Aqueous antimicrobial composition, 16. use of a composition and method to clean and hygiene and / or disinfect a surface and provide residual inhibition against microbes

Description

1 / 36 “AQUEOUS ANTIMICROBIAL COMPOSITION, 16. USE OF A COMPOSITION AND METHOD FOR CLEANING AND SANITIZING AND / OR DISINFECTING A SURFACE AND PROVIDING RESIDUAL INHIBITION AGAINST MICROBES” Cross-referencing related requests

[001] The present invention relates to antiseptic formulations, in particular to antiseptic conditioning formulations, as well as to their uses and methods of manufacturing corresponding formulations. Field of invention

[002] There has been a growing focus on compositions that impart antimicrobial properties to products and surfaces, especially during the pandemic. Some pathogenic bacteria have evolved to become resistant to most, if not all, antibiotics currently available on the market. These drug-resistant bacteria pose a challenge to the healthcare sector. Furthermore, in the food industry, recalls related to bacterial infestation have become more frequent due to the inadequacy of current cleaning methods. Even consumers are seeking solutions to this problem with products that impart antibacterial properties to architectural paints and household and laundry care products.

[003] Before or after implantation, cleaning the implants and removing biofilms are important to prevent or reduce the risks of inflammation, respectively.

[004] In addition, antiseptic formulations are used to rinse parts of the body, for example, for mouthwashes, Petition 870260071585, dated 07 / 20 / 2026, page 7 / 42 2 / 36 wound treatment, skin disinfection and similar.

[005] To this end, several different antiseptic formulations are known to the expert. Typical antiseptic formulations include aqueous or at least partially aqueous formulations of antiseptic agents such as octenidine (1,1'-(decane-1,10-diyl)bis(noctylpyridin-4(1H)-imine)-hydrogen chloride), chlorhexidine (1,6bis(4-chlorophenylbiguanido)hexane) or polyhexanide (polyhexamethylene biguanide), but also strongly basic or acidic solutions or solutions based on inorganic antiseptic systems.

[006] Document JP-A-2007045732 provides a mild disinfectant solution for the skin and effective in inactivating Norovirus. The antiseptic solution contains 0.05-0.5% by weight of a polyhexamethylene biguanide-based compound and has a pH in the range of 9-12.

[007] Patent EP-A-1049763 refers to aqueous disinfectant solutions containing biguanide, containing an improved buffer system comprising a phosphate buffer and a borate buffer. Preferred embodiments include methods and compositions for simultaneous cleaning and disinfection of contact lenses. Summary of the invention

[008] Therefore, an objective of the present invention is to provide improved antiseptic formulations, in particular for conditioning, for example, implants, that utilize the lowest possible concentration of organic antiseptic compounds, that are compatible with the body and / or surfaces to which the formulations are applied, and that exhibit the greatest antiseptic effect. Petition 870260071585, dated 07 / 20 / 2026, p. 8 / 42 3 / 36 possible, while also achieving the highest possible biocompatibility.

[009] This objective is achieved by the subject matter claimed, more specifically by the compositions claimed, by the uses of such compositions and by the methods for their preparation.

[010] According to a first aspect of the present invention, it relates to an aqueous antimicrobial composition comprising or consisting of the following components: (a) polyhexanide (PHMB) or a salt thereof at a concentration in the range of 0.001-0.2% w / v; (b) an organic or inorganic buffer other than a) with at least one pK value in the range of 8.5 to 11.5 at a concentration in the range of 0.01 to 1 M; (c) additional additives at a concentration in the range of 0 to 20% w / v; (d) water; with the pH value of the composition being in the range of 8 to 11.5.

[011] Polyhexanide is polyhexamethylene biguanide, which typically has a weight-average molecular weight (Mw) in the range of 1500 to 4000 g / mol, preferably in the range of 2400 to 3000 g / mol.

[012] In addition, the polyhexanide (PHMB) of component (a) typically has a polydispersity index (PDI) in the range of 1.4 to 2.2, preferably in the range of 1.7 to 2.8.

[013] Typically, the starting material for the preparation of the corresponding composition is a polyhexanide salt, in particular the hydrochloride of Petition 870260071585, dated 07 / 20 / 2026, page 9 / 42 4 / 36 polyhexamethylene biguanide, and the percentage by weight is expressed relative to the total weight of polyhexamethylene biguanide hydrochloride.

[014] In general, when giving percentages in w / v, this means the weight of the corresponding substance measured in g per 100 mL, and the unit is g / cm3, which is equivalent to 1 kg / dm3, which is equivalent to 1000 kg / m3. 1% v / w therefore corresponds to 1 g / 100 mL, which is equivalent to 0.01 g / cm3.

[015] Furthermore, the pK values ​​provided are for water under standard laboratory conditions, i.e., 20 °C and 101 325 Pa.

[016] When pH values ​​are given, they are normally measured using calibrated pH / conductometers, after calibration, under standard laboratory conditions, for example, using a device of the 914 pH / conductometer type - Metrohm.

[017] The water in the composition is typically pure nano water, that is, water in accordance with the ASTM type I standard.

[018] In fact, it was unexpectedly discovered, as will be evidenced in the experimental section below, that the antibacterial efficacy of polyhexanide can be significantly increased by adjusting the pH in the claimed range and stabilizing it in that range by providing a corresponding buffer, preferably at the claimed concentration.

[019] Without relying on any scientific explanation, it appears, based on experimental evidence, that this is not merely a combination of the antibacterial effect due to the high pH value and the activity of polyhexanide, but that the Petition 870260071585, dated 07 / 20 / 2026, page 10 / 42 5 / 36 Experimental evidence demonstrates that there is a synergistic effect in the claimed concentration window for polyhexanide, allowing, to achieve the same disinfectant or preventive effect, a reduction in the concentration of polyhexanide or, using the same concentration, to achieve a more pronounced disinfectant or preventive effect.

[020] Preferably, the aqueous antimicrobial composition is ethanol-free or comprises less than 10% w / v or less than 5% w / v or less than 2% w / v of ethanol.

[021] Alternatively, and also preferably, the aqueous antimicrobial composition is free of linear or branched monohydric alkyl or arylalkyl alcohols with 1 to 9 or 1 to 6 carbon atoms (in particular, free of methanol, ethanol, propanol, 1-phenyl-1-propanol or a combination thereof) or comprises less than 10% w / v or less than 5% w / v or less than 2% w / v or less than 1% w / v of such linear or branched monohydric alkyl or arylalkyl alcohols.

[022] As defined above, the composition may consist of components (a) to (d). However, it may also comprise other components, in particular, according to a preferred embodiment, it further comprises component (e), preferably in this case, it then consists of components (a) to (e).

[023] Thus, according to a preferred embodiment, in addition to components (a) to (d), the composition also comprises the following component (preferably, this component (e) is the only additional component, so that the formulation consists of (a) to (e), also including means for adjusting the pH to the claimed value in the range of 8 Petition 870260071585, dated 07 / 20 / 2026, page 11 / 42 6 / 36 to 11.5, preferably in the form of NaOH): (e) at least one sugar alcohol with at least three carbon atoms.

[024] Sugar alcohols (also called polyhydric alcohols, polyalcohols, alditols or glycols) should be understood as organic compounds, typically derived from sugars, containing a hydroxyl group (-OH) attached to each carbon atom. They are usually white, water-soluble solids that can occur naturally or be produced industrially by the hydrogenation of sugars. Because they contain multiple -OH groups, they are classified as polyols. Sugar alcohols, according to this description, are systems of this type with at least three carbon atoms, preferably from 4 to 12 carbon atoms, particularly preferably from 4 to 6 carbon atoms or exactly 6 carbon atoms. Sugar alcohols may be added to influence the flavor and / or viscosity of the composition and, due to this effect, may synergistically influence the effectiveness of the other components, in particular component (a) combined with (b).

[025] Preferably, at least one sugar alcohol with at least three carbon atoms is present in the composition at a concentration of up to 75% w / v, preferably at a concentration of up to 70% w / v or in the range of 2-50% or 5-50% w / v or in the range of 3-40% or 10-40% w / v or 4-10% or 20-30%.

[026] Preferably, the sugar alcohol of component (e) is selected from the group consisting of glycerol, erythritol, threitol, arabitol, ribitol, mannitol, sorbitol, Petition 870260071585, dated 07 / 20 / 2026, page 12 / 42 7 / 36 xylitol, galactitol, fucitol, iditol, inositol, volemitol, isomalt, maltitol, lactitol, maltotriitol, maltotetraitol or a combination thereof.

[027] Correspondingly, according to a preferred embodiment, the concentration of polyhexanide (PHMB) of component (a) in the composition is in the range of 0.002-0.15% w / v or 0.002-0.1% w / v, preferably in the range of 0.005-0.07% w / v or 0.005-0.05% w / v, in particular in the range of 0.007-0.03% w / v or 0.007-0.02% w / v or 0.007-0.015% w / v. A particularly high synergistic effect of the high pH value and the presence of polyhexanide can be achieved if the pH and B concentration is in the range of 0.01% w / v, using polyhexanide hydrochloride as a starting material. Preferably, the organic or inorganic buffer of component (b) is selected from the following systems: • amino acid, in particular selected from the group consisting of alanine, glycine, asparagine, isoleucine, leucine, serine, threonine, valine or a combination thereof, in particular glycine; • bicarbonate buffer, in particular selected from the group consisting of: carbonate bicarbonate buffer, triethylammonium bicarbonate buffer; • borate buffer, in particular sodium borate buffer; • buffer (acetic acid) ethanolamine; or a combination or mixture thereof.

[028] In particular preference, the organic or inorganic buffer of component (b) is selected from the group consisting of short-chain amino acids without side chains that leads to a pKs value and with a pKs value in Petition 870260071585, dated 07 / 20 / 2026, page 13 / 42 8 / 36 claimed range, preferably glycine, alanine, valine, leucine, isoleucine or a combination or mixture thereof.

[029] In particular, the buffer is selected as a glycine buffer, in particular at a concentration in the range of 0.05-0.2 M. This range appears to be sufficient to stabilize the pH in the desired range and to cooperate optimally with the other components of the composition.

[030] In general, preferably, the organic or inorganic buffer of component (b) has at least one pK value in the range of 9-10.5, preferably in the range of 9.5-10.

[031] Also generally, the concentration of the organic or inorganic buffer of component (b) in the composition is in the range of 0.05-0.8 M, preferably in the range of 0.08-0.5 M, in particular in the range of 0.1-0.2 M.

[032] The pH value of the buffer of component (b) or, better yet, of the whole composition is, if necessary, adapted to the claimed pH value in the range of 8 to 11.5, preferably by the addition of a preferably strong base, preferably inorganic, said base being preferably selected from the group consisting of NaOH, KOH, Ca(OH)2, NaClO, KClO or a combination thereof. Typically, the concentration of this base in the composition is in the range of 0.001-0.07 M or 0.04-0.06 M or in the range of 0.045 M to 0.055 M, therefore, in particular, around 0.05 M.

[033] Preferably, the buffer (b) is glycine at a concentration in the range of 0.05-0.2 M, with the pH adjusted with an alkali metal hydroxide, in particular selected as Petition 870260071585, dated 07 / 20 / 2026, page 14 / 42 9 / 36 NaOH, KOH, or a mixture of these, at a concentration in the range of 0.04–0.06 M.

[034] It is even preferable that, as component (c), the additives be selected from the group: poloxamer, polyethylene glycol, flavorings, colorings, sugar alcohol, calcium donor or derivatives or combinations thereof, in a concentration in the range of 0-10% w / v.

[035] Particularly preferred is a composition in which the additives of component (c) are selected as: 1-3% w / v of poloxamer, preferably poloxamer with the structure EOn-POm-EOn, where n is in the range of 80-120 and e is in the range of 50-60; 1-3% w / v of polyethylene glycol and / or ethoxylated vegetable oil, in particular both, with ethoxylated castor oil being preferably used; 0.1-0.4% w / v of flavoring substances, preferably selected as flavoring oil; 2-10% w / v of sugar alcohol, preferably xylitol; 0.0-0.05% w / v or 0.01-0.05% w / v of calcium donor; Preferably, the composition has a pH in the range of 9.5-10.

[036] As indicated above, the composition may comprise additives / auxiliaries other than components (a) and (b) or (a) - (e).

[037] The additives of component (c) may be selected from the group consisting of: surfactants, anticorrosive agents, flavor modifiers (other than the sugar alcohols mentioned above), perfumes, colorants, stabilizers, thickeners (other than the sugar alcohols mentioned above), complexing agents, organic solvents, in particular alcohols (other than Petition 870260071585, dated 07 / 20 / 2026, page 15 / 42 10 / 36 sugar alcohols mentioned above), other disinfectants, preservatives, flavor modifiers (other than the sugar alcohols mentioned above), thickeners (other than the sugar alcohols mentioned above) or mixtures or combinations thereof.

[038] Examples of flavor modifiers (other than the sugar alcohols mentioned above) are one or a combination of: natural or synthetic fragrances, essential oils, tannic acid, menthol, sodium cyclamate, fructose, galactose, thymol. Flavor modifiers (other than the sugar alcohols mentioned above) are typically in a concentration in the range of 0.01 to 10% w / v, preferably in the range of 0.1 to 5%, relative to the aqueous antimicrobial composition.

[039] Examples of thickeners (other than the sugar alcohols mentioned above) are one or a combination of: gelatin, polyethylene glycol, for example, polyethylene glycol 1000, polyvinyl alcohol, silicon dioxide, starch, poloxamer (for example, 188, 407), carrageenan. Thickeners (other than the sugar alcohols mentioned above) are typically at a concentration in the range of 1 to 20% w / v relative to the aqueous antimicrobial composition.

[040] Surfactants can be ionic, non-ionic or amphoteric surfactants.

[041] The anionic surfactants of these additives are selected in particular from among alkyl sulfates, alkyl ether sulfates, alkyl sulfonates, alkylbenzyl sulfonates, α-olefin sulfonates, alkylamide sulfonates, alkyl polyether sulfates, alkylamidoether sulfates, alkyl monoglyceryl ether sulfates, alkyl monoglycerides Petition 870260071585, dated 07 / 20 / 2026, page 16 / 42 11 / 36 sulfates, alkyl monoglyceride sulfonates, alkyl succinates, alkyl sulfossuccinates, alkyl ether sulfossuccinates, alkyl sulfossuccinamates, alkyl amidosulfossuccinates; Alkyl sulfoacetates, alkyl phosphates, alkyl ether phosphates, alkyl ether carboxylates, alkyl amido-ether carboxylates, acyl lactylates, alkyl isethionates, acyl isethionates, carboxylate salts and surfactants derived from amino acids, such as N-alkyl amino acids, N-acyl amino acids, alkyl peptides and mixtures thereof.

[042] The amphoteric surfactants of these additives can be selected, in particular, from alkyl betaines; alkylamido betaines; alkylamido sultaines; alkyl mono and di-amphocarboxylates; amine oxides; and mixtures of these.

[043] The non-ionic surfactants of these additives, for example, may be selected from ethoxylated fatty alcohols, ethoxylated alkylphenols, ethoxylated Guerbet alcohols, ethoxylated vegetable oils (e.g., ethoxylated castor oil, for example, of the PEG-40 type; such compounds may, for example, assist in the dissolution of other components), polypropylene glycol, polyethylene glycol, propylene glycol and / or ethylene glycol block copolymers, ethoxylated sorbitan esters, sorbitan esters, alkyl polyglycosides, alkyl glucamides and mixtures thereof.

[044] The surfactant concentration of component (c) may be zero or greater than zero and be in the range of less than 10% w / v or less than 5% w / v relative to the total composition.

[045] The concentration of additives in component (c) may Petition 870260071585, dated 07 / 20 / 2026, p. 17 / 42 12 / 36 being in the range of less than 10% p / v or less than 5% p / v.

[046] As for the colorants in component (c), these are typically present in the composition at a concentration of less than 0.2% w / v, preferably in the range of less than 0.1% w / v, particularly preferably in the range of 0.001-0.05% w / v relative to the total composition.

[047] Preferably, additives other than thickeners (other than the sugar alcohols mentioned above) and flavor modifiers (other than the sugar alcohols mentioned above) are in the range of less than 0.2% w / v, preferably in the range of less than 0.1% w / v, particularly preferably in the range of 0.001-0.05% w / v, for example, in combination with a proportion of thickeners (other than the sugar alcohols mentioned above) in the range of 1-19.8% w / v in relation to the total composition.

[048] For some applications, it is preferable that there are no additives, so that the additive content is essentially 0.0 w / v in relation to the total composition.

[049] According to another preferred embodiment of the proposed composition, the composition has a calcium content as an additional component or as part of component (c) in the range of 1 to 5 mmol / L, preferably 2 to 3 mmol / L, which is the total, and the calcium may be in ionic form or in bound ionic form (in particular, bound to albumin).

[050] Typically, Ca2+ is introduced into the composition as CaCl2, calcium citrate, Ca(OH)2, calcium lactate, calcium carbonate or calcium phosphate, or combinations thereof.

[051] As noted above, an important characteristic of the composition is the pH value, which adapts to the most pKs Petition 870260071585, dated 07 / 20 / 2026, page 18 / 42 13 / 36 elevated from the polyhexanide system, which is in the range of 12. The pH value of the composition must be lower than this value.

[052] According to a preferred embodiment, the composition has a pH value in the range of 8 to 11, preferably in the range of 8.5 - 10.5, in particular in the range of 9.5 - 10. An optimal effect is achieved if the pH is around 9.8.

[053] According to a particularly preferred embodiment, the composition consists of (a) - (d) or (a) - (e), the polyhexanide (PHMB) of component (a) has a weight-average molecular weight (Mw) in the range of 2400-3000 g / mol, the polyhexanide (PHMB) of component (a) takes the form of polyhexamethylene biguanide hydrochloride and the concentration of the polyhexanide (PHMB) of component (a) in the composition is in the range of 0.007-0.015% w / v. In addition, the organic or inorganic buffer of component (b) is selected as glycine at a concentration of the organic or inorganic buffer of component (b) in the composition in the range of 0.1-0.2 M, and the concentration of the additional additives of component (c) is less than 0.01% w / v, preferably in the range of 0.001-0.01% w / v. According to a further aspect of the present invention, it relates to specific uses of such a composition.

[054] The therapeutic uses of such a composition (also formulated as Composition for use as...) include wound treatment (generic), including use as a cleansing solution, debridement solution, wound cleansing solution, wound treatment solution, wound treatment, in particular the promotion of Petition 870260071585, dated 07 / 20 / 2026, page 19 / 42 14 / 36 Wound healing in the initial phase of the oral cavity, preparation / conditioning of the wound / injured area, support for the purification phase during the primary phase of healing, cleaning and moistening of wounds, or combinations thereof.

[055] The therapeutic uses of such a composition (also formulated as Composition for use as...) also include wound irrigation (as a wound irrigation solution), uses for antimicrobial decontamination and infection prevention, infection prophylaxis and for reducing the antimicrobial load in the oral cavity.

[056] The therapeutic uses described may be for the treatment of humans or animals, and may be preventive or curative uses.

[057] According to this aspect of the invention, what is proposed is a composition, as defined above, for the treatment or prevention of bacterial infection or bacterial colonization, or both, in particular of the skin, wounds, preferably of the mouth.

[058] Or, in other words, the invention relates to the use of such a composition for the treatment or prevention of bacterial infection or bacterial colonization, or both, in particular of the skin, wounds, preferably of the mouth.

[059] According to a first preferred embodiment, the composition is for the treatment and / or prevention of patients during and / or after implantation, in particular during and / or after dental implantation, preferably in the form of a rinsing or washing composition or drip solution, including post-treatment treatment over days or even weeks after implantation. Petition 870260071585, dated 07 / 20 / 2026, page 20 / 42 15 / 36

[060] Non-therapeutic uses are intended for application to non-living objects, for example, hard surfaces or implants, and, if applied to humans or animals, are applied to healthy humans or animals without the need for therapy or prevention for purely disinfection reasons. According to this additional aspect of the present invention, it relates to the use of a composition, as detailed above, for the non-therapeutic disinfectant treatment of surfaces, in particular non-living surfaces, preferably by immersion, spraying, dripping, or for storage and / or preparation for implantation of devices, in particular implants, but also biomaterials for soft or hard tissue regeneration.

[061] According to a first preferred embodiment of this aspect, the composition is used to store an implant or a biomaterial, for example, for soft or hard tissue regeneration, in particular a dental implant, in immersion before use, or is packaged together or as a kit with an implant or biomaterial, for example, for soft or hard tissue regeneration, in particular a dental implant, so that the implant or biomaterial is immersed or moistened by the composition shortly before implantation.

[062] The present invention also relates to a method for cleaning, sanitizing and / or disinfecting a surface and providing residual inhibition against microbes, comprising the method: a) applying the composition, as detailed above, to a surface, article and / or substrate.

[063] In addition, the present invention relates to a method for preparing a composition as detailed. Petition 870260071585, dated 07 / 20 / 2026, page 21 / 42 16 / 36 above, preferably comprising the steps of providing an aqueous solution of sodium hydroxide and adding polyhexanide and buffer to it in the necessary proportions and adding additives, if necessary, before or after, followed by mixing.

[064] The compositions can be stored in containers that do not require refrigeration, preferably glass or plastic containers, and such containers can be combined with objects to which the corresponding compositions must be applied before use. For example, combined packaging is possible that includes one container for the composition, as described above, and another separate container for the corresponding object, for example, an implant. Furthermore, such packaging can be structured so that the container with the composition can be opened by manipulation, so that the composition flows into the packaging compartment of the object for direct wetting and / or immersion.

[065] Other embodiments of the invention are presented in the dependent claims. Brief description of the drawings

[066] Preferred embodiments of the invention are described below with reference to the drawings, which are intended to illustrate preferred embodiments of the invention and not to limit them. In the drawings:

[067] Figure 1 shows the results of initial in vitro tests with biofilm models of the synergistic antimicrobial effect of test reagents in alkaline buffers for low concentrations of PHMB;

[068] Figure 2 shows the results of initial tests. Petition 870260071585, dated 07 / 20 / 2026, p. 22 / 42 17 / 36 in vitro with biofilm models of the synergistic antimicrobial effect of test reagents in alkaline buffers for medium concentrations of PHMB;

[069] Figure 3 shows the results of initial in vitro tests with biofilm models of the synergistic antimicrobial effect of test reagents in alkaline buffers for higher concentrations of PHMB;

[070] Fig. 4 (a) shows the results of tests on denser and older biofilm models of the synergistic antimicrobial effect of the test reagents in alkaline buffers for higher concentrations of PHMB, and (b) shows the behavior of Streptococcus sanguinis cultured in neutral PBS (pH 7.2) or in alkaline T buffer (pH 9.7) or in alkaline T buffer enriched with 0.015% PHMB;

[071] Fig. 5 shows the results of the biocompatibility tests of the different alkaline buffers;

[072] Fig. 6 shows the results of the biocompatibility tests of PHMB and CHX for different concentrations;

[073] Fig. 7 shows the cell viability of primary human cells after treatment with various antiseptics at concentrations similar to those used in products currently available on the market;

[074] Fig. 8 shows the effect of additive ethanol at 10, 20 and 50% on the cellular viability of human primary cells. Detailed description of the invention Preparation of test reagents: As test reagents, different solutions of chlorhexidine and polyhexamethylene biguanide (PHMB) were prepared with different concentrations and buffers. The solution of Petition 870260071585, dated 07 / 20 / 2026, page 23 / 42 18 / 36 Chlorhexidine digluconate (CHX, 20% w / v in water, CAS 18472-51-0) was purchased from Sigma-Aldrich, Switzerland (article number C9394). Glycine was purchased as a solid from Honeywell Fluka, Switzerland (CAS: 56-40-6). Polyhexamethylene biguanide was purchased as a solid from Biosynth, United Kingdom (article number FP76704, batch number: B20V06202). As a stock solution, a 10% (w / v) PHMB solution in water was prepared. All PHMB test reagents were prepared based on the 10% stock solution.

[075] The buffers, as per Table 1 below, were prepared using a 0.05 M NaOH solution and adding solid glycine in an amount that would lead to the indicated glycine concentration, with pH measured using a pH electrode (914 pH / Metrohm Conductometers, 3-point calibration).

[076] Table 1: Solutions and Buffers used (all concentrations are given for the final composition) Solution / Buffer pH 0.05 M NaOH 12.51 0.05 M NaOH / 0.05 M Glycine 11.06 0.05 M NaOH / 0.1 M Glycine 9.78 0.05 M NaOH / 0.2 M Glycine 9.27 0.05 M NaOH / 0.4 M Glycine 8.94

[077] The polyhexanide test reagents (polyhexamethylene biguanide hydrochloride, PHMB, CAS number 28757-47-3, also 32289-58-0) were prepared from a 10% PHMB stock solution in ultrapure water (weight by volume, ASTM Type 1, as supplied by a Sartorius Arium Pro Water System, e.g., ultrapure, as per ASTM D1193-06 (2018)) and using the solutions and buffers presented in Table 1 and tested in different Petition 870260071585, dated 07 / 20 / 2026, page 24 / 42 19 / 36 experiments, according to Table 2 below.

[078] Table 2: PHMB solutions used in the experiments (all concentrations are given for the final composition) Experiment No. Concentration (% by weight by volume) PHMB Buffer Observation 1 0.05% • Water • 0.05 M NaOH / 0.05 M Glycine • 0.05 M NaOH / 0.1 M Glycine • 0.05 M NaOH / 0.2 M Glycine • 0.05 M NaOH / 0.4 M Glycine Initial biofilm formation 2 0.015% • Water • 0.05 M NaOH • 0.05 M NaOH / 0.1 M Glycine 3 0.01% • Water • 0.05 M NaOH / 0.05 M Glycine • 0.05 M NaOH / 0.2 M Glycine • 0.05 M NaOH / 0.4 M Glycine 4 0.0002% • Water • 0.05 M NaOH • 0.05 M NaOH / 0. 0.05 M Glycine • 0.05 M NaOH / 0.2 M Glycine 5 0.05 & 0.1% • Water • 0.05 M NaOH / 0.1 M Glycine Dense biofilm Petition 870260071585, dated 07 / 20 / 2026, page 25 / 42 20 / 36

[079] Table 3: Initial biofilm formation Concentration CHX Buffer 0.06% • Water • 0.05 M NaOH / 0.4 M Glycine Initial biofilm formation 0.12% • Water

[080] In addition, a 0.9% (weight by volume) NaCl solution in ultrapure water was used as a state-of-the-art rinsing solution.

[081] In addition to the solutions mentioned above, used in the in vitro experiments reported below, the following final formulation, presented in Table 3a, for in vivo use, was prepared and considered stable and effective:

[082] Table 3a: PHMB solution used for in vivo tests (measured pH = 9.8) Substance Concentration (w / v) Water >70% NaOH 0.2% Glycerol 5-10% Poloxamer 1-3% Glycine 0.75% (0.1 M) Preservative 0.1% PHMB (Polyhexanide) 0.01-0.1% Xylitol 10% PEG dye 1-2% Aromatic oil 0.3% Calcium donor 0.03%

[083] Another formulation for in vivo use was prepared and found to be stable and effective, and is shown in Table 3b below. Petition 870260071585, dated 07 / 20 / 2026, page 26 / 42 21 / 36

[084] Table 3b: PHMB solution used for in vivo tests (pH = 9.8 as measured) Substance Concentration (w / v) Water >70% NaOH 0.2% Poloxamer 1-3% Glycine 0.75% (0.1M) PHMB (Polyhexanide) 0.01-0.1% Xylitol 4% PEG 1-2.5% Aromatic oil 0.2%

[085] Determination of the antimicrobial effect of various test reagents in an in vitro biofilm model:

[086] To test the antimicrobial effect of various test reagents, an oral strain of Streptococcus sanguinis, acquired from ATCC, was used. All experiments began with the preparation of an overnight culture of S. sanguinis in brain-heart infusion (BHI) medium at 37°C under aerobic conditions. After 16 hours, the bacterial suspension was centrifuged at 3000 rpm for 5 minutes. The supernatant was removed and the pellet was resuspended in 1 ml of phosphate-buffered saline (PBS, Sigma-Aldrich, Switzerland). The bacterial suspension was adjusted with PBS to achieve an optical density of 0.5, equivalent to 10⁸ CFU / ml. For the antimicrobial experiments, an initial inoculum of 10³ CFU / ml in BHI medium was used.

[087] To model the initial formation of biofilm after the fixation of a dental implant, round titanium discs (16 mm diameter, supplied by Thommen Medical AG) were cleaned, autoclaved and placed on a 24 plate. Petition 870260071585, dated 07 / 20 / 2026, page 27 / 42 22 / 36 wells.

[088] In a subsequent step, these titanium discs were preconditioned with the test reagents or BHI / 0.9% NaCl medium (control group).

[089] The conditioned titanium discs were subsequently incubated with 1 ml / well of bacterial suspension (103 CFU / ml) in BHI medium.

[090] After 24 hours, the BHI medium was removed and the titanium discs were washed with the test reagents for 10 minutes at 37°C and 70 rpm.

[091] To quantify the biofilm mass on the titanium discs, staining with crystal violet was performed. For this purpose, the test reagents were removed with a pipette and 300 μL / well of crystal violet solution (0.1% in ultrapure water, acquired from Sigma-Aldrich Switzerland) were added and incubated for 10 minutes at room temperature.

[092] After the incubation time, the titanium discs were washed four times with distilled water (600 μl / well) and transferred to a new 24-well plate. To release the crystal violet staining from the titanium discs, 1 ml / well of acetic acid (33%) was added and incubated for 5 minutes at room temperature and 70 rpm. Duplicates of 100 μ^ were transferred from each well to a new 96-well plate and the absorbance at 590 nm was measured using a BioTek Epoch2 microplate reader.

[093] To evaluate the antimicrobial effect of test reagents not only on the initial formation of biofilm, but also on an older and denser biofilm, an experiment Petition 870260071585, dated 07 / 20 / 2026, page 28 / 42 23 / 36 similar to that described in the section above was repeated using an initial inoculum of 106 CFU / ml and an incubation time of 72 hours.

[094] To study the effect of alkaline pH on bacterial growth, a bacterial suspension of S. sanguinis with an optical density (600 nm) of 0.2 was prepared in PBS medium with pH 7.2 or in T buffer (0.05 M NaOH / 0.1 M glycine, 0.03% CaCl2) with pH 9.7 ± addition of 0.015% PHMB. Then, 100 µl of the bacterial suspension were transferred to a 96-well plate and placed in a pre-warmed (37 °C) Biotek reader. The optical density at 600 nm was measured at 2-hour intervals over 24 hours to assess bacterial behavior in varying buffers. Cell culture and cytocompatibility testing of test reagents and alkaline buffers:

[095] Mouse fibroblasts (cell line L929) were used in this study as the recommended cell line for cytotoxicity tests, according to the EN ISO 10993-5 standard.

[096] Fibroblasts were acquired from ATCC and cultured in Dulbecco's modified Eagle medium with high glucose content, supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin (Sigma-Aldrich, Switzerland) in T75 flasks at 37°C and 5% CO2. For cytotoxicity experiments, L929 cells were seeded in 96-well cell culture plates, 100 μl / well, at a density of 1x10⁶ cells / mL to achieve approximately 80% confluence after 24 hours. The following day, 100 μl of fresh medium enriched with varying concentrations (0.001, 0.005, 0.01, 0.05 and 0.1%) of Petition 870260071585, dated 07 / 20 / 2026, page 29 / 42 24 / 36 CHX or PHMB.

[097] As a control, the cells were incubated with fresh medium without antiseptic agent.

[098] Six wells replicated by concentration were tested.

[099] After 60 minutes of incubation at 37°C, the medium was removed and the wells were washed once with 100 µl of fresh medium.

[100] Next, the cells were incubated with 100 pl of fresh medium containing 0.015 mg / ml of AlamarBlue dye, for 2 hours at 37°C.

[101] Finally, 100 pL of the medium were removed from each well and transferred to a new 96-well plate, and the absorbance was measured using a BioTek microplate reader (test wavelength: 570 nm; reference wavelength: 630 nm).

[102] To test the cytocompatibility of alkaline buffers, a similar experiment was performed. However, the alkaline buffers were incubated for 10 minutes instead of 60 minutes.

[103] The following alkaline buffers were tested: a) NaOH 0.025 M, NaOH 0.025 M / glycine 0.05 M, NaOH 0.025 M / glycine 0.05 M / CaCl2O, 0.03% (weight by volume in the final composition); b) 0.05 M NaOH, 0.05 M NaOH / 0.05 M glycine, 0.05 M NaOH / 0.05 M glycine / 0.03% CaCl2 (weight by volume in the final composition); c) 0.1 M NaOH, 0.05 M NaOH / 0.1 M glycine, 0.05 M NaOH / 0.1 M glycine / 0.03% CaCl2 (weight by volume in the final composition); Petition 870260071585, dated 07 / 20 / 2026, page 30 / 42 25 / 36 d) 0.05 M NaOH / 0.1 M glycine; e) 0.05 M NaOH / 0.1 M glycine / 0.03% CaCl2 (weight by volume in the final composition). Human primary cells and the effect of antiseptic substances and ethanol on cell vitality: Human periodontal ligament fibroblasts (HPDLF) were used in this study as a test system, more closely simulating the in vivo situation. Fibroblasts were acquired from ATCC and cultured in ScienCell FM medium, supplemented with 2% fetal bovine serum, 1% fibroblast growth factor, and 1% penicillin-streptomycin (Sigma-Aldrich, Switzerland) in T75 flasks at 37°C and 5% CO2.

[104] To study the effect of antiseptic substances, at concentrations currently used in commercial products, the viability of HPDLF cells was evaluated. For this purpose, cells were seeded in 24-well cell culture plates, 1 ml / well, at a density of 3x104 cells / mL to achieve approximately 80% confluence after 48 hours. In a subsequent step, 300 µl of buffer T. (0.05 M NaOH / 0.1 M glycine, 0.03% CaCl2) enriched with varying concentrations (0.06 and 0.12%) of CHX or (0.015, 0.025, 0.05 and 0.15%) PHMB were added. As a control, cells were incubated with buffer T. without antiseptic agent. Three replicated wells per concentration were tested. After 1 minute of incubation at 37°C, the supernatant was removed and the wells were washed once with 600 µL of fresh medium. Then, the cells were incubated with 600 µL of fresh medium containing 0.015 mg / ml of Alamar Blue dye for 4 hours at 37°C. Finally, 100 µL Petition 870260071585, dated 07 / 20 / 2026, pp. 31 / 42 26 / 36 of the medium was removed from each well and transferred to a new 96-well plate, and absorbance was measured using a BioTek microplate reader (test wavelength: 570 nm; reference wavelength: 630 nm).

[105] To evaluate the effect of ethanol on cell viability, HPDLF cells were seeded in 96-well cell culture plates, 100 µL / well, at a density of 6x103 cells / mL to achieve approximately 80% confluence after 24 hours. The following day, 100 µL of fresh medium contaminated with varying concentrations (10, 20 and 50% ethanol. In addition, the developed T buffer was contaminated with 10, 20, and 50% ethanol. As a control, cells were incubated with fresh medium without ethanol. Three wells replicated by concentration were tested. After 5 minutes of incubation at 37°C, the medium was removed and the wells were washed once with 100 µl of fresh medium. Then, the cells were incubated with 100 µl of fresh medium containing 0.015 mg / ml of AlamarBlue dye for 2 hours at 37°C. Finally, 100 µl of medium were removed from each well and transferred to a new 96-well plate, and the absorbance was measured using a BioTek microplate reader (test wavelength: 570 nm; reference wavelength: 630 nm). Results and discussion Compatibility of test reagents in alkaline buffers: Polyhexamethylene biguanide hydrochloride (Polyhexanide, PHMB) is a chemical biocide.There are six different types of PHMB products identified, which have combinations of amine, guanidine, and cyanoguanidine terminal groups. Petition 870260071585, dated 07 / 20 / 2026, pages 32 / 42 27 / 36

[106] The antibacterial activity of PHMB depends on the molecular structure.

[107] The minimum requirements are met by more than 2 biguanide groups and 5 to 7 methylene groups as spacers.

[108] The biguanide groups of PHMB are strong bases and monoprotonated at a pH value of 7 (pKal d 2-3; pKa2 d 10.5-11.5), resulting in a positively charged polycation in each biguanide group.

[109] It is believed that the positively charged fractions bind to the negatively charged phosphate groups of phospholipids in the cell walls of bacteria, leading to increased fluidity, permeability and loss of integrity, followed by the death of the organism.

[110] PHMB can be seen as virtually detoxified CHX, since the molecular structure of PHMB monomers closely resembles the structure of CHX molecules, except for the terminal NH group of CHX, which consists of 4-chloroaniline (4-CA). Similar to PHMB, the biguanide moiety of CHX with a pKa d 10.8 makes it a strong base.

[111] In general, inorganic bases such as sodium hydroxide lead to the precipitation of PHMB and CHX if pH > pka value, for example, for 0.05 M NaOH with pH 12.7.

[112] Sodium hydroxide, on the other hand, is known to induce an alkaline saponification reaction with the phospholipid membrane of bacteria, causing membrane rupture and ultimately cell death.

[113] Surprisingly, there is a synergistic effect between PHMB and sodium hydroxide, leading to increased bacterial death or allowing a reduced active concentration. Petition 870260071585, dated 07 / 20 / 2026, pp. 33 / 42 28 / 36 to achieve the same effect.

[114] To implement this in practice, however, it is also necessary to formulate a buffer that prevents PHMB precipitation, but still allows sufficient availability of hydroxyl ions to achieve alkaline saponification.

[115] Therefore, different buffer compositions were tested for PHMB and CHX precipitation, as shown in Table 1.

[116] The results of these precipitation tests were as follows (precipitation determined by visual inspection / visible turbidity, pH measured): NaOH 0.05 M / glycine 0.05 M / CHX 0.06%, pH 11.06: Precipitation NaOH 0.05 M / glycine 0.1 M / CHX 0.06%, pH 9.9: No precipitation NaOH 0.05 M / PHMB 0.05%, pH 12.51: Precipitation NaOH 0.05 M / glycine 0.05 M / PHMB 0.05%, pH 11.06: No precipitation NaOH 0.05 M / glycine 0.1 M / PHMB 0.05%, pH 9.9: No precipitation.

[117] The 0.06% and 0.12% w / v concentrations of CHX were based on common mouthwash solutions (e.g., products with Chlorhexamed), and 0.05% PHMB was based on mouthwash solutions with concentrations in the range of 0.04% w / v.

[118] Based on the pKa value of 10.8 for the strongest base within the CHX molecule, precipitation occurs using a 0.05 M NaOH / 0.05 M glycine buffer with a pH of 11.2 (pH > pKa). Petition 870260071585, dated 07 / 20 / 2026, pages 34 / 42 29 / 36

[119] Increasing the glycine concentration to 0.1 M and consequently decreasing the pH does not cause CHX precipitation.

[120] Unlike CHX, the pKa value of the strongest base within the PHMB molecule is in the range of 10.5 to 11.5. Thus, no PHMB precipitation was observed using a 0.05 M NaOH / 0.05 M glycine buffer (pH 11.06). However, a 0.05 M NaOH solution (pH 12.5) without the addition of glycine also induces PHMB precipitation.

[121] Therefore, it can be concluded that, for PHMB, a minimum of 0.05 M glycine is appropriate. As the pH of 11.06 for a 0.05 M NaOH / 0.05 M glycine solution is close to the pKa value of the strongest PHMB base, for some applications it is advisable to use a 0.05 M NaOH solution enriched with 0.1 M glycine, i.e., a solution with a pH in the range of 10 or below. Synergistic antimicrobial effect of test reagents in alkaline buffers:

[122] In a study by Ojan Assadian et al., 2011 (Assadian O, Wehse K, Hübner NO, Koburger T, Bagel S, Jethon F, Kramer A. Minimum inhibitory concentration (MIC) and minimum microbicidal concentration (MMC) of polyhexanide and triclosan against antibiotic-sensitive and antibiotic-resistant strains of Staphylococcus aureus and Escherichia coli. GMS Krankenhhyg Interdiszip. 2011;6(1)), a minimum inhibitory concentration (MIC) in the range of 1-2 pg / ml (0.002% w / v) was reported for Streptococcus aureus and Escherichia coli treated with PHMB solutions.

[123] To detect a possible synergistic effect of PHMB and an alkaline buffer, an initial biofilm of Petition 870260071585, dated 07 / 20 / 2026, pages 35 / 42 30 / 36 Streptococcus sanguinis was cultured on titanium discs for 24 hours before the discs were rinsed with different PHMB solutions containing 0.002% PHMB, similar to the reported MIC. As a control, the biofilm was cultured in BHI medium.

[124] The PHMB solution (0.002% in nanopure water) did not result in a reduction in bacterial quantity, suggesting that 0.0002% is below the MIC for Streptococcus sanguinis in this assay.

[125] However, if PHMB (0.002%) was prepared with 0.05 M NaOH / 0.05 M glycine buffer at pH 11.06, a significant reduction in absorbance and therefore in bacterial quantity was observed.

[126] Similar results were obtained for PHMB (0.002%) prepared with 0.05 M NaOH / 0.2 M glycine buffer at pH 9.27.

[127] In contrast, PHMB (0.002% in distilled water) added to a 0.05 M NaOH solution with a pH of 12.5 (no visible precipitation, probably due to the low concentration of PHMB used) did not result in a reduced bacterial count, since the pH > pKa did not allow the formation of a positively charged PHMB molecule.

[128] Figure 1 graphically illustrates the absorbance results for the situation of low PHMB concentrations, with higher absorbance indicating less effect on the biofilm.

[129] In a subsequent experiment, a higher concentration of PHMB (0.01% in ultrapure water) was tested on the initial biofilm formation of Streptococcus sanguinis, grown on titanium discs. Petition 870260071585, dated 07 / 20 / 2026, pp. 36 / 42 31 / 36

[130] In this experiment, a synergistic effect of 0.01% PHMB and NaOH / glycine buffer was also observed.

[131] The best results, in terms of reducing the amount of bacteria, were obtained using a 0.05 M NaOH / 0.05 M glycine buffer (pH 11.06) and a 0.05 M NaOH / 0.2 M glycine buffer (pH 9.27). Higher concentrations of glycine, for example, 0.4 M (pH 8.9), did not lead to a significant reduction in the amount of bacteria, compared to lower concentrations of glycine, for example, 0.05 M-0.2 M.

[132] Figure 2 graphically illustrates the absorbance results for the situation of low PHMB concentrations; a higher absorbance indicates a smaller effect on the Biofilm.

[133] As can be seen from the results illustrated in Fig. 3, in the initial biofilm model, no pronounced synergistic effect can be observed if 0.05% PHMB is added to alkaline buffers.

[134] To verify whether the initial in vitro biofilm model, configured with a low concentration initial bacterial inoculum and short incubation time, can prevent a possible synergistic effect of a higher concentration of PHMB, for example, 0.05% in alkaline buffers, the experiment was repeated using a denser and older biofilm model (initial inoculum of 106 CFU / ml, incubation time of 72 hours). Using the adapted experimental design, a synergistic effect of 0.05% PHMB in 0.05 M NaOH / 0.1 M glycine buffer was obtained (see results in Fig. 4a).

[135] To assess the effect of pH and substance Petition 870260071585, dated 07 / 20 / 2026, pp. 37 / 42 32 / 36 antiseptic on bacterial behavior, a bacterial suspension of S. sanguinis was measured for 24 hours at an optical density of 600 nm and 37 °C. Incubation in neutral PBS demonstrated significant growth of S. sanguinis over time, while cultivation of S. sanguinis in T. buffer with pH 9.7 showed a significant inhibitory effect. The addition of 0.015% PHMB to alkaline T. buffer with pH 9.7 resulted in the elimination of bacteria over time, see the results in Figure 4b. Biocompatibility of test reagents:

[136] To test the biocompatibility of the different alkaline buffers, murine L929 fibroblasts were used.

[137] The cells were seeded in a 96-well plate at a density of 1x106 cells / ml and cultured for 24 hours before being treated with the different alkaline buffers for 30 minutes.

[138] As a control, the cells were grown in fresh medium.

[139] After the incubation period, metabolic activity, used as an indirect marker of cytotoxicity, was measured with an AlamarBlue assay. Vital cells were able to transform the dye by redox reaction into a fluorescent product. Therefore, the highest metabolic activity was observed in the control. A decrease in metabolic activity is associated with possible toxic effects.

[140] Incubation with pure sodium hydroxide (0.025 M, 0.05 M and 0.1 M) led to a significant reduction in metabolic activity. Petition 870260071585, dated 07 / 20 / 2026, pp. 38 / 42 33 / 36

[141] In addition, alkaline buffers containing 0.05 M NaOH / 0.05 M glycine (pH 11.06) and 0.1 M NaOH / 0.05 M glycine showed evident toxic effects.

[142] Compared to the buffers mentioned above, alkaline buffers containing 0.025 M NaOH / 0.05 M glycine or 0.05 M NaOH / 0.1 M glycine showed metabolic activities comparable to the control.

[143] Furthermore, it was observed that the addition of 0.03% CaCl2 to alkaline buffers may have additional beneficial effects on metabolic activity.

[144] Figure 5 graphically illustrates the results of the biocompatibility tests of the different alkaline buffers. The pH values ​​of the samples in this figure are presented in Table 4.

[145] Table 4: pH values ​​measured from the samples illustrated in Figure 5 Sample pH 0.025 M NaOH 12.15 0.025 M NaOH / 0.05 M glycine 10.42 0.025 M NaOH / 0.05 M glycine / 0.03% CaCl2 10.42 0.05 M NaOH 12.51 0.05 M NaOH / 0.05 M glycine 11.06 0.05 M NaOH / 0.05 M glycine / 0.03% CaCl2 11.06 0.1 M NaOH 12.71 0.1 M NaOH / 0.05 M glycine 11.24 0.1 M NaOH / 0.05 M glycine / 0.03% CaCl2 11.24 0.05 M NaOH / 0.1 M glycine 9. 78 0.05 M NaOH / 0.1 M glycine / 0.03% CaCl2 9, 78

[146] To evaluate the biocompatibility of PHMB compared to that of CHX, murine L929 fibroblasts were treated for 60 minutes with different concentrations of PHMB and CHX, respectively, supplemented in the medium.

[147] After incubation with antiseptic agents, metabolic activity was measured using an assay Petition 870260071585, dated 07 / 20 / 2026, pp. 39 / 42 34 / 36 AlamarBlue. The data were normalized to the control (cells in fresh medium) and therefore adjusted for 100% metabolic activity. To compare the two antiseptic agents, the IC50 value, representing the concentration that allows 50% cell survival (horizontal line in bold), was calculated for each. The calculation revealed an IC50 value of 0.1% for PHMB and 0.01% for CHX, see Figure 6.

[148] To evaluate the cell viability of the antiseptic agents PHMB and CHX, human primary cells (HPDLF) were treated for 1 minute with different concentrations of PHMB and CHX, supplemented in buffer T. After incubation with the antiseptic agents, metabolic activity was measured using an AlamarBlue assay (after 4 h). Low cell viability was observed for cells treated with CHX at the tested concentrations of 0.06 and 0.12% in buffer T. For PHMB, low cell viability was obtained for cells treated with PHMB at a concentration of 0.15%, while PHMB at 0.05% led only to reduced cell viability. PHMB at 0.025% and 0.015% revealed cell viability in the range of the buffer T control. Cells treated with the negative control (5% DMSO) showed low cell viability. See Figure 7.

[149] Note that this is valid for antiseptic applications. If the application is more anti-inflammatory (for example, in the form to be used against mucositis), higher concentrations of PHMB, up to 0.2%, can also be used. The additional advantage over the use of CHX is that, unlike the use of CHX-based formulations, there is no staining effect on the teeth with the application of Petition 870260071585, dated 07 / 20 / 2026, pages 40 / 42 35 / 36 formulation of PHMB, and PHMB exhibits greater biocompatibility than CHX.

[150] Finally, the influence of ethanol on cell vitality was investigated. HPDLF cells were cultured in FM cell culture medium as a control. The effect of adding ethanol was studied by adding ethanol to 10, 20, and 50% of the cell culture medium or by adding the developed T. buffer and corresponding amounts of ethanol. After 5 minutes of incubation with the different test substances, cell vitality was measured by the AlamarrBlue assay, performed as described previously. Ethanol added to the cell culture medium at concentrations of 20 and 50% showed a significant decrease in cell vitality. Ethanol added to T. buffer also caused a negative effect on cell vitality, see Figure 8. Key findings:

[151] Some of the main findings from the experimental evidence can be summarized as follows: • A particularly pronounced synergistic effect of PHMB and alkaline buffers was obtained for PHMB concentrations in the range of 0.002% to 0.04% in an initial biofilm formation model (initial inoculum of 103 CFU / ml, incubation time of 24 hours) with Streptococcus sanguinis. In experiments using an initial biofilm formation configuration, little synergistic effect of PHMB and alkaline buffers was observed with PHMB concentrations > 0.05%. • Alkaline buffers containing 0.05 M NaOH and < 0.05 M glycine tend to result in a very high pH and pH > pKa, resulting Petition 870260071585, dated 07 / 20 / 2026, pp. 41 / 42 36 / 36 showed lower antimicrobial activity. Buffers containing 0.05 M NaOH and glycine > 0.4 M also resulted in lower antimicrobial activity. The best performance in terms of synergistic effect (antimicrobial activity) was achieved with buffers containing 0.05 M NaOH and 0.05 M-0.2 M glycine (pH range 11.06 to 9.8). • A synergistic effect with PHMB (0.05%) and an alkaline buffer (0.05 M NaOH / 0.1 M glycine) was demonstrated in an in vitro biofilm model with a higher bacterial load (106 CFU / ml) and a longer incubation time (72 hours). • Biocompatibility tests with murine L929 fibroblasts demonstrated good tolerance to alkaline buffers containing 0.025 M NaOH / 0.05 M glycine or 0.05 M NaOH / 0.1 M glycine + 0.03% CaCl2 (pH 9.78). This indicates that alkaline buffers composed of NaOH / glycine with pH < 10 can be considered biocompatible. • Biocompatibility tests with murine L929 fibroblasts and PHMB and CHX revealed the following IC50 values: PHMB < 0.1%, CHX < 0.01%. These data indicate greater biocompatibility for PHMB than for CHX. • Cellular vitality after exposure to the proposed formulations is very good compared to other products; the presence of ethanol negatively affects cellular vitality, therefore, alcoholic components should be avoided. • The proposal provides an ideal balance between high antimicrobial activity and biocompatibility for oral tissue cells. Petition 870260071585, dated 07 / 20 / 2026, page 42 / 42

Claims

1 / 7 CLAIMS 1. An aqueous antimicrobial composition, characterized in that it comprises or consists of the following components: (a) polyhexanide (PHMB) or a salt thereof at a concentration in the range of 0.001-0.2% w / v; (b) an organic or inorganic buffer other than a) with at least a pK value in the range of 8.5-11.5 at a concentration in the range of 0.01-1 M; (c) additional additives at a concentration in the range of 0.20% w / v; (d) water; wherein the pH value of the composition is in the range of 8 to 11.

5.

2. Composition according to claim 1, characterized in that the polyhexanide (PHMB) of component (a) has a weight-average molecular weight Mw in the range of 1500-4000 g / mol, preferably in the range of 2400-3000 g / mol; and / or in that the polyhexanide (PHMB) of component (a) has a polydispersity index (PDI) in the range of 1.4 to 2.2, preferably in the range of 1.7 to 2.8; and / or in that the polyhexanide (PHMB) of component (a) takes the form of polyhexamethylene biguanide hydrochloride.

3. Composition, according to any of the preceding claims, characterized in that the concentration of polyhexanide (PHMB) of component (a) in the composition is in the range of 0.002 to 0.15% w / v or 0.002 to 0.1% w / v, preferably in the range of 0.005 to 0.07% w / v or 0.005 to 0.05% w / v, in particular in the range of 0.007 to 0.03% w / v or 0.007 to 0.02% w / v or 0.007 to 0.015% w / v. Petition 870250067189, dated 01 / 08 / 2025, page 49 / 64 2 / 7 4. Composition, according to any of the preceding claims, characterized in that the organic or inorganic buffer of component (b) is selected from the group consisting of: amino acid, in particular selected from the group consisting of alanine, glycine, asparagine, isoleucine, leucine, serine, threonine, valine or a combination thereof, in particular glycine; bicarbonate buffer, in particular selected from the group consisting of: carbonate bicarbonate buffer, triethylammonium bicarbonate buffer; borate buffer, in particular sodium borate buffer; ethanolamine (acetic acid) buffer; or a combination or mixture thereof, wherein, preferably, the organic or inorganic buffer of component (b) is selected from the group consisting of: glycine, alanine, valine, leucine, isoleucine or a combination or mixture thereof.

5. Composition, according to any of the preceding claims, characterized in that the organic or inorganic buffer of component (b) has at least one pK value in the range of 9 to 10.5, preferably in the range of 9.5 to 10.

6. Composition, according to any of the preceding claims, characterized in that the concentration of the organic or inorganic buffer of component (b) in the composition is in the range of 0.05-0.8 M, preferably in the range of 0.08-0.5 M, in particular in the range of 0.1-0.2 M.

7. Composition, according to any of the preceding claims, characterized in that the additives of component (c) are selected from the group that Petition 870250067189, dated 01 / 08 / 2025, page. 50 / 64 3 / 7 consists of: surfactants, including ionic and non-ionic surfactants, thickeners, compatibilizers, in particular anionic surfactants, in particular selected from alkyl sulfates, alkyl ether sulfates, alkyl sulfonates, alkyl benzyl sulfonates, α-olefin sulfonates, alkylamide sulfonates, alkyl polyether sulfates, alkyl amido-ether sulfates, alkyl monoglyceryl ether sulfates, alkyl monoglyceride sulfates, alkyl monoglyceride sulfonates, alkyl succinates, alkyl sulfosuccinates, alkyl ether sulfosuccinates, alkyl sulfosuccinamates, alkyl amidosulfosuccinates;alkyl sulfoacetates, alkyl ether phosphates, alkyl ether carboxylates, alkyl amide ether carboxylates, alkyl lactylates, alkyl isethionates, alkyl isethionates, carboxylate salts, and surfactants derived from amino acids, such as N-alkyl amino acids, N-acyl amino acids, alkyl peptides and mixtures thereof; amphoteric surfactants, in particular selected from alkyl betaines; alkylamido betaines; alkylamido sultains; mono-alkyl and di-amphocarboxylates; amine oxides; and mixtures thereof, non-ionic surfactants, thickeners or compatibilizers, in particular selected from ethoxylated fatty alcohols, ethoxylated alkylphenols, ethoxylated Guerbet alcohols, ethoxylated vegetable oils, polypropylene glycol, polyethylene glycol, propylene glycol and / or ethylene glycol block copolymers, ethoxylated sorbitan esters, sorbitan esters, alkyl polyglycosides, alkyl glucamides and mixtures thereof;Petition 870250067189, dated 01 / 08 / 2025, page 51 / 64 4 / 7 anti-corrosive agents, perfumes, dyes, stabilizers, complexing agents, organic solvents, in particular alcohols, disinfectants, preservatives or mixtures or combinations thereof.; 8. Composition, according to any of the preceding claims, characterized in that the concentration of the additives of component (c) is in the range of less than 10% w / v or less than 5% w / v, wherein the colorants, as part of component (c), are preferably present in a concentration in the range of less than 0.2% w / v, preferably less than 0.1% w / v, particularly preferably in the range of 0.001-0.05% w / v or 0.0 w / v.

9. Composition, according to any of the preceding claims, characterized in that the composition has a pH value in the range of 8 to 11, preferably in the range of 8.5 to 10.5, in particular in the range of 9.5 to 10.

10. Composition, according to any of the preceding claims, characterized in that the polyhexanide (PHMB) of component (a) has a weight-average molecular weight (Mw) in the range of 2,400 to 3,000 g / mol, wherein the polyhexanide (PHMB) of component (a) takes the form of polyhexamethylene biguanide hydrochloride and wherein the concentration of the polyhexanide (PHMB) of component (a) in the composition is in the range of 0.007 to 0.015% w / v; wherein the organic or inorganic buffer of component (b) is selected as glycine at a concentration of the organic or inorganic buffer of component (b) in the composition in the range of 0.1 to 0.2 M; Petition 870250067189, dated 01 / 08 / 2025, p. 52 / 64 5 / 7 and wherein the concentration of additional additives of component (c) is less than 0.01% w / v, preferably in the range of 0.001 to 0.01% w / v.

11. A composition, according to any of the preceding claims, characterized by being for treating or preventing bacterial infection or bacterial colonization, or both, particularly of the skin, wounds, preferably of the mouth.

12. Composition, according to claim 11, characterized by treating and / or preventing problems in patients, during and / or after an implant, in particular, during and / or after dental implantation, preferably in the form of a rinsing or washing composition or drip solution, including post-treatment during the weeks following implantation.

13. Composition, according to any of the preceding claims, characterized in that, in addition to components (a) to (d), the composition further comprises the following component: (e) at least one sugar alcohol having at least three carbon atoms, preferably in a concentration of up to 75% w / v, preferably in a concentration of up to 70% w / v or in the range of 5 to 50% w / v or in the range of 10 to 40% w / v or 20 to 30%, and preferably said sugar alcohol of component (e) is selected from the group consisting of glycerol, erythritol, threitol, arabitol, ribitol, mannitol, sorbitol, xylitol, galactitol, fucitol, iditol, inositol, volemitol, isomalt, maltitol, lactitol, maltotriitol, maltotetraitol or a combination thereof. Petition 870250067189, dated 01 / 08 / 2025, pp. 53 / 64 6 / 7 14. Composition, according to any of the preceding claims, characterized in that it is free of ethanol or contains less than 10% w / v, less than 5% w / v or less than 2% w / v of ethanol.

15. Composition, according to any of the preceding claims, characterized in that it is free of linear or branched monohydric alkyl or arylalkyl alcohols with 1 to 9 or 1 to 6 carbon atoms, in particular, free of methanol, ethanol, propanol, 1-phenyl-1-propanol or a combination thereof, or contains less than 10% w / v, less than 5% w / v, less than 2% w / v or less than 1% w / v of such linear or branched monohydric alkyl or arylalkyl alcohols.

16. Use of a composition, according to any one of claims 1 to 15, characterized by being directed to the non-therapeutic disinfectant treatment of surfaces, in particular non-living surfaces or biomaterials, preferably by immersion, spraying, dripping, or for storage and / or preparation for implantation of devices, in particular implants or biomaterials, in particular for soft or hard tissue regeneration.

17. Use, according to claim 16, characterized in that the composition is used to store an implant or biomaterial, in particular for soft or hard tissue regeneration, in particular a dental implant, in immersion before use, or is packaged together or as a kit with an implant or biomaterial, in particular for soft or hard tissue regeneration, in particular a dental implant, so that the implant or material is immersed or moistened by the composition shortly before implantation.

18. Method for cleaning and sanitizing and / or disinfecting a surface and providing residual inhibition against microbes, the method being characterized by comprising: a) applying the composition of any of the preceding claims to a surface, article and / or substrate. Petition 870250067189, dated 01 / 08 / 2025, pp. 55 / 64

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