Disturbance of planktonic bacterial metabolism

Abstract

A composition comprising a 2-hydroxycarboxylic acid for deactivating the metabolism of planktonic bacteria and uses thereof.

Description

Technical Field

[0001] This disclosure relates to compositions comprising preferred enantiomers of 2-hydroxycarboxylic acids for the dysregulation of planktonic bacterial metabolism and the sensitization of planktonic bacteria to antimicrobial compounds, and methods for using the aforementioned compositions to dysregulate the metabolism of planktonic bacteria and sensitize planktonic bacteria to antimicrobial compounds. Background Technology

[0002] In nature, bacteria can alternate between two main growth modes: a unicellular life stage, in which cells move freely (plankton); and a multicellular life stage, in which cells are sessile and live in communities, such as biofilms. The unicellular life stage allows bacteria to spread and colonize new environments, while biofilms allow sessile cells to survive in a coordinated and more persistent manner, which is beneficial for survival and continuous proliferation. In this alternating cycle, bacteria complete two physiological transitions through differential gene expression: (i) from planktonic cells to sessile cells within biofilms, and (ii) from sessile cells to newly detached planktonic cells.

[0003] Because sessile bacteria possess physicochemical conditions and expressed physiological characteristics that facilitate their survival in confined environments such as biofilms, they differ metabolically from planktonic bacteria. Estimates vary, but some results suggest that up to 40% of bacterial genes may be upregulated or downregulated during the transition from a planktonic to a biofilm state, and vice versa. Furthermore, bacteria can develop and initiate drug resistance mechanisms against treatments (e.g., antibiotics and bactericides), thereby disrupting treatment and purification efforts. The drug resistance mechanisms initiated by sessile biofilm bacteria may differ from those initiated by planktonic bacteria.

[0004] The presence of planktonic bacteria can pose risks to healthcare, drinking water distribution systems, the food industry, and the marine industry. For example, in the food industry, pathogenic bacteria can be present inside processing facilities, causing food spoilage and endangering consumer health. In hospital settings, both drug-resistant and non-drug-resistant planktonic bacteria have been shown to persist on the surfaces of medical devices, in patient tissues, or as airborne or fluid-borne single-celled bacteria, causing persistent infections. Furthermore, planktonic bacteria can cause life-threatening infections and diseases in humans and animals, such as sepsis, waterborne or foodborne diarrhea, urinary tract infections, lung infections, otitis media, periodontitis, sinusitis, chronic wounds, eye infections, sexually transmitted diseases, bacterial meningitis and osteomyelitis, and vaginitis. When these bacteria are resistant to disinfectants, they can cause significant healthcare and pollution problems.

[0005] There is a need to eliminate persistent bacterial infections that are prevalent in nature, including those caused by human diseases, crop maintenance, commercial animal farms, animal diseases, and biofouling (including marine biofouling, biofouling of membranes, sensors and pipes, wind turbines, etc.).

[0006] Antimicrobial resistance can lead to a decrease in the efficacy of applied therapeutic agents, which can be antibiotics or agents with different modes of antimicrobial action, such as bacteriophages used to help eliminate planktonic bacteria, preservatives, saline, chlorine-based compounds (e.g., bleach), iodine-based compounds, copper-based compounds, heavy metals, etc.

[0007] Bacterial biofilms are complex bacterial communities that adhere to surfaces and / or bind together through a polymer matrix (primarily composed of polysaccharides, secreted proteins, and extracellular DNA). Biofilm formation can begin with free-floating planktonic microorganisms attaching to surfaces. Once colonization begins, the biofilm grows through a combination of cell division and recruitment. If planktonic bacteria can be removed or killed before biofilm formation, biofilm formation and the persistence of other forms of bacteria can be avoided. Furthermore, since biofilms can release individual bacterial cells to colonize in other spaces where new biofilms can form, it is advantageous to inhibit, disrupt, or kill these released bacteria before they form new biofilms.

[0008] Alternative treatments are needed to help disrupt, inhibit, and / or kill planktonic bacteria; or at least to provide sensitizers to supplement additional bacterial treatments, which can prevent or disrupt the formation of antibiotic resistance mechanisms initiated by the bacteria. This disclosure aims to provide an improved or alternative method for disrupting, inhibiting, and killing planktonic bacteria, particularly a method that can be used in combination with antimicrobial compounds, such as antibiotics or bactericides.

[0009] The preceding discussion of the background art is intended only to facilitate understanding of this disclosure. This discussion is not an admission or endorsement that any material mentioned was or was part of common general knowledge at the priority date of this application. Summary of the Invention

[0010] This disclosure provides a composition for treating metabolic disorders in planktonic bacteria, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid.

[0011] This disclosure also provides a composition for treating metabolic disorders of planktonic bacteria, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid and an antimicrobial compound.

[0012] Dysregulation of planktonic bacterial metabolism (e.g., as measured by, for example, a decrease in antimicrobial MIC, an increase in in vivo antimicrobial efficacy, or aerobic respiration in metabolically active cells (reduction of non-fluorescent resalicylate to highly fluorescent halogen)) can lead to sensitization of planktonic bacteria to antimicrobial compounds and / or disruption of planktonic bacteria's resistance to antimicrobial compounds. When antimicrobial compounds are introduced into planktonic bacteria—whether while the preferred enantiomer of 2-hydroxycarboxylic acid is dysregulating (preferably leading to sensitization) the planktonic bacteria's metabolism, or after the preferred enantiomer of 2-hydroxycarboxylic acid has already dysregulated (preferably leading to sensitization) the planktonic bacteria's metabolism—it can also lead to enhanced bactericidal activity or enhanced infection relief. Dysregulation of planktonic bacterial metabolism can be measured as metabolic inhibition, metabolic enhancement, or metabolic inhibition followed by metabolic enhancement.

[0013] This disclosure also provides a composition for the metabolic dysregulation of planktonic bacteria that are not resistant to one or more antimicrobial compounds (i.e., planktonic bacteria sensitive to antimicrobial compounds, referred to as "non-resistant planktonic bacteria"), the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation of the planktonic bacteria and makes the bacteria sensitive to the antimicrobial compounds.

[0014] This disclosure also provides a composition for dysregulation of planktonic bacteria (referred to as "drug-resistant planktonic bacteria") that are resistant to one or more antimicrobial compounds, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces dysregulation of planktonic bacterial metabolism and susceptes the bacteria to the antimicrobial compound.

[0015] This disclosure also provides a composition for sensitizing planktonic bacteria against microbial compounds, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria.

[0016] This disclosure also provides a composition for sensitizing planktonic bacteria to an antimicrobial compound, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid and an antimicrobial compound, wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0017] This disclosure also provides a composition for enhancing the efficacy of antimicrobial compounds in purifying surfaces containing planktonic bacteria, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0018] This disclosure also provides a composition for enhancing the efficacy of an antimicrobial compound in purifying surfaces containing planktonic bacteria, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid and an antimicrobial compound, wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0019] The antimicrobial compound in the composition of the present invention may be added together with a preferred enantiomer of 2-hydroxycarboxylic acid, or after the preferred enantiomer of 2-hydroxycarboxylic acid has been introduced into the planktonic bacteria. Optionally, if the antimicrobial compound is added after the preferred enantiomer of 2-hydroxycarboxylic acid, the planktonic bacteria have already become sensitive to the antimicrobial compound through the preferred enantiomer of 2-hydroxycarboxylic acid.

[0020] In the compositions of the present invention, optionally, dysregulation of planktonic bacterial metabolism leads to bacterial sensitization against microbial compounds. Alternatively, in the compositions of the present invention, dysregulation of planktonic bacterial metabolism may be an inhibition of planktonic bacterial metabolism, or an inhibition of bacterial metabolism followed by an enhancement of bacterial metabolism.

[0021] Planktonic bacteria can be drug-resistant, non-drug-resistant, or a combination of drug-resistant and non-drug-resistant planktonic bacteria.

[0022] The compositions disclosed herein may contain 0.001% to 100% total 2-hydroxycarboxylic acids. The remainder of the composition may contain a carrier, diluent, or excipient and / or other active agents. In one aspect, the percentage of non-preferred enantiomers of the 2-hydroxycarboxylic acids in the composition is less than 20%.

[0023] Antimicrobial compounds can be bactericidal, bacteriostatic, antibiofilm, or preservative-antimicrobial compounds. Antimicrobial compounds can be antibiotics or agents with different antimicrobial modes of action, such as bacteriophages, preservatives, saline solutions, chlorine-based compounds (e.g., bleach), iodine-based compounds, copper-based compounds, heavy metals, etc.

[0024] The compositions of the present disclosure containing preferred enantiomers of 2-hydroxycarboxylic acid can be used to dysregulate the metabolism of planktonic bacteria (e.g., by inhibition) and optionally make planktonic bacteria sensitive to microbial compounds, including biological and non-biological surfaces and free-floating planktonic bacteria in fluids and gases (including air).

[0025] This disclosure also provides a method for treating metabolic disorders in planktonic bacteria, comprising the following steps: i. Preferred enantiomers of 2-hydroxycarboxylic acid for application to planktonic bacteria.

[0026] This disclosure also provides a method for treating metabolic disorders in planktonic bacteria, comprising the following steps: i. Preferred enantiomeric forms of 2-hydroxycarboxylic acid for application to planktonic bacteria The preferred enantiomer of the 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0027] This disclosure also provides a method for making planktonic bacteria sensitive to microbial compounds, comprising the following steps: i. Preferred enantiomeric forms of 2-hydroxycarboxylic acid for application to planktonic bacteria The preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic disorders in planktonic bacteria.

[0028] This disclosure also provides a method for making planktonic bacteria sensitive to microbial compounds, comprising the following steps: i. Preferred enantiomeric forms of 2-hydroxycarboxylic acid for application to planktonic bacteria The preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0029] This disclosure also provides a method for enhancing the effectiveness of antimicrobial compounds in purifying surfaces containing planktonic bacteria, comprising the following steps: i. Preferred enantiomeric forms of 2-hydroxycarboxylic acid for application to planktonic bacteria The preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic disorders in planktonic bacteria.

[0030] This disclosure also provides a method for enhancing the effectiveness of antimicrobial compounds in purifying surfaces containing planktonic bacteria, comprising the following steps: i. Preferred enantiomeric forms of 2-hydroxycarboxylic acid for application to planktonic bacteria The preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0031] This disclosure also provides a method for treating or preventing bacterial infections caused by airborne bacteria, comprising the following steps: i. Preferred enantiomers of 2-hydroxycarboxylic acid applied to the site of infection The preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic disorders in planktonic bacteria.

[0032] This disclosure also provides a method for treating or preventing bacterial infections caused by airborne bacteria, comprising the following steps: i. Preferred enantiomers of 2-hydroxycarboxylic acid applied to the site of infection The preferred enantiomer of the 2-hydroxycarboxylic acid is combined with an antimicrobial compound, and the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria.

[0033] This disclosure also provides a composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid for treating or preventing bacterial infections in a subject, wherein the bacterial infection is caused by planktonic bacteria, and wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0034] This disclosure also provides a composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid in combination with an antimicrobial compound for treating or preventing infection in a subject, wherein the infection is caused by planktonic bacteria, and wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0035] This disclosure also provides a composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid for treating or preventing bacterial infections in a subject, wherein the bacterial infection is caused by planktonic bacteria, wherein the composition sensitizes the planktonic bacteria to an antimicrobial compound, and wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0036] This disclosure also provides a composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid in combination with an antimicrobial compound for treating or preventing infection in a subject, wherein the infection is caused by planktonic bacteria, wherein the composition sensitizes the planktonic bacteria to the antimicrobial compound, and wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0037] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in the manufacture of compositions for the treatment of metabolic disorders in planktonic bacteria.

[0038] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in combination with antimicrobial compounds in the manufacture of compositions for treating metabolic disorders of planktonic bacteria.

[0039] This disclosure also provides the use of preferred enantiomers of 2-hydroxycarboxylic acids for the treatment of metabolic disorders in planktonic bacteria.

[0040] This disclosure also provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in combination with antimicrobial compounds for the treatment of metabolic disorders in planktonic bacteria.

[0041] This disclosure also provides the use of preferred enantiomers of 2-hydroxycarboxylic acids for sensitizing planktonic bacteria against microbial compounds, wherein the preferred enantiomers of 2-hydroxycarboxylic acids induce metabolic dysregulation in planktonic bacteria.

[0042] This disclosure also provides the use of preferred enantiomers of 2-hydroxycarboxylic acids for sensitizing planktonic bacteria to antimicrobial compounds, wherein the preferred enantiomers of 2-hydroxycarboxylic acids are combined with antimicrobial compounds, and wherein the preferred enantiomers of 2-hydroxycarboxylic acids induce metabolic dysregulation in planktonic bacteria.

[0043] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in the manufacture of compositions for making planktonic bacteria sensitive to microbial compounds, wherein the preferred enantiomers of 2-hydroxycarboxylic acids induce metabolic dysregulation in planktonic bacteria.

[0044] This disclosure provides the use of a preferred enantiomer of a 2-hydroxycarboxylic acid in the manufacture of compositions for making planktonic bacteria sensitive to antimicrobial compounds, wherein the preferred enantiomer of the 2-hydroxycarboxylic acid is combined with an antimicrobial compound, and wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria.

[0045] In the above applications, optionally, the dysregulation of planktonic bacterial metabolism is the inhibition of planktonic bacterial metabolism.

[0046] Preferred enantiomers of 2-hydroxycarboxylic acids may be in the same composition as the antimicrobial compound or in a separate composition. The antimicrobial compound may be a bactericidal compound, a bacteriostatic compound, an antibiofilm compound, or a preservative-antimicrobial compound. The antimicrobial compound may be a bactericidal or bacteriostatic antibiotic, or an agent with a different antimicrobial mode of action, such as a bacteriophage, an antibiofilm agent, a quorum sensing inhibitor, a preservative, saline solution, a chlorine-based compound (e.g., a bleach), an iodine-based compound, a copper-based compound, or a heavy metal.

[0047] This disclosure provides a kit for inhibiting the metabolism of planktonic bacteria, comprising: a) Preferred enantiomers of 2-hydroxycarboxylic acids; and b) Instructions for use.

[0048] This disclosure provides a kit for inhibiting the metabolism of planktonic bacteria, comprising: a) Preferred enantiomers of 2-hydroxycarboxylic acids; and b) Instructions for use The preferred enantiomer of the 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0049] This disclosure provides a kit for sensitizing planktonic bacteria against microbial compounds, comprising: a) Preferred enantiomers of 2-hydroxycarboxylic acids; and b) Instructions for use.

[0050] The preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic disorders in planktonic bacteria.

[0051] This disclosure provides a kit for sensitizing planktonic bacteria against microbial compounds, comprising: a) Preferred enantiomers of 2-hydroxycarboxylic acids; and b) Instructions for use The preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0052] In the methods, uses, and kits of the present invention, the antimicrobial compound may be added together with a preferred enantiomer of the 2-hydroxycarboxylic acid, or added after the preferred enantiomer of the 2-hydroxycarboxylic acid has been introduced into the planktonic bacteria. Optionally, if the antimicrobial compound is added after the preferred enantiomer of the 2-hydroxycarboxylic acid, then drug-resistant and / or non-drug-resistant planktonic bacteria are already sensitive to the antimicrobial compound via the preferred enantiomer of the 2-hydroxycarboxylic acid.

[0053] In the methods, uses, and kits of the present invention, optionally, dysregulation of planktonic bacterial metabolism leads to bacterial sensitization to antimicrobial compounds. Alternatively, in the methods, uses, and kits of the present invention, dysregulation of planktonic bacterial metabolism may be an inhibition of planktonic bacterial metabolism, or an inhibition of bacterial metabolism followed by an enhancement of bacterial metabolism.

[0054] Planktonic bacteria can be drug-resistant, non-drug-resistant, or a combination of drug-resistant and non-drug-resistant planktonic bacteria.

[0055] In the above embodiments, the preferred enantiomeric form of 2-hydroxycarboxylic acid is D-lactic acid or a pharmaceutically acceptable salt thereof. Attached Figure Description

[0056] Other features of this disclosure are described more fully in the following description of several non-limiting embodiments thereof. This description is included for illustrative purposes only and should not be construed as a limitation of the broad overview, disclosure, or description of this disclosure as described above. The description will be made with reference to the accompanying drawings, in which: Figures 1A-1B This involves adding D-lactic acid at a concentration reduced by half to the mixed phenotype. Pseudomonas aeruginosa (P.) aeruginosa) A graph showing the results from the planktonic culture of the clinical isolates. Figure 1A , Figure 1C and Figure 1E The results of culture absorbance measurements as an indicator of suspended bacterial density are shown. As can be seen from each histogram, treatment did not lead to a significant decrease in culture absorbance within the assessed D-lactic acid concentration range. However, Figure 1B , Figure 1D and Figure 1F The inflection point between 0.25 mg / mL and 0.5 mg / mL D-lactic acid is shown when the metabolic rate (resazurin reduction to Resorufin) is used as the reading. Figure 1AThe image shows smooth colony morphology after treatment with the indicated D-lactic acid concentration. Pseudomonas aeruginosa Absorbance (culture density) of clinical isolates (n=8). Figure 1B This demonstrates adding razor violet to the same species with a smooth colony morphology. Pseudomonas aeruginosa (P.) aeruginosa) Relative fluorescence measured after clinical isolates (n=8). The metabolic breakpoints of these cultures were evident between treatments with 0.25 mg / mL and 0.5 mg / mL D-lactic acid. Figure 1C The images show rough colony morphology after treatment with the indicated D-lactic acid concentration range. Pseudomonas aeruginosa Absorbance (culture density) of clinical isolates (n=5). Figure 1D This demonstrates adding razor violet to the same species with a rough colony morphology. Pseudomonas aeruginosa Relative fluorescence was measured after clinical isolates (n=5). The inflection point of the metabolic response in these cultures was evident between 0.25 mg / mL and 0.5 mg / mL D-lactic acid treatment. Figure 1E The images show the slime-like colony morphology exhibited after treatment with the indicated D-lactic acid concentration range. Pseudomonas aeruginosa Absorbance (culture density) of clinical isolates (n=11). Figure 1F This demonstrates the addition of razor violet to the same species exhibiting a slime-like colony morphology. Pseudomonas aeruginosa (P.) aeruginosa) Relative fluorescence was measured after clinical isolates (n=11). The breakpoint of the metabolic response was evident between treatments with 0.25 mg / mL and 0.5 mg / mL D-lactic acid.

[0057] Figure 2 This is a graph showing the enhanced metabolic activity of D-lactic acid in the presence of ceftriaxone. A: Golden grapes Cocci (S. aureus) 29213 (MSSA). B: Staphylococcus aureus 510 (MRSA). Relative metabolic activity of ceftriaxone treatment alone is shown in black, while relative metabolic activity of ceftriaxone plus A: 10 mM D-lactic acid or B: 0.1 mM D-lactic acid is shown in gray.

[0058] Figure 3 This is a graph showing the enhanced metabolic activity of D-lactic acid in the presence of meropenem. A: Staphylococcus aureus 29213 (MSSA). B: Staphylococcus aureus 510 (MRSA). Relative metabolic activity of meropenem treatment alone is shown in black, and relative metabolic activity of meropenem plus 10 mM D-lactic acid is shown in gray.

[0059] Figure 4This is a graph showing the enhanced metabolic activity of D-lactic acid in the presence of methicillin. A: Golden grape balls Bacteria (S. aureus) 29213 (MSSA). B: Staphylococcus aureus 510 (MRSA). Relative metabolic activity of methicillin alone is shown in black, and relative metabolic activity of methicillin plus 10 mM D-lactic acid is shown in gray.

[0060] Figure 5 This is a graph showing the enhanced metabolic activity of D-lactic acid in the presence of rifampicin. A: Staphylococcus aureus (S. aureus) MSSA293 (MSSA). B: Staphylococcus aureus 510 (MRSA). Relative metabolic activity of rifampicin treatment alone is shown in black, and relative metabolic activity of rifampicin plus 0.16 mM D-lactic acid is shown in gray.

[0061] Figure 6 This shows that D-lactic acid in the presence of tetracycline... Staphylococcus aureus Chart of enhanced metabolic activity inhibition in 510 (MRSA). Relative metabolic activity of tetracycline treatment alone is shown in black, and relative metabolic activity of tetracycline plus 10 mM D-lactic acid is shown in gray.

[0062] Figure 7 This is a graph showing the enhanced metabolic activity of D-lactic acid in the presence of vancomycin. A: Golden grape balls Bacteria (S. aureus) 29213 (MSSA). B: Staphylococcus aureus 510 (MRSA). Relative metabolic activity of vancomycin treatment alone is shown in black, while relative metabolic activity of vancomycin plus A: 10 mM D-lactic acid or B: 1.0 mM D-lactic acid is shown in gray.

[0063] Figure 8 This shows that D-lactic acid in the presence of antibiotics... Klebsiella pneumoniae ( K. pneumoniae (Charts showing the enhancement of metabolic activity inhibition in WACC 790. A: Ciprofloxacin. B: Colistin. Relative metabolic activity of antibiotic treatment alone is shown in black, and relative metabolic activity of antibiotic plus A: 0.1 mM D-lactic acid or B: 10 mM D-lactic acid is shown in gray.)

[0064] Figure 9 D-lactic acid in the presence of antibiotics Streptococcus pneumoniae Chart showing the enhancement of metabolic activity inhibition in D39. A: Amoxicillin. B: Ampicillin. Relative metabolic activity of antibiotic treatment alone is shown in black, and relative metabolic activity of antibiotic plus 10 mM D-lactate is shown in gray.

[0065] Figure 10 D-lactic acid in the presence of antibiotics Streptococcus pneumoniae Chart showing the enhancement of metabolic activity inhibition in D39. A: Ceftriaxone. B: Tetracycline. C: Vancomycin. Relative metabolic activity of antibiotic treatment alone is shown in black, and relative metabolic activity of antibiotic plus 10 mM D-lactic acid is shown in gray.

[0066] Figure 11 This involves adding D-lactic acid or L-lactic acid at a concentration reduced by half. Pseudomonas aeruginosa Histogram of results from WACC91 planar culture.

[0067] Figure 12 This is a graph showing the total bacterial count in the blood of control and treated mice 4 hours post-infection. The results were analyzed using an unpaired t-test (two-tailed). = p<0.05; = p<0.01; ns=not significant) A significant difference was obtained between the control group and the treatment group. The horizontal dashed line indicates the limit of detection.

[0068] Figure 13 Two male CD1 mice were provided for use in col R E. coli Representative images (ventral and dorsal) taken at specified times after Xen14 intraperitoneal injection attack demonstrate that treatment with D-lactate and colistin is more effective than colistin alone.

[0069] Figure 14 This is a graph showing the survival analysis of infected mice. Five out of eight mice in the D-lactate + 1 mg / kg colistin combination group and three out of eight mice in the 1 mg / kg colistin treatment group showed survival in [the following data is missing from the original text]. R E. coli Xen14-infected mice survived until the end of the experiment (72 hours post-infection). However, all control mice and mice treated with D-lactate only died from infection 12 hours post-challenge.

[0070] Invention Description Detailed Implementation

[0071] 2-Hydroxycarboxylic acids (such as lactic acid) are known to have bactericidal properties due to their ability to lower environmental pH and disrupt cell membrane integrity. These antimicrobial properties have been used in food production.

[0072] However, this disclosure unexpectedly reveals that one enantiomer of the enantiomer of 2-hydroxycarboxylic acid (referred to herein as "preferred enantiomer of 2-hydroxycarboxylic acid," "preferred enantiomer," or "preferred 2-hydroxycarboxylic acid") is effective against metabolic dysregulation in planktonic bacteria. This metabolic dysregulation effect is not provided by other enantiomers of the enantiomer of 2-hydroxycarboxylic acid (also referred to as "non-preferred enantiomer of 2-hydroxycarboxylic acid," "non-preferred enantiomer," or "non-preferred 2-hydroxycarboxylic acid"). It has been found that, in some cases, the non-preferred enantiomer is not only inactive in metabolic dysregulation but may also potentially prevent the preferred enantiomer from functioning. Preferably, the dysregulation of planktonic bacterial metabolism is an inhibition of planktonic bacterial metabolism.

[0073] Metabolic dysregulation effects are not bactericidal; it is known that two enantiomers of 2-hydroxycarboxylic acids (e.g., lactic acid) have the same bactericidal activity. In some cases, the bactericidal activity of the two enantiomers is due to their effect on pH. In this disclosure, the concentration of the preferred enantiomer used is lower than the concentration used if the preferred enantiomer were used for its bactericidal activity.

[0074] The term "metabolic dysregulation" encompasses both inhibition and enhancement of bacterial activity. Dysregulation can be caused by changes in aerobic respiration, metabolic activity, inhibition of NADH or NADPH production, changes in bacterial viability, changes in the formation of resistance to antimicrobial compounds (bacteriostatic and bactericidal compounds), and / or changes in bacterial sensitivity to antimicrobial compounds (bacteriostatic and bactericidal compounds).

[0075] It has been found that the addition of a preferred enantiomer of a 2-hydroxycarboxylic acid in the absence of antimicrobial compounds generally induces inhibition of planktonic bacterial metabolism; however, in some cases, it leads to enhancement of metabolism. This bacterial metabolic dysregulation has been found to make planktonic bacteria more sensitive to antimicrobial compounds. Alternatively, planktonic bacterial metabolic dysregulation leads to bacterial sensitization to antimicrobial compounds.

[0076] In some cases, the metabolism of planktonic bacteria remains inhibited when they are simultaneously exposed to both a preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound. In other cases, the metabolism of planktonic bacteria may be enhanced in the presence of both the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound.

[0077] The preferred enantiomer of 2-hydroxycarboxylic acid can cause metabolic dysregulation in planktonic bacteria in one of the following ways: - It inhibits metabolism in the presence of 2-hydroxycarboxylic acid alone, and in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; - It inhibits metabolism in the presence of 2-hydroxycarboxylic acid alone, but enhances metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; - Enhanced metabolism in the presence of 2-hydroxycarboxylic acid alone, and enhanced metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; - It enhances metabolism in the presence of 2-hydroxycarboxylic acid alone, but inhibits metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; - It has no effect on metabolism in the presence of 2-hydroxycarboxylic acid alone, but inhibits metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; - It has no effect on metabolism in the presence of 2-hydroxycarboxylic acid alone, but enhances metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds.

[0078] The bacteria can be exposed to 2-hydroxycarboxylic acid at any time: before, simultaneously with, or after the introduction of the planktonic bacteria into the antimicrobial compound.

[0079] The term "inhibition of planktonic bacterial metabolism" includes: inhibition of bacterial viability, such as inhibition of aerobic respiration, inhibition of mitochondrial metabolic activity, and inhibition of NADH or NADPH production; inhibition of bacterial survivability; inhibition of resistance formation to antimicrobial compounds (bacteriostatic and bactericidal compounds); and / or enhancement of bacterial sensitivity to antimicrobial compounds (bacteriostatic and bactericidal compounds). Inhibition can be measured by comparing bacterial metabolism in the presence of a preferred enantiomer of 2-hydroxycarboxylic acid with that of the same bacteria in the absence of a preferred enantiomer of 2-hydroxycarboxylic acid. If bacterial metabolism is lower in the presence of 2-hydroxycarboxylic acid, planktonic bacterial metabolism is inhibited. The percentage change depends on the metabolic parameter used (e.g., a decrease in resazurin, or a decrease in antimicrobial MIC, or an increase in in vivo antimicrobial efficacy), and can be 1%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100%, or any range thereof.

[0080] The term "enhanced planktonic bacterial metabolism" includes: enhanced bacterial activity, such as enhanced aerobic respiration, enhanced mitochondrial metabolic activity, enhanced NADH or NADPH production; enhanced bacterial viability; and / or enhanced bacterial sensitivity to microbial compounds (antimicrobial and bactericidal compounds). This enhancement can be measured by comparing bacterial metabolism in the presence of a preferred enantiomer of 2-hydroxycarboxylic acid with that of the same bacteria in the absence of the preferred enantiomer. If bacterial metabolism is higher in the presence of 2-hydroxycarboxylic acid, then planktonic bacterial metabolism is enhanced. The percentage change depends on the metabolic parameters used (e.g., a decrease in resveratrol, or a concurrent decrease in antimicrobial MIC, or an increase in in vivo antimicrobial potency), and can be 1%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100%, or any range thereof. Enhancement of planktonic bacterial metabolism preferably occurs after metabolic inhibition has occurred, preferably after the addition of an antimicrobial compound. However, in some cases, enhancement of planktonic bacterial metabolism can occur only upon exposure to a preferred enantiomer of 2-hydroxycarboxylic acid, without the prerequisite of inhibition and / or the presence of an antimicrobial compound.

[0081] This disclosure has revealed that preferred enantiomers of 2-hydroxycarboxylic acids (including D-lactic acid) are small, water-soluble compounds that can be used to inhibit the metabolism of planktonic bacteria. Planktonic bacterial infection and contamination can be treated alone by the preferred enantiomers of 2-hydroxycarboxylic acids, by disrupting the bacteria's metabolic capacity. In this case, the metabolic disturbance is preferably metabolic inhibition.

[0082] Alternatively, preferred enantiomers of 2-hydroxycarboxylic acids can be used in combination with other antimicrobial compounds or therapies (both bactericidal and bacteriostatic antimicrobial compounds). The antimicrobial compound can be an antibiotic or a non-antibiotic antimicrobial compound. The metabolic dysregulation activity of the preferred enantiomers of 2-hydroxycarboxylic acids enhances the bactericidal, bacteriostatic, antibiofilm, or antiseptic efficacy of the antimicrobial compound or therapy. In this case, the metabolic disturbance can be either metabolic inhibition or metabolic enhancement. For example, preferred enantiomers of 2-hydroxycarboxylic acids can cause metabolic inhibition, followed by metabolic enhancement upon introduction of the antimicrobial compound. Alternatively, preferred enantiomers of 2-hydroxycarboxylic acids can cause metabolic inhibition, which is maintained in the presence of the antimicrobial compound. However, in some cases, enhancement of planktonic bacterial metabolism can occur upon exposure to only the preferred enantiomers of 2-hydroxycarboxylic acids, without the prerequisite of inhibition and / or the presence of the antimicrobial compound. In this case, metabolic enhancement followed by metabolic inhibition can occur in the presence of both 2-hydroxycarboxylic acid and the antimicrobial compound.

[0083] In this disclosure, a preferred enantiomer of a 2-hydroxycarboxylic acid can act as an "antimicrobial resistance breaker" (ARB). An ARB is a compound administered in conjunction with an antibiotic to overcome bacterial resistance to antibiotics by enhancing the efficacy of the antimicrobial agent or by inhibiting bacterial resistance mechanisms. If a preferred enantiomer of a 2-hydroxycarboxylic acid acts as an ARB, it can enhance the efficacy of the antibiotic by lowering the minimum inhibitory concentration (MIC); for example, the preferred enantiomer of a 2-hydroxycarboxylic acid can reduce the MIC of the antibiotic to below that of antibiotic monotherapy.

[0084] Without adhering to any particular theory, it is believed that metabolic dysregulation induced by the preferred enantiomer of 2-hydroxycarboxylic acid can enable 2-hydroxycarboxylic acid to act as an ARB and increase bacterial sensitivity to antibiotics, and / or increase the efficacy of antibiotics.

[0085] If the metabolic dysregulation induced by the preferred enantiomer of 2-hydroxycarboxylic acid is a metabolic inhibition, then the role of 2-hydroxycarboxylic acid as an ARB could be to increase the amount of antibiotic retained in bacterial cells by reducing efflux (once the antibiotic is internalized, the bacteria cannot modify it), or through other mechanisms.

[0086] Once bacteria are simultaneously exposed to a preferred enantiomer of a 2-hydroxycarboxylic acid and an antimicrobial compound, the role of the 2-hydroxycarboxylic acid acting as an ARB can be to increase the uptake of the antimicrobial compound or to increase the generation of free radicals if the metabolism of the planktonic bacteria is enhanced.

[0087] As used herein, the term "planktonic bacteria" refers to free-swimming or free-floating bacteria that are not located within a biofilm. Although bacteria are free-swimming or free-floating, this movement or floating can occur on a solid surface. Thus, planktonic bacteria can be present on a surface. However, their presence on a surface does not mean they are within or bound to a biofilm, and planktonic bacteria do not attach to the surface. At the micrometer scale, planktonic habitats for prokaryotes can also include water films around soil particles, oral saliva, intestinal fluid, serum in blood vessels, urine in the bladder and urethra, aqueous compositions flowing through the xylem and / or phloem of plants, surface water on plant leaves, soil matrix, etc. Planktonic bacteria can settle to surfaces and eventually develop into biofilms by attaching and then secreting extracellular matrix; however, the bacteria targeted in this disclosure are those that have not yet formed biofilm characteristics, or individual bacteria released from a biofilm as free-floating bacteria that can settle elsewhere to restart the biofilm process.

[0088] The planktonic bacteria disclosed herein can be bacteria that cannot develop into biofilms and exist in a planktonic form throughout their lives.

[0089] The planktonic bacteria disclosed herein can be bacteria released from biofilm-bound colonies in a spontaneous or induced manner (e.g., due to disruption of the biofilm matrix).

[0090] Compared to bacteria within biofilms, planktonic bacteria typically possess upregulated DNA repair genes (such as bacterial SOS response genes). There are also reports that bacteria released from biofilms and becoming planktonic may retain some or all characteristics of the "parental" population associated with the biofilm, thus exhibiting enhanced antibiotic resistance. These released planktonic bacterial populations can be difficult to remove from sterile environments and often serve as sources of reinfection.

[0091] Composition

[0092] This disclosure provides a composition for treating metabolic disorders in planktonic bacteria, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid, such as D-lactic acid.

[0093] This disclosure also provides a composition for treating metabolic disorders of planktonic bacteria, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid and an antimicrobial compound.

[0094] This disclosure also provides a composition for sensitizing planktonic bacteria against microbial compounds, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria.

[0095] This disclosure also provides a composition for sensitizing planktonic bacteria to an antimicrobial compound, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid and an antimicrobial compound, wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0096] Bacterial metabolic dysregulation can lead to sensitization. Sensitization can occur through enhancement or synergistic effects. In one aspect, dysregulation of planktonic bacterial metabolism is inhibition, in another aspect it is enhancement, and in yet another it is either inhibition or enhancement, followed by the other.

[0097] 2-Hydroxycarboxylic acid

[0098] The 2-hydroxycarboxylic acid of the present invention has a stereochemistry, and preferably an enantiomer of the stereochemistry. The 2-hydroxycarboxylic acid having the preferred stereochemistry disclosed herein is referred to herein as a "preferred enantiomer of 2-hydroxycarboxylic acid," "preferred enantiomer," or "preferred 2-hydroxycarboxylic acid." The 2-hydroxycarboxylic acid is optionally a hydrophilic 2-hydroxycarboxylic acid. In one aspect, a preferred enantiomer of the 2-hydroxycarboxylic acid is D-lactic acid. The preferred 2-hydroxycarboxylic acid of the present invention can be a salt of a preferred 2-hydroxycarboxylic acid, optionally a pharmaceutically acceptable salt.

[0099] In the case where the preferred 2-hydroxycarboxylic acid is not D-lactic acid, the preferred 2-hydroxycarboxylic acid to be used in this disclosure may be: wherein, with regard to the three-dimensional orientation of the hydroxyl group, the absolute stereochemistry of the chiral center corresponding to the hydroxyl group of the 2-hydroxycarboxylic acid corresponds to the absolute stereochemistry of the corresponding chiral center in D-lactic acid, as follows: The preferred 2-hydroxycarboxylic acid of the above general formula can be any of the substituents R selected from the group consisting of: hydrogen, halogen (F, Cl, Br, I), methyl, ethyl, n-propyl, isopropyl, cyclopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, cyclobutyl, n-pentyl, isopentyl, neopentyl, cyclopentyl, n-hexyl, 2-methylpentyl, 3-methylpentyl, 2,3-dimethylbutyl, 2,2-dimethylbutyl, cyclohexyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, 2,3-dimethylpentyl, 2,4-dimethylpentyl, 2,2-dimethylpentyl, 3,3-dimethylpentyl, 3-ethylpentyl, 2,2,3-trimethylbutyl, cycloheptyl, phenyl, benzyl, furanyl, tetrahydrofuranyl, vinyl (ethenyl), vinyl, allyl, crotonyl, isopentenyl, 1-propenyl, 2-propenyl, 1-butenyl, 2-butenyl, 2-methylprop-2-enyl, hexenyl, butadienyl, pentadienyl, hexadienyl, 2,3-dimethyl-2-butenyl, heptenyl, octenyl, octtrienyl, butadienyl, 1,3-pentadienyl, 1,4-pentadienyl, cyclopentadienyl, and cyclohexadienyl, wherein each of the substituents (other than hydrogen and halogen) may be unsubstituted or substituted with one or more substituents selected from the group consisting of: hydrogen, halogen, hydroxyl, methoxy, ethoxy, propoxy, butoxy, carboxylic acid, amide, or ester substituents, and pharmaceutically acceptable salts thereof.

[0100] The preferred base addition salts of 2-hydroxycarboxylic acids of this invention can be prepared from inorganic and organic bases. Salts derived from inorganic bases include (by way of example only) sodium, potassium, lithium, ammonium, calcium, magnesium, copper, zinc, gallium, and silver salts. Salts derived from organic bases include, but are not limited to, salts of primary, secondary, and tertiary amines, such as (by way of example only) alkylamines, dialkylamines, trialkylamines, substituted alkylamines, di(substituted alkyl)amines, tri(substituted alkyl)amines, alkenylamines, dienylamines, trienylamines, substituted alkenylamines, di(substituted alkenyl)amines, tri(substituted alkenyl)amines, cycloalkylamines, di(cycloalkyl)amines, tri(cycloalkyl)amines, substituted cycloalkylamines, disubstituted cycloalkylamines, trisubstituted cycloalkylamines, cycloalkenylamines, di(cycloalkyl)amines, tri(cycloalkyl)amines, substituted cycloalkylamines, disubstituted cycloalkylamines, trisubstituted cycloalkylamines, cycloalkenylamines, di(cycloalkyl)amines, trisubstituted cycloalkylamines, di(cycloalkyl)amines, trisubstituted cycloalkylamines, cycloalkenylamines, di(cycloalkyl) ... Amines, including tri(cycloalkenyl)amines, substituted cycloalkenylamines, disubstituted cycloalkenylamines, trisubstituted cycloalkenylamines, arylamines, diarylamines, triarylamines, heteroarylamines, diheteroarylamines, triheteroarylamines, heterocyclic amines, diheterocyclic amines, triheterocyclic amines, mixed diamines, and triamines, wherein at least two substituents on the amine are different and selected from the group consisting of: alkyl, substituted alkyl, alkenyl, substituted alkenyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, heteroaryl, heterocyclic, etc. Also including amines in which two or three substituents together with the amino nitrogen form a heterocycle or heteroaryl group.

[0101] The preferred acid addition salts of the 2-hydroxycarboxylic acid of this invention can be prepared from inorganic and organic acids. Usable inorganic acids include (by way of example only) hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc. Usable organic acids include (by way of example only) acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, malic acid, malonic acid, succinic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, etc.

[0102] Examples of acceptable salts also include iodides, acetates, phenylacetates, trifluoroacetates, acrylates, ascorbic acid salts, benzoates, chlorobenzoates, dinitrobenzoates, hydroxybenzoates, methoxybenzoates, methylbenzoates, o-acetoxybenzoates, naphthalene-2-benzoates, bromides, isobutyrate, phenylbutyrate, γ-hydroxybutyrate, β-hydroxybutyrate, butyne-1,4-diacidate, hexyne-1,4-diacidate, hexyne-1,6-diacidate, hexanoate, octanoate, chlorides, cinnamates, citrates, decanoates, formates, fumarates, glycolates, heptanoates, hippurate, lactates, and malates. Maleate, hydroxymaleate, malonate, mandelate, methanesulfonate, nicotinate, isonicotinate, nitrate, oxalate, phthalate, terephthalate, phosphate, hydrogen phosphate, dihydrogen phosphate, metaphosphate, pyrophosphate, propynate, propionate, phenylpropionate, salicylate, sebate, succinate, octanoate, sulfate, hydrogen sulfate, pyrosulfonate, sulfite, bisulfite, sulfonate, benzenesulfonate, p-bromobenzenesulfonate, chlorobenzenesulfonate, propanesulfonate, ethanesulfonate, 2-hydroxyethanesulfonate, methanesulfonate, naphthalene-1-sulfonate, naphthalene-2-sulfonate, p-toluenesulfonate, xylenesulfonate, tartrate, etc.

[0103] The compositions disclosed herein may contain a 2-hydroxycarboxylic acid with a non-preferred stereochemistry, such as L-lactic acid. The 2-hydroxycarboxylic acid with a non-preferred stereochemistry disclosed herein is referred to herein as a "non-preferred enantiomer of a 2-hydroxycarboxylic acid," "non-preferred enantiomer," or "non-preferred 2-hydroxycarboxylic acid." The percentage of the non-preferred 2-hydroxycarboxylic acid may be less than 20%. Alternatively, the compositions disclosed herein may not contain a non-preferred 2-hydroxycarboxylic acid. Therefore, the composition may contain only a 2-hydroxycarboxylic acid with a preferred stereochemistry as the 2-hydroxycarboxylic acid.

[0104] In the case where the non-preferred 2-hydroxycarboxylic acid is not L-lactic acid, the non-preferred 2-hydroxycarboxylic acid according to this disclosure is one in which, with respect to the three-dimensional orientation of the hydroxyl group, the absolute stereochemistry of the chiral center corresponding to the hydroxyl group of the 2-hydroxycarboxylic acid corresponds to the absolute stereochemistry of the corresponding chiral center in L-lactic acid, as shown below: Typically, most lactic acid bacteria primarily produce L-lactic acid. Without adhering to any single theory, it is believed that many bacteria have evolved to avoid or denature L-lactic acid to prevent the effects of organic acids on the bacterial membrane. Without adhering to any single theory, it is possible that bacteria are less able to avoid or denature the D-isomer of lactic acid, thus enabling them to have metabolic disruptive effects.

[0105] High-purity L-lactic acid can be produced by a variety of microorganisms, such as bacteria, fungi, algae and cyanobacteria. On the other hand, most microorganisms that produce D-lactic acid either produce racemic mixtures or other organic acids, such as acetic acid or succinic acid (Alexandri et al. (2019) Food Tech & Biotech 57(3):293-304).

[0106] The compositions disclosed herein may contain 0.001% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid, such as D-lactic acid. The compositions may contain 0.001%, 0.01%, 0.1%, 1%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, diluent, or excipient and / or other active agent. For example, ophthalmic compositions may contain 0.001% to 1% of a preferred enantiomer of a 2-hydroxycarboxylic acid; wash and soak, dressing and wipe, topical, inhalation, or oral compositions may contain 0.2% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid.

[0107] The compositions disclosed herein may comprise a non-preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the percentage of the non-preferred enantiomer of the 2-hydroxycarboxylic acid is less than 50%, 40%, 30%, 20%, 10%, 5%, or 1%. For example, the percentage of the non-preferred enantiomer of the 2-hydroxycarboxylic acid is less than 20%, 18%, 16%, 14%, 12%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1%. The percentage of the non-preferred enantiomer of the 2-hydroxycarboxylic acid may be less than 20%.

[0108] Alternatively, the compositions of this disclosure may not contain any non-preferred enantiomers of 2-hydroxycarboxylic acids. Therefore, the compositions may contain only preferred enantiomers of 2-hydroxycarboxylic acids as the 2-hydroxycarboxylic acid. For example, the compositions may contain only D-lactic acid as lactic acid. It has been found that compositions containing enantiomerically pure amounts of preferred enantiomers of 2-hydroxycarboxylic acids have metabolic dysregulatory effects on planktonic bacteria, compositions containing enantiomerically pure amounts of non-preferred enantiomers of 2-hydroxycarboxylic acids have little to no metabolic dysregulatory effect, and compositions containing a 1:1 ratio of preferred enantiomers of 2-hydroxycarboxylic acids to non-preferred enantiomers of 2-hydroxycarboxylic acids have reduced metabolic dysregulatory effects or no metabolic dysregulatory effects on planktonic bacteria compared to the application of pure preferred enantiomers of 2-hydroxycarboxylic acids.

[0109] The compositions disclosed herein optionally comprise an enantiomeric enrichment of a preferred enantiomeric form of a 2-hydroxycarboxylic acid. The compositions may comprise an enantiomeric pure preferred enantiomeric form of a 2-hydroxycarboxylic acid.

[0110] Optionally, the concentration of the preferred enantiomer of 2-hydroxycarboxylic acid used in the compositions of this disclosure is less than the concentration of the preferred enantiomer of 2-hydroxycarboxylic acid required to kill bacteria.

[0111] Without adhering to any particular theory, this disclosure requires a 2-hydroxycarboxylic acid with a chiral center at position 2. It should be understood that if an aliphatic chain has at least three carbons in length, the chiral center is located at position 2; by this definition, glycolic acid [2-hydroxyacetic acid] is not chiral because its length is less than three carbons.

[0112] 2-Hydroxycarboxylic acids can be hydrophilic. That is, 2-hydroxycarboxylic acids can optionally be soluble in water. Without adhering to any particular theory, it is believed that the water solubility of 2-hydroxycarboxylic acids contributes to their ability to penetrate planktonic bacterial films and their metabolically disruptive activities.

[0113] It has been found that 2-hydroxycarboxylic acids, which typically have seven or fewer carbons (including the carbon in the hydroxycarboxylic acid moiety), are more soluble in water. Therefore, optionally, the 2-hydroxycarboxylic acids of this disclosure contain seven or fewer carbons. For example, 2-hydroxypropionic acid (also known as lactic acid) contains three carbons, 2-hydroxypentanoic acid (also known as 2-hydroxyvaleric acid) contains five carbons, and 2-hydroxycaproic acid (also known as 2-hydroxyhexanoic acid) contains six carbons. For example, the 2-hydroxycarboxylic acids of this disclosure contain six or fewer carbons. In one example, the 2-hydroxycarboxylic acids of this disclosure contain five or fewer carbons.

[0114] Optionally, the molecular weight of the 2-hydroxycarboxylic acid is less than 800, less than 700, less than 600, or less than 500. For example, the molecular weight of the 2-hydroxycarboxylic acid is less than 200, less than 180, less than 150, or less than 100. Alternatively, the absolute water solubility of 2-hydroxycarboxylic acid is greater than about 0.5 g / L, 0.75 g / L, or 1.0 g / L at pH values ​​of about 5 to 10, about 9 to 6, or about 6 to 8.

[0115] Alternatively, at a temperature of about 20°C, the absolute water solubility of 2-hydroxycarboxylic acid is greater than about 0.5 g / L, 0.75 g / L, or 1.0 g / L.

[0116] Alternatively, the water solubility of 2-hydroxycarboxylic acids can be determined by their Log... P Definition: This refers to its octanol-water partition coefficient. (Log) P Also known as LogK ow or n -Octanol-water partition ratio. The general rule is Log POctyl alcohol - water < 1.0 = water-soluble; Log P Octyl alcohol-water > 1.0 = hydrophobic. Alternatively, the Log of the 2-hydroxycarboxylic acid disclosed herein... P Octyl alcohol-water < 1.0. Log P Some compounds with a carbon content <1.0 can have more than seven carbons; however, despite the additional carbons, their structure allows for water solubility. For example, 2-hydroxy-2-phenylacetic acid (also known as mandelic acid) has eight carbons, but due to its cyclic structure, its Log0 is much lower than that of other compounds. P Its value is 0.67, therefore it is soluble in water.

[0117] Some 2-hydroxycarboxylic acids with special structures, such as 2-hydroxy-2-phenylacetic acid (also known as mandelic acid), which contains a six-membered ring and a total of eight carbons, can still be water-soluble and have a logarithmic property. P Octyl alcohol-water < 1.0. Such 2-hydroxycarboxylic acids are considered suitable for this disclosure.

[0118] 2-Hydroxycarboxylic acids can have substituents, such as hydrogen, halogen, hydroxyl, methoxy, ethoxy, propoxy, butoxy, carboxylic acid, amide, or ester substituents.

[0119] 2-Hydroxycarboxylic acids can be pharmaceutically acceptable 2-hydroxycarboxylic acids. For example, 2-hydroxycarboxylic acids are GRAS (“Generally Recognized As Safe”), as defined in Sections 201(s) and 409 of the United States Federal Food, Drug, and Cosmetic Act or equivalent regulations.

[0120] 2-Hydroxycarboxylic acids may be selected from acids or esters, salts, amides or other derivatives of the group consisting of: lactic acid, glycolic acid (2-hydroxyacetic acid), tartaric acid (2,3-dihydroxysuccinic acid), mandelic acid, 1-hydroxycyclohexane-1-carboxylic acid, 2-hydroxy-2-(tetrahydrofuran-2-yl)acetic acid, 2-hydroxy-2-(2-furanyl)acetic acid, 2-hydroxy-2-phenylpropionic acid, 2-hydroxy-2-methylpropionic acid, 2-hydroxy-2-methylbutyric acid, 2-hydroxybutyric acid, 2-hydroxyvalerate, or mixtures thereof.

[0121] 2-Hydroxycarboxylic acids may be selected from the following list: 2-hydroxypropionic acid (also known as lactic acid), 2-hydroxypentanoic acid (also known as 2-hydroxyvaleric acid), 2-hydroxybutyric acid, 2-hydroxyacetic acid (also known as glycolic acid), 2-hydroxyhexanoic acid (also known as 2-hydroxycaproic acid), and 2-hydroxy-2-phenylacetic acid (also known as mandelic acid).

[0122] 2-Hydroxycarboxylic acids may be selected from the following list: D-lactic acid, 2-hydroxyvalerate, 2-hydroxybutyric acid, 2-hydroxyhexanoic acid, (2)-hydroxyphenylacetic acid, and 2-hydroxy-2-phenylacetic acid. Preferred 2-hydroxycarboxylic acids include those in Table 1 and / or Table 2.

[0123] Table 1: Solubility of selected 2-hydroxycarboxylic acids

[0124] Table 2: Solubility of selected 2-hydroxycarboxylic acids

[0125] Optionally, the 2-hydroxycarboxylic acid has fewer than seven carbon atoms and Log P Octyl alcohol-water < 1.0.

[0126] Optionally, the 2-hydroxycarboxylic acid disclosed herein is lactic acid, which has two enantiomers. Optionally, the preferred enantiomer of the 2-hydroxycarboxylic acid is D-lactic acid, also known as (2R)-2-hydroxypropionic acid and (R)-lactic acid.

[0127]

[0128] Preferred enantiomers of 2-hydroxycarboxylic acids can be synthesized via: cell-free pathways (e.g., enzyme-catalyzed reactions); or from lactic acid bacteria, including bacteria such as... Lactobacillus species or Bifidobacterium (Bifidobacterium) The species; derived from the supernatant of cultured bacteria (e.g., multiple strains of Gram-positive and / or Gram-negative probiotics); derived from natural sources, such as 2-hydroxy-2-phenylacetic acid (mandelic acid) from almonds, olive oil and beer; or derived from other starchy biomass.

[0129] If the preferred enantiomer of 2-hydroxycarboxylic acid is D-lactic acid, then D-lactic acid can be synthesized via: cell-free pathways (e.g., enzyme-catalyzed reactions), or from... LactobacillusOr from the supernatant derived from cultured bacteria (e.g., multiple strains of Gram-negative probiotics).

[0130] The D-lactic acid disclosed herein is optionally a monomer. Therefore, D-lactic acid is optionally not polylactic acid (also known as PLA, poly-D-lactic acid [PDLA], or poly-L-lactic acid [PLLA]) or similar polymers. PLA is a polymer obtained by ring-opening polymerization of the monomer lactide (a cyclic dimer of lactic acid).

[0131] Antimicrobial compounds

[0132] The method disclosed herein also provides a preferred enantiomer of a 2-hydroxycarboxylic acid, which is co-administered in combination with an antimicrobial compound, wherein the preferred enantiomer of the 2-hydroxycarboxylic acid enhances the activity of the antimicrobial compound against planktonic bacteria. The preferred enantiomer of the 2-hydroxycarboxylic acid may be in the same composition as the antimicrobial compound or may be in a separate composition. Planktonic bacteria can cause infection in a subject, in or on plants or in the soil in which plants grow, or may be present on surfaces, such as abiotic surfaces. Planktonic bacteria can be freely floating in liquids or gases.

[0133] The term "antimicrobial compound" refers to a compound that can kill or reduce the viability of bacteria. Antimicrobial compounds can be antibiotics or agents with different modes of antimicrobial action, such as bacteriophages, preservatives, saline solutions, chlorine-based compounds (e.g., bleach), iodine-based compounds, copper-based compounds, or heavy metals. The term includes bactericidal compounds, bacteriostatic compounds, antibiofilm compounds, or preservative-antimicrobial compounds. Antimicrobial compounds can be antibiofilm compounds.

[0134] The term "antibiotic" refers to antimicrobial compounds that have antibacterial activity and are used for prevention or treatment in medical or veterinary settings. They can kill bacteria (bactericidal) or inhibit bacterial growth (bacteriostatic). Antibiotics can be of natural origin (compounds obtained by screening bacteria and fungi for antibiotic compounds; e.g., penicillin) or of laboratory origin (compounds obtained by chemically identifying the target antibiotic compound; e.g., sulfamethoxazole). Antibiotics can be derivatives of compounds of natural or laboratory origin.

[0135] If the antimicrobial compound administered with the preferred enantiomer of the 2-hydroxycarboxylic acid of this disclosure is an antibiotic, it may be selected from one or more of the following categories:

[0136] The term "sensitization" refers to the enhanced effect of an antimicrobial compound on planktonic bacteria, for example, by lowering the minimum inhibitory concentration (MIC) of the antimicrobial compound. The MIC is the lowest antibiotic concentration at which visible bacterial growth is inhibited. Sensitization can occur through either enhancement or synergistic effects.

[0137] Preferably, the planktonic bacteria are resistant to one or more antimicrobial compounds.

[0138] As used herein, the term "synergistic effect" refers to the combined action of a 2-hydroxycarboxylic acid and an antimicrobial compound (e.g., an antibiotic) to produce a more potent effect than each active substance when administered alone. For example, in this disclosure, a synergistic effect can occur when the combined effect of the 2-hydroxycarboxylic acid and the antimicrobial compound on planktonic bacteria is greater than the sum of their effects when administered alone. This synergistic effect can occur if the planktonic bacteria are sensitive to the antimicrobial compound (e.g., by enhancing the effect of the antimicrobial compound through lowering the MIC, etc., so that less antimicrobial compound can be used to achieve the same effect), or if the planktonic bacteria are resistant to the antimicrobial compound (e.g., by enhancing the effectiveness of the antimicrobial compound against planktonic bacteria by lowering the MIC to an effective value that allows for safe, easy, and cost-effective administration, so that the antimicrobial compound can now be used effectively in cases where it was previously ineffective due to resistance).

[0139] The synergistic effect in combinations of preferred enantiomers of 2-hydroxycarboxylic acids and antimicrobial compounds can be measured using a simple plate assay, in which the compounds are applied to separate paper discs and diffused onto agar plates inoculated with target bacteria. The synergistic effect manifests as enhanced bactericidal activity at the boundary of the diffusion zone. Quantitative measurement of the synergistic effect can be performed using a simple checkerboard strategy, in which the concentrations of both reagents are systematically diluted to determine the concentrations of the two drugs that achieve the strongest interaction.

[0140] Synergistic effects are typically measured by calculating the fractional inhibitory concentration index (FICI), where: FICI = (MIC AB / MIC A ) + (MIC BA / MICM B ) MIC AB It is the minimum inhibitory concentration (MIC) of drug A during the combined test. A It is the MIC of drug A when tested alone. BA It is the MIC of drug B during the combined test, and the MIC BThis is the MIC of drug B when tested alone. FICI ≤ 0.50 is defined as synergistic effect (i.e., MIC reduced by 4-fold), FICI of 0.50 to 1.00 represents additive effect, and FICI of 1.00 to 4.00 is defined as no difference. FICI above 4.00 represents antagonistic effect.

[0141] However, it can be difficult to determine the FICI value of the current combination of a preferred enantiomer of a 2-hydroxycarboxylic acid and an antimicrobial compound, because the 2-hydroxycarboxylic acid itself does not have antimicrobial activity but merely enhances the activity of the antimicrobial compound. In other cases, the antimicrobial compound may not have a MIC in the absence of a preferred enantiomer of the 2-hydroxycarboxylic acid. When neither the preferred enantiomer of the 2-hydroxycarboxylic acid nor the antimicrobial compound has a measurable MIC, the concentration of the 2-hydroxycarboxylic acid that determines the MIC of the antimicrobial compound can be used. In one example, the concentration of the 2-hydroxycarboxylic acid that reduces the MIC of the antimicrobial compound can be used as an indicator of potency. For example, a four-fold reduction in MIC can be used as an indicator of potency (i.e., a four-fold reduction in MIC). More relevantly, the concentration of the preferred enantiomer of the 2-hydroxycarboxylic acid reduces the MIC of the antimicrobial compound in resistant bacteria to equal or below the breakpoint concentration, i.e., the concentration of the antimicrobial compound that defines whether the strain is resistant or susceptible. Therefore, the rescue concentration (RC) is defined as the concentration of 2-hydroxycarboxylic acid required to reduce the effective concentration of an antimicrobial compound to less than or equal to the breakpoint (Wright et al. (2016) Trends Microbiol. 24(11):862-871).

[0142] As used herein, the term "enhancing effect" refers to the ability of a 2-hydroxycarboxylic acid to enhance the activity of an antimicrobial compound (e.g., an antibiotic). For example, in this disclosure, an enhancing effect can occur when planktonic bacteria are resistant to an antimicrobial compound (and therefore the antimicrobial compound alone has no effect, i.e., no MIC), but when the antimicrobial compound is combined with a 2-hydroxycarboxylic acid, the planktonic bacteria's response to the antimicrobial compound is enhanced (i.e., the antimicrobial compound has a more significant effect on the bacteria, whether bacteriostatic or bactericidal, i.e., the MIC can be determined at this point).

[0143] Preferred enantiomers of the 2-hydroxycarboxylic acid disclosed herein can combat bacterial resistance through several mechanisms, such as increasing the uptake of antimicrobial compounds, hindering the efflux of antimicrobial compounds, disrupting signal transduction pathways, preventing the modification of antimicrobial compounds, and / or preventing the modification of antimicrobial compound targets.

[0144] If the dysregulation of bacterial metabolism is a metabolic inhibition caused by a preferred enantiomer of the 2-hydroxycarboxylic acid of this disclosure, such metabolic inhibition can function, for example, by slowing the efflux of the antimicrobial compound (because metabolism, including efflux pumps, is inhibited) to retain the antimicrobial compound in the planktonic bacteria. Alternatively, inhibition can also result in the retention of the antimicrobial compound in the planktonic bacteria by slowing the rate at which the antimicrobial compound is modified (because metabolism, including enzyme production and activity, is inhibited).

[0145] If the bacterial metabolic dysregulation is an enhanced metabolism caused by a preferred enantiomer of the 2-hydroxycarboxylic acid of this disclosure, this enhancement can function, for example, by increasing the uptake of antimicrobial compounds. Alternatively, the enhanced metabolism can lead to increased free radical production, thereby making the bacterial cells more sensitive to antimicrobial compounds.

[0146] Optionally, the pH of the compositions disclosed herein is from about 6.5 to 8.0, for example, about 7.0 and 7.4. It has previously been found that bacterial resistance to antimicrobial therapies increases as pH decreases. A preferred pH helps prevent bacteria from developing resistance to the effects of a preferred enantiomer of 2-hydroxycarboxylic acid. Buffers can be added to adjust the pH level of the composition. Optionally, the compositions disclosed herein contain tris(hydroxymethyl)aminomethane (TRIS, also known as THAM or tromethamine) or phosphate-buffered saline (PBS) as a buffer. The potency of the preferred enantiomer of 2-hydroxycarboxylic acid at a pH of about 6.5 to 8.0 further indicates that the effect on bacterial metabolism is a result of the metabolically dysregulatory activity of 2-hydroxycarboxylic acid itself, and not merely a bactericidal effect of low pH.

[0147] Without adhering to any particular theory, it is believed that the preferred enantiomers of the 2-hydroxycarboxylic acid disclosed herein function by dysregulating the metabolic rate of planktonic bacteria. Dysregulation can be either inhibition or enhancement; or one of inhibition or enhancement followed by inhibition or enhancement of the other.

[0148] Metabolic inhibition can lead to increased sensitivity of planktonic bacteria to antimicrobial compounds. Metabolic inhibition is distinct from the activity of any antimicrobial compound and can occur in the absence of any other antimicrobial compound.

[0149] Enhanced metabolism can lead to increased susceptibility of planktonic bacteria to antimicrobial compounds. Metabolic enhancement is distinct from the activity of any antimicrobial compound and can occur in the absence of any other antimicrobial compound. However, it typically occurs in the presence of both a preferred enantiomer of the 2-hydroxycarboxylic acid and the antimicrobial compound. In some cases, however, enhanced planktonic bacterial metabolism can occur upon exposure to only the preferred enantiomer of the 2-hydroxycarboxylic acid, without the prerequisite of inhibition and / or the presence of an antimicrobial compound. In such cases, metabolic enhancement in the presence of both the 2-hydroxycarboxylic acid and the antimicrobial compound can be followed by metabolic inhibition.

[0150] When the metabolism of planktonic bacteria is disrupted, the bacteria may undergo one or more of the following processes: - Increased susceptibility of bacteria to host immune system factors, other microorganisms, or other antimicrobial compounds (including but not limited to antibiotics and bacteriophages); - Bacteria develop resistance to antimicrobial compounds (whether from external sources or from the host); - Decreased cell membrane potential or impaired cell membrane integrity; and / or - Bacterial growth slows down.

[0151] Dysregulation of bacterial metabolism can be measured as a decrease in the MIC (enhanced potency) of antimicrobial compounds, alterations in respiratory metabolism (measured by resazurin reduction), or any other measure of bacterial metabolism. All measurements can be performed in vitro or in vivo. The extent of the metabolic dysregulation must be quantifiable, preferably in a clinical setting, and must be clinically significant. Regardless of how the dysregulation is measured, bacterial metabolic dysregulation should result in bacterial sensitization to antimicrobial compounds and / or a decrease in the MIC of antimicrobial compounds.

[0152] Optionally, the treatment regimen of this disclosure induces bacterial metabolic dysregulation in the form of at least two-fold metabolic inhibition, as determined by, for example, the reduction of resazurin to halogen. Alternatively, the metabolic dysregulation of planktonic bacteria can be measured by sensitization to an antimicrobial agent, wherein sensitization is a reduction of the MIC or bMIC concentration of the antimicrobial agent by at least 20%, 30%, or 50%.

[0153] Optionally, the treatment regimen of this disclosure induces bacterial metabolic dysregulation in the form of at least a twofold increase in metabolism, as determined by, for example, the reduction of resazurin to halogen. Alternatively, the metabolic dysregulation in the form of increased metabolism in planktonic bacteria can be measured by sensitization to an antimicrobial agent, wherein sensitization is a reduction in the MIC or bMIC concentration of the antimicrobial agent by at least 20%, 30%, or 50%.

[0154] airborne bacteria

[0155] Planktonic bacteria can be Gram-positive or Gram-negative. For example, planktonic bacteria can be from the Enterobacteriaceae family, or can be selected from the following genera: Bacillus species, Clostridium species, Campylobacter (Campylobacter) species, Pseudomonas species, Streptococcus species, Actinobacillus species, Species of the genus *Staphylococcus* and *Escherichia coli* (Escherichia) species, Acinetobacter species, Klebsiella species, Aeromonas species, Enterococcus species, Legionella The species, and Salmonella species, Species of the genus Shigella, genus Gardnerella (Gardnerella) species, Haemophilus genus species, Helicobacter species, Mora Moraxella species, Mycobacterium species, Neisseria species, Anaerococcus species, Atopobium (genus) species, Bacteroides species, Leptotrichia species, Mobiluncus species, Peptostreptococcus (Peptostreptococcus) species, Prevotella species, Sneathia species, Sphingomonas species, Nitrospira species, Mycobacterium (Mycobacterium) species or Hyphomicrobium species and Vibrio species 。 Planktonic bacteria can be cyanobacteria or nontuberculous mycobacteria (NTM).

[0156] Planktonic bacteria can be selected from the following bacteria: Pseudomonas aeruginosa Streptococcus pneumoniae, Streptococcus mutans, pleuropneumoniae Actinobacillus pleuropneumoniae and Staphylococcus epidermidis Epidermidis, Escherichia coli, Staphylococcus aureus aureus) , Acinetobacter baumannii and Aeromonas hydrophila Hydrophila, Aeromonas salmonicida, Clostridium difficile difficile, Enterococcus faecium, Gardnerella vaginalis vaginalis) , Haemophilus influenzae and Helicobacter pylori pylori, Moraxella bovis , Moraxella catarrhalis, bird branch Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium tuberculosis Mycobacterium tuberculosis, Neisseria gonorrhoeae, Neisseria meningitidis Neisseria meningitidis and Streptococcus pyogenes and Vibrio cholerae (Vibrio cholerae).

[0157] For example, Bacillus species, Clostridium species and Pseudomonas (Pseudomonas) These substances are often found in natural gas and oil in pipelines. Sphingosomalidosis (Sphingomonas) species, Nitrospira species, Legionella species, branch Mycobacterium species or Hyphomicrobium These species are often found in water pipes and air in air conditioning units. Eubacteria (including cyanobacteria) can exist in seawater and, once attached, can form the basis for biofilm fouling on ship hulls.

[0158] Bacterial vaginosis can be caused by Gardnerella vaginalis Other taxa (e.g.) Anaerobic cocci genus Anaerococcus, Atopobium, Bacteroides , genus *Ciliophytum* (Leptotrichia) , Mobiluncus, Peptostreptococcus, Prevotella Prevotella and Sneathia and Clostridium It is caused by or associated with metabolic disorders and can be treated with the metabolic disorder compositions disclosed herein.

[0159] Planktonic bacteria can be antibiotic-resistant strains of the bacteria listed above. For example, planktonic bacteria can be antibiotic-resistant... Pseudomonas aeruginosa or antibiotic resistance Staphylococcus aureus (Staphylococcus aureus) (e.g., MRSA).

[0160] Planktonic diarrhea pathogens are usually transmitted through the "oral-fecal route," which includes contaminated drinking water, ice in cocktails (or soft drinks), and inadequate handwashing.

[0161] Planktonic bacteria can be associated with symptoms or diseases such as sepsis, cystic fibrosis-associated pneumonia, periodontitis, diabetic ulcers and other wound infections, otitis media, periodontitis, sinusitis, infective endocarditis, eye infections, glue ear, dysentery, respiratory infections, sexually transmitted diseases (STDs), bacterial vaginosis, and osteomyelitis.

[0162] Alternatively, planktonic bacteria can be associated with industrial and non-biological surfaces, such as hydroelectric turbines, oil and gas pipelines, air conditioning units, soap dispensers, industrial water systems, and ship hulls.

[0163] In other examples, airborne bacteria can be associated with medical devices and instruments, such as bags for storing donated blood and indwelling medical devices.

[0164] Planktonic bacteria can also be associated with food production and storage. For example, planktonic bacteria can be found on food preparation surfaces, storage containers, in water used to wash food that will no longer be cooked (such as salads), or on the food itself.

[0165] Planktonic bacteria can also be associated with animal husbandry facilities or materials and equipment related to animal husbandry.

[0166] As used herein, “treating” or “treatment” means suppressing a disease or condition, that is, preventing or reducing its development or at least one of its clinical or subclinical symptoms. “Treatment” or “treatment” also means alleviating a disease or condition, that is, the disappearance of at least one of the disease or condition or its clinical or subclinical symptoms. The benefit to the subject being treated is either statistically significant or at least perceptible to the subject and / or physician.

[0167] Rezazon (7-hydroxy-3H-phen) can be used This disclosure assesses or measures the ability of preferred enantiomers of 2-hydroxycarboxylic acids to cause metabolic dysregulation in planktonic bacteria. Aerobic bacteria possess reducing cytoplasm, under which conditions resazurin is irreversibly reduced to highly fluorescent halogen. Therefore, the number of "relative fluorescence units" (RFUs) is a reading of aerobic respiration / oxidative phosphorylation. This disclosure has found that, in some respects, administration of preferred enantiomers of 2-hydroxycarboxylic acids to planktonic bacteria results in significant changes (decreases or increases) in the rate of resazurin reduction, and thus, administration of preferred enantiomers of 2-hydroxycarboxylic acids to planktonic bacteria results in significant changes (decreases or increases) in the rate of bacterial aerobic respiration / oxidative phosphorylation. The assays used in this disclosure determine the minimum inhibitory concentration (MIC) of the preferred enantiomers of 2-hydroxycarboxylic acids (rather than determining the dose / response).

[0168] Optionally, the metabolic dysregulation induced by the preferred enantiomer of the 2-hydroxycarboxylic acid does not lead to the death of planktonic bacteria. It is noted that studies have shown that application of bactericidal compounds may lead to increased metabolic rates and increased free radical production (oxidative stress leading to cell death). Therefore, the preferred enantiomer of the 2-hydroxycarboxylic acid disclosed herein is optionally not a bactericidal compound.

[0169] Application of the preferred enantiomer of the 2-hydroxycarboxylic acid disclosed herein will optionally sufficiently dysregulate (inhibit or enhance) the metabolism of planktonic bacteria to allow other mechanisms (e.g., host defense mechanisms or bactericidal compounds) to exert their effects in killing the metabolically dysregulated planktonic bacteria. When the preferred enantiomer of the 2-hydroxycarboxylic acid is provided alone, bacterial metabolism is dysregulated (inhibited or enhanced) in one direction, and then when the preferred enantiomer of the 2-hydroxycarboxylic acid is provided in combination with an antimicrobial compound, bacterial metabolism is dysregulated (enhanced or inhibited) in the other direction.

[0170] Planktonic bacteria treated with the preferred enantiomer of the 2-hydroxycarboxylic acid disclosed herein may develop resistance to one or more antimicrobial compounds. The resistance mechanism may take one or more of the following forms: - Prevents antimicrobial compounds from entering bacterial cells; - Modify the structure of antimicrobial compound targets; - Increased efflux of antimicrobial compounds from bacterial cells; and / or - Increases the metabolism of antimicrobial compounds within bacterial cells.

[0171] Metabolic dysregulation induced by the preferred enantiomer of the 2-hydroxycarboxylic acid of this disclosure can contribute to sensitizing planktonic bacteria to antimicrobial compounds (e.g., antibiotic forms). This dysregulation can be an inhibition of bacterial metabolism, which can lead to, for example, preventing the efflux of antimicrobial compounds from bacterial cells and / or reducing the metabolism of antimicrobial compounds within bacterial cells. Alternatively, if the dysregulation is an enhancement of bacterial metabolism induced by the preferred enantiomer of the 2-hydroxycarboxylic acid of this disclosure, this can contribute to sensitizing planktonic bacteria to antimicrobial compounds, for example by increasing the ability of antimicrobial compounds to penetrate planktonic bacteria, increasing the amount of free radicals in the cell and making them sensitized to antimicrobial compounds, and / or increasing the ability of antimicrobial compounds to bind to their targets (even if the planktonic bacteria alter the structure of the antimicrobial compound's target).

[0172] Methods to address metabolic disorders

[0173] This disclosure also provides a method for treating metabolic disorders in planktonic bacteria, comprising the following steps: i. Preferred enantiomers of 2-hydroxycarboxylic acid for application to planktonic bacteria.

[0174] This disclosure also provides a method for treating metabolic disorders in planktonic bacteria, comprising the following steps: i. Preferred enantiomeric forms of 2-hydroxycarboxylic acid for application to planktonic bacteria The preferred enantiomer of the 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0175] This disclosure also provides a method for treating or preventing bacterial infections caused by airborne bacteria, comprising the following steps: i. Preferred enantiomers of 2-hydroxycarboxylic acid to be applied to the site of infection.

[0176] This disclosure also provides a method for treating or preventing bacterial infections caused by airborne bacteria, comprising the following steps: i. Preferred enantiomers of 2-hydroxycarboxylic acid applied to the site of infection The preferred enantiomer of the 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0177] This disclosure also provides a method for making planktonic bacteria sensitive to microbial compounds, comprising the following steps: i. Preferred enantiomeric forms of 2-hydroxycarboxylic acid for application to planktonic bacteria The preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic disorders in planktonic bacteria.

[0178] This disclosure also provides a method for making planktonic bacteria sensitive to microbial compounds, comprising the following steps: i. Preferred enantiomeric forms of 2-hydroxycarboxylic acid for application to planktonic bacteria The preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0179] Preferably, the dysregulation of planktonic bacterial metabolism is an inhibition of planktonic bacterial metabolism. If a preferred enantiomer of a 2-hydroxycarboxylic acid is administered in combination with an antimicrobial compound, the dysregulation can be an inhibition followed by enhancement of planktonic bacterial metabolism. However, in some cases, enhancement of planktonic bacterial metabolism can occur upon exposure to only the preferred enantiomer of a 2-hydroxycarboxylic acid, without the prerequisite of inhibition and / or the presence of an antimicrobial compound. In this case, in the presence of both the 2-hydroxycarboxylic acid and the antimicrobial compound, metabolic enhancement can be followed by metabolic inhibition.

[0180] The term "infection" encompasses the growth of microorganisms on and within living and non-living surfaces, tissues, liquids, and gases. For example, an infection can occur in lung tissue. Pseudomonas In the growth of apple and pear trees Pear fire blight fungus (Erwinia amylovora) (Fire plague) infection, in water pipes Legionella bacteria The growth of bacteria or bacterial infection of the blood (septicemia).

[0181] The compositions disclosed herein may contain 0.001% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. For example, ophthalmic compositions may contain 0.001% to 1% of a preferred enantiomer of a 2-hydroxycarboxylic acid; washes and soaks, dressings and wipes, topical, inhaled, or oral compositions may contain 0.2% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, diluent, or excipient and / or other active agent. Optionally, a preferred enantiomer of a 2-hydroxycarboxylic acid is D-lactic acid.

[0182] The compositions disclosed herein may contain a non-preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the percentage of the non-preferred enantiomer of the 2-hydroxycarboxylic acid may be less than 20%. Alternatively, the compositions disclosed herein may not contain any non-preferred enantiomers of a 2-hydroxycarboxylic acid. Thus, the compositions may contain only the preferred enantiomers of a 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0183] This disclosure also provides a composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid for treating or preventing bacterial infections in a subject, wherein the bacterial infection is caused by planktonic bacteria, and wherein the composition causes metabolic dysregulation of the planktonic bacteria.

[0184] This disclosure also provides a composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid in combination with an antimicrobial compound for treating or preventing infection in a subject, wherein the infection is caused by planktonic bacteria, and wherein the composition causes metabolic dysregulation of the planktonic bacteria.

[0185] This disclosure also provides a composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid for treating or preventing bacterial infections in a subject, wherein the bacterial infection is caused by planktonic bacteria, wherein the composition sensitizes the planktonic bacteria to an antimicrobial compound, and wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0186] This disclosure also provides a composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid in combination with an antimicrobial compound for treating or preventing infection in a subject, wherein the infection is caused by planktonic bacteria, wherein the composition sensitizes the planktonic bacteria to the antimicrobial compound, and wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0187] The efficacy of a method for treating or preventing infection in a subject by administering a preferred enantiomer of a 2-hydroxycarboxylic acid can be determined by assessing results other than measuring bacterial metabolism. For example, efficacy can be determined by a reduction in the bacterial load of a sputum sample from a subject with an infectious lung disease after administration of a preferred enantiomer of a 2-hydroxycarboxylic acid. In another example, the efficacy of treating breast infection with a preferred enantiomer of a 2-hydroxycarboxylic acid can be determined by a reduction in visible inflammation on the outer surface of breast tissue, or a reduction in swelling, fever, pain, and / or redness. In the lung or breast tissue examples, other tests could include a microbiological assessment of biological fluids (e.g., sputum, breast milk) from the site of infection to determine the microbial load. If the preferred enantiomer of a 2-hydroxycarboxylic acid causes metabolic dysregulation of the planktonic bacteria causing the infection, the microbial load at the site of infection should be reduced.

[0188] surface

[0189] The preferred enantiomeric compositions of the 2-hydroxycarboxylic acid disclosed herein can be used to dysregulate the metabolism of planktonic bacteria and optionally susceptible them to antagonistic microbial compounds, including planktonic bacteria on biological and non-biological surfaces and in biological and non-biological materials (e.g., liquids and gases). For the purposes of this invention and for ease of discussion, the presence of planktonic bacteria in liquids and gases will be considered as "on a surface".

[0190] Planktonic bacteria can exist on and in almost any type of surface, such as organic surfaces (biological or abiotic), plastics, metals, glass, soil particles, wood, and food, as well as in biological and abiotic liquids and gases, such as blood, air, lymph, and drinking water. This disclosure can be used to cause metabolic disorders in planktonic bacteria in such locations.

[0191] Biological surfaces that can serve as hosts for planktonic bacteria (which can be metabolically dysregulated using the compositions disclosed herein) include: lung mucosal surfaces (particularly the lungs of subjects with cystic fibrosis or bronchiectasis); chronic wounds (including ulcers, such as diabetic ulcers); skin surfaces (particularly skin surfaces with skin flora imbalance, acne, pruritus, warts, psoriasis, and atopic dermatitis); vaginal surfaces (including bacterial vaginosis and vulvovaginal candidiasis); and gastrointestinal surfaces (e.g., small intestinal bacterial overgrowth). Clostridium difficile Surfaces that can serve as hosts for planktonic bacteria (whose metabolism can be inhibited using the compositions disclosed herein) include: blood, mucus, gastric juice, tears, and lymph. Other ocular surfaces include those affected by infection, gastric ulcers, middle ear (especially those with otitis media), periodontal surfaces (especially those with periodontitis), sinus surfaces (especially those with infected sinusitis), heart surfaces (especially those with infective endocarditis), ocular surfaces (especially those with postoperative endophthalmitis, microbial keratitis, or infectious crystalline keratitis), other ocular surfaces (especially those associated with scleral buckling implantation, conjunctival plug implantation, and lacrimal duct intubation), and bone surfaces (especially those with osteomyelitis). Biological materials that can serve as hosts for planktonic bacteria (whose metabolism can be inhibited using the compositions disclosed herein) include: blood, mucus, gastric juice, tears, and lymph.

[0192] The presence of airborne bacteria is very common in the food industry and deserves attention because airborne bacteria can be pathogenic.

[0193] Medical non-biological surfaces that can serve as hosts for planktonic bacteria (whose metabolism can be inhibited using the compositions disclosed herein) include indwelling medical devices such as urinary catheters; intravenous catheters; intrauterine devices; prostheses (including artificial hip joints, knee joints, heart valves, etc.; cochlear implants; intraocular lenses; breast implants, etc.); vascular access devices; endotracheal tubes; tracheostomies; enteral feeding tubes; wound drainage tubes; ear ventilation tubes; soap dispensers; and surface medical devices such as contact lenses, orthodontic retainers, and mouthguards. Medical non-biological substances that can serve as hosts for planktonic bacteria (whose metabolism can be disrupted using the compositions disclosed herein) include: oxygen or other air sources used to assist the breathing of subjects, and hand sanitizers.

[0194] Industrial surfaces that can serve as hosts for planktonic bacteria (causing metabolic disruption) include hydroelectric turbines, oil and gas pipelines, air conditioning units, soap dispensers, industrial water systems, ship hulls, dairy processing equipment, etc. Planktonic bacteria can also be present in animal husbandry facilities or on materials and equipment associated with animal husbandry. Industrial substances that can serve as hosts for planktonic bacteria (causing metabolic disruption using the compositions disclosed herein) include: lubricating oils, oxygen or other air sources, and hand sanitizers.

[0195] plant

[0196] The preferred enantiomeric compositions of the 2-hydroxycarboxylic acid disclosed herein can be used to induce metabolic dysregulation in planktonic bacteria in and on plant tissues and in the soil in which the plant grows, and optionally to sensitize the planktonic bacteria to antimicrobial compounds. Many plant infections are treated with antimicrobial compounds, and resistance to these compounds develops in a manner similar to that encountered in infections in animal subjects. Resistance can be counteracted by sensitizing planktonic bacteria to antimicrobial compounds using the preferred enantiomeric compositions of the 2-hydroxycarboxylic acid of the present invention.

[0197] Plants can be field crops, vegetable crops, fruit crops, forest plantations, orchards, potted plants, ornamental gardens, parks, forests, lawns, etc.

[0198] The preferred enantiomeric compositions of the 2-hydroxycarboxylic acid disclosed herein can be used to induce metabolic disorders in planktonic bacteria on food crops, field crops, and vegetable and fruit crops in fields, orchards, vineyards, and nurseries; as well as on plants, potted plants, lawns, sports fields, etc., in home garden environments. The 2-hydroxycarboxylic acid compositions can also be used to induce metabolic disorders in planktonic bacteria in densely planted environments (e.g., greenhouses). The 2-hydroxycarboxylic acid compositions can also be used to induce metabolic disorders in planktonic bacteria in environments with fewer plants but higher bacterial levels (e.g., backyards, around pig farms, chicken coops and sheds, and livestock pens).

[0199] Bacterial infections of plants that can be treated with the preferred enantiomeric compositions of the 2-hydroxycarboxylic acids disclosed herein include infections caused by bacteria, such as: Agrobacterium species Xanthomonas species Pseudomonas species Erwinia species Corynebacterium (Corynebacterium) species Ralstonia, the pathogen of bacterial wilt species and Streptomyces For example, it can treat the following diseases:

[0200] deliver

[0201] The compositions disclosed herein may be provided, for example, in the following forms: topical compositions, such as ointments, creams, foams, adhesives or lotions, eye ointments, eye drops or ear drops, or nasal sprays; impregnated dressings, patches and wipes; internal topical compositions, such as vaginal suppositories and suppositories; inhalation compositions, such as nebulized or dry powder compositions; injectable formulations for intravenous and local injection (e.g., injection into an abscess), or oral formulations in the form of tablets, liquid capsules, oral gels, etc. The compositions may also be in the form of washes or soaking solutions. The compositions may also be in the form of foliar sprays or soil sprays.

[0202] As used herein, the term "therapeutic effective amount" refers to an amount of composition sufficient to at least partially achieve the desired effect when administered according to the required dosage or administration regimen. Optionally, the desired effect is a disruption of the metabolic processes of planktonic bacteria. However, the effect may be to delay the onset of a planktonic bacterial infection (including infections on animals, surfaces, or plants), inhibit its progression, or partially or completely prevent its onset or progression.

[0203] As used herein, the term "preventative effective amount" refers to an amount of composition which, when applied according to the desired dosage or administration regimen, is sufficient to cause at least partial metabolic dysregulation of planktonic bacteria.

[0204] Dysregulation of planktonic bacterial metabolism can be either inhibition or enhancement in the presence of a preferred enantiomer of 2-hydroxycarboxylic acid alone. Upon introduction of an antimicrobial compound, the dysregulation (inhibition or enhancement) can persist, and the degree of dysregulation may increase. However, once planktonic bacteria are simultaneously exposed to both a preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound, the dysregulation can change: either inhibition becomes enhancement, or enhancement becomes inhibition. Alternatively, metabolic dysregulation may not be present in the presence of 2-hydroxycarboxylic acid alone, but significant dysregulation may occur in the presence of both a preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound, particularly when the planktonic bacteria are resistant to the antimicrobial compound in the absence of a preferred enantiomer of 2-hydroxycarboxylic acid.

[0205] Based on the foregoing, those skilled in the art will understand that a variety of different treatment and administration methods can be used to treat individual subjects or surfaces. For example, subjects already receiving drug treatment (e.g., intravenous antibiotics) may benefit from oral delivery, intravenous delivery, inhalation, or topical application of the compositions of this disclosure. Some subjects may receive only the compositions of this disclosure containing a preferred enantiomer of a 2-hydroxycarboxylic acid through oral, inhalation, or topical application. For example, a subject may have symptoms of cystic fibrosis, be diagnosed with a lung infection, or have symptoms of a medical condition that may benefit from administration of an inhaled composition containing a preferred 2-hydroxycarboxylic acid to the subject. Alternatively, a subject may have a local infection, such as a chronic wound, an eye infection, or periodontitis, and may receive topical application of a preferred enantiomer of a 2-hydroxycarboxylic acid. A subject may have a gut microbiota disorder (“dysbiosis”) that may benefit from oral administration of the preferred enantiomer composition of the 2-hydroxycarboxylic acid of this disclosure. These dysbiosis can lead to diseases caused by intestinal pathogens, inflammatory bowel disease (IBS), Crohn's disease, ulcerative colitis, or colorectal cancer.

[0206] The compositions disclosed herein can be used on plants to treat plant infections on and in plant tissues and in the soil in which the plants grow.

[0207] The compositions disclosed herein can be used on non-biological surfaces, for example, in the form of wet wipes, as a washing or soaking solution for non-biological surfaces (e.g., indwelling instruments and water pipes), and as a cleaning composition for pipes, food preparation machines, and medical devices. If the compositions are applied to non-biological surfaces, they can be washed with conventional detergents, UV disinfectants, or bleach after wiping, washing, or soaking with 2-hydroxycarboxylic acid.

[0208] The compositions disclosed herein can also be used for diagnosis. In one embodiment, for example, a subject may receive a dose of the compositions disclosed herein as part of a procedure for diagnosing an infection associated with planktonic bacteria (e.g., a lung infection), wherein one or more of the subject's symptoms improve in response to the compositions.

[0209] According to certain embodiments, the composition is administered periodically until treatment is achieved. In a preferred embodiment, the composition is administered to the subject requiring this treatment using a dosing regimen selected from the group consisting of: hourly, every 2 hours, every 3 hours, once daily, twice daily, three times daily, four times daily, five times daily, once weekly, twice weekly, once every two weeks, and once monthly. However, other application schedules may be used according to this disclosure. Optionally, the composition of the treatment regimen is administered to the subject once to five times daily, for example, once or twice daily.

[0210] If applied to a non-biological surface, the composition can be applied to the surface periodically until airborne bacteria are removed or destroyed, optionally by dysregulating their metabolism. In a preferred embodiment, the composition is applied to the surface hourly, every 2 hours, every 3 hours, once daily, twice daily, three times daily, four times daily, five times daily, once weekly, twice weekly, once every two weeks, and once monthly. However, other application schedules can be used according to this disclosure. The composition can be applied to the indwelling device before insertion and after removal.

[0211] Alternatively, the compositions of this disclosure may be administered, for example, orally, topically (to the eyes, cheeks, and sublingually, rectally, vaginally, or intranasally), or by aerosol. If the composition is to be delivered to a non-biological surface, it may be applied as a wash, soaking solution, or wipe. The manner of application or administration may optionally be adapted to the form in which the composition has been prepared. The manner of application that yields the most effective response can be determined empirically, and the methods of application or administration described below are given as examples and do not in any way limit the methods of delivery of the compositions of this disclosure.

[0212] The compositions disclosed herein may optionally include pharmaceutically acceptable nontoxic excipients and carriers. As used herein, a “drug carrier” is a pharmaceutically acceptable solvent, suspending agent, excipient, or medium for delivering a compound to a subject. The carrier may be liquid or solid and is selected according to the planned administration method.

[0213] The compositions disclosed herein may be selected from the group consisting of: immediate-release compositions, delayed-release compositions, controlled-release compositions, and rapid-release compositions.

[0214] The compositions disclosed herein may also contain anti-inflammatory agents (e.g., corticosteroids).

[0215] The compositions described herein can be formulated as oil-in-water emulsions and / or water-in-oil emulsions. In such compositions, the immediate-release formulation can be in the continuous phase, while the sustained-release formulation can be in the discontinuous phase. The compositions can also be prepared in a manner suitable for delivering the three dosage forms described above. For example, an oil-in-water-in-oil emulsion can be provided, wherein the oil is the continuous phase containing the immediate-release component, the water is dispersed in the oil containing the first sustained-release formulation, and the oil is dispersed in the water containing the third sustained-release formulation.

[0216] The compositions described herein may be in the form of liquid compositions. Liquid compositions may comprise a solution containing a therapeutic agent dissolved in a solvent. Generally, any solvent with the desired effect can be used, wherein the therapeutic agent is dissolved and can be administered to the subject. Generally, any concentration of the therapeutic agent with the desired effect can be used. In some variations, the composition is an unsaturated solution, a saturated solution, or a supersaturated solution. The solvent may be a pure solvent or a mixture of liquid solvent components. In some variations, the resulting solution is an in-situ gelling composition. The types of solvents and solutions that can be used are well known to those skilled in such drug delivery technologies.

[0217] The composition may or may not contain water. Optionally, the composition contains water, i.e., it is aqueous. In another embodiment, the composition does not contain a preservative. In one embodiment, the composition described herein may be aqueous and contain 0%-90% water. In other embodiments, the aqueous composition described herein may contain 20%-80% water. In yet another embodiment, the aqueous composition may contain 50%-70% water. The water may also include tap water, distilled water, sterile water, demineralized water, or deionized water. Alternatively, the composition may be non-aqueous and anhydrous, or contain a negligible amount of water (e.g., less than 1%, less than 0.1%, less than 0.01%).

[0218] Drug or veterinary compositions may be formulated in accordance with routine drug or veterinary practice (see, for example, Remington: The Science and Practice of Pharmacy, 20th edition, 2000, edited by AR Gennaro, Lippincott Williams & Wilkins, Philadelphia; and Encyclopedia of Pharmaceutical Technology, edited by J. Swarbrick and J.C. Boylan, 1988–1999, Marcel Dekker, New York; and Remington's Pharmaceutical Sciences, 18th edition, Mack Publishing Company, Easton, PA).

[0219] Examples of suitable carriers, excipients, and diluents typically include, but are not limited to, water, brine, ethanol, dextrose, glycerol, lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum arabic, calcium phosphate, alginate, tragacanth gum, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, aqueous syrup, methylcellulose, methylparaben and propylparaben, polysorbate, magnesium talc stearate, mineral oil, or combinations thereof. The composition may additionally contain lubricants, pH buffers, wetting agents, emulsifiers and suspending agents, preservatives, sweeteners, or flavoring agents.

[0220] The composition may be in the form of a controlled-release composition and may comprise a degradable or non-degradable polymer, hydrogel, organic gel, or other physical construct that modifies the release of a preferred enantiomer of a 2-hydroxycarboxylic acid. It should be understood that such compositions may contain additional inactive ingredients added to provide desired color, stability, buffering capacity, dispersibility, or other known desired characteristics. Such compositions may also comprise liposomes, such as emulsions, foams, micelles, insoluble monolayers, liquid crystals, phospholipid dispersions, lamellar layers, etc. Liposomes used in this disclosure may be formed from standard vesicle-forming lipids and typically comprise neutral and negatively charged phospholipids and sterols (e.g., cholesterol).

[0221] Washing and soaking composition

[0222] The compositions of this disclosure, containing a preferred enantiomer of a 2-hydroxycarboxylic acid, can be used for washing and / or soaking devices, such as indwelling medical devices or water pipe systems. Washing or soaking such devices with a composition containing a preferred enantiomer of a 2-hydroxycarboxylic acid helps to disrupt the metabolism of any planktonic bacteria present on the surface of the device, and thus facilitates sterilization or cleaning of the device by making it easier to remove and / or kill any microorganisms present. The treatment may optionally sufficiently disrupt the metabolism to enhance the effect of antimicrobial compounds (e.g., bactericidal compounds, bacteriostatic compounds, antibiofilm compounds, or antiseptic compounds).

[0223] In one form, the composition for washing or soaking may contain 0.1% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, diluent or excipient and / or other surfactants.

[0224] In one form, the composition for washing or soaking may contain a non-preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the percentage of the non-preferred enantiomer of the 2-hydroxycarboxylic acid may be less than 20%. Alternatively, the compositions of this disclosure may not contain any non-preferred enantiomers of a 2-hydroxycarboxylic acid. Thus, the composition may contain only the preferred enantiomer of a 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0225] Dressings and wipes

[0226] The compositions of this disclosure, comprising a preferred enantiomer of a 2-hydroxycarboxylic acid, can be provided in the form of an impregnated dressing, bandage, or wipe. Dressings can be applied to wounds or localized surfaces that may be infected with airborne bacteria and can remain in place for hours, days, or weeks. Wipes can be used to reduce the presence of airborne bacteria on surfaces such as wounds or mucous membranes (e.g., vaginal or rectal surfaces) or non-biological surfaces. Wipes can be used hourly, daily, weekly, or as needed. For example, wipes can be used after each urination or defecation to disrupt the metabolism of airborne bacteria present on vaginal or rectal mucosal tissue. Alternatively, wipes can be used before using food preparation surfaces.

[0227] In one form, the composition for dressings or wipes may contain 0.2% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, diluent or excipient and / or other active agents.

[0228] In one form, the composition for dressings or wipes may contain a non-preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the percentage of the non-preferred enantiomer of the 2-hydroxycarboxylic acid may be less than 20%. Alternatively, the compositions of this disclosure may not contain any non-preferred enantiomers of a 2-hydroxycarboxylic acid. Thus, the composition may contain only the preferred enantiomer of a 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0229] Inhalation delivery

[0230] The compositions disclosed herein can be delivered via a nebulizer or other aerosol delivery device. This is particularly suitable for respiratory and ear, nose, and throat (ENT) diseases associated with airborne bacteria.

[0231] Optionally, the composition may be administered to the subject in need about once a day to about six times a day (e.g., about once or twice a day).

[0232] Alternatively, the composition can be administered to the subject in need via a nebulizer through continuous inhalation. The nebulized composition can be delivered for 24 hours, 12 hours, 8 hours, 6 hours, 4 hours, 2 hours, or 1 hour, and each of these deliveries (except for 24 hours and 12 hours) can be repeated several times within 24 hours.

[0233] In one form, the composition delivered via an atomizer or aerosol delivery device may contain 0.2% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, a diluent or excipient, and / or other active agents.

[0234] In one form, the composition delivered by an atomizer or aerosol delivery device may contain a non-preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the percentage of the non-preferred enantiomer of the 2-hydroxycarboxylic acid may be less than 20%. Alternatively, the compositions of this disclosure may not contain any non-preferred enantiomers of a 2-hydroxycarboxylic acid. Thus, the composition may contain only the preferred enantiomer of a 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0235] Subjects may typically receive a preferred enantiomer of 2-hydroxycarboxylic acid at doses ranging from 1 mg / mL to 500 mg / mL (±20% or ±10%). The dose may be delivered over various time intervals, such as by a single inhalation (several seconds) or over several minutes to several hours. The dose may be delivered over 5 minutes, 10 minutes, 15 minutes, 20 minutes, 30 minutes, 45 minutes, 60 minutes, 120 minutes, or longer. The dose is typically administered via a nebulizer or by at least one, preferably multiple, inhalations from an aerosol device. For example, subjects may receive a preferred enantiomer of 2-hydroxycarboxylic acid at a single daily dose or several doses throughout the day.

[0236] The total daily dose may optionally be administered at least once daily, but may be divided into two or more daily doses. Some subjects may benefit from a “loading” period, which involves administering the preferred enantiomer of 2-hydroxycarboxylic acid at a higher dose or more frequently over several days or weeks, followed by a reduction or maintenance dose. Because cystic fibrosis, COPD (chronic obstructive pulmonary disease), and other conditions are typically chronic, long-term treatment is expected in subjects.

[0237] There are various mechanical devices designed for the pulmonary delivery of therapeutic products, including but not limited to nebulizers, metered-dose inhalers, and powder inhalers, all of which are well known to those skilled in the art. Some specific examples suitable for practicing the commercially available devices disclosed herein are: the Ultravent nebulizer manufactured by Mallinckrodt, Inc. of St. Louis, Missouri; the Acorn II nebulizer manufactured by Marquest Medical Products, Inc. of Englewood, Colorado; the Ventolin metered-dose inhaler manufactured by Glaxo, Inc. of Research Triangle Park, North Carolina; and the Spinhaler powder inhaler manufactured by Fisons, Inc. of Bedford, Massachusetts.

[0238] All of these devices require the use of compositions suitable for dispensing preferred enantiomers of 2-hydroxycarboxylic acids. Typically, each composition is specific to the type of device employed and may involve the use of appropriate propellant materials in addition to the commonly used diluents, adjuvants, and / or carriers for treatment.

[0239] Regardless of the form of the pharmaceutical composition, it is preferably produced as droplets or particles for inhalation with a size of about 0.1 μm to 12 μm, or about 0.25 μm to 6 μm, optionally 1 μm to 6 μm (e.g., about 2 μm to 4 μm). Alternatively, the particles may be 0.1 μm to 1.0 μm, 0.2 μm to 0.9 μm, 0.3 μm to 0.8 μm, 0.4 μm to 0.7 μm, or 0.5 μm. By producing inhaled particles with a relatively narrow size range, the efficiency of the drug delivery system can be further improved and the repeatability of administration can be enhanced. Therefore, it is preferred that the particles not only have a size of 0.1 µm to 12 µm, or 2 µm to 6 µm, or about 3 µm to 4 µm, but also an average particle size within a narrow range, such that 80% or more of the particles delivered to the subject have a particle size within ±20% or ±10% of the average particle size, for example, ±5% of the average particle size.

[0240] "Particle size" is a concept introduced to compare the size of solid particles and liquid particles (droplets). For droplets and aerosols, terms such as "aerodynamic diameter" and "mass median aerodynamic diameter" (MMAD) are used.

[0241] "Aerodynamic diameter" is the diameter of a sphere of unit density with the same terminal settling velocity as the particle in question. It is used to predict where such particles will deposit in the respiratory tract.

[0242] "Median Mass Aerodynamic Diameter" (MMAD) is the geometrically average aerodynamic diameter. By weight, 50% of particles will be smaller than MMAD, and 50% of particles will be larger than MMAD.

[0243] In particle size distribution experiments, the suspension contains numerous moving particles of varying sizes. When a particle size analyzer analyzes these particles, it forms a particle size distribution curve that covers the entire particle size range, from the smallest particle (which may be 1 nm) to the largest particle (which may be 100 µm). The cumulative frequency of the particles is calculated from this particle size distribution curve. 10 This refers to a specific particle diameter in which 10% of the particles in the suspension have a diameter less than or equal to that of a specific particle.

[0244] D 50 Similar to D 10 D 50 It is the cutoff diameter of 50% of the particle group in the composition, and refers to the specific particle diameter in which the diameter of 50% of the particles in the suspension is less than or equal to the specific particle diameter.

[0245] D 90 :D 90It is the cutoff diameter of 90% of the particle group in the composition, and refers to the specific particle diameter in which the diameter of 90% of the particles in the suspension is less than or equal to the diameter of a specific particle.

[0246] The term "respiratory tract" should be understood to refer to the cellular and organ systems that function in respiration, particularly the organs, tissues, and cells of the respiratory tract, including the lungs, nose, nasal passages, sinuses, nasopharynx, larynx, trachea, bronchi, bronchioles, respiratory bronchioles, alveolar ducts, alveolar sacs, alveoli, lung cells (type 1 and type 2), ciliated mucosal epithelium, mucosal epithelium, squamous epithelial cells, mast cells, goblet cells, neutrophils, macrophages, and intraepithelial dendritic cells.

[0247] In one form of this disclosure, a method for treating pulmonary planktonic bacterial metabolic disorders in a subject comprises administering a preferred enantiomer of a 2-hydroxycarboxylic acid at one or more doses of 1 mg / mL to 1000 mg / mL (e.g., 5 mg / mL to 500 mg / mL) to a therapeutically effective or preventatively effective concentration.

[0248] In one form of this disclosure, a method for treating pulmonary planktonic bacterial metabolic disorder in a subject comprises administering an inhaled composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid at a therapeutically effective concentration in one or more doses ranging from 1 mg / kg / day to 1000 mg / kg / day (e.g., 6 mg / kg / day to 600 mg / kg / day).

[0249] In one form of this disclosure, a method for achieving pulmonary dysbiosis of a subject is described by administering an inhaled composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid at one or more doses of 1 mg / mL to 1000 mg / mL (e.g., 5 mg / mL to 500 mg / mL).

[0250] The compositions disclosed herein can be administered to a subject using disposable packaging and portable, handheld, battery-powered devices, such as the AERx device (U.S. Patent No. 5,823,178, Aradigm, Hayward, Calif.) Alternatively, the compositions disclosed herein can be administered using mechanical (non-electronic) devices. Other inhalation devices can be used to deliver the compositions, including conventional jet nebulizers, ultrasonic nebulizers, soft fog inhalers, dry powder inhalers (DPIs), metered-dose inhalers (MDIs), condensation aerosol generators, and other systems.

[0251] For use as an aerosol, the disclosed compounds in solution or suspension can be packaged together with a suitable propellant (e.g., a hydrocarbon propellant such as propane, butane, or isobutane) and conventional adjuvants in a pressurized aerosol container. A dry powder inhaler is a system that can be operated with a pressurized air source to produce dry powder particles of a pharmaceutical composition compacted into very small volumes. For inhalation, the system has multiple chambers or blisters, each containing a single dose of the pharmaceutical composition and a selection element for the release of that single dose.

[0252] Aerosols can be generated by forcing a drug through pores in a membrane, the pore size ranging from about 0.25 µm to 6 µm (U.S. Patent No. 5,823,178). When the pores have this size, the diameter of the particles escaping through the pores to generate the aerosol will be in the range of 0.5 µm to 12 µm. The drug particles can be released using an airflow designed to keep the particles within this size range. The generation of small particles can be facilitated by using a vibrating device that provides a vibration frequency in the range of about 800 kHz to about 4000 kHz. Those skilled in the art will recognize that some adjustments can be made to parameters such as the pore size from which the drug is released, the vibration frequency, pressure, and other parameters based on the density and viscosity of the composition, while keeping in mind that the aim of some embodiments is to provide atomized particles with diameters in the range of about 0.5 μm to 12 μm.

[0253] Local delivery

[0254] The compositions disclosed herein can be delivered topically. Topical application may include applying a therapeutically effective amount of a preferred enantiomer of a 2-hydroxycarboxylic acid directly to the skin, eyes, or mucous membranes of a subject. Alternatively, a preferred enantiomer of a 2-hydroxycarboxylic acid may be topically applied to the skin, mucous membranes (oral cavity, nose, vagina, rectum), or eyes of a subject. Use may include administering a therapeutically effective amount of a preferred enantiomer of a 2-hydroxycarboxylic acid to the skin, mucous membranes (oral cavity, nose, vagina, rectum), or eyes of a subject.

[0255] The compositions disclosed herein can be applied topically. Therefore, compositions suitable for direct application to the skin are contemplated herein.

[0256] In one form, the composition for local delivery may contain 0.2% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, a diluent or excipient, and / or other active agents.

[0257] In one form, the composition for local delivery may comprise a non-preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the percentage of the non-preferred enantiomer of the 2-hydroxycarboxylic acid may be less than 20% of the total 2-hydroxycarboxylic acid content. Alternatively, the compositions of this disclosure may not contain any non-preferred enantiomers of a 2-hydroxycarboxylic acid. Thus, the composition may comprise only the preferred enantiomer of a 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0258] The compositions can be in the form of suspensions, emulsions, liquids, creams, oils, lotions, ointments, gels, hydrogels, pastes, plasters, roll-on liquids, skin patches, sprays, glass bead dressings, synthetic polymer dressings, and solids. For example, the compositions disclosed herein can be provided as water-based compositions or ointments based on organic solvents (e.g., oils). Alternatively, the compositions disclosed herein can be applied by a liquid spray comprising a film-forming component and at least one solvent in which a preferred enantiomer of a 2-hydroxycarboxylic acid is dispersed or dissolved.

[0259] The compositions disclosed herein may be provided in the form of, but not limited to, the group consisting of: rinses, shampoos, lotions, gels, leave-on formulations, rinse-off formulations, and ointments.

[0260] Various local delivery systems can be adapted to administer the compositions of this disclosure, depending on the preferred treatment regimen. The local compositions can be prepared by dissolving or mixing a preferred enantiomer of the 2-hydroxycarboxylic acid of this disclosure in an aqueous or non-aqueous carrier. Generally, any liquid, cream, gel, or similar substance that does not react significantly with the compound or any other active ingredient and is non-irritating is suitable and can be incorporated into the composition. Suitable non-sprayable viscous, semi-solid, or solid forms may also be used, comprising a carrier compatible with local application and having a dynamic viscosity greater than that of water.

[0261] Suitable compositions are well known to those skilled in the art and include, but are not limited to, solutions, suspensions, emulsions, creams, gels, ointments, powders, liniments, ointments, aerosols, transdermal patches, etc., which may be sterilized or mixed with adjuvants, such as preservatives, stabilizers, emulsifiers, wetting agents, fragrances, colorants, odor control agents, thickeners (e.g., natural gums), if necessary. Particularly preferred topical compositions include ointments, creams, or gels.

[0262] Ointments are typically prepared using (1) an oily matrix, i.e., a matrix composed of fixed oils or hydrocarbons, such as white petroleum or mineral oil; or (2) an absorbent matrix, i.e., a matrix composed of one or more anhydrous substances that can absorb water (e.g., anhydrous lanolin). Typically, after the matrix (whether oily or absorbent) is formed, a preferred 2-hydroxycarboxylic acid is added to provide the desired concentration.

[0263] Creams are oil / water emulsions. They consist of an oil phase (internal phase) and an aqueous phase (continuous phase). The oil phase typically contains fixed oils, hydrocarbons, waxes, petroleum, mineral oils, etc., while the aqueous phase contains water and any water-soluble substances, such as added salts. These two phases are stabilized using emulsifiers, such as surfactants (e.g., sodium dodecyl sulfite); hydrophilic colloids (e.g., gum arabic); plant gums (e.g., veegum). After the emulsion is formed, a preferred enantiomeric form of a 2-hydroxycarboxylic acid can be added in an amount to achieve the desired concentration.

[0264] The gel comprises a matrix selected from oily, aqueous, or emulsion-suspension matrices. A gelling agent is added to the matrix, which forms a gel matrix within the matrix, increasing its viscosity. Examples of gelling agents are hydroxypropyl cellulose, acrylic polymers, etc. Typically, a preferred enantiomeric form of a 2-hydroxycarboxylic acid is added to the composition at the desired concentration prior to the addition of the gelling agent.

[0265] Topical delivery compositions for application to mucosal surfaces (e.g., oral, vaginal, nasal, or rectal mucosa) or skin wounds may contain 5% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, diluent, or excipient and / or other active agents. In one aspect, higher doses may be used in medically monitored situations, while lower doses may be used for non-life-threatening wounds treated at home.

[0266] In one form, the composition for application to mucosal surfaces and / or skin wounds may contain a non-preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the percentage of the non-preferred enantiomer of the 2-hydroxycarboxylic acid may be less than 20%. Alternatively, the compositions of this disclosure may not contain any non-preferred enantiomers of a 2-hydroxycarboxylic acid. Thus, the composition may contain only the preferred enantiomer of a 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0267] The compositions disclosed herein can be applied via topical ocular delivery. Optionally, the ocular composition comprises 0.001% to 1% of a preferred enantiomer of a 2-hydroxycarboxylic acid.

[0268] Ocular delivery includes delivery to the sclera, retina, intraocular fluid, and periocular tissues. For example, delivery can be topical (cream, gel, ointment, spray, eye drops), intraocular implants, or other methods.

[0269] Ocular delivery may also include injecting a preferred enantiomer of a 2-hydroxycarboxylic acid into the sclera, intraocular space, or retroocular region. Compositions suitable for ocular injection may optionally include sterile aqueous solutions (where soluble in water) or dispersions, as well as sterile powders for the ad hoc preparation of sterile injectable solutions or dispersions. Alternatively, in some aspects, the compounds of this disclosure are encapsulated in liposomes and delivered as injectable solutions to facilitate their transmembrane transport. Alternatively, or additionally, such formulations contain components with self-assembling pore structures to promote transmembrane transport.

[0270] Oral delivery composition

[0271] The compositions disclosed herein can be administered orally.

[0272] In one form, the orally delivered composition may contain 5% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, a diluent or excipient, and / or other active agents.

[0273] In one form, the orally delivered composition may comprise a non-preferred enantiomer of a 2-hydroxycarboxylic acid, wherein the percentage of the non-preferred enantiomer of the 2-hydroxycarboxylic acid may be less than 20%. Alternatively, the compositions of this disclosure may not contain any non-preferred enantiomers of a 2-hydroxycarboxylic acid. Thus, the composition may comprise only the preferred enantiomer of a 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0274] Subjects can usually receive preferred enantiomers of 2-hydroxycarboxylic acids at doses ranging from 0.1 mg / kg / day to 2 g / kg / day (±20% or ±10%).

[0275] Despite the varying pH levels in the digestive system (acidic in the stomach, alkaline in the small intestine), the potency of the preferred enantiomers of 2-hydroxycarboxylic acids against planktonic bacteria remains. The pH of the surrounding fluid does not affect the metabolic dysregulation capacity of the preferred enantiomers of 2-hydroxycarboxylic acids. This is because 2-hydroxycarboxylic acids remain stable in the gastrointestinal tract and have been shown to maintain their function, as observed in numerous studies of the acid tolerance of lactic acid bacteria (e.g., Wang et al., 2018, Archives of Microbiology, 200, 195-201) and lactic acidosis (a metabolic acidosis caused by excessive fermentation in the gastrointestinal tract).

[0276] The oral compositions disclosed herein can be delivered together with an immune enhancer or modulator. Examples of immune enhancers or modulators include specific immune stimulants (e.g., vaccines and antigens) and nonspecific agents such as bee products (including propolis and honey), probiotics and prebiotics, hormones, vitamins (vitamin C, vitamin D), minerals (zinc oxide), antioxidants (including glutathione), interferons (including INF-α), interleukins (including interleukin-10), colchicine, thalidomide, and imiquimod. The immune enhancer can be delivered simultaneously with or after a composition containing a preferred enantiomer of a 2-hydroxycarboxylic acid.

[0277] Plant delivery

[0278] The application of a composition containing a preferred enantiomer of a 2-hydroxycarboxylic acid to plants and / or soil can be in the form of spraying using a pump-pack, manual sprayer, or boom sprayer; using a hose or jet or an aircraft equipped with a crop sprayer; and through an irrigation system, i.e., drip irrigation, micro-sprinkler irrigation, flood irrigation, etc. The composition can also be injected into plants or trees.

[0279] Subjects

[0280] If a preferred enantiomer of a 2-hydroxycarboxylic acid is administered to a subject, the subject can be any subject capable of being infected by bacteria, viruses, fungi, or archaea. The subject can be any animal, including fish, crustaceans, mollusks, mammals, reptiles, or birds. Optionally, the subject is a mammal selected from the group consisting of: humans, canines, birds, pigs, cattle, sheep, horses, rodents, mustelids, rabbits, and felines. The subject can be a companion animal, a domesticated animal, or an animal of importance in aquaculture or agriculture. In one example, the subject is a human.

[0281] excipient

[0282] The exemplary forms of the compositions described herein can be manufactured by methods well known to those skilled in the art of compositional science. Furthermore, the compositions described herein may contain other optional excipients to aid in the manufacture and / or application of the compositions. In one embodiment, the composition further comprises one or more pharmaceutically or physiologically acceptable carriers, diluents, or excipients. Non-limiting examples of such excipients are well known in the art and include flavoring agents, coloring agents, palatability agents, antioxidants, viscosity modifiers, tension agents, drug carriers, sustained-release agents, comfort enhancers, emulsifiers, solubilizers, lubricants, binders, and other stabilizers to aid in the manufacture and / or application of the compositions.

[0283] The compositions disclosed herein are intended for use in pharmaceutical or veterinary applications, or for use in cleaning equipment, such as medical devices, water pipes, or food preparation equipment.

[0284] Optionally, the compositions of this disclosure are sterile. In addition to or instead of sterilization, the compositions of this disclosure may contain pharmaceutically acceptable preservatives to minimize the possibility of microbial contamination. In another embodiment, the compositions of this disclosure are stable. Pharmaceutically acceptable preservatives may be used in the compositions of this disclosure to increase the stability of the composition. However, it should be noted that any preservative must be selected for safety reasons, as treated tissues may be sensitive to irritants. Preservatives suitable for use herein include, but are not limited to, preservatives that protect solutions from pathogen contamination, including phenylethyl alcohol, benzalkonium chloride, or benzoic acid, or benzoates such as sodium benzoate and phenylethyl alcohol. In some embodiments, the compositions herein contain about 0.001% w / w to about 10.0% w / w benzalkonium chloride, or about 0.01% v / w phenylethyl alcohol. Preservatives may also be present in amounts from about 0.001% w / w to about 1% w / w, for example, from about 0.002% w / w to about 0.02% w / w or about 0.02% w / w.

[0285] The compositions provided herein may also contain one or more emulsifiers, wetting agents or suspending agents in amounts of about 0.001% to about 90%, about 0.001% to about 50%, about 0.001% to about 25%, about 0.001% to about 10%, or about 0.001% to about 1%. The reagents used in this article include, but are not limited to, polyoxyethylene sorbitan fatty acid esters or polysorbate esters, including but not limited to, polyethylene sorbitan monooleate (polysorbate 80), polysorbate 20 (polyoxyethylene (20) sorbitan monolaurate), polysorbate 65 (polyoxyethylene (20) sorbitan tristearate), polyoxyethylene (20) sorbitan monooleate, polyoxyethylene (20) sorbitan monopalmitate, polyoxyethylene (20) sorbitan monostearate; lecithin; agar; carrageenan; locust bean gum; guar gum; tragacanth gum; gum arabic; xanthan gum; black privet gum; pectin; amidated pectin; ammonium phospholipids; microcrystalline cellulose; methylcellulose; hydroxypropyl Cellulose; Hydroxypropyl methylcellulose; Ethyl methylcellulose; Carboxymethylcellulose; Sodium, potassium, and calcium salts of fatty acids; Monoglycerides and diglycerides of fatty acids; Acetates of monoglycerides and diglycerides of fatty acids; Lactates of monoglycerides and diglycerides of fatty acids; Citrates of monoglycerides and diglycerides of fatty acids; Tartrates of monoglycerides and diglycerides of fatty acids; Monoacetyl tartrates and diacetyl tartrates of monoglycerides and diglycerides of fatty acids; Mixed acetates and tartrates of monoglycerides and diglycerides of fatty acids; Sucrose esters of fatty acids; Sucrose glycerides; Polyglycerides of fatty acids; Castor oil condensation polyglycerides of fatty acids; Propane-1,2-diol esters of fatty acids; Sodium stearoyl-21-propionate (sodium...) Stearoyl-21 actylate); calcium stearoyl-2-lactic acid; stearoyl tartrate; sorbitan monostearate; sorbitan tristearate; sorbitan monolaurate; sorbitan monooleate; sorbitan monopalmitate; soap tree extract; soybean oil dimer fatty acid polyglycerol ester; oxidized polymerized soybean oil; and pectin extract.

[0286] The compositions disclosed herein may contain about 0.001% by weight to about 5% by weight of a humectant to prevent mucosal dryness and irritation. Any of a variety of pharmaceutically acceptable humectants may be used, including, for example, sorbitol, propylene glycol, polyethylene glycol, glycerin, or mixtures thereof.

[0287] This disclosure covers variations of the above compositions, as the amounts of the individual compounds can vary by ±5%, ±7.5%, ±10%, ±15%, ±17.5%, or ±20%.

[0288] This disclosure covers compositions in which the relative proportions of the active ingredient and / or each excipient independently differ from those described above. In one form of this disclosure, the relative proportions of the active ingredient and / or each excipient independently differ from those described above by up to 50%. In one form of this disclosure, the relative proportions of the active ingredient and / or each excipient independently differ from those described above by up to 40%. In one form of this disclosure, the relative proportions of the active ingredient and / or each excipient independently differ from those described above by up to 30%. In one form of this disclosure, the relative proportions of the active ingredient and / or each excipient independently differ from those described above by up to 20%. In one form of this disclosure, the relative proportions of the active ingredient and / or each excipient independently differ from those described above by up to 10%. In one form of this disclosure, the relative proportions of the active ingredient and / or each excipient independently differ from those described above by up to 5%. In one form of this disclosure, the relative proportions independently differ from those described above by up to 10%. In one form of this disclosure, the relative proportions of the active ingredient and / or each excipient differ independently from those described above by up to 2%.

[0289] As those skilled in the art will understand, the sum of the percentages of excipients and active ingredients cannot exceed 100%, and the above variations are subject to this limitation. As those skilled in the art will understand, since this disclosure includes components other than those described above, the sum of the percentages of excipients and active ingredients can be less than 100%.

[0290] The changes described above are percentage changes in relative proportions. For example, a 20% change in the relative proportion of a component (excipient or active ingredient) specified as 1% means that the relative proportion of that component could be between 0.8% and 1.2%.

[0291] Inhalation delivery excipients

[0292] Compositions suitable for use with nebulizers (jet or ultrasonic) typically contain preferred enantiomers of 2-hydroxycarboxylic acids suspended in water or non-aqueous solvents. The composition may also contain buffers and monosaccharides (e.g., for stabilizing and regulating osmotic pressure). Nebulizer compositions may also contain surfactants to reduce or prevent surface-induced aggregation of the preferred enantiomers of 2-hydroxycarboxylic acids during solution atomization during aerosol formation. Furthermore, the use of liposomes, microcapsules or microspheres, inclusion complexes, or other types of carriers is considered.

[0293] Compositions used with metered-dose inhaler devices typically comprise finely ground powder containing a preferred enantiomer of a 2-hydroxycarboxylic acid suspended in the propellant by means of a surfactant. The propellant can be any conventional material used for this purpose, such as chlorofluorocarbons, hydrochlorofluorocarbons, hydrofluorocarbons, or hydrocarbons, including trichlorofluoromethane, dichlorodifluoromethane, dichlorotetrafluoroethanol, and 1,1,1,2-tetrafluoroethane or combinations thereof. Suitable surfactants include sorbitan trioleate and soybean lecithin. Oleic acid can also be used as a surfactant.

[0294] The composition for dispensing from a powder inhaler device will comprise a finely ground dry powder containing a preferred enantiomer of a 2-hydroxycarboxylic acid, and may also contain a filler, such as lactose, sorbitol, sucrose, or mannitol, in an amount that facilitates the dispersion of the powder from the device, for example, 50% to 90% by weight of the composition. The preferred enantiomer of the 2-hydroxycarboxylic acid should most advantageously be prepared in particulate form with an average particle size of less than 10 micrometers, for example, 0.5 to 5 micrometers, for most efficient delivery to the distal lung.

[0295] In one embodiment, the compositions of this disclosure may comprise a preservative, a suspending agent, a wetting agent, a tensioning agent, and / or a diluent. The compositions provided herein may comprise one or more pharmacologically suitable suspensions that are physiologically acceptable when administered by inhalation, ranging from about 0.01% to about 90%, or from about 0.01% to about 50%, or from about 0.01% to about 25%, or from about 0.01% to about 10%, or from about 0.01% to about 5%. Pharmacologically suitable fluids used herein include, but are not limited to, polar solvents, including, but not limited to, compounds containing hydroxyl or other polar groups. Solvents include, but are not limited to, water or alcohols, such as ethanol, isopropanol, and glycols, including propylene glycol, polyethylene glycol, polypropylene glycol, glycol ethers, glycerol, and polyoxyethylene alcohol. Polar solvents also include proton solvents, including, but not limited to, water, saline solutions having one or more pharmaceutically acceptable salts, alcohols, glycols, or mixtures thereof. In an alternative embodiment, the water used in the compositions of this disclosure shall meet or exceed applicable regulatory requirements for inhaled medications.

[0296] In one embodiment, the composition described herein may be aqueous and contain 0%-90% water. In other embodiments, the aqueous composition described herein may contain 20%-80% water. In yet another embodiment, the aqueous composition may contain 50%-70% water. The water may also include tap water, distilled water, sterile water, demineralized water, or deionized water. Alternatively, the composition may be non-aqueous and water-free, or contain a negligible amount of water (e.g., less than 1%, less than 0.1%, less than 0.01%).

[0297] The compositions disclosed herein may also contain adjuvants, such as bronchodilators, anti-inflammatory agents, surfactants, aspirin, or ethanol.

[0298] Optional bronchodilators used in the compositions disclosed herein include, but are not limited to, β2-adrenergic receptor agonists (e.g., salbutamol, bamboorol, salmeterol, formoterol, artoformoterol, levosalbutamol, procaterol, indacaterol, carmoterol, milveterol, procaterol, terbutaline, etc.) and antimuscarinic agents (e.g., troxetine, ipratropium bromide, glycopyrronium bromide, adecyl bromide, etc.). Combination therapy is possible.

[0299] Anti-inflammatory agents that may optionally be used in the compositions disclosed herein include, but are not limited to, inhaled corticosteroids (e.g., beclometasone, budesonide, ciclesonide, fluticasone, etiprednol, mometasone, etc.), leukotriene receptor antagonists and leukotriene synthesis inhibitors (e.g., montelukast, zileuton, ibudilast, zafirlukast, pranlukast, amelubant, tipelukast, etc.), and cyclooxygenase inhibitors (e.g., ibuprofen). (These medications include) ketoprofen, ketoroxyfen, indomethacin, naproxen, zaltoprofen, lornoxicam, meloxicam, celecoxib, lumiracoxib, etoricoxib, piroxicam, ampiroxicam, cinnoxicam, diclofenac, biphenylacetic acid, lornoxicam, mesalazine, trifluralin, tinoridine, iguratimod, pamigraf, etc. These can be used in combination. Aspirin can also be added as an anti-inflammatory agent.

[0300] The surfactants covered in this disclosure include, but are not limited to, synthetic surfactants (Exosurf®), dipalmitoylphosphatidylcholine, and oleic acid. They can be used in combination. Antioxidants such as glutathione and vitamin E, zinc, and zinc salts of EDTA can be added.

[0301] Ethanol vapor acts as an antifoaming agent in the lungs, making sputum more fluid, which can aid breathing and reduce pulmonary edema. Ethanol can be added to the compositions disclosed herein at 0.5% to 60% (e.g., 1% to 40%, 1% to 20%, or 1% to 10%). Ethanol can be added at 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, or 60%.

[0302] This disclosure also relates to the use of preferred enantiomers of 2-hydroxycarboxylic acids in combination with other drugs administered by inhalation. These other drugs may include nucleotide sequences that can be incorporated into suitable delivery vectors (e.g., plasmids or viral vectors). Other drugs may be therapeutic nucleotide sequences (DNA, RNA, siRNA), enzymes that reduce mucus viscoelasticity such as DNase and other mucolytics, chemicals that upregulate chloride channels or promote ion transcellular flow, nicotine, P2Y2 agonists, elastase inhibitors (including α-1 antitrypsin (AAT)), N-acetylcysteine, antibiotics and cationic peptides (e.g., lanthanum antibiotics, particularly nifedipine), short-acting bronchodilators (e.g., β2-adrenergic receptor agonists such as salbutamol or indacaterol), M3 muscarinic antagonists (e.g., ipratropium bromide), K +Channel openers, long-acting bronchodilators (e.g., formoterol, salmeterol), steroids (e.g., budesonide, fluticasone, triamcinolone, beclomethasone, ciclesonide, etc.), xanthine, leukotriene antagonists (e.g., montelukast sodium), phosphodiesterase 4 inhibitors, adenosine receptor antagonists, various other anti-inflammatory drugs (e.g., Syk kinase inhibitors (AVE-0950), trypsin inhibitors (AVE-8923 and AVE-5638), tachykinin antagonists (AVE-5883), inducible nitric oxide synthase inhibitors (GW-274150), and others), transcription factor inducers The following are considered as potential agents: α, TLR-9 agonists, antisense oligonucleotides, siRNA, DNA, CGRP, lidocaine, inverse β2-agonists, anti-infective oxidative therapies, cytokine modulators (e.g., CCR3 receptor antagonists (GSK-766994, DPC-168, AZD-3778), TNF-α production inhibitors (LMP-160 and YS-TH2) and IL-4 antagonists (AVE-0309)), small molecule IgE inhibitors, cell adhesion molecule (CAM) inhibitors, small molecules targeting VLA4 receptors or integrin α4β1 (e.g., R-411, PS-460644, DW-908e and CDP-323), and immunomodulators, including those that block T cell signaling by inhibiting calcineurin (tacrolimus), heparin neutralizers (Talactoferrin α), cytoplasmic PLA2 inhibitors (Efipladib), or combinations thereof. If a subject with CF is in need, standard medications such as ivacaftor, pulmozyme, mannitol, or other approved medications according to standard practice may also be administered in combination with the compositions disclosed herein.

[0303] Local delivery excipients

[0304] The compositions of this disclosure, comprising a preferred enantiomer of a 2-hydroxycarboxylic acid, can be delivered to a topical surface. For example, the topical formulation can be applied topically to the skin, mucous membranes (oral cavity, nose, vagina, rectum) or eyes of a subject.

[0305] The compositions disclosed herein may contain water (aqueous) or may be non-aqueous.

[0306] The compositions disclosed herein may also include small amounts of conventional additives, such as viscosity modifiers (e.g., xanthan gum) and preservatives (e.g., phenoxyethanol or benzyl alcohol, including mixtures thereof). For some therapeutic agents, a buffer may need to be incorporated to maintain a suitable pH.

[0307] Suitable preservatives for such compositions or pharmaceuticals include, for example, phenoxyethanol and other preservatives commonly used in pharmaceutical preparations, particularly creams. Suitable preservatives include methylparaben, chlorocresol, sorbic acid, and benzoic acid.

[0308] The compositions disclosed herein can be prepared using conventional pharmaceutical techniques. Therefore, ointments and creams can be conveniently prepared by mixing the components constituting the medium together at a high temperature (optionally 60°C-70°C) until an emulsion is formed. The mixture can then be cooled to room temperature and, after the addition of a preferred enantiomer of the 2-hydroxycarboxylic acid and any other components, stirred to ensure adequate dispersion.

[0309] Liquid formulations, such as ear drops and eye drops, are prepared by dissolving the therapeutic agent in the components that make up the medium and then adding other ingredients. The resulting solution or suspension is then dispensed into glass or plastic bottles, or packaged in single-dose packs (e.g., soft gelatin capsules) and then heat-sealed.

[0310] Artificial tear mediators can be used for the delivery of ocular compositions containing preferred enantiomers of 2-hydroxycarboxylic acids. Thicker artificial tears use high concentrations of thickeners such as Celluvisc®, high-viscosity carboxymethyl cellulose (CMC), and Refresh Liquigel® (a mixture of 0.35% high-viscosity CMC and 0.65% low-viscosity CMC).

[0311] Gelling agents can be used in compositions containing preferred enantiomers of 2-hydroxycarboxylic acids for delivery to the eye. Such agents can be instilled as liquid drops, triggering a gel phase almost immediately. Examples of such agents include Timoptic gel (gellan gel), AzaSite® (polycarbophil, poloxamer), Besivance® (polycarbophil, poloxamer), and 0.3% Keltrol® alginate. Another type of gelling agent is a polycarbophil-poloxamer gel (e.g., Durasite®).

[0312] In various aspects, ocular carriers are solvents or dispersion media containing, for example, water, ethanol, polyols (such as glycerol, propylene glycol, and liquid polyethylene glycol), suitable mixtures thereof, and vegetable oils. This is achieved, for example but not limited to, by using coatings (such as lecithin), by maintaining a desired particle size in the dispersed state, and by maintaining appropriate flowability through the use of surfactants. In some aspects, the absorption of injectable compositions is prolonged by using agents that delay absorption (such as aluminum monostearate and gelatin) in the composition.

[0313] Nasal delivery of preferred enantiomers of 2-hydroxycarboxylic acids is also considered. Nasal delivery allows preferred enantiomers of 2-hydroxycarboxylic acids to enter the bloodstream directly after the therapeutic product is applied to the nose, without depositing them in the lungs. Compositions for nasal delivery include compositions containing dextran or cyclodextrin.

[0314] Oral delivery excipients

[0315] The compositions of this disclosure, which contain a preferred enantiomer of a 2-hydroxycarboxylic acid, can be delivered orally. For example, oral formulations can be used for topical application to a specific site in the gastrointestinal system (e.g., the esophagus, stomach, or large or small intestine) or for systemic administration, and can be delivered in a variety of forms, including simple solutions, syrups, suspensions, tablets, or capsules.

[0316] The compositions disclosed herein may contain water (aqueous) or may be non-aqueous.

[0317] The compositions disclosed herein may also include small amounts of conventional additives, such as preservatives, solubilizers (including ethanol), complexing agents (e.g., cyclodextrin), flow enhancers and gliding agents (including colloidal silica), flavor enhancers and compaction enhancers, including mixtures thereof. For some therapeutic agents, a buffer may be incorporated to maintain a suitable pH.

[0318] The oral compositions disclosed herein can be delivered together with an immune enhancer or modulator. Examples of immune enhancers or modulators include specific immune stimulants (e.g., vaccines and antigens) and nonspecific agents such as bee products (including propolis and honey), probiotics and prebiotics, hormones, vitamins (vitamin C, vitamin D), minerals (zinc oxide), antioxidants (including glutathione), interferons (including INF-α), interleukins (including interleukin-10), colchicine, thalidomide, and imiquimod. The immune enhancer can be delivered simultaneously with or after a preferred enantiomer composition of a 2-hydroxycarboxylic acid.

[0319] In some cases, it is believed that the administration of a preferred enantiomer of 2-hydroxycarboxylic acid can sufficiently dysregulate the metabolism of planktonic bacteria in the body, so that the simultaneous or subsequent administration of an immunostimulant can clear the planktonic bacteria from the body without the use of other antimicrobial agents (such as antibiotics).

[0320] Plant delivery excipients

[0321] The composition may contain active agrochemical compounds other than the preferred enantiomers of the 2-hydroxycarboxylic acid of the present invention.

[0322] The composition may also contain one or more of the following auxiliary components: an inert carrier, a surfactant (e.g., a binder or spreader), a stabilizer, and / or a dye. The composition may also be suspended in a carrier liquid, such as air, nitrogen, carbon dioxide, or a fumigant. The composition preferably contains adjuvants, such as extenders, solvents, spontaneous promoters, carriers, emulsifiers, dispersants, antifreeze agents, biocides, thickeners, and / or other adjuvants, such as adjuvants. In this context, an adjuvant is a component that enhances the biological activity of the composition, even if the component itself does not have biological activity. Examples of adjuvants are agents that promote the retention, spreading, adhesion to leaf surfaces, or penetration into plant tissues of preferred enantiomers of 2-hydroxycarboxylic acids. Generally, the active compound can be combined with any solid or liquid additive commonly used for formulation purposes.

[0323] The compositions of the present invention may also contain formulation adjuvants and additives known to those skilled in the art as formulation adjuvants (some of which may also be considered as solid diluents, liquid diluents, or surfactants). Such formulation adjuvants and additives can control: pH (buffers), foaming during processing (defoamers, such as polysiloxanes), sedimentation of active ingredients (suspending agents), viscosity (thixotropic or pseudoplastic thickeners), microbial growth within the container (antimicrobial agents), product freezing (antifreeze agents), color (dye / pigment dispersions), elution (film-forming agents or binders), evaporation (evaporation inhibitors), and other formulation properties. Film-forming agents include, for example, polyvinyl acetate, polyvinyl acetate copolymers, polyvinylpyrrolidone-vinyl acetate copolymers, polyvinyl alcohol, polyvinyl alcohol copolymers, and waxes.

[0324] stability

[0325] Optionally, the compositions disclosed herein are stable. As used herein, the stability of the compositions provided herein refers to the length of time that the composition retains more than 80%, 85%, 90%, or 95% of the initial amount of antibiotic-lactic acid at a given temperature. For example, the compositions provided herein can be stored at about 15°C to about 30°C and remain stable for at least 1, 2, 12, 18, 24, or 36 months. Furthermore, the compositions can be adapted for administration to subjects in need after storage at 25°C for more than 1, 2, 12, 18, 24, or 36 months. Additionally, in another alternative embodiment, Arrhenius kinetics is used to retain more than 80%, or more than 85%, or more than 90%, or more than 95% of the initial amount of active substance (e.g., a preferred enantiomer of 2-hydroxycarboxylic acid) after storage at about 15°C to about 30°C for more than 1, 2, 12, 18, 24, or 36 months.

[0326] As used herein, the expression "stable during long-term storage" means that the composition is suitable for administration to subjects in need when it has an estimated shelf life of more than 1, 2, or 3 months of use at 25°C and more than or equal to 1, 2, or 3 years of storage at 5°C. In some embodiments herein, Arrhenius kinetics is used to estimate the retention of >80% or >85% or >90% or >95% of the preferred enantiomeric 2-hydroxycarboxylic acid after such storage.

[0327] antimicrobial agents

[0328] Other active agents may also be incorporated into the compositions disclosed herein. For example, additional antimicrobial compounds, such as antibacterial agents, antifungal agents, etc., may be incorporated.

[0329] The adjunct active agent provided with the preferred enantiomer of the 2-hydroxycarboxylic acid of this disclosure can be a bactericidal active agent, but not an antibiotic. For example, the adjunct active agent can be a chlorine-based compound (e.g., a bleach), an iodine-based compound, a copper-based compound, etc.

[0330] Preferably, the preferred enantiomers of the 2-hydroxycarboxylic acid of this disclosure cause metabolic dysregulation in planktonic bacteria, allowing the adjuvant surfactant (antibiotic or other compound capable of killing bacteria) to exert its bactericidal effect more effectively. Optionally, the metabolic dysregulation in planktonic bacteria is to the extent that the bacteria cannot resist the action of the adjuvant surfactant. More preferably, the preferred enantiomers of the 2-hydroxycarboxylic acid of this disclosure cause metabolic dysregulation in planktonic bacteria such that the planktonic bacteria do not develop resistance to the adjuvant surfactant.

[0331] Dysregulation of planktonic bacterial metabolism can be either inhibition or enhancement in the presence of a preferred enantiomer of 2-hydroxycarboxylic acid alone. Upon introduction of an antimicrobial compound (exposing bacteria to both the antimicrobial compound and 2-hydroxycarboxylic acid), the dysregulation (inhibition or enhancement) can persist, and the degree of dysregulation may increase. However, once planktonic bacteria are simultaneously exposed to both the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound, the dysregulation of planktonic bacterial metabolism can change: a previous inhibition becomes enhancement, or a previous enhancement becomes inhibition. Alternatively, metabolic dysregulation may not exist in the presence of 2-hydroxycarboxylic acid alone, but significant dysregulation exists in the presence of both the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound, particularly when the planktonic bacteria are resistant to the antimicrobial compound in the absence of the preferred enantiomer of 2-hydroxycarboxylic acid.

[0332] The composition may also contain benzoyl peroxide, or antibiotics such as erythromycin, clindamycin, doxycycline, or meclocycline.

[0333] Additional antimicrobial compounds that may be used include, but are not limited to, silver compounds (e.g., silver chloride, silver nitrate, silver oxide), silver ions, silver particles, iodine, povidone / iodine, chlorhexidine, 2-p-sulfanilyanilinoethanol, 4,4'-sulfinyldiphenylamine, 4-sulfasalic acid, sulfadiazine, acetylsulfone, amikacin, amoxicillin, amphotericin B, ampicillin, apalcillin, apicycline, apramycin, arbekacin, and apocytin. xicillin), chloramphenicol, azithromycin, aztreonam, bacitracin, flavomycin, biapenem, bromopyrrin, butirosin, capreomycin, carbenicillin, carbamate, carumonam, cefamandole, cefadroxil, cefadroxil, cefadroxil, cefotaxime, cefotiam, cefotaxime, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam, cefotiam Cefotaxime, Ceftazidime, Cefotaxime, Cefotiam, Cefoxitin, Ceftriaxone, Cefazolin, Cefotaxime, Cefotaxime C, Cefradine, Chloramphenicol, Aureomycin, Ciprofloxacin, Clarithromycin, Clindamycin, Clomocycline, Colistin, Cyclopemycin, Dapsone, Demethylchlortetracycline, Diathymosulfone, Dibekacin, Dihydrostreptomycin, Dierythromycin, Doxycycline, Enoxacin, Enviomycin, Epicillin, Erythromycin, Fluoxetine, Fortimycin Fortimicin(s), Gentamicin, Sulfone Glucoside, Sulfone Phenylacetyl, Bacitracin S, Bacitracin, Grepafloxacin, Guanicocycline, Hetacillin, Imipenem, Isepamicin, Josamycin, Kanamycin(s), Leukomycin, Lincomycin, Lomefloxacin, Lucentomycin, Lysylcycline, Meclocycline, Meropenem, Methoxycycline, MicronomicinMidecamycin(s), minocycline, moxalactam, mupirocin, nadifloxacin, natamycin, neomycin, netilmicin, norfloxacin, pyruvicin, oxytetracycline, p-sulfanilylbenzylamine, panipenem, paromomycin, pazufloxacin In), penicillin N, piperacillin, pipemidic acid, polymyxin, primamycin, quinacillin, ribosomycin, rifamide, rifampin, rifamycin SV, rifapentine, rifaximin, ristocetin, ritipenem, rokitamycin, hydropyridine, rosaramycin, roxithromycin, sulfasalazine, kasugamycin, sisomicin, sparfloxacin, and more. Spectinomycin, Spiramycin, Streptomycin, Dapsone succinate, Sulfacoic acid, Sulfamethoxazole, Sulfamicin, P-aminobenzenesulfonic acid, Sodium sulfadiazine, Teicoplanin, Temafloxacin, Temocillin, Tetracycline, Tetraoxorphine, Thiazole sulfone, Thioxetine, Ticarcillin, Tigemonam, Tobramycin, Tosufloxacin, Trimethoprim, Trospectomycin, Trovafloxacin, Tuberculosis Cytosin, vancomycin, azoserine, candidacloprid, chlorpheniramine, nystatin, filipin, nystatin, mepartricin, nystatin, oligomycin, ciprofloxacin, norfloxacin, ofloxacin, pefloxacin, enoxacin, rosoxacin, amoflavone, fleroxacin, temafloxacin, lomefloxacin.Epimycin A or tuberculin, etc.

[0334] use

[0335] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in the manufacture of compositions for the treatment of metabolic disorders in planktonic bacteria.

[0336] This disclosure also provides the use of preferred enantiomers of 2-hydroxycarboxylic acids for the treatment of metabolic disorders in planktonic bacteria.

[0337] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in combination with antimicrobial compounds in the manufacture of compositions for treating metabolic disorders of planktonic bacteria.

[0338] This disclosure also provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in combination with antimicrobial compounds for the treatment of metabolic disorders in planktonic bacteria.

[0339] This disclosure also provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in the manufacture of compositions for treating or preventing infection in a subject, wherein the infection is caused by planktonic bacteria, and wherein the composition causes metabolic dysregulation of the planktonic bacteria.

[0340] This disclosure also provides the use of a preferred enantiomer of a 2-hydroxycarboxylic acid in combination with an antimicrobial compound in the manufacture of a composition for treating or preventing infection in a subject, wherein the infection is caused by planktonic bacteria, and wherein the composition causes metabolic dysregulation of the planktonic bacteria.

[0341] This disclosure also provides the use of preferred enantiomers of 2-hydroxycarboxylic acids for the treatment or prevention of infection in a subject, wherein the infection is caused by planktonic bacteria, and wherein the composition causes metabolic dysregulation of the planktonic bacteria.

[0342] This disclosure also provides the use of a preferred enantiomer of a 2-hydroxycarboxylic acid in combination with an antimicrobial compound for the treatment or prevention of infection in a subject, wherein the infection is caused by planktonic bacteria, and wherein the composition causes metabolic dysregulation of the planktonic bacteria.

[0343] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in the manufacture of compositions for making planktonic bacteria sensitive to microbial compounds, wherein the preferred enantiomers of 2-hydroxycarboxylic acids induce metabolic dysregulation in planktonic bacteria.

[0344] This disclosure also provides the use of preferred enantiomers of 2-hydroxycarboxylic acids for sensitizing planktonic bacteria against microbial compounds, wherein the preferred enantiomers of 2-hydroxycarboxylic acids induce metabolic dysregulation in planktonic bacteria.

[0345] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in the manufacture of compositions for making planktonic bacteria sensitive to antimicrobial compounds, wherein the preferred enantiomers of 2-hydroxycarboxylic acids induce metabolic dysregulation in planktonic bacteria, and wherein the preferred enantiomers of 2-hydroxycarboxylic acids are used in combination with antimicrobial compounds.

[0346] This disclosure also provides the use of preferred enantiomers of 2-hydroxycarboxylic acids for sensitizing planktonic bacteria to antimicrobial compounds, wherein the preferred enantiomers of 2-hydroxycarboxylic acids induce metabolic dysregulation in planktonic bacteria, and wherein the preferred enantiomers of 2-hydroxycarboxylic acids are combined with antimicrobial compounds.

[0347] This disclosure also provides the use of preferred enantiomers of 2-hydroxycarboxylic acid in the manufacture of compositions for treating or preventing infection in a subject, wherein the infection is caused by planktonic bacteria, wherein the composition sensitizes the planktonic bacteria to an antimicrobial compound, and wherein the preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in the planktonic bacteria.

[0348] This disclosure also provides the use of a preferred enantiomer of a 2-hydroxycarboxylic acid in combination with an antimicrobial compound in the manufacture of a composition for treating or preventing infection in a subject, wherein the infection is caused by planktonic bacteria, wherein the composition sensitizes the planktonic bacteria to the antimicrobial compound, and wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0349] This disclosure also provides the use of a preferred enantiomer of a 2-hydroxycarboxylic acid for the treatment or prevention of infection in a subject, wherein the infection is caused by planktonic bacteria, wherein the composition sensitizes the planktonic bacteria to an antimicrobial compound, and wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0350] This disclosure also provides the use of a preferred enantiomer of a 2-hydroxycarboxylic acid in combination with an antimicrobial compound for the treatment or prevention of infection in a subject, wherein the infection is caused by planktonic bacteria, wherein the composition sensitizes the planktonic bacteria to the antimicrobial compound, and wherein the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0351] In the aforementioned applications, the dysregulation of planktonic bacterial metabolism in the presence of a preferred enantiomer of 2-hydroxycarboxylic acid alone can be either an inhibition or enhancement of planktonic bacterial metabolism. Upon introduction of an antimicrobial compound, the dysregulation (inhibition or enhancement) can persist, and the degree of dysregulation may increase. However, once planktonic bacteria are simultaneously exposed to both the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound, the dysregulation of planktonic bacterial metabolism can change: the original inhibition becomes enhancement, or the original enhancement becomes inhibition. Alternatively, metabolic dysregulation may not exist in the presence of 2-hydroxycarboxylic acid alone, but significant dysregulation exists in the presence of both the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound, particularly when the planktonic bacteria are resistant to the antimicrobial compound in the absence of the preferred enantiomer of 2-hydroxycarboxylic acid.

[0352] Preferred enantiomers of 2-hydroxycarboxylic acids may be in the same composition as the antimicrobial compound, or may be in a separate composition. Planktonic bacteria may be associated with or cause infection in the subject, or planktonic bacteria may be present on surfaces, such as non-biological surfaces. Planktonic bacteria may be present in liquids (biological or non-biological) or gases.

[0353] If a preferred enantiomer of a 2-hydroxycarboxylic acid is in a separate composition with an antimicrobial compound, the preferred enantiomer of the 2-hydroxycarboxylic acid may be administered simultaneously with the antimicrobial compound, or may be administered at a different time from the antimicrobial compound.

[0354] For the above-described uses, the preferred enantiomeric form of the 2-hydroxycarboxylic acid may be D-lactic acid. Alternatively, the infection or colonization may be bacterial infection or colonization.

[0355] Reagent test kit

[0356] This disclosure provides a kit for treating metabolic disorders of planktonic bacteria, comprising: a) Preferred enantiomers of 2-hydroxycarboxylic acids; and b) Instructions for use.

[0357] This disclosure provides a kit for treating metabolic disorders of planktonic bacteria, comprising: a) Preferred enantiomers of 2-hydroxycarboxylic acids; and b) Instructions for use The preferred enantiomer of the 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0358] This disclosure provides a kit for sensitizing planktonic bacteria against microbial compounds, comprising: a) Preferred enantiomers of 2-hydroxycarboxylic acids; and b) Instructions for use.

[0359] The preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic disorders in planktonic bacteria.

[0360] This disclosure provides a kit for sensitizing planktonic bacteria against microbial compounds, comprising: a) Preferred enantiomers of 2-hydroxycarboxylic acids; and b) Instructions for use The preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0361] In the aforementioned kit, the dysregulation of planktonic bacterial metabolism in the presence of only the preferred enantiomer of 2-hydroxycarboxylic acid can be either inhibition or enhancement of planktonic bacterial metabolism. Upon introduction of an antimicrobial compound, the dysregulation (inhibition or enhancement) can persist, and the degree of dysregulation may increase. However, once planktonic bacteria are simultaneously exposed to both the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound, the dysregulation of planktonic bacterial metabolism can change: the original inhibition becomes enhancement, or the original enhancement becomes inhibition. Alternatively, metabolic dysregulation may not exist in the presence of 2-hydroxycarboxylic acid alone, but significant dysregulation may exist in the presence of both the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound, particularly when the planktonic bacteria are resistant to the antimicrobial compound in the absence of the preferred enantiomer of 2-hydroxycarboxylic acid.

[0362] The preferred enantiomeric form of the 2-hydroxycarboxylic acid in the kit is D-lactic acid.

[0363] When the components of the kit are provided as one or more liquid solutions, the liquid solutions may be aqueous solutions, such as sterile aqueous solutions. For in vivo use, the expression construct may be formulated into a pharmaceutically acceptable injectable composition. In this case, the container device itself may be an inhaler, syringe, pipette, eye dropper, or other similar device, which may be used to apply the composition to the affected area of ​​an animal, such as the lungs, to inject it into the animal, or even to be applied to and mixed with other components of the kit.

[0364] In one embodiment, the kit of this disclosure comprises a composition containing a preferred enantiomer of a 2-hydroxycarboxylic acid, optionally in combination with an antimicrobial compound. In an alternative embodiment, the composition is pre-measured, pre-mixed, and / or pre-packaged.

[0365] The kits disclosed herein may also include instructions for use designed to promote user compliance. As used herein, instructions for use refers to any label, insert, etc., and may be located on one or more surfaces of the packaging material, or may be provided on a separate sheet of paper, or any combination thereof. For example, in one embodiment, the kits disclosed herein include instructions for use of the compositions of the disclosed invention. In one embodiment, the instructions for use indicate that the compositions of the disclosed invention are suitable for metabolic disorders of planktonic bacteria. Such instructions for use may also include dosage instructions and administration instructions.

[0366] Preferred enantiomers of 2-hydroxycarboxylic acids and suitable excipients can be packaged separately so that practitioners or users can formulate pharmaceutically acceptable compositions as needed. Alternatively, antisense oligomers and suitable excipients can be packaged together, thereby requiring minimal combination work from the practitioner or user. In any case, the packaging should maintain the chemical, physical, and aesthetic integrity of the active ingredient.

[0367] General instructions

[0368] Those skilled in the art will understand that the invention described herein is readily adaptable to variations and modifications beyond those specifically described. This invention includes all such variations and modifications. It also includes all steps, features, compositions, and compounds individually or collectively mentioned or indicated in the specification, as well as any and all combinations or any two or more steps or features.

[0369] Every document, reference, patent application, or patent cited in this article is explicitly incorporated in its entirety by reference, meaning that the reader should read and consider it as part of this article. References, patent applications, or patents cited in this article are not repeated here solely for the sake of brevity.

[0370] Descriptions, specifications, and instructions of any product by any manufacturer mentioned in any document incorporated herein by reference are hereby incorporated by reference and may be used in the practice of this invention.

[0371] The scope of this invention is not limited to any of the specific embodiments described herein. These embodiments are for illustrative purposes only. Functionally equivalent products, compositions, and methods are obviously within the scope of the invention described herein.

[0372] The invention described herein may include one or more ranges of values ​​(e.g., dimensions, displacements, and field strengths). A range of values ​​will be understood to include all values ​​within that range, including the value defining the range and values ​​adjacent to the range that produce the same or substantially the same result as the value immediately adjacent to the value defining the boundary of the range. Therefore, unless stated to the contrary, the numerical parameters set forth in the specification and claims are approximate values ​​that may vary depending on the desired characteristics sought to be obtained according to the invention. Thus, "about 80%" means both "about 80%" and "80%". At a minimum, each numerical parameter should be interpreted according to the number of significant figures and common rounding methods.

[0373] Throughout this specification, unless the context otherwise requires, the word “comprise” or variations thereof, such as “comprises” or “comprising”, shall be understood to imply inclusion of the stated integers or groups of integers, but not to exclude any other integers or groups of integers. It should also be noted that in this disclosure, particularly in the claims and / or paragraphs, terms such as “comprises,” “comprised,” “comprising,” etc., may have the meanings conferred upon them by U.S. patent law; for example, they may mean “includes,” “included,” “including,” etc.; and terms such as “consisting essentially of” and “consists essentially of” have the meanings conferred upon them by U.S. patent law, for example, they allow elements not expressly stated, but exclude elements found in the prior art or elements affecting the essential or novel features of the invention.

[0374] Other definitions of the selected terms used herein can be found and are consistent throughout the specific embodiments of the invention. Unless otherwise defined, all other scientific and technical terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. The term "activator" may refer to one activator or may encompass two or more activators.

[0375] The following examples are provided to more fully describe how the invention is used and to illustrate the best modes of carrying out various aspects of the invention. It should be understood that these methods are not intended to limit the true scope of the invention, but are presented for illustrative purposes.

[0376] Example

[0377] Other features of the invention are described more fully in the following non-limiting embodiments. This description is included for illustrative purposes only and should not be construed as limiting the broad description of the invention as described above.

[0378] Example 1: Inhibition of planktonic bacterial metabolism This study shows that D-lactic acid inhibits the metabolism of planktonic bacteria.

[0379] Materials and methods

[0380] Planktonic bacteria culture

[0381] Collected from patients with cystic fibrosis Pseudomonas aeruginosa Phenotypic analysis was performed on clinical isolates, which were then stored as a 50% glycerol stock solution at -70°C ± 2°C. Clinical isolates were thawed and streaked onto blood agar plates for growth at 37°C and 5% CO2 for 12–24 hours, or until single colonies large enough for further culture were selected. Single colonies were picked and inoculated into 5 mL of LB broth and incubated overnight with gentle shaking at 37°C and 5% CO2 to reach the stationary phase (B0).

[0382] The B0 culture was diluted 1:1000 in Pseudomonas basal medium (referred to as PMM in this paper) to achieve approximately 10 8 CFU / mL, then 150 µL of each diluted culture was transferred to the wells of a 96-well microplate and incubated overnight at 37°C and 5% CO2. D-lactic acid was then applied by adding 50 µL of stock solution appropriately diluted in PMM (or using only the medium in the case of control wells) and incubating for another 24 hours at 37°C and 5% CO2. 20 µL of culture supernatant was then transferred to wells containing 180 µL of fresh PMM, and the culture density was determined by measuring absorbance at 595 nm (using a ThermoFisher Multiskan FC microplate reader). Metabolic activity was then determined using these diluted cultures by resazurite staining.

[0383] Blade Blue Dye

[0384] Changes in bacterial metabolic activity after treatment were measured using rezamidium staining (Kirchner et al., 2012). 10 μl of 0.02% (v / v) rezamidium (diluted in distilled water) was added to each well, and the microplate was incubated aerobically at 37°C for 1–2 h with shaking at 150 rpm. Live cells reduced the blue rezamidium dye to the pink fluorescent halogen form. After incubation, fluorescence was quantified using an Enspire microplate reader at an excitation wavelength of 540 nm and an emission wavelength of 590 nm.

[0385] Statistical analysis

[0386] Unless otherwise stated, all data provided represent data from 24 plants. Pseudomonas aeruginosa aeruginosa) Data from clinical isolates, including two technical replicates, were used. Student's t-test was employed to assess significance compared to untreated control biofilms, with a significance level set at p < 0.05.

[0387] result

[0388] The fluorescence of halogen was measured to quantify the effect of D-lactic acid on the metabolic activity of planktonic cultures. A 90% inhibition of metabolic activity was observed in cultures treated with D-lactic acid, with the most significant changes observed in D-lactic acid concentrations ranging from 0.25 mg / mL to 0.5 mg / mL (Figure 1).

[0389] D-lactic acid does not decrease significantly. Pseudomonas aeruginosa It affects planktonic biomass, but has a significant impact on baseline metabolic activity.

[0390] Example 2: Inhibition of planktonic bacterial metabolism increases the sensitivity of microorganisms to antimicrobial compounds. This study shows that D-lactic acid treatment increases the sensitivity of planktonic bacteria to antibiotics to such an extent that D-lactic acid treatment can reverse antimicrobial resistance.

[0391] Materials and methods

[0392] airborne bacteria culture

[0393] The bacterial strain was maintained as a suspension in 20% (v / v) glycerol / brain heart extract at -80°C. Before use, a ring of ice was scraped from the stock solution and streaked onto a rich medium containing 1.8% agar to obtain single colonies (for...). verdigris fake order Pseudomonas aeruginosa or Klebsiella pneumoniae Luria-Bertani medium; for Staphylococcus aureus or Streptococcus pneumoniae (Streptococcus pneumoniae)(trypsin-soybean agar) and incubated overnight at 37°C.

[0394] Determination of minimum inhibitory concentration by microdilution method of broth

[0395] Standard inoculum was prepared from broth cultures. One to three colonies from an overnight agar plate were transferred to 5 ml of broth and incubated at 37°C with shaking (200 rpm) until the turbidity was greater than 0.5 McFarland standard (overnight). The following broth media were used: cationic-regulated Mueller-Hinton broth (caMHB - all strains); Roswell Park Memorial Institute medium (RPMI 1640, phenol red-free). Staphylococcus aureus Trypsin-enriched soy broth (TSB) Staphylococcus aureus and Streptococcus pneumoniae ); Todd Hewitt Broth (THB – Streptococcus pneumoniae M63 basal medium supplemented with 0.2% glucose (M63 – Klebsiella pneumoniae .

[0396] The broth culture was diluted in broth to a turbidity equal to 0.5 McFarland standard at 620 nm using a spectrophotometer. The 0.5 McFarland standard was further diluted in broth at a 1:100 ratio to produce a standard inoculum. 50 µl of this standard inoculum was dispensed into each well of a microdropper dish, except for the negative control, which was dispensed with 50 µl of broth.

[0397] Next, working concentrations of each antimicrobial agent were generated in broth using the stock solution, twice the highest target concentration. 100 µl of this working concentration was pipetted into the first well, and 50 µl of broth culture was pipetted into all other wells. 50 µl was taken from the first well and serially diluted in all wells of the row until the final concentration of the range was reached. 50 µl was taken from the last well and discarded. This produced a concentration range where each well contained twice the target concentration. All reagents were performed in duplicate.

[0398] The 96-well plates were incubated at 37°C for 18 ± 2 hours. After incubation, the MIC was visually assessed and recorded as the lowest concentration of antimicrobial agent that completely inhibited bacterial growth (determined by complete absence of turbidity).

[0399] Razorazine reduction used to assess metabolic activity

[0400] After incubation and visual recording of the MIC, or in separate studies, aerobic respiration was measured in the presence or absence of D-lactic acid and in the presence or absence of accompanying antibiotics. For evaluation after MIC determination, the entire volume (100 µl) of the MIC well was mixed by pipetting back and forth and then transferred to the corresponding well in a 96-well plate suitable for fluorescence-absorbance detection. Alternatively, the cultures were pipetted together as described above and incubated overnight at 37°C in the absence of any antimicrobial compounds. The culture supernatant was removed, adherent cells were washed, and then treated with 100 µl of fresh broth, 50 µl of broth containing 4 times the required concentration of antibiotic, and 50 µl of control containing 4 times the required concentration of D-lactic acid or only the medium, followed by a re-incubation at 37°C for 18 ± 2 h. After both incubation protocols were completed, resazurin reduction was assessed by adding 10 µl of 0.02% resazurin solution to each well and incubating the plate again for 90 min with gentle shaking (40 rpm) at 37°C. 96-hole plates and azure blades should be kept out of light as much as possible.

[0401] After incubation, the 96-well plate was read on a fluorescence-absorption spectrophotometer using an excitation frequency of 565 nm and an emission frequency of 590 nm. Blank correction was performed on the values ​​of each well by subtracting the values ​​of the negative controls. Each value was then converted to a ratio relative to the mean of the positive controls.

[0402] result

[0403] The combined administration of D-lactic acid reduced the minimum inhibitory concentration (MIC) of multiple antibiotics.

[0404] First, for medically relevant bacterial strains / antibiotic combinations, the MICs of multiple antibiotics (used alone or in combination with D-lactic acid) were determined. The results, shown in Table 3, indicate that D-lactic acid treatment resulted in a minimum two-fold reduction in MICs, meaning that the efficacy of the antibiotics against the bacterial strains in question was at least twice as strong.

[0405] Table 3: Effects of combined administration of D-lactic acid on antibiotic MIC

[0406] Antibiotic treatment further enhanced the inhibition of D-lactic acid on the metabolism of planktonic bacteria.

[0407] The fluorescence of halogenated thiocyanate was measured to quantify the effect of D-lactic acid combined with bactericidal or bacteriostatic antibiotics on the metabolic activity of planktonic cultures. Data are summarized in Table 4 and shown below. Figures 2-10 middle.

[0408] Table 4: Effects of D-lactic acid administration on bacterial metabolic activity with or without accompanying antimicrobial agents, as measured by certain metabolic dysregulation indicators, such as fluctuations in resazurin activity or a decrease in the MIC of the accompanying antimicrobial agent.

[0409] 1. Resazu activity refers to the increase (↑) or decrease (↓) in resazu reduction when treated with D-lactic acid in the absence of any accompanying antimicrobial agents, compared to treatment with the medium alone.

[0410] 2. Resazu activity refers to the increase (↑) or decrease (↓) in resazu reduction when treated with D-lactic acid and the indicated accompanying antimicrobial agent, compared to treatment with the medium alone.

[0411] 3. Enhanced antimicrobial efficacy refers to the increase (↑) or decrease (↓) in the efficacy of the indicated antimicrobial agent (MIC measured in vitro) compared to the efficacy in the absence of D-lactic acid (i.e., its MIC decreases or increases, respectively).

[0412] In all cases, "NT" means "untested".

[0413] The decreased metabolic activity observed after antibiotic treatment was further enhanced upon combination with D-lactic acid. Figures 2-10 In each figure, the histogram shows the amount of metabolic activity measured by halogen fluorescence relative to a control of the medium alone. Relative metabolic activity for antibiotic treatment alone is shown in black, and relative metabolic activity for antibiotic treatment with D-lactic acid is shown in gray.

[0414] Example 3: Dysregulation of planktonic bacterial metabolism is specific for D-enantiomers. This study demonstrates that the metabolic dysregulation of Pseudomonas aeruginosa observed in the presence of D-lactic acid does not recur in the presence of L-lactic acid.

[0415] Materials and methods

[0416] airborne bacteria culture

[0417] At -80℃, Pseudomonas aeruginosa Strain WACC91 was maintained as a suspension in 20% (v / v) glycerol / brain-heart extract. Before use, a ring of ice was scraped from the stock solution and streaked onto LB medium containing 1.8% agar to obtain a single colony, which was then incubated overnight at 37°C. A single colony was picked and inoculated into 5 mL of LB broth, and gently shaken at 37°C until the turbidity was greater than 1.0 McFarland standard (3–4 hours).

[0418] The broth culture was diluted to a turbidity equal to 1.0 McFarland standard in M63 basal medium containing 0.2% (w / v) glucose as the sole carbon source at 595 nm using a spectrophotometer. The culture was then diluted 1:100 in M63 (glucose), and 100 μL along with an additional 50 μL was transferred to each well of a 96-well microdroplet and incubated overnight at 37°C and 5% CO2. D-lactic acid, L-lactic acid, or mediator-only (sterile H2O) was then applied by adding 50 μL of the appropriate stock solution diluted in M63 (glucose) and incubating for another 24 hours at 37°C and 5% CO2. The metabolic activity of the culture was then determined using the resazure reduction method as described in Example 1 above.

[0419] result

[0420] The fluorescence of halogenated halogen was measured to quantify the effects of D-lactic acid or L-lactic acid on the metabolic activity of planktonic cultures. >50% inhibition of metabolic activity was observed in cultures treated with D-lactic acid (compared to media-only treatment), with the most significant changes observed in D-lactic acid concentrations ranging from 12.5 mM to 25 mM (Figure 1). No changes in metabolic activity were observed in cultures treated with L-lactic acid. Therefore, the observed metabolic dysregulation is specific to the D-enantiomer of lactate.

[0421] Figure 11 This involves adding D-lactic acid or L-lactic acid at a concentration reduced by half. Pseudomonas aeruginosa Histogram of results from WACC91 planktonic culture. This plot shows that the treatment range resulted in a breakpoint between 12.5 mM and 25 mM D-lactic acid when the metabolic rate (reduction of resamarium to fluorescent halogen) was used as a reading compared to mediator-only treatment. No comparable breakpoint was observed when L-lactic acid was added at the same concentration as observed for D-lactic acid.

[0422] Example 4: D-lactate enhances the antibacterial activity of metal ions. This study shows that the antibacterial activity against planktonic bacteria is enhanced when metal ions are present in the form of D-lactate.

[0423] Materials and methods

[0424] airborne bacteria culture

[0425] The bacterial strain was maintained as a suspension in 20% (v / v) glycerol / brain heart extract at -80°C. Before use, a ring of ice was scraped from the stock solution and streaked onto trypsin-soy agar containing 1.8% agar to obtain a single colony, which was then incubated overnight at 37°C.

[0426] Determination of minimum inhibitory concentration by microdilution method of broth

[0427] Standard inoculum was prepared from broth culture. One to three colonies from an overnight agar plate were transferred to 5 ml of broth and incubated at 37°C with shaking (200 rpm) until the turbidity was greater than 0.5 McFarland standard (overnight). All assays were performed in trypsin-treated soy broth.

[0428] The broth culture was diluted in broth to a turbidity equal to 0.5 McFarland standard at 620 nm using a spectrophotometer. The 0.5 McFarland standard was further diluted in broth at a 1:100 ratio to produce a standard inoculum. 50 µl of this standard inoculum was dispensed into each well of a microdropper dish, except for the negative control, which was dispensed with 50 µl of broth.

[0429] Next, working concentrations of each salt (in aqueous solution) were generated in the broth using the stock solution, which were twice the highest target concentration. 100 µl of this working concentration was pipetted into the first well, and 50 µl of the broth culture medium was pipetted into all other wells. 50 µl was taken from the first well and serially diluted in all wells of the row until the final concentration of the range was reached. 50 µl was taken from the last well and discarded. This produced a concentration range where the concentration in each well was twice the target concentration. All reagents were performed in duplicate.

[0430] The 96-well plates were incubated at 37°C for 18 ± 2 hours. After incubation, the MIC was visually assessed and recorded as the lowest concentration of antimicrobial agent that completely inhibited bacterial growth (determined by complete absence of turbidity).

[0431] result

[0432] The minimum inhibitory concentration (MIC) decreases in the presence of D-lactate.

[0433] The minimum inhibitory concentrations (MICs) of zinc (Zn) or copper (Cu) salts, present in the form of nitrate or D-lactate, were determined. The results are shown in Table 4, indicating that the presence of D-lactate resulted in a minimum two-fold reduction in the MIC, meaning that the metal ion efficacy against the bacterial strains in question was at least twice enhanced.

[0434] Table 5: Effects of application as nitrate or D-lactate on the MIC of metal ions

[0435] Example 6: Inhibition of planktonic bacterial metabolism increases the susceptibility of microorganisms to antimicrobial agents. This study demonstrates that in a mouse model of peritonosesepticemia, D-lactic acid treatment increased colistin resistance when compared to colistin treatment alone. Escherichia coli The susceptibility of the strain to colistin: (1) the severity of disease was reduced, as measured by viable counts in blood samples and bacterial load imaging in live animals; and (ii) the survival rate of mice was improved.

[0436] Materials and methods

[0437] Preparation of bacterial inoculum

[0438] bioluminescent col R E. coli Xen14 was streaked onto 5% sheep blood agar plates containing 30 µg / ml kanamycin and incubated overnight (o / n) at 37°C in normal air. The plates were first checked on a Lumina XRMS III instrument to ensure bioluminescence, and then a loop of the o / n growth was suspended in 10 mL of Luria Bertani (LB) broth until A was reached. 600 nm = 0.05. Incubate the suspension at 37°C for approximately 3 hours until A 600 nm = 0.5 (equivalent to approximately 5 x 10) 8 CFU / mL). The bacterial suspension was prepared at 4000 x g Centrifuge for 5 minutes, resuspend in phosphate-buffered saline (PBS, pH 7.2) and wash twice, resuspend again in PBS, then place on wet ice and use for mouse infection within 10 minutes of preparation, as described below.

[0439] Mouse infection study

[0440] These experiments used 6-7 week old outbred Swiss male (CD1) mice (approximately 26-33 g, 8 mice per group) obtained from the University of Adelaide Laboratory Animal Service Breeding Facility. The mice had free access to food and water. All animal experiments were reviewed and approved by the University of Adelaide Animal Ethics Committee (Approval No. S-2022-044). The study was conducted in accordance with the Australian Guidelines for the Care and Use of Animals for Scientific Purposes (8th Edition, 2013) and the South Australian Animal Welfare Act (1985).

[0441] Approximately 1 x 10⁻⁶ in 200 μL PBS containing 3% porcine gastric mucin type III (Sigma Aldrich). 8 CFU bioluminescence R E. coliXen14 was administered intraperitoneally (IP) to each mouse, followed by bioluminescence imaging at the ventral and dorsal positions using the IVIS Lumina XRMS Series III system under anesthesia. Next, the potency of sodium D-lactate (1000 mg / kg; pH 7.03) was tested as follows: Group: Group 1: Single oral administration of the medium only (sterile water only) Group 2: Single oral dose of 1000 mg / kg of D-lactic acid sodium (active ingredient 1) aqueous solution Group 3: A single oral dose of 1000 mg / kg D-lactic acid sodium at 0 hours, followed by four intraperitoneal injections of 1 mg / kg colistin (col) (active ingredient 2) at 0, 4, 8, and 12 hours. Group 4: Four intraperitoneal injections of 1 mg / kg colistin (subtherapy control) were administered at 0, 4, 8, and 12 hours. At 4 hours, blood samples (approximately 50 µL) were collected from individual mice in all groups via submandibular (facial vein) sampling for bacterial counting. Mice were frequently monitored for signs of distress, their clinical condition was recorded and imaged within 72 hours post-infection, and mice that were near death or showed any signs of distress were humanely euthanized by CO2 asphyxiation. Signals were collected from defined regions of interest in all experiments, and total flux intensity (photons / second) was analyzed using Living Image Software 4.7.3. Survival rates were recorded for each group. Differences in bioluminescent signals between groups were compared using an unpaired t-test (two-tailed).

[0442] result

[0443] As described above, approximately 1 x 10 in 200 μL PBS 8 CFU bioluminescence R E. coli Each mouse was attacked by intraperitoneal injection (IP) of Xen14 and then treated with a single oral dose of sterile water (negative control), a single oral dose of D-lactate sodium (1000 mg / kg), a single oral dose of D-lactate sodium (1000 mg / kg) combined with four intraperitoneal doses of colistin (1 mg / kg at 0, 4, 8 and 12 hours), or an intraperitoneal dose of four colistin (1 mg / kg at 0, 4, 8 and 12 hours) alone.

[0444] Treatment effectiveness was assessed using the following methods: colony count in a blood sample collected 4 hours after infection; and colony count against a fluorescently labeled colony. R E. coli Imaging of infected mice at concentrations; and survival rates ( Figures 12 to 14The colony counts in blood samples from mice treated with a single dose of 1000 mg / kg D-lactate followed by four doses of 1 mg / kg colistin were significantly lower than those from mice treated with four doses of 1 mg / kg colistin alone. A single dose of 1000 mg / kg D-lactate alone had no protective effect. (Regarding fluorescently labeled colonies...) R E. coli Comparable effects were observed in imaging and survival rates of infected mice at the same concentration.

Claims

1. A composition for inducing metabolic dysregulation in planktonic bacteria, said composition comprising a 2-hydroxycarboxylic acid of the following formula: 。 2. The composition according to claim 1, wherein the 2-hydroxycarboxylic acid is a preferred enantiomeric form of a 2-hydroxycarboxylic acid having the following formula: 。 3. The composition according to claim 2, further comprising a non-preferred enantiomeric form of a 2-hydroxycarboxylic acid having the following formula: 。 4. The composition according to claim 3, wherein the percentage of the non-preferred enantiomeric form of the 2-hydroxycarboxylic acid is less than 20% of the total 2-hydroxycarboxylic acid.

5. The composition according to claim 2, wherein the composition does not contain a non-preferred enantiomer of 2-hydroxycarboxylic acid.

6. The composition according to claim 1, wherein the imbalance is: (i) Inhibition of metabolism in the presence of 2-hydroxycarboxylic acid alone, and inhibition of metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; (ii) It inhibits metabolism in the presence of 2-hydroxycarboxylic acid alone, but enhances metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; (iii) Enhanced metabolism in the presence of 2-hydroxycarboxylic acid alone, and enhanced metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; (iv) It enhances metabolism in the presence of 2-hydroxycarboxylic acid alone, but inhibits metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; (v) It has no effect on metabolism in the presence of 2-hydroxycarboxylic acid alone, but inhibits metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; (vi) It has no effect on metabolism in the presence of 2-hydroxycarboxylic acid alone, but enhances metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds.

7. The composition of claim 1, wherein the dysregulation results in sensitization of the bacteria against the microbial compound.

8. The composition according to claim 1, wherein the 2-hydroxycarboxylic acid is D-lactic acid or a pharmaceutically acceptable salt thereof.

9. The composition according to claim 1, further comprising an antimicrobial compound.

10. A method for inducing metabolic disorder in planktonic bacteria, comprising the following steps: The planktonic bacteria are given a composition containing 2-hydroxycarboxylic acid according to any one of claims 1 or 2.

11. A composition comprising the 2-hydroxycarboxylic acid according to any one of claims 1 or 2, for treating or preventing a microbial infection in a subject, wherein the microbial infection is caused by planktonic bacteria, and wherein the 2-hydroxycarboxylic acid causes metabolic dysregulation of the planktonic bacteria.

12. Use of the 2-hydroxycarboxylic acid according to any one of claims 1 or 2 in the manufacture of a composition for causing metabolic disorder in planktonic bacteria.

13. Use of a composition comprising the 2-hydroxycarboxylic acid according to any one of claims 1 or 2 for the purpose of causing metabolic disorders in planktonic bacteria.

14. Use of 2-hydroxycarboxylic acid according to any one of claims 1 or 2 in the manufacture of a composition for treating or preventing a microbial infection in a subject, wherein the microbial infection is caused by planktonic bacteria, and wherein the composition causes metabolic dysregulation of the planktonic bacteria.

15. Use of a composition comprising the 2-hydroxycarboxylic acid according to any one of claims 1 or 2 for the treatment or prevention of a microbial infection in a subject, wherein the microbial infection is caused by planktonic bacteria, and wherein the 2-hydroxycarboxylic acid causes metabolic dysregulation of the planktonic bacteria.

16. Use of a composition comprising the 2-hydroxycarboxylic acid according to any one of claims 1 or 2 for causing metabolic dysregulation of planktonic bacteria on or in non-biological surfaces or in non-biological materials.

17. A kit for inducing metabolic dysregulation in planktonic bacteria, the kit comprising: a) a composition comprising the 2-hydroxycarboxylic acid according to any one of claims 1 or 2; and b) Instructions for use.

18. The method according to claim 10, the use according to any one of claims 11 to 16, or the kit according to claim 17, wherein, The imbalance is: (i) Inhibition of metabolism in the presence of 2-hydroxycarboxylic acid alone, and inhibition of metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; (ii) It inhibits metabolism in the presence of 2-hydroxycarboxylic acid alone, but enhances metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; (iii) Enhanced metabolism in the presence of 2-hydroxycarboxylic acid alone, and enhanced metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; (iv) It enhances metabolism in the presence of 2-hydroxycarboxylic acid alone, but inhibits metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; (v) It has no effect on metabolism in the presence of 2-hydroxycarboxylic acid alone, but inhibits metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds; (vi) It has no effect on metabolism in the presence of 2-hydroxycarboxylic acid alone, but enhances metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds.

19. The method of claim 10, the use of any one of claims 11 to 16, or the kit of claim 17, wherein the dysregulation results in sensitization of the bacteria against the antimicrobial compound.

20. The method of claim 10, the use of any one of claims 11 to 16, or the kit of claim 17, wherein the 2-hydroxycarboxylic acid is D-lactic acid or a pharmaceutically acceptable salt thereof.

21. The method of claim 10, the use of any one of claims 11 to 16, or the kit of claim 17, wherein the composition further comprises an antimicrobial compound.

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