WO2025191282A1

WO2025191282A1 - Antimicrobial composition

Info

Publication number: WO2025191282A1
Application number: PCT/GB2025/050533
Authority: WO
Inventors: Gareth WYNN-JONES; Ewa King; Graeme Kettlewell
Original assignee: Speciality Fibres and Materials Ltd
Current assignee: Speciality Fibres and Materials Ltd
Priority date: 2024-03-14
Filing date: 2025-03-14
Publication date: 2025-09-18
Anticipated expiration: 2026-09-14
Also published as: GB202403729D0; GB2639579A

Abstract

There is provided an antimicrobial composition comprising: an antimicrobial compound; and a population of metal nanoparticles. Also described is a method of producing such an antimicrobial composition, a coating or treatment for a surface or substrate comprising such an antimicrobial composition, and a medical device comprising such a coating or treatment.

Description

Antimicrobial Composition

Technical Field

The present invention relates to antimicrobial compositions and to methods for producing antimicrobial compositions.

Metals including silver, copper, zinc and mercury are known for their antimicrobial properties. A renewed interest has developed in the use of metallic silver as an antimicrobial agent, especially in wound dressings, driven in part by the development of antibiotic resistant bacteria. Metallic silver is a broad-spectrum antimicrobial agent which has been proven to be effective against such resistant bacteria as well as other microorganisms. Current research suggests that due to its mode of action, metallic silver does not allow for the development of bacterial resistance. WO2015/040435 and W02023/017240 by the present Applicant describe processes for preparing cellulose fibres impregnated with metal nanoparticles, and a process for preparing a solution of metal nanoparticles. As used herein, the term “metal nanoparticles” means particles of elemental metal having an average (i.e. mean) diameter of no more than 100 nm.

Growing antibiotic and antimicrobial resistance has led to a demand for new or more effective antibiotic and antimicrobial drugs.

It is an aim of the present invention to provide novel or improved antimicrobial compositions.

Summary of Invention

The present inventors have surprisingly found that existing antimicrobial and antibacterial compositions are enhanced by the presence of a population of metal nanoparticles to a greater extent than would be expected than by combining their individual effects.

37983926-1 Disclosed herein is an improved antimicrobial composition comprising an antimicrobial compound and a population of metal nanoparticles.

According to a first aspect of the invention, there is provided an improved antimicrobial composition comprising an antimicrobial compound and a population of metal nanoparticles comprising a polymer coating, wherein the population of metal nanoparticles has a mean diameter from 5 nm to less than 20 nm, and the polymer coating comprises poly(N-vinylpyrrolidone).

The antimicrobial composition may further comprise a carrier. The carrier may comprise a fluid. The fluid may be water and/or an organic compound. The carrier may comprise a colloid e.g. a gel or an emulsion. The emulsion may comprise a water-in-oil or an oil- in-water emulsion. The emulsion may comprise an emulsifier and/or a stabiliser. The antimicrobial composition may be present within either the continuous or the discontinuous phase. The carrier may comprise a cream. The carrier may comprise a gel e.g. a hydrogel.

The antimicrobial composition may be surfactant-free.

The antimicrobial compound and/or the population of metal nanoparticles may be present at concentrations below their respective individual minimum bactericidal concentration (MBC). For example, the antimicrobial compound and/or the population of metal nanoparticles may be present at concentrations of no greater than 50% of their MBCs. In some embodiments, the antimicrobial compound and/or the population of metal nanoparticles may be present at concentrations of no greater than 45, 40, 35, 30, 25, or 20% of their individual MBCs. In some embodiments, the antimicrobial compound and/or the population of metal nanoparticles may be present at concentrations of at least 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, or 25% of their individual MBCs.

The antimicrobial compound and/or the population of metal nanoparticles may be present at concentrations below their respective minimum inhibitory concentration (MIC). For example, the antimicrobial compound and/or the metal nanoparticles may be present at a concentration of less than 50% of their MIC. In some embodiments, the antimicrobial compound and/or the population of metal nanoparticles may be present at concentrations of no greater than 45, 40, 35, 30, 25, or 20% of their individual MICs. In

37983926-1 some embodiments, the antimicrobial compound and/or the population of metal nanoparticles may be present at concentrations of at least 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, or 25% of their individual MICs.

The antimicrobial composition may have a weight ratio of the antimicrobial compound to the metal nanoparticles from 1 :4 to 4: 1 , such as 3: 1 , 2: 1 , 1 :1 , 1 :2, 1 :3.

In some embodiments, the population of metal nanoparticles may have a concentration of less than 20ppm, or preferably, less than 15ppm. In some embodiments, the population of metal nanoparticles may have a concentration of less than 14, 13, 12, 11 , 10, 9, 8, 7, 6, 5, 4, 3, 2, 1 , 0.75, 0.5, 0.25, or 0.1ppm. In some embodiments, the population of metal nanoparticles has a concentration between 0.1 and 10ppm, or 0.5 and 5ppm.

The metal nanoparticles may comprise a metal selected from the group consisting of: silver, copper, zinc, selenium, gold, cobalt, nickel, zirconium, molybdenum, gallium, iron, or any combination thereof. In a preferred series of embodiments, the metal is silver and/or copper, and more preferably, silver.

The metal nanoparticles may comprise a polymer coating. The polymer may be chosen from a group consisting of: a polyamide, polyimide, polyethyleneimine, polyvinylalcohol, pectin, albumin, gelatin, carrageenan, a gum, cellulose or a derivative thereof, poly (N- vinylpyrrolidone), poly (N-vinylcaprolactam), and mixtures thereof. For example, the gum may be xanthan, guar, Arabic, acacia etc. For example, the cellulose derivative may be hydroxyethylcellulose, hydroxypropylcellulose, methylcellulose, hydroxypropylmethylcellulose etc. In a preferred series of embodiments, the polymer is poly (N-vinylpyrrolidone). Poly (N-vinylpyrrolidone) is also known as Povidone, Polyvidone, or PVP.

The polymer may have a weight average molecular weight (M_w) of 8 to 360kg/mol, or from 20 to 80 kg/mol. The polymer may have a M_w of more than 10, 15, 20, 25, 30, 32, 34, 36, 38, or 40 kg/mol. The polymer may have a M_w of less than 360, 300, 250, 200, 150, 100, 80, 70, 60, 50, 45, 40, 38, 36, 34, 32, or 30 kg/mol. In a series of embodiments, the polymer may have a weight average molecular weight (M_w) of 25 to 45 kg/mol, 30 to 40 kg/mol, 32 to 38, or 34 to 36 k/mol.

37983926-1 For example, in a series of embodiments, the polymer is poly (N-vinylpyrrolidone) and wherein the polymer has a weight average molecular weight (M_w) of 30 to 40kg/mol.

The population of polymer-coated metal nanoparticles may comprise metal nanoparticles having a polymer coating with an average thickness of from 40 to 100 nm. Optionally, the polymer-coated metal nanoparticles may have a polymer coating with an average thickness between 50 and 90nm, 55 to 85nm, 60 to 80nm, or 65 to 75nm.

The metal nanoparticles may have a zeta potential with a magnitude of less than 10mV. Optionally, the zeta potential may have a magnitude less than 9, less than 8, less than

7, or less than 6mV. In some embodiments, the zeta potential may have a magnitude between 4 and 6mV, or may be 5mV.

The population of metal nanoparticles may comprise metal nanoparticles having an average (mean) diameter of from 2 to 50nm. In a preferred series of embodiments, the mean diameter may be from 5 to 20nm, optionally 6 to 19 nm, 7 to 18, 8 to 17, 9 to 16nm, or 10 to 15nm. In some embodiments, the mean diameter may be at least 5, 6, 7, 8, 9, 10, or 11 nm. In some embodiments, the mean diameter may be less than 20, 19, 18,

17, 16, 15, 14, 13, or 12nm.

The median diameter may be between 5 and 15nm, optionally from 6 to 14, 7 to 13, 8 to 12, or 9 to 11 nm. In some embodiments, the median diameter may be at least 5, 6, 7,

8, 9, 10, or 11 nm. In some embodiments, the median diameter may be less than 20, 19,

18, 17, 16, 15, 14, 13, 12, or 11nm.

The range of nanoparticle diameters within the population may have a standard deviation of 4 to 15nm, from 5 to 14nm, from 6 to 13nm, from 7 to 12nm. In some embodiments, the population of metal nanoparticles may have a standard deviation greater than 4, 5, 6, or 7nm. In some embodiments, the population of metal nanoparticles may have a standard deviation less than 15, 14, 13, 12, or 11nm.

The metal nanoparticles may have a D10 particle size of 6nm or less, 5nm or less, or 4nm or less. The metal nanoparticles may have a D90 particle size of 15nm or greater. In some embodiments, the D90 particle size is 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26,

37983926-1 27, 28, 29, 30nm, or greater than 30nm. In some embodiments, the D90 particle size is between 15-25nm, 16-24nm, 17-23nm, 18-22nm, 19-21nm.

The population of metal nanoparticles may comprise metal nanoparticles with a diameter greater than 20nm, greater than 25nm, greater than 30nm, greater than 35nm, or greater than 40nm.

The population of metal nanoparticles may comprise less than 15% of nanoparticles with diameters greater than 25nm. Optionally, the population may comprise less than 10%, 8%, 6%, or 5% of nanoparticles with diameters greater than 25nm. In some embodiments, the population comprises at least s, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, or 15% of nanoparticles with diameters greater than 25nm.

The population of metal nanoparticles may be formed by: a) mixing a first aqueous alkaline solution with an aqueous polymer solution to form an aqueous alkaline polymer solution; and b) mixing the aqueous alkaline polymer solution with an aqueous solution of a metal salt to form a solution of polymer-coated metal nanoparticles.

The first aqueous alkali solution may comprises a Group I hydroxide (e.g. sodium or potassium hydroxide), a Group I carbonate (e.g. Na2COs or K2CO3), a Group I bicarbonate (e.g. NaHCOs or KHCO3), a tetraalkylammonium hydroxide (e.g. tetraethylammonium hydroxide), or mixtures thereof. In a preferred series of embodiments, the first aqueous solution comprises sodium hydroxide and sodium carbonate.

The metal salt may comprise a metal selected from the group consisting of: silver, copper, zinc, selenium, gold, cobalt, nickel, zirconium, molybdenum, gallium, iron, or any combination thereof. In a preferred series of embodiments, the metal is silver.

The metal salt may be a nitrate, an acetate, a carbonate, a bicarbonate, a sulphate, or mixtures thereof. In a preferred series of embodiments, the metal salt is a nitrate. In a preferred series of embodiments, the metal salt is silver nitrate.

37983926-1 In a series of embodiments, the solution of polymer-coated metal nanoparticles is obtainable in the absence of any additional reducing agent and/or in the absence of a surfactant.

In step (b), the mixing may be carried out at a temperature of from 20 °C to 120°C. For example, the temperature may be at least 20, 30, 40, 50, 60, 70, 80, 90, 100, or 110°C. The temperature may be less than 110, 100, 90, 80, 70, 60, 50, 40, or 30°C. In a preferred series of embodiments, the temperature is from 60 to 100°C.

The metal nanoparticles may be UV-stable. As used herein, the term “UV-stable” refers to the ability of the metal nanoparticles to withstand prolonged exposure to ultraviolet (UV) radiation over a defined period without sustaining permanent damage. It will be appreciated that UV radiation emitted from sunlight can be included in the definition. As such, the UV stability of the metal nanoparticles may correlate with their anticipated shelflife. The metal nanoparticles may be UV-stable for a period of one month or more. The metal nanoparticles may be UV-stable for at least 14 days. The metal nanoparticles may be UV-stable for at least 7 days. As such, the metal nanoparticles may be stable when exposed to sunlight for at least 7 days.

The antimicrobial compound may comprise an antibiotic.

The antimicrobial compound may comprise one or more compounds from a group comprising beta-lactams, carbapenems, ansamycins, macrolides, quinolones, tetracyclines, glycopeptides, aminoglycosides, chloramphenicol, lipopeptide, bispyridine, ebselen, pseudomonic acid, polychloro phenoxy phenol, hypochlorite anion, antimicrobial peptides, and quaternary amine compounds.

The antimicrobial compound may comprise one or more of: cefotetan, rifampicin, azithromycin, nalidixic acid, tetracycline, ceftazidime, minocycline, doxycycline, ampicillin, amoxicillin, ebselen, spectinomycin, cefaclor, aztreonam, doripenem, ertapenem, imipenem, meropenem, kanamycin, gentamicin, chloramphenicol, streptomycin, nitrofurantoin, octenidine, mupirocin, triclosan, sodium hypochlorite, benzalkonium chloride, and chlorohexidine.

37983926-1 The antimicrobial composition may comprise one or more of: a cream, paste, ointment, or medicament for topical or internal use.

According to a second aspect of the invention, there is provided a method of producing an antimicrobial composition as described herein. The method may comprise combining an antimicrobial compound with a population of metal nanoparticles. The population of metal nanoparticles may be in solution. The population of metal nanoparticles may be formed as described above with respect to the first aspect of the invention. The antimicrobial compound may be as described above.

According to a third aspect of the invention, there is provided a coating or treatment for a surface or substrate comprising an antimicrobial composition as described herein.

According to a fourth aspect of the invention, there is provided a medical device comprising a coating or treatment as described herein.

The medical device may comprise an absorbent article. The absorbent article may comprise at least one fibre, wherein the fibre is impregnated with the antimicrobial composition. The fibre may comprise cellulose and/or a derivative thereof, and optionally may comprise at least one other type of fibre blended therewith. In some embodiments, the at least one other type of fibre is: a gelling fibre based on alginate, cellulose and modified cellulose, modified chitosan, guar gum, carrageenan, pectin, starch, polyacrylates or copolymers thereof, polyethyleneoxides or polyacrylamides, or mixtures thereof; and/or a non-gelling fibre based on polyester, polyethylene, polyamide, cellulose, thermoplastic bicomponent fibres, glass fibres, or mixtures thereof. In one series of embodiments, the at least one other type of fibre comprises carboxymethyl cellulose (CMC) and a semi-synthetic fiber comprising regenerated cellulose, generically referred to as lyocell.

The medical device and/or absorbent article may comprise a wound care dressing and/or bandage. The absorbent article may comprise a woven or non-woven fabric.

In one series of embodiments, the medical device comprises an absorbent article which is impregnated with the composition as described herein.

37983926-1 The medical device may comprise a wearable item e.g. an apron, glove, mask and/or item of personal protective equipment.

Brief description of the figures

Embodiments of the present invention will now be described by way of example and with reference to the accompanying Figures, in which:

Figure 1 is a table showing the size distribution for a nanoparticle composition; Figure 2 is a table showing the results of a microdilution assay to investigate MIC and MBC;

Figure 3 is a table of the MIC and MBC of a range of antimicrobial compounds for E.colr,

Figure 4 is a table of the MIC and MBC of a range of antimicrobial compounds for S. aureus,

Figures 5a and 5b are tables of the MIC and MBC for a range of antimicrobial compositions.

Examples and Experiments

Example 1 - Silver nanoparticles synthesis

A silver nanoparticle solution was prepared as follows:

1 . 1459g of deionised (DI) water was placed in a first vessel, such as a 3L beaker. The first vessel was placed in a water bath set to the temperature in Table 1 .

2. 625g of polyvinylpyrrolidone (PVP 40 - Sigma Aldrich) was added to the beaker gradually with mixing to form a PVP solution. The molecular weight of the PVP was as follows: M_n=14,100 gmol’¹, M_w=35,300 gmol’¹, and M_w/M_n = 2.5.

3. In a second vessel, 0.86 moles of sodium hydroxide and 0.20 moles of sodium carbonate were dissolved in 1096g of DI water to form a sodium hydroxide and sodium carbonate solution. The second vessel was also placed in the water bath.

4. In a third vessel, a silver nitrate solution was prepared by adding 0.93 moles of AgNOs to 371g of DI water. The third vessel was also placed in the water bath.

37983926-1 5. The first, second, and third vessels were all maintained in the water bath until they had reached the temperature of the water bath.

6. Once the first, second and third vessels had reached 90°C, the sodium hydroxide and sodium carbonate solution in the second vessel was added to the PVP solution in the first vessel to form an intermediate solution.

7. Subsequently, the silver nitrate solution was added slowly to the intermediate solution and gently stirred. After all of the silver nitrate solution had been added, the reaction was left to run for 20 minutes with constant gentle stirring to yield the silver nanoparticle solution.

8. The silver nanoparticles in the silver nanoparticle solution had a coating comprising a polymer shell formed by the PVP. The nanoparticle concentration was determined to be 7800 ppm.

Example 2 - Silver nanoparticle size statistics

The silver nanoparticle solution formed in Example 1 was analysed by Scanning Transmission Electron Microscopy (STEM) using Imaged Fiji software to determine the size of the silver core and the PVP coating. The mean diameter of the silver core was found to be 18nm, and the mean PVP shell thickness was 73nm.

A further three samples of nanoparticle solution formed as per Example 1 were analysed in the same manner to determine the size distribution of the cores of the silver nanoparticles. The results are set out in the table of Figure 1 .

The samples A1 to A3 displayed a range of nanoparticle core sizes, with mean nanoparticle diameters of 11.85nm, 14.6nm and 11.94nm respectively. The standard deviations were quite broad, being 8.2nm, 10.76nm and 7.97nm respectively. The D10, D50 and D90 percentiles are shown in Table 1 below. In all three samples, less than 10% of the nanoparticles had core diameters of 4nm or less. Similarly, a significant number of larger nanoparticles were observed, spread broadly between the median and D90 percentile and even with particles as large as 50nm observed.

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Three commercially available nanoparticle solutions were obtained for comparative use. The data below is taken from the published Certificate of Analysis for each. Detailed size distribution data was not available, but the graphs provided in the CoA display no particles below 4nm or above 14nm for sample C1 , or below approximately 17nm or above approx. 40nm for sample C2.

Table 2

Example 3 - Determining MIC of nanoparticle solutions

The Minimum Inhibitory Concentrations (MIC) of the nanoparticles prepared according to Example 1 (sample A) according to and comparative samples B to D were investigated using a broth microdilution assay according to ELICAST best practice guidelines. MIC was determined against both E.coli 8739 and S. aureus 6538.

The results of the test are shown in Figure 2, with the MIC identified in the upper table as a percentage of the initial sample concentration and in ppm (equal to pg/ml) in the lower table. The MIC of Sample A was found to be 3.096 ppm with respect to E.coli, and 6.191 ppm for S. aureus, thereby showing extremely high efficacy against both gramnegative and gram-positive bacteria.

Surprisingly, the nanoparticle solution Sample A was found to be significantly more efficacious than any of the three commercially available solutions B to D. For E.coli, the MICs of samples B to D were 312.5ppm, 78.125ppm, and 18.750ppm respectively,

37983926-1 thereby showing an approximate six-fold improvement over the best performing comparative sample D. In comparison to sample B, the sample A was two orders of magnitude more effective at inhibiting E.coli growth.

Sample A was found to be inhibitory for E.coli at half the concentration required for inhibition of S. aureus. This finding was repeated for Samples C and D. It is possible that sample B could also be modestly more effective against E.coli compared to S. aureus, but this was not observed in this test and was not investigated further. Without wishing to be bound by theory, it is suspected that the difference in cell structure between grampositive and gram-negative bacteria accounts for the increase in efficacy with gramnegative E.coli.

Furthermore, and without wishing to be bound by theory, it is suspected that the greater efficacy of sample A compared to samples B to D may be due to the broader distribution of particles sizes within the solution. Sample B and C comprise nanoparticles with a mean diameter which is respectively smaller than and greater than the mean diameter of sample A. However, the standard deviations of 1 ,5nm and 4nm are less than a quarter and half respectively of the standard deviations observed in Samples A1 to A3 in Example 2. It is suspected that nanoparticles of different sizes have distinct targets on the bacterial cell.

Example 4 - Charge

The charge of the metal nanoparticles formed by the process described in Example 1 was measured and recorded in Table 3 below. The measurements were performed using an Anton-Paar Litesizer 500. Samples were diluted at a rate of 1 drop into 1ml of MilliCi® water and gently agitated by hand to homogenise the dispersion, then transferred into an omega zeta potential cuvette. Tests were run at 25°C with a 5 second temperature stabilisation time. Smoluchowski approximation was used as the approximation with a Henry factor of 1.5. Power was set to automatic with a maximum voltage set at 200V. Data quality was set to manual which performs 100 measurements. Each sample was tested 3 times and the mean average recorded in Table 3.

The value for sample C was obtained from the Certificate of Analysis for this product, and the range for sample D is obtained from the supplier’s marketing literature.

37983926-1 Nanoparticles produced by the process of Example 1 have been found to have a charge with a much lower magnitude than the commercially available product sample C.

Table 3

Without wishing to be bound by theory, it is believed that this reduction in charge is due to the relatively mild conditions in which the nanoparticles are formed in solution. In Example 1 , a relatively low concentration solution of NaOH and NaCCh is added to the PVP solution, and thus has a limited effect on charging the polymer. Conventional processes are known to use higher concentrations of base, or use citric acid to reduce the silver salts. This was confirmed via FTI R spectroscopy, which showed that the charge on the PVP did not significantly change during the production of the silver nanoparticles.

Furthermore, and without wishing to be bound by theory, it is suggested that the lower charge of the present nanoparticles in solution may be partially responsible for the greater efficacy of the present nanoparticles compared to the comparative products. It is believed that any repulsive effect due to the charges of the particles is minimised, and thus leads to better contact with bacterial cells. It is understood that the bacterial cell walls are negatively charged, and thus strongly negative nanoparticles are likely to be repelled, lessening their effectiveness. The combination of the near neutral charge is also suspected to directly relate the broader distribution of particle sizes as a result of increased agglomeration during formation, and both factors correlate with the improvement in efficacy.

Example 5 - Antimicrobial synergy

A further experiment was conducted to determine the effectiveness of the nanoparticles in combination with a range of antimicrobial compounds with respect to E.coli 8739 and S. aureus 6538. A broth microdilution assay according to ELICAST best practice guidelines was carried out to determine the MIC and the Minimum Bactericidal Concentration (MBC) for each antimicrobial compound in the tables in Figures 3 and 4.

37983926-1 The tests were then repeated by adding silver nanoparticles produced according to Example 1 (Sample A) to each of the test wells. The silver nanoparticles were added to provide a concentration of 25% of the MIC for the silver nanoparticles (as determined in Example 3) in each of the wells. For the E.coli tests, the concentration of silver nanoparticles in the wells was 0.77ppm, and in the S. aureus tests, the concentration of silver nanoparticles in each well was 1.55ppm. The silver nanoparticle solution concentration was selected to represent a minimally effective concentration whereby the addition of the silver nanoparticles alone would be expected to have no impact on the MBC or MIC of the antimicrobial compounds. The results of the test for E.coli were recorded in Figure 3, and for S. aureus in Figure 4.

Figure 3 shows that for 18 of the antimicrobial compounds tested on E.coli, a synergistic effect with at least a two-fold reduction in both the MIC and MBC was observed due to the addition of silver nanoparticles (Sample A) at a concentration of 0.77ppm (25% of the MIC). For the remaining 11 antimicrobial compounds, there was observed either no change or a modest improvement in one of the MIC or MBC.

The combination of silver nanoparticles to the antimicrobial compounds was found to be particularly effective for aminoglycosides. The addition of 0.77ppm of silver nanoparticle solution (Sample A) caused a 64-fold reduction in the MIC and MBC of spectinomycin on E.coli, a 66-fold to 128-fold reduction in MIC and MBC respectively for kanamycin, a 32-fold reduction in the MIC and MBC of streptomycin, and a 1300-fold reduction in the MIC and MBC for gentamicin.

In addition to the above, the addition of silver nanoparticles (Sample A) was particularly effective with ebselen. The addition of 0.77ppm of silver nanoparticle solution (Sample A) caused a reduction in the MIC and MBC for E.coli of approximately 5 orders of magnitude. This data was limited by the assay test, and could demonstrate even greater improvements if it could be tested and measured reliably at such low concentrations.

As shown in Figure 4, similar effects were observed for 17 antimicrobial compounds with respect to testing on S. aureus. Each antimicrobial compound was tested in combination with 1.55ppm of the silver nanoparticles produced according to Example 1.

37983926-1 Again, the combination of silver nanoparticles with aminoglycosides showed consistent improvements, albeit of a lesser magnitude than for E.coli. Ebselen again showed the best results, with a 2000-fold reduction in the MBC and 500-fold reduction in the MIC, a result limited primarily by the greater initial efficacy of ebselen against the gram-positive S. aureus.

The synergy observed between the antimicrobial compound and the silver nanoparticles was not consistent between gram-positive and gram-negative bacteria - without wishing to be bound by theory, it is suspected this variation in efficacy reflects the mode of attack for the respective antimicrobial compounds.

Example 6 - Comparison against commercial nanoparticles

The synergy of the antimicrobial compounds with silver nanoparticles was investigated further by comparison with a commercially available silver nanoparticle solution. A shortlist of antimicrobial compounds tested in Example 5 was selected and the microdilution assay was repeated with Sample C (NanoComposix, 25nm Silver Nanoparticles, PVP, Econix™). The process of Example 5 was repeated by adding Sample C to each of the test wells to provide a concentration of 25% of the MIC of Sample C.

Figure 5a shows the comparative results for the MIC and MBC analysis for 8 antimicrobial compounds with respect to E.coli. The table shows the MIC and MBC of the antimicrobial compounds in isolation, in combination with 0.77ppm of Sample A (as produced according to Example 1), and lastly in combination with 19.5ppm of Sample C. The concentrations of Sample A and Sample C used is 25% of their respective MIC’s in isolation, and represent a concentration which is expected to have no inhibitory effect.

Figure 5b shows the equivalent data with respect to S. aureus. Sufficient Sample C was added to the cells to produce a concentration of 39ppm, 25% of the MIC identified in Example 3.

For both E.coli and S. aureus, the addition of nanoparticle solution Sample C produced a reduction in the MIC and MBC for all of the antimicrobial compounds tested. Neither Sample A nor Sample C caused an improvement in the efficacy of nalidixic acid with

37983926-1 respect to S. aureus, although both were effective in reducing the MIC and MBC for E.coli. The combination of Sample C and Ebselen was found to be even more efficacious against E.coli than Sample A, although the final values were undetermined due to reaching the limit of the assay plate. This improvement was not repeated for S. aureus, wherein Sample A caused a greater reduction in the MIC and MBC for Ebselen.

The nanoparticle solution Sample C was found to demonstrate a similar synergy with the antimicrobial compounds tested, but of a lesser magnitude than Sample A. These results were achieved with a significantly greater concentration of Sample C in the test wells; 19.5 ppm and 39 ppm compared to 0.77ppm and 1.55ppm used for Sample A. Without wishing to be bound by theory, it is believed that the improved result with respect to Ebselen and E.coli for Sample C is a result of the increased concentration rather than due to the composition of Sample C.

37983926-1

Claims

CLAIMS:

1. An antimicrobial composition comprising: an antimicrobial compound; and a population of metal nanoparticles comprising a polymer coating, wherein the population of metal nanoparticles has a mean diameter from 5 nm to less than 20 nm, and the polymer coating comprises poly (N-vinylpyrrolidone).

2. The antimicrobial composition according to claim 1 , wherein the antimicrobial compound is an antibiotic.

3. The antimicrobial composition according to claim 1 or 2, further comprising a carrier, and optionally wherein the carrier comprises a fluid.

4. The antimicrobial composition according to any one of the preceding claims, wherein the antimicrobial compound and/or the population of metal nanoparticles are present at concentrations below their respective minimum bactericidal concentration (MBC); preferably wherein the antimicrobial compound and/or the population of metal nanoparticles have a concentration of less than or equal to 50% of their MBCs.

5. The antimicrobial composition according to any one of the preceding claims, wherein the antimicrobial compound and/or the population of metal nanoparticles are present at concentrations below their respective minimum inhibitory concentration (MIC); preferably wherein the antimicrobial compound and/or the population of metal nanoparticles have a concentration of less than 50% of their MIC.

6. The antimicrobial composition according to any one of the preceding claims, wherein the population of metal nanoparticles has a concentration of less than 15ppm.

7. The antimicrobial composition according to any one of the preceding claims, wherein the metal nanoparticles comprise silver and/or copper.

8. The antimicrobial composition according to any one of the preceding claims, wherein the polymer has a weight average molecular weight (M_w) of 20 to 80 kg/mol.

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9. The antimicrobial composition according to any one of the preceding claims, wherein the population of metal nanoparticles have a zeta potential with a magnitude of less than 10mV, preferably less than 5mV.

10. The antimicrobial composition according to any one of the preceding claims, wherein the population of metal nanoparticles has a mean diameter of 10 to 15nm, and/or wherein the population of metal nanoparticles has a standard deviation of 4 to 15nm.

11. The antimicrobial composition according to any one of the preceding claims, wherein the metal nanoparticles have a D10 particle size of 4nm or less, and/or wherein the metal nanoparticles have a D90 particle size of 15 nm or greater.

12. The antimicrobial composition according to any one of the preceding claims, wherein the metal nanoparticles are UV-stable; optionally, wherein the metal nanoparticles are UV-stable for at least one week, at least two weeks, or at least one month.

13. The antimicrobial composition according to any one of the preceding claims, wherein the population of silver nanoparticles is formed by: a) mixing a first aqueous alkaline solution with an aqueous polymer solution to form an aqueous alkaline polymer solution; and b) mixing the aqueous alkaline polymer solution with an aqueous solution of a metal salt to form a solution of polymer-coated metal nanoparticles.

14. The antimicrobial composition according to any one of the preceding claims, wherein the antimicrobial compound comprises one or more compounds from a group comprising beta-lactams, carbapenems, ansamycins, macrolides, quinolones, tetracyclines, glycopeptides, aminoglycosides, chloramphenicol, lipopeptide, bispyridine, ebselen, pseudomonic acid, polychloro phenoxy phenol, hypochlorite anion, antimicrobial peptides, and quaternary amine compound.

15. The antimicrobial composition according to any one of the preceding claims, wherein the antimicrobial compound comprises one or more of: cefotetan, rifampicin, azithromycin, nalidixic acid, tetracycline, ceftazidime, minocycline, doxycycline, ampicillin, amoxicillin, ebselen, spectinomycin, cefaclor, aztreonam, doripenem,

37983926-1 ertapenem, imipenem, meropenem, kanamycin, gentamicin, chloramphenicol, streptomycin, nitrofurantoin, octenidine, mupirocin, triclosan, sodium hypochlorite, benzalkonium chloride, and chlorohexidine.

16. The antimicrobial composition according to any one of the preceding claims, wherein the antimicrobial composition has a greater antimicrobial efficacy against gramnegative and/or gram-positive bacteria than the population of metal nanoparticles.

17. The antimicrobial composition according to any one of the preceding claims, wherein the antimicrobial compound and the population of metal nanoparticles comprising a polymer coating have a synergistic effect in that the MIC and/or MBC of the antimicrobial compound is reduced.

18. The antimicrobial composition according to any one of the preceding claims, wherein the composition comprises one or more of: a cream, paste, ointment, or medicament for topical or internal use.

19. A method of producing an antimicrobial composition according to any one of the preceding claims, comprising combining an antimicrobial compound with a population of silver nanoparticles.

20. A coating or treatment for a surface or substrate comprising an antimicrobial composition according to any one of claims 1 to 17.

21 . A medical device comprising a coating or treatment according to claim 20.

22. The medical device according to claim 21 , wherein the medical device is a wound care dressing or a bandage.

23. The medical device according to claim 21 or claim 22, wherein the medical device comprises an absorbent article, optionally wherein the absorbent article comprises at least one fibre impregnated with the antimicrobial composition.

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24. The medical device according to claim 23, wherein the at least one fibre comprises cellulose or a derivative thereof, the device optionally further comprising at least one other type of fibre blended therewith.

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