Lactobacillus plantarum and application thereof in protecting nerves, improving memory and relieving anxiety
By screening and validating Lactobacillus plantarum GXU-U6, the problem of proving the specific effects of existing probiotic strains has been solved, and significant neuroprotective and symptom-improving effects have been achieved on neurodegenerative diseases such as Parkinson's disease.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- GUANGXI UNIV
- Filing Date
- 2026-04-16
- Publication Date
- 2026-06-16
AI Technical Summary
There is a lack of probiotic strains in the current technology that can significantly improve neurodegenerative diseases such as Parkinson's disease, and the functions of existing probiotics are highly strain-specific, making it difficult to prove their specific effects.
A strain of Lactobacillus plantarum GXU-U6 was screened out, and its neuroprotective effects were verified through in vitro cell experiments and in vivo animal experiments. Its fermentation supernatant and fermented food were prepared for application in the prevention and treatment of neurodegenerative diseases.
Lactobacillus plantarum GXU-U6 significantly improved the viability of MPP+-induced damaged SH-SY5Y cells, reduced LDH release rate, ROS level and NO level, and improved MPTP-induced motor dysfunction, cognitive dysfunction and depressive-like behavior in Parkinson's disease mice.
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Figure CN122214201A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the fields of microbial technology and biomedicine, and in particular to a strain of Lactobacillus plantarum and its applications in protecting nerves, improving memory, and relieving anxiety. Background Technology
[0002] The core characteristic of neurodegenerative diseases is irreversible neuronal damage or death, accompanied by reduced neurotransmitters, brain atrophy, or abnormal protein deposition. As the disease progresses, it can lead to systemic dysfunction and even endanger life. Parkinson's disease (PD) is the second most common neurodegenerative disease worldwide. Its main pathological features are the progressive loss of dopaminergic neurons in the substantia nigra pars compacta of the midbrain and the abnormal accumulation of α-synuclein, leading to motor dysfunctions such as resting tremor, rigidity, and bradykinesia, often accompanied by nonmotor symptoms such as depression and cognitive impairment. Current treatments mainly involve drug replacement therapy such as levodopa, which can improve symptoms but cannot slow disease progression, and long-term use can cause serious side effects.
[0003] In recent years, the discovery of the gut-brain axis has provided new insights into the prevention and treatment of neurodegenerative diseases. Studies have shown that gut microbiota dysbiosis is closely related to the occurrence and development of Parkinson's disease. Probiotics can exert neuroprotective effects by regulating the gut microbiota, enhancing intestinal barrier function, and reducing systemic inflammatory responses and oxidative stress. However, the functions of probiotics are highly strain-specific. Even different strains of the same species may exhibit significant differences in metabolite secretion, environmental tolerance, and the ability to modulate the host's immune system. Therefore, screening for probiotics with excellent therapeutic effects on neurodegenerative diseases such as Parkinson's disease has significant application value for developing novel anti-neurodegenerative disease products. Summary of the Invention
[0004] The first objective of this invention is to overcome the shortcomings and deficiencies of the prior art and provide a strain of *Lactobacillus plantarum*.
[0005] A second objective of this invention is to provide a method for culturing the *Lactobacillus plantarum*.
[0006] A third objective of this invention is to provide a fermentation supernatant of Lactobacillus plantarum.
[0007] A fourth object of the present invention is to provide the use of the *Lactobacillus plantarum* and / or the fermentation supernatant of the *Lactobacillus plantarum* in the preparation of products for the prevention, treatment and / or relief of neurodegenerative diseases.
[0008] The fifth objective of this invention is to provide a fermented dairy product.
[0009] The sixth objective of this invention is to provide a fermented fruit and vegetable product.
[0010] The objective of this invention is achieved through the following technical solution:
[0011] A strain of Lactiplantibacillus plantarum, named Lactiplantibacillus plantarum GXU-U6, was deposited on November 17, 2025, at the Guangdong Provincial Microbial Culture Collection Center (GDMCC), located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC No: 67308.
[0012] The morphological characteristics of *Lactobacillus plantarum* GXU-U6 are as follows: it is a Gram-positive bacterium that does not form spores and has rod-shaped cells; on MRS solid medium, the colonies are milky white, round, with neat edges and smooth surfaces.
[0013] A method for culturing the aforementioned *Lactobacillus plantarum* comprises the following steps: inoculating the *Lactobacillus plantarum* into a culture medium and culturing it at 35°C–37°C for 16–48 hours.
[0014] Preferably, the culture medium is an MRS culture medium, such as MRS broth medium, MRS liquid medium, and MRS solid medium.
[0015] Preferably, the culture is a static culture.
[0016] Preferably, the culture time is 16 to 24 hours; more preferably 24 hours.
[0017] A fermentation supernatant of Lactobacillus plantarum, obtained by fermentation of the aforementioned Lactobacillus plantarum.
[0018] The fermentation supernatant of *Lactobacillus plantarum* is prepared by the following method: *Lactobacillus plantarum* is inoculated into a fermentation medium and fermented at 35℃~37℃. The fermentation supernatant is collected by centrifugation, the pH is adjusted to 7.4±0.1, and the mixture is filtered to remove bacteria.
[0019] Preferably, the fermentation medium is MRS liquid medium.
[0020] Preferably, the fermentation culture time is 16 to 48 hours;
[0021] More preferably, the fermentation culture time is 16 to 24 hours.
[0022] Preferably, the centrifugation conditions are: 4°C, 12000 rpm for 8-12 minutes.
[0023] More preferably, the centrifugation conditions are: 4°C, 12000 rpm for 10 minutes.
[0024] Preferably, the filtration sterilization is performed using a sterile filter with a pore size of 0.22 μm or a microporous membrane.
[0025] A composition containing Lactobacillus plantarum, comprising an effective amount of the aforementioned Lactobacillus plantarum cells and / or its fermentation supernatant, and a pharmaceutically or food-grade acceptable carrier.
[0026] The composition may be a pharmaceutical composition, a health food, or a dietary supplement.
[0027] The application of the above-mentioned Lactobacillus plantarum or its combination in the preparation of fermented products (food).
[0028] A fermented product containing Lactobacillus plantarum is produced by fermentation using the aforementioned Lactobacillus plantarum.
[0029] The fermented foods include solid, liquid, and semi-liquid foods; preferably fermented dairy products (such as fermented yogurt), fermented fruit and vegetable products, etc.
[0030] A fermented dairy product is prepared by the following method: after activating the above-mentioned Lactobacillus plantarum, it is inoculated into pure milk and fermented at a constant temperature of 35℃~37℃ for 12~18 hours. After the milk coagulates, it is transferred to 4℃ for refrigeration and maturation.
[0031] Preferably, the inoculation amount of *Lactobacillus plantarum* is calculated based on a final concentration of 2-3% by volume in the system.
[0032] Preferably, the fermentation temperature is 37°C.
[0033] A fermented fruit and vegetable product is prepared by the following method: fresh fruits and vegetables are washed, cut, and mixed evenly with salt. Then, the above-mentioned Lactobacillus plantarum is inoculated, sterile water is added to submerge the vegetables, and the mixture is sealed and fermented at 20-25°C.
[0034] Preferably, the amount of salt used accounts for 2-3% of the weight of the fresh fruits and vegetables.
[0035] Preferably, the inoculum size of *Lactobacillus plantarum* is 10. 5 CFU / g ~ 10 7 CFU / g (based on the weight of fresh fruits and vegetables); preferably 10 6 CFU / g.
[0036] Preferably, the fermentation time is 5 to 7 days.
[0037] The use of at least one of the above-mentioned Lactobacillus plantarum, fermentation supernatant of Lactobacillus plantarum, composition containing Lactobacillus plantarum, and fermented product containing Lactobacillus plantarum in the preparation of products for the prevention, treatment, and / or relief of neurodegenerative diseases.
[0038] Preferably, the neurodegenerative disease includes Parkinson's disease.
[0039] Preferably, the products include pharmaceuticals, functional foods, health (food) products, dietary supplements, etc.
[0040] The above-mentioned use of Lactobacillus plantarum, fermentation supernatant of Lactobacillus plantarum, composition containing Lactobacillus plantarum, and fermentation product containing Lactobacillus plantarum in the preparation of products for protecting nerves, improving or assisting in improving memory, motor dysfunction, cognitive dysfunction, depressive-like behavior, and relieving or assisting in relieving anxiety.
[0041] Preferably, the products include pharmaceuticals, functional foods, health (food) products, and dietary supplements.
[0042] The present invention has the following advantages and effects compared with the prior art:
[0043] 1. The *Lactobacillus plantarum* GXU-U6 strain of this invention is a beneficial active microorganism screened from the feces of healthy humans. This invention employs a multi-strain parallel comparison strategy to screen out a single strain with significant neuroprotective effects from nine strains of the same species. This was comprehensively verified through in vitro cell experiments and in vivo animal experiments, demonstrating the uniqueness and unpredictability of its efficacy, overcoming the deficiency in existing technologies where reporting only a single strain is insufficient to prove its specificity.
[0044] 2. The fermentation supernatant of *Lactobacillus plantarum* GXU-U6 of this invention significantly improved the viability of MPP+-induced SH-SY5Y cells and reduced LDH release rate, ROS level, NO level, and apoptosis rate in an in vitro cell model, indicating that it has a good neuroprotective effect. Comparative experiments showed that eight other strains of the same species did not have significant protective effects under the same conditions.
[0045] 3. The *Lactobacillus plantarum* GXU-U6 of the present invention can significantly improve motor dysfunction (shortened pole climbing time, increased open field movement distance), cognitive dysfunction (improved new object recognition index), and depressive-like behavior (shortened tail immobility time) in an MPTP-induced Parkinson's disease mouse model in vivo, indicating that it has an ameliorative effect on both motor and non-motor symptoms of neurodegenerative diseases such as Parkinson's disease and Alzheimer's disease.
[0046] 4. The Lactobacillus plantarum GXU-U6 of the present invention is derived from the human body, has high safety, and can be widely used in the fields of medicines, health products, and food for the prevention or treatment of Parkinson's disease, and has good development and application prospects. Attached Figure Description
[0047] Figure 1 These are images showing the colony morphology and Gram staining microscopic examination results of Lactobacillus plantarum GXU-U6 of the present invention on MRS agar medium.
[0048] Figure 2 It is the fermentation supernatant of 9 strains of Lactobacillus plantarum against MPP + The effect of induced SH-SY5Y cell viability.
[0049] Figure 3 The fermentation supernatant of *Lactobacillus plantarum* GXU-U6 and eight control strains (GXU-U2, GXU-U3, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, GXU-G6) was compared with that of MPP. + The effect of induced LDH release rate in SH-SY5Y cells.
[0050] Figure 4 The fermentation supernatant of *Lactobacillus plantarum* GXU-U6 and eight control strains (GXU-U2, GXU-U3, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, GXU-G6) was compared with that of MPP. + The effect of induced ROS levels in SH-SY5Y cells.
[0051] Figure 5 The fermentation supernatant of *Lactobacillus plantarum* GXU-U6 and eight control strains (GXU-U2, GXU-U3, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, GXU-G6) was compared with that of MPP. + The effect of induced NO release in SH-SY5Y cells.
[0052] Figure 6 The fermentation supernatant of *Lactobacillus plantarum* GXU-U6 and eight control strains (GXU-U2, GXU-U3, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, GXU-G6) was compared with that of MPP. + The effect of induced apoptosis in SH-SY5Y cells.
[0053] Figure 7This is a graph showing the effect of Lactobacillus plantarum GXU-U6 on the behavior of MPTP-induced Parkinson's disease model mice; where A is the time for the pole climbing test; B is the statistical graph of the recognition index in the new object recognition test; C is the statistical graph of the movement distance in the open field test; and D is the statistical graph of the immobility time in the tail suspension test.
[0054] Figure 8 This is a graph showing the hemolytic activity assessment results of Lactobacillus plantarum GXU-U6 on blood agar plates. Detailed Implementation
[0055] The present invention will be further described in detail below with reference to embodiments, but the embodiments of the present invention are not limited thereto. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in this technical field. Test methods in the following embodiments that do not specify specific experimental conditions are generally performed under conventional experimental conditions. Unless otherwise specified, the reagents and raw materials used in the present invention are all commercially available.
[0056] In the following examples, Excel 2019 and Microsoft Origin 2019 were used to process and analyze the experimental data. SPSS Statistics 27 was used to perform one-way ANOVA and significance tests on the data, and letter notations (such as letters a, b, c, etc.) were used to indicate significant differences between groups. Compared with the model group, *: indicates p < 0.05; **: indicates p < 0.01; ***: indicates p < 0.001; compared with the control group, #: indicates p < 0.05; ##: indicates p < 0.01; ###: indicates p < 0.001.
[0057] Example 1: Screening, isolation, purification, identification, and preservation of Lactobacillus plantarum GXU-U6
[0058] (a) The method for screening strains includes the following steps:
[0059] (1) Sample collection and processing: Fresh fecal samples were collected from healthy volunteers, placed in sterile sampling tubes, stored at 4°C, and transported to the laboratory. 1g of fecal sample was weighed, 9mL of sterile physiological saline was added, and the mixture was vortexed to prepare 10... -1 Diluent.
[0060] (2) Gradient dilution and coating: Take the above 10 -1 The diluent was serially diluted 10-fold with sterile physiological saline to obtain 10 -2 10 -3 10 -4 10 -5 10 -6Series of dilutions. Take 100 μL of each serial dilution and spread it onto MRS solid culture plates, with three replicates for each dilution. Incubate the plates upside down at 37°C for 48 hours.
[0061] (3) Initial screening: Observe the colony morphology on the plate, pick typical single colonies (milky white, round, with neat edges), and purify them by streaking on MRS solid plates. Incubate at 37℃ for 48 hours, and repeat the purification 2-3 times until pure cultures are obtained. Multiple pure cultures of suspected lactic acid bacteria were isolated from the samples.
[0062] (4) Gram staining identification: Select purified single colonies for Gram staining and observe the morphology and staining reaction under an optical microscope. Lactic acid bacteria should be Gram-positive bacteria, with rod-shaped or coccobacillus-shaped cells, arranged singly, in pairs, or in short chains. Select strains that show a Gram-positive reaction for subsequent experiments.
[0063] (5) Peroxidase identification: Pick a single Gram-positive colony, add 3% (v / v) hydrogen peroxide solution to the colony, and observe the formation of bubbles. Lactic acid bacteria are peroxidase-negative and should not produce bubbles. Screen for peroxidase-negative strains for subsequent identification.
[0064] (6) Molecular biological identification and species confirmation: Genomic DNA was extracted from each strain, and PCR amplification was performed using universal primers for bacterial 16S rDNA. The PCR products, which amplified approximately 1500 bp target bands, were sequenced. The obtained 16S rDNA sequences were compared and analyzed using BLAST in the NCBI database. The results showed that the 16S rDNA sequences of 9 strains had more than 99% homology with the standard strain of *Lactiplantibacillus plantarum*, and they were identified as *Lactiplantibacillus plantarum*. These 9 strains were named GXU-U2, GXU-U3, GXU-U6, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, and GXU-G6, respectively. Figure 1 The images show the colony morphology and Gram staining microscopic results of the GXU-U6 of this invention on MRS agar medium.
[0065] The 16S rRNA gene sequence (1564 bp) (SEQ ID NO.1) of the strain GXU-U6, which is the main focus of this patent application, is as follows:
[0066]
[0067] (7) Preservation of strains:
[0068] The selected Lactobacillus plantarum strain GXU-U6 was named Lactobacillus plantarum GXU-U6. This strain was deposited on November 17, 2025, at the Guangdong Provincial Microbial Culture Collection Center (GDMCC), 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC No: 67308.
[0069] (8) Safety assessment of strains
[0070] Fresh cultures of the test strain were collected, and single colonies were picked up using a sterile inoculation loop and streaked onto blood agar plates. The plates were then incubated upside down in a 36℃±1℃ incubator for 18–24 hours. The presence of a clear hemolytic zone (β-hemolysis), a grass-green hemolytic zone (α-hemolysis), or no hemolytic zone (γ-hemolysis) around the colonies was observed to assess the hemolytic activity and potential pathogenicity of the test strain. Results are as follows: Figure 8 As shown, the strain GXU-U6 of this invention is non-hemolytic.
[0071] Example 2: Preparation method of fermentation supernatant for in vitro functional verification and construction of a Parkinson's disease cell model.
[0072] This embodiment describes the preparation method of the fermentation supernatant of the strain for in vitro functional verification, and the construction process of the Parkinson's disease cell model:
[0073] (1) The above-mentioned Lactobacillus plantarum GXU-U6 (GDMCC No: 67308) and eight other Lactobacillus plantarum strains GXU-U2, GXU-U3, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, and GXU-G6 as controls were inoculated into 5 mL of MRS liquid medium and incubated at 37°C for 24 hours. The culture medium was centrifuged at 12,000 rpm for 10 minutes at 4°C, the supernatant was collected, the pH was adjusted to 7.4, and the supernatant was filtered through a sterile filter with a pore size of 0.22 μm to obtain sterile bacterial fermentation supernatant, which was aliquoted and stored at -20°C for later use.
[0074] (2) Human neuroblastoma cells SH-SY5Y (purchased from the Cell Bank of the Chinese Academy of Sciences) were cultured in DMEM / F12 complete medium (containing 10% (v / v) fetal bovine serum and 1% penicillin-streptomycin) at 37℃ and 5% CO2. Logarithmic growth phase cells were harvested and cultured at 5 × 10⁶ cells per well. 5 Cells were seeded at a density of 1,000 cells per well in 96-well plates and cultured for 24 hours to allow for full cell adhesion before use in subsequent experiments.
[0075] Example 3: Strains Screening Experiment Based on Cell Viability Detection
[0076] This embodiment screens for MPP by detecting cell viability. + The strain exhibits protective effects against induced neurotoxicity. The specific steps are as follows:
[0077] The SH-SY5Y cells prepared in Example 2 were divided into multiple experimental groups: normal control group (DMEM / F12 complete culture medium), model group (MPP) + (Contains 500 μM MPP) + The study included DMEM / F12 medium and nine intervention groups (containing 5% (v / v) fermentation supernatant of GXU-U2, GXU-U3, GXU-U6, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, and GXU-G6 strains, respectively, all containing 500 μM MPP). + (DMEM / F12 medium).
[0078] All intervention groups adopted a pretreatment-induction experimental procedure: cells were first incubated for 12 hours in a medium containing 5% (v / v) fermentation supernatant of the corresponding strain, and then the medium was replaced with a medium containing the same concentration of supernatant and 500 μM MPP. + The cells were cultured in DMEM / F12 medium for 24 hours. After culturing, CCK-8 reagent was added to each well, and the reaction was carried out for 2 hours. The absorbance was read at 450 nm. The relative survival rate of cells in each group was calculated with the survival rate of the normal control group defined as 100%. This experiment was independently repeated three times.
[0079] The results are as follows Figure 2 As shown: MPP + The cell viability in the model group was significantly decreased. Among the nine strain intervention groups, only the GXU-U6 strain intervention group showed a highly significant increase in cell viability compared to the model group, while the fermentation supernatant interventions of strains GXU-U2, GXU-U3, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, and GXU-G6 did not show significant protective effects. This indicates that not all *Lactobacillus plantarum* strains possess neuroprotective effects, and that strain GXU-U6 exhibits a unique protective effect.
[0080] Example 4: Cell damage index detection experiment
[0081] In this embodiment, lactate dehydrogenase release rate was used as the detection index to evaluate the MPP response of each tested strain. +The protective ability against nerve damage was assessed. The experimental grouping and cell processing procedures were the same as in Example 3. After the intervention, cell culture supernatants from each group were collected, and the absorbance of each sample was measured at 490 nm according to the operating procedures of the lactate dehydrogenase assay kit to calculate the lactate dehydrogenase release rate. The experiment was conducted in triplicate.
[0082] The results are as follows Figure 3 As shown: MPP + The LDH release rate was significantly increased in the model group. Among the nine strain intervention groups, only the LDH release rate of the *Lactobacillus plantarum* GXU-U6 strain intervention group was significantly lower than that of the model group, while the fermentation supernatant intervention of strains GXU-U2, GXU-U3, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, and GXU-G6 did not show significant protective effects. The experiment confirmed that *Lactobacillus plantarum* GXU-U6 has a unique protective effect on neuronal cell membrane integrity, while the other tested strains were ineffective.
[0083] Example 5: Experiment on the detection of oxidative stress indicators
[0084] This embodiment screens for MPP by detecting intracellular reactive oxygen species (ROS) levels. + The strain exhibited protective effects against induced neuronal damage. Specific grouping and treatment methods are detailed in Example 3. After treatment, cells were incubated with the DCFH-DA probe at 37°C in the dark for 30 minutes. Fluorescence signal intensity was recorded using a microplate reader; fluorescence intensity was positively correlated with reactive oxygen species levels. All experiments were repeated three times.
[0085] The results are as follows Figure 4 As shown: MPP + The ROS level in the model group was significantly increased. Among the nine strain intervention groups, only the *Lactobacillus plantarum* GXU-U6 strain intervention group showed a significant decrease in ROS level compared to the model group, while the fermentation supernatant interventions of strains GXU-U2, GXU-U3, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, and GXU-G6 did not show significant protective effects. The experiment confirmed that *Lactobacillus plantarum* GXU-U6 can specifically reduce intracellular ROS levels in neurons, a function not observed in other tested strains.
[0086] Example 6: Inflammatory Marker Detection Experiment
[0087] This embodiment uses nitric oxide release as the detection index to evaluate the MPP response of each tested strain. +The protective ability against nerve damage was assessed. The experimental grouping and cell processing procedures were the same as in Example 3. After the intervention, cell culture supernatants from each group were collected, and the absorbance at 540 nm was measured according to the instructions of the nitric oxide assay kit (Griess method). The concentration of nitric oxide in the sample was calculated based on the sodium nitrite standard curve. Each treatment group underwent three independent replicate experiments. The experiment was set up with three replicates.
[0088] The results are as follows Figure 5 As shown: MPP + NO release was significantly increased in the model group. Among the nine intervention groups, only the *Lactobacillus plantarum* GXU-U6 strain showed a significant decrease in NO release compared to the model group, while the fermentation supernatant interventions of strains GXU-U2, GXU-U3, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, and GXU-G6 did not show significant protective effects. The experiment confirmed that *Lactobacillus plantarum* GXU-U6 is effective against MPP. + The induced neuroinflammation had a unique inhibitory effect, while other tested strains were ineffective.
[0089] Example 7 Apoptosis Detection Experiment
[0090] This embodiment screens for MPP by detecting cell apoptosis rate. + The strain exhibited protective effects against induced neurotoxicity. The grouping and experimental methods were followed as described in Example 3. After treatment, cells were stained with Annexin V-FITC / PI apoptosis detection kit, and the apoptosis rate was detected by flow cytometry. The experiment was conducted in triplicate.
[0091] The results are as follows Figure 6 As shown: MPP + The apoptosis rate in the model group was significantly increased. Among the nine strain intervention groups, only the *Lactobacillus plantarum* GXU-U6 strain intervention group showed a significantly lower apoptosis rate compared to the model group, while the fermentation supernatant interventions of strains GXU-U2, GXU-U3, GXU-A10, GXU-A11, GXU-ES7, GXU-ES10, GXU-G5, and GXU-G6 did not show significant protective effects. The experiment confirmed that *Lactobacillus plantarum* GXU-U6 can specifically reduce the apoptosis rate of neuronal cells, a function not observed in other tested strains.
[0092] Example 8: Therapeutic effect of Lactobacillus plantarum GXU-U6 on MPTP-induced Parkinson's disease model mice
[0093] (1) Preparation of bacterial agent: Lactobacillus plantarum GXU-U6 was inoculated into MRS liquid medium and cultured at 37℃ for 12 hours. The bacterial cells were collected by centrifugation at 8000rpm for 10 minutes at 4℃, washed twice with sterile physiological saline, resuspended in sterile physiological saline, and the concentration was adjusted to 1×10⁻⁶. 9 The bacterial agent for gavage is obtained by measuring CFU / mL.
[0094] (2) Animal grouping and treatment: Eight-week-old male C57BL / 6J mice, weighing 20-25g, were selected and randomly divided into four groups (n=10 per group) after one week of acclimatization: blank control group (Control), model group (MPTP), GXU-U6 intervention group (U6+MPTP), and positive control group (L-DOPA+MPTP). The blank control group received an equal volume of physiological saline intraperitoneally daily for 6 consecutive days, followed by gavage administration of 0.2mL sterile physiological saline for 42 consecutive days. The model group received an intraperitoneal injection of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, 30mg / kg body weight) daily for 6 consecutive days, followed by gavage administration of 0.2mL sterile physiological saline for 42 consecutive days. The GXU-U6 intervention group received an intraperitoneal injection of MPTP (30mg / kg body weight) daily for 6 consecutive days, followed by gavage administration of 0.2mL GXU-U6 bacterial agent (1×10⁻⁶) for 6 consecutive days. 9 CFU / mL), for 42 consecutive days. Positive control group: levodopa / benserazide combination preparation was administered by gavage daily (equivalent dose calculated based on mouse body weight, equivalent to levodopa 25 mg / kg / d combined with benserazide 6.25 mg / kg / d).
[0095] Example 9 Animal behavioral testing
[0096] This embodiment uses pole climbing, novel object recognition, open field, and tail suspension tests. Specific experimental procedures were conducted according to the methods described in the literature (Reference 1: Zou Renying. Efficacy evaluation and mechanism study of compound probiotics in relieving depression [D]. Jiangnan University, 2021. Reference 2: Zhang Tao. Effects and mechanism of probiotic Probio-M8 on Alzheimer's disease mice [D]. Inner Mongolia Agricultural University, 2026.). The effects of the GXU-U6 strain on motor coordination, cognitive function, voluntary motor ability, and depressive-like behavior in Parkinson's disease model mice were comprehensively evaluated. The experimental subjects were the mice in the intervention groups described in Example 8.
[0097] The results of the pole climbing experiment show that ( Figure 7 In A): The head-turning time and bar-down time of the MPTP model group mice were significantly longer than those of the control group; while the two indicators of the GXU-U6 strain intervention group mice were significantly shortened and close to the level of the control group, indicating that the strain can effectively improve the motor coordination ability of the model mice.
[0098] The results of the new object recognition experiment show that ( Figure 7 In B): The control group mice showed a clear preference for new objects and had a high discrimination index; the model group mice had a significantly lower discrimination index; the GXU-U6 strain intervention group mice had a significantly higher discrimination index than the model group, indicating that their cognitive function was improved.
[0099] The results of the open field experiment show that ( Figure 7 In the C group, the total movement distance of mice in the model group was significantly reduced compared with that of the control group, and their activity was limited to the peripheral area; the total movement distance of mice in the GXU-U6 strain intervention group was significantly restored, and the number of times they explored the central area increased, indicating that this strain can effectively improve the voluntary motor dysfunction of model mice.
[0100] The results of the tail suspension experiment showed that ( Figure 7 D): The latency period for the first immobile state in the model group mice was significantly shorter than that in the control group; the latency period for the first immobile state in the GXU-U6 strain intervention group mice was significantly longer, indicating that the strain can effectively improve the depressive-like behavior associated with the model mice.
[0101] Example 10 Preparation of fermented food from Lactobacillus plantarum GXU-U6
[0102] Preparation of fermented dairy products: After activating Lactobacillus plantarum GXU-U6, it is inoculated into commercially available pure milk at an inoculation rate of 2% (v / v), mixed well, and then dispensed into sterile containers. It is fermented in a constant temperature incubator at 37°C for 12-18 hours. After the milk coagulates, it is transferred to 4°C for refrigeration and ripening to obtain fermented yogurt containing live Lactobacillus plantarum of the present invention.
[0103] Preparation of fermented fruit and vegetable products: Fresh vegetables are washed, cut, and mixed with 2-3% (by weight) salt. The mixture is then packed tightly into a fermentation tank. Lactobacillus plantarum GXU-U6 is activated and fermented at 10... 6 Inoculate vegetables with an inoculum of CFU / g, add sterile water to submerge the vegetables, seal and ferment at 20-25℃ for 5-7 days to obtain fermented kimchi containing the Lactobacillus plantarum of this invention.
[0104] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.
Claims
1. A strain of *Lactobacillus plantarum*, characterized by: The strain, named Lactiplantibacillusplantarum GXU-U6, was deposited on November 17, 2025, at the Guangdong Provincial Center for Microbial Culture Collection, 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC No: 67308.
2. A method for culturing *Lactobacillus plantarum* as described in claim 1, characterized in that, The specific steps are as follows: inoculate the above-mentioned Lactobacillus plantarum into the culture medium and culture it at 35℃~37℃ for 16~48 hours.
3. A fermentation supernatant of Lactobacillus plantarum, characterized in that: Obtained by fermentation of Lactobacillus plantarum as described in claim 1.
4. A composition containing Lactobacillus plantarum, characterized in that: It comprises an effective amount of the *Lactobacillus plantarum* cells and / or its fermentation supernatant as described in claim 1, and a pharmaceutically or food-grade acceptable carrier.
5. The use of the *Lactobacillus plantarum* of claim 1 or the composition containing *Lactobacillus plantarum* of claim 4 in the preparation of fermented products.
6. A fermented product containing Lactobacillus plantarum, characterized in that: It is produced by fermentation using Lactobacillus plantarum as described in claim 1.
7. A fermented dairy product, characterized in that, The following method was used to prepare the product: after activating the Lactobacillus plantarum described in claim 1, it was inoculated into pure milk and fermented at a constant temperature of 35℃~37℃ for 12~18 hours. After the milk coagulated, it was transferred to 4℃ for refrigeration and ripening. The inoculation amount of *Lactobacillus plantarum* is calculated based on a final concentration of 2-3% by volume in the system.
8. A fermented fruit and vegetable product, characterized in that, The following method is used to prepare the product: Fresh fruits and vegetables are washed, cut, and mixed evenly with salt. Then, the plant lactobacillus described in claim 1 is inoculated, and sterile water is added to submerge the vegetables. After sealing, the mixture is fermented at 20-25°C to obtain the product. The amount of salt used is 2-3% of the weight of the fresh fruits and vegetables; The inoculation amount of *Lactobacillus plantarum* is 10. 5 CFU / g ~ 10 7 CFU / g; The fermentation time is 5 to 7 days.
9. The use of at least one of the following in the preparation of products for the prevention, treatment and / or relief of neurodegenerative diseases: the *Lactobacillus plantarum* of claim 1, the fermentation supernatant of *Lactobacillus plantarum* of claim 3, the composition containing *Lactobacillus plantarum* of claim 4, and the fermented product containing *Lactobacillus plantarum* of claim 6.
10. The application according to claim 9, characterized in that: The neurodegenerative diseases mentioned include Parkinson's disease; The products mentioned include pharmaceuticals, functional foods, health products, or dietary supplements.