Cross-linkable antibiotic compounds and durable self-decontaminating antibiotic coating materials and coated substrates containing the same

Abstract

Antibiotic compounds that are reaction products of an antibiotic molecule and a cross-linkable silane molecule are provided which are useful for incorporation into a liquid base coating material. The antibiotic molecule may be a fluoroquinolone, penicillin, cephalosporin, tetracycline, macrolide and / or aminoglycoside, while the cross-linkable silane molecule may be an epoxy-functionalized silane, an aldehyde-functionalized silane and / or a halogenated silane. Coating formulations including the antibioitic compounds and coated substrates which are coated with such coating formulations are also provided.

Description

CROSS-LINKABLE ANTIBIOTIC COMPOUNDS AND DURABLE SELF-DECONTAMINATING ANTIBIOTIC COATING MATERIALS AND COATED SUBSTRATES CONTAINING THE SAMECROSS-REFERENCE TO RELATED APPLICATION

[0001] This application is based on and claims priority benefits from U.S. Provisional Patent Application Serial No. 63 / 701 ,302 filed on September 30, 2024, the entire content of which is expressly incorporated hereinto by reference.GOVERNMENT RIGHTS

[0002] This invention was made with support by the Defense Advanced Research Projects Agency (DARPA) under contract number W31 P4Q-22-C-0001 . The Government has certain rights in the invention.FIELD

[0003] The embodiments disclosed herein relate generally to antibiotic compounds that are capable of crosslinking with substrate coating materials (e g., paint coatings) and to substrates coated with such coating materials that provide self-decontamination antibiotic properties upon contact with harmful microbial agents, e.g., gram-positive and gramnegative bacteria.BACKGROUND AND SUMMARY

[0004] Given the recent pandemic caused by the outbreak of coronavirus (COVID-19) and the persistent threat of biological warfare to deployed U.S forces, the need for active decontamination methods to neutralize biological threats is at an all-time high. Decontamination of surfaces comprises removal or neutralization of bacteria is labor intensive, must be continually applied and potentially use materials that are notenvironmentally safe. The ideal approach would be to provide surface coatings with an additive compound that will rapidly self-decontaminate the harmful bacteria upon contact. However, there are no commercially available products that offer both the required durability and antibacterial activity.

[0005] The embodiments described herein relate to a cross-linkable antibiotic additive compound that is compatible with sol-gel coatings and paint products and is capable of decontaminating a variety of microbial agents, e.g., gram-positive and gram-negative bacteria, upon contact. The antibiotic additive compound in accordance with the embodiments disclosed herein is generally structurally comprised of an antibiotic head and a cross-linkable silane tail. In certain embodiments, the cross-linkable antibiotic compound is preferably the reaction product of norfloxacin and a cross-linkable silane (hereinafter referred to as a “NOR-Silane” compound which is shown in Figure 1 ).

[0006] The embodiments disclosed herein are thus also directed toward coating additives that are compatible with durable sol-gel coating platforms, fabrics, and paints to create a robust, multi-functional, and selfdecontaminating surface coating. Utilizing a modular small-molecule incorporated in a topcoat paint system or thin coating as schematically shown in Figure 2 that provides biological activity, the additive delivers active biological decontamination and against bacterial threats on heavily used non-porous surfaces. The antimicrobial silane tail of the compound may therefore be covalently tethered to the coating material so that the antibiotic head is available for decontamination thereby providing a very durable antibiotic coating material with no leaching of the active antibiotic agent and ensuring long service life. The coating materials according to the embodiments disclosed herein thereby provide durable antibiotic functionality, e.g., providing active >99% kill of bacteria at the surface for sustained use over a sustained period.

[0007] These and other aspects of the present invention will become more clear after careful consideration is given to the following detailed description of a presently preferred exemplary embodiment thereof.BRIEF DESCRIPTION OF THE DRAWINGS

[0008] Reference will be made to the accompanying drawing Figures, wherein:

[0009] FIG. 1 shows the chemical structure of the exemplary NOR- Silane molecule in accordance with an embodiment of the invention;

[0010] FIG. 2 schematically shows the chemical structure and workings of the exemplary NOR-Silane molecule with crosslinking functionality provided by the cross-linkable silane tail of the molecule and decontamination functionality provided by the norfloxacin antibiotic head of the molecule;

[0011] FIG. 3 shows an exemplary chemical reaction scheme for synthesizing the NOR-Silane compound disclosed herein;

[0012] FIG. 4 is a nuclear magnetic resonance (NMR) spectrum of the NOR-Silane molecule taken in deuterated chloroform;

[0013] FIG. 5 is a tabular presentation of the crosshatch adhesion classification according to ASTM D3359;

[0014] FIG. 6 are photographs showing the crosscut adhesion analysis of self-decontaminating coating materials in varying concentrations of the NOR-Silane compound from 5 to 10 wt.% (based on total weight of the coating material);

[0015] FIG. 7 is a graphic illustration of various exemplary silane coupling units that may be employed to covalently bond with norfloxacin toprovide the NOR-Silane compounds according to the disclosed embodiments; and

[0016] FIG. 8 is a graphic illustrating various silane coupling functionalities to allow the cross-linkable silane tail of the antimicrobial silane compound to be covalently tethered (bonded) to the coating material, whereby the wavy lines in FIG. 8 represent the antimicrobial head components of the molecules.DETAILED DESCRIPTION

[0017] The cross-linkable antibiotic molecules described herein are provided with i) a crosslinking functionality for long term durability and non-leachability, and ii) a chemically (preferably covalently) bound active antibiotic agent.

[0018] The disclosed material can incorporate a variety of silane coupling units to enhance its properties and facilitate bonding with different substrates. Exemplary silane coupling units include, for example, epoxy-functional ized silanes such as 2-(3,4- epoxycyclohexyl)ethyltriethoxysilane and 2-(3,4- epoxycyclohexyl)ethyltriethoxysilane, which provide functionality by virtue of their reactive epoxy groups. Aldehyde-functionalized silanes, such as, for example, triethoxysilylbutyraldehyde and triethoxysilylbutyraldehyde, may also be employed and offer different reactive sites for chemical attachment. Additionally, halogenated silanes, such as chloromethyltriethoxysilane and chloromethyltriethoxysilane, can be employed for their versatile reactivity. For applications requiring robust covalent bonding through an isocyanate linkage, 3-isocyanatopropyl triethoxysilane represents an especially preferred silane coupling unit that may be employed in the embodiments of this invention. The cross-linking functionality is most preferably a silane coupling agent, such as 3-isacyanatopropyltrithoxysilane which work by cross-linking the decontaminating materials to the bulk of the coating material (see e.g., Figure 1). Other examples of various silane coupling units are shown in Figure 7 and Figure 8.

[0019] Multiple antibiotic molecules may be employed in the embodiments disclosed herein. Preferably the antibiotic molecules are biocidal molecules in the chemical class of fluoroquinolones can be used to synthesize the cross-linkable antibiotic silane compounds of the invention having the silane coupling units as described hereinabove. Such biocidal molecules can therefore provide the antibiotic head of the crosslinkable antibiotic silane compounds and can thereby provide the “antibacterial function” as schematically shown in Figure 2. Examples of biocidal molecules which can be used for antibiotic functionality include at least one fluoroquinolone selected from the group consisting of norfloxacin, ciprofloxacin, levofloxacin, moxifloxacin, ofloxacin, delafloxacin, and gemifloxacin. The antibiotic tethering approach using a cross-linkable silane tail can also be used with any other antibiotic or biocide materials, such as penicillin, cephalosporins, tetracyclines, macrolides, and aminoglycosides.

[0020] The embodiments disclosed herein will be better understood by reference to the following Examples.EXAMPLESA. Exemplary Synthesis of NOR-Silane Molecule

[0021] An exemplary synthesis procedure of the NOR-silane molecule shown in Figure 1 is depicted in Figure 3. More specifically, Norfloxacin (12.17 g, 38.12 mmol) and anhydrous dichloromethane (DCM, 50 mL) are added to a 100 mL round bottom flask equipped with a magnetic stir bar. Dichloromethane (DCM) is used to wash any residual Norfloxacin down the side of the flask. A rubber septum is added, and thesolution is sparged with argon. After 15 minutes, 3- isocyanatopropyltriethoxysilane (10.0 mL, 40.021 mmol, 1.05 excess) is added in one charge. The solution is then allowed to stir at 25 °C for 1 hour, after which point it is concentrated on a roto-evaporator. Unreacted starting materials are removed by concentrating the reaction solution under reduced pressure.

[0022] Nuclear magnetic resonance (NMR) provides a spectral confirmation and characterization of molecules. NMR spectra were taken of freshly-synthesized NOR-silane molecule in deuterated chloroform, and can be used to confirm correct chemical synthesis. An example NMR spectrum of the NOR-silane molecule in deuterated chloroform is shown in Figure 4.B. Exemplary Coating Formulations

[0023] The antibiotic coating formulations as described herein may be applied to virtually any substrate to which the base coating material can be applied. Thus, the antibiotic coating formulations can be applied to flexible woven or nonwoven fabrics or films that may be employed to construct protective apparel or other equipment, such as tents, awnings, covers and the like. The antibiotic coating formulations may also be used to coat a variety of rigid substrate surfaces, such as surfaces formed of metal, polymeric, ceramic, refractory materials and the like.

[0024] The antibiotic coating may be applied by any convenient coating technique, such as spraying, brush or roll application, padding, or the like. Once applied onto the substrate surface, the antibiotic coating is allowed to cure, e.g., by acid-catalyzed curing at ambient air temperature or at accelerated elevated temperature.

[0025] While the antibiotic efficacy has been demonstrated in the exemplified sol-gel base coating system, it is understood that the biocidalNOR-Silane compound could be included in any other liquid base coating material, such as acrylic architectural paint, and the like.B1. Sol-Gel Coating Formulations

[0026] An exemplary sol-gel coating formulation in which the biocidal NOR-Silane molecule may be employed in shown in Table 1 below.Table 1. Base Sol-Gel Formulation

[0027] The preferred NOR-Silane molecule as shown in Figure 1 was loaded into a base sol-gel coating formulation at a loading level from 5 to 10 wt % (based on total weight of the coting formulation) to test efficacy. The thus formed antibiotic coating formulation showed complete kill (6 log kill, >99.9999%) of S. aureus (Table 2) and E. coli bacteria . As demonstrated, therefore, the highly loaded coatings performed very well against biocidal challenges.

[0028] In order to make a viable commercial antibiotic coating product, it is also necessary to determine the lowest percent of biocide needed to afford activity. As such, a ladder study was performed wherecoupons coated with 5 wt.%, 6.25 wt.%, 7.5 wt.%, 8.75 wt.%, and 10.0 wt. % (based on total weight of the coting formulation) solids selfdecontamination biocidal coatings were tested using the test method ISO 22196. The results of such testing appear in Tables 2 and 3 below. As can be seen in Table 2 and Table 3, 10 wt.% of the NOR-silane molecule was shown to provide >4 logio bacterial reduction in the coating material.Table 2. ISO 22196 antibacterial efficacy results against S. aureus ATCC 6538 with norfloxacin silane utilizing a NOR-Silane molecule.Table 3. ISO 22196 antibacterial efficacy results against E. Coli ATCC 8739 with norfloxacin silane utilizing a NOR-Silane molecule.B2. Acrylic Coating Formulations

[0029] An alternative coating material of standard acrylic interior paint with the NOR-Silane material incorporated also provides surfacedecontaminating with antimicrobial performance as described below.

[0030] A ladder study was conducted to determine the biocidal efficacy of the NOR-silane in standard acrylic interior paint. NOR-silane in paint show complete bacterial kill against E. coli (Table 4), and shows limiting antibacterial effect against P. aeruginosa (Table 5).Table 4. Biocidal efficacy of NOR-Silane material incorporated in standard acrylic interior paint versus E. coli bacteria.*Limit of detection. Plate counts fell below the limit of detection of 1.25E+00 CFU / cmA2Table 5. Biocidal efficacy of NOR-Silane material incorporated in standard acrylic interior paint versus S. aureus bacteria.*Limit of detection. Plate counts fell below the limit of detection of 1.25E+00 CFU / cmA2C. Crosshatch Adhesion

[0031] It is imperative that a commercially successful coating material provides proper adhesion to a variety of substrate surfaces to achieve a useful commercial product. A crosshatch adhesion test is conducted using pressure-sensitive tape to determine the adhesion quality of a coating. In order for the developed coating to perform properly, it must adhere to the surface on which the coatings are applied, and the crosshatch adhesion test can determine whether or not a coating is properly adhering to its substrate. ASTM D3359 represents the Standard Test Method for Measuring Adhesion by Tape Test, and a rating classification of “5B” represents a perfect adhesion rating, meaning no delamination.

[0032] Example appearances and the classification of the adhesion performance of the test can be visualized from the ASTM D3359 crosshatch adhesion-classification chart shown in Figure 5. In preferred embodiments, the coatings disclosed herein will comply with Classification 5B under the ASTM D3359 guidelines. Under this classification, theedges of the cuts are completely smooth, and none of the squares of the lattice is detached after application and removal of the adhesion tape.

[0033] Utilizing a ladder study of varying concentrations of the down selected NOR-Silane, all samples showed excellent adhesion to glass. The NOR-Silane was selected due to its excellent performance as a disinfectant molecule. The results for NOR-silane in sol-gel with a varying ladder concentration appear in Table 6, and the NOR-silane in standard acrylic paint at 10 wt.% concentration is shown in Table 7 below. Example result microscope images of post-cross-hatch damage is shown in Figure 6.Table 6. X-Hatch adhesion (ASTM D3359) results of NOR-silane in sol-gel at varying concentrations.Table 7. X-Hatch adhesion (ASTM D3359) results of NOR-silane in standard acrylic paint at 10 wt.% concentration.

[0034] As is shown in Tables 6 and 7 above, all selfdecontaminating additive (NOR-Silane) in sol-gel formulations and acrylic paint in a variety of loadings performed to the 5B adhesion standardaccording to ASTM D3359. Furthermore, when compared to the adhesion of the base sol-gel formula and no-additive paint samples that acts as the matrix of the coating, there is no visual change in the adhesion when the cross-linkable antimicrobial silane molecule is added to the formula. Examples of the crosscut adhesion test for each formulation can be seen in Figure 6, where each formulation exhibited zero flaking of the coating after application and removal of the adhesive tape.

[0035] Additionally, these tests represent quantities of the biocidal molecule that are expected to far exceed the wt. % incorporation that would likely be present in a finalized formulation. Thus, the cross-linkable antimicrobial silane compound can be incorporated into a liquid coating carrier material in amounts, based on total weight of the coating formulation including the liquid coating carrier material and the crosslinkable antimicrobial silane compound, ranging from at least about 0.1 wt.% up to about 30 wt.%, for example up to about 5 wt.%, about 6.5 wt.%, up to about 7.5 wt.%, or even up to about 8.5 wt.%, it will usually be present in amounts to provide effective antimicrobial (e.g., antibiocidal) effects of between about 5 wt.% to about 10 wt.%.**********

[0036] While the invention has been described in connection with what is presently considered to be the most practical and preferred embodiment, it is to be understood that the invention is not to be limited to the disclosed embodiment, but on the contrary, is intended to cover various modifications and equivalent arrangements included within the spirit and scope thereof.

Claims

WHAT IS CLAIMED IS:1 . An antibiotic compound which is a reaction product of an antibiotic molecule and a cross-linkable silane molecule, wherein the antibiotic molecule is selected from the group consisting of fluoroquinolones, penicillin, cephalosporins, tetracyclines, macrolides and aminoglycosides.

2. The antibiotic compound according to claim 1 , wherein the antibiotic molecule is a fluoroquinolone selected from the group consisting of norfloxacin, ciprofloxacin, levofloxacin, moxifloxacin, ofloxacin, delafloxacin and Gemifloxacin.

3. The antibiotic compound according to any one of claims 1-2, wherein cross-linkable silane molecule is selected from the group consisting of epoxy-functionalized silanes, aldehyde-functionalized slianes and halogenated silanes.

4. The antibiotic compound according to any one of claims 1-3, wherein the cross-linkable silane molecule is selected from the group consisting of 3-isocyanatopropyl triethoxysilane, 2-(3,4- epoxycyclohexyl) ethyltriethoxysilane, 2-(3,4-epoxycyclohexyl) ethyltriethoxysilane, triethoxysilylbutyraldehyde, triethoxysilylbutyraldehyde, chloroethyltriethoxysilane and chloromethyltriethoxysilane.

5. The antibiotic compound according to any one of claims 1-4, wherein the antimicrobial compound is the reaction product of norfloxacin and 3-isocyanatopropyl triethoxysilane.

6. A cross-linkable antibiotic compound of the formula:

7. A coating formulation which comprises the antibiotic compound according to any one of the preceding claims.

8. The coating formulation according to claim 7, wherein the coating formulation comprises a liquid base coating material and an antibiotic effective amount of the antibiotic compound.

9. The coating formulation according to claim 8, wherein the coating formulation comprises a sol-gel coating material or an acrylic paint material.

10. The coating formulation according to any one of claims 7-9, wherein the antibiotic compound is present in an effective antibiotic amount, based on total weight of the coating formulation, of at least about 0.1 wt.% up to about 10 wt.%, preferably up to about 5 wt.%, about 6.5 wt.%, up to about 7.5 wt.%, or up to about 8.5 wt.%.11 . The coating formulation according to claim 10, wherein the antibiotic compound is present in an effective antibiotic amount, based on total weight of the coating formulation, of between about 5 wt.% to about 10 wt%.

12. A coated substrate which comprises a coating of the coating formulation according to any one of claims 7-11 .

13. The coated substrate according to claim 12, wherein the coating formulation is cured such that the antibiotic compound is chemically anchored to the cured coating formulation by silane linkages of the silane molecule.

14. The coated substrate according to any one of claims 12-13, wherein the substrate is selected from substrates formed of materials selected from the group consisting of woven or nonwoven fabrics or films, metals, polymers, ceramics and refractory materials.

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