Application of lactobacillus rhamnosus SSN-13 in preparation of anti-glycosylation and anti-aging drugs

Abstract

The invention discloses an application of rhamnolactobacilli SSN-13 in preparation of anti-glycosylation and anti-aging drugs, and is characterized in that the preservation number of the rhamnolactobacilli SSN-13 is CGMCC No: 27227, and the application of the rhamnolactobacilli SSN-13 in preparation of preparations for improving degenerative changes of skin, intestinal tracts, liver and brain tissues and cognitive function decline caused by individual aging is provided, and the application of the rhamnolactobacilli SSN-13 in preparation of anti-glycosylation and anti-aging drugs in preparation of anti-glycosylation and anti-aging drugs in preparation of anti-glycosylation and anti-aging drugs in preparation of anti-glycosylation and anti-aging drugs. According to the present invention, the application in the preparation of the preparation for improving the composition and the structure of the aging individual intestinal flora has the advantages of significantly improving the inhibition ability of glycosylation reaction products AGEs, pentosan, fructosamine and dityrosine, reducing the levels of IL-1beta, IL-6, TNF-alpha, 8-OHdG and AGEs in serum, improving the enzymatic activity of SOD and GSH-Px, improving the HA content and the moisture content in skin, reducing the MDA content, and improving the cognitive ability.

Description

TECHNICAL FIELD

[0001] The present application relates to a lactobacillus rhamnosus, in particular to the application of lactobacillus rhamnosus SSN-13 in the preparation of anti-glycosylation and anti-aging drugs. BACKGROUND

[0002] Aging is a biological process accompanied by the decline of physiological integrity of many tissues and the loss of regenerative capacity, which is the result of the combined effects of excessive oxidative stress damage, stem cell decline, DNA degradation, dietary factors, and active aging groups, and can lead to an increase in morbidity and mortality. At present, more and more anti-aging drugs and related research are carried out, which plays a positive role in promoting human health, preventing aging-related diseases, and improving host metabolism. Global aging intensifies (WHO predicts that the proportion of the elderly population will reach 22% in 2050) and metabolic diseases (liver damage and glycosylation damage) become the core pathological axis that synergistically accelerates aging, forming a superimposed effect, and the core pathological correlation lies in the imbalance of the liver-sugar metabolism axis, which drives systemic aging through pathways such as oxidative stress and AGEs accumulation. Current intervention methods have multiple limitations, such as probiotic therapy, which is limited by low survival rate of live bacteria in gastric acid and uncertainty of colonization; synthetic drugs only target a single point and cannot block AGEs accumulation to cause collagen cross-linking and inflammatory response; and the bioavailability of plant and medicinal active ingredients is insufficient.

[0003] Postbiotics are microecological preparations made of non-living microorganism cells and / or their components that are beneficial to the health of the host, and their metabolites. Compared with probiotics composed of active cells, postbiotics are safer, more stable, more effective, and more convenient and economical to prepare, store, and transport. Postbiotic products do not contain live bacteria and do not need to be refrigerated, making them easy to carry. In addition, postbiotics are not inhibited by antibiotics and can greatly reduce the risk of infection. Using postbiotics can achieve similar effects to live probiotics, while avoiding problems such as low bioavailability of live bacteria, unstable effects, and easy transmission of drug resistance genes. The inactivation process can also kill a small amount of possible impurities, making postbiotics safer and not limited by food form, which can be taken internally or externally. Although the process of aging is irreversible, it can be intervened by appropriate means and the related metabolic diseases caused thereby can be prevented. With the development of the health industry, there is an increasing demand for functional anti-aging products, which can prevent and treat various diseases by improving the body's resistance to disease through the intake of some antioxidants. Using topical antioxidants can overcome some of these effects and delay aging, but there are limitations such as low stability, high production cost, and side effects. Therefore, it is urgent to develop a safe and efficient anti-aging lactic acid bacteria and its postbiotic product to improve health through food. SUMMARY

[0004] The technical problem to be solved by the present application is to provide a rhamnolactic casein bacillus SSN-13 for preparing an anti-glycosylation and anti-aging drug, which can improve the inhibitory capacity of glycosylation products AGEs, pentosidine, fructosamine and di-tyrosine, reduce the beta-galactosidase activity of Hacat cells of aging skin, regulate the abnormal expression of aging-related genes of HepG2 cells in liver MAPK1 、 PSEN1 、 Sirt1 、 Collagen Ⅲ , reduce the levels of IL-1beta, IL-6, TNF-alpha, 8-OHdG and AGEs in serum, improve the enzyme activities of SOD and GSH-Px, reduce the MDA content in skin, and improve the skin, intestinal, liver and brain tissue degradation and cognitive learning ability decline caused by aging of individuals.

[0005] The technical solution adopted by the present application to solve the above technical problem is that the rhamnolactic casein bacillus SSN-13 is used for preparing an anti-glycosylation and anti-aging drug, and the preservation number of the rhamnolactic casein bacillus SSN-13 is CGMCC No: 27227.

[0006] Further, the rhamnolactic casein bacillus SSN-13 is used for preparing a preparation for inhibiting the generation of fluorescent AGEs, fructosamine, di-tyrosine and pentosidine.

[0007] Further, the rhamnolactic casein bacillus SSN-13 inhibits the increase of beta-galactosidase activity of human immortalized keratinocytes (Hacat) under oxidative damage conditions.

[0008] Further, the rhamnolactic casein bacillus SSN-13 regulates the abnormal mRNA expression of key target points MAPK1 、 PSEN1 、 Sirt1 、 Collagen Ⅲ for regulating the growth of human liver cancer cells (HepG2) under oxidative stress and glycosylation damage conditions.

[0009] Further, the rhamnolactic casein bacillus SSN-13 is used for preparing a preparation for improving the skin, intestinal, liver and brain tissue degenerative changes and cognitive function decline caused by aging of individuals.

[0010] Further, the rhamnolactic casein bacillus SSN-13 is used for preparing a preparation for improving the composition and structure of intestinal flora of aging individuals.

[0011] Further, the rhamnolactic casein bacillus SSN-13 is used for preparing a preparation for reducing the AGEs content and the contents of inflammatory markers IL-6, IL-1beta and TNF-alpha in serum of aging individuals.

[0012] Furthermore, the application of *Lactobacillus rhamnosus* SSN-13 in the preparation of formulations that reduce the decrease in hyaluronic acid content in the skin of aging individuals caused by glycation damage.

[0013] Furthermore, the viable count of the *Lactobacillus rhamnosus* SSN-13 prepared was 1 × 10⁻⁶. 7 ~1×10 9 CFU / mL of probiotic preparations or bacterial cell concentration of 1×10 7 ~1×10 9 A CFU / mL postbiotic formulation, wherein the postbiotic comprises lactic acid bacteria cell metabolites.

[0014] Furthermore, the application of the aforementioned microecological preparations or postbiotic preparations in the preparation of food, health products, cosmetics, or therapeutic drugs.

[0015] Compared with the prior art, the advantages of this invention are as follows: This invention discloses for the first time the application of *Lactobacillus rhamnosus* SSN-13 in the preparation of anti-glycation and anti-aging drugs. The active ingredient of the probiotic microecological preparation includes the above-mentioned *Lactobacillus rhamnosus* SSN-13 cells, and the lactic acid bacteria postbiotic preparation is obtained by fermentation, heat inactivation, filtration sterilization, and freeze-drying of the above-mentioned *Lactobacillus rhamnosus* SSN-13. It can significantly improve the inhibitory ability of glycosylation products AGEs, pentosaccharides, fructosamine, and dityrosine, reduce the β-galactosidase activity of senescent Hacat cells, and regulate senescence-related genes in senescent HepG2 cells. MAPK1 , PSEN1 , Sirt1 , Collagen Ⅲ The expression of the substance and the results of the tissue section showed that it had a significant effect on improving the degenerative changes of the intestine, skin and brain organs caused by aging. At the same time, it effectively reduced the levels of IL-1β, IL-6, TNF-α, 8-OHdG and AGEs in mouse serum, increased the enzyme activities of SOD and GSH-Px, as well as the HA and water content in the skin, reduced the MDA content, improved intestinal flora disorder, reduced the Firmicutes / Bacteroidetes ratio, and alleviated anxiety-like behavior and cognitive impairment caused by aging in mice. It has a comprehensive anti-aging effect on mice with D-galactose-induced accumulation of glycation end products, filling the gap in comprehensive anti-aging functional products and overcoming the defects of traditional special drugs such as large side effects and low bioavailability.

[0016] The above-mentioned Lactobacillus rhamnosus ( Lacticaseibacillus rhamnosus Strain SSN-13, with accession number CGMCC No: 27227, was deposited on April 27, 2023, at the China General Microbiological Culture Collection Center, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing. Attached Figure Description

[0017] Figure 1 Inhibition rate of AGEs, fructosamine, pentosidine and di-tyrosine by SSN-13 and its postbiotic; Figure 2 Growth curve and acid production curve of SSN-13; Figure 3 Adhesion ability evaluation of SSN-13; Figure 4 Phylogenetic tree of SSN-13; Figure 5 Effect of SSN-13 postbiotic on the staining level of β-galactosidase in senescent human immortalized keratinocytes (Hacat); Figure 6 Effect of SSN-13 postbiotic on the mRNA expression level of senescence-related genes in senescent human hepatocarcinoma cells (HepG2) (A) MAPK1 ; (B) PSEN1 ; (C) Sirt1 ; (D) Collagen Ⅲ ; Figure 7 HE staining of mouse skin tissue; Figure 8 HE staining of mouse colon tissue; Figure 9 HE staining of mouse liver tissue; Figure 10 HE staining of mouse brain tissue hippocampus; Figure 11 Effect of SSN-13 microecological preparation and its postbiotic preparation on the relative abundance of species at the phylum level of intestinal flora; Figure 12 Effect of SSN-13 microecological preparation and its postbiotic preparation on the relative abundance of species at the genus level of intestinal flora; Figure 13 Open field movement trajectory (A), total movement distance (B), and center area movement time (C) of mice; Figure 14 Serum IL-1β (A), IL-6 (B), TNF-α (C), 8-OHdG (D), and AGEs (E) levels of mice; Figure 15 SOD activity (A), GSH-Px activity (B), and MDA content (C) of mouse serum; Figure 16 Effect of SSN group and SSN postbiotic group on the hyaluronic acid content (A) and water content (B) of the skin of aging mice; Note: CONT group: blank group; MOD group: aging model group; SSN group: rhamnolactic lactobacillus SSN-13 group; SSNH: rhamnolactic lactobacillus SSN-13 probiotic high-dose group; SSNM: rhamnolactic lactobacillus SSN-13 probiotic medium-dose group; SSNL: rhamnolactic lactobacillus SSN-13 probiotic low-dose group. P <0.05, P <0.01, P <0.001, P <0.0001. DETAILED DESCRIPTION

[0018] The application will be further described in detail below with reference to the embodiments combined with the accompanying drawings.

[0019] I. Experimental methods 1. Preparation of culture medium and solution Complete cell culture medium: 9% fetal bovine serum (FBS), 90% DMEM medium, 1% penicillin-streptomycin double antibody.

[0020] MRS broth: 10.0 g / L proteose peptone, 8.0 g / L beef extract, 4.0 g / L yeast extract, 20.0 g / L glucose, 2.0 g / L potassium phosphate dibasic, 2.0 g / L ammonium citrate dibasic, 5.0 g / L sodium acetate, 0.2 g / L magnesium sulfate, 0.04 g / L manganese sulfate, 1.0 g / L Tween 80; prepared according to the proportion of 52.24 g stirred and dissolved in 1 L distilled water, 121 ℃ high pressure sterilization for 15 min.

[0021] MRS agar: formula is 10.0 g / L proteose peptone, 8.0 g / L beef extract, 4.0 g / L yeast extract, 20.0 g / L glucose, 2.0 g / L potassium phosphate dibasic, 2.0 g / L ammonium citrate dibasic, 5.0 g / L sodium acetate, 0.2 g / L magnesium sulfate, 0.04 g / L manganese sulfate, 14.0 g / L agar, 1.0 g / L Tween 80; prepared according to the proportion of 66.2 g stirred and dissolved in 1 L distilled water, 121 ℃ high pressure sterilization for 15 min.

[0022] The fermentation medium is dissolved with 1 L distilled water, sterilized at 121 ℃ for 15 min, and the formula of the MRS solid medium is 10 g proteose peptone, 10 g beef extract, 5 g yeast extract, 2 g diammonium citrate, 5 g sodium acetate, 20 g glucose, 80 mL Tween 80, 0.5 g magnesium sulfate, 0.25 g manganese sulfate and 15 g agar powder.

[0023] 2. Activation and preservation of strains The strain cryopreserved in -80 °C refrigerator was inoculated in MRS broth medium and incubated at 37 °C for 18 h to obtain seed liquid. The seed liquid was diluted and spread on MRS agar medium, and continuously cultured for 2 generations until single colonies appeared. The colony morphology was observed and inverted in 4 °C refrigerator for standby.

[0024] (1) Lactic acid bacteria culture solution: the single colony of lactic acid bacteria preserved above was inoculated in MRS broth medium and incubated at 37 °C for 18 h to activate for three generations to obtain lactic acid bacteria culture, and the bacterial concentration was adjusted to 1×10 9 CFU / mL. The bacterial solution was stored at 4 °C for standby.

[0025] (2) Lactic acid bacteria postbiotic sample solution: the lactic acid bacteria culture above was inactivated by heating in 100 °C water bath for 25 min, centrifuged at 4000 rpm for 15 min, and the supernatant was collected after sterilization through 0.22 μm water filter membrane in a clean bench to obtain postbiotic sample solution, which was stored in -80 °C refrigerator.

[0026] 3. Determination of anti-glycosylation ability of lactic acid bacteria and its postbiotics (1) Establishment of BSA-Fru (bovine serum albumin-fructose) simulation system BSA (bovine serum albumin) and Fru (fructose) were weighed and dissolved in 3 mL phosphate buffer solution (0.01 M, pH 7.4) to obtain a final concentration of 15 mg / mL and 0.5 M, respectively. Then, penicillin-streptomycin was added to a final concentration of 1% (v / v). In each experimental group, 3 mL of lactic acid bacteria culture solution and lactic acid bacteria postbiotic sample solution prepared in the above-mentioned experimental method 2 were added. The experimental group without adding fructose, lactic acid bacteria and postbiotics was added with the same amount of sterile water as a blank control group. The positive control group was added with the same amount of aminoguanidine with a concentration of 10 mmol / L. Each group was placed in a 65 °C water bath for 24 h in the dark, and then stored at -20 °C for standby.

[0027] (2) Inhibitory effect of lactic acid bacteria and its postbiotics on fructosamine The content of fructosamine, an early glycosylation product, was determined by NBT reduction method. The glycosylated BSA sample (40 μL) in 3(1) above was mixed with 800 μL NBT color developing solution (0.3 mM, dissolved in 0.1 M sodium carbonate buffer) and 160 μL ultrapure water, and incubated at 25 °C for 15 min. The absorbance at 530 nm was determined by ultraviolet spectrophotometer, and the inhibition rate of fructosamine generation was calculated as follows: , wherein A0 is the absorbance of the blank control group, and A is the absorbance of the experimental group.

[0028] (3) Inhibition of AGEs by lactic acid bacteria and postbiotics The fluorescence intensity of each sample was measured by setting the excitation (ex) and emission (em) wavelengths at 360 nm and 460 nm, respectively, and the fluorescence AGEs generation inhibition rate was calculated: where F0is the fluorescence intensity of the blank control group and F is the fluorescence intensity of the experimental group.

[0029] (4) Inhibition of di-tyrosine by lactic acid bacteria and postbiotics The fluorescence intensity of each sample was measured by setting the ex and em wavelengths at 330 nm and 415 nm, respectively, and the di-tyrosine generation inhibition rate was calculated: where F0is the fluorescence intensity of the blank control group and F is the fluorescence intensity of the experimental group.

[0030] (5) Inhibition of pentosidine by lactic acid bacteria and postbiotics The fluorescence intensity of each sample was measured by setting the ex and em wavelengths at 330 nm and 415 nm, respectively, and the pentosidine generation inhibition rate was calculated: where F0is the fluorescence intensity of the blank control group and F is the fluorescence intensity of the experimental group.

[0031] 4. Growth curve and acid production curve of lactic acid bacteria The lactic acid bacteria were cultured in MRS broth medium and inoculated into MRS broth liquid medium at a 2 vt% inoculation amount. The bacterial solution was cultured in a 37°C constant temperature biochemical incubator for 24 h, and the absorbance of the bacterial solution at 600 nm and the pH value of the fermentation broth were measured every 2 h. The average value was taken after parallel measurement for 3 times, and the curve was drawn.

[0032] 5. Self-aggregation and surface hydrophobicity of lactic acid bacteria The bacterial solution was inoculated into MRS broth medium at a 3% inoculation amount and cultured for 24 h. The bacterial cells were collected by centrifugation at 4°C and 5000 r / min for 10 min, washed twice with PBS, and resuspended in PBS. The absorbance of the bacterial suspension at 600 nm was measured, and the OD value of the bacterial suspension was adjusted to be in the range of 0.25±0.05. The initial OD was recorded as A0. 4 ml of the bacterial suspension was transferred into an EP tube, and after standing at 37°C for 20 h, 3 mL of the supernatant was taken and added to another centrifuge tube. The OD 600 was measured, which was recorded as A1. The self-aggregation rate was calculated according to the following formula. The independent test was repeated for 3 times, and the average value was taken.

[0033] .

[0034] Prepare a bacterial suspension as described above. Add 1 mL of xylene to 3 mL of the bacterial suspension, vortex to mix, and let stand at 37°C for 1 h. Take the aqueous phase and measure its absorbance at 600 nm, denoted as A. Calculate the hydrophobicity of the strain using the following formula. Repeat the independent experiment three times and take the average value.

[0035] .

[0036] 6. Antibiotic susceptibility testing of lactic acid bacteria Antimicrobial susceptibility testing was conducted using the double-layer soft agar diffusion method with discs to assess the resistance of lactic acid bacteria to 10 commonly used clinical antibiotics. 10 mL of MRS solid medium was poured into a petri dish and allowed to solidify. Then, soft agar containing 1% lactic acid bacteria inoculum was poured in and allowed to solidify again. Antimicrobial susceptibility discs were then affixed to the surface, grouped, numbered, and recorded. The discs used were for: amikacin, neomycin, tetracycline, erythromycin, vancomycin, polymyxin B, ciprofloxacin, norfloxacin, penicillin, and rifampin. The plates with the discs affixed were incubated at 37°C for 48 h, and the size of the inhibition zone produced by each disc was measured and recorded.

[0037] 7. Identification and evolutionary analysis of 16S rDNA of lactic acid bacteria (1) DNA extraction Take fresh bacterial culture, centrifuge at 8000 rpm for 10 min to obtain bacterial cells, and extract total DNA from the strain using a kit.

[0038] (2) PCR amplification and product detection The 16S rRNA gene was amplified using universal primers. The primer sequences are as follows: Upstream primer 27F: 5′-AGAGTTTGATCCTGGCTCAG-3′ Downstream primer 1495R: 5′-CTACGGCTACCTTGTTACGA-3′, The PCR cycling parameters were: 94℃ pre-denaturation for 5 min, 94℃ denaturation for 1 min, 64℃ annealing for 1 min, 72℃ extension for 2 min, and 4℃ final extension for 10 min, for 34 cycles. The amplified PCR products were subjected to nucleic acid electrophoresis, and the products were sent to Sangon Biotech Co., Ltd. for gene sequencing.

[0039] (3) Phylogenetic tree The obtained strain sequence was subjected to 16S rDNA gene fragment alignment on the NCBI website, the 16S rDNA gene sequences of strains with high similarity were extracted, a phylogenetic tree was constructed using software MEGAX 64, and a map was drawn according to the results.

[0040] 8. Lactobacillus rhamnosus SSN-13 probiotic and senescent cells co-culture (1) Hacat cell culture: 2 mM H2O2 solution prepared with complete culture medium as the solution was used as the senescence modeling solution, and Hacat cells were seeded in a 6-well plate at a density of 3×10 6 cells / mL, incubated for 24 h, then the culture medium was discarded, the cells were washed with 1 mL PBS buffer, then 1 mL complete culture medium containing the modeling solution was added, 200 μL of lactobacillus probiotic sample solution was added, and incubation was carried out at 37°C for 24-48 h.

[0041] (2) HepG2 cell culture: 300 mM D-gal solution prepared with complete culture medium as the solution was used as the senescence modeling solution, and HepG2 cells were seeded in a 6-well plate at a density of 1×10 5 cells / mL, incubated for 24 h, then the culture medium was discarded, the cells were washed with 1 mL PBS buffer, then 1 mL complete culture medium containing the modeling solution was added, 200 μL of lactobacillus probiotic sample solution was added, and incubation was carried out at 37°C for 24 h.

[0042] (3) β-galactosidase activity determination After Hacat cell culture was completed, the cell culture solution was carefully aspirated, washed once with PBS, 1 mL of β-galactosidase staining and fixing solution was added, and after 15 min of room temperature fixing, the cell fixing solution was aspirated, the cells were washed with PBS 3 times, each time for 3 min, the PBS was aspirated, 1 mL of staining working solution was added to each well, and incubation was carried out overnight in a 37°C incubator.

[0043] (4) Senescence-related gene expression After HepG2 cell culture was completed, the cell culture solution was carefully aspirated, washed once with PBS, the cells were collected to extract RNA, and reverse transcription was carried out in time to obtain a cDNA sample, which was reacted in an RT-qPCR system, and the relative expression level of each target gene was calculated using the 2 -△△CT method, and each group was repeated 3 times. The RT-qPCR reaction system was: cDNA sample 2 μL, upstream primer 0.5 μL, downstream primer 0.5 μL, 2×Taq Pro Universal SYBR qPCR Master Mix enzyme 5 μL, and ddH2O 2 μL. The fluorescence quantitative primer sequences of the target genes are shown in the following table.

[0044] Table 1. Fluorescence quantitative primer table of aging-related genes

[0045] 9. Data analysis The data obtained in this study were statistically analyzed and plotted using GraphPad Prism 9.4.0 and SPSS 19.0. Multiple comparisons and significance analysis were performed using one-way ANOVA and Tukey's test.

[0046] II. Functional analysis of lactic acid bacteria 1. Anti-glycation ability of lactic acid bacteria and postbiotics During the growth and metabolism of the body, a large number of free radicals accumulate in the body, excessive oxidative stress is damaged, and glycation occurs, all of which can lead to cell function decline and degenerative changes. Various active substances produced by lactic acid bacteria fermentation, such as lactic acid, exopolysaccharides, and bacteriocins, can play an anti-aging role through the pathways of free radical scavenging, regulation of cell metabolism, and improvement of intestinal barrier function, effectively improving the overall metabolic function of the host and helping to restore the balance of the host homeostasis. In the application of lactic acid bacteria and postbiotics in anti-aging research, anti-glycation and anti-oxidation can be used as criteria for screening lactic acid bacteria with potential anti-aging efficacy. In this study, lactic acid bacteria strains were selected from the Ningbo University Laboratory of Animal Products Processing strain preservation library, and the anti-glycation ability of lactic acid bacteria and postbiotics was detected to determine a strain of lactic acid bacteria with potential anti-aging efficacy.

[0047] From Figure 1 It was found that the inhibition rates of lactic acid bacteria SSN-13 on AGEs, pentosidine, fructosamine, and di-tyrosine in the BSA-Fru glycosylation reaction system were 35.8%, 36.25%, 47.56%, and 35.58%, respectively. The inhibition rates of SSN-13 postbiotics on the above indicators were 42.35%, 33.86%, 50.61%, and 37.42%, respectively. The inhibition rates of postbiotics on AGEs, fructosamine, and di-tyrosine were higher than those of lactic acid bacteria SSN-13, close to the 51.27%, 64.85%, and 42.25% of the positive control aminoguanidine group, which could reflect the glycosylation inhibition ability of SSN-13 and its postbiotics at each stage of the BSA-Fru system, especially in the early stage of the reaction to reduce the generation of fructosamine.

[0048] 2. Growth curve and acid production curve By drawing the growth curve, the growth state of lactic acid bacteria at different stages of cultivation can be determined, which helps to determine the optimal cultivation time of lactic acid bacteria and provides experimental basis for the production of microecological preparations. The acid production curve can reflect the ability and dynamic changes of lactic acid bacteria in the growth process, and can be used to evaluate the acid production ability of lactic acid bacteria strains at different stages.

[0049] As shown in Figure 2 , under the condition of 37℃ incubation temperature, the strains entered the logarithmic phase after 2 h and entered the stationary phase after 14 h; the initial pH value of SSN-13 fermentation broth was about 5.40, which rapidly decreased to about 4.00 within 2-14 h, and then slowly decreased, and tended to be flat after 18 h of inoculation; the experimental results showed that the strains had good acid-producing capacity, and could quickly enter the logarithmic phase, so that the pH value rapidly decreased, which had a good effect on inhibiting the growth of pathogenic bacteria.

[0050] 3. Self-aggregation ability and surface hydrophobicity determination Self-aggregation refers to the ability of lactic acid bacteria to aggregate with each other to form clumps, and surface hydrophobicity is one of the key factors driving self-aggregation. Hydrophobic groups (such as lipoteichoic acid) promote bacterial adhesion between bacteria through hydrophobic interaction. Lactic acid bacteria with appropriate hydrophobicity can better bind to epithelial cells, other microbial cells, etc. with specificity or non-specificity, which is conducive to the colonization of lactic acid bacteria in the human body. The self-aggregation of lactic acid bacteria is about 16-35%, which is low self-aggregation ability, 36-50% is moderate self-aggregation ability, and more than 51% is high self-aggregation ability. According to Figure 3 , the self-aggregation ability of lactic acid bacteria SSN-13 is 52.56%, and the hydrophobicity is 38.2%.

[0051] 4. Antibiotic sensitivity As probiotic preparations (such as yogurt, probiotic supplements) or fermentation agents, lactic acid bacteria may transfer drug-resistant genes to pathogenic bacteria through the food chain or intestinal environment, exacerbating the global drug resistance crisis. Drug sensitivity test can screen for drug-resistant strains to ensure the biological safety of the strain.

[0052] According to the results in Table 2, SSN-13 is highly sensitive to tetracycline, erythromycin, and penicillin, moderately sensitive to lincomycin, and resistant to amikacin, neomycin, vancomycin, polymyxin B, ciprofloxacin, and norfloxacin.

[0053] Table 2. Results of lactic acid bacteria drug sensitivity test

[0054] Note: S-susceptible, highly sensitive; I-intermediate, moderately sensitive; R-resistance, drug resistance.

[0055] 5. Lactic acid bacteria strain identification results Through 16S rDNA sequencing identification, the phylogenetic tree of strain SSN-13 is shown in Figure 4 , and the results show that the strain SSN-13 is Lactobacillus rhamnosusLacticaseibacillus rhamnosus The growth curves and acid production curves showed that the strains could grow rapidly and produce acid stably, entered the exponential growth phase after 2 h and had a long stationary phase, indicating good growth stability. The strains had high self-aggregation ability and were moderately hydrophobic. The in vitro safety evaluation showed that the strains met the safety evaluation conditions and could be used to prepare probiotic microecological preparations.

[0056] Effect of Lactic Acid Bacteria Bioinoculant on β-galactosidase Activity of Senescent Hacat Cells Cellular senescence is a phenomenon of functional decline and gradual death of cells under normal environmental conditions. Although senescent cells grow irreversibly, they still have metabolic functions. The characteristics of senescent cells include cell volume enlargement, flat shape, large nucleus, nuclear membrane invagination, chromatin aggregation, pyknosis, and lysis, increased intracellular granules, and vacuole formation. The number and shape of mitochondria change, membrane fluidity decreases, and lysosomal content increases, leading to increased β-galactosidase activity. SA-β-Gal is a reliable biomarker for detecting senescent cells. Based on the up-regulation of senescence-associated β-galactosidase activity during cell aging, a deep blue-green product is generated, and senescent cells or tissues are stained.

[0057] From Figure 5 As shown in (B), senescent Hacat cells had strong SA-β-Gal staining and abnormal morphological changes, such as increased volume, flattening, and increased granules. Figure 5 The data in (D) showed that the SA-β-Gal staining rate of senescent cells in the model group reached 69.31%, indicating the occurrence of senescence. Figure 5 (A) and (C) reflect that the overall cells in the SSN-13 bioinoculant treatment group were more similar to the normal control group, with 25.36% and 30.04%, respectively. Although senescent cells still existed, there was a highly significant difference between the treatment group and the model group (P<0.0001), indicating that the intervention of R. rhamnosus SSN-13 bioinoculant could improve cell senescence to some extent. P

[0058] Effect of Lactic Acid Bacteria Bioinoculant on Expression of Senescence Genes in HepG2 Cells Genes MAPK1 , Sirt1 , PSEN1 and Collagen Ⅲ Maintain body homeostasis from the dimensions of signal transduction, epigenetic regulation, protein processing, and structural support. Functional imbalance points to aging and degenerative diseases. During the aging process of HepG2 cells, the expression of genes changes. Understanding these changes helps to explore the molecular mechanisms of cell aging. Figure 6 (A), Figure 6 (B), Figure 6 ​(C) and Figure 6 (D) It can be seen that, compared with senescent cells damaged by oxidation, the SSN-13 post-biotic can significantly downregulate the gene encoding progerin. PSEN1 The expression level of [the substance] was 0.975, significantly lower than that of the senescent cell group (1.72). P <0.001), significantly upregulated type III collagen gene Collagen Ⅲ Longevity-related genes Sirt1 , MAPK1 The mRNA expression levels were 2.51, 1.28, and 1.64, respectively, indicating that Lactobacillus rhamnosus SSN-13 postbiotic can slow down the process of cellular senescence by altering the expression of multiple senescence-related genes.

[0059] III. Product Application Examples 1. Specific Implementation Examples Example 1 A probiotic strain with anti-glycation and anti-aging effects has been classified and named *Lactobacillus rhamnosus*. Lacticaseibacillus rhamnosus SSN-13 was deposited on April 27, 2023, at the China General Microbiological Culture Collection Center (CGMCC), with accession number CGMCC No: 27227.

[0060] Example 2 A method for preparing a probiotic microecological preparation with anti-glycation and anti-aging effects includes the following steps: (1) Lactic acid bacteria fermentation: Lactobacillus rhamnosus SSN-13 was inoculated into the fermentation medium at a volume ratio of 3% and cultured at 37℃ for 18 h to obtain lactic acid bacteria culture; (2) Freeze-drying: Centrifuge the obtained lactic acid bacteria culture to collect the lactic acid bacteria pellet, and adjust the bacterial concentration to 1×10⁻⁶ using sterile PBS buffer. 9 Add CFU / mL of lyophilization protectant to an equal volume of sterile PBS buffer, mix well, and pre-freeze overnight at -80℃. Then freeze-dry the pre-frozen sample at -50℃ and 0.01 mbar for 48 h to obtain the lactic acid bacteria microecological preparation. The lyophilization protectant formulation consists of 118.2 g / L trehalose, 17.1 g / L L-cysteine, 10.3 g / L sorbitol, 1.7 g / L sodium acetate, and 120 g / L skim milk powder.

[0061] (3) Administration method: oral gavage, bacterial concentration of 1×10 9 CFU / mL, named Lactobacillus rhamnosus SSN-13 microecological preparation group (SSN group).

[0062] Example 3 A preparation method of a lactic acid bacteria postbiotic with anti-glycosylation and anti-aging effects, comprising the following steps: The lactic acid bacteria postbiotic: the lactic acid bacteria cell concentration is adjusted to 1×10 9 CFU / mL, inactivated by heating in a 100℃ water bath for 25min, centrifuged at 4000 rpm for 15min, and the supernatant was collected, sterilized by 0.22 μm water filter membrane in a clean bench, and then the liquid was collected to obtain the postbiotic sample liquid; Freeze-drying: the postbiotic sample liquid obtained in (1) is pre-frozen in a -80℃ refrigerator, and the pre-frozen sample is freeze-dried at -50℃ and 0.01mbar for 48h to obtain a lactic acid bacteria postbiotic preparation; Dosage form: oral gavage, concentration of 200 mg / kg / d per mouse, named SSN postbiotic high-dose group (SSNH).

[0063] Example 4 The same as Example 3 above, except that the lactic acid bacteria cell concentration in step (1) is 1×10 8 CFU / mL, and the concentration in step (3) is 100 mg / kg / d per mouse, named SSN postbiotic medium-dose group (SSNM).

[0064] Example 5 The same as Example 3 above, except that the lactic acid bacteria cell concentration in step (1) is 1×10 7 CFU / mL, and the concentration in step (3) is 50 mg / kg / d per mouse, named SSN postbiotic low-dose group (SSNL).

[0065] 2. Improvement effect of lactic acid bacteria and its postbiotic preparation on aging mice (1) Experimental materials: SPF C57BL / 6J mice, male, 60 in total, 6-8 weeks old, weighing about 20.00 ± 2 g per mouse, were purchased from Zhejiang Weitong Lihua Experimental Animal Technology Co., Ltd. The mice were sent to the Ningbo University Experimental Animal Center, and the feeding environment was barrier level, with license information: SYXK (Zhe) 2024-0002. The experimental plan has been approved by the Ningbo University Animal Experiment Ethics Committee (approval number: 14026), in line with the principles of animal protection and relevant regulations of the national experimental animal experiment ethics, and all experimental operations were strictly in accordance with the internationally effective guidelines and procedures.

[0066] (2) Experimental plan The mice were sent to the animal experiment center barrier for 7-day adaptive culture, the temperature was 25±1℃, the humidity was 25±1%, the light cycle was 12 h light, 12 h dark, and the animals could freely take water and food in their respective cages. After the adaptation period, 60 mice were randomly divided into groups according to the body weight, 10 mice in each group, and the grouping information was: blank control group (CONT group), aging model group (MOD), Lactobacillus rhamnosus SSN-13 group (SSN), Lactobacillus rhamnosus SSN-13 postbiotic high-dose group (SSNH group), Lactobacillus rhamnosus postbiotic medium-dose group (SSNM group), and Lactobacillus rhamnosus postbiotic low-dose group (SSNL group). From the next day of grouping, the CONT group was injected with 0.2 mL of normal saline at a relatively fixed time every day, and the rest of the groups of mice were injected with 5% D-galactose 0.2 mL at a dose of 600 mg / kg / d subcutaneously on the nape to simulate aging caused by oxidative stress, inducing mice to have memory decline, skin relaxation, and decreased immune function, etc. 4 h after each subcutaneous injection of D-galactose, the mice in each group were gavaged, the SSN group had a bacterial concentration of 1×10 9 CFU / mL, the postbiotic SSNH high-dose group was prepared when the bacterial concentration was 1×10 9 CFU / mL, the gavage concentration was 200 mg / kg / d, the postbiotic SSNM medium-dose group was prepared when the bacterial concentration was 1×10 8 CFU / mL, the gavage concentration was 100 mg / kg / d, the SSNL low-dose group was prepared when the bacterial concentration was 1×10 7 CFU / mL, the gavage concentration was 50 mg / kg / d, and the blank and model groups were gavaged with normal saline. The gavage volume of each group of mice was 0.2 mL / d. The injection and administration were performed once a day for 8 consecutive weeks, and the food intake, mental state, and activity ability of the mice were observed. Before the end of the animal experiment, the mice were subjected to open field behavior test; after the experiment, the colon, skin, liver, and brain tissues of the mice were sampled and subjected to HE staining; the levels of IL-1β, IL-6, TNF-α, AGEs, 8-OHdG, SOD, GSH-Px, and MDA in the serum of the mice and the hyaluronic acid content and water content in the skin tissue were determined by using an Elisa kit; and the relative abundance of intestinal flora of the mice was determined.

[0067] (3) Experimental results As Figure 7The skin tissue condition of mice was detected by H&E staining. The skin of normal group mice showed complete structure, thin epidermis layer, tight connection between dermis layer and epidermis layer, and full and orderly arranged sebaceous glands and hair follicles. The skin of mice treated with long-term D-gal showed disordered tissue structure, uneven epidermis, fewer sweat pores than other groups, unclear tissue boundary, loose level, incomplete local epidermis keratinization and atrophic sebaceous glands. After intragastric administration of SSN and its postbiotics, the skin tissue of mice was layered more clearly, the subcutaneous tissue structure was much clearer, and the number of hair follicle sweat pores also increased. Especially in the SSNH group, the skin tissue structure was arranged more regularly than other groups, the structure between tissues was clear, the number of hair follicles and sweat pores increased significantly, and the SSNM and SSNL groups also showed more regular tissue structure, showing their role in promoting the recovery of aging skin tissue.

[0068] Figure 8 The results were the colon tissue morphology of mice in each group. The colon tissue structure of the CONT group was normal, the intrinsic layer of the intestinal gland was arranged neatly and closely, the gland structure was clear and complete, there was no deformation and atrophy, and no inflammatory cell infiltration was observed. The MOD group showed more severe inflammatory lesions, mucosal epithelial cells were shed, colon gland structure was changed, glands were deformed and atrophied, interstitial space was widened, and irregular crypts were present on the surface. The colon glands of mice treated with SSN were arranged more regularly, the destruction of intestinal gland structure was reduced to some extent, and inflammatory cell aggregation was less. After treatment with SSN-13 postbiotics, the colon tissue structure of mice was improved and was closer to that of the CONT group. The colon tissue crypt structure of the SSNH group was restored well, the number increased, the mucosal epithelium and crypt were relatively complete, the submucosal space was small and regular, the goblet cells were relatively rich, and the inflammatory cell infiltration was less. It was revealed that aging of the body could damage the intestinal barrier and cause inflammatory lesions, and Lactobacillus rhamnosus SSN-13 and its postbiotics could improve the integrity of the intestinal mucosa and the degenerative changes of the intestinal tissue.

[0069] Continuous administration of D-gal can cause damage to liver cell structure, cell apoptosis, degeneration and necrosis, and these structural changes are closely related to the aging of the liver. Figure 9The liver cells of the MOD group were swollen, arranged in disorder, and abnormal in shape, and the number of binucleate cells was reduced. The cytoplasm was deeply stained in eosinophilic blue-purple. Necrosis of liver cells and inflammatory cell infiltration were observed around the central vein. The liver cells of the SSN group were less vacuolar degenerative, and no necrotic liver cells were observed. The cells were arranged in order, close to the CONT group, and basically restored to normal structure. No obvious inflammatory cell infiltration was observed in the SSNH / M / L group. The cytoplasm was loose and lightly stained. There was mild steatosis of liver cells around the central vein. Among them, the liver cells of the SSNH group were relatively more complete, indicating that the high-dose probiotic of R. lactis SSN-13 had a better effect on improving liver cell aging.

[0070] The hippocampus is a structure in the brain that is primarily responsible for processing and storing memories, as shown in Fig. 1. Figure 10 As shown in Fig. 2, the nuclei of the brain tissue of the CONT group of mice were uniform in size and regular in shape, the nucleoli were clear, and the chromatin was evenly distributed. The neurons of the MOD group of mice were arranged loosely and in disorder, the cell layer was reduced, the nuclei were shrunk, and the neurons were necrotic. The layout was disordered. After intervention of R. lactis SSN-13, the mouse nerve cells had no obvious changes, and the morphology and distribution were more regular. The nuclei of the SSNH / M / L group were obviously better than those of the MOD group in morphology and distribution, the neuron density was improved, the nucleoli were clear, the arrangement was orderly, the outline was clear, and there was no obvious pathological damage.

[0071] Through 16S rRNA gene sequencing analysis, it was found that Figure 11 It can be seen that the relative abundance of the Firmicutes phylum in the MOD group is the highest. After gavage of SSN and its metaplast, the relative abundance of Bacteroidetes in the mouse intestine increases, and the relative abundance of Firmicutes decreases. The ratio of Firmicutes to Bacteroidetes is an important indicator reflecting the stability of intestinal flora and is directly related to the occurrence of various diseases. The experimental results show that the relative abundance ratio of Firmicutes / Bacteroidetes in the MOD group is significantly higher than that in the CONT group, the SSN group, the SSNH group, the SSNM group, the SSNL group and the SSML group, reaching 1.67, 0.35, 0.42, 0.59, 0.99, 1.17 respectively, indicating that aging may lead to the disorder of the composition and structure of intestinal flora, and the supplement of SSN-13 can reverse the changes in intestinal flora structure caused by aging. In addition, the SSNH group and the SSN group are closer to the CONT group, and lower than the SSNM group and the SSNL group, indicating that the effect of R. lactis SSN-13 and its metaplast on improving the composition and structure of intestinal flora in aging mice is enhanced with the increase of the dose.

[0072] Figure 12The relative abundance of intestinal flora of each group of mice at genus level was analyzed, Muribaculaceae, Bacteroides 、 Clostridium 、 Prevotella and Ileibacterium The relative abundance of each group of mice in the intestine was high. Among them, Akkermansia in SSN and SSN(H / M / L) groups was higher than 2%, MOD group Muribaculaceae was 24% lower than CONT group, SSN group, SSNH group, SSNM group and SSML were 6.5%, 23.7%, 16.3%, 7.7% higher than MOD group, respectively. The relative abundance structure of each group of mice at genus level, the genus composition of SSN group and SSNH group was more close to CONT group, indicating that the improvement effect on intestinal flora of aging mice was relatively better; SSNL and SSNM groups had significant differences in relative abundance and composition structure of genus from CONT group, the effect was second, but still better than MOD group.

[0073] Aging is closely related to the occurrence of chronic diseases, physiological function and cognitive ability decline. Open field test is a common method to evaluate the autonomous behavior, exploratory behavior and tension of experimental animals in a new and unfamiliar environment. Figure 13 (A) is the behavior trajectory of each group of mice in the open field. MOD group mice have less trajectory in the center area. In contrast, SSN group, SSN(L / M / H) group of aging mice have more and denser trajectory lines, which are more close to CONT group. Figure 13 (B) is the total trajectory distance of each group of mice in the open field area within 5 minutes. MOD group mice move 5.18m, which is significantly lower than CONT group mice of 14.93m ( P <0.0001), and also has significant difference with SSN group (13.26m), SSNH group (12.98m), SSNM group (12.00m) and SSNL group (12.04m); Figure 13 (C) is the residence time of each group of mice in the center area. Compared with CONT group (47.04s), MOD group only stays in the area for 7.03s, while SSN group (37.17s) and SSNH group (39.8s) are more close to CONT group, and the residence time in the center area presents a gradient decrease with the decrease of SSN-13 postbiotic concentration, SSNM group is 36.07s, SSNL group is 25.3s, which can show that Lactobacillus rhamnosus SSN-13 and its postbiotic supplementation can make mice more active and improve the autonomous movement ability of mice.

[0074] The inflammatory microenvironment during the aging process is closely related to the increased inflammatory cytokines in circulation. Senescent cells secrete various pro-inflammatory cytokines, chemokines, and interleukins, such as IL-6, TNF-a, and IL-1b, etc. As shown in Figure 14 (A), the IL-6 content of the MOD group was 108.38 ng / L, which was significantly higher than that of the CONT group (50.45 ng / L). After supplementing the microecological preparation, the IL-6 content of the SSN group was significantly reduced (P < 0.001). P <0.001), the postbiotic preparation also showed the ability to reduce IL-6 levels, and with the increase of postbiotic preparation concentration, the IL-6 content was closer to the CONT group and the SSN group, and the SSNH group had the best effect, with an IL-6 content of 56.79 ng / L. IL-1b inhibits the expression of longevity genes through the NF-κB pathway, and its excessive release can induce circulatory disorders and organ plasticity damage. SIRT1 Figure 14 (B) is the change of IL-1b content in each group of mice. Compared with the MOD group (101.99 ng / L), the IL-1b levels in the SSN treatment and SSN postbiotic high / medium / low dose groups were significantly reduced (P < 0.0001). P <0.0001), the IL-1b levels of the SSN group and the SSNH group were 67.66 ng / L and 55.9 ng / L, respectively, which were close to the CONT group of 44.96 ng / L. In terms of TNF-a levels, according to Figure 14 (C) shows that the TNF-a content in the serum of the MOD group was the highest, reaching 627.36 ng / L, which was significantly higher than that of the CONT group (270.47 ng / L). After administration, the lactic acid bacteria microecological preparation SSN group could significantly reduce the TNF-a content in the serum of mice, which was close to the SSNH group. With the increase of the dose of postbiotic preparation, TNF-a showed a downward trend, and after reaching the high-dose group, the TNF-a content was significantly reduced compared with the MOD group (P < 0.01). P <0.01); there was no significant correlation between the low-dose postbiotic preparation and the MOD group, indicating that different doses of postbiotic preparations have different effects on the regulation of TNF-a levels.

[0075] 8-hydroxydeoxyguanosine (8-OHdG) is not only a "molecular scar" of oxidative damage, but also a key landmark for disease warning and anti-aging intervention. As a specific biomarker of DNA oxidative damage, it can be used to assess the aging process, oxidative stress level, and related disease risk. For example, Figure 14 ​As shown in (D), the 8-OHdG content in the CONT group was 32.01 ng / L, significantly lower than that in the MOD group (69.74 ng / L). Similarly, the 8-OHdG content in the SSN group (46.10 ng / L) and the SSNH group (36.09 ng / L) was also significantly lower than that in the MOD group. P <0.0001), and closer to the CONT group. The SSNM and SSNL groups also reduced 8-OHdG levels in the serum of aging mice, indicating that supplementation with lactic acid bacteria microecological preparations and post-biotic preparations can improve aging in mice. Advanced glycation end products (AGEs) in mouse serum showed a strong positive correlation with 8-OHdG, jointly constructing an oxidative damage network, which is a core quantitative indicator of metabolic damage. For example... Figure 14 As shown in (E), the serum AGEs content in the MOD group of aging mice was significantly increased. P <0.0001), reaching 564.35 ng / L, the SSNH group (297.53 ng / L) was closer to the CONT group (226.32 ng / L), and the SSN group (371.38 ng / L) showed a significant difference from the MOD group. P <0.0001), the AGEs content in the SSNM group (376.37 ng / L), SSNL group (411.47 ng / L), and SSNH group showed a dose-gradient decrease. Although it did not reach the AGEs level in normal mouse serum, it could effectively reverse the increase in AGEs level in aging mice caused by D-galactose.

[0076] Superoxide dismutase (SOD) is an important antioxidant enzyme in the host body, and its activity level is an important indicator of aging. SOD can protect cells from free radical damage and plays a role in delaying aging and inhibiting bacteria. Figure 15 (A) The results showed that the SOD enzyme activity in the CONT group reached 74.25 U / mL, while the activity in the MOD group (34.79 U / mL) was significantly lower than that in the CONT group. P <0.001), the SSN group and the SSNH / M / L group showed significant improvement compared to the MOD group. Among them, the SSNH group reached 75.23 U / mL, which was better than the other groups. Furthermore, the SOD enzyme activity in the serum of mice in the postbiotic preparation group showed a concentration gradient change.

[0077] Glutathione peroxidase (GSH-Px) is an important peroxide-degrading enzyme widely present in the body. GSH-Px is the most abundant selenoprotein in most cells. As a type of peroxidase, it can decompose peroxides, thus protecting cells from peroxide damage. Increasing GSH-Px activity can effectively delay organ aging and prevent related diseases. Figure 15(B) shows that the GSH-Px enzyme activity in the serum of the MOD group (48.63 U / L) is significantly lower than that of the CONT group (98.56 U / L), and the SSN group (91.46 U / L) is closer to the CONT group. The supplementation of the postbiotic preparation effectively inhibits the attenuation of GSH-Px activity in the serum of the aging mice, and the effect is enhanced with the increase of the concentration. The GSH-Px activities of the SSNH group, the SSNM group and the SSNL group are 77.39 U / L, 72.43 U / L and 61.57 U / L, respectively.

[0078] Malondialdehyde is one of the core end products of lipid peroxidation, which can interact with biological molecules such as nucleic acids and proteins, leading to enzyme inactivation, collagen hardening, and irreversible damage to cell function, and is an important indicator of aging. For example Figure 15 (C) shows that the MDA content in the serum of the CONT group, the SSN group, the SSNH / M / L group is 2.49, 2.79, 2.60, 2.62, 4.64 nmol / mL, respectively, which is significantly lower than 5.59 nmol / mL of the MOD group, and the SSN group, the SSNH group and the SSNM group all reach extremely significant difference (P<0.001), and are closer to the CONT group. The experimental results show that D-galactose significantly accelerates the aging process and increases the MDA level in the mice. The supplementation of the SSN lactic acid bacteria microecological preparation and the postbiotic preparation significantly reduces the MDA content in the serum of the aging mice, which is of great significance for the maintenance of host health. P <0.001), and is closer to the CONT group. The experimental results show that D-galactose significantly accelerates the aging process and increases the MDA level in the mice. The supplementation of the SSN lactic acid bacteria microecological preparation and the postbiotic preparation significantly reduces the MDA content in the serum of the aging mice, which is of great significance for the maintenance of host health.

[0079] Hyaluronic acid has high viscoelasticity and can maintain skin elasticity and keep skin moisture, which is another key factor reflecting skin aging. The excessive accumulation of D-galactose can cause oxidative stress and glycosylation damage in the skin, affect the synthesis of hyaluronic acid, and trigger signs of impaired skin barrier function. The effects of the SSN group and the SSN postbiotic group on the hyaluronic acid content in the skin of the aging mice are shown in Figure 16 (A) shows that the hyaluronic acid content in the skin tissue of the mice increases after the supplementation of SSN and its postbiotic high / middle / low dose, which is 8.76, 8.14, 9.21, 14.72 ng / mg, respectively, and the CONT group is 13.55 ng / mg, which is higher than the MOD group (5.18 ng / mg), indicating that the supplementation of Rhamnosus lactis SSN-13 and its postbiotic preparation can improve the decrease of hyaluronic acid in the aging mice, and the postbiotic low dose group shows a higher hyaluronic acid content.

[0080] The water content of the body gradually decreases with age, and the water content in the skin mainly exists in the dermis. Compared with young individuals, the skin water content of elderly individuals is often lower. For example Figure 16As shown in (B), the moisture content of the CONT group was 85.08%, which was significantly different from the 59.73% of the MOD group. P <0.001), indicating that D-galactose accelerates aging in mice, leading to accelerated loss of skin moisture. After intervention with SSN-13 and its post-biotic, the skin moisture content of mice increased. The moisture content of the SSN group and the SSNH / M / L group were 76.22%, 79.71%, 76.23%, and 81.23%, respectively, all of which were significantly different from the MOD group. P <0.001). The results showed that intervention with Lactobacillus rhamnosus SSN-13 and its post-biotics increased the water content in the skin of aging mice, slowed down the skin aging process, and inhibited skin moisture loss caused by cumulative glycation damage.

[0081] The foregoing description is not intended to limit the invention, nor is the invention limited to the examples given. Any changes, modifications, additions, or substitutions made by those skilled in the art within the scope of the invention should also be considered within the protection scope of the invention.

Claims

1. Use of Lactobacillus rhamnosus SSN-13 in the preparation of an anti-glycation, anti-aging medicament, characterized in that: The preservation number of the Lactobacillus rhamnosus SSN-13 is CGMCC No: 27227.

2. Use according to claim 1, characterized in that: The application of the Lactobacillus rhamnosus SSN-13 in the preparation of a preparation for inhibiting the generation of fluorescent AGEs, fructosamine, bis-tyrosine and pentosidine.

3. Use according to claim 1, characterized in that: The Lactobacillus rhamnosus SSN-13 inhibits the increase of β-galactosidase activity of human immortalized keratinocytes under oxidative damage conditions.

4. Use according to claim 1, characterized in that: The Lactobacillus rhamnosus SSN-13 regulates the mRNA abnormal expression of the key target points for regulating the senescent growth of human liver cancer cells under oxidative stress and glycosylation damage conditions MAPK1 、 PSEN1 、 Sirt1 、 Collagen Ⅲ ​ 5. The use according to claim 1, characterized in that: The application of the Lactobacillus rhamnosus SSN-13 in the preparation of a preparation for improving the degenerative changes of skin, intestinal tract, liver and brain tissue and the decline of cognitive function caused by aging of individuals.

6. Use according to claim 1, characterized in that: The application of the Lactobacillus rhamnosus SSN-13 in the preparation of a preparation for improving the composition and structure of intestinal flora of aging individuals.

7. Use according to claim 1, characterized in that: The application of the Lactobacillus rhamnosus SSN-13 in the preparation of a preparation for reducing the content of AGEs and the content of inflammatory markers IL-6, IL-1β and TNF-α in serum of aging individuals.

8. The use according to claim 1, characterized in that: The application of the Lactobacillus rhamnosus SSN-13 in the preparation of a preparation for reducing the decrease of hyaluronic acid and moisture content of skin of aging individuals caused by glycosylation damage.

9. Use according to any one of claims 1 to 8, characterized in that: The Lactobacillus rhamnosus SSN-13 is prepared into a microecological preparation with a viable cell count of 1 x 10 7 ~1 x 10 9 CFU / mL or a microbial body preparation with a concentration of 1 x 10 7 ~1 x 10 9 CFU / mL, wherein the metaplasma contains lactic acid bacterial cell metabolites.

10. Use according to claim 9, characterized in that: The application of the microecological preparation or metagenomic preparation in the preparation of food, health care products, cosmetics or therapeutic drugs.

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