolite nanoparticles can exhibit a monodisperse pore size distribution. As used herein, a monodisperse pore size distribution refers to pore size distributions in which 80% of the distribution (e.g., 85% of the distribution, 90% of the distribution, or 95% of the distribution) lies within 20% of the median pore size (e.g., within 15% of the median pore size, within 10% of the median pore size, or within 5% of the median pore size).

In certain embodiments, the zeolite nanoparticles can exhibit an external pore size of 75 angstroms or less (e.g., 70 angstroms or less, 65 angstroms or less, 60 angstroms or less, 55 angstroms or less, 50 angstroms or less, 45 angstroms or less, 40 angstroms or less, 35 angstroms or less, 30 angstroms or less, 25 angstroms or less, 20 angstroms or less, or 15 angstroms or less). In certain embodiments, the zeolite nanoparticles can exhibit an external pore size of at least 10 angstroms (e.g., at least 15 angstroms, at least 20 angstroms, at least 25 angstroms, at least 30 angstroms, at least 35 angstroms, at least 40 angstroms, at least 45 angstroms, at least 50 angstroms, at least 55 angstroms, at least 60 angstroms, at least 65 angstroms, or at least 70 angstroms).

The zeolite nanoparticles can exhibit an external pore size of from any of the minimum values described above to any of the maximum values described above. For example, in some embodiments, the zeolite nanoparticles can exhibit an external pore size of from 10 to 75 angstroms (e.g., from 10 to 50 angstroms). In certain embodiments, the zeolite nanoparticles can exhibit an internal pore size of 8 angstroms or less (e.g., an internal pore size of from 2 to 8 angstroms).

The zeolite nanoparticles can also possess a high internal surface area. For example, in some embodiments, the zeolite nanoparticles can exhibit an average internal surface area of from 100 to 1,000 m²/g (e.g., from 200 to 1,000 m²/g, from 100 to 800 m²/g, from 200 to 800 m²/g, from 300 to 800 m²/g, from 300 to 700 m²/g, from 100 to 500 m²/g, from 200 to 500 m²/g, or from 400 to 800 m²/g).

The ion-exchange capacities of the zeolite nanoparticles may depend on the silica/aluminum ratio in their formulation. Zeolite types with low silica/aluminum ratios generally exhibit high ion-exchange capacities. In some embodiments, the SiCF/AhCh mole ratio in the zeolite nanoparticles is 14 or less (e.g., 13 or less, 12 or less, 11 or less, 10 or less, 9 or less, 8 or less, 7 or less, 6 or less, or 5 or less). In some embodiments, the zeolite nanoparticles can retain a metal ion in an amount as large as or less than an ion-exchange saturation capacity of the zeolite nanoparticles.

Antimicrobial Metal Ions

As described above, in some embodiments, the zeolite nanoparticles can comprise an effective amount of antimicrobial metal ions to kill or inhibit the growth of a microbe. Suitable antimicrobial metal ions are known in the art, and include silver, copper, zinc, and combinations thereof.

In some embodiments, the antimicrobial metal ions can comprise nanoparticles formed from an antimicrobial metal (e.g., Cu, Zn, Ag, or a combination thereof).

The nanoparticles can have an average particle size of 15 nm or less (e.g., 10 nm or less, 9 nm or less, 8 nm or less, 7 nm or less, 6 nm or less, 5 nm or less, 4 nm or less, 3 nm or less, 2 nm or less, or even 1 nm). In certain embodiments, the nanoparticles can have an average particle size of at least 1 nm (e.g., at least 2 nm, at least 3 nm, at least 4 nm, at least 5 nm, at least 6 nm, at least 7 nm, at least 8 nm, at least 9 nm, or up to 10 nm).

The nanoparticles can have an average particle size ranging from any of the minimum values described above to any of the maximum values described above. For example, in some embodiments, the nanoparticles can have an average particle size of from 1 to 10 nm (e.g., from 1 to 8 nm, or from 1 to 5 nm).

The nanoparticles can be present in an amount of at least 1 % by weight (e.g., at least 5% by weight, at least 10% by weight, at least 15% by weight, at least 20% by weight, or at least 25% by weight), based on the total weight of the zeolite nanoparticles and metal nanoparticles. In certain embodiments, the nanoparticles can be present in an amount of 25% by weight or less (e.g., 22% by weight or less, 20% by weight or less, 15% by weight or less, 10% by weight or less, or 5% by weight or less), based on the total weight of the zeolite nanoparticles and metal nanoparticles.

The nanoparticles can be present in an amount ranging from any of the minimum values described above to any of the maximum values described above. For example, in some embodiments, the nanoparticles can be present in an amount from 1% to 25% by weight (e.g., from 5% to 20% by weight, from 5% to 25% by weight, from 10% to 20% by weight, or from 15% to 25% by weight), based on the total weight of the zeolite nanoparticles and metal nanoparticles.

In some embodiments, the antimicrobial metal ions comprise antimicrobial metal ions (copper ions, zinc ions, silver ions, or a combination thereof) retained at ion-exchangeable sites within the zeolite nanoparticles. That is, the ion-exchangeable ions such as sodium ions, calcium ions, potassium ions, magnesium ions and/or iron ions in the zeolite nanoparticles can be partially or wholly replaced with the antimicrobial metal ions.

The antimicrobial metal ions can be present in an amount as large as or less than the ion-exchange saturation capacity of the zeolite nanoparticles. In some embodiments, the zeolite nanoparticles retain antimicrobial metal ions in an amount of 10% or greater, 15% or greater, 20% or greater, 25% or greater, 30% or greater, 40% or greater, 50% or greater, 60% or greater, 75% or greater, 80% or greater, 90% or greater, 95% or greater, or up to 100%, of the ion exchange capacity of the zeolite nanoparticles. In some embodiments, the zeolite nanoparticles can retain the antimicrobial metal ions in an amount of 100% or less, 95% or less, 90% or less, 85% or less, 75% or less, 50% or less, 40% or less, or 25% or less, of the ion exchange capacity of the zeolite nanoparticles. The antimicrobial metal ions can be present in an amount ranging from any of the minimum values described above to any of the maximum values described above. For example, in some embodiments, the antimicrobial metal ions can be retained in an amount from 10% up to 100% by weight (e.g., from 20% up to 100%, from 30% up to 100%, from 40% up to 100%, or from 50% up to 100%), of the ion exchange capacity of the zeolite nanoparticles.

In some embodiments, the zeolite nanoparticles can further comprise an additional bioactive metal ion (e.g., calcium ions) retained at ion-exchangeable sites within the zeolite nanoparticles. That is, the ion-exchangeable ions such as sodium ions, calcium ions, potassium ions, magnesium ions and/or iron ions in the zeolite nanoparticles can be partially or wholly replaced with the calcium ions. The calcium can induce clotting in the case of hemostatic compositions.

The additional bioactive metal ions can be present in an amount as large as or less than the ion-exchange saturation capacity of the zeolite nanoparticles. In some embodiments, the zeolite nanoparticles retain antimicrobial metal ions in an amount of 10% or greater, 15% or greater, 20% or greater, 25% or greater, 30% or greater, 40% or greater, 50% or greater, 60% or greater, 75% or greater, 80% or greater, 90% or greater, 95% or greater, or up to 100%, of the ion exchange capacity of the zeolite nanoparticles. In some embodiments, the zeolite nanoparticles can retain the antimicrobial metal ions in an amount of 100% or less, 95% or less, 90% or less, 85% or less, 75% or less, 50% or less, 40% or less, or 25% or less, of the ion exchange capacity of the zeolite nanoparticles.

The antimicrobial metal ions can be present in an amount ranging from any of the minimum values described above to any of the maximum values described above. For example, in some embodiments, the antimicrobial metal ions can be retained in an amount from 10% up to 100% by weight (e.g., from 20% up to 100%, from 30% up to 100%, from 40% up to 100%, or from 50% up to 100%), of the ion exchange capacity of the zeolite nanoparticles.

Quaternary Ammonium Compounds (Quats)

As described above, the compositions and methods described herein can comprise a quaternary ammonium compound. Quaternary ammonium compounds (quats) are an effective class of biocides. Quats are considered contact kill agents. The long hydrophobic chains of quats can penetrate the polysaccharide-based peptidoglycan layer into the phospholipid cytoplasmic membrane of bacteria, and causse cell death. In some embodiments, the quaternary ammonium compound can be defined by the Formula I or Formula II below

Formula I Formula II wherein

In some embodiments, the quaternary ammonium compound is present in the composition in an amount of from 10 ppm to 500 ppm.

Wound Dressings and Substrate Materials

As described above, the compositions and methods described herein can comprise wound dressings. Wounds in particular that can get infected are of the greatest interest. Both acute and chronic wounds are of interest. Examples of wounds of particular interest include, but are not limited to, bums, diabetic ulcers (e.g., diabetic foot ulcers), pressure ulcers (e.g., bed sores), and surgical incisions (e.g., surgical site infections). As discussed above, these wound dressings can comprise a substrate material having a patient contacting surface, and a population of zeolite nanoparticles and a quaternary ammonium compound disposed on the patient contacting surface.

The wound dressing can comprise any suitable wound dressing known in the art (formed from any suitable substrate material). Conventional dressings include, for example, absorbent pads, absorbent cotton, gauze (e.g., gauze bandages and/or gauze pads), wrap (e.g., elastic wraps and/or gauze wraps), dermal patches, surgical drapes, bandage, tapes, cotton- tipped stick, adhesive bandages, or other support wrap or medical bandage or wound cover.

The term “dressing” as used herein is also intended to cover casts (e.g., orthopedic cast, body cast, plaster cast, or surgical cast), frequently made from plaster, encasing a limb (or, in some cases, large portions of the body) to stabilize and hold anatomical structures, most often a broken bone (or bones), in place until healing is confirmed. In some embodiments, the dressing is a sterile dressing. The dressing may be disposable, washable and/or reusable.

In some embodiments, the dressing is an adhesive bandage. An adhesive bandage is usually covered by a woven fabric, plastic, or latex strip which has an adhesive. Adhesive bandages usually have an absorbent pad, which is sometimes medicated with an antiseptic solution. Some bandages have a thin, porous-polymer coating over the pad to keep it from sticking to the wound. The bandage is applied such that the pad covers the wound, and the fabric or plastic sticks to the surrounding skin to hold the dressing in place and prevent dirt from entering the wound. Adhesive bandages may include, but are not limited to, strip bandages, winged bandages, fingertip bandages, butterfly bandages, knuckle bandages, triangular bandages, tube bandages, compression bandages, elastic bandages, gauze bandages, donut bandages, pressure bandages, sterile-strips, eye bandages, sterile bum sheets, and adhesive tape.

In some embodiments, the substrate material comprises an absorbent or water- permeable material. The absorbent or water-permeable material is capable of absorbing wound exudate. The dressing can be composed of a substrate material comprising woven materials, non-woven materials or both. In some embodiments, the dressing comprises a fabric, cloth or sponge material. The dressing may be composed of natural and synthetic materials (e.g., natural or synthetic fibers). For example, the substrate material may be composed of natural or synthetic fibers selected from the group consisting of rayon, polyester, polyurethane, polyolefin, cellulose, cellulose derivatives, cotton, orlon, nylon, hydrogel polymeric material, and combinations thereof. In use, the substrate material can be an elastic substrate. Materials suitable for use as an elastic substrate in the present invention include materials which are elastic, conformable, porous, provide adequate compression and which are self-adhering. In general, the material is sufficiently porous if the material allows for the transmission of air and moisture vapor through the material.

Methods

As described above, the methods described herein can relate to inhibiting, preventing, and/or disrupting biofilms. The term “biofilm” refers to a mucilaginous community of microorganisms such as bacteria, archaea, fungi, molds, algae or protozoa or mixtures thereof that grow on various surfaces when the microorganisms establish themselves on a surface and activate genes involved in producing a matrix that includes polysaccharides.

In certain embodiments, the methods described herein can comprise methods for disrupting biofilms. In some embodiments, the method can comprise contacting a surface or substrate with a therapeutically effective amount of a composition described above. In certain embodiments, the surface or substrate is treated with a composition described above so as to leave a residue or deposit of the quart and antimicrobial metal ions on the surface or substrate that remains of the surface or substrate.

In certain embodiments, a composition described above can be used to treat articles, devices, substrates and surfaces (mammalian or inanimate) to disrupt the formation of or disrupt already formed biofilms.

In some embodiments, the surface to be treated with a composition described above includes medical devices such as catheters, respirators, and ventilators. In other embodiments, the surface can be that of implanted medical devices, including stents, artificial valves, joints, pins, bone implants, sutures, staples, pacemakers, and other temporary or permanent medical devices.

In other embodiments, substrates used for wound dressings can be treated with a composition described above to generate wound dressings that will inhibit biofilm formation, and disrupt already formed biofilms on wounds and kill the bacteria in the biofilm.

In other embodiments, the surface to be treated with a composition described above includes articles such as drains, tubs, kitchen appliances, countertops, shower curtains, grout, toilets, industrial food and beverage production facilities, flooring, and food processing equipment and the like. The surface to be treated with a composition described above in yet another embodiment includes article surfaces such as filter or heat exchanger surfaces, providing means for reducing and/or eliminating biofouling of heat exchangers or filters.

Other embodiments, relate to use on or application of a composition described above to articles, devices, substrates or surfaces associated marine structures including, but not limited to, boats, piers, oil platforms, water intake ports, sieves, and viewing ports.

In certain embodiments, the articles, substrate or device surface being treated with a composition described above can alternatively be associated with a system for water treatment and/or distribution (like drinking water treatment and/or distributing systems, pool and spa water treatment systems, water treatment and/or distribution systems in manufacturing operations, and a system for dental water treatment and/or distribution). In some embodiments, the biofilm disruptor article, substrate or device surface treated with a composition described above can also be associated with a system for petroleum drilling, storage, separation, refining and/or distribution (like petroleum separation trains, a petroleum container, petroleum distributing pipes, and petroleum drilling equipment). In other embodiments, a composition described above can also be included in formulations directed at reducing or eliminating biofilm deposits or biofouling in porous medium, such as with oil and gas bearing geological formations. In particular embodiments, a composition described above may be accomplished by applying a coating of a composition described above, such as by painting, to the surface of articles, substrate or device.

By way of non-limiting illustration, examples of certain embodiments of the present disclosure are given below.

EXAMPLES

The invention will be described in greater detail by way of specific examples. The following examples are offered for illustrative purposes, and are not intended to limit the invention in any manner. Those of skill in the art will readily recognize a variety of non- critical parameters which can be changed or modified to yield essentially the same results.

Example 1: Combinations of Quat and Antimicrobial Metal Ions for Wound Dressings to Promote Wound Healing by Disrupting Biofilms

We are interested in developing wound dressings that will promote wound healing. Substrates used for wound dressings can be treated with a nanozeolite + quat compositions described above to generate wound dressings that will inhibit biofilm formation and disrupt already formed biofilms on wounds and kill the bacteria in the biofilm and promote wound healing. Figure 2 shows an illustration of how a wound becomes non-healing due to the formation of a protective layer, known as extracellular polymeric substances (EPS) that surrounds the bacteria and prevents drugs from reaching the bacteria.

Chronic wounds last from 2 weeks to 3 months, and often associated with vascular, diabetic and pressure ulcers. In 2014, 15% of Medicare patients in the US had a wound infection, and 4% had surgical site infections. In the healthcare area, $96.8B is spent on wound care for humans. Part of this expenditure is for chronic wounds, with biofilms being associated with 78.2% of chronic wounds and 6% of acute wounds, with about $7.2B for chronic wound care. This delayed healing of chronic wounds is due to presence of bacterial biofilms in the wound, leaving the wound in a chronic inflammatory state, with concurrent tissue damage. Since biofilms resist antibiotics, antibiotic use just leads to increased antibiotic resistance. Antimicrobial resistance deaths in 2019 were 1.9 million and expected to reach 10 million by 2050. Biofilms are diverse, dependent on the bacteria, combinations of bacteria and with different strains. Antibiofilm wound dressings in 2022 was $714.7 million market and expected to expand at a compound annual growth rate (CAGR) of 9.6% from 2023 to 2030.

Since biofilms are structurally robust, and behave like viscoelastic solids, strategies to remove biofilms involve mechanical and chemical disruption (targeting eDNA, polysaccharides, and proteinaceous adhesins. Mechanical debridement can be effective, but can cause damage to healthy tissue, patient pain and spread of bacteria. For wound dressings, polymeric nanoparticles such as gelatin, alginate, chitosan, polyglycolic acids with antimicrobial encapsulants have been studied. Though there are many studies reporting improvement in wound healing with silver-based dressings, clinical evidence (Cohcrane and VULCAN reviews) suggests that silver dressings provide minimal advantage in wound healing for ulcers. Our hypothesis is that if there are mature biofilms in the wound, then silver-based dressings will not be as effective.

Nanozeolites including both silver ions and zinc ions retained at ion-exchangeable sites within the zeolite nanoparticles (termed “AM-30”) exhibit the ability to inhibit biofilm formation. The control experiment involved creating Pseudomonas aeruginosa (PA01) biofilms (growth time 24h at 37°C) on the walls of a test tube followed by treatment with crystal violet. The crystal violet-stained biofilm can be dissolved in acetic acid, followed by measurement of absorbance at 590 nm to indicate a quantitative measure of the biofilm formed. Growth of the PA01 biofilm was monitored in the presence of different concentrations of metal-zeolite AM-30 (in PBS and broth) for 24h, and then the amount of biofilm quantitated by the crystal violet assy. Figure 3 shows the results. Above concentrations of metal-zeolite over 5 ppm (Ag-200 ppb) in PBS and 50 ppm AM30 (Ag-2 ppm) in LA broth, there is complete biofilm inhibition.

Further, by delivering the antimicrobial transition metal ions using zeolite nanoparticles, the antimicrobial transition metal ions can be protected from deactivation by proteins present in biological media. To demonstrate this, we compared the ability of free silver ions and silver ions protected within zeolite nanoparticles at the same concentration of silver (2.5 ppm Ag) to kill planktonic cells of Pseudomonas aeruginosa upon a two hour exposure. Figure 4 shows that though both exposures have the same level of Ag⁺ ions, since the free silver ion in AgNCL can precipitate with the constituents of the broth, its antimicrobial ability is diminished (3-log decrease), whereas the encapsulated Ag⁺ in zeolite nanoparticles is protected from inactivation and leads to a 4-log decrease. This aspect of the protection of the active transition metal ion will be important in wound environments because of the presence of proteins and other ions that can precipitate free silver ions. Moreover, this also allows for the nanozeolite to penetrate deeper into the wound, carrying its cargo of transition metal ions.

In order to create wound dressings, zeolite nanoparticles need to be deposited on a suitable substrate material. Extracellular matrix (ECM)-based wound dressings are widely used in clinical practice. ECM-based wound dressings promote wound healing via promotion of cell proliferation and angiogenesis with minimal immune response. However, these dressing are not antimicrobial, and biofilm-infested wounds cannot take advantage of the ECM-assisted wound healing prowess. The beneficial proteins and scaffold structure in ECM collagen matrix assists in wound healing, as a tendon protector sheet, and for use as a biomaterial for stem cell delivery. Such an ECM matrix would be ideal for deposition of zeolite nanoparticles including antimicrobial transition metal ions retained at ionexchangeable sites within the zeolite nanoparticles.

Figure 5 shows a Pseudomonas aeruginosa biofilm grown on a cellulose membrane (M). On this membrane, we placed a series of ECM-based dressings for evaluation: a control ECM wound dressing (B), as well as ECM-based dressings containing AM-30 (Silver ion), silver nanoparticles (Silver NPs), and a mixture of a quat (Benzalkonium nitrate, BZN) and nanozeolites including both silver ions and zinc ions retained at ion-exchangeable sites within the zeolite nanoparticles (ZeoVation) (all designated as B) and exposed for 24 hours. After 24 hours, both the cellulose membrane M and the wound dressing B were analyzed for bacteria. As shown in Figure 6, only a wound dressing containing quat + nanozeolites including both silver ions and zinc ions retained at ion-exchangeable sites within the zeolite nanoparticles (labeled ZeoVation) eliminated bacteria on both B and M. The silver metals alone exhibit a decrease in bacteria, but not the complete killing observed with the ZeoVation formulation. Similar results were obtained with methicillin-resistant Staphylococcus aureus (MRSA) biofilms.

Example 2. Virucidal Activity of Combinations of Quat and Antimicrobial Metal Ions

Viruses can be classified into envelope viruses, which have an envelope and nonenveloped viruses, as shown in Figure 7.

Combinations of a quat and nanozeolites including antimicrobial transition metal ions retained at ion-exchangeable sites within the zeolite nanoparticles were very effective with envelope viruses, whereas the metal zeolite alone is effective against non-enveloped virus.

Figure 8 shows that quat (BZN) spread on a textile at 22 pg/cm² reduces the SARS- CoV-2 virus load (PFU/ml = plaques forming units) by 0.5 log in 10 minutes (10 mCT), but along with nanozeolites including both silver ions and zinc ions retained at ion-exchangeable sites within the zeolite nanoparticles (AM 30, 100 pg/cm²) and nanozeolites including both silver ions and copper ions retained at ion-exchangeable sites within the zeolite nanoparticles (AV30, 100 pg/cm²) reduces the SARS-CoV-2 virus load by 1.5 log. At 66 pg/cm² quat, there is a reduction of about 1.5-2 log in SARS-CoV-2 plaques, but along with AM30 (100 pg/cm²) and AV30 (100 pg/cm²) reduces the SARS-CoV-2 virus load by 5-6 log, and no evidence of virus infection is observed. At 110 pg/cm² quat, with or without the zeolite, no infected cells are observed. Thus, the 66 pg/cm² quat loading data demonstrates that combinations of a quat and nanozeolites including antimicrobial transition metal ions retained at ion-exchangeable sites within the zeolite nanoparticles have a marked effect on SARS- CoV-2 virus viability.

Example 3: Antifungal Activity of Combinations of Quat and Antimicrobial Metal Ions

Fungi, as shown in Figure 9, also have cell walls. Experiments using the following fungi: Aspergillus brasiliensis ATCC 9642, Aureobasidium pullulans ATCC 15233, Chaetomium globosum ATCC 6205, Talaromyces pinophilus ATCC 11797 and Trichoderma virens ATCC 9645 were carried out. These fungi were grown on control textile (Figure 10), on a textile containing a quat (Figure 11), and on a textile containing a quat in combination with nanozeolites including both silver ions and zinc ions retained at ion-exchangeable sites within the zeolite nanoparticles (AM 30, 100 pg/cnr) (Figure 12). The quat in combination with nanozeolites including both silver ions and zinc ions retained at ion-exchangeable sites within the zeolite nanoparticles zeolite was very effective at disrupting growth of fungi in the textile.

Claims

CLAIMS WHAT IS CLAIMED IS:

1. A wound dressing comprising a substrate material having a patient contacting surface, and a population of zeolite nanoparticles and a quaternary ammonium compound disposed on the patient contacting surface; wherein the zeolite nanoparticles comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles.

2. The wound dressing of claim 1 , wherein the substrate material comprises a dry or impregnated gauze, a film, a gel, a foam, a hydrocolloid, a woven or non-woven fabric, a hydrogel, a paste, granules, beads, or a combination thereof.

3. The wound dressing of any of claims 1-2, wherein the substrate material comprises a woven or non-woven fabric.

4. The wound dressing of any of claims 1-3, wherein the substrate material comprises collagen, cellulose, carboxymethylcellulose, oxidized regenerated cellulose (ORC), an alginate, extracellular matrix, a polyester, a polyurethane, or combinations thereof.

5. The wound dressing of any of claims 1-4, wherein the quaternary ammonium compound is deposited on the patient contacting surface at a concentration of from 10-500 pg/cm².

6. The wound dressing of any of claims 1-5, wherein the zeolite nanoparticles are deposited on the patient contacting surface at a concentration of from 10-1200 pg/cm².

7. The wound dressing of any of claims 1-6, wherein the zeolite nanoparticles are deposited on the patient contacting surface such that the antimicrobial metal ions are present on the patient contacting surface at a concentration of from 10-500 pg/cm².

8. The wound dressing of any of claims 1-7, wherein the quaternary ammonium compound is defined by the Formula I or Formula II below

Formula I Formula II wherein

9. The wound dressing of any of claims 1-8, wherein the antimicrobial metal ions comprise silver, zinc, copper, or any combination thereof.

10. The wound dressing of any of claims 1-9, wherein the antimicrobial metal ions are present in an amount of from 1% up to 50% of the ion exchange capacity of the zeolite nanoparticles, such as from 1% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 1% up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 1% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 25% of the ion exchange capacity of the zeolite nanoparticles, or from 10% up to 20% of the ion exchange capacity of the zeolite nanoparticles.

11. The wound dressing of any of claims 1-10, wherein the antimicrobial metal ions comprise silver and zinc, or wherein the antimicrobial metal ions comprise silver and copper.

12. The wound dressing of any of claims 1-11 , wherein the zeolite nanoparticles are deposited on the patient contacting surface such that silver ions are present on the patient contacting surface at a concentration of from 1-100 pg/cm², such as from 5-50 pg/cm², from 10-40 pg/cm², from 20-40 pg/cm², or from 25-35 pg/cm².

13. The wound dressing of any of claims 1-12, wherein the zeolite nanoparticles are deposited on the patient contacting surface such that zinc ions are present on the patient contacting surface at a concentration of from 0.25-25 pg/cm², such as from 1.25-12.5 pg/cm², from 2.5-10 pg/cm², from 5-10 pg/cm², or from 6.25-8.75 pg/cm².

14. The wound dressing of any of claims 1-12, wherein the antimicrobial metal ions comprise silver and zinc, and the silver and zinc are present on the patient contacting surface at a weight ratio of silver:zinc of from 1:1 to 10: 1, such as from 2:1 to 8:1, or from 3:1 to 5:1.

15. The wound dressing of any of claims 1-12, wherein the antimicrobial metal ions comprise silver and zinc, and the silver and zinc are present on the patient contacting surface at a weight ratio of silverzinc of about 4: 1.

16. A method of inhibiting or disrupting a biofilm, the method comprising contacting the biofilm with a composition comprising a population of zeolite nanoparticles and a quaternary ammonium compound dissolved or dispersed within or on the surface of a carrier; wherein the zeolite nanoparticles comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles.

17. The method of claim 16, wherein the method comprises disrupting an existing biofilm.

18. A method of preventing or inhibiting the formation of a biofilm on a surface, the method comprising contacting the surface with a composition comprising a population of zeolite nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier; wherein the zeolite nanoparticles comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles.

19. The method of claim 18, wherein the surface comprises a fomite surface.

20. The method of any of claims 18-19, wherein the surface comprises a surface of a medical device or a wound dressing.

21. The method of any of claims 18-20, wherein the quaternary ammonium compound is deposited on the surface at a concentration of from 10-500 pg/cm².

22. The method of any of claims 18-21 , wherein the zeolite nanoparticles are deposited on the surface at a concentration of from 10-1200 pg/cm².

23. The method of any of claims 18-22, wherein the zeolite nanoparticles are deposited on the surface such that the antimicrobial metal ions are present on the surface of the carrier at a concentration of from 10-500 pg/cm².

24. The method of any of claims 18-23, wherein the zeolite nanoparticles are deposited on the surface such that silver ions are present on the surface at a concentration of from 1-100 pg/cm², such as from 5-50 pg/cm², from 10-40 pg/cm², from 20-40 pg/cm², or from 25-35 pg/cm².

25. The method of any of claims 18-24, wherein the zeolite nanoparticles are deposited on the surface such that zinc ions are present on the surface at a concentration of from 0.25-25 pg/cm², such as from 1.25-12.5 pg/cm², from 2.5-10 pg/cm², from 5- 10 pg/cm², or from 6.25-8.75 pg/cm².

26. The method of any of claims 18-25, wherein the antimicrobial metal ions comprise silver and zinc, and the silver and zinc are present on the surface at a weight ratio of silverzinc of from 1:1 to 10:1, such as from 2:1 to 8:1, or from 3:1 to 5:1.

27. The method of any of claims 18-26, wherein the antimicrobial metal ions comprise silver and zinc, and the silver and zinc are present on the surface at a weight ratio of silver: zinc of about 4: 1.

28. A method of killing, inhibiting, and/or affecting a virus, the method comprising contacting the virus with a composition comprising a population of zeolite nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier; wherein the zeolite nanoparticles comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles.

29. The method of claim 28, wherein the virus comprises an enveloped virus.

30. The method of claim 29, wherein the enveloped virus comprises a DNA virus, such as a virus in the family Herpesviridae, Poxviridae, Hepadnaviridae, or Asfarviridae.

31. The method of claim 28, wherein the enveloped virus comprises an RNA virus, such as a flavivirus, an alphavirus, a togavirus, a coronavirus, a hepatitis virus, an orthomyxovirus, a paramyxovirus, a rhabdovirus, a bunyavirus, or a filovirus.

32. A method of killing, inhibiting, and/or affecting a fungus, the method comprising contacting the fungus with a composition comprising a population of zeolite nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier; wherein the zeolite nanoparticles comprise an antimicrobially effective amount of antimicrobial metal ions retained at ion-exchangeable sites within the zeolite nanoparticles.

33. The method of any of claims 16-32, wherein the quaternary ammonium compound is defined by the Formula I or Formula II below

R

R¹— N-R' R¹— NH₃ R'

Formula I Formula II wherein

R¹ is selected from C1-20 alkyl, C2-20 alkenyl, C2-20 alkynyl, Ci-2ohaloalkyl, C3-10 cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 4-20 membered heterocycloalkyl, C iocycloalkyl-Ci-io alkylene, 4-10 membered heterocycloalkyl-Ci-io alkylene, 6-10 membered aryl-Ci-10 alkylene, and 5-10 membered heteroaryl-Ci-10 alkylene, each optionally substituted with 1, 2, 3, or 4 independently selected R^x groups;

R' is, individually for each occurrence, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-4 haloalkyl, C3-10 cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 4-10 membered heterocycloalkyl, C 10 cycloalkyl -C 1-4 alkylene, 4-10 membered heterocycloalkyl-Ci-4 alkylene, 6-10 membered aryl-Ci-4 alkylene, and 5-10 membered heteroaryl-Ci.4 alkylene, each optionally substituted with 1, 2, 3, or 4 independently selected R^x groups; and each R^x, when present, is independently selected from OH, NO2, CN, halo, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, CM haloalkyl, C1-6 alkoxy, C 1-6 haloalkoxy, cyano-Ci-3 alkyl, HO- C1-3 alkyl, amino, C 1-6 alkylamino, di(Ci-6 alkyl)amino, thio, C 1-6 alkylthio, C 1-6 alkylsulfinyl, C1-6 alkylsulfonyl, carbamyl, Ci-6 alkylcarbamyl, di(Ci-6 lkyl)carbamyl, carboxy, Ci-6 alkylcarbonyl, C 1-6 alkoxycarbonyl, C 1-6 alkylcarbonylamino, C 1-6 alkylsulfonylamino, aminosulfonyl, C 1-6 alkylaminosulfonyl, di(C 1-6 alky l)aminosulfonyl, aminosulfonylamino, C1-6 alkylaminosulfonylamino, di(C 1-6 alky l)aminosulfonylamino, aminocarbonylamino, C1-6 alkylaminocarbonylamino, and di(Ci-6 alkyl)aminocarbonylamino.

34. The method of any of claims 16-33, wherein the quaternary ammonium compound is present in the composition in an amount of from 10 ppm to 500 ppm.

35. The method of any of claims 16-34, wherein the antimicrobial metal ions comprise silver, zinc, copper, or any combination thereof.

36. The method of any of claims 16-35, wherein the antimicrobial metal ions are present in an amount of from 1% up to 50% of the ion exchange capacity of the zeolite nanoparticles, such as from 1% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 1% up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 1% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 25% of the ion exchange capacity of the zeolite nanoparticles, or from 10% up to 20% of the ion exchange capacity of the zeolite nanoparticles.

37. The method of any of claims 16-36, wherein the antimicrobial metal ions comprise silver and zinc, or wherein the antimicrobial metal ions comprise silver and copper.

38. The method of any of claims 16-37, wherein the zeolite nanoparticles are present in the composition in an amount of from 10 ppm to 10,000 ppm.

39. A method of inhibiting or disrupting a biofilm, the method comprising contacting the biofilm with a composition comprising an antimicrobially effective amount of antimicrobial metal ions from a metal salt or metal nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier.

40. The method of claim 39, wherein the method comprises disrupting an existing biofilm.

41. A method of preventing or inhibiting the formation of a biofilm on a surface, the method comprising contacting the surface with a composition comprising an antimicrobially effective amount of antimicrobial metal ions from a metal salt or metal nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier.

42. The method of claim 41 , wherein the surface comprises a fomite surface.

43. The method of any of claims 41-42, wherein the surface comprises a surface of a medical device.

44. The method of any of claims 41-43, wherein the quaternary ammonium compound is deposited on the surface at a concentration of from 10-600 pg/cm².

45. The method of any of claims 41-44, wherein the antimicrobial metal ions are deposited on the surface at a concentration of from 10-1200 pg/cnr.

46. The method of any of claims 41-45, wherein the antimicrobial metal ions comprise silver ions, and the silver ions are deposited on the surface at a concentration of from 1-100 pg/cm², such as from 5-50 pg/cm², from 10-40 pg/cm², from 20-40 pg/cm², or from 25-35 pg/cm².

47. The method of any of claims 41-46, wherein the antimicrobial metal ions comprise zinc ions, and the zinc ions are deposited on the surface at a concentration of from 0.25-25 pg/cm², such as from 1.25-12.5 pg/cm², from 2.5-10 pg/cm², from 5-10 pg/cm², or from 6.25-8.75 pg/cm².

48. The method of any of claims 41-47, wherein the antimicrobial metal ions comprise silver and zinc, and the silver and zinc are present on the surface at a weight ratio of silverzinc of from 1:1 to 10:1, such as from 2: 1 to 8: 1, or from 3:1 to 5: 1.

49. The method of any of claims 41-48, wherein the antimicrobial metal ions comprise silver and zinc, and the silver and zinc are present on the surface at a weight ratio of silver: zinc of about 4: 1.

50. A method of killing, inhibiting, and/or affecting a virus, the method comprising contacting the virus with a composition comprising an antimicrobially effective amount of antimicrobial metal ions from a metal salt or metal nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier.

51. The method of claim 50, wherein the virus comprises an enveloped virus.

52. The method of claim 51, wherein the enveloped virus comprises a DNA virus, such as a virus in the family Herpesviridae, Poxviridae, Hepadnaviridae, or Asfarviridae.

53. The method of claim 51 , wherein the enveloped virus comprises an RNA virus, such as a flavivirus, an alphavirus, a togavirus, a coronavirus, a hepatitis virus, an orthomyxovirus, a paramyxovirus, a rhabdovirus, a bunyavirus, or a filovirus.

54 A method of killing, inhibiting, and/or affecting a fungus, the method comprising contacting the fungus with a composition comprising an antimicrobially effective amount of antimicrobial metal ions from a metal salt or metal nanoparticles and a quaternary ammonium compound dissolved or dispersed in a carrier.

55. The method of any of claims 39-54, wherein the quaternary ammonium compound is defined by the Formula I or Formula II below

R

R¹— N-R'

R¹— NH₃ R'

Formula I Formula II wherein

R¹ is selected from C1-20 alkyl, C2-20 alkenyl, C2-20 alkynyl, Ci-2ohaloalkyl, C3-10 cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 4-20 membered heterocycloalkyl, Cs-iocycloalkyl-Ci-io alkylene, 4-10 membered heterocycloalkyl-Ci-io alkylene, 6-10 membered aryl-Ci-10 alkylene, and 5-10 membered heteroaryl-Ci-10 alkylene, each optionally substituted with 1, 2, 3, or 4 independently selected R^x groups;

R' is, individually for each occurrence, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-4 haloalkyl, C3-10 cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 4-10 membered heterocycloalkyl, Cv m cycloalky I -C 1-4 alkylene, 4-10 membered heterocycloalkyl-Ci-4 alkylene, 6-10 membered aryl-Ci-4 alkylene, and 5-10 membered heteroaryl-Ci-4 alkylene, each optionally substituted with 1, 2, 3, or 4 independently selected R^x groups; and each R^x, when present, is independently selected from OH, NO2, CN, halo, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-4 haloalkyl, C1-6 alkoxy, C 1-6 haloalkoxy, cyano-Ci-3 alkyl, HO- C1-3 alkyl, amino, C 1-6 alkylamino, di(C 1-6 alky l)amino, thio, C 1-6 alkylthio, C 1-6 alkylsulfinyl, C1-6 alkylsulfonyl, carbamyl, C 1-6 alkylcarbamyl, di(Ci-6 alkyl)carbamyl, carboxy, Ci-6 alkylcarbonyl, C 1-6 alkoxycarbonyl, C 1-6 alkylcarbonylamino, C 1-6 alkylsulfonylamino, aminosulfonyl, Ci-6 alkylaminosulfonyl, di(Ci-6 alkyl)aminosulfonyl, aminosulfonylamino, C1-6 alkylaminosulfonylamino, di(C 1-6 alky l)aminosulfonylamino, aminocarbonylamino, C1-6 alkylaminocarbonylamino, and di(Ci-6 alkyl)aminocarbonylamino.

56. The method of any of claims 39-55, wherein the quaternary ammonium compound is present in the composition in an amount of from 10 ppm to 500 ppm.

57. The method of any of claims 39-56, wherein the antimicrobial metal ions comprise silver, zinc, copper, or any combination thereof.

58. The method of any of claims 39-57, wherein the antimicrobial metal ions comprise antimicrobial metal nanoparticles.

59. The method of any of claims 39-58, wherein the antimicrobial metal ions comprise antimicrobial metal nanoparticles disposed on or within zeolite nanoparticles.

60. The method of any of claims 39-59, wherein the antimicrobial metal ions are retained at ion-exchangeable sites within zeolite nanoparticles

61. The method of claim 60, wherein the antimicrobial metal ions are present in an amount of from 1% up to 50% of the ion exchange capacity of the zeolite nanoparticles, such as from 1% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 1% up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 1% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 25% of the ion exchange capacity of the zeolite nanoparticles, from 5% up to 20% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 50% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 30% of the ion exchange capacity of the zeolite nanoparticles, from 10% up to 25% of the ion exchange capacity of the zeolite nanoparticles, or from 10% up to 20% of the ion exchange capacity of the zeolite nanoparticles.

62. The method of any of claims 39-61, wherein the antimicrobial metal ions comprise silver and zinc, or silver and copper.

63. The method of any of claims 39-62, wherein the antimicrobial metal ions are present in the composition in an amount of from 10 ppm to 10,000 ppm.

64. The wound dressing of any of claims 1-15, wherein the population of zeolite nanoparticles and the quaternary ammonium compound are Coulombically associated with one another.

65. The method of any of claims 16-38, wherein the population of zeolite nanoparticles and the quaternary ammonium compound are Coulombically associated with one another.

66. A method for protecting antimicrobial metal ions within a biological environment, the method comprising retaining the antimicrobial metal ions at ion-exchangeable sites within zeolite nanoparticles.

67. The method of claim 66, wherein the biological environment comprises a wound.

68. The method of any of claims 66-67, wherein the method comprises protecting the antimicrobial metal ions from deactivation, thereby preserving their antimicrobial activity.

69. A method of increasing the penetration depth of antimicrobial metal ions with a wound, the method comprising retaining the antimicrobial metal ions at ion-exchangeable sites within zeolite nanoparticles, and contacting the wound with the zeolite nanoparticles.

70. The method of claim 69, further comprising Coulombically associating the zeolite nanoparticles with a quaternary ammonium compound.

71. The method of any of claims 69-70, wherein Brownian motion of the zeolite nanoparticles results in the increase in penetration depth.