Urease inhibitor composition for agricultural or pharmaceutical applications, and associated production methods

Abstract

The present invention relates to a urease inhibitor composition, which can be used in agricultural and pharmaceutical sectors. The composition comprises a urease inhibitor, specifically dihydrolevoglucosenone, and can include co-solvents, plant extracts, living microorganisms or extracts thereof.

Description

Urease Inhibiting Composition for Agricultural or Pharmaceutical Applications, and Associated Manufacturing Methods

[0001] The present invention relates to the use of dihydrolevoglucosenone (also called cyrene) as a urease inhibitor and to compositions comprising dihydrolevoglucosenone.

[0002] STATE OF THE ART

[0003] Urease inhibitors play a crucial role in various fields, such as medicine, where they can be used to treat infections and medical conditions related to excessive ammonia production, and agriculture, where they help reduce ammonia volatilization, thereby improving fertilizer efficiency and minimizing negative environmental impacts such as eutrophication of water bodies. Despite their importance, currently available urease inhibitors, such as hydroxylamine acetate, phosphorus thiophosphate, and NBPT (N-(n-butyl) thiophosphoric triamide), have several drawbacks, including their potential toxicity and adverse environmental impact. These substances can have ecotoxic effects, polluting aquatic and terrestrial ecosystems and posing risks to human and animal health.

[0004] NBPT, for example, can break down into byproducts (such as N-butyl phosphorothioic triamide) that are potentially harmful to the environment. Furthermore, studies have shown that NBPT and its breakdown products can persist in treated crops, raising questions about potential impacts on human health throughout the food chain. Finally, the cost of using NBPT can be significant, especially when high doses are required to achieve the desired efficacy, thus increasing the overall production cost for farmers.

[0005] In this context, it becomes imperative to develop safer, more economical and more ecological alternatives to reduce ammonia volatilization.

[0006] DESCRIPTION OF THE INVENTION

[0007] The present invention relates to an agricultural or pharmaceutical urease-inhibiting composition comprising at least one urease inhibitor being dihydrolevoglucosenone.

[0008] The present invention offers an innovative solution to the problems mentioned above by introducing the use of dihydrolevoglucosenone (also known as cyrene), a green solvent derived from biomass, as a urease inhibitor in medicine or agriculture. Indeed, the inventors have discovered, surprisingly, that cyrene, previously known primarily for its non-toxic and biodegradable solvent properties, offers a promising alternative to urease inhibitors due to its ability to effectively inhibit urease.

[0009] Furthermore, in the agricultural field, the inventors have discovered a significant synergy when Cyrene is used in combination with an additional urease inhibitor such as NBPT (N-(n-butyl) thiophosphoric triamide), considerably improving inhibitory efficacy compared to using the compounds alone. This synergy allows for a reduction in the required dosages and a corresponding decrease in the environmental impact of their use.

[0010] This application therefore aims to establish the use of dihydrolevoglucosenone, alone or in combination with NBPT, to inhibit urease, reducing the environmental impacts associated with traditional inhibitors while maintaining optimal agronomic efficiency.

[0011] According to a first aspect of the invention, the invention therefore relates to an agricultural urease-inhibiting composition, comprising at least one urease inhibitor being dihydrolevoglucosenone.

[0012] In the context of the present invention, dihydrolevoglucosenone refers to a chemical compound also known as cyrene, which is a green solvent derived from biomass. This compound is characterized by its non-toxicity and biodegradability, thus offering an environmentally friendly alternative to traditional urease inhibitors.

[0013] In the context of the present invention, a urease inhibitor refers to any substance or chemical composition capable of reducing or preventing the activity of the urease enzyme. This enzyme catalyzes the breakdown of urea into ammonia and carbon dioxide, and its inhibition reduces ammonia volatilization, thereby improving the efficiency of nitrogen fertilizers and reducing negative environmental impacts.

[0014] According to one embodiment, dihydrolevoglucosenone is present in an amount between 20% and 99.9% by total weight of the composition, with preferred concentrations ranging from 40% to 99.5% by total weight of the composition, and more preferably between 50% and 99% by total weight of the composition.

[0015] One of the advantages of the invention is that the use of dihydrolevoglucosenone eliminates the need for additional solvent(s) and / or urease inhibitor(s), since dihydrolevoglucosenone already acts as both a solvent and a urease inhibitor, thereby simplifying production and reducing costs and the complexity of the formulation. According to one embodiment of the invention, the agricultural formulation therefore comprises no solvent other than dihydrolevoglucosenone and / or no urease inhibitor other than dihydrolevoglucosenone.

[0016] According to one embodiment of the invention, the inventors have also surprisingly demonstrated a notable synergy when dihydrolevoglucosenone is used in combination with an additional urease inhibitor such as NBPT (N-(n-butyl) thiophosphoric triamide). According to another embodiment, the agricultural composition can therefore include an additional urease inhibitor, such as NBPT. In this configuration, the dihydrolevoglucosenone can be present in an amount of between 20% and 99%, preferably between 35% and 98%, and more preferably between 45% and 95% by weight of the total composition, while the additional urease inhibitor, preferably NBPT, can be present in an amount of between 1% and 65%, preferably between 2% and 60%, and more preferably between 5% and 55% by weight of the total composition.

[0017] According to one embodiment, the additional urease inhibitor is selected from: N-butyl thiophosphoric triamide (NBPT), miophosphoryl triamide, cyclohexyl phosphoric triamide, cyclohexyl thiophosphoric triamide, phosphoric triamide, hydroquinone, p-benzoquinone, hexamidocyclotriphosphazene, thiopyridines, thiopyrimidines, thiopyridine-N-oxides, N,A'-dihalo-2-imidazolidinone, N-halo-2-oxazolidinone, ammonium thiosulfate (ATS), N-cyclohexyl phosphoric triamide (CHPT), phenyl phosphorodiamidate (PPT) and 2-nitrophenyl phosphoric triamide (2-NPT), and mixtures thereof.

[0018] According to one embodiment, the agricultural composition may further comprise at least one plant extract(s), preferably having biostimulant properties and / or at least one live microorganism(s), and / or at least one extract(s) of microorganism(s) and / or at least one co-solvent or a combination thereof.

[0019] These additional components can provide synergistic benefits by enhancing the efficacy of the urease-inhibiting compound. They can further improve soil fertility by increasing nutrient availability, stimulating beneficial microbial activity, and improving soil structure. For plants, these components can enhance growth, strengthen resistance to disease and environmental stresses, and promote better nutrient uptake.

[0020] According to one embodiment, the plant extracts are selected from red clover extract (Trifolium pretense), neem extract (Azadirachta indica), clover extract (Equisetum arvense), consoude extract (Symphytum officinale), chamomile extract (Matricla), nettle extract (Urtica dioica), willow extract (Salix spp.), fern extract (Pteridium aquilinum), burdock extract (Arctium lappa), millepertuis extract (Hypericum perforatum), pissenlit (Taracum perforatum) extract, lavender extract (Lavandula angustifolia), thyme extract (Thymus vulgaris), rosemary extract (Rosmarinus officinalis), calendula extract (Calendula officinalis), peppermint extract (Mentha piperita), ginseng extract (Panax ginseng), ginseng extract officinale), aloe vera extract (Aloe barbadensis), turmeric extract (Curcuma longa), licorice extract (Glycyrrhiza glabra),fenugreek extract (Trigonella foenum-graecum), marigold extract (Calendula officinalis), horse chestnut extract (Aesculus hippocastanum), cypress extract (Cupressus sempervirens), juniper extract (Juniperus communis), garlic extract (Allium sativum), echinacea extract (Echinacea purpurea), chili pepper extract (Capsicum annuum), green tea extract (Camellia sinensis), and valerian root extract (Valeriana officinalis), and mixtures thereof.

[0021] According to one embodiment, plant extracts with biostimulating properties are selected from kelp extract, brown algae extract, comfrey extract, horsetail extract, nettle extract, aloe extract, green tea extract, dandelion extract, yucca extract, neem extract, chamomile extract, and red clover extract and mixtures thereof, preferably red clover extract.

[0022] According to one embodiment, the live microorganisms or extracts of microorganisms are selected from the families of Thiorhodaceae, Athiorhodaceae, Chlorobacterium, Nitrobacteraceae (preferably of the genus Nitrosomonas or Nitrobacter), Thiobacterium (preferably of the genus Thiobacillus), Methanomonas (preferably of the genera Hydrogenomonas, Carboxydomonas or Methanomonas), Caulobacterium, Siderocapsaceae, Pseudomonaceae (preferably of the genera Pseudomonas, Xanthomonas, Acetobacter or Azotomonas), Spirillaceae (preferably of the genera Vibrio, Cellvibrio, Cellfalcicula, Desulfovibrio or Spirillum), Azotobacteraceae (preferably of the genera Azotobacter, Beijerinckia or Derxia), Rhizobacteraceae (preferably of the genera Rhizobium, Chromobacterium or Agrobacterium), and Achrobacterium. preferably of the genus Achromobacter or Flavobacterium, Enterobacteriaceae preferably of the genera Escherichia, Proteus, Aerobacter or Serratia,Micrococcaceae, preferably of the genera Micrococcus or Sarcina; Brevibacteraceae, preferably of the genus Brevibacterium; Lactobacillaceae, preferably of the genera Streptococcus, Leuconostoc, or Lactobacillus; Corynebacteraceae, preferably of the genera Corynebacterium, Cellulomonas, or Arthrobacter; Bacillaceae, preferably of the genera Bacillus or Clostridium, and mixtures thereof. Preferably, live microorganisms or extracts of microorganisms are selected from the species Pichia guilliermondii, Azotobacter chroococcum, Azotobacter vinelandii, Azospirillum lipoferum, Bacillus megaterium, Bacillus aryabhattai, Bacillus amiloliquefaciens, Bacillus licheniformis, and Oceanobacillus picturae, and mixtures thereof.

[0023] Within the scope of the present invention, the concentration ranges for plant extracts may vary between 0.01% and 10% by total weight of the composition, with preferred concentrations ranging from 0.1% to 5% by total weight of the composition to optimize bioactive effects without causing phytotoxicity. Live microorganisms (MOs) or their extracts may be present in concentrations ranging from 0.01% to 5% by total weight of the composition, with preferences for ranges of 1% to 5% or 3% to 4% by total weight of the composition to ensure beneficial microbial activity and a positive impact on soil and plant health. These concentrations are chosen to maximize synergistic benefits, enhancing the efficacy of the urease-inhibiting composition while promoting soil fertility and plant vigor.

[0024] In one embodiment, the composition further comprises at least one nitrification inhibitor, preferably dicyandiamide. By adding a nitrification inhibitor, the liquid agricultural composition benefits from an additional action that further reduces nitrogen loss in gaseous form, thus increasing the overall efficiency of the fertilizer.

[0025] In the context of the present invention, a nitrification inhibitor refers to a chemical substance that slows the conversion of ammonia to nitrate by soil bacteria. This inhibition is crucial for reducing nitrogen losses in gaseous form (particularly nitrous oxide, a greenhouse gas) and for maintaining nitrogen in a form more available to plants over a longer period.

[0026] In one embodiment, non-limiting examples of nitrification inhibitors include one or more of the following: N-2,5-dichlorophenyl succinamic acid, dicyandiamide (DCD), zinc ethylene-bis-dithiocarbamate, 2,4,6-triehloroaniline, pentachlorophenol, thiourea, ammonium thiosulfate (ATS) or 3,4-dimethylpyrazole phosphate (DMPP), and mixtures thereof.

[0027] According to one embodiment, at least one nitrification inhibitor may be present in the liquid agricultural composition in a range of 0.5% to 75%, preferably between 5% and 50%, more preferably between 10% and 40% by total weight of the composition.

[0028] To stabilize and improve the performance of the composition, co-solvents, such as gamma-valerolactone (GVL), delta-valerolactone, gamma-butyrolactone (GBL), ethanol, methanol, isopropanol, propylene glycol, ethylene glycol, butanol, glycol butyl ether, dimethyl sulfoxide (DMSO), acetone, toluene, xylene, benzene, 1,4-dioxane, tetrahydrofuran (THF), n-hexane, cyclohexane, dichloromethane, chloroform, N-methyl-2-pyrrolidone (NMP), and dimethylformamide (DMF) and stabilizers, such as amine-based stabilizers, may be included.

[0029] Preferably, one of the co-solvents used is gamma-valerolactone (GVL). Gamma-valerolactone is an organic solvent known for its stability, low toxicity, and effective solvating properties. It can improve the solubility and stability of dihydrolevoglucosenone in the composition, which is beneficial for agricultural and pharmaceutical applications. In one embodiment, the agricultural composition does not include any solvent other than dihydrolevoglucosenone and gamma-valerolactone (GVL).

[0030] Additional elements such as buffers, flow aids, silicas, surfactants and dyes may also be added to improve the physical properties and application of the composition.

[0031] According to another aspect of the invention, the invention relates to a solid or liquid fertilizer composition comprising, based on the total weight of the composition: - between 90% and 99.99%, preferably between 99% and 99.99% by total weight of the composition, of one or more nitrogen fertilizer compounds; - between 0.01% and 10%, preferably between 0.01% and 1.5% by total weight of the composition, of the urease-inhibiting composition according to the invention or of a urease inhibitor being dihydrolevoglucosenone; - between 0% and 1.5% by total weight of the composition, of an additional urease inhibitor being preferably NBPT.

[0032] This dosage optimizes agronomic efficiency while minimizing environmental impacts.

[0033] Fertilizer composition can be solid or liquid.

[0034] In its solid form, nitrogen fertilizer is generally in the form of urea. Urea is one of the most commonly used nitrogen fertilizers due to its high nitrogen content, approximately 46%, and its ease of handling and application. It is applied directly to the soil, where it dissolves slowly, releasing ammonia which is then converted to nitrate by soil microorganisms. This solid form can be granulated or powdered, facilitating transport and storage. Urea granules can be coated with urease inhibitors such as dihydrolevoglucosenone, thus prolonging the availability of nitrogen to plants and reducing losses through ammonia volatilization.

[0035] In its liquid form, fertilizer is generally a nitrogen solution, such as urea-ammonium nitrate (UAN). Liquid nitrogen solutions are popular because they are easy to apply via drip irrigation, foliar sprays, or direct injection into the soil. These solutions allow for uniform nitrogen distribution and rapid absorption by plants, promoting fast and efficient growth. Liquid solutions can also be enriched with dihydrolevoglucosenone, as well as co-solvents and stabilizers, to improve solution stability and the effectiveness of urease inhibition. This helps reduce nitrogen losses and improve the availability of this essential nutrient for crops.

[0036] Whether in solid or liquid form, fertilizer composition can be adapted to include additives such as nitrification inhibitors, co-solvents, stabilizers, and plant and / or microorganism extracts to improve agronomic and environmental performance. The goal is to maximize nitrogen use efficiency, minimize losses through volatilization and leaching, and reduce the environmental impact of fertilization practices.

[0037] According to another aspect of the invention, the invention relates to a method of manufacturing a liquid or solid fertilizer composition, comprising a step of treating one or more nitrogen fertilizer compounds with a composition of urease inhibitors according to the invention comprising a urease inhibitor solvent being dihydrolevoglucosenone, and optionally at least one co-solvent.

[0038] This method can be implemented in two main ways:

[0039] Coating on urea (solid granules): Dihydrolevoglucosenone is applied as a coating layer to solid urea granules. This process involves spraying or dipping the granules in a solution containing dihydrolevoglucosenone, followed by drying to fix the urease inhibitor to the surface of the granules.

[0040] Mixing with nitrogen solution (liquid): Dihydrolevoglucosenone is also compatible with liquid nitrogen fertilizer formulations. In this case, the inhibitor is dissolved or dispersed in the nitrogen solution, creating a homogeneous mixture. The addition of co-solvents can be considered to improve the solubility and stability of dihydrolevoglucosenone in the solution.

[0041] According to another aspect of the invention, the agricultural composition according to the invention is a formulation in two separate preparations.

[0042] According to one embodiment, the first preparation comprises: gamma-valerolactone (GVL) and / or N-(n-butyl)-thiophosphoric triamide (NBPT) and optionally dihydrolevoglucosenone. This first preparation is stable and can be stored for extended periods without loss of efficacy.

[0043] According to one embodiment, the first preparation comprises, in total weight of the first preparation: between 0% and 70% by weight of dihydrolevoglucosenone, between 20% and 90% by weight of gamma-valerolactone (GVL) and / or between 15% and 70% by weight of N-(n-butyl)-thiophosphoric triamide (NBPT).

[0044] According to one embodiment, the second preparation comprises dihydrolevoglucosenone, water, and / or at least one plant extract, preferably at least one plant extract having biostimulating properties, and / or at least one live microorganism or their extracts.

[0045] Dihydrolevoglucosenone ensures compatibility and homogeneity in the final mixture when present in both preparations, or improves the stability of the final mixture when present only in the second preparation. Water acts as the primary solvent for the biological components, facilitating their dispersion and activity. Separating these components from those in the first preparation avoids undesirable interactions that could affect their stability and efficacy.

[0046] According to one embodiment, the second preparation comprises, by total weight of the second preparation: between 1% and 70% by weight of dihydrolevoglucosenone, between 20% and 80% by weight of water, and between 2% and 20% by weight of at least one plant extract, preferably red clover, and / or between 10% and 50% by weight of at least one live microorganism or their extracts.

[0047] Before spraying, the two preparations are mixed in specific proportions and then applied to the nitrogen fertilizer (either coated onto urea or mixed with the nitrogen solution) to form the final composition. This mixing step can be carried out beforehand or in the field, using standard spraying equipment fitted with mixing systems. Once mixed, the composition is applied directly to the crops or, preferably, to the soil, providing effective urease inhibition, reduced nitrogen losses through volatilization, and additional agronomic benefits from plant extracts and microorganisms.

[0048] According to one embodiment, the first preparation and the second preparation are mixed in a volume ratio of 6:1, 5.75:1, 5.5:1, 5.25:1, 5:1, 4.75:1, 4.5:1, 4.25:1, 4:1, 3.75:1, 3.5:1, 3.25:1, 3:1, 2.75:1, 2.5:1, 2.25:1, 2:1, 1.75:1, 1.5:1, 1.25:1, 1:1, 1:1.25, 1:1.5, 1:1.75, 1:2, 1:2.25, 1:2.5, 1:2.75, 1:3, 1:3.25, 1:3.5, 1:3.75, 1:4, 1:4,25, 1:4,5, 1:4,75, 1:5, 1:5,25, 1:5,5, 1:5,75, 1:6.

[0049] By separating the components into two distinct preparations, the stability problems often encountered with complex mixtures are resolved. This also extends the shelf life of the products, thus reducing losses and associated costs. Furthermore, the ability to mix the two preparations just before application ensures that the composition is in its most effective and reactive state, thereby optimizing its performance during spraying.

[0050] According to another aspect of the invention, the invention relates to a method of fertilizing target plants, comprising the application of a liquid or solid fertilizer composition which includes: one or more nitrogen fertilizer compounds, and a urease-inhibiting composition according to the invention or a urease-inhibiting solvent being dihydrolevoglucosenone.

[0051] Finally, according to another aspect of the invention, the invention relates to the use of dihydrolevoglucosenone as a urease inhibitor to reduce ammonia volatilization in agricultural applications. Dihydrolevoglucosenone can be used as the sole solvent and / or sole urease inhibitor.

[0052] According to one embodiment, dihydrolevoglucosenone is used as a urease inhibitor in combination with NBPT, in agricultural applications to significantly reduce ammonia volatilization, offering a more sustainable and environmentally friendly solution for crop fertilization.

[0053] Pharmaceutical Applications of the Invention:

[0054] Urease inhibition has significant benefits for both agriculture and medicine, where it can be used to treat infections and medical conditions related to excessive ammonia production.

[0055] In this respect, the present application also relates to a urease-inhibiting pharmaceutical composition, comprising at least one urease inhibitor being dihydrolevoglucosenone.

[0056] In a medical setting, this composition can be formulated in various dosage forms, such as tablets, capsules, injectable solutions, or oral suspensions.

[0057] For pharmaceutical applications, the pharmaceutical composition may include at least 1, 5, 10, 25, 50, 75, 95, 97, 99, 99.5, or 99.9% by total weight of dihydrolevoglucosenone, with specific preferences depending on the dosage form and therapeutic indication. High concentrations ensure effective urease inhibition, essential for treatments requiring rapid and substantial ammonia reduction, such as in severe urinary tract infections or ammonia-related metabolic disorders.

[0058] The formulation of a suitable composition can be achieved using standard pharmaceutical chemicals and formulation methodologies, all of which are readily available to a person skilled in the art. For example, the urease inhibitor can be combined with one or more pharmaceutically acceptable excipients or vehicles.

[0059] Auxiliary substances, such as wetting or emulsifying agents, pH buffers, reducing agents, and the like, may be present in the excipient or vehicle. Suitable reducing agents include cysteine, thioglycerol, thioreducin, glutathione, and the like. Excipients, vehicles, and auxiliary substances are generally pharmaceutical agents that do not induce an immune response in the individual receiving the composition and can be administered without excessive toxicity.

[0060] Pharmaceutically acceptable excipients include, but are not limited to, liquids such as water, saline solution, polyethylene glycol, hyaluronic acid, glycerol, thioglycerol, and ethanol. Pharmaceutically acceptable salts may also be included, for example, salts of mineral acids such as hydrochlorides, hydrobromides, phosphates, sulfates, and the like; and salts of organic acids such as acetates, propionates, malonates, benzoates, and the like.

[0061] Such compositions may be prepared, packaged, or sold in a form suitable for bolus or continuous administration. Injectable compositions may be prepared, packaged, or sold in unit-dose form, for example, in ampoules or preservative-containing multidose containers. Compositions include, but are not limited to, suspensions, solutions, emulsions in oily or aqueous vehicles, pastes, and extended-release or biodegradable implantable formulations.

[0062] Such compositions may also include one or more additional ingredients including, but not limited to, suspending, stabilizing or dispersing agents.

[0063] In one embodiment of a parenteral composition, the active ingredient is supplied in dry form (e.g., a powder or granules) for reconstitution with a suitable vehicle (e.g., sterile, pyrogen-free water) prior to parenteral administration of the reconstituted composition. Compositions may be prepared, packaged, or sold as a sterile injectable suspension or aqueous or oily solution. This suspension or solution may be formulated according to the known art and may include, in addition to the active ingredient, supplementary ingredients such as dispersing agents, wetting agents, or suspending agents as described herein. Such sterile injectable formulations may be prepared using a non-toxic diluent or solvent acceptable for parenteral use, such as water or 1,3-butanediol, for example.Other acceptable diluents and solvents include, but are not limited to, Ringer's solution, isotonic sodium chloride solution, and fixed oils such as synthetic mono- or diglycerides.

[0064] Other useful parenteral formulations include those containing the active ingredient in microcrystalline form, in a liposomal preparation, or as a component of a biodegradable polymer system. Extended-release or implantable formulations may contain pharmaceutically acceptable polymeric or hydrophobic materials such as emulsions, ion-exchange resins, sparingly soluble polymers, or sparingly soluble salts. These formulations may be suitable for administration via any appropriate route, including, for example, intradermal, subcutaneous, percutaneous, intramuscular, intra-arterial, intraperitoneal, intra-articular, intraosseous, or other suitable routes.

[0065] According to another aspect, the present invention relates to a urease-inhibiting pharmaceutical composition, comprising at least one urease inhibitor being dihydrolevoglucosenone for its use in the treatment of urinary tract infections and / or gastric ulcers related to H. pylori and / or hyperammonemia, and / or the prevention of recurrent urinary stones in said subject.

[0066] As used here, the term "subject" refers to an animal, preferably a mammal, including humans, pigs, chimpanzees, dogs, cats, cows, mice, rabbits, or rats. Even more preferably, the subject is a human being, including an adult, a child, and a human being in the prenatal stage.

[0067] As used herein, the term "treatment" refers to any act intended to improve the health status of patients, such as therapy, prevention, prophylaxis, and disease delay. In some embodiments, this term refers to the improvement or eradication of a disease or symptoms associated with a disease. In other embodiments, this term refers to the minimization of the spread or worsening of a disease resulting from the administration of one or more therapeutic agents to a subject suffering from such a disease.

[0068] In another aspect, the present invention relates to a method of treatment in a subject, comprising the administration of a therapeutically effective amount of urease inhibitor being dihydrolevoglucosenone in said subject.

[0069] By "therapeutically effective amount" is meant an amount of the pharmaceutical composition administered to a subject that is sufficient to reduce urease and / or ammonia levels in said subject, preferably to normal levels and / or to treat an infection, preferably H. pylori.

[0070] Urinary tract infections, caused by bacteria such as Proteus mirabilis, Klebsiella pneumoniae, and Staphylococcus saprophyticus, can produce urease. This enzyme catalyzes the breakdown of urea into ammonia, increasing the pH of urine and contributing to the formation of urinary stones (struvite stones). By inhibiting urease, ammonia production can be reduced, thus preventing the formation of these stones and associated complications.

[0071] Helicobacter pylori is a bacterium responsible for many cases of gastric and duodenal ulcers. This bacterium produces urease to neutralize gastric acid, allowing it to survive in the acidic environment of the stomach. By inhibiting urease, the ability of H. pylori to survive and colonize the stomach can be reduced, thus helping to eradicate the infection and heal the ulcers.

[0072] Hyperammonemia is a condition characterized by excessive levels of ammonia in the blood, often due to liver failure or inherited metabolic disorders. Ammonia is toxic to the brain and can lead to hepatic encephalopathy, a serious condition that affects brain function. Urease inhibitors can be used to reduce ammonia production in the intestine in patients with advanced liver disease or urea cycle disorders.

[0073] Patients with recurrent kidney stones, particularly those composed of struvite, may benefit from urease inhibitors. By reducing ammonia formation, these inhibitors can prevent the precipitation of struvite crystals and decrease the frequency of recurrences.

[0074] According to another aspect of the invention, the present invention relates to the use of a urease inhibitor, namely dihydrolevoglucosenone, in a subject to reduce urease and / or ammonia levels in said subject, preferably to normal levels, and / or to treat an infection, preferably H. pylori, in said subject.

[0075] According to another aspect of the invention, the present invention relates to the use of a urease inhibitor, namely dihydrolevoglucosenone, in a subject for the treatment of urinary tract infections and / or gastric ulcers related to H. pylori and / or hyperammonemia, and / or the prevention of recurrent urinary stones in said subject.

[0076] EXAMPLES

[0077] Example 1: Measurement of the CI 50 Cyrene on urease

[0078] The primary objective of this experiment was to measure the IC 50(50% inhibitory concentration) of NBPT (N-butylphosphorothioic triamide) in a solvent composed of Cyrene and water, to determine whether Cyrene denatured the inhibitor NBPT. Surprisingly, the inventors observed that all NBPT concentrations appeared to inhibit urease, even at very low concentrations, and that the water-Cyrene control also exhibited inhibitory activity. Analyses of the inhibition of Cyrene alone were therefore carried out to verify this surprising effect.

[0079] Materials and Methods:

[0080] Preparing the Solutions:

[0081] Preparation of Cyrene and NBPT solutions at various concentrations (see table 1).

[0082] Ammonia Measurement:

[0083] Ammonia levels were measured to determine urease activity.

[0084] The measurements were compared to controls: using only water, a water control with NBPT and controls with different concentrations of cyrene in water.

[0085] InhibitorSolventAdditive concentrationIC 50 compared to the solvent control IC 50 compared to the control: water, Cyrene, WaterX 270 μM, Water + NBPT 10 nM, 180 μM < 10 μM, BPTE, WaterX 80 nM, 80 nM, Water + Cyrene 1 mM, 3 nM < 1 nM, 100 μM, 20 nM < 1 nM, 50 μM, 20 nM, 10 nM, 10 μM, 60 nM, 40 nM

[0086] Results :

[0087] Inhibition by Cyrene alone:

[0088] Cyrene at 270 μM in aqueous solution showed a 50% inhibition of urease (IC50). 50 =270 μM), indicating an inhibitory effect on urease.

[0089] Inhibition by NBPT alone:

[0090] NBPT in aqueous solution showed an IC 50 of 80 nM, confirming its powerful inhibitory effect on urease.

[0091] Combined inhibition of Cyrene and NBPT:

[0092] When Cyrene and NBPT were combined, the results revealed a synergistic inhibition:

[0093] NBPT at 10 nM in water with 1 mM of Cyrene showed an IC 50 of < 1 nM, indicating a significant increase in the inhibitory effect.

[0094] Conclusion: The results show that Cyrene has a direct inhibitory effect on urease. Furthermore, in combination with NBPT, it synergistically enhances the inhibitory efficacy of NBPT.

Claims

Agricultural urease inhibitor composition comprising at least one urease inhibitor being dihydrolevoglucosenone and an additional urease inhibitor being N-(n-butyl)-thiophosphoric triamide (NBPT). Agricultural composition according to claim 1, further comprising at least one plant extract(s) and / or at least one live microorganism(s) and / or at least one microorganism extract(s) and / or at least one co-solvent or a combination thereof. Agricultural composition according to any one of claims 1 or 2, not comprising any other urease inhibitor than dihydrolevoglucosenone and N-(n-butyl)-thiophosphoric triamide (NBPT). Agricultural composition according to any one of claims 1 to 3, wherein dihydrolevoglucosenone is present in an amount of between 20% and 99%, preferably between 35% and 98%, more preferably between 45% and 95% by total weight of the composition and N-(n-butyl)-thiophosphoric triamide (NBPT) is present in an amount of between 1% and 65%, preferably between 2% and 60%, more preferably between 5% and 55% by total weight of the composition. Liquid agricultural composition of any one of claims 1 to 4, further comprising at least one nitrification inhibitor, preferably dicyandiamide. Solid or liquid fertilizer composition comprising, on the basis of the total weight of the composition: - between 90% and 99.99%, preferably between 99% and 99.99% by total weight of the composition, of one or more nitrogen fertilizer compounds; - between 0.01% and 10%, preferably between 0.01% and 1.5% by total weight of the composition, of the agricultural composition according to any one of claims 1 to 5. Use of dihydrolevoglucosenone in combination with NBPT as urease inhibitors to reduce ammonia volatilization in agricultural applications and / or the composition according to any one of claims 1 to 6 as a urease inhibitor to reduce ammonia volatilization in agricultural applications.