Composition

Abstract

The present invention relates to compositions for use in the treatment or prevention of pathogenic infections, rosacea, eczema and psoriasis in humans or animals. The compositions of the invention are also useful for wound healing in humans or animals and for killing or inactivating surface viruses.

Description

[Technical Field]

[0001] This invention relates to compositions for use in the treatment or prevention of pathogenic infections in humans or animals, such as bacterial, fungal, and / or viral infections. Specific examples of such infections include pathogenic infections within / on the surface of wounds, bacterial foot disease, impetigo, and tinea. The compositions of this invention can be used at very low concentrations and have been shown to be non-irritating to human or animal skin. The compositions of this invention are used as disinfectants.

[0002] The compositions of the present invention are also useful for the treatment or prevention of rosacea, eczema and / or psoriasis in humans or animals.

[0003] Furthermore, the compositions of the present invention can be used to heal wounds in humans or animals, or to heal wounds while also treating or preventing pathogenic infections within or on the surface of the wound.

[0004] Furthermore, the present invention relates to a method for killing or inactivating viruses on a surface by applying the composition of the present invention to the surface. Examples of such viruses include norovirus and coronavirus. [Background technology]

[0005] Pathogenic infections, such as bacterial infections, can develop into serious infections that, if left untreated, can progress to life-threatening conditions such as sepsis. Traditional treatments for pathogenic infections include antibiotics and disinfectants such as chlorohexidine. However, medical professionals are trying to avoid the use of antibiotics as much as possible to prevent the development of antibiotic resistance. Disinfectants are used to destroy or inhibit pathogenic microorganisms on tissue surfaces without causing harmful effects on living tissue. However, many known disinfectants cause adverse effects such as allergic reactions, and many pathogenic infections have become resistant to treatment with known disinfectants. Therefore, there is a need for novel, antibiotic-free agents to treat pathogenic infections.

[0006] Pathogenic infections within or on the surface of wounds are particularly problematic because pathogens can lead to local tissue damage and hinder wound healing (leading to chronic wounds such as venous ulcers in the lower extremities). Therefore, effective treatment of pathogenic infections within or on the surface of wounds may be essential for wound healing. The environment of a wound, especially a chronic wound, is very different from that of a hard surface or intact skin. A wound contains a complex biological mixture consisting of damaged tissue, cells necessary for healing, protein-rich exudate, and microorganisms. Therefore, while disinfectants may be used to cleanse hard surfaces or intact skin, these disinfectants are often inactivated in the wound environment. Thus, while a drug may prove to be an excellent disinfectant, this does not mean that a disinfectant will be a useful disinfectant for wounds. In fact, many agents that can be used to disinfect woundless skin before medical treatment (e.g., alcohol before venipuncture) can be proven ineffective as wound disinfectants; that is, they are not useful for treating pathogenic infections within or on the surface of a wound. The same is true even for some known disinfectants; that is, some known disinfectants are inactivated in the wound environment.

[0007] Chronic wounds are even more difficult to treat because they typically contain biofilms; in fact, current estimates suggest that approximately 60% to 100% of chronic, non-healing wounds contain biofilms. The presence of biofilms in chronic wounds means that the wound often appears to heal but eventually becomes congested again. Biofilms are complex structures invisible to the naked eye, composed of mixed strains of pathogens, and are typically formed when certain types of microorganisms on the wound surface adhere to them, followed by the secretion of viscous substances. Pathogens within biofilms are considerably more difficult to eradicate with known treatments such as antibiotics and disinfectants than those within free-floating cells. The complex structures within biofilms create a stressful microenvironment and introduce spatio-physiological heterogeneity into the population within the biofilm. This likely explains why viable-but-non-culturable (VBNC) cells and persister cells are frequently found within biofilms. Current recommendations for managing chronic wounds include biofilm removal by physical debridement or "aggressive" cleansing, acknowledging that these actions are not effective in removing the entire biofilm and therefore need to be repeated.

[0008] Given the significantly high morbidity and mortality rates associated with biofilm-related infections, effective treatments for pathogenic infections within and on the wound surface, particularly in chronic wounds containing biofilms, are desirable. Therefore, the development of novel strategies to inhibit and treat biofilms is crucial.

[0009] Surfaces often come into contact with potentially harmful viruses, providing an environment for them and creating a breeding ground. It is common practice to clean surfaces with agents that work to reduce and / or destroy potentially harmful viruses. Cleaning surfaces in this manner is beneficial to human and animal health because it inhibits the spread of viruses and reduces the chances of subjects(s) becoming infected with viral infections through contact with potentially harmful viruses. Particularly important surfaces are those in hospital and veterinary environments, such as those in veterinary or hospital operating rooms. Therefore, general cleaning is a cornerstone of infection control practices in healthcare settings. However, many of the agents currently used, at the concentrations they are used, pose a serious exposure risk to human / animal health, meaning that the clinical areas being decontaminated often have to be closed for several days. Similarly, agents are often insufficiently effective against some of the viruses causing common problems, lack any residual antimicrobial activity, and are prone to recontamination. Therefore, there is a need for effective and safe agents that can be used in a variety of situations for cleaning and decontaminating virus-containing surfaces.

[0010] Two specific viruses of particular importance are norovirus and coronavirus. It is estimated that there are over 267 million norovirus infections worldwide each year (including 23 million in the US and up to 1 million in the UK). Norovirus outbreaks cost National Health Insurance over £184 million (based on 2002-2003 figures) in hospitalizations, and an estimated $2 billion annually in the US. While infection is self-limiting, the symptoms are unpleasant, and patients continue to shed numerous viral particles for days after becoming ill. Human coronavirus 229E is a human respiratory pathogen that causes mild respiratory illness in healthy individuals, but can cause serious illness in immunocompromised patients and those with multiple sclerosis. Human coronavirus 229E is a hazard group 2 pathogenic virus that is used as a surrogate for coronaviruses that cause more serious respiratory illnesses such as severe acute respiratory disease syndrome (SARS) and Middle East respiratory syndrome (MERS), requiring higher levels of isolation facilities.

[0011] International Publication No. 2015 / 028806 describes a cleaning solution for disinfecting surfaces and water supply, and an aqueous mixture containing the cleaning solution. The cleaning solution and aqueous mixture in International Publication No. 2015 / 028806 are compositions according to the present invention. The cleaning solution and aqueous mixture in International Publication No. 2015 / 028806 are also described as useful for disinfecting skin areas. Disinfecting skin areas means that the cleaning solution / aqueous mixture kills or inhibits microorganisms on intact skin surfaces within minutes, for example, to ensure a “clean” area before the skin barrier is destroyed by medical or surgical intervention. The World Health Organization defines disinfectants as chemical agents applied during cleaning of animate objects and substances to destroy or inhibit pathogenic microorganisms.

[0012] General definition The term "comprising" encompasses "including" and "consisting," for example, a composition "comprising" X may consist solely of X or may include something additional, such as X + Y. As used herein, the term "comprising" also encompasses "consisting essentially of," for example, a composition "comprising" X may consist of X and any other components that do not substantially affect the essential characteristics of the composition.

[0013] The numerical term "approximately" is optional and can mean, for example, x + 10%.

[0014] The term "animal" refers to mammals or birds.

[0015] "Pathogenic infection of the skin" as described herein refers to an infection of the skin caused by a pathogen invading from the normal skin barrier due to the destruction of skin integrity caused by injury (e.g., wound) or disease (e.g., impetigo) that impairs the skin barrier. This invasion leads to an inflammatory response, and if the pathogen is sufficiently pathogenic, it can lead to widespread tissue infection, such as sepsis. Treatment of such pathogenic infections of the skin is distinguished from "disinfecting" an area of ​​skin, as disinfection involves killing or inhibiting microorganisms on the surface of intact skin for a short period of time (i.e., simply sterilizing the surface of intact skin) to ensure a "clean" area, for example, before the skin barrier is destroyed by medical or surgical intervention.

[0016] The term "skin" includes the skin on the surface and inside of the mouth.

[0017] The term "wound" means an injury to living tissue (e.g., skin, muscle, etc.) caused by a cut, blow, or other impact, typically where the skin is cut or damaged. Examples of wounds include burns, ulcers, cuts, abrasions (friction), and punctures.

[0018] The term "healing" with respect to a wound means accelerating the closure / repair of the wound.

[0019] "Accelerating the closure of a wound" means that the wound closes / repairs faster when the composition of the present invention is used than it would in the absence of the composition of the present invention.

[0020] The term "chronic wound" means a wound that does not progress in an orderly and timely manner to the healing stage and does not show significant progress towards healing within 30 days.

[0021] "Inactivating" a virus means that the virus enters a dormant state.

Prior Art Documents

Patent Documents

[0022]

Patent Document 1

Brief Description of the Drawings

[0023] The present invention will now be described by way of example only, with reference to the accompanying drawings. [Figure 1-1] ~ [Figure 1-3] Images before and after the treatment of impetigo using the composition of the present invention. [Figure 2a] ~ [Figure 2j] Images before or after multiple spectra showing the treatment and healing of Gram-positive and Gram-negative viable but non-culturable (VBNC) biofilm wounds using the composition of the present invention. [Figure 3a] ~ [Figure 3b] These are before and after images of a dog bite wound treated with the composition of the present invention. [Figure 4a] ~ [Figure 4b] These are before and after images of a horse leg wound treated with the composition of the present invention. [Figure 5a] ~ [Figure 5c] These are images of a nail fungal infection before and after treatment using the composition of the present invention. [Figure 6] These are the results of a skin irritation test related to the composition of the present invention. [Figure 7a] ~ [Figure 7h] This figure shows human coronavirus 229E killed and inactivated using the composition of the present invention. [Figure 8a] ~ [Figure 8c] This figure shows the killing and inactivation of murine norovirus using the composition of the present invention. [Figure 9] The present invention provides the MICs for various Gram-positive and Gram-negative bacteria using the composition of this invention. [Figure 10a-1] ~ [Figure 10b-2] The present invention relates to the treatment of psoriasis and eczema on human skin using the composition of the present invention. [Overview of the Initiative]

[0024] These drawings are further explained in the section "Examples".

[0025] This application is written section by section to aid in readability. However, it is not intended that each section be read in isolation. On the contrary, unless otherwise specified, each section should be read in cross-reference with the other sections, that is, in consideration of the entire application as a whole. Unless explicitly stated, no artificial separation of embodiments is intended. Any of the compositions of the present invention described herein may be used in any of the uses described herein.

[0026] If, for example, the composition of the present invention is stated to contain at least 99.2% by weight of water, then the other components in the composition should be selected such that their total amount does not exceed 0.8% by weight, i.e., the total weight percentages in the composition are 100%. Selecting the amounts of components in such a technically reasonable manner is within the scope of the skills of those skilled in the art. [Means for solving the problem]

[0027] In a first aspect, compositions of the present invention are provided for use in the treatment or prevention of pathogenic infections in humans or animals, preferably in humans, namely compositions comprising benzalkonium halide; didecyldimethylammonium halide; polyhexamethylene biguanide salt; bronopol; and p-chloro-m-cresol. Thus, the compositions of the present invention are used as disinfectants and are useful in the treatment or prevention of diseases / conditions associated with, i.e., caused by, pathogenic infections, preferably of the skin, muscle tissue, connective tissue and / or skeletal tissue. Specific examples include pathogenic infections of the inside / surface of wounds and impetigo.

[0028] As shown in the examples, the compositions of the present invention are not only capable of treating or preventing a variety of pathogenic infections, but are also useful for treating or preventing Gram-positive and Gram-negative pathogenic infections containing biofilms with VBNC cells, such as chronic wounds (e.g., chronic burns). The compositions of the present invention can also be used at low concentrations that are non-irritating to the skin.

[0029] In a second aspect, compositions of the present invention are provided for use in the treatment or prevention of rosacea, psoriasis and / or eczema in humans or animals, preferably in humans. Thus, compositions of the present invention can further treat or prevent other skin conditions, namely skin conditions that are not or not caused by pathogenic infections.

[0030] In a third aspect, compositions of the present invention are provided for use in healing wounds in humans or animals, preferably in humans, i.e., in accelerating the time it takes for a wound to close. Compositions of the present invention can also heal wounds while simultaneously treating or preventing pathogenic infections inside / on the wound surface, i.e., killing pathogens such as bacteria inside / on the wound surface.

[0031] A fourth aspect provides a method for killing or inactivating viruses on a surface, comprising the step of applying a composition of the present invention to the surface. Preferred viruses are norovirus and coronavirus. It has been found that the composition of the present invention only needs to be used at low concentrations to achieve this effect. Furthermore, the composition of the present invention can be pre-coated to a surface, i.e., the surface can be pre-treated for up to 48 hours to prevent viral infection on the surface.

[0032] In a fifth aspect, the present invention relates to a composition of the present invention for use as a pharmaceutical. In a sixth aspect, the present invention relates to a composition of the present invention for use as a disinfectant, that is, a composition of the present invention is used as a disinfectant on the body of a human or animal, preferably on the surface of a human body. [Modes for carrying out the invention]

[0033] The present invention relates to compositions comprising benzalkonium halide; didecyldimethylammonium halide; polyhexamethylene biguanide salt; bronopol; and p-chloro-m-cresol for use in the treatment or prevention of pathogenic infections in humans or animals. Accordingly, the compositions of the present invention are useful for the treatment or prevention of diseases / conditions associated with, i.e., caused by, pathogenic infections.

[0034] The compositions of the present invention are also useful for treating or preventing rosacea, psoriasis and / or eczema, accelerating wound healing, i.e., the time it takes for wounds to close, and killing or inactivating viruses on surfaces.

[0035] Composition of the present invention The compositions of the present invention may be any of the compositions disclosed in International Publication No. 2015 / 028806 (which is incorporated herein by reference), specifically in pages 4, line 11 to 7, line 2, pages 8, line 7 to 13, line 31, and in claims 1 to 16, that is, the compositions of the present invention may be any of the “cleaning liquids” or “aqueous mixtures” disclosed in International Publication No. 2015 / 028806. The compositions of the present invention may also be any one of the compositions defined herein.

[0036] The composition of the present invention comprises benzalkonium halide, didecyldimethylammonium halide, polyhexamethylene biguanide salt, bronopol, and p-chloro-m-cresol.

[0037] In one embodiment, the composition of the present invention comprises 0.04 to 0.2% by weight of benzalkonium halide; 0.04 to 0.2% by weight of decyldimethylammonium halide; 0.04 to 0.2% by weight of polyhexamethylene biguanide salt; 0.01 to 0.06% by weight of bronopol; and 0.0005 to 0.005% by weight of p-chloro-m-cresol.

[0038] In one embodiment, benzalkonium halide is benzalkonium chloride, and / or didecyldimethylammonium halide is didecyldimethylammonium chloride, and / or polyhexamethylene biguanide salt is polyhexamethylene biguanide hydrochloride. In a preferred embodiment, benzalkonium halide is benzalkonium chloride, didecyldimethylammonium halide is didecyldimethylammonium chloride, and polyhexamethylene biguanide salt is polyhexamethylene biguanide hydrochloride.

[0039] This composition may further contain a solvent. Preferably, the solvent is ethanol. The solvent is preferably present in the composition at a concentration of 0.05 to 0.3% by weight.

[0040] The composition may further contain alkylene glycol. Suitable alkylene glycols that can be used in the composition of the present invention include ethylene glycol, propylene glycol, diethylene glycol, block copolymers of ethylene oxide and propylene oxide, any other alkylene glycol formed by combining alkylene oxide, and / or any combination of alkylene glycols. In a preferred embodiment, the alkylene glycol is ethylene glycol. The alkylene glycol is preferably present in the composition at a concentration of 0.01 to 0.07% by weight.

[0041] Alkylene glycol may be substituted in whole or in part with other alkylene glycols, such as ethylene glycol, propylene glycol, diethylene glycol, block copolymers of ethylene oxide and propylene oxide (e.g., various types of Pluronic® sold by BASF®), any other alkylene glycol formed together with alkylene oxide, and / or any combination of alkylene glycols.

[0042] In a preferred embodiment, the composition further comprises ethanol and ethylene glycol, preferably 0.05 to 0.3% by weight of ethanol and 0.01 to 0.07% by weight of ethylene glycol.

[0043] Preferably, the composition of the present invention does not contain one or more siloxanes.

[0044] Preferably, the compositions of the present invention do not contain antibiotics. In fact, the advantage of the compositions of the present invention is that they can rapidly and at low concentrations kill a wide range of pathogenic infections without the need to use antibiotics, which are often ineffective. Similarly, the development of antimicrobial resistance can be avoided by using the compositions of the present invention instead of antibiotics.

[0045] In one embodiment, the composition of the present invention further comprises water. In one embodiment, the composition comprises at least 83.9% by weight of water, or at least 96% by weight of water, or at least 97% by weight of water, or at least 98% by weight of water, or at least 98.7% by weight of water, or at least 99.0% by weight of water, or at least 99.2% by weight of water, or at least 99.5% by weight of water, or at least 99.6% by weight of water, or at least 99.7% by weight of water, or at least 99.76% by weight of water. Preferably, the composition comprises at least 99.2% by weight of water or at least 99.76% by weight of water.

[0046] As shown in the examples, the inventors have found that the compositions of the present invention can treat various pathogenic infections and skin conditions such as psoriasis, heal wounds, and kill or inactivate viruses even when used at surprisingly low concentrations, i.e., when the water content in the composition is high, for example, at least 99.2% by weight of water or even at least 99.76% by weight of water.

[0047] Furthermore, as described in Example 7, an additional advantage of the compositions of the present invention is that they can be effectively used even at very low concentrations to be non-cytotoxic. For example, when the water content in the composition is higher than about 99.5% by weight, e.g., higher than 99.76% by weight, the compositions of the present invention are non-cytotoxic and, even more surprisingly, still capable of treating pathogenic infections, psoriasis, eczema and rosacea, and healing wounds in humans or animals, as described herein. The combination of non-cytotoxicity and maintaining the effectiveness for the uses described herein is remarkable.

[0048] Therefore, in one embodiment, the composition of the present invention comprises at least 99.76% by weight of water and is for use in the treatment of impetigo. In one embodiment, the composition of the present invention comprises at least 99.76% by weight of water and is for use in the treatment of nail fungal infection. In one embodiment, the composition of the present invention comprises at least 99.76% by weight of water and is for use in the treatment of bacterial infection inside / on the surface of a wound. In one embodiment, the composition of the present invention comprises at least 99.76% by weight of water and is for use in wound healing. In one embodiment, the composition of the present invention comprises at least 99.76% by weight of water and is for use in the treatment of psoriasis.

[0049] In one embodiment, the composition of the present invention is Benzalkonium halide Decyldimethylammonium halogenate Polyhexamethylene biguanide salt Bronopol p-chloro-m-cresol solvent Alkylene glycol, and water Includes.

[0050] In another embodiment, the composition of the present invention is 0.04-0.2% by weight of benzalkonium halide 0.04-0.2% by weight of decyldimethylammonium halide 0.04-0.2% by weight of polyhexamethylene biguanide salt 0.01-0.06% by weight of Bronopol 0.0005 to 0.005% by weight of p-chloro-m-cresol 0.05-0.3% by weight of solvent 0.01 to 0.07% by weight of alkylene glycol Includes, The remainder is water.

[0051] In a preferred embodiment, the composition of the present invention is Benzalkonium chloride Decyldimethylammonium chloride Polyhexamethylene biguanide hydrochloride Bronopol p-chloro-m-cresol ethanol Ethylene glycol, and water Includes.

[0052] In a particularly preferred embodiment, the composition of the present invention is 0.04-0.2% by weight of benzalkonium chloride 0.04-0.2% by weight of decyldimethylammonium chloride 0.04-0.2% by weight of polyhexamethylene biguanide hydrochloride 0.01-0.06% by weight of Bronopol 0.0005 to 0.005% by weight of p-chloro-m-cresol 0.05-0.3% by weight of ethanol 0.01 to 0.07% by weight of ethylene glycol Includes, The remainder is water.

[0053] In a more preferred embodiment, the composition of the present invention is 0.15% by weight of benzalkonium chloride 0.15% by weight of decyldimethylammonium chloride 0.165% by weight of polyhexamethylene biguanide hydrochloride 0.045% by weight of Bronopol 0.002% by weight of p-chloro-m-cresol 0.245% by weight ethanol 0.05% by weight of ethylene glycol, and 99.2% by weight of water Includes.

[0054] In another, more preferred embodiment, the composition of the present invention is 0.045% by weight of benzalkonium chloride 0.045% by weight of decyldimethylammonium chloride 0.0495% by weight of polyhexamethylene biguanide hydrochloride 0.0135% by weight of Bronopol 0.0006% by weight of p-chloro-m-cresol 0.0735% by weight ethanol 0.015% by weight of ethylene glycol, and 99.76% by weight of water Includes.

[0055] The compositions of the present invention may be formulated in any manner suitable for administration to humans or animals. Preferred examples include liquids, foams, humectants, and gels.

[0056] The compositions of the present invention may be applied to humans or animals by any suitable means, for example, by wipes, sprays, sponges, cloths, towels, mouthwash, or dressings impregnated with the compositions. Preferably, the compositions are applied to humans or animals by wipes, sprays, or dressings impregnated with the compositions. In another preferred embodiment, the compositions of the present invention are applied to the skin of humans or animals in the oral cavity / oral surface by mouthwash, i.e., the compositions of the present invention are used as mouthwash.

[0057] Use of the composition of the present invention (i) Treatment or prevention of pathogenic infections The compositions of the present invention are useful for the treatment or prevention of pathogenic infections in humans or animals, preferably in humans. Most preferably, the compositions of the present invention as defined herein are useful for the treatment of pathogenic infections in humans or animals, preferably in humans. Therefore, the compositions of the present invention are useful for the treatment or prevention of diseases / conditions associated with, i.e., caused by, pathogenic infections. The compositions are also useful for preventing the transmission of pathogenic infections from one human or animal to another, preferably from human to human.

[0058] Pathogenic infections that can be treated or prevented by the compositions of the present invention include bacterial infections, fungal infections, and viral infections.

[0059] Examples of bacterial infections that can be treated or prevented by the compositions of the present invention include bacterial infections inside / on the surface of wounds, impetigo, ulcers (including diabetic foot ulcers and bedsores / pressure ulcers), boils, leprosy, bacterial foot disease, cellulitis, abscesses, interdigital dermatitis, and erysipelas. Particularly preferred bacterial infections are bacterial infections inside / on the surface of wounds (i.e., the wound contains at least one type of bacteria) and impetigo. Treatment of bacterial infections inside / on the surface of wounds means that the bacteria inside / on the surface of the wound are killed, i.e., removed in whole or in part, by the compositions of the present invention.

[0060] Other particularly preferred bacterial infections that can be treated or prevented by the compositions of the present invention include bacterial infections involving Staphylococcus aureus (e.g., MRSA), Pseudomonas aeruginosa, Escherichia coli, E. hirae, A. baumannii, Corynebacterium amycolatum, Corynebacterium striatum, and / or Klebsiella sp., preferably Staphylococcus aureus, Pseudomonas aeruginosa and / or Klebsiella sp., i.e., bacterial infections at least partially caused by these.

[0061] In preferred embodiments, the bacterial infection includes rod-shaped (e.g., Klebsiella) and / or spherical (e.g., Staphylococcus aureus) bacteria. More preferably, the bacterial infection includes rod-shaped bacteria.

[0062] Examples of fungal infections that can be treated or prevented by the compositions of the present invention include fungal infections inside / on the wound, nail fungal infections, lower extremity fungal infections, ulcers, yeast infections, tinea pedis, and tinea. Particularly preferred fungal infections are fungal infections inside / on the wound (i.e., the wound contains at least one type of fungus) and nail fungal infections. Treatment of a fungal wound means that the fungus inside / on the wound is killed, i.e., removed (in whole or in part) by the compositions of the present invention.

[0063] Examples of viral infections that can be treated or prevented by the compositions of the present invention include norovirus, coronavirus, herpes, cold sore virus, HPV, chickenpox, herpes zoster, measles, warts, and molluscum contagiosum. Particularly preferred viruses are norovirus and coronavirus.

[0064] In one embodiment, the composition of the present invention is used for the treatment or prevention of bacterial and / or fungal infections. In a preferred embodiment, the pathogenic infection is a bacterial infection. In another preferred embodiment, the pathogenic infection is a fungal infection.

[0065] In one embodiment, the pathogenic infection is a pathogenic infection of human or animal skin, muscle tissue, connective tissue, and / or skeletal tissue. In a preferred embodiment, the pathogenic infection is a pathogenic infection of human or animal skin.

[0066] In preferred embodiments, the compositions of the present invention are applied topically to humans or animals. If the pathogenic infection is a cutaneous pathogenic infection, the compositions may be applied directly to the skin of humans or animals. Therefore, in preferred embodiments, the compositions of the present invention are applied topically to the skin of humans or animals.

[0067] In preferred embodiments, the pathogenic infection is internal to the wound / on the wound surface, and therefore the compositions of the present invention are for treating or preventing pathogenic infections internal to the wound / on the wound surface, i.e., for killing pathogenic infections internal to the wound / on the wound surface. In preferred embodiments, the wound is a bacterial wound and / or a fungal wound. In more preferred embodiments, the wound is a bacterial wound, i.e., the wound contains bacteria that are killed by the compositions of the present invention, i.e., bacteria that are removed, either whole or in part. Preferably, the bacteria internal to the wound / on the wound surface include one or more of Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, E. hyalae, A. baumannii, Corynebacterium amycolatum, Corynebacterium striatum, and / or Klebsiella species, more preferably Staphylococcus aureus, Pseudomonas aeruginosa, and / or Klebsiella species. This example demonstrates that the compositions of the present invention, when tested under “contaminated conditions” and in real-life wounds, possess bactericidal activity against bacteria in these common wounds. In another preferred embodiment, the bacterial infection inside / on the wound surface includes rod-shaped and / or spherical bacteria, more preferably rod-shaped bacteria. In some embodiments, the wound is a cut, burn, puncture, or ulcer.

[0068] The wound may be a chronic wound. As shown in the examples, the compositions of the present invention can treat chronic wounds because their activity spectrum encompasses Gram-positive and Gram-negative bacteria, and they have rapid bactericidal and fungicidal activity with excellent residual activity. The compositions of the present invention act rapidly and exhibit broad-spectrum bactericidal activity, thereby reducing the possibility of resistance development, and are therefore suitable for treating or preventing chronic wounds.

[0069] This example also demonstrates that the compositions of the present invention can treat pathogenic infections containing biofilms, particularly biofilms containing VBNC cells, i.e., pathogenic infections containing pathogens in a VBNC state. In preferred embodiments, the compositions of the present invention are for use in treating bacterial infections containing Gram-positive and / or Gram-negative bacterial VBNC biofilms. In particularly preferred embodiments, the compositions of the present invention are for use in treating bacterial infections within / on the surface of a wound, where the wound contains a biofilm, and the biofilm contains Gram-positive and / or Gram-negative bacteria, which may be in a VBNC state.

[0070] The compositions of the present invention may also be used to heal wounds and treat pathogenic infections in wounds as described herein, which may be done simultaneously or sequentially. "Healing wounds" means that the compositions of the present invention promote wound closure, i.e., accelerate healing.

[0071] In certain embodiments, the present invention is For use in the treatment or prevention of bacterial and / or fungal infections of the skin of humans or animals, Benzalkonium chloride Decyldimethylammonium chloride Polyhexamethylene biguanide hydrochloride Bronopol p-chloro-m-cresol ethanol Ethylene glycol, and water A composition comprising, which is applied topically to the skin of a human or animal, or The above composition for use in the treatment or prevention of bacterial and / or fungal infections inside / on the surface of a wound, and optionally the above composition for further healing of the wound, The present invention relates to a composition for use in the treatment or prevention of bacterial infection inside or on the surface of a wound, wherein the wound is a chronic wound comprising a biofilm which may contain VBNC cells.

[0072] In another embodiment, the present invention is for use in the treatment or prevention of bacterial and / or fungal infections of human or animal skin. 0.04-0.2% by weight of benzalkonium chloride 0.04-0.2% by weight of decyldimethylammonium chloride 0.04-0.2% by weight of polyhexamethylene biguanide hydrochloride 0.01-0.06% by weight of Bronopol 0.0005 to 0.005% by weight of p-chloro-m-cresol 0.05-0.3% by weight of ethanol 0.01 to 0.07% by weight of ethylene glycol A composition comprising, with the remainder being water, A composition to be applied topically to the skin of a human or animal, or The above composition for use in the treatment or prevention of bacterial and / or fungal infections inside / on the surface of a wound, and optionally the above composition for further healing of the wound, The present invention relates to a composition for use in the treatment or prevention of bacterial infection inside or on the surface of a wound, wherein the wound is a chronic wound comprising a biofilm which may contain VBNC cells.

[0073] In another embodiment, the present invention is for use in the treatment or prevention of bacterial and / or fungal infections of human or animal skin. 0.15% by weight of benzalkonium chloride 0.15% by weight of decyldimethylammonium chloride 0.165% by weight of polyhexamethylene biguanide hydrochloride 0.045% by weight of Bronopol 0.002% by weight of p-chloro-m-cresol 0.245% by weight ethanol 0.05% by weight of ethylene glycol, and 99.2% by weight of water A composition comprising, A composition to be applied topically to the skin of a human or animal, or The above composition for use in the treatment or prevention of bacterial and / or fungal infections inside / on the surface of a wound, and optionally the above composition for further healing of the wound, The present invention relates to a composition for use in the treatment or prevention of bacterial infection inside or on the surface of a wound, wherein the wound is a chronic wound comprising a biofilm which may contain VBNC cells.

[0074] In another embodiment, the present invention is for use in the treatment or prevention of bacterial and / or fungal infections of human or animal skin. 0.045% by weight of benzalkonium chloride 0.045% by weight of decyldimethylammonium chloride 0.0495% by weight of polyhexamethylene biguanide hydrochloride 0.0135% by weight of Bronopol 0.0006% by weight of p-chloro-m-cresol 0.0735% by weight ethanol 0.015% by weight of ethylene glycol, and 99.76% by weight of water A composition comprising, A composition to be applied topically to the skin of a human or animal, or The above composition for use in the treatment or prevention of bacterial and / or fungal infections inside / on the surface of a wound, and optionally the above composition for further healing of the wound, The present invention relates to a composition for use in the treatment or prevention of bacterial infection inside or on the surface of a wound, wherein the wound is a chronic wound comprising a biofilm which may contain VBNC cells.

[0075] (ii) Treatment or prevention of rosacea, eczema and / or psoriasis Any one of the compositions defined herein may be used to treat or prevent, most preferably to treat, rosacea, eczema, or psoriasis. Accordingly, the present invention also relates to compositions defined herein for use in the treatment of rosacea, eczema, or psoriasis, preferably psoriasis and / or eczema. As shown in the examples, the inventors have surprisingly found that the compositions of the present invention also treat skin infections that are not caused by or resulting from pathogenic infections.

[0076] (iii) Wound healing Any one of the compositions of the present invention described herein can also be used to heal wounds, i.e., to accelerate wound closure. Therefore, the compositions of the present invention can be used to heal wounds (without necessarily treating pathogenic infection inside / on the wound surface).

[0077] However, the treatment of pathogenic infections within / on the wound surface contributes to wound healing; that is, the successful treatment of pathogenic infections within / on the wound surface using the compositions of the present invention contributes to wound healing. Therefore, in preferred embodiments, the compositions of the present invention are used to heal wounds, that is, to promote wound healing / closure, while simultaneously treating pathogenic infections within / on the wound surface as described herein, which may be done simultaneously or sequentially. In particularly preferred embodiments, the compositions of the present invention are used to treat or prevent bacterial and / or fungal infections within / on the wound surface, and to heal wounds.

[0078] The wound may be any wound described herein, such as a cut, burn, puncture, or ulcer. In a preferred embodiment, the wound is a chronic wound. In a more preferred embodiment, the wound is a chronic wound comprising a biofilm. In another preferred embodiment, the composition of the present invention used to heal the wound comprises at least 99.76% by weight of water.

[0079] (iv) The virus on the surface is killed or inactivated. Any one of the compositions of the present invention described herein can be used to kill or inactivate viruses on a surface. Accordingly, the present invention also relates to a method for killing or inactivating viruses on a surface, comprising the step of applying a composition of the present invention to the surface.

[0080] As shown in Examples 8 and 9, the compositions of the present invention can kill or inactivate viruses on surfaces even at low concentrations, that is, even when the composition contains a large amount of water, for example, more than 99% by weight.

[0081] The surface may be any surface that contains viruses. Non-limiting examples of surfaces include floors, tabletops, veterinary operating rooms, hospital operating rooms, and kitchen sideboards, and may have any angle to the ground surface and any shape; that is, reference to a surface is not limited to a flat surface. The surface may be made of any material, such as plastic, wood, or metal. In preferred embodiments, the surface is a plastic or metal surface, such as steel.

[0082] In one embodiment, a method for killing or inactivating a virus is not a method for treating the body of a human or animal by therapy.

[0083] Viruses that can be killed or inactivated on a surface by the composition of the present invention include coronaviruses, such as human coronavirus 229E (HuCoV-229E), noroviruses, such as murine norovirus 1 (MNV-1) strain CW1, and herpes. Preferred viruses that can be killed or inactivated on a surface by the composition of the present invention are coronavirus 229E (HuCoV-229E) and murine norovirus 1 (MNV-1) strain CW1.

[0084] As shown in Examples 8 and 9, the compositions of the present invention kill or inactivate even the most robust surface viruses, such as murine norovirus 1 (MNV-1) strain CW1.

[0085] In one embodiment, the surface is pre-treated with the composition. Pre-treatment means that the composition is applied to the surface before viruses are identified on the surface; that is, the composition of the present invention is used as a preventive measure against viruses on a surface.

[0086] When applied to a surface, the composition of the present invention prevents the proliferation / activation of viruses on the surface for a long period of time after application of the composition to the surface. In one embodiment, the composition kills or inactivates viruses on the surface at least 1 hour after application of the composition to the surface. In a preferred embodiment, the composition kills or inactivates viruses on the surface at least 24 hours after application of the composition to the surface. In another preferred embodiment, the composition kills or inactivates viruses on the surface at least 48 hours after application of the composition to the surface.

[0087] Numerous modifications and variations of the embodiments described herein may be made without departing from the spirit and scope of the invention, as will be obvious to those skilled in the art. The specific embodiments described herein are presented merely as examples. [Examples]

[0088] The composition of the present invention used in this embodiment contained the following components.

[0089] Composition of the present invention (83.9% by weight of water): 3% by weight of benzalkonium chloride 3% by weight of decyldimethylammonium chloride 3.3% by weight of polyhexamethylene biguanide hydrochloride 0.9% by weight of Bronopol 0.04% by weight of p-chloro-m-cresol 4.9% by weight ethanol 1.0% by weight of ethylene glycol, and 83.9% by weight of water.

[0090] Composition of the present invention (99.2% by weight of water): 0.15% by weight of benzalkonium chloride 0.15% by weight of decyldimethylammonium chloride 0.165% by weight of polyhexamethylene biguanide hydrochloride 0.045% by weight of Bronopol 0.02% by weight of p-chloro-m-cresol 0.245% by weight ethanol 0.05% by weight of ethylene glycol, and 99.2% by weight of water.

[0091] Composition of the present invention (99.76% by weight of water): 0.045% by weight of benzalkonium chloride 0.045% by weight of decyldimethylammonium chloride 0.0495% by weight of polyhexamethylene biguanide hydrochloride 0.0135% by weight of Bronopol 0.0006% by weight of p-chloro-m-cresol 0.0735% by weight ethanol 0.015% by weight of ethylene glycol, and 99.76% by weight of water.

[0092] All examples presented herein include experiments conducted under confidentiality agreements. [Examples]

[0093] Treatment of widespread acute impetigo A 55-year-old female patient developed widespread acute impetigo. Initially, she was misdiagnosed by a local pharmacist with a tinea pedis infection, which allowed the infection to proliferate and spread uncontrollably for two weeks. The pharmacist prescribed clotrimazole / Canestin cream twice daily, along with thorough washing and drying procedures. The patient strictly followed these instructions, using dressings and cleaning four times daily for several days. Her condition continued to worsen, and her legs became so swollen that she could not wear closed shoes. Furthermore, rash-like symptoms similar to those of an infection began to appear on her arms, chest, neck, and face, accompanied by a low-grade fever.

[0094] Next, the patient went to University College Hospital A&E, where she was immediately diagnosed with impetigo. Due to the acute and widespread nature of the infection, oral antibiotics were advised and prescribed. The patient requested a parenteral alternative because she had a history of anaphylactic reactions, primarily to penicillin-class antibiotics. The resident general practitioner stated that while antibiotic cream was not related to the patient's allergy, it was likely insufficient to adequately treat the patient's widespread infection. Clindamycin ointment was prescribed, and the patient was advised to return to A&E for oral clindamycin if symptoms did not improve within 48 hours.

[0095] Clindamycin ointment initially appeared effective, and the exudation seemed to be slightly reduced. The treated area appeared inflamed, but the infection seemed to be decreasing, and application was continued. Subsequently, redness / inflammation and swelling in the area increased. Topical treatment was discontinued after the first morning application. Swelling persisted around the clock, accompanied by contractions of breathing and swallowing, as well as chest pain. This is a typical anaphylactic reaction. The National Health Service (NHS) was contacted directly, and the patient was treated by ambulance, but they were told they did not have an epinephrine pen.

[0096] Next, the patient made an appointment with a local general practitioner, who immediately administered epinephrine, which stopped further allergic reactions. However, the impetigo infection persisted. The impetigo infection was located on the upper part of the patient's feet, ankles, forearms, chest, neck, and face. Corticosteroids were prescribed by the general practitioner, which helped to reduce some of the swelling, but the infection did not subside.

[0097] A wipe containing the composition of the present invention (99.2% by weight of water) was received from the applicant under a confidentiality agreement, and the entire infected area was immediately wiped. The patient felt an immediate cooling / sedation sensation, and the itching subsided significantly. The raised, oozing rash began to dry and lessen. The composition was applied to the entire area several times a day after thorough cleaning and drying of the affected area.

[0098] Significant improvement was observed, with the previous exudation visibly drying up, although swelling remained. Redness and itching decreased exponentially. The uncontrolled spread of infection ceased. After one week of treatment, some redness and swelling were still present, but the exudate had decreased significantly, and there was no itching.

[0099] The composition of the present invention was used for 4 weeks, gradually decreasing the frequency of application from 3-4 times per day to 2 times per day to 1 time per day. After 4 weeks, the patient was completely free of impetigo infection.

[0100] In summary, the composition of the present invention stopped and controlled widespread impetigo bacterial infections within one week and promoted complete recovery within three to four weeks. However, the patients did not show improvement with known medicines such as antibiotics and corticosteroids, thus illustrating the superiority of the composition of the present invention over known medicines. Photographs of the patient's legs before, during, and after treatment with the composition of the present invention are shown in Figure 1. [Examples]

[0101] Treatment and healing of Gram-positive and Gram-negative VBNC bacterial (biofilm) wounds. The trials were conducted in a confidential, approved clinical study in India using a non-invasive fluorescence-based multispectral imaging device, capturing pre- and post-treatment images of trauma with the composition of the present invention (99.2% by weight water). This method enabled automated detection of the presence (or absence) of specific Gram-positive and Gram-negative VBNC (biofilm) pathogens and wound dimensions, as well as clinical classification, within two minutes of imaging, without the addition of any external reagents. The device uses multispectral imaging in combination with advanced computational algorithms and a proprietary state-of-the-art ML engine for spatial mapping and detection of pathogens.

[0102] The results showed that, prior to the application of the composition of the present invention to the wound (by wiping), the wound contained VBNC Gram-negative bacteria (in this case, Klebsiella) in the form of a biofilm (Figure 2a). However, two minutes after application of the composition of the present invention, the bacteria and biofilm were completely absent (Figure 2b).

[0103] Similar results are shown in Figures 2c–h. In Figures 2c–e, a visible reduction in rod-shaped bacteria, specifically Klebsiella and Pseudomonas aeruginosa, can be seen after application of the compositions of the present invention to wounds, and in Figures 2f–h, a visible reduction in Klebsiella can be seen after application of the compositions of the present invention to different wounds. The bacteria in these examples initially appear to be present in the muscle and connective tissue inside / on the wound surface, and can then be dramatically reduced after treatment with the compositions of the present invention.

[0104] Figures 2i-j show that MRSA, or Staphylococcus aureus, is reduced in wounds after application of the composition of the present invention.

[0105] These remarkable results are significant because even when wounds are cultured "clean," healing may still be hindered by the presence of biofilm. However, the compositions of the present invention completely removed biofilm (e.g., Figures 2a-b) and thus promoted improved healing without the use of systemic antibiotics.

[0106] Therefore, since the compositions of the present invention can remove biofilms containing Gram-positive and Gram-negative bacteria in a VBNC state, they can be used to promote the treatment of chronic wounds (which are prone to biofilm formation), as well as trauma, ulcers, burns, and the like. Considering that biofilm-related infections lead to significantly high morbidity and mortality rates, the compositions of the present invention are considered particularly important. [Examples]

[0107] Treatment of dog bites and horse leg wounds: Cure and prevention of pathogenic infections An 82-year-old man sustained a dog bite puncture wound on his finger. The composition of the present invention (99.2% by weight of water) was applied immediately and daily (under a confidentiality agreement), and the wound healed rapidly without infection, and then completely healed within three weeks. The man strongly asserted that the use of the composition of the present invention significantly contributed to the acceleration of the healing process.

[0108] Photographs of the wound before and after healing are shown in Figure 3a and Figure 3b (1 month later), respectively.

[0109] A 24-year-old horse suffered a trauma that penetrated the tendon sheath of its foreleg. Based on past experience, the veterinarian recommended immediate euthanasia, citing the high likelihood of infection and the very poor prognosis for horses of that age. The owner refused and instead chose to suture the wound and (under a confidentiality agreement) apply the composition of the present invention daily. To the veterinarian's surprise, the horse recovered completely without any apparent infection.

[0110] Photographs of the horse's legs from the front and back (at 5-week intervals) are shown in Figures 4a and 4b, respectively. [Examples]

[0111] Treatment of nail fungal infections The patient tried various treatments for their nail fungal infection over a three-year period. After numerous visits to the doctor, nothing proved effective.

[0112] Under a confidentiality agreement, the patient was introduced to the composition of the present invention (99.2% by weight of water) and began applying it to their nails. Just one year later, despite previous treatments being ineffective, their nails were healthy. The patient rated the composition "10 out of 10" and commented that it had no odor and no greasy feeling.

[0113] Photographs of patients' thumbs before, during, and after treatment with the composition of the present invention are shown in Figures 5a, 5b, and 5c, respectively. [Examples]

[0114] Treatment of psoriasis and eczema in human skin within 48 hours The patient was 20 years old and suffered from a long-term skin condition diagnosed as psoriasis and eczema. The patient had used various over-the-counter (OTC) and RX medications with limited effectiveness. Following an invasive procedure, the patient, under a confidentiality agreement, chose to undergo an experiment using the composition of the present invention (in this case, containing 99.76% by weight of water). The patient applied it only to the rash on their hands and compared the results with the rash on their face. The first application was at 0 hours, and at 24 hours, the patient reported a reduction in skin irritation on their hands, with less redness and less pain. At that time (24 hours), the patient applied the composition of the present invention again, and thereafter, at 48 hours, observed a significant reduction in the symptoms on their hands, with the rash no longer damp, no itching, and the skin color almost returning to normal. The untreated area, namely the patient's face, still retained dampness, pain, itching, and redness.

[0115] Figures 10a and 10b show front and rear photographs of the patient's hand and face, respectively. These figures show that the untreated skin (i.e., face) worsened over a 48-hour period, while the skin treated with the composition of the present invention (i.e., hand) showed dramatic improvement over the same period.

[0116] Therefore, the compositions of the present invention not only treat pathogenic infections (e.g., skin), but also treat skin conditions not caused by pathogenic infections (e.g., psoriasis and / or eczema). [Examples]

[0117] Skin irritation test An in vitro evaluation of the irritancy potential of the compositions of the present invention, using a modified human skin model, was conducted under a confidentiality agreement by Dr. David Voegeli, Associate Professor of Nursing at the Skin Health Research Group.

[0118] The overall objective of this project was to investigate the effects of several compositions of the present invention on a modified human skin model of approved tissue (European Centre for Evaluation of Alternatives to Animal Experiments, EURL-ECVAM).

[0119] The purpose of this test is as follows: 1. Determination of the degree of skin irritation caused by various compositions of the present invention. 2. Determination of the in vitro toxicity of various compositions of the present invention to human keratinocytes.

[0120] result • Macroscopic and microscopic examination of implants All received tissues were found to be intact and viable upon visual inspection and were used. After tissue administration, randomly selected inserts were examined by low-power optical microscopy to confirm that no damage had occurred during the procedure. No cellular damage was observed in any of the examined inserts.

[0121] • MTT assay The raw optical density of each well at 570 nm is shown in Table 1 below:

[0122] [Table 1]

[0123] Relative survival rate The calculation of relative survival rates for the tested controls and compositions is shown below (Table 2) and summarized in Figure 6:

[0124] [Table 2]

[0125] conclusion This study was designed to investigate the potential skin irritation of various compositions of the present invention intended for topical application in humans / animals. A total of four products, as well as positive and negative controls, were tested. The calculation of RV% as a measure of irritation showed that all of the compositions of the present invention tested had an RV% greater than 50, with the majority (75%) having an RV% greater than 100.

[0126] These data demonstrate that, unlike many known disinfectants in the art, none of the compositions tested are likely to cause skin irritation, even at higher concentrations, such as the composition according to the present invention containing 16% by weight of water. [Examples]

[0127] non-cytotoxic The compositions of the present invention were studied for cytotoxicity (in accordance with ISO 10993-5) under a confidentiality agreement. While it is not essential for the compositions of the present invention to be non-cytotoxic according to this test, it is an advantageous characteristic.

[0128] The composition according to the present invention (containing 99.2% culture medium) was found to be cytotoxic, i.e., to cause complete cell death. However, the composition according to the present invention containing 99.76% culture medium was found to be non-cytotoxic (26% reduction in cell density compared to the negative control; ISO 10993-5 allows for a 30% inhibition of cell proliferation).

[0129] Therefore, if the composition of the present invention contains at least about 99.5% by weight of water (e.g., at least 99.76% by weight of water), the composition of the present invention is non-cytotoxic. As shown in Examples 5 and 11, the composition of the present invention, at this non-cytotoxic concentration, is remarkably capable of treating skin conditions such as psoriasis and eczema, and also possesses bactericidal activity against various bacteria in a contaminated state (i.e., in vitro conditions simulating the conditions inside / on the surface of a human / animal wound), and is therefore capable of treating pathogenic infections such as pathogenic infections inside / on the surface of a wound and infections illustrated in Examples 1-4, for example. [Examples]

[0130] Killing and inactivating coronavirus on surfaces protocol The two methods were performed under confidentiality agreements at 21°C: i) A stainless steel (S30400) surface was pre-coated with the composition of the present invention (in this case, containing 99.2% by weight of water), and then sensitized with a virus. ii) The virus was dried on a stainless steel surface and then sensitized with the composition of the present invention (in this case, containing 99.2% by weight of water).

[0131] Next, the virus was recovered from all surfaces and tested to see how much infectious virus (i.e., capable of causing disease) remained after treatment.

[0132] • Preparation of various reagents Preservation solution of HuCoV-229E virus (lysate of crude infected cells): This virus was originally isolated from the upper respiratory tract of humans with bronchitis in the 1960s. This virus has been used as a substitute for highly pathogenic coronaviruses that cause SARS and MERS, requiring specialized, more advanced containment facilities. Unlike other substitutes such as TGEV, HuCoV-229E is a hazard group 2 (HG2) virus that causes respiratory infections in humans and affects the intestinal mucosa. In most individuals, the immune response is unable to eradicate the virus over time, so HuCoV-229E infections occur every few years. The viral preparation contains a complex mixture of dead human lung cells and biomolecules (including components of the growth medium) to mimic viral contamination in sputum and natural respiratory secretions.

[0133] The host cell line is MRC-5 normal human lung fibroblasts (HuCov-229E can attach to and replicate in this cell line).

[0134] Throughout, stainless steel test piece (1cm 2 A sample of a common surface material (×0.5 mm) was used.

[0135] Phosphate-buffered saline (PBS), cell culture growth medium, and supplements such as trypsin / EDTA, low-melting-point agarose solution for overlay, and a neutral red dye.

[0136] · consumables Disposable tubes (6 mL, 30 mL, and 50 mL), glass beads, tissue culture flasks and 6-well trays, loops and spreaders, petri dishes, culture boxes, pipettes, tips, pastettes, syringes, and syringe filters.

[0137] · Device Microbial safety cabinet Microbiological incubator (37°C, carbon dioxide pumped into the incubator and maintained at 5% for optimal cell growth). Microcentrifuge Inverted microscope Lightbox and magnifying lens

[0138] • Test Procedure The effectiveness of the composition of the present invention in inactivating HuCoV-229E was determined by two methods: 1. Non-biocidal surfaces were pre-coated with the compositions of the present invention for 1, 24, and 48 hours to investigate whether viruses were inactivated upon contact, i.e., whether, if a surface was washed with the compositions of the present invention, the untreated layer would inactivate any viral contamination of that surface, for example, in respiratory secretions during coughing and sneezing that occurred later. 2. We investigated whether the composition of the present invention, applied to a non-biocidal surface pre-coated with a virus for 0, 30 seconds, 5 minutes, or 10 minutes, inactivates the virus, that is, whether cleaning an already contaminated surface with the composition of the present invention inactivates the virus.

[0139] • Experimental protocol Preliminary experiments and preparation: - Passage cell line MRC-5 to a sufficient quantity for the experiment. This includes passaging every 3 days for each additional flask (cells will adhere to plastic and need to be removed using trypsin / EDTA). On the day before the experiment, place the cells into approximately 10 tissue culture flasks (with a base of 75 cm). 2 Remove the cells from the wells, resuspend them in fresh culture medium, and seed them in a 6-well tray, where the cells settle and adhere to the bottom of each well, forming a monolayer of cells. - Coat the test specimens with the composition of the present invention 24 hours and 48 hours before the experiment.

[0140] On the day of the experiment: - Before starting the experiment, confirm that the cells appear healthy and subconfluent. - Inoculate the stainless steel test piece with one of the following (triple increment): (1) Spread 20 μL of the composition of the present invention (99.2% by weight water) over the entire surface of the test specimen for 1 hour - Method 1 (2) For 1 hour, coronavirus (20 μL of HuCo-229E, approximately 103 plaque-forming units, on the surface of the test specimen (1 cm) 2 (Spread it evenly) - Method 2 ○ Add the virus to the test specimen for Method 1 for 0, 30 seconds, 5 minutes, and 10 minutes, or add the composition of the present invention to the test specimen for Method 2 for the same duration. ○ Each test specimen is placed in a separate tube containing 5 mL of growth medium (5% fetal bovine serum) / glass beads (2 mm), and the virus is removed in the same manner for each method. ○ Vortex each for 15 seconds. Prepare a 10-fold dilution of each sample in the growth medium. Remove the culture medium from each well of the 6-well tray containing MRC-5 cells, add 1 mL of test dilution, and incubate at 37°C and 5% CO2 for 90 minutes (the virus will attach to the cells during this time). Carefully remove the virus and add the overlay. Allow to cool for 15 minutes to allow the overlay to harden, then transfer to the incubator. Incubate at 37°C and 5% CO2 for 6 days.

[0141] Observation and calculation of infectious viruses recovered from each test specimen: Add 3 mL of a 3% solution of sterile, filtered neutral red vital stain to each well and incubate for 2 hours. Remove the liquid stain from the top of the well and incubate for another hour. This stain is taken up by living cells and stains them red. Dead cells do not take up this stain. If the virus replicates inside the cell, the plaque is visible as an unstained area. In a cell monolayer, this actually becomes a hole, in which case the infected cell ruptures and dies (lyses), and is surrounded by the residue of the dead infected cell. The overlay prevents the virus from spreading beyond the attachment time, resulting in clearer plaques that are easier to count.

[0142] - Count the plaque * The amount of infectious virus recovered from each test specimen is calculated. Photographs are taken as needed. Regarding plaque counting, the virus count is often expressed as plaque-forming units (pfus) (not all viruses produce plaques), and this is difficult to represent as the actual virus count, so it is an estimate of the amount of infectious virus present. - Use graphic software to analyze the data.

[0143] result The results are shown in Figure 7 and are summarized below: The tables in Figures 7a and 7b include raw data for each contact time (average pfu recovered per surface area of ​​the test specimen) and the Log10 reduction for infectious human coronavirus 229E (HuCoV-229E) at 1 cm when initiating vaccination, i.e., when treatment is not performed. 2 The maximum amount of infectious virus per unit is 2500 plaque-forming units (pfu).

[0144] Figures 7a and 7c-7e (Method (1)): 1 cm at 48 hours, 24 hours, or 1 hour before virus application. 2 The surface is pre-coated with 10 μL of the composition of the present invention.

[0145] Figures 7b and 7f-7g (Method (2)): One hour before applying the composition of the present invention, the surface is pre-coated with 20 μL of HuCoV-229E (2500 pfu).

[0146] Figure 7h = Inactivation rate at 30 seconds for methods (1) and (2).

[0147] The log reduction for infectious viruses was calculated according to the following formula: Log reduction = log10(pfu before processing) - log10(pfu after processing) A 1-log decrease = a 90% decrease. 2log decrease = 99% decrease 3log decrease = 99.9% decrease 4log decrease = 99.99% decrease A 5log decrease = a 99.999% decrease. 6log decrease = 99.9999% decrease

[0148] The summarized results show that the composition of the present invention is highly effective against human coronavirus 229E, even when used at very low concentrations.

[0149] Furthermore, these results demonstrate that coating a surface with the composition of the present invention keeps viruses inactive for extended periods. There was no difference between coatings after 1 hour and 24 hours, and after 48 hours, only a slight decrease in effectiveness was observed on the coated surface. All viruses were inactivated within 30 seconds to 5 minutes.

[0150] The composition of the present invention is highly effective against dried viruses, and also inactivated dried viruses within 30 seconds to 5 minutes.

[0151] Therefore, the compositions of the present invention can remain inactive on any surface for a long period of time, which is useful, for example, for contamination that occurs between two cleaning regimens. [Examples]

[0152] Killing and inactivating norovirus on the surface. The purpose of these experiments was to determine whether the composition of the present invention (in this case, containing 99.2% by weight of water) could kill or inactivate murine norovirus. Three experiments were conducted under confidentiality agreements, as summarized below.

[0153] Experimental protocol Virus strains and cell lines The mouse norovirus 1 (MNV-1) strain CW1 and the mouse monocyte macrophage strain RAW264.7 were provided by Dr. Herbert Virgin IV (Washington University, USA). The semi-adherent cell line was maintained at sub-confluence to prevent loss of the characteristic phenotype and was maintained at 37 °C in a humidified atmosphere of 5% CO2 in HEPES-buffered Dulbecco's modified Eagle's medium (DMEM) containing GlutaMAX, 25 mM D-glucose, 10% fetal bovine serum, and no sodium pyruvate. The cells attach to tissue culture grade plastic via cation-dependent and independent receptors and can be easily detached by rubbing.

[0154] Preparation of the sample surface Stainless steel specimens (10×10×0.5 mm) were degreased in acetone, stored in absolute ethanol, and flamed before use.

[0155] Inoculation of metal specimens with MNV-1 (to mimic contamination by wet contaminated interferences) and assessment of infectious virus by detection of the cytopathic effect in a mouse cell line (plaque assay) To present a wet contaminated interference, 20 μL of MNV-1 (1.2×10 6 PFU / ml) was inoculated onto the surface of the specimen (dried at 22 °C for 30 - 40 minutes). The drying time was included in the exposure time. The virus was removed from the specimen in 2 hours by vortexing for 30 seconds in 2 ml of complete DMEM containing glass beads approximately 50×2 mm in diameter. Various dilutions were immediately prepared in complete DMEM and 1 mL aliquots were added to each well of a 6-well plate (3.5 cm diameter) 3 hours before at a rate of 10 5 plaque-forming units (pfu) per well. 6Individual cells were seeded into a monolayer of RAW264.7 and incubated at 37°C and 5% CO2 for 90 minutes. The inoculum was aspirated, and 3 mL of 3% low-melting-point (LMP) agarose overlay in complete medium was added per well to prevent the virus from spreading to other cells. The plates were incubated at 4°C for 15 minutes until hardened, then at 37°C and 5% CO2 for 72 hours. The monolayer was stained with 2 mL per well of filtered 0.01% Neutral Red, a supervivicultural stain that accumulates lysosomes that stain cells red when pinocytized by living cells in PBS at 37°C and 5% CO2 for 2 hours. Excess stain was removed, and the plates were incubated again for a further 1 hour. The plates were stored overnight at 4°C to increase plaque resolution, which was then counted and used to calculate the pfu recovered per specimen.

[0156] Modifications to the above protocol 1. Add the virus to a test piece that has been pre-coated with the composition of the present invention. Twenty hours prior to the experiment, steel specimens were coated with either the composition of the present invention or 20 μl of sterile distilled water. These liquids were spread over the entire surface and allowed to dry. The specimens were then stored at room temperature until required for the assay, and the virus was added to the top of this pre-coated layer. The assay was then carried out as detailed above.

[0157] 2. Add the composition of the present invention to a test piece that has been pre-coated with the virus. 40 minutes before starting the assay, apply 20 μL of MNV-1 1.2 × 10⁶ to the surface of the test specimen. 5 Plaque-forming units (PFUs) were inoculated. Subsequently, once the surface had dried, it was coated with the composition of the present invention or sterile distilled water. After incubation for 2 hours, the assay was continued as described above.

[0158] 3. The test specimen is pre-coated with the composition of the present invention, then the virus is applied, and then the composition of the present invention is applied again. Two hours before starting the assay, the steel specimens were coated with 20 μl of the composition of the present invention or sterile distilled and coated with the composition. Forty minutes before starting the assay, the surface of the specimens was coated with 20 μl of MNV-1 1.2 × 10⁶. 5 Plaque-forming units (PFUs) were inoculated. Subsequently, once the surface was dry, it was coated with the composition of the present invention or sterile distilled water. After incubation for 2 hours, the assay was continued as described above. Each specimen underwent only one type of treatment, and the composition of the present invention or distilled water was in two doses.

[0159] Experiment and Results • Effects of the composition of the present invention, pre-coated on the surface, on murine norovirus A fixed amount of 20 μl of the composition of the present invention was applied to a stainless steel surface. To mimic the regular, predetermined cleaning of the surface, it was dried overnight at room temperature. Next, a virus suspension was applied to the pre-coated surface to mimic droplets spreading through human infection during a norovirus outbreak. After 2 hours of incubation on the steel surface pre-coated with the composition of the present invention, the infectivity of the virus decreased by 1 log and 2 log, respectively (Table 3 and Figure 8a). Since the composition of the present invention had been applied to the steel surface 20 hours earlier, the above indicates that it remains active for a certain period of time while the surface dries.

[0160] [Table 3]

[0161] • Effects of the composition of the present invention when applied directly to murine norovirus A fixed amount of 20 μl of the virus suspension was dried on a steel surface to mimic virus-containing droplets that spread through human infection. Next, this was treated with 20 μl of the composition of the present invention to mimic the surface cleaning procedure after infection. After 2 hours of incubation, the infectivity of the virus decreased by 2 log (Table 3 and Figure 8b).

[0162] • Effects of the composition of the present invention when applied directly to a dry surface and murine norovirus. A stainless steel surface was pre-coated with the composition of the present invention, a virus suspension was applied to the surface, and then the composition of the present invention was reapplied to the surface and incubated for 2 hours. This study was intended to mimic a set cleaning procedure, norovirus outbreak, and a second surface decontamination. After incubation at room temperature for 2 hours with these applications of the composition of the present invention, it was observed that the infectivity of the virus decreased by at least 3 log in the case of the composition of the present invention (Table 3 and Figure 8b).

[0163] Summary of results The composition of the present invention (containing 99.2% by weight of water) was able to kill or inactivate murine norovirus (MNV-1) by 2 log by directly treating a contaminated surface. When the surface was pre-treated with the composition of the present invention, the long-lasting effect of the composition on the surface could reduce the viral load on the surface by 1 log, and when the surface was treated both before and after a viral contamination event, a reduction of more than 3 log of the virus was observed (Figure 8c). [Examples]

[0164] The present invention relates to the MICs of various Gram-positive and Gram-negative bacteria. The compositions of the present invention have a broad spectrum of activity at low concentrations and encompass a wide range of Gram-positive and Gram-negative bacteria, as shown in Figure 9. Figure 9 shows the MICs of the compositions of the present invention against various bacteria in a standard bacteriological culture environment.

[0165] Therefore, this example illustrates that the composition of the present invention has a broad spectrum of activity, encompassing Gram-positive bacteria, Gram-negative bacteria, and spore-forming bacteria, which are required, for example, in the treatment of chronic wounds. When combined with Examples 1-7 and 11, this example illustrates that the composition of the present invention has a broad spectrum of activity against bacteria in the treatment of pathogenic infections, such as pathogenic infections inside / on the surface of wounds in humans or animals. [Examples]

[0166] Bactericidal activity of the composition of the present invention under contaminated conditions The tests were conducted by Abbott Analytical Ltd. under a confidentiality agreement to evaluate the activity of the compositions of the present invention against various bacteria under contaminated conditions (as defined below).

[0167] Test method and its verification Method = Dilution - Neutralization Neutralizing agent = 30.0 g / l polysorbate 80 + 3.0 g / l lecithin + 1.0 g / l L-histidine + 1.0 g / l L-cysteine ​​(Neutralizing agent A) • Verification of the neutralizing agent = Verification in accordance with EN 13727:2012+A2:2015(5.5.2) Experimental conditions • Contact time = 5 minutes ± 10 seconds • Test temperature = 20±1℃ • Interfering substances = 3.0 g / l bovine albumin + 3.0 ml / l sheep red blood cells ("contamination state") • Incubation temperature Requirements Bactericidal activity was defined as a decrease of at least 5.00 log(lg).

[0168] result

[0169] [Table 4]

[0170] conclusion The compositions of the present invention exhibit bactericidal activity against various bacteria under contaminated conditions, in accordance with EN 13727:2012+A2:2015. This contaminated condition (as defined above) was chosen because it corresponds to the conditions found inside / on the surface of human / animal wounds. The range of bacteria tested in this study includes those most commonly found in wounds, particularly chronic wounds. For example, several studies highlight that the majority of chronic wounds contain Staphylococcus aureus, and more than half also contain Pseudomonas aeruginosa. Both of these bacteria, as well as other bacteria commonly found in wounds, were tested in this study.

[0171] These results indicate that when the compositions of the present invention are tested in an environment that mimics real-life wound environments, they exhibit a reduction of more than 5 lb against the bacteria most commonly found in wounds. Furthermore, the compositions of the present invention have been shown to be active against these bacteria even at very low concentrations, i.e., even when the compositions contain at least 99.2% by weight of water (e.g., 99.76% by weight of water).

[0172] Therefore, it was surprisingly found that the compositions of the present invention can treat various bacterial infections inside / on the surface of wounds, even when used at very low concentrations.

Claims

1. For use in healing wounds in humans or animals, 0.04 to 0.2% by weight of benzalkonium halide; 0.04 to 0.2% by weight of didecyldimethylammonium halide; 0.04–0.2% by weight of polyhexamethylene biguanide salt; 0.01–0.06% by weight of bronopol; 0.0005 to 0.005% by weight of p-chloro-m-cresol; and At least 99.2% by weight of water A composition containing the following:

2. The composition for use according to claim 1, wherein the benzalkonium halide is benzalkonium chloride.

3. The composition for use according to claim 1 or 2, wherein didecyldimethylammonium halide is didecyldimethylammonium chloride.

4. The composition for use according to any one of claims 1 to 3, wherein the polyhexamethylene biguanide salt is polyhexamethylene biguanide hydrochloride.

5. The composition for use according to any one of claims 1 to 4, further comprising a solvent.

6. The composition for use according to claim 5, comprising 0.05 to 0.3% by weight of a solvent.

7. The composition for use according to claim 5 or 6, wherein the solvent is ethanol.

8. The composition for use according to any one of claims 1 to 7, further comprising alkylene glycol.

9. The composition for use according to claim 8, comprising 0.01 to 0.07% by weight of alkylene glycol.

10. The composition for use according to claim 8 or 9, wherein the alkylene glycol is ethylene glycol.

11. The composition for use according to any one of claims 1 to 10, further comprising at least 99.76% by weight of water.

12. 0.04 to 0.2% by weight of benzalkonium chloride; 0.04 to 0.2% by weight of didecyldimethylammonium chloride; 0.04–0.2% by weight of polyhexamethylene biguanide hydrochloride; 0.01–0.06% by weight of bronopol; 0.0005 to 0.005% by weight of p-chloro-m-cresol; 0.05 to 0.3% by weight of ethanol; 0.01 to 0.07% by weight of ethylene glycol; Includes, The remainder is water. A composition for use according to claim 1.

13. A composition for use according to any one of claims 1 to 12, wherein the wound comprises a pathogenic infection, preferably the wound comprises a bacterial infection and / or a fungal infection.

14. The composition for use according to claim 13, wherein the wound includes a bacterial infection.

15. The composition for use according to claim 13 or 14, wherein the wound is a chronic wound.

16. A composition for use according to any one of claims 13 to 15, wherein the wound is a cut, burn, puncture, or ulcer.

17. Use of the composition according to any one of claims 1 to 16 for manufacturing a pharmaceutical product for healing wounds in humans or animals.

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