Glutamicobacterium nicotianae, uric acid reducing microbial agent and application thereof

Abstract

The invention belongs to the technical field of microbial uric acid reduction, and particularly relates to bacillus nicotianae, a uric acid reducing microbial agent and application of the bacillus nicotianae and the uric acid reducing microbial agent. According to the present invention, the bacillus nicotianae is an FL1-1 strain, the preservation number of the bacillus nicotianae is CGMCC (China General Microbiological Culture Collection Center) No.37236, and the preservation date is December 29, 2025. The FL1-1 strain can grow in a bile salt environment with the pH being greater than or equal to 3.0 and the bile salt concentration being less than or equal to 0.3%. The uric acid reduction rate of FL1-1 in 72 hours reaches 95.96%, and the uric acid degradation capacity of FL1-1 can be improved by adding a proper amount of Fe < 2 + > into the culture medium. The FL1-1 has good gastrointestinal tract environment tolerance, remarkable uric acid reducing capacity, antibacterial activity and antibiotic resistance spectrum.

Description

Technical Field

[0001] This invention belongs to the field of microbial uric acid lowering technology, specifically relating to a tobacco glutamate bacterium, a uric acid lowering bacterial agent, and their applications. Background Technology

[0002] Hyperuricemia in poultry is a common metabolic disease caused by purine metabolism disorder, characterized by abnormally high levels of uric acid in the blood. This condition is particularly prominent in intensive poultry farming, often leading to visceral or articular gout, resulting in stunted growth, decreased feed conversion ratio, organ damage, and even death, causing significant economic losses to the poultry industry. Poultry physiologically lack endogenous uricase, making it unable to effectively break down uric acid into water-soluble allantoin. Therefore, the end product of purine metabolism accumulates as uric acid, which is the fundamental reason for the prevalence of hyperuricemia in poultry. Currently, production mainly addresses this by adjusting to low-protein diets and using chemical drugs such as allopurinol. However, the former may affect production performance, while the latter carries risks of drug residues, potential toxic side effects, and bacterial resistance.

[0003] Utilizing microbial uricase to degrade uric acid offers a unique advantage over directly adding enzyme preparations. Probiotics can be orally administered directly to the intestines, continuously generating enzyme activity locally. They may also possess multiple beneficial functions, such as regulating gut microbiota, inhibiting pathogens, and enhancing immunity, achieving "multiple benefits from a single strain." However, existing uric acid-lowering microorganisms struggle to survive in the pH environment of gastric acid and bile salts, and some are pathogenic or hemolytic, making them unsuitable as feed additives. Summary of the Invention

[0004] To address the aforementioned technical problems, this invention provides *Bacillus glutamate*, a uric acid-lowering bacterium, and their applications.

[0005] To facilitate understanding of this invention, the materials used in this invention and their abbreviations are listed below: Gentamicin (GEN). Ceftriaxone (CTR). Ciprofloxacin (CIP). Clindamycin (CC). Chloramphenicol (C). Cephalexin (CN). Minocycline (MI). Levofloxacin (LVX). Erythromycin (E). Vancomycin (VAN). Piperacillin (PIP). Amikacin (AMK). Oxacillin (OX). Tetracycline (TET). Imipenem (IPM). Sulfamethoxazole / Trimethoprim (SXT).

[0006] The first objective of this invention is to provide a Tobacco Glutamate Bacterium (TGA) Glutamicibacter nicotianaeThe *Tobacco glutamate* strain mentioned is FL1-1, with accession number CGMCC No. 37236 and accession date of December 29, 2025. The FL1-1 strain can grow in bile salt environments with a pH greater than or equal to 3.0 and a bile salt concentration less than or equal to 0.3%.

[0007] The second objective of this invention is to provide a uric acid-lowering bacterial agent based on *Bacillus glutamicum*, comprising a *Bacillus glutamicum* bacterial agent and excipients. The *Bacillus glutamicum* bacterial agent is a bacterial suspension or freeze-dried *Bacillus glutamicum* bacterial powder, and the excipients are Fe-containing... 2+ Salts or solutions thereof.

[0008] Preferably, the viable count of the *Bacillus glutamate* bacterial suspension is greater than or equal to 10. 8 CFU / mL.

[0009] Preferably, the viable count of the *Bacillus glutamate* bacterial solution is 10. 8 CFU / mL ~10 9 CFU / mL.

[0010] Preferably, the viable bacteria count in the uric acid-lowering bacterial agent and the Fe content in the excipients are... 2+ The ratio is 5×10 8 CFU~5×10 9 CFU: 3.66mg~10.96mg.

[0011] Preferably, the viable bacteria count in the uric acid-lowering bacterial agent and the Fe content in the excipients are... 2+ The ratio is 5×10 9 CFU: 5.48mg.

[0012] Preferably, the Fe-containing 2+ The salt is ferrous sulfate.

[0013] Preferably, the concentration of the ferrous sulfate is 0.010 g / L to 0.03 g / L.

[0014] Preferably, the concentration of the ferrous sulfate is 0.015 g / L.

[0015] The third objective of this invention is to provide an application of a uric acid-lowering bacterial agent based on *Bacillus glutamate* in lowering uric acid levels.

[0016] Bioinformatics Preservation Instructions The strains involved in this invention are as follows: *Bacillus nicotine* strain FL1-1 was deposited at the China General Microbiological Culture Collection Center on December 29, 2025. The recommended classification name is... Glutamicibacter nicotianaeThe collection number is CGMCC No. 37236, and the collection address is No. 1, Beichen West Road, Chaoyang District, Beijing.

[0017] Compared with the prior art, the present invention has the following beneficial effects: 1. The *Tobacco glutamate* strain of this invention is FL1-1, with accession number CGMCC No. 37236 and accession date of December 29, 2025. The optimal growth temperature of strain FL1-1 is 37°C, and the optimal pH value is 6. Strain FL1-1 can grow in bile salt environments with a pH greater than or equal to 3.0 and a bile salt concentration less than or equal to 0.3%. For example, strain FL1-1 can grow in environments with pH 3 and pH 4, and at bile salt concentrations of 3 g / L, 2 g / L, and 1 g / L. In uric acid culture medium, strain FL1-1 exhibits a uric acid degradation rate of 55.35% after 24 hours, 94.75% after 48 hours, and 95.96% after 72 hours. Therefore, strain FL1-1 has a strong uric acid degradation ability. Furthermore, strain FL1-1 exhibits antibacterial activity against Staphylococcus aureus and Salmonella, and its DPPH scavenging ability in the fermentation broth is stronger than that of vitamin C, demonstrating antioxidant capacity. Strain FL1-1 did not show hemolysis. On the other hand, strain FL1-1 is sensitive to antibiotics GEN, CTR, CIP, CC, C, CN, MI, LVX, E, VAN, PIP, and AMK, but insensitive to antibiotics OX, TET, IPM, and SXT, which facilitates the use of antibiotics to control the growth and development of strain FL1-1.

[0018] In summary, the FL1-1 strain of the present invention, while possessing high uric acid degradation capacity, also exhibits acid resistance, high bile salt tolerance, antibacterial properties, antioxidant activity, antibiotic sensitivity, and non-hemolytic activity. Therefore, the FL1-1 strain can provide a new material for preparing formulations to prevent and alleviate hyperuricemia in poultry.

[0019] 2. The uric acid-lowering bacterial agent based on *Bacillus nicotineus* of this invention is made from *Bacillus nicotineus* and excipients. The FL1-1 strain of this invention exhibits a uric acid degradation rate of 86.78% after 24 hours in a culture environment of 0.015 g / L FeSO4, significantly higher than the 55.35% without FeSO4, representing a 57% increase in uric acid degradation rate. The FL1-1 strain in this uric acid-lowering bacterial agent can efficiently degrade uric acid and, due to its acid and bile salt resistance, maintains its activity and function in the complex environment of the poultry digestive tract. The addition of FeSO4 can further significantly enhance uric acid degradation efficiency, thereby effectively preventing and alleviating hyperuricemia in poultry, demonstrating good application potential. Attached Figure Description

[0020] Figure 1 This is a diagram showing the screening results of the urate oxidase-producing strains of the present invention.

[0021] Figure 2 This is the growth curve of FL1-1 according to the present invention.

[0022] Figure 3 This is a colony morphology diagram of FL1-1 of the present invention. In the diagram, A represents the colony morphology, and B represents Gram staining.

[0023] Figure 4 This is a phylogenetic tree diagram of the FL1-1 bacterium of the present invention.

[0024] Figure 5 This is a diagram showing the uric acid degradation of FL1-1 according to the present invention.

[0025] Figure 6 Fe of the present invention 2+ Chart showing the uric acid-lowering ability of FL1-1.

[0026] Figure 7 This is a diagram showing the acid resistance of FL1-1 according to the present invention.

[0027] Figure 8 This is a diagram showing the bile salt resistance of FL1-1 of the present invention.

[0028] Figure 9 This is a graph showing the antioxidant activity of the FL1-1 fermentation supernatant of the present invention.

[0029] Figure 10 This diagram illustrates the hemolysis of FL1-1 and Staphylococcus aureus according to the present invention. In the diagram, A represents Staphylococcus aureus, and B represents FL1-1 strain.

[0030] Figure 11 This is a diagram of the antibiotic resistance of FL1-1 in this invention. Detailed Implementation

[0031] To enable those skilled in the art to better understand and implement the technical solutions of the present invention, the following detailed description, in conjunction with preferred embodiments and accompanying drawings, provides a clear and complete account of the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, and not all of them. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.

[0032] It should be noted that all technical terms used in this invention are for the purpose of describing specific embodiments only and are not intended to limit the scope of protection of this invention. Unless otherwise specified, all raw materials, reagents, instruments and equipment used in the following embodiments of this invention can be purchased from the market or prepared by existing methods.

[0033] The main materials used in this invention are fresh goose feces, uric acid, uric acid test kit, Columbia blood agar medium, and antimicrobial susceptibility test strips.

[0034] Fresh goose droppings were collected from healthy Pingba grey geese at the Baiyi Base Goose Farm of Guizhou Agricultural Vocational College and stored at 4℃. Uric acid, 99% purity, CAS number 69-93-2, was purchased from Aladdin. A uric acid test kit, model ADS-W-KY010, was purchased from Nanjing Jiancheng Bioengineering Institute; the product name is Uric Acid Content (Uric Acid Enzyme Method) Detection Kit. Columbia blood agar medium, 90mm, was purchased from Jingxi In Vitro Diagnostic Reagents Co., Ltd. Antimicrobial susceptibility testing discs, a complete set of 30 models, were purchased from Bickman Biotechnology Co., Ltd.

[0035] The culture medium used in this invention has the following formulation: LB liquid medium: 10g peptone, 5g sodium chloride, 10g yeast extract, add double-distilled water to 1000mL, and adjust the pH to 7.2.

[0036] LB solid medium: 10g peptone, 5g sodium chloride, 10g yeast extract, 15g agar, add double-distilled water to 1000mL, and adjust the pH to 7.2.

[0037] Enrichment medium: 5g uric acid, 0.5g yeast extract, 0.5g sodium chloride, 0.5g magnesium sulfate, 6.5g dipotassium hydrogen phosphate, 0.5g potassium hydrogen phosphate, add double-distilled water to 1000mL, and adjust the pH to 7.4.

[0038] Uric acid culture medium: 3g uric acid, 0.5g yeast extract, 0.5g sodium chloride, 0.5g magnesium sulfate, 6.5g dipotassium hydrogen phosphate, 0.5g potassium hydrogen phosphate, 15g agar, add double-distilled water to 1000mL, and adjust the pH to 7.4.

[0039] Example 1 The method for obtaining uric acid-lowering bacterial strains includes the following steps: 1. Experimental Methods 1.1 Initial screening of uric acid-lowering strains In a sterile laminar flow hood, 1g of fresh goose feces sample was weighed and added to 10mL of sterile physiological saline. After thorough mixing and standing at room temperature, 2mL of the supernatant was added to 50mL of enrichment medium and incubated at 37℃ and 200rpm for 2 days using a constant temperature shaker. The bacterial concentration was adjusted to 10 using a turbidimetric method. 4 CFU / mL, 10 3 CFU / mL, 10 2CFU / mL, pipette 100 μL of bacterial suspension onto selection medium, spread, and incubate at 37°C until visible colonies are formed. Pick single colonies with a clear zone around the colony and further isolate and purify the strain using LB agar. Incubate the purified strain overnight at 37°C on LB liquid medium, then adjust the bacterial concentration to 10. 5 CFU / mL, the bacterial suspension was inoculated onto uric acid medium, with 3 spots for each strain, and incubated statically at 37℃ for 24 h; the diameter of the clear zone and the colony diameter were measured. The diameter of the clear zone was denoted as H and the colony diameter as C. The H / C ratio was calculated, and the colony with the highest H / C ratio was selected as strain FL1-1.

[0040] 1.2 Growth performance determination The FL1-1 strain was inoculated into LB broth medium and cultured at 37°C for 24 h. The 24-h growth line was plotted.

[0041] 1.3 Strain Identification Colonies were picked from the selected plates using an inoculation loop and streaked onto LB agar plates, then incubated at 37°C for 24 hours. Colony size, shape, color, transparency, edge regularity, and surface roughness were observed. Single colonies were picked for Gram staining. The isolated bacteria were inoculated into LB medium for activation, followed by extraction of bacterial DNA according to the instructions of a bacterial genomic DNA extraction kit. Amplification primers were synthesized by Shanghai Sangon Biotech Co., Ltd., with the nucleotide sequence of 27F as shown in SEQ ID NO.1: AGAGTTTGATCMTGGCTCAG. The nucleotide sequence of 1492R is shown in SEQ ID NO.2: GGTTACCTTGTTACGACTT. The extracted genomic DNA was used as a template for PCR amplification, with denaturation at 95°C for 5 min, followed by 30 cycles of 94°C, 30 s, 57°C, 30 s, 72°C, and 90 s, and an extension at 72°C for 10 min. The PCR products were detected by 1.5% agarose gel electrophoresis, and the amplified target bands were purified and sequenced. The obtained sequences were subjected to BLAST alignment analysis in the NCBI database.

[0042] 2. Experimental Results 2.1 Screening results of uricase-producing strains The screening results of uricase-producing strains are as follows: Figure 1 As shown in the figure. The results showed that on the uricase-producing screening medium, a clear zone formed around the positive strains, while no clear zone formed around the negative strains. The experiment screened out a strain with strong uricase production, FL1-1, with an H / C value as high as 7.72 mm.

[0043] 2.2 Growth Curve The growth curve of strain FL1-1 is as follows Figure 2As shown in the figure. The results showed that strain FL1-1 entered the logarithmic growth phase 2 hours after inoculation, and was in the stationary phase from 10 to 24 hours. At 24 hours, the OD600nm value of strain FL1-1 reached 1.83, indicating strong growth and reproduction capabilities.

[0044] 2.3 Strain Identification FL1-1 colony morphology as follows Figure 3 As shown in the figure. The results showed that strain FL1-1, isolated from the droppings of Pingba grey geese, appeared as yellowish-white colonies on LB agar plates. The colonies were round, slightly raised, with a smooth, moist surface, neat and smooth edges, and were generally opaque. Gram staining was purple, indicating it was a Gram-positive bacterium. The phylogenetic tree of FL1-1 is shown below. Figure 4 As shown in the figure. The results showed that the sequencing results, after BLAST comparison, identified the FL1-1 strain as *Bacillus nicotine*, GenBank number PX735911.

[0045] Comparative Example 1 The in vitro uric acid-lowering method of FL1-1 strain includes the following steps: The FL1-1 strain was cultured in LB liquid medium for 24 h. 2 mL of the culture was centrifuged at 3000 rpm for 10 min, and the precipitate was collected. The precipitate was washed twice with 1 mL of 0.9% sodium chloride solution, resuspended in 0.9% sodium chloride solution, and the bacterial concentration was adjusted to 1 × 10⁻⁶. 9 CFU / mL was added to uric acid culture medium at a volume fraction of 0.5% and cultured at 37℃ and 180 rpm. 2 mL of uric acid culture medium was collected at 24h, 48h, and 72h, and the bacterial concentration was measured simultaneously. The uric acid culture medium was centrifuged at 3000 rpm for 10 min to obtain the supernatant, which was then frozen and stored. After sample collection was completed, the UA concentration in the supernatant was measured using a uric acid detection kit (uricase method). The degradation rate of uric acid was calculated according to formula (1).

[0046] Formula (1): Degradation rate = [original uric acid concentration – uric acid concentration after degradation] / original uric acid concentration × 100%.

[0047] The uric acid-lowering properties of strain FL1-1 are as follows Figure 5 As shown in the figure. The results showed that after culturing in uric acid medium for 3 days, the uric acid degradation rates of this bacterium at 24h, 48h, and 72h were 55.35%, 94.75%, and 95.96%, respectively. The degradation rate increased with the increase of bacterial biomass, and reached a plateau at 48h.

[0048] Example 2 The viable count and Fe of a uric acid-lowering bacterial agent based on *Bacillus glutamicum* 2+ The ratio is 5×10 9 CFU: 3.66mg.

[0049] The application method of the uric acid-lowering bacterial agent includes the following steps: Inoculate FL1-1 strain into LB broth medium and culture at 37℃ for 24 hours. Take 2 mL of the culture medium, centrifuge at 3000 rpm for 10 min, then wash twice with 1 mL of 0.9% sodium chloride solution, resuspend in 0.9% sodium chloride solution, and adjust the bacterial concentration to 1×10⁻⁶. 9 CFU / mL was used to obtain the bacterial suspension of strain FL1-1. The FL1-1 bacterial suspension was inoculated into uric acid medium containing 0.010 g / L FeSO4 at a volume fraction of 0.5%. After incubation at 37℃ and 180 rpm for 24 h, 2 mL of uric acid medium was collected, and the solution was centrifuged at 3000 rpm for 10 min to obtain the supernatant. The UA concentration in the supernatant was measured using a uric acid detection kit. The results showed that the uric acid degradation rate of FL1-1 was 70.89% after 24 h at a FeSO4 concentration of 0.010 g / L.

[0050] Example 3 The viable count and Fe of a uric acid-lowering bacterial agent based on *Bacillus glutamicum* 2+ The ratio is 1×10 9 CFU: 5.48mg.

[0051] The method of applying the uric acid-lowering bacterial agent includes the following steps: The FL1-1 strain was inoculated into LB broth medium and cultured at 37°C for 24 h. 2 mL of the culture was centrifuged at 3000 rpm for 10 min, then washed twice with 1 mL of 0.9% sodium chloride solution, resuspended in 0.9% sodium chloride solution, and the bacterial concentration was adjusted to 2 × 10⁻⁶. 8 CFU / mL was used to obtain the bacterial suspension of strain FL1-1. The FL1-1 bacterial suspension was inoculated into uric acid medium containing 0.015 g / L FeSO4 at a volume fraction of 0.5% and cultured at 37℃ and 180 rpm for 24 h. 2 mL of uric acid medium was collected, and the solution was centrifuged at 3000 rpm for 10 min to obtain the supernatant. The UA concentration in the supernatant was measured using a uric acid detection kit. The results showed that the uric acid degradation rate of FL1-1 was 86.78% after 24 h at a FeSO4 concentration of 0.015 g / L.

[0052] Example 4 The viable count and Fe of a uric acid-lowering bacterial agent based on *Bacillus glutamicum* 2+ The ratio is 5×10 8 CFU: 7.31mg.

[0053] The method of applying the uric acid-lowering bacterial agent includes the following steps: The FL1-1 strain was inoculated into LB broth medium and cultured at 37°C for 24 h. 2 mL of the culture was centrifuged at 3000 rpm for 10 min, then washed twice with 1 mL of 0.9% sodium chloride solution, resuspended in 0.9% sodium chloride solution, and the bacterial concentration was adjusted to 5 × 10⁻⁶. 8 CFU / mL was used to obtain the bacterial suspension of strain FL1-1. The FL1-1 bacterial suspension was inoculated into uric acid medium containing 0.020 g / L FeSO4 at a volume fraction of 0.5% and cultured at 37℃ and 180 rpm for 24 h. 2 mL of uric acid medium was collected, and the solution was centrifuged at 3000 rpm for 10 min to obtain the supernatant. The UA concentration in the supernatant was measured using a uric acid detection kit. The results showed that the uric acid degradation rate of FL1-1 was 76.85% after 24 h at a FeSO4 concentration of 0.020 g / L.

[0054] Example 5 The viable count and Fe of a uric acid-lowering bacterial agent based on *Bacillus glutamicum* 2+ The ratio is 5×10 9 CFU: 10.96mg.

[0055] The method of applying the uric acid-lowering bacterial agent includes the following steps: The FL1-1 strain was inoculated into LB broth medium and cultured at 37°C for 24 h. 2 mL of the culture was centrifuged at 3000 rpm for 10 min, then washed twice with 1 mL of 0.9% sodium chloride solution, resuspended in 0.9% sodium chloride solution, and the bacterial concentration was adjusted to 1 × 10⁻⁶. 9 CFU / mL was used to obtain the bacterial suspension of strain FL1-1. The bacterial suspension of strain FL1-1 was inoculated into uric acid medium containing 0.03 g / L FeSO4 at a volume fraction of 0.5%. After incubation at 37℃ and 180 rpm for 24 h, 2 mL of uric acid medium was collected, and the solution was centrifuged at 3000 rpm for 10 min to obtain the supernatant. The UA concentration in the supernatant was measured using a uric acid detection kit. The results showed that the uric acid degradation rate was 75.12% after 24 h.

[0056] The results of comparing the uric acid-lowering ability of FL1-1 with those of Examples 1 to 5 are as follows: Figure 6 As shown. The results show that, compared with Example 2, under the same conditions, Fe 2+It can significantly enhance the uric acid-lowering ability of FL1-1. The degradation rates after 24 hours at FeSO4 concentrations of 0 g / L, 0.010 g / L, 0.015 g / L, 0.020 g / L, and 0.03 g / L were 55.35%, 70.89%, 86.78%, 76.85%, and 75.12%, respectively. The optimal FeSO4 concentration was 0.015 g / L, which increased the degradation rate to over 157%. Therefore, Fe... 2+ It can improve the uric acid-lowering efficiency of FL1-1.

[0057] To illustrate the beneficial effects of the present invention, the following experiments were also conducted.

[0058] 1. Experimental Methods 1.1 Acid and bile salt resistance The pH of the LB liquid medium was adjusted to 2.5, 3.0, 4.0, and 6.0, respectively, and the bile salt concentrations were set to 0 g / L, 1 g / L, 2 g / L, 3 g / L, and 4 g / L, respectively. FL1-1 was inoculated into the LB liquid medium at a 1% (v / v) inoculum and cultured at 37°C for 24 h. The absorbance of each experimental group at 600 nm was measured using a spectrophotometer. Acid tolerance curves and bile salt tolerance curves of the strains were plotted with pH and bile salt concentration on the x-axis and absorbance on the y-axis, respectively.

[0059] 1.2 Determination of antibacterial properties The antibacterial activity of the strain was determined using the perforation diffusion method, as follows: FL1-1 strain was inoculated into LB broth and cultured for 24 h. 2 mL of the bacterial culture was centrifuged at 8000 rpm for 5 min, and the collected supernatant was filtered through a 0.22 μm sterile filter for sterilization. Staphylococcus aureus and Salmonella typhi were activated and evenly spread onto LB agar plates. After the plate surface was slightly dry, perforations were made, and 100 μL of the treated fermentation broth was added to each well. The plates were then incubated overnight under suitable conditions. The diameter of the inhibition zone was measured using calipers, with three replicates for each strain. Uninoculated LB broth was added to the wells as a negative control, and the experiment was conducted simultaneously.

[0060] 1.3 Determination of DPPH free radical scavenging ability Weigh 0.0394 g of DPPH, add 10 mL of anhydrous ethanol, and sonicate for 2 min to completely dissolve it, preparing a 10 mmol DPPH stock solution. Store the stock solution in a sealed container at 4°C, protected from light. Dilute the DPPH stock solution with anhydrous ethanol to obtain a 0.1 mmol / L DPPH working solution. Inoculate FL1-1 strain into LB liquid medium and collect the supernatant after 12 h, 24 h, and 36 h of culture in LB liquid medium. Take 2 mL of the supernatant from each time period, add 2 mL of DPPH working solution, mix well, and let stand at room temperature in the dark for 30 min. Measure the absorbance at 517 nm using a spectrophotometer. Use 1 mg / mL of Vc as a positive control and LB liquid medium as a blank control. Each time period sample and control was collected three times, and the average value was taken. Calculate the DPPH scavenging rate of the fermentation broth according to formula (2).

[0061] Formula (2): DPPH free radical scavenging rate = [1-(ab) / c]x100%.

[0062] In the formula, a is the absorbance of FL1-1 supernatant and DPPH working solution, b is the absorbance of FL1-1 supernatant and LB medium, and c is the absorbance of LB medium and DPPH working solution.

[0063] 1.4 Safety Measurement 1.4.1 Hemolysis test Bacteria were streaked onto Columbia agar plates, with Staphylococcus aureus as a positive control, and incubated in an anaerobic environment for 24 hours. The presence of hemolytic zones around the colonies was observed, and the type of hemolysis (α-hemolysis, β-hemolysis, or γ-hemolysis) was recorded.

[0064] 1.4.2 Antibiotic susceptibility analysis The test strains were evenly spread on the surface of LB solid medium, and drug sensitivity test discs were attached to them. After incubation at 37°C for 24 hours, the diameter of the inhibition zone was measured, and the sensitivity of the strains to antimicrobial drugs was evaluated based on the size of the inhibition zone.

[0065] 2. Experimental Results 2.1 Acid and bile salt resistance The acid resistance of FL1-1 is as follows: Figure 7 As shown in the figure, the growth of strain FL1-1 was essentially inhibited at an ambient pH of 2, indicating that strongly acidic conditions significantly affected the proliferation of this bacterium. The strain could still grow at pH 3 and 4, but at a slower rate. However, at pH 6.0, the growth of the strain was not significantly affected and essentially returned to normal. This suggests that the bacterium possesses certain acid-resistant characteristics.

[0066] The bile salt tolerance of FL1-1 is as follows: Figure 8As shown in the figure. The results showed that the OD600nm values ​​of FL1-1 bacteria were 0.95, 1.1, 1.07, and 0.96 under bile salt concentrations of 0 g / L, 1 g / L, 2 g / L, and 3 g / L, respectively. FL1-1 could grow well under the 3 g / L bile salt condition, indicating that the FL1-1 strain can tolerate a high bile salt environment.

[0067] 2.2 Antibacterial properties As shown in Table 1, the sterile supernatant of FL1-1 has inhibitory activity against Staphylococcus aureus and Salmonella, and the degree of inhibition varies, with the inhibitory effect on Staphylococcus aureus being the most significant.

[0068] Table 1 Antibacterial effect of FL1-1 2.3 The scavenging effect of FL1-1 fermentation supernatant on DPPH like Figure 9 As shown, the DPPH free radical scavenging rate of the fermentation supernatant of strain FL1-1 showed a trend of first increasing and then decreasing with fermentation time, reaching 83.25% after 24 hours. The scavenging rate of the positive control Vc was approximately 77.11%. This indicates that the fermentation supernatant of strain FL1-1 has DPPH free radical scavenging ability, and its antioxidant activity is relatively more significant at a fermentation time of 24 hours, exceeding that of Vc, demonstrating strong antioxidant potential.

[0069] 2.4 Security 2.4.1 Hemolysis test The hemolytic activity of FL1-1, such as Figure 10 As shown in the figure. The results showed that FL1-1 did not exhibit hemolysis after 48 hours of incubation, with Staphylococcus aureus as a positive control.

[0070] 2.4.2 FL1-1 antibiotic sensitivity FL1-1 sensitivity to antibiotics, such as Figure 11 As shown in the figure. The results show that FL1-1 is sensitive to GEN, CTR, CIP, CC, C, CN, MI, LVX, E, VAN, PIP, and AMK, but not sensitive to OX, TET, IPM, and SXT.

[0071] It should be noted that when numerical ranges are involved in this invention, it should be understood that both endpoints of each numerical range and any value between the two endpoints can be selected. Since the steps and methods used are the same as in the embodiments, preferred embodiments are described in this invention to avoid redundancy. Although preferred embodiments of this invention have been described, those skilled in the art, once they understand the inventive concept of this invention, can make other changes and modifications to these embodiments, and all such changes and modifications fall within the scope of this invention.

[0072] Obviously, those skilled in the art can make various modifications and variations to this invention without departing from its spirit and scope. If such modifications and variations fall within the scope of equivalents of this invention, then this invention is also intended to include such modifications and variations.

Claims

1. A type of Tobacco Glutamate Bacillus ( Glutamicibacter nicotianae ), characterized in that, The *Tobacco glutamate* strain is FL1-1, with accession number CGMCC No. 37236 and accession date of December 29, 2025. The FL1-1 strain can grow in a bile salt environment with a pH greater than or equal to 3.0 and a bile salt concentration less than or equal to 0.3%.

2. The uric acid-lowering bacterial agent based on *Bacillus glutamicum* according to claim 1, characterized in that, It is composed of *Bacillus glutamicum* inoculum and excipients; wherein, the *Bacillus glutamicum* inoculum is a bacterial suspension or freeze-dried powder of *Bacillus glutamicum*, and the excipients are Fe-containing... 2+ Salts or solutions thereof; 3. The uric acid-lowering bacterial agent based on *Bacillus glutamate* according to claim 2, characterized in that, The viable count of the *Bacillus glutamate* bacterial suspension is greater than or equal to 10. 8 CFU / mL.

4. The uric acid-lowering bacterial agent based on *Bacillus glutamicum* according to claim 3, characterized in that, The viable count of the *Bacillus glutamate* bacterial suspension was 10. 8 CFU / mL ~10 9 CFU / mL.

5. The uric acid-lowering bacterial agent based on *Bacillus glutamate* according to claim 2, characterized in that, The number of live bacteria in the uric acid-lowering bacterial agent and the Fe in the excipients 2+ The ratio is 5×10 8 CFU~5×10 9 CFU: 3.66mg~10.96mg.

6. The uric acid-lowering bacterial agent based on *Bacillus glutamate* according to claim 5, characterized in that, The number of live bacteria in the uric acid-lowering bacterial agent and the Fe in the excipients 2+ The ratio is 5×10 9 CFU: 5.48mg.

7. A uric acid-lowering preparation based on *Bacillus nicotine* according to claim 2, characterized in that, The Fe-containing 2+ The salt is ferrous sulfate.

8. The application of the uric acid-lowering bacterial agent based on *Bacillus glutamate* according to claim 2 in the preparation of uric acid-lowering drugs.

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