Lactobacillus rhamnosus strain lra06 and application thereof in intervention of uric acid synthesis

Abstract

The application relates to the technical field of Lactobacillus rhamnosus, in particular to a Lactobacillus rhamnosus and application of the Lactobacillus rhamnosus in intervention of uric acid synthesis. The strain is Lactobacillus rhamnosus LRa06 with a preservation number of CGMCC NO.33730. The strain has an inhibiting effect on metabolic enzymes of a uric acid synthesis pathway, especially on adenosine deaminase and xanthine oxidase. The strain inhibits uric acid synthesis by inhibiting the two metabolic enzymes of the uric acid synthesis pathway, reduces serum urate, and has application prospects in preparation of a uric acid-lowering preparation.

Description

Technical Field

[0001] This application relates to the technical field of Lactobacillus rhamnosus, specifically to a strain of Lactobacillus rhamnosus LRa06 and its application in intervening in uric acid synthesis. Background Technology

[0002] *Lactobacillus rhamnosus* belongs to the phylum Firmicutes, class Lactobacillus, order Lactobacilliles, family Lactobacillusaceae, and genus *Lactobacillus*. It is a Gram-positive bacillus with colonies 0.5-1 mm in diameter, round in shape, with regular edges, opaque, milky white on the front, convex in the center, smooth, moist, and viscous in texture. *Lactobacillus rhamnosus* exhibits good resistance to gastric acid and bile salts; for example, *Lactobacillus rhamnosus* R7041 can survive and exert beneficial effects in the gastrointestinal tract. It also demonstrates high tolerance to bile, acid, and salts.

[0003] Lactobacillus rhamnosus can be used as a probiotic additive and is widely used in yogurt, fermented dairy products, functional beverages, etc.

[0004] For example, Lactobacillus rhamnosus R7041 provided by CN116396910A can be used to reduce the expression of inflammatory cytokines related to skin allergies. Lactobacillus rhamnosus can also be used as a feed additive. For example, CN116836889A discloses a Lactobacillus rhamnosus JL-1 that alleviates hyperuricemia, which can reduce the serum uric acid, urea nitrogen and creatinine content in mice with high purine diet-induced hyperuricemia, and reduce the content of xanthine oxidase in liver tissue. Wei-Ting Tseng et al. (Lacticaseibacillus paracasei LT12-A Probiotic Strain That Reduces Hyperuricemia via Inhibiting XO Activity and Regulating Renal Uric Acid Transportation Protein, Fermentation 2025, 11(2), 96) studied the inhibitory activity of three lactic acid bacteria strains on xanthine oxidase (XO), among which Lactobacillus rhamnosus Lr-32 showed 4.85% XO inhibitory activity. However, Lactobacillus rhamnosus, which provides metabolic enzymes that can inhibit the uric acid synthesis pathway, still plays a very important and positive role in reducing uric acid synthesis in a broader and more direct way. Summary of the Invention

[0005] This application provides a strain of *Lactobacillus rhamnosus*, specifically strain LRa06, with accession number CGMCC NO.33730. This LRa06 strain was deposited on March 6, 2025, at the China General Microbiological Culture Collection Center, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing. This LRa06 strain exhibits inhibitory activity against metabolic enzymes involved in uric acid synthesis, particularly adenosine deaminase (ADA, EC 3.5.4.4) and xanthine oxidase (XOD, EC 1.17.3.2). By inhibiting these two metabolic enzymes in the uric acid synthesis pathway, this strain suppresses uric acid synthesis, lowers serum urate levels, and shows promise for application in the preparation of uric acid-lowering preparations.

[0006] This application also provides a composition comprising at least one or more of the following: dead, live, and inactivated cells of *Lactobacillus rhamnosus* LRa06; lysates of one or more *Lactobacillus rhamnosus* LRa06 strains; metabolites of one or more *Lactobacillus rhamnosus* LRa06 strains; analogs of one or more *Lactobacillus rhamnosus* LRa06 strains; derivatives of one or more *Lactobacillus rhamnosus* LRa06 strains; fragments of one or more *Lactobacillus rhamnosus* LRa06 strains; or combinations thereof. This composition inhibits uric acid synthesis and lowers serum urate levels by inhibiting the metabolic enzymes of these two uric acid synthesis pathways, and shows promise for application in the preparation of uric acid-lowering agents.

[0007] This application also provides a formulation containing *Lactobacillus rhamnosus* LRa06 as the active ingredient, and excipients for forming the formulation. This formulation inhibits uric acid synthesis and lowers serum urate levels by inhibiting metabolic enzymes in both uric acid synthesis pathways, and shows promise for application in the preparation of uric acid-lowering formulations.

[0008] In some embodiments, the formulation is selected from at least one of the following: uric acid synthesis pathway metabolic enzyme inhibitors, uric acid-lowering agents, and intestinal microbiota regulators.

[0009] In some embodiments, the metabolic enzyme inhibitor of the uric acid synthesis pathway is at least one of an adenosine deaminase inhibitor and a xanthine oxidase inhibitor.

[0010] This application also provides a probiotic composition containing Lactobacillus rhamnosus LRa06 as an active ingredient, and one or more prebiotics.

[0011] This application also provides a culture comprising Lactobacillus rhamnosus LRa06 as described above or a composition as described above.

[0012] This application also provides the application of a strain of *Lactobacillus rhamnosus* in the preparation of a formulation. The *Lactobacillus rhamnosus* is *Lactobacillus rhamnosus* LRa06, with accession number CGMCC NO.33730. This LRa06 strain was deposited on March 6, 2025, at the China General Microbiological Culture Collection Center, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing. The formulation is selected from at least one of the following: the formulation is selected from at least one of the following: uric acid synthesis pathway metabolic enzyme inhibitors, uric acid-lowering agents, and intestinal microbiota regulators.

[0013] According to the technical solution provided in this application, Lactobacillus rhamnosus LRa06 has been found to be able to degrade uric acid, inosine and guanosine in vitro, indicating that it has an intervention effect on the in vivo uric acid synthesis pathway.

[0014] According to the technical solution provided in this application, Lactobacillus rhamnosus LRa06 has been found to be able to intervene in hyperuricemia model mice, reduce uric acid content, counteract weight loss, liver and kidney damage, and effectively improve the health status of hyperuricemia mice.

[0015] According to the technical solution provided in this application, Lactobacillus rhamnosus LRa06 has been found to inhibit the activity of adenosine deaminase (ADA, EC 3.5.4.4) and xanthine oxidase (XOD, EC 1.17.3.2), thereby inhibiting uric acid synthesis and metabolism and reducing uric acid production.

[0016] According to the technical solution provided in this application, Lactobacillus rhamnosus LRa06 can counteract kidney inflammation and stress damage in hyperuricemia model mice and significantly reduce the levels of TNF-α, IL-1β, and IL-18.

[0017] According to the technical solution provided in this application, Lactobacillus rhamnosus LRa06 can improve intestinal flora disorder caused by hyperuricemia and promote the increase of beneficial bacteria abundance in the intestine of hyperuricemia model mice. Attached Figure Description

[0018] Figure 1 The graph shows the changes in body weight of mice in the NC, MOD, LRa06, and ADC groups during the experiment.

[0019] Figure 2 The graph shows the serum uric acid levels of mice in the NC, MOD, LRa06, and ADC groups provided for the experimental example.

[0020] Figure 3 The figure shows the statistical results of adenosine deaminase activity in the serum of mice in the NC group, MOD group, LRa06 group and ADC group provided for the experimental example.

[0021] Figure 4The graph shows the statistical results of xanthine oxidase activity in the serum of mice in the NC group, MOD group, LRa06 group and ADC group provided for the experimental example.

[0022] Figure 5 The graph shows the statistical results of liver index in mice in the NC group, MOD group, LRa06 group and ADC group provided for the experimental case.

[0023] Figure 6 The graph shows the statistical results of the kidney index of mice in the NC group, MOD group, LRa06 group and ADC group provided for the experimental case.

[0024] Figure 7 The graph shows the statistical results of serum creatinine levels in mice from the NC group, MOD group, LRa06 group, and ADC group provided for the experimental case.

[0025] Figure 8 The graph shows the statistical results of urea nitrogen content in mice from the NC group, MOD group, LRa06 group and ADC group provided for the experimental case.

[0026] Figure 9 Morphological images of kidney tissue sections from mice in the NC group (A), MOD group (B), LRa06 group (C), and ADC group (D) provided for the experimental examples.

[0027] Figure 10 The figure shows the statistical results of the content of inflammatory factors (IL-1β, IL-18, TNF-α) and malondialdehyde (MDA) in the kidney tissues of mice in the NC group, MOD group, LRa06 group and ADC group provided for the experimental case.

[0028] Figure 11 The statistical results of the α-diversity index (ACE index) of the intestinal flora of mice in the NC group, MOD group, LRa06 group and ADC group provided for the experimental cases are shown in the figure.

[0029] Figure 12 The statistical results of the α-diversity index (Shannon index) of the intestinal flora of mice in the NC group, MOD group, LRa06 group and ADC group provided for the experimental cases are shown in the figure.

[0030] Figure 13 The NMDS diagrams for the β-diversity analysis of the gut microbiota of mice in the NC, MOD, LRa06, and ADC groups provided for the experimental examples.

[0031] Figure 14 The graph shows the changes in gut microbiota composition at the phylum level in mice from the NC, MOD, LRa06, and ADC groups, which are provided for experimental examples.

[0032] Figure 15The graph shows the changes in gut microbiota composition at the genus level in mice from the NC, MOD, LRa06, and ADC groups provided for the experimental cases. Detailed Implementation

[0033] To make the objectives, technical solutions, and advantages of this application clearer, the following detailed description is provided in conjunction with embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the scope of this application. Reagents not specifically described in detail herein are all conventional reagents and are commercially available; methods not specifically described in detail are all conventional experimental methods and can be learned from the prior art.

[0034] As used in this article, “hyperuricemia” is characterized by abnormally high concentrations of urate or uric acid in the blood. Hyperuricemia is known to be associated with many diseases and conditions, including cardiovascular disease, metabolic syndrome, non-alcoholic fatty liver disease (e.g., non-alcoholic steatohepatitis (NASH)), chronic kidney disease, gout, insulin resistance, hypertension, dyslipidemia, renal insufficiency, obesity, prediabetes, and diabetes, particularly type II diabetes.

[0035] Therefore, this application still provides a strain of *Lactobacillus rhamnosus*, specifically *Lactobacillus rhamnosus* LRa06 with accession number CGMCC NO.33730. This LRa06 strain was deposited on March 6, 2025, at the China General Microbiological Culture Collection Center (CGMCC), accession number CGMCC NO.33730, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing. This LRa06 strain exhibits inhibitory activity against metabolic enzymes involved in uric acid synthesis, particularly adenosine deaminase (ADA, EC 3.5.4.4) and xanthine oxidase (XOD, EC 1.17.3.2). By inhibiting these two metabolic enzymes in the uric acid synthesis pathway, this strain suppresses uric acid synthesis, lowers serum urate levels, and shows promise for application in the preparation of uric acid-lowering preparations.

[0036] In some embodiments, the Lactobacillus rhamnosus strain according to this application may be an isolated bacterial strain.

[0037] This application discloses Lactobacillus rhamnosus LRa06, including pure cultures of Lactobacillus rhamnosus deposited at the China General Microbiological Culture Collection Center or their analogues, fragments, lysates, or combinations thereof.

[0038] This application provides a composition comprising at least one or more of the following: dead, live, and inactivated cells of *Lactobacillus rhamnosus* LRa06; lysates of one or more *Lactobacillus rhamnosus* LRa06 strains; metabolites of one or more *Lactobacillus rhamnosus* LRa06; analogs of one or more *Lactobacillus rhamnosus* LRa06; derivatives of one or more *Lactobacillus rhamnosus* LRa06; fragments of one or more *Lactobacillus rhamnosus* LRa06; or combinations thereof. This composition can inhibit the enzymatic activity of adenosine deaminase (ADA, EC 3.5.4.4) and xanthine oxidase (XOD, EC 1.17.3.2), thereby inhibiting uric acid synthesis.

[0039] In some embodiments of the provided compositions, the concentration of at least one or more of the dead bacterial cells, live bacterial cells, and inactivated bacterial cells is 10. 3 Up to 10 17 Within the range of colony-forming units per gram or per milliliter (CFU / g or CFU / mL), for example, in 10 5 -10 17 Within the range of CFU / g or CFU / mL, for example, in the range of 10 6 -10 17 Within the range of CFU / g or CFU / mL, for example, in the range of 10 7 -10 17 Within the range of CFU / g or CFU / mL, for example, in the range of 10 8 -10 17 Within the range of CFU / g or CFU / mL, for example, in the range of 10 9 -10 17 Within the range of CFU / g or CFU / mL, for example, in the range of 10 10 -10 17 Within the range of CFU / g or CFU / mL, for example, in the range of 10 11 -10 16 Within the range of CFU / g or CFU / mL, for example, in the range of 10 12 -10 16 Within the range of CFU / g or CFU / mL, for example, in the range of 10 13 -10 16 Within the range of CFU / g or CFU / mL, for example, in the range of 10 7 -10 16 Within the range of CFU / g or CFU / mL, for example, in the range of 10 8 -10 15 Within the range of CFU / g or CFU / mL, for example, in the range of 10 9 -10 15 Within the range of CFU / g or CFU / mL, for example, in the range of 1010 -10 15 Within the range of CFU / g or CFU / mL, for example, in the range of 10 11 -10 15 Within the range of CFU / g or CFU / mL, for example, in the range of 10 12 -10 15 Within the range of CFU / g or CFU / mL.

[0040] This application also provides a formulation containing Lactobacillus rhamnosus LRa06 as an active ingredient, and excipients for forming the formulation.

[0041] In some embodiments of the formulation provided, Lactobacillus rhamnosus LRa06 is present as dead or live cells.

[0042] In some of the formulations provided, *Lactobacillus rhamnosus* LRa06 is used as an active ingredient in a mixture of at least one or more of dead, live, and inactivated bacterial cells, or lysates of one or more *Lactobacillus rhamnosus* LRa06 strains, or metabolites of one or more *Lactobacillus rhamnosus* LRa06, or analogs of one or more *Lactobacillus rhamnosus* LRa06, or derivatives of one or more *Lactobacillus rhamnosus* LRa06, or fragments of one or more *Lactobacillus rhamnosus* LRa06, or combinations thereof. The concentration of the active ingredient is from 0.0001% (w / w) to 99% (w / w).

[0043] In the context of this application, *Lactobacillus rhamnosus* LRa06 as defined herein can be provided in the composition according to this application in the form of dead cells, live cells, and inactivated cells, or a mixture of at least one or more of these. The term "live cell" refers to a live *Lactobacillus reuteri* bacterium with intact cell structure, capable of normal metabolism and reproduction, for example, cultured in a medium (such as MRS medium), centrifuged, washed, and then preserving the viability, typically in lyophilized form (such as lyophilized bacterial powder). The term "dead cell" refers to cells that have died naturally or have lost activity through physical / chemical treatment (such as high temperature, ultraviolet light), and whose cell structure may be intact or partially destroyed. The term "inactivated cell" specifically refers to cells that have been killed by controlled methods (such as heat inactivation, formaldehyde treatment, autoclaving) but retain their cell surface structures (such as cell walls, capsules). Inactivated cells emphasize "structural preservation," while dead cells may suffer structural destruction due to the treatment method.

[0044] In this document, the terms "lysate" and "extract" specifically refer to a solution or suspension of microbial cells in an aqueous medium according to this application, and contain, for example, macromolecules (e.g., DNA, RNA, proteins, peptides, lipids, carbohydrates, etc.) and cell debris. The lysate preferably includes the cell wall or cell wall components, including binding receptors. Methods for producing the lysate are well known to those skilled in the art, including, for example, using a French press or enzymatic lysis, or a ball mill with glass or iron beads. Cells can be lysed by enzymatic, physical, or chemical methods. Examples of enzymatic cell lysis include individual enzymes and mixtures of enzymes, such as proteases, proteinase K, lipases, and glycosidases; chemical lysis can be induced by ionophores, detergents (e.g., SDS), acids, or bases; physical methods can also be implemented using, for example, high pressure, osmotic pressure, temperature variations, or alternating hot and cold treatments during a French press. Furthermore, chemical, physical, and enzymatic methods can, of course, be combined.

[0045] In this document, the term "metabolite" refers to various substances produced by the microbial cells according to this application during their growth, reproduction and metabolism. These metabolites are closely related to their physiological functions, probiotic properties and applications, such as organic acids, bacteriocins, extracellular polysaccharides, amino acids and their derivatives, vitamins and coenzymes, secondary metabolites and signaling molecules.

[0046] In this document, the term "derivative" refers to a substance with a specific structure or function generated from microbial cells as raw materials through chemical modification, biotransformation or other treatments. It may be a modified product of cell components or a metabolic intermediate, such as a derivative of peptidoglycan, a component of bacterial cell walls.

[0047] In this document, the term "analyte" refers to a substance whose structure is similar to, but not identical to, a component in the dead cells of a microorganism according to this application, and which may be artificially synthesized or obtained from other sources and can mimic the function of the original component. For example, based on the structure of the active component in the cell, a compound with a similar structure is designed and synthesized (such as a peptide that mimics a bacterial surface antigen), or a substance extracted from other organisms that has a structure similar to that of a bacterial component (such as a polysaccharide analog from a plant).

[0048] In this document, the term "fragment" refers to a partial structural or component fragment of a microbial cell according to this application, which is usually obtained by physical cutting, enzymatic digestion or genetic engineering, such as cell wall fragments, protein fragments, nucleic acid fragments, etc.

[0049] Some embodiments provide formulations containing pharmaceutically, health-promoting, or food-grade carriers or excipients. In some embodiments, the formulation may be provided in solid, liquid, viscous, emulsion, or as a dry form.

[0050] Some of the formulations provided in these embodiments may preferably be formulated as pastes, soft gelatin capsules, hard gelatin capsules, powders, talc, granules, beads, tablets, effervescent tablets, lozenges, chewable tablets, sublingual tablets, oils, liquids, solutions, tinctures, emulsions, fruit juices, concentrates, syrups, sprays, mists, drinking ampoules, gels, gels, tablets, coated pills, or as food or feed products or beverages.

[0051] In some embodiments, the formulation is in the form of pills, powders, capsules, tablets, granules, film-coated tablets, creams, ointments, gels, lotions, foams, suppositories, sachets, or sugar-coated pills.

[0052] This application also provides a culture comprising Lactobacillus rhamnosus LRa06 as described above or a composition as described above.

[0053] In some embodiments, the culture is a bacterial suspension of Lactobacillus rhamnosus LRa06.

[0054] In some embodiments, the culture also contains a nutrient-providing component (e.g., a solid or liquid culture medium, or a feeder cell or bacteria).

[0055] In some embodiments, the nutritional components are selected from proteins, carbohydrates, fats, probiotics, enzymes, vitamins, immunomodulators, milk substitutes, minerals, amino acids, or any combination thereof.

[0056] In some embodiments, the culture also comprises a cell-free culture filtrate of Lactobacillus rhamnosus LRa06.

[0057] In some embodiments, the culture also contains a derivative of Lactobacillus rhamnosus LRa06.

[0058] In some embodiments, the derivative is selected from metabolites, enzymes, cellular structural components (e.g., cell walls or components thereof), extracellular polysaccharides, bacteriocins, compounds containing immunogenic components, or any combination thereof.

[0059] According to another aspect of this application, a prebiotic composition comprising Lactobacillus rhamnosus LRa06 and at least one or more prebiotics is provided.

[0060] In the context of this application, a "prebiotic" is a non-digestible food ingredient that promotes the growth of specific microorganisms. A "synbiotic" is a composition comprising at least one probiotic and at least one prebiotic. Such compositions are understood to promote the growth of beneficial bacteria (e.g., probiotics). Therefore, a potent synbiotic is based on a combination of specific strains of probiotics and carefully selected prebiotics. These can provide significant health benefits to mammals.

[0061] Prebiotics are chemical products that induce the growth and / or activity of symbiotic microorganisms (such as bacteria and fungi) that contribute to the health of the host. Prebiotics are indigestible carbohydrates that pass through the upper gastrointestinal tract undigested and stimulate the growth and / or activity of beneficial bacteria that colonize the gut or skin microbiota.

[0062] Some oligosaccharides used as prebiotics are fructooligosaccharides (FOS), xylooligosaccharides (XOS), polydextrose, pectin, galactooligosaccharides (GOS), or human milk oligosaccharides (HMOs). In addition, disaccharides such as lactulose or some monosaccharides such as lactose or tagatose can also be used as prebiotics.

[0063] In one embodiment of this application, at least one prebiotic compound may be included in the composition of this application. In a very broad concept, prebiotics are all compounds that can be metabolized by probiotics.

[0064] Preferably, prebiotics are indigestible or poorly digestible by mammals. Therefore, after being ingested by mammals, indigestible prebiotics can pass through the small intestine and enter the large intestine to stimulate the growth of probiotics in that compartment. Thus, prebiotics can serve as a food source for probiotics. It is believed that prebiotics (many of which are poorly digestible carbohydrates) promote the growth of probiotics. Prebiotics are naturally found in, for example, cabbage, onions, whole grains, bananas, garlic, honey, leeks, artichokes, fortified foods and beverages, and dietary supplements. Prebiotics are well known in the art, and there are no particular limitations on prebiotics themselves when used in this application.

[0065] In one embodiment, at least one prebiotic product in the composition is selected from the following compounds and compositions: indigestible carbohydrates, β-glucan, mannooligosaccharides, inulin, fructooligosaccharides, human milk oligosaccharides (HMO), galactooligosaccharides (GOS), lactulose, lactulose oligosaccharides, galactotriose, fructooligosaccharides (FOS), cellobiose, cellodextrin, cyclodextrin, maltitol, lactitol, glycosilsucrose, betaine, vitamin E, or variants thereof (wherein the variants are selected from α, β, γ, δ tocopherols, tocotrienols, and tocomonenophenols). Optionally, mannooligosaccharides and / or inulin may be preferred. HMOs may include lact-N-tetrasaccharide, lact-N-fucopentose, lact-N-triose, 3'-sialyllactose, lact-N-neofucopentose, sialic acid, L-fucose, 2-fucosyllactose, 6'-sialyllactose, lact-N-neotetrasaccharide, and 3-fucosyllactose.

[0066] In the context of this application, acceptable additives and / or excipients for pharmaceutical or food applications include adjuvant substances known to those skilled in the art for preparing solid, semi-solid, or liquid forms, such as diluents, solvents (including water, glycerol, and ethanol), solubilizers, acidifiers, thickeners, sweeteners, flavor enhancers, colorants, lubricants, surfactants, preservatives, pH-stabilizing buffers, and mixtures thereof.

[0067] In various embodiments of this application, the composition of this application containing Lactobacillus rhamnosus LRa06 can be a pharmaceutical composition (or live biological therapeutic product), a medical device composition, a dietary supplement, a food (or a novel food or medical food), a composition for use as a dietary supplement or food, a cosmetic composition, or a composition for use in a food for special medical purposes (FSMP).

[0068] For example, the inventors of this application found through testing that the Lactobacillus rhamnosus LRa06 provided in this application can degrade uric acid, inosine and guanosine in vitro, indicating that it has an intervention effect on the in vivo uric acid synthesis pathway.

[0069] For example, the inventors of this application found through testing that the Lactobacillus rhamnosus LRa06 provided in this application was able to intervene in hyperuricemia model mice, reduce uric acid content, counteract their weight loss, liver and kidney damage, and effectively improve the health status of hyperuricemia mice.

[0070] For example, the inventors of this application discovered through testing that the Lactobacillus rhamnosus LRa06 provided in this application can inhibit the activity of adenosine deaminase (ADA, EC 3.5.4.4) and xanthine oxidase (XOD, EC 1.17.3.2), thereby inhibiting uric acid synthesis and metabolism and reducing uric acid production.

[0071] For example, the inventors of this application found through testing that the Lactobacillus rhamnosus LRa06 provided in this application can combat kidney inflammation and stress damage in hyperuricemia model mice and significantly reduce the levels of TNF-α, IL-1β, and IL-18.

[0072] For example, the inventors of this application found through testing that the Lactobacillus rhamnosus LRa06 provided in this application can improve the intestinal flora disorder caused by hyperuricemia and promote the increase of beneficial bacteria abundance in the intestines of hyperuricemia model mice.

[0073] To aid in understanding the screening and identification of Lactobacillus rhamnosus LRa06, hyperuricemia, and intestinal flora imbalance caused by hyperuricemia, detailed experimental descriptions will be provided below, but these descriptions do not constitute a limitation on the implementation methods of this application.

[0074] I. Isolation and Identification of Strains

[0075] 1. Isolation of strains

[0076] Naturally fermented dairy products were serially diluted with 0.85% physiological saline under aseptic conditions. The diluted products were then spread onto LBS agar plates and incubated at 37°C for 48-72 hours. Colony morphology was observed visually. Suspected single colonies were picked for microscopic examination, followed by preliminary screening and purification. The purified strains were incubated in MRS broth at 37°C for 8-12 hours. After centrifugation to remove the supernatant, the cultures were resuspended in sterile 30% glycerol aqueous solution and stored at -80°C.

[0077] 2. Identification of strains

[0078] The selected target strain was cultured in liquid medium, the bacterial cells were collected, genomic DNA was extracted, and PCR amplification was performed on the genomic DNA. The content and purity of the PCR amplification products were detected, and those that passed the test were sent to Wuhan Jinkairui Biotechnology Co., Ltd. for sequencing. Based on the sequencing results, homology comparison was performed using the BLAST tool in the NCBI database, and the obtained strain was identified as *Lactobacillus rhamnosus*. This strain was named *Lactobacillus rhamnosus* LRa06 and deposited for preservation. Its preservation information is as follows:

[0079] Accession number: CGMCC NO.33730, Accession date: March 6, 2025

[0080] Classification and nomenclature: Lacticaseibacillus rhamnosus

[0081] Preservation Institution: China General Microbiological Culture Collection Center, China Microbiological Culture Collection Committee

[0082] Address of the depository: No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing.

[0083] II. In vitro degradation capacity of Lactobacillus rhamnosus LRa06 for uric acid and nucleosides

[0084] The preserved *Lactobacillus rhamnosus* strain LRa06 was passaged three times and inoculated into liquid MRS medium at a 3% (v / v) inoculum. All bacterial cultures were collected into 50 mL centrifuge tubes, centrifuged at 8000 rpm for 5 min, the supernatant was discarded, and the bacterial pellet was washed with 5 mL PBS, repeated twice. The strain was resuspended in 5 mL of 0.6 mM inosine and 0.6 mM guanosine solutions, respectively, and anaerobically cultured at 37 °C for 6 h. 900 μL of culture medium was transferred to 100 μL of perchloric acid (0.1 mol / L), vortexed, and centrifuged at 4000 x g for 10 min. The supernatant was collected and filtered through a 0.22 μm filter into a liquid chromatography vial. The mobile phase was 20 mmol / L potassium dihydrogen phosphate:methanol 96:4, UV wavelength 254 nm, column temperature 25 °C, isocratic elution for 20 min. Results were quantified using the external standard method.

[0085] The results showed that the inosine degradation rate of Lactobacillus rhamnosus LRa06 was 43.14%, and the guanosine degradation rate was 78.14%.

[0086] III. In vivo intervention effect of Lactobacillus rhamnosus LRa06 on hyperuricemia

[0087] 1. Sources of biological materials

[0088] Six-week-old male BALB / c mice were purchased from Spifort (Beijing) Biotechnology Co., Ltd. and housed in the animal facility of Hubei Provincial Center for Disease Control and Prevention. Animal experiment ethics number: Safety Evaluation Center Animal (Fu) No. 202410369.

[0089] 2. Establishment of a hyperuricemia model and grouped intervention experiment

[0090] Male BALB / c mice were housed in an environment with a room temperature of 25°C ± 2°C, humidity of 50% ± 5%, and a 12-hour light-dark cycle. After a 7-day acclimatization period, the mice were randomly divided into four groups (n=10 per group): NC group, MOD group, LRA06 group, and ADC group. Mice in the MOD, LRA06, and ADC groups underwent hyperuricemia modeling. The model was established by suspending potassium oxonate in 0.5% sodium carboxymethyl cellulose solution and adenine in physiological saline. Mice were administered 0.2 mL of adenine (50 mg / kg) by gavage and 0.2 mL of potassium oxonate solution (250 mg / kg) by intraperitoneal injection daily. Mice in the NC group were administered the same volume of physiological saline by gavage and the same volume of 0.5% sodium carboxymethyl cellulose solution by intraperitoneal injection. Mice in the LRA06 group were treated with adenine and potassium oxonate for 4 hours, followed by administration of 0.2 mL of potassium oxonate solution containing 1×10⁻⁶ mg / kg of potassium oxonate.10 Mice in the ADC group were given physiological saline containing CFU / mL Lactobacillus rhamnosus LRa06. Mice in the NC group were given 0.2 mL of physiological saline containing 5 mg / kg allopurinol. Mice in the NC group and MOD group were given the same volume of physiological saline.

[0091] 3. Body weight, uric acid, blood urea nitrogen, and creatinine levels in each group of mice.

[0092] Mice were weighed every 3 days, and orbital blood was collected every 7 days to measure uric acid levels. The experiment lasted for 14 days. After 14 days, the mice were weighed and euthanized.

[0093] like Figure 1 As shown, the body weight of mice in the MOD and ADC groups gradually decreased, showing a significant difference compared to the NC group. After intervention with *Lactobacillus rhamnosus* LRa06, the body weight of mice gradually recovered, significantly increasing compared to the MOD group. This indicates that *Lactobacillus rhamnosus* LRa06 can effectively improve the health status of mice with hyperuricemia.

[0094] like Figure 2 As shown, on day 7 of the experiment, the serum uric acid level in the MOD group mice was significantly higher than that in the NC group, while LRa06 intervention significantly reduced the serum uric acid level. On day 14 of the experiment, the serum uric acid level in the MOD group mice was still significantly different from that in the NC group, and LRa06 was still able to reduce the serum uric acid level. However, the ability of LRa06 to reduce serum uric acid was not as strong as that of allopurinol. These results indicate that *Lactobacillus rhamnosus* LRa06 can effectively reduce the serum uric acid level in hyperuricemic mice.

[0095] 4. Inhibitory effect of Lactobacillus rhamnosus LRa06 on metabolic enzymes

[0096] After the experiment, blood was collected from each group of mice to detect the activity of adenosine deaminase (ADA) and xanthine oxidase (XOD).

[0097] ADA (EC 3.5.4.4) and XOD (EC 1.17.3.2) are important metabolic enzymes in the uric acid production pathway. ADA deamination converts adenosine to inosine. Inosine is further converted to hypoxanthine. XOD catalyzes the conversion of hypoxanthine to xanthine, which is then further catalyzed to uric acid. When the activities of ADA and XOD increase, uric acid production increases; when the activities of ADA and XOD decrease, uric acid production decreases.

[0098] like Figure 3 and Figure 4As shown, the activities of ADA and XOD in the MOD group mice were significantly higher than those in the NC group. However, after LP305 intervention, the activities of ADA and XOD decreased significantly compared with the MOD group, indicating that Lactobacillus rhamnosus LRa06 can inhibit the activities of ADA and XOD to reduce the serum uric acid level in hyperuricemic mice.

[0099] 5. Lactobacillus rhamnosus LRa06 improves kidney damage caused by high uric acid.

[0100] After the experiment, liver and kidney tissues were collected from mice in each group, and liver and kidney indices were measured. Liver index (%) = (liver weight / mouse body weight) × 100%. Kidney index (%) = (kidney weight / mouse body weight) × 100%.

[0101] like Figure 5 and Figure 6 As shown, the liver and kidney indices in the MOD and ADC groups were significantly higher than those in the NC group, indicating lesions such as liver and kidney enlargement. The kidney index in the LRa06 group was significantly lower, with no significant difference from the NC group.

[0102] After the experiment, blood was collected from each group of mice to test the urea nitrogen and creatinine levels.

[0103] like Figure 7 and Figure 8 As shown, with increasing experimental time, the creatinine and blood urea nitrogen levels in the MOD and ADC groups were significantly higher than those in the NC group. However, after intervention with *Lactobacillus rhamnosus* LRa06, the creatinine and blood urea nitrogen levels in the LRa06 group were significantly lower, showing no significant difference from the NC group. Creatinine and blood urea nitrogen are indicators for evaluating kidney function. Elevated levels of creatinine and blood urea nitrogen in the blood indicate renal insufficiency, potentially suggesting kidney failure, nephritis, or acute tubular necrosis. Figures 5-8 The results showed that Lactobacillus rhamnosus LRa06 could reduce kidney damage in mice with hyperuricemia.

[0104] 8. Lactobacillus rhamnosus LRa06 improves kidney inflammation and oxidative stress caused by hyperuricemia.

[0105] After the experiment, kidney tissues from mice in each group were collected, HE sections were prepared, and photographs were taken under an optical microscope.

[0106] like Figure 9 In the NC group, the renal tubules and glomeruli of mice showed clear morphology and no obvious lesions. In the MOD group, the glomeruli of mice were atrophied, the renal tubules were enlarged, and the renal interstitium was filled with inflammatory cells. The renal pathological condition of mice in the LRa06 group was significantly improved compared to the MOD group, with normal glomerular and tubular morphology and reduced inflammatory cells in the renal interstitium. However, the renal pathological condition of the ADC group showed no significant improvement compared to the MOD group.

[0107] After the experiment, kidney tissues from mice in each group were collected, kidney tissue homogenates were prepared, and the contents of IL-1β, IL-18, TNF-α and MDA were detected.

[0108] like Figure 10 As shown, compared with the NC group, the MOD group mice had significantly higher levels of TNF-α, IL-1β, IL-18, and MDA in their kidneys, indicating that hyperuricemia leads to inflammatory responses and stress damage in the kidneys of hyperuricemic mice. However, intervention with *Lactobacillus rhamnosus* LRa06 significantly reduced the levels of TNF-α, IL-1β, and IL-18, demonstrating that LRa06 can alleviate kidney inflammation and stress damage caused by hyperuricemia.

[0109] 9. Lactobacillus rhamnosus LRa06 improves intestinal flora imbalance caused by hyperuricemia.

[0110] After the experiment, cecal tissues of mice in each group were collected, and 16S rRNA amplicon sequencing was performed on the contents of the cecal tissues. The abundance of the mouse gut microbiota at the α-diversity, β-diversity, phylum level and genus level was analyzed.

[0111] like Figure 11 and Figure 12 The α-diversity analysis results showed that, compared with the NC group, the ACE and Shannon indices of the intestines of MOD group mice were significantly decreased, indicating that hyperuricemia leads to a reduction in the richness and diversity of gut microbiota. After intervention with *Lactobacillus rhamnosus* LRa06, the ACE and Shannon indices of hyperuricemic mice significantly increased. This indicates that *Lactobacillus rhamnosus* LRa06 can improve the richness and diversity of the gut microbiota in hyperuricemic mice.

[0112] like Figure 13 The β-diversity analysis results showed that the MOD group and the NC group were far apart on the NMDS map, and the Stress value was less than 0.2, indicating that the intestinal community composition of the MOD and NC groups was different. Meanwhile, the LRa06 group had overlap with the NC group but no overlap with the MOD group, indicating that the intestinal microbiota composition of the LRa06 group was similar to that of the NC group. Combining the α and β-diversity analysis results, it is suggested that LRa06 can alleviate the damage to the intestinal microecology caused by hyperuricemia in mice.

[0113] like Figure 14The phylum-level intestinal community abundance analysis results for each mouse group showed that, compared with the NC group, the abundance of Firmicutes and Verrucous microbes in the MOD group decreased, while the abundance of Bacteroidetes increased. Intervention with *Lactobacillus rhamnosus* LRa06 significantly restored the abundance of Firmicutes and Verrucous microbes and decreased the abundance of Bacteroidetes. Furthermore, *Lactobacillus rhamnosus* LRa06 also promoted an increase in the abundance of Actinobacteria.

[0114] like Figure 15 The results of the genus-level intestinal community abundance analysis for each group of mice showed that the relative abundance of *Eurotium*, *Bacteroides*, *Parabacteroides*, and *Helicobacter* was significantly higher in the MOD group than in the NC group, while the relative abundance of *Ackermania* and *Lactobacillus* was lower. Related studies have reported that the abundance of *Eurotium*, *Bacteroides*, *Parabacteroides*, and *Helicobacter* significantly increases during inflammation. This indicates that high uric acid leads to an increase in the abundance of harmful bacteria in the gut. In the LRa06 group, the relative abundance of *Eurotium*, *Bacteroides*, *Parabacteroides*, and *Helicobacter* was significantly lower than in the MOD group, while the relative abundance of *Ackermania*, *Lactobacillus*, *Lactobacillus*, and *Bifidobacterium* was significantly higher. Related studies have shown that the abundance of *Ackermania*, *Lactobacillus*, *Lactobacillus*, and *Bifidobacterium* is negatively correlated with hyperuricemia. These results all indicate that the Lactobacillus rhamnosus LRa06 strain provided in this application can promote the increase of beneficial bacteria abundance and reduce the abundance of harmful bacteria in the intestine.

[0115] The above description is merely a preferred embodiment of this application, but the scope of protection of this application is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the scope of the technology disclosed in this application should be included within the scope of protection of this application.

Claims

1. A strain of Lactobacillus rhamnosus, characterized in that, It is Lacticaseibacillus rhamnosus LRa06 with accession number CGMCC NO. 33730.

2. A uric acid-lowering preparation comprising, as an active ingredient, *Lactobacillus rhamnosus* LRa06 as described in claim 1, and excipients for forming the preparation.

3. A metabolic enzyme inhibitor of the uric acid synthesis pathway, comprising Lactobacillus rhamnosus LRa06 as an active ingredient as described in claim 1, and excipients for forming the inhibitor.

4. The inhibitor according to claim 3, characterized in that, The metabolic enzyme inhibitors of the uric acid synthesis pathway are at least one of adenosine deaminase inhibitors and xanthine oxidase inhibitors.

5. An intestinal microbiota regulating preparation, comprising Lactobacillus rhamnosus LRa06 as an active ingredient as described in claim 1, and excipients for forming the preparation.

6. A probiotic composition, characterized in that, It contains Lactobacillus rhamnosus LRa06 as an active ingredient as described in claim 1, and one or more prebiotics.

7. The use of Lacticaseibacillus rhamnosus as described in claim 1 in the preparation of formulations.

8. The application according to claim 7, characterized in that, The formulation is selected from at least one of the following: uric acid synthesis pathway metabolic enzyme inhibitors, uric acid-lowering agents, and intestinal microbiota regulators.

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