Lactobacillus sleivae harbinensis z152 strain, its probiotics and application in preparation of food allergy relief preparation

Abstract

This invention discloses a strain of *Lactobacillus schreiberensis* Z152 from Harbin, its metabiotic, and its application in the preparation of food allergy relieving agents. This strain was deposited at the Guangdong Provincial Microbial Culture Collection Center (GDMCC) on May 8, 2026, with accession number GDMCC No: 68230. This strain exhibits excellent self-aggregation characteristics, surface hydrophobicity, cell adhesion, and gastrointestinal fluid tolerance, colonizing the host intestine with high safety. This strain can alleviate food allergies by inhibiting hyaluronidase and β-HEX inhibitory activity, reducing the levels of allergy-related cytokines. Animal experiments have also confirmed that it can increase mouse body weight, reduce spleen index, alleviate hypothermia, and reduce immunoglobulin levels, significantly alleviating food allergy symptoms; simultaneously, it can also lower blood lipids. Therefore, the *Lactobacillus schreiberensis* Z152 strain from Harbin and its metabiotic provided by this invention can be used in anti-food allergy preparations, showing promising application prospects.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology, specifically relating to a strain of *Lactobacillus schleiferus* Z152 from Harbin, its postbiotics, and its application in the preparation of food allergy relieving agents. Background Technology

[0002] Food allergy (FA) is an immune stress response triggered by specific protein antigens in food. Clinically, it can manifest as oral itching, diarrhea, asthma, and difficulty breathing. In severe cases, it can lead to shock and death. Its prevalence is increasing annually and has become a global public health problem. Currently, there is no cure for food allergy. The core prevention method is to avoid contact with allergens. While clinical anti-allergy drugs such as antihistamines and corticosteroids are fast-acting, they are prone to drug resistance and have side effects. Therefore, developing natural and safe anti-allergy substances has significant clinical and social value.

[0003] Probiotics, prebiotics, and post-biotics are research hotspots in the field of lactic acid bacteria. my country's abundant fermented food resources provide a foundation for the development of probiotic functional lactic acid bacteria in the fight against food allergies. Probiotics need to be in live form at a sufficient dosage (≥10). 6 To exert a probiotic effect, probiotics must contain at least 100 CFU / g of live bacteria and possess characteristics such as resistance to gastrointestinal fluids, lack of hemolytic activity, and easy intestinal colonization. However, the activity of live probiotics is easily affected by storage, processing, and the gastrointestinal environment, limiting their application. Compared to probiotics, metabiotics have better stability and safety, are easier to store and transport, and have a lower risk of antimicrobial resistance. Metabiotics include inactivated bacterial cells, metabolites, and / or bacterial lysate components. Currently, [the relevant regulations / standards are not provided in the original text]. Schleiferilactobacillus harbinensisThe article "JNDM Postbiotics Alleviate Atopic Dermatitis with Concurrent Changes in Gut Microbiota and Fecal SCFAs" discloses the efficacy of cell-free supernatant and lysates derived from Lactobacillus schefflera JNDM strain in a mouse model of DNFB-induced atopic dermatitis. The results showed that, at the immunological level, treatment inhibited the Th2-dominated inflammatory cascade and reduced serum IgE and IFN-γ levels. The lysates exhibited superior systemic effects, significantly inhibiting mast cell infiltration and reducing the spleen index. Furthermore, postbiotic intervention can remotely remodel the gut microbiota, particularly reversing the depletion of beneficial bacteria and inhibiting their compensatory increase. In other words, postbiotics from Lactobacillus schefflera JNDM strain, especially its lysates, can improve atopic dermatitis through a dual mechanism of local barrier repair and systemic metabolic regulation via the gut-skin axis. Although this article discloses the role of Lactobacillus schleiferus JNDM strain and its postbiotics in regulating Th1 / Th2 immune balance, inhibiting mast cell activation, and remodeling gut microbiota, providing a strain with better efficacy and a clear ability to alleviate food allergies still has important application value. Summary of the Invention

[0004] The purpose of this invention is to overcome the aforementioned defects and deficiencies in the prior art and to provide a strain of *Lactobacillus schleiferus* from Harbin (…). Schleiferilactobacillus harbinensis Z152 strain.

[0005] The second objective of this invention is to provide a metabiotic prepared using the above-mentioned Harbin Schleifer Lactobacillus Z152 strain.

[0006] A third objective of this invention is to provide the use of the aforementioned Harbin Schleifer Lactobacillus Z152 strain or the aforementioned metabiotic in the preparation of drugs for relieving / preventing food allergies and / or lowering blood lipids.

[0007] A fourth objective of this invention is to provide a drug comprising the above-mentioned Harbin Schleifer Lactobacillus Z152 strain or the above-mentioned metabiotic.

[0008] The above-mentioned objective of this invention is achieved through the following technical solution: This invention provides a strain of *Lactobacillus schleifera* from Harbin (…). Schleiferilactobacillus harbinensis The strain Z152 was deposited at the Guangdong Provincial Microbial Culture Collection Center (GDMCC) on May 8, 2026, with accession number GDMCC No: 68230.

[0009] This invention isolates and purifies a strain capable of hydrolyzing CaCO3 from fermented cowpeas in Hezhou, Guangxi, and identifies it as *Lactobacillus schleifera* (Harbin). Schleiferilactobacillus harbinensis The Z152 strain, deposited at the Guangdong Provincial Microbial Culture Collection Center (GDMCC), located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, Guangdong Province, was deposited on May 8, 2026, with accession number GDMCC No. 68230. This strain exhibits excellent self-aggregation characteristics, hydrophobic surface, cell adhesion, and gastrointestinal fluid tolerance, enabling it to successfully colonize the host gut, exert stable probiotic effects, and demonstrate high safety due to sensitivity to 10 common antibiotics and γ-hemolysis.

[0010] This invention also provides a metabiotic prepared using the aforementioned *Lactobacillus schreiformis* Z152 strain from Harbin. The metabiotic comprises one or more of the following: fermentation / metabolites of *Lactobacillus schreiformis* Z152 strain, abiotic *Lactobacillus schreiformis* Z152 strain, and lysates of *Lactobacillus schreiformis* Z152 strain. The metabiotic (fermentation / metabolites of *Lactobacillus schreiformis* Z152 strain) has a total sugar content of 58.03 ± 1.26 μg / mL and a total protein content of 55.45 ± 2.72 μg / mL; its main components include phenyllactic acid, hydroxyisocaproic acid, citrate, bilirubin, 2-piperidinic acid, L-pyroglutamic acid, D-lactic acid, and N-acetylglucosamine.

[0011] Furthermore, the preparation method of the post-genetic agent is to inoculate the above-mentioned Harbin Schleifer Lactobacillus Z152 strain into a culture medium, culture it, sterilize it, and filter it to obtain the product.

[0012] Furthermore, the method for preparing the post-genetic agent includes the following steps: S1. The activated Harbin Schleifer Lactobacillus Z152 strain was inoculated into MRS liquid medium at a volume inoculation rate of 2% and cultured at 37℃ for 24 h. S2. Place the fermentation system in a 100℃ water bath for 15 min to inactivate it, cool it to room temperature, filter it under reduced pressure, and collect the filtrate and cell precipitate. S3. After rotary evaporation of the filtrate, it was pre-cooled at -80℃ for 24 h, and then freeze-dried at -80℃ under vacuum of 15 Pa for 48 h to obtain the post-biotic (fermentation / metabolite of Lactobacillus schleiferus Z152 strain from Harbin). The freeze-dried product was stored at -20℃. The bacterial precipitate was pre-cooled at -80℃ for 24 h, vacuumed at 15 Pa, and freeze-dried at -80℃ for 48 h to obtain inactivated bacterial cells, which were then stored at -20℃.

[0013] Preferably, the MRS liquid culture medium contains 17.6 g glucose, 5.4 g yeast extract, 2.2 g triammonium citrate, 10.0 g tryptone, 5.0 g anhydrous sodium acetate, 2.0 g dipotassium hydrogen phosphate, 0.2 g anhydrous magnesium sulfate, 1.5 mL Tween 80, and 0.05 g manganese sulfate, with an initial pH of 6.5.

[0014] The standardized preparation method of Z152 strain metabiotic provided by this invention yields metabiotic products with high stability and convenient storage and transportation, overcoming the limitation that the activity of live probiotics is easily affected by the environment. Furthermore, the metabiotic also has functions such as lowering blood lipids, anti-oxidation, and food thickening / stabilizing, making it suitable for a wide range of applications.

[0015] The present invention also provides the application of the above-mentioned Harbin Schleifer Lactobacillus Z152 strain or the above-mentioned metabiotic in the preparation of antioxidant products.

[0016] The present invention also provides the application of the above-mentioned Harbin Schleifer Lactobacillus Z152 strain or the above-mentioned postbiotic in the preparation of food additives, wherein the food additives are food thickeners, thickeners or stabilizers.

[0017] This invention also investigated the effects of strain Z152, inactivated bacterial cells, and postbiotics (fermentation / metabolites of *Lactobacillus schleiferus* strain Z152 from Harbin). The results showed that strain Z152, inactivated bacterial cells, and postbiotics (fermentation / metabolites of *Lactobacillus schleiferus* strain Z152 from Harbin) all have high hyaluronidase and β-HEX inhibitory activities, and can significantly reduce the levels of allergy-related cytokines such as IL-4, IL-5, IL-13, and TNF-α in RBL-2H3 cells. Animal experiments confirmed that it can significantly alleviate food allergy symptoms, increase weight gain in mice, reduce spleen index, alleviate body temperature drop, and reduce the levels of immunoglobulins such as IgE, histamine, and IgG1. It can be used as a probiotic / postbiotic to prepare anti-food allergy preparations.

[0018] Therefore, the present invention also provides the use of the above-mentioned Harbin Schleifer Lactobacillus Z152 strain or the above-mentioned metabiotic in the preparation of drugs for relieving / preventing food allergies and / or lowering blood lipids.

[0019] Furthermore, the drug achieves relief / prevention by inhibiting hyaluronidase and β-HEX.

[0020] Furthermore, the drug achieves relief / prevention by reducing and downregulating the levels of cytokines IL-4, IL-5, IL-13, and TNF-α.

[0021] Furthermore, the drug achieves relief / prevention by downregulating serum levels of immunoglobulin IgE, IgG1, histamine, and β-LG-specific IgE.

[0022] Furthermore, the food allergy is a food allergy induced by ovalbumin.

[0023] The present invention also provides a drug comprising the above-mentioned Harbin Schleifer Lactobacillus Z152 strain or the above-mentioned metabiotic.

[0024] Furthermore, the drug may also contain other pharmaceutically acceptable carriers.

[0025] Furthermore, the concentration of *Lactobacillus schleiferus* Z152 strain (live or inactivated) in the drug is 10. 8 ~10 10 .

[0026] Furthermore, the concentration of *Lactobacillus schleiferus* Z152 strain (live or inactivated) in the drug is 10. 9 .

[0027] Furthermore, the dosage of the post-biotic (fermentation / metabolite of Lactobacillus schleiferus Z152 strain from Harbin) is 300 mg BW / 1000g.

[0028] Compared with the prior art, the present invention has the following beneficial effects: This invention provides a strain of *Lactobacillus schleifera* Z152 from Harbin, its postbiotic, and its application in the preparation of a food allergy relieving agent. This strain was deposited at the Guangdong Provincial Microbial Culture Collection Center (GDMCC) on May 8, 2026, with accession number GDMCC No: 68230. This strain exhibits excellent self-aggregation characteristics, surface hydrophobicity, cell adhesion, and gastrointestinal fluid tolerance, enabling it to successfully colonize the host intestine and exert stable probiotic effects. It is sensitive to 10 common antibiotics, exhibits γ-hemolysis, and has high safety. Furthermore, it can alleviate food allergies by inhibiting hyaluronidase and β-HEX inhibitory activity, reducing the levels of allergy-related cytokines such as IL-4, IL-5, IL-13, and TNF-α in RBL-2H3 cells. Animal experiments have also confirmed that it can increase weight gain in mice, reduce spleen index, alleviate hypothermia, and reduce the levels of immunoglobulins such as IgE, histamine, and IgG1, significantly alleviating food allergy symptoms. Simultaneously, it can lower blood lipids and possess antioxidant properties. The Harbin Schleifer Lactobacillus Z152 strain and its postbiotic provided by this invention can be used in anti-food allergy preparations, showing promising application prospects. Attached Figure Description

[0029] Figure 1 Colony morphology and Gram staining microscopic images of Lactobacillus schleiferus Z152 strain from Harbin.

[0030] Figure 2Phylogenetic tree of 16S rRNA of Lactobacillus schleiferus Z152 strain from Harbin.

[0031] Figure 3 The results show the tolerance of the Harbin Schleifer Lactobacillus Z152 strain to artificial simulated gastric and intestinal fluids.

[0032] Figure 4 Hemolytic plate image of Lactobacillus schreiberensis strain Z152 from Harbin.

[0033] Figure 5 The in vitro binding rates of bile salts and cholesterol of different concentrations of *Lactobacillus schleifera* Z152 postbiotic were determined.

[0034] Figure 6 To investigate the in vitro antioxidant activity of different concentrations of *Lactobacillus schleifera* Z152 postbiotic.

[0035] Figure 7 The results show the detection rates of inhibition of hyaluronidase (A) and β-HEX (B) by strains Z152, LGG, and JNDM, inactivated bacterial cells, and metabiotics. Note: Different groups... This indicates a significant difference. p <0.05.

[0036] Figure 8 The results show the effects of Z152, LGG, and JNDM strains, inactivated bacterial cells, and metabiotics on the levels of TNF-α, IL-4, IL-5, and IL-13 in RBL-2H3 cells. Note: Different letters between different groups indicate significant differences. p <0.05.

[0037] Figure 9 This study presents the effects of strain Z152, inactivated bacterial cells, metabiotics, and JNDM metabiotics on serum IgE, histamine, β-LG-specific IgE, and IgG1 levels in allergic mice. Note: Different letters between different groups indicate significant differences. p <0.05. Detailed Implementation

[0038] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, but the embodiments do not limit the present invention in any way. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in this technical field.

[0039] Unless otherwise specified, all reagents and materials used in the following examples are commercially available.

[0040] Example 1: Isolation, purification and identification of Lactobacillus schleiferus strain Z152 from Harbin 1. Isolation and purification of strain Z152 The isolation and purification process of the Z152 strain described in this invention is as follows: A sample of pickled cowpeas purchased from a market in Hezhou, Guangxi, was crushed, filtered through sterile gauze, and the filtrate was diluted 10 times in a serial gradient. 2 10 3 10 4 Take 300 μL of different concentrations of dilution and spread them on MRS medium plates containing 0.5% (v / m) CaCO3. Incubate at 37℃ for 48 h. After the incubation, pick out the colonies with larger CaCO3 hydrolysis zones from the plates where the colonies are evenly dispersed. Then, streak the colonies three times until the colony morphology is uniform. Then preserve the colonies with 20% (v / v) glycerol and number them. Store them in an ultra-low temperature freezer at -80℃.

[0041] 2. Identification of strain Z152 Single colonies of strain Z152 isolated by streak plating were cultured on MRS solid medium for 48 h, and the colony morphology was observed. The results are as follows: Figure 1 As shown in Figure A, the colonies of strain Z152 are 2–4 mm in size, round in shape, slightly raised in the center, with relatively neat edges, and a moist, milky-white, opaque surface. Gram staining microscopic examination results are as follows. Figure 1 As shown in B, strain Z152 is a Gram-positive bacillus.

[0042] This invention also used strain Z152 as a template for 16S molecular biological identification, the method of which is as follows: PCR amplification was performed using universal primers 1492R (5'-CTACGGCTACCTTGTTACGA-3') and 27F (5'-AGAGTTTGATCCTGGCTCAG-3') for bacterial rRNA gene; the amplification system was: 9.5 μL sterile water, 1.0 μL primer 1492R, 1.0 μL primer 27F, 12.0 μL Taq enzyme, and 1.5 μL template, totaling 25 μL; the amplification program was: 95℃ pre-denaturation for 2 min; 95℃ denaturation for 20 s, 65℃ annealing for 1 min 20 s, 70℃ extension for 1 min 35 s, 40 cycles; 70℃ extension for 12 min; the PCR amplification product was detected by electrophoresis on a 1.5% agarose gel and then sent to Shanghai Sangon Biotech Co., Ltd. for sequencing.

[0043] The sequencing results of the 16S rRNA of strain Z152 are shown below:

[0044] The 16S rRNA sequence obtained from sequencing was compared with the GeneBank database, and a phylogenetic tree of the strain was drawn as follows: Figure 2 As shown, the 16S rRNA alignment results of strain Z152 indicate that its 16S rDNA sequence is similar to that of *Lactobacillus schleifera* from Harbin. Schleiferilactobacillus harbinensis The homology rate of SBT10908 (NR041263.1) was 99.79%, therefore strain Z152 was identified as *Lactobacillus hystericus*. The *Lactobacillus hystericus* strain described in this invention (…) Schleiferilactobacillus harbinensis Strain Z152 is deposited at the Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, Guangdong Province. The deposit date is May 8, 2026, and the accession number is GDMCC No. 68230.

[0045] Example 2 Evaluation of the biological characteristics of Lactobacillus schleiferus strain Z152 from Harbin I. Experimental Methods 1. Self-aggregation of strain Z152 The specific operation procedure is as follows: Z152 strain and commercially available *Lactobacillus schleifera* JNDM (GDMCC No. 64143, with a 2.36% heterologous sequence to Z152) were inoculated at a 2% (v / v) in MRS medium for three generations. After centrifugation for 8 min (4℃, 12000 r / min), the supernatant was discarded. The bacterial cells were then washed twice with PBS buffer (0.1 mol / L, pH 6.86), followed by centrifugation to collect the bacterial pellet. The bacterial pellet was resuspended in PBS buffer, and the bacterial concentration was adjusted to 10. 8 CFU / mL (OD) 600nm (Approximately 0.60); Take 6 mL of bacterial culture, shake thoroughly, and let stand at room temperature for 6 h. Take 200 μL of the supernatant and measure the OD. 600nm The formula for calculating the self-condensation rate is as follows: Self-cohesion rate (%) = [(A0-A j ) / A0] / ×100%; where A0 is the OD at 0 h. 600nm A j OD at 6 h 600nm .

[0046] 2. Surface hydrophobicity of strain Z152 Adjusting the bacterial culture OD of the strain 600nm The value was 0.60. Two groups of 6 mL bacterial cultures were taken, and 2 mL of chloroform and toluene were added to each, respectively. The control group received neither chloroform nor toluene. The cultures were vortexed for 120 s and allowed to stand for 8–10 min until separation. The lower aqueous phase was collected and analyzed at OD0.05.600nm The values ​​of H and H0 were measured and recorded. The experiment was repeated three times. The formula for calculating the hydrophobicity is as follows: D(%) = [(H0-H) / H0] × 100%; where: D represents hydrophobicity; H0 and H are the hydrophobicity of the bacterial culture before and after mixing with organic solvent, respectively. 600nm Measure the absorbance below.

[0047] 3. Caco-2 cell adhesion ability assay Construction of in vitro cell adhesion model: Commercially available Caco-2 cells were cultured in DMEM high glucose medium containing 12% fetal bovine serum at 37°C under 5% CO2 conditions until the cells reached about 80% confluence. The cells were then passaged into 6-well plates at a concentration of 600,000 cells / mL. The medium was changed every 48 hours until the cells were fully differentiated. The culture time was about 15 days. At this point, the in vitro cell adhesion model was successfully established.

[0048] Adhesion assay: Two bacterial suspensions resuspended in DMEM medium were added to the in vitro adhesion model of differentiated Caco-2 cells, with the viable cell count adjusted to approximately 1 × 10⁻⁶ cells / year. 8 The concentration of CFU / mL was recorded as N0, and the cells were incubated in a cell culture environment for 2 hours. The supernatant was removed, and the cells were washed twice with sterile PBS. Cells and bacterial cells were then digested with trypsin, collected, and counted for viability, recorded as N1. Adhesion rate was calculated using the following formula: Adhesion rate (%) = (N1 / N0) × 100%.

[0049] II. Experimental Results The adhesion properties of *Lactobacillus scheffleriana* Z152 and commercially available *Lactobacillus scheffleriana* JNDM from Harbin are shown in Table 1. The self-aggregation and surface hydrophobicity of *Lactobacillus scheffleriana* Z152 in toluene and chloroform are significantly higher than those of commercially available *Lactobacillus scheffleriana* JNDM. The in vitro adhesion rate to Caco-2 cells is also much higher than that of commercially available *Lactobacillus scheffleriana* JNDM. These results indicate that *Lactobacillus scheffleriana* Z152 has better self-aggregation, surface hydrophobicity in toluene and chloroform, and adhesion properties.

[0050] Table 1. Adhesion properties of *Lactobacillus schleiferus* Z152 from Harbin and commercially available *Lactobacillus schleiferus* from Harbin.

[0051] Note: There are significant differences between different letter groups in the same row of data in the table (p < 0.05); results are expressed as mean ± standard deviation.

[0052] Example 3: Tolerance of Harbin Schleifer Lactobacillus Z152 strain to artificial gastric juice I. Experimental Methods 1. Tolerance of strain Z152 to artificial gastric juice The Z152 strain was activated for three generations in MRS medium at an inoculum of 2% (v / v). 1.0 mL of the bacterial culture was centrifuged for 6 min (4℃, 12000 r / min), and the supernatant was discarded. The cells were washed three times with PBS buffer (0.1 mol / L, pH 6.86) and then inoculated into 9.0 mL of artificial gastric fluid, with PBS buffer as a control. After incubation at 37℃ for 2 h, viable cell counts were performed using the dilution plate method. The artificial gastric fluid consisted of 50 mmol / L NaHCO3, 8 mmol / L KCl, 120 mmol / L NaCl, and 5 g / L pepsin, prepared with deionized water, pH adjusted to 2.0, and filtered through a 0.22 μm filter to remove contaminants.

[0053] 2. Tolerance of strain Z152 to artificial intestinal fluid Take 1.0 mL of the bacterial culture after treatment with the simulated gastric fluid for 2 h, and then treat it in 9.0 mL of artificial simulated intestinal fluid for 8 h, taking samples every 2 h. Measure the viable bacterial count after incubation in the artificial simulated intestinal fluid for 2, 4, 6, and 8 h. The artificial simulated intestinal fluid was prepared by adding 3 g / L ox bile salt and 1 g / L trypsin to deionized water, adjusting the pH to 7.5, and filtering with a 0.22 μm filter for sterilization.

[0054] II. Experimental Results The results of the tolerance test of strain Z152 to artificial gastric and intestinal fluids are as follows: Figure 3 As shown, the initial viable count of strain Z152 was 8.81 × 10⁻⁶. 8 The CFU / mL count was 8.83 × 10⁻⁶ after treatment with simulated gastric fluid for 2 hours. 8 The viable bacterial count after treatment with simulated gastric fluid for 2 hours and simulated intestinal fluid for 8 hours was 7.26 × 10⁻⁶ CFU / mL. 7 The CFU / mL count indicates that strain Z152 exhibits good resistance to artificial gastric and intestinal fluids, consistent with the requirement that the live probiotic count reaches 10^6 in the colon. 6 The standard is above CFU / g.

[0055] Example 4: Safety evaluation of Lactobacillus schleifer strain Z152 from Harbin. I. Experimental Methods 1. Evaluation of hemolytic activity Single colonies of strain Z152 were inoculated onto sheep blood agar plates and incubated at 37°C for 48 h to observe their hemolytic state. Three characteristics of hemolysis were observed on sheep blood agar plates: ① α-hemolysis: a grass-green hemolytic ring appears around the colony, indicating conditional pathogenicity; ② β-hemolysis: a clear hemolytic ring appears around the colony, indicating strong pathogenicity; ③ γ-hemolysis: no hemolytic ring appears around the colony, indicating no pathogenicity.

[0056] 2. Antibiotic sensitivity test Adjusting the OD of bacterial culture of strain Z152 600nm The bacterial suspension was approximately 0.80. 300 μL of bacterial suspension was evenly spread onto an MRS plate, and different drug sensitivity test strips were attached. For a 9 cm diameter plate, 3-4 drug sensitivity test strips were attached, with the center-to-center distance between each strip greater than 24 mm. After incubation at 37°C for 24 h, the diameter of the inhibition zone was measured, with each strip measured three times. Specifically, the drug sensitivity test strips used in this invention include: gentamicin, cefazolin, penicillin G, norfloxacin, ampicillin, amikacin, ciprofloxacin, erythromycin, chloramphenicol, and trimethoprim-sulfamethoxazole.

[0057] II. Experimental Results Experimental results are as follows Figure 4 As shown in Table 2, the blood plate of the Z152 strain described in this invention showed γ-hemolysis and was sensitive to all 10 tested antibiotics, indicating that it has good safety.

[0058] Table 2. Sensitive diameter and sensitivity of *Lactobacillus schreiberensis* Z152 to 10 antibiotics.

[0059] Note: Letters indicate drug resistance assessment results, S for sensitive and I for moderately sensitive; the diameter of the drug susceptibility test strips is 6.00 mm, and the results are expressed as Mean±SEM.

[0060] Example 5: Preparation of *Lactobacillus schleiferus* Z152 strain, inactivated bacterial cells, and metabiotics from Harbin, and analysis of metabiotic components. I. Experimental Methods 1. Preparation of inactivated bacterial cells and metabiotics The activated Z152 strain was inoculated into MRS liquid medium at a volume of 2% and cultured at 37°C for 24 h. The fermentation system was then inactivated by heating in a 100°C water bath for 15 minutes. After cooling to room temperature, the fermentation broth was filtered under reduced pressure to collect the bacterial cells. The filtrate was then rotary evaporated and placed in sterile glassware. The bacterial cells and filtrate were pre-cooled at -80°C for 24 h and then freeze-dried in a vacuum freeze dryer for 48 h (15 Pa, -80°C). After freeze-drying, the filtrate was stored at -20°C. Inactivated Z152 cells and metabiotics (fermentation / metabolites of *Lactobacillus schleifera* Z152 strain, hereinafter the same) were obtained.

[0061] 2. Analysis of postgenetic components The present invention also determined the obtained metagenic components. Specifically, the total sugar content of the metagenic components was determined using the sulfuric acid-phenol method, and the protein content was detected using the Bradford reagent kit according to the instruction manual.

[0062] Untargeted metabolomics analysis was performed on the postbiotics: First, the lactic acid bacteria postbiotics were slowly thawed at 4°C. A certain amount was then added to a 2:2:1 (v / v) methanol / acetonitrile / water solution, followed by vortex mixing and sonication at low temperature for 30 min. After sonication, the mixture was allowed to stand at -20°C for 10 min, and finally centrifuged at 4°C and 4000g for 20 min. The supernatant was then freeze-dried. Before mass spectrometry analysis, 100 μL of acetonitrile aqueous solution (acetonitrile:water = 1:1, v / v) was added to the dried sample and vortexed to reconstitute it. The sample was then centrifuged at 4°C and 14000g for 15 min, and the supernatant was injected for analysis. The raw data obtained after mass spectrometry were converted to .mzXML format using ProteoWizard. After conversion, peak alignment, retention time correction, and peak area extraction were performed using XCMS software. Finally, the data extracted by XCMS were used for metabolite structure identification and data processing.

[0063] II. Component Analysis Results The total sugar content of the metagenic component of *Lactobacillus schleiferus* strain Z152 from Harbin was 58.03 ± 1.26 μg / mL, and the total protein content was 55.45 ± 2.72 μg / mL. Non-target metabolomics analysis revealed that the main components of the metagenic component were phenyllactic acid, hydroxyisocaproic acid, citrate, bilirubin, 2-piperidinic acid, L-pyroglutamic acid, and D-lactic acid N-acetylglucosamine.

[0064] Example 6: Evaluation of the in vitro cholesterol-lowering activity and antioxidant properties of Z152 postbiotic. I. Experimental Methods 1. Determination of post-genetic bile salt binding rate The in vitro cholesterol-lowering activity of the postbiotic was evaluated by measuring its ability to bind sodium glycocholate, sodium taurocholate, and sodium deoxycholate solutions. Distilled water was used as a blank control, and cholestyramine as a positive control. 1.0 mL of postbiotic solutions at different concentrations (2.0, 4.0, 6.0, 8.0, and 10.0 mg / mL) and cholestyramine solutions of the same concentration were placed in test tubes. 1.0 mL of simulated gastric juice (prepared with 0.01 mol / L hydrochloric acid to form a 10 mg / mL pepsin solution) was added, and the mixture was shaken at 120 r / min at 37℃ for 1 h. The pH was then adjusted to 6.2 with NaOH solution. 4.0 mL of simulated intestinal juice (prepared with 0.1 mol / L pH 6.2 phosphate buffer to form a 10 mg / mL trypsin solution) was added, and the mixture was shaken for 1 h. Finally, 4.0 mL of 0.3 mmol / L bile salt solution was added to each sample, and the mixture was incubated at 37℃ with shaking for 1 h. The sample was then centrifuged (6000 r / min, 20 min), and 2.0 mL of the supernatant was collected. 6.0 mL of H₂SO₄ (60%, v / v) was added, and the sample was incubated in a 70℃ water bath for 20 min, followed by an ice bath for 5 min. The absorbance was measured at 387 nm. Each sample was tested in triplicate to compare the binding rate of each component to bile salts. The calculation method is as follows.

[0065]

[0066] In the formula: A0 is the absorbance at 337 nm of the blank group; A1 is the absorbance at 337 nm of the biogenic sample groups after adding different mass concentrations.

[0067] 2. The effect of metageners on cholesterol micelles The in vitro cholesterol-lowering activity of EPS produced by fermentation with different carbon sources was evaluated by measuring its effect on cholesterol micelles. Cholesterol micelle solutions were prepared according to the method described in the reference (Wang Yihan et al., 2025). Cholestyramine was used as a positive control, and distilled water as a blank control. 1.0 mL of 2, 4, 6, 8, and 10 mg / mL postbiotic solutions and cholestyramine solutions of the same concentration were added to 3.0 mL of cholesterol micelle solution, and the mixtures were incubated at 37 ℃ with shaking for 2 h. The supernatant was collected by centrifugation (6000 r / min, 20 min), and the cholesterol concentration was determined using a cholesterol kit. The cholesterol binding rate was calculated using the following formula.

[0068]

[0069] In the formula, D1 represents the cholesterol concentration in the supernatant of the blank group, mmol / L; D0 represents the cholesterol concentration in the supernatant of the sample group, mmol / L.

[0070] 3. Antioxidant activity of post-biotics The metabiotics were dissolved in primary water to prepare solutions of different concentrations (0.5 mg / mL, 1.0 mg / mL, 2.0 mg / mL, 4.0 mg / mL, and 8.0 mg / mL), with ascorbic acid (Vc) of the same concentration used as a positive control. Following the method of Wu et al. (Wu et al., 2024), the DPPH, •OH, and ABTS of the samples were analyzed. + The free radical scavenging rate was determined.

[0071] 4. Viscosity Measurement The viscosity of EPS at 5.0 mg / L was measured using a digital viscometer at a rotation speed of 60 r / min, and the viscosity value was recorded in mPa·s.

[0072] II. Experimental Results In vitro lipid-lowering activity is often assessed based on the binding rate of a substance to bile salts and cholesterol. Bile salts include sodium taurocholate, sodium glycocholate, and sodium deoxycholate, among others. Figure 5 It can be seen that the binding rate of Z152 postbiotic with sodium taurocholate, sodium glycocholate, sodium deoxycholate and cholesterol increases with increasing mass concentration, showing a clear dose-effect relationship. This indicates that Z152 postbiotic has a good ability to bind bile salts and cholesterol and has the potential to lower blood lipids.

[0073] For DPPH radicals, OH radicals and ABTS + The free radical scavenging rate is an important indicator for measuring in vitro antioxidant activity, such as... Figure 6 As shown, the scavenging rates of the Z152 metabiotic for the three free radicals all increased with increasing concentration, exhibiting a significant dose-response relationship. When the concentration exceeded 4.0 mg / mL, the metabiotic showed significant scavenging rates for DPPH, •OH, and ABTS. + The free radical scavenging rate was higher than 50%, indicating that Z152 post-biotic has good in vitro antioxidant activity.

[0074] Furthermore, the viscosity of Z152 post-biotic at a concentration of 5.0 mg / mL reached (387.5±33.2) mPa·s, indicating that this post-biotic component has the potential to act as a thickener and stabilizer.

[0075] Example 7: Evaluation of in vitro anti-allergic activity of Z152 strain, inactivated bacterial cells, and metabiotics I. Experimental Methods 1. Hyaluronidase inhibition test This invention measured the hyaluronidase activity inhibition of live and inactivated *Lactobacillus schleifera* JNDM and Z152 strains from Harbin. Following the testing method of Zheng Miao et al. (Zheng Miao et al., 2024), four test tubes (A, B, C, and D) were filled with 0.1 mL of 2.5 mmol / L calcium chloride solution. First, 0.50 mL of 800–1200 U / mL hyaluronidase solution was added to tubes A and C. Then, acetate buffer (pH 5.60) was added to tubes B and D. The tubes were then treated at 37°C for 20 min. The bacterial suspensions (OD values ​​of live and inactivated bacteria) were collected. 600nm 0.50 mL of 1.2 mg / mL sodium hyaluronate solution was added to tubes A and B, and 0.50 mL of deionized water was added to tubes C and D. The tubes were then treated at 37°C for 20 min. 0.50 mL of 0.4 mg / mL sodium hyaluronate solution was added to tubes A and C, and 0.10 mL of pH 5.6 acetate buffer solution was added to tubes B and D. The tubes were treated at 37°C for 40 min and then allowed to stand at room temperature for 10 min. Simultaneously, 0.50 mL of acetylacetone solution and 0.10 mL of 0.4 mol / mL sodium hydroxide solution were added sequentially to the four tubes, vortexed to mix, and then incubated in a boiling water bath for 15 min, followed by an ice bath for 15 min, and cooled to room temperature. Finally, 1.0 mL of Ehrlich's reagent was slowly added to the four tubes, vortexed to mix, and allowed to stand at room temperature for 30 min for color development. The absorbance of each tube was measured at 530 nm. *Lactobacillus rhamnosus* LGG was used as a control strain. The inhibition rate of hyaluronidase in the sample was calculated according to formula (1).

[0076]

[0077] In the formula: A is the absorbance of the experimental group (hyaluronidase solution + sample + sodium hyaluronate solution); B is the absorbance of the blank group (acetic acid buffer + sample + acetate buffer); C is the absorbance of the control group (hyaluronidase solution + distilled water + sodium hyaluronate solution); D is the absorbance of the blank group (acetic acid buffer + distilled water + acetate buffer).

[0078] 2. Measurement of β-HEX release during cell degranulation Following a slight modification of the method described by Zheng et al. (Zheng et al., 2024), logarithmic growth phase RBL-2H3 cells were prepared into a suspension and seeded into 48-well plates. 2.0 mL of cell suspension was added to each well, and the cell concentration was adjusted to 1 × 10⁻⁶ cells / well. 5Cells / well were collected and incubated for 24 h. DNP-IgE monoclonal antibody was added to a final concentration of 0.6 μg / mL, and the cells were incubated for 12 h. The supernatant was removed by centrifugation, and the cells were washed twice with sterile PBS. Normal, sensitized, and sample groups were established sequentially. The sample group was treated with *Lactobacillus schleifera* JNDM and *Lactobacillus schleifera* Z152 cells at a concentration of 10... 9 CFU / mL bacterial cells, inactivated strains, and 2.0 mg / mL post-biotic solution were used, with *Lactobacillus rhamnosus* LGG serving as the positive control. The remaining groups were cultured for 1 h using an equal volume of DMEM medium as a reference. Except for the normal group, all other groups were added with 0.25 μg / mL DNP-BSA and incubated for 1 h. The normal group was replaced with an equal volume of PBS buffer, and the incubation was stopped by incubating on ice for 10 min. After centrifugation, the supernatant of each culture system was collected into EP tubes, and 100 μL of cell lysis buffer was added to each well to lyse the cells, reacting for 5 min. 100 μL of supernatant and lysis buffer from each group were collected into sterile new 96-well plates, and 100 μL of chromogenic solution was added to each well. The plates were incubated for 1.5 h, and then 300 μL of stop solution was added to each well to terminate the reaction. The absorbance at 405 nm was measured, and the degranulation efficiency and degranulation inhibition rate of each group were calculated using the following formulas.

[0079]

[0080]

[0081] 3. Measurement of cytokine levels After cells were stimulated by DNP-BSA, cell supernatant was collected from each sample group, and the release of TNF-α, IL-4, IL-5 and IL-13 was detected according to the ELISA kit instructions.

[0082] II. Experimental Results The higher the inhibition rate of hyaluronidase, the stronger the anti-allergic activity. The inhibition rates of hyaluronidase in different sample groups are as follows: Figure 7 As shown in Figure A, the inhibition rates of live and inactivated *Lactobacillus schleiferus* Z152 from Harbin, as well as post-genetic hyaluronidase, were significantly higher than those of the control group, *Lactobacillus schleiferus* JNDM from Harbin. P <0.05); while the inhibition rates of live and inactivated *Lactobacillus rhamnosus* LGG cells and post-genetic hyaluronidase in the positive control group were not significantly different ( P >0.05), indicating that the Harbin Schleifer Lactobacillus Z152 strain, inactivated cells, and metabiotics all have high anti-allergic activity.

[0083] The inhibition rates of β-HEX in different sample groups are as follows: Figure 7As shown in Figure B, the inhibition rates of β-HEX were not significantly different between live and inactivated Z152 bacteria, inactivated bacteria, and metabiotics and the positive control group *Lactobacillus rhamnosus* LGG bacteria, inactivated bacteria, and metabiotics. P >0.05), but all were significantly higher than JNDM ( P <0.05), indicating that the live bacteria, inactivated bacteria and post-biotics of Z152 have strong β-HEX inhibitory activity and have the potential to become anti-allergy biological agents.

[0084] When RBL-2H3 cells are activated and degranulated, they release cytokines such as IL-4, IL-5, IL-13, and TNF-α, triggering an allergic reaction. Cytokine levels in each sample group were detected using an ELISA kit, and the results are as follows: Figure 8 As shown, compared with the model group, the levels of IL-4, IL-5, IL-13, and TNF-α cytokines in strain Z152, inactivated cells, and metabiotics were significantly lower than those in the model group. p <0.05), and also significantly lower than JNDM strain, inactivated cells and metabiotics ( p <0.05), and the levels of IL-4, IL-5, IL-13 and TNF-α cytokines in live Z152 bacteria and metagener were close to those in the normal group and the positive control group (Lactobacillus rhamnosus LGG and its metagener). These results indicate that Z152 strain and metagenerer have excellent anti-allergic activity, and their anti-allergic activity is significantly better than that of JNDM strain and its metagenerer.

[0085] Example 8: Z152 strain, inactivated bacterial cells, and their postbiotics to alleviate ovalbumin-induced food allergy. I. Experimental Methods 1. Grouping of experimental animals and establishment of experimental models Five-week-old healthy, clean-grade male BALB / c mice were selected and fed at (23±2)℃ and (50±5)% humidity, with a 12-hour day-night cycle. Mice had free access to food and water. After one week of acclimatization, the mice were randomly divided into 6 groups (n=8 mice / group). These groups included a normal diet group (ND group), an ovalbumin-induced food allergy group (OVA group), and a Z152 live bacteria intervention group (bacterial concentration 10...). 9 CFU / mL, dissolved in 0.2mL PBS solution, HLAB group), Z152 inactivated bacteria intervention group (bacterial concentration 10). 9Mice in the ND group were intraperitoneally injected with 0.2 mL PBS solution on days 0 and 14. Mice in the other groups were intraperitoneally injected with 50 μg ovalbumin and 2 mg Al(OH)3 adjuvant (dissolved in 0.2 mL PBS solution) on days 0 and 14, respectively. Except for the ND group, mice in the other groups were challenged by oral gavage with 50 mg OVA on days 28, 31, 34, 37, 40, and 43, for a total of 6 times. Mice in the ND and OVA groups were gavaged once a day at the same time in the morning with 0.2 mL of PBS, while mice in the other groups were gavaged once a day at the same time in the morning with 0.2 mL of different doses of bacterial suspension or postbiotic solution.

[0086] 2. Mouse body weight determination and evaluation of allergic symptoms The body weight and spleen weight of mice in each group were measured and recorded at days 0, 14, 28, 31, 34, 37, 40, and 43. At the last stimulation, rectal temperature was measured and recorded before and after stimulation using a rectal thermometer. Changes in body temperature before and after stimulation were detected in each group, with 5 mice from each group used for measurement. Allergic symptoms were observed and scored within 2 hours of gavage. A total of 5 levels were assigned: 0 = no symptoms; 1 = scratching nose and ears; 2 = swollen ears and eyes, rapid breathing, and lethargy; 3 = rashes on tail and mouth, accompanied by asthma and difficulty breathing; 4 = inactivity, muscle contraction, and convulsions after gavage; 5 = death and shock (Wang, et al., 2022).

[0087] 3. Detection of serum immunoglobulin and cytokine levels The collected venous blood was centrifuged at 4000 r / min for 10 min to separate the serum. The immunoglobulins in the serum included IgE, IgG1, histamine, and β-LG-specific IgE, and their levels were detected according to the ELISA kit instructions.

[0088] II. Experimental Results Table 3 shows the results of the effects of strains, inactivated bacteria, and postbiotics on allergic symptoms in mice. The weight gain in the ovalbumin-induced food allergy group (OVA group, model group) was significantly lower than that in the normal feeding group (ND group). p <0.05), after intervention with gavage of Z152 strain (HLAB), inactivated bacterial cells (MLAB), and postbiotics (Z-POS group and J-POS group), the weight gain was significantly increased ( p<0.05, among which the weight gain of the Z152 post-natal group was significantly higher than that of the JNDM post-natal group ( p <0.05).

[0089] Regarding the spleen index, the OVA group showed a significant increase compared to the ND group, while the index significantly decreased after intervention with Z152 strain, inactivated bacteria, and post-biotics. p <0.05), among which the spleen index reduction effect of the Z152 post-natal group was significantly better than that of the JNDM post-natal group ( p <0.05).

[0090] The body temperature of mice in each group was measured with a rectal thermometer 2 hours after OVA stimulation. The body temperature of mice in the OVA group decreased by 0.89±0.04℃, which was the most significant. The body temperature of mice in all intervention groups was significantly higher than that in the model group. Among them, the body temperature changes in the HLAB group tended to be similar to those in the normal group, with no significant difference. p >0.05), the increase in body temperature in the Z152 post-genetic group was significantly higher than that in the JNDM post-genetic group ( p <0.05).

[0091] The results of the allergy symptom scoring in mice showed that there was no obvious allergic reaction in the ND group. Mice in the OVA group exhibited allergic symptoms such as convulsions, nose scratching, and asthma after gavage challenge. However, the allergic symptoms were alleviated after gavage administration of Z152 strain, inactivated bacteria, and metabiotics. This indicates that Z152 strain, inactivated bacteria, and metabiotics can effectively alleviate allergic symptoms in mice, and the allergy relief effect of the Z152 metabiotic group is significantly better than that of the JNDM metabiotic group. p <0.05).

[0092] The effects of Z152 strain, inactivated bacterial cells, and their postbiotics on immunoglobulin levels in allergic mice are as follows: Figure 9 As shown, compared with the ND group, the levels of IgE, histamine, β-LG-specific IgE, and IgG1 in the OVA group were significantly increased, while those after intervention with Z152 strain, inactivated bacteria, and metabiotics were significantly decreased. p <0.05), and the intervention effects of strain Z152 and metabiotics were significantly better than those of inactivated bacteria and JNDM metabiotics ( p <0.05). This indicates that strain Z152, inactivated bacterial cells, and metabiotics all have the effect of alleviating allergic symptoms in mice.

[0093] Table 3. Z152 strain, inactivated bacterial cells, and postbiotics alleviate allergic symptoms in mice.

[0094] Note: Different letters following the data in the same column of the table indicate significant differences between treatments. P <0.05); results are expressed as mean ± standard deviation.

Claims

1. A strain of *Lactobacillus schleifera* from Harbin ( Schleiferilactobacillus harbinensis Z152 strain, characterized in that, The strain was deposited at the Guangdong Provincial Microbial Culture Collection Center (GDMCC) on May 8, 2026, with accession number GDMCC No: 68230.

2. A metabiotic prepared using the *Lactobacillus schleiferus* Z152 strain from Harbin as described in claim 1, characterized in that... The metabiotic includes one or more of the following: fermentation / metabolites of *Lactobacillus schreiformis* Z152 strain, abiotic *Lactobacillus schreiformis* Z152 strain, and lysates of *Lactobacillus schreiformis* Z152 strain.

3. The epigenetic agent according to claim 2, characterized in that, The method for preparing the post-genetic agent is to inoculate the Harbin Schleifer Lactobacillus Z152 strain as described in claim 1 into a culture medium, culture it, sterilize it, and filter it to obtain the product.

4. The use of the Harbin Schleifer Lactobacillus Z152 strain as described in claim 1 or the metabiotic as described in claim 2 or 3 in the preparation of a drug for relieving / preventing food allergies and / or lowering blood lipids.

5. The application according to claim 4, characterized in that, The drug achieves relief / prevention by inhibiting hyaluronidase and β-HEX.

6. The application according to claim 4, characterized in that, The drug achieves relief / prevention by lowering and downregulating the levels of cytokines IL-4, IL-5, IL-13, and TNF-α.

7. The application according to claim 4, characterized in that, The drug achieves relief / prevention by downregulating serum levels of immunoglobulin IgE, IgG1, histamine, and β-LG-specific IgE.

8. The application according to claim 4, characterized in that, The food allergy mentioned is a food allergy induced by ovalbumin.

9. A drug, characterized in that, The drug comprises the Harbin Schleifer Lactobacillus Z152 strain as described in claim 1 or the metabiotic as described in claim 2 or 3.

10. The drug according to claim 9, characterized in that, The drug may also contain other pharmaceutically acceptable carriers.

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