Metagen prepared from bifidobacterium adolescentis and used for promoting synthesis of host collagen

Abstract

The invention discloses a metagen prepared from bifidobacterium adolescentis and used for promoting synthesis of host collagen, and belongs to the technical field of microorganisms and medicines. The metagen of the bifidobacterium adolescentis CCFM1386 provided by the invention has a good effect of promoting synthesis of host collagen, improves moisture and elasticity of skin, and meanwhile, can improve the antioxidant capacity of the skin and maintain the stable state of collagen. The method specifically comprises the following steps: promoting the enzyme activity of the HSF cell LH1 in vitro by a thallus lysate, and improving the content of COL I and COL III and the enzyme activity of the LH1 in vitro from a fermented supernatant; the bifidobacterium adolescentis has the advantages that the content of collagen is increased by orally taking inactivated thalli, the enzyme activity of LH1, P4H and LOX can be improved by orally taking self-fermentation supernate, the synthesis of the collagen is promoted, and the content of the collagen is increased, so that the bifidobacterium adolescentis has huge application prospects in preparation of oral medicines for promoting the synthesis of host collagen and preparation of foods, health-care products or cosmetics.

Description

Technical Field

[0001] The present invention relates to a postbiotic prepared from a strain of Bifidobacterium adolescentis and capable of promoting host collagen synthesis, belonging to the technical field of microorganisms and medicine. Background Art

[0002] Collagen, the most abundant structural protein in the body, is widely distributed in various tissues, including skin, bone, cartilage, tendons, blood vessels, and internal organs. It plays a crucial role in maintaining tissue structural integrity, imparting mechanical properties to tissues, and regulating cellular physiological behavior. Maintaining a dynamic balance between collagen synthesis and degradation is crucial for the proper functioning of tissues under various physiological and pathological conditions. In particular, promoting the host's own collagen synthesis has become a key strategy for enhancing tissue regeneration, repairing damage, and improving function in areas such as skin and bone repair, cartilage regeneration, and chronic wound healing.

[0003] At present, there are a number of patented technologies that attempt to improve the collagen content in host tissues by means of oral and topical intervention, small molecule drugs or plant extract applications. For example, CN119700752A discloses the application of Bergapten or its isomers in the preparation of products that improve skin aging and / or promote skin repair. However, existing research mostly stays at the level of total collagen detection. In particular, most current studies have failed to systematically evaluate the expression and activity changes of key enzymes in the process of collagen biosynthesis, such as prolyl hydroxylase (P4H), lysyl hydroxylase (LH1), lysyl oxidase (LOX), etc. These enzymes play a core role in the maturation, stability and fiber formation of collagen molecules. The key regulatory nodes in the anabolic process are not detected, which makes it difficult for the existing technology to clarify the mechanism of action, limiting the scientific evaluation and optimization of the intervention effect. At the same time, the identification and verification of regulatory targets (such as key molecules in upstream signaling pathways) are also relatively lacking, making the strategy of promoting collagen synthesis have obvious deficiencies in terms of mechanism clarity and precise intervention. Summary of the Invention

[0004] In order to solve the above technical problems, the present invention provides a use of Bifidobacterium adolescentis CCFM1386 and its postbiotics in the preparation of a product that promotes host collagen synthesis.

[0005] The first technical solution provided by the present invention is a strain of Bifidobacterium adolescentis CCFM1386, which has been deposited in the Guangdong Provincial Microbial Culture Collection Center on January 10, 2025, with the deposit number: GDMCC No: 65746.

[0006] The Bifidobacterium adolescentis CCFM1386 was derived from samples of healthy people. The strain was sequenced and analyzed, and the obtained sequence was compared with the nucleic acid sequence in NCBI. The result showed that it was Bifidobacterium adolescentis, and was named Bifidobacterium adolescentis CCFM1386.

[0007] The colonies of Bifidobacterium adolescentis CCFM1386 on the MRS solid culture medium are white, round, and raised, relatively small, with neat edges.

[0008] The second technical solution provided by the present invention is a microbial preparation containing Bifidobacterium adolescentis CCFM1386 described in the first technical solution.

[0009] In certain embodiments, the concentration of Bifidobacterium adolescentis CCFM1386 in the microbial preparation is not less than 1×10 6 CFU / mL or ×10 6 CFU / g.

[0010] The third technical solution provided by the present invention is a postbiotic, which is prepared using the Bifidobacterium adolescentis CCFM1386 described in the first technical solution or the microbial preparation described in the second technical solution.

[0011] In certain embodiments, the postbiotics include bacterial lysate, inactivated or inactivated cells, fermentation supernatant, or a powder prepared by drying any of the above.

[0012] In certain embodiments, the inactivated or inactivated cells are prepared as follows: the Bifidobacterium adolescentis CCFM1386 is cultured in a culture medium, bacterial cells in the cell culture medium are collected, and the cells are heat-treated to obtain inactivated bacterial cells.

[0013] In certain embodiments, the heat treatment conditions are: 65-95°C, 10-30 min; preferably 65°C, 30 min.

[0014] In certain embodiments, the bacterial lysate is prepared by culturing the Bifidobacterium adolescentis CCFM1386 in a culture medium, collecting bacterial cells, homogenizing under high pressure, and obtaining the bacterial lysate from the supernatant of the centrifugation.

[0015] In certain embodiments, the fermentation supernatant is the supernatant obtained by centrifuging the Bifidobacterium adolescentis CCFM1386 in a culture medium.

[0016] The fourth technical solution provided by the present invention is a product containing the Bifidobacterium adolescentis CCFM1386 described in the first technical solution and / or the postbiotics described in the third technical solution.

[0017] In certain embodiments, the products include but are not limited to food, medicine, health care products or daily chemical products.

[0018] In certain embodiments, the food comprises the above composition and conventional auxiliary materials.

[0019] In certain embodiments, the conventional excipients include one or more of fillers, flavoring agents, binders, disintegrants, lubricants, antacids, and nutritional enhancers.

[0020] In certain embodiments, the health care product comprises the above composition and conventional excipients.

[0021] In certain embodiments, the conventional excipients include one or more of fillers, flavoring agents, binders, disintegrants, lubricants, antacids, and nutritional enhancers.

[0022] In certain embodiments, the dosage form of the product includes at least one of a cream, an emulsion, an oil, an aqueous solution, a gel, a powder, and a freeze-dried formulation.

[0023] In certain embodiments, the product is a probiotic powder.

[0024] In certain embodiments, the bacterial powder is a solid powder postbiotic prepared by drying the prepared liquid postbiotic of Bifidobacterium adolescentis CCFM1386.

[0025] In certain embodiments, drying includes but is not limited to spray drying, vacuum freeze drying, fluidized bed drying, and vacuum drying.

[0026] The fifth technical solution also provided by the present invention is the use of Bifidobacterium adolescentis CCFM1386 described in the first technical solution, the microbial preparation described in the second technical solution, or the postbiotic described in the third technical solution in the preparation of a product that promotes collagen synthesis in skin fibroblasts.

[0027] In certain embodiments, the product comprises at least one of the following effects:

[0028] (1) Increase the collagen content secreted by skin fibroblasts in vitro;

[0029] (2) Increase the activity of collagen synthase in skin fibroblasts in vitro.

[0030] The sixth technical solution provided by the present invention is the use of Bifidobacterium adolescentis CCFM1386 described in the first technical solution, the microbial preparation described in the second technical solution, or the postbiotics described in the third technical solution in the preparation of anti-aging products.

[0031] In some embodiments, the application includes at least one of the following effects:

[0032] (1) Increase the moisture and elasticity of skin tissue in aging individuals;

[0033] (2) Increase the antioxidant enzyme activity in the skin tissue of aging individuals;

[0034] (3) reduce inflammation levels in aging individuals;

[0035] (4) Increase collagen content in skin tissue of aging individuals;

[0036] (5) Increase the activity of collagen synthase in the skin tissue of aging individuals.

[0037] In certain embodiments, the symptoms associated with skin collagen loss include skin appearance, decreased skin collagen content, slowed collagen synthesis, and accelerated collagen degradation.

[0038] In certain embodiments, the aging comprises skin aging.

[0039] In certain embodiments, the skin aging includes skin dryness, decreased elasticity, sagging, wrinkling, oxidative damage, or collagen loss.

[0040] In certain embodiments, the content of Bifidobacterium adolescentis CCFM1386 in the product is not less than 5×10 7 CFU / mL.

[0041] In certain embodiments, the dosage of the postbiotics prepared by Bifidobacterium adolescentis CCFM1386 in the product is not less than 250 μg / kg.

[0042] In certain embodiments, the product is a pharmaceutical or cosmetic product.

[0043] In certain embodiments, the drug comprises the Bifidobacterium adolescentis CCFM1386, a drug carrier and / or a pharmaceutical excipient.

[0044] In certain embodiments, the pharmaceutical excipients include excipients and additives.

[0045] In certain embodiments, the pharmaceutical excipients include solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavoring agents, preservatives, suspending agents, coating materials, fragrances, anti-adhesive agents, integrities, penetration enhancers, pH regulators, buffers, plasticizers, surfactants, foaming agents, defoaming agents, thickeners, inclusion agents, humectants, absorbents, diluents, flocculants and deflocculating agents, filter aids, and release retardants.

[0046] In certain embodiments, the cosmetic contains Bifidobacterium adolescentis CCFM1386, matrix raw materials and / or conventional excipients.

[0047] In certain embodiments, the matrix raw materials include oil raw materials, wax raw materials, synthetic oil raw materials, powdery raw materials, colloid raw materials, coagulants, and surfactants.

[0048] In certain embodiments, the conventional excipients include one or more of moisturizers, whitening agents, flavoring agents, adhesives, lubricants, preservatives, film agents, antioxidants, emulsifiers, and cosmetic nutritional additives.

[0049] The seventh technical solution provided by the present invention is a method for promoting collagen synthesis by skin fibroblasts in vitro, wherein the method comprises mixing the postbiotics described in the third technical solution with skin fibroblasts and incubating them.

[0050] Compared with the prior art, the present invention has the following beneficial effects:

[0051] The Bifidobacterium adolescentis CCFM1386 of the present invention and the postbiotics prepared therefrom have the ability to promote host collagen synthesis, which is specifically embodied in:

[0052] (1) Increase the content of type I and type III collagen (COLⅠ, Ⅲ) secreted by human skin fibroblasts (HSF) in vitro;

[0053] (2) Increase LH1 enzyme activity in human skin fibroblasts (HSF) in vitro;

[0054] (3) Improve skin moisture and elasticity in aging individuals;

[0055] (4) Improve the enzyme activities of CAT and SOD in the skin of aging individuals;

[0056] (5) Reduce the levels of CRP, IL-6, and TNF-α in the serum of aging individuals;

[0057] (6) Increase the levels of COL I, III, IV, and VII in the skin of aging individuals;

[0058] (7) Increase the enzyme activities of LH1, P4H and LOX in the skin of aging individuals;

[0059] Therefore, postbiotics prepared from Bifidobacterium adolescentis CCFM1386 have great application prospects in products that promote host collagen synthesis.

[0060] Biomaterial Deposit

[0061] Bifidobacterium adolescentis CCFM1386, taxonomically named Bifidobacterium adolescentis, was deposited in the Guangdong Provincial Microbial Culture Collection on January 10, 2025, with the collection number GDMCC No: 65746, and the collection address is Building 59, No. 100 Xianlie Middle Road, Guangzhou. BRIEF DESCRIPTION OF THE DRAWINGS

[0062] Figure 1 Effects of postbiotics prepared for CCFM1386 on the secretion of COLⅠ and COLⅢ by HSF cells;

[0063] Figure 2 Effects of postbiotics prepared for CCFM1386 on LH1 enzyme activity in HSF cells;

[0064] Figure 3 This is the flow chart of the mouse experiment;

[0065] Figure 4 Effects of postbiotics prepared for CCFM1386 on the levels of COLⅠ, Ⅲ, Ⅳ and Ⅶ in mouse skin tissue;

[0066] Figure 5 Effects of postbiotics prepared for CCFM1386 on LH1, P4H, and LOX enzyme activities in mouse skin tissue;

[0067] Figure 6 Effects of postbiotics prepared for CCFM1386 on CAT and SOD enzyme activities in mouse skin tissue;

[0068] Figure 7 Effects of postbiotics prepared for CCFM1386 on the levels of CRP, IL-6, and TNF-α in mouse serum;

[0069] Figure 8 Effects of postbiotics prepared for CCFM1386 on skin hydration and elasticity in mice;

[0070] Figure 9 Masson stain for skin.

[0071] “**” indicates a significant statistical difference compared with the control group (P<0.01), and “***” indicates an extremely significant statistical difference compared with the control group (P<0.001).

[0072] “#” indicates statistical difference compared with the model group (P<0.05), “##” indicates significant statistical difference compared with the model group (P<0.01), “###” indicates extremely significant statistical difference compared with the model group (P<0.001); “####” indicates extremely significant statistical difference compared with the model group (P<0.0001). DETAILED DESCRIPTION

[0073] The following describes preferred embodiments of the present invention. It should be understood that the embodiments are for better explanation of the present invention and are not intended to limit the present invention.

[0074] The raw materials used in the embodiment are:

[0075] The human skin fibroblasts (HSF) involved in the following examples were purchased from Kunming Cell Bank.

[0076] The BALB / c mice involved in the following examples were purchased from Weitonglihua Company.

[0077] The Bifidobacterium adolescentis CCFM1386 involved in the following examples is a strain screened by the Food Biotechnology Center of Jiangnan University.

[0078] The ELISA kits involved in the following examples were purchased from Nanjing Senbeijia Biotechnology Co., Ltd.

[0079] The D-galactose involved in the following examples was purchased from Shanghai Aladdin Biochemical Technology Co., Ltd.

[0080] The BCA protein concentration determination kit involved in the following examples was purchased from Shanghai Beyotime Biotechnology Co., Ltd. The culture medium involved in the following examples is as follows:

[0081] Modified MRS liquid medium: yeast extract 5.0 g / L, peptone 10.0 g / L, glucose 20.0 g / L, anhydrous sodium acetate 2.0 g / L, diammonium hydrogen citrate 2.0 g / L, dipotassium hydrogen phosphate 2.6 g / L, manganese sulfate monohydrate 0.25 g / L, magnesium sulfate heptahydrate 0.5 g / L, cysteine ​​1 g / L, and Tween-80 1 mL / L, pH 6.2-6.4.

[0082] Modified MRS solid medium: yeast powder 5.0 g / L, peptone 10.0 g / L, glucose 20.0 g / L, anhydrous sodium acetate 2.0 g / L, diammonium hydrogen citrate 2.0 g / L, dipotassium hydrogen phosphate 2.6 g / L, manganese sulfate monohydrate 0.25 g / L, magnesium sulfate heptahydrate 0.5 g / L, Tween-80 1 mL / L, cysteine ​​1 g / L, and agar 20.0 g / L, pH 6.2-6.4.

[0083] Cell culture medium: 89% (v / v) DMEM medium + 10% (v / v) fetal bovine serum + 1% (v / v) 100× mixed solution of penicillin and streptomycin (the penicillin content in the mixed solution is 10000 U / mL, and the streptomycin concentration is 10 mg / mL).

[0084] Example 1: Cell recovery and culture

[0085] First, take out the frozen human skin fibroblast cell line (HSF), quickly thaw it in a 37℃ water bath, then centrifuge it at 1000r / min for 3min, discard the supernatant, add an appropriate volume of cell culture medium to resuspend the cells, and place them in a culture dish. Place them in a 37℃ incubator containing 5% CO2 for culture. When the cells recover their vitality and grow for 1-2 days to reach 70%-80% fusion, cell passage is performed.

[0086] Example 2: Preparation of postbiotics using Bifidobacterium adolescentis CCFM1386

[0087] 1. Screening and identification of Bifidobacterium adolescentis CCFM1386

[0088] The strain samples screened were from healthy people. After pretreatment, they were stored in a -80°C refrigerator in 20% glycerol. After thawing, they were mixed and 0.5 mL of sample was added to 4.5 mL of normal saline. The sample was gradient diluted with normal saline. The appropriate gradient dilution solution was selected and spread on MRS solid culture medium. It was anaerobically cultured at 37°C for 48 hours. The typical colonies of Bifidobacterium adolescentis were picked and streaked on MRS solid culture medium for purification. Single colonies were picked and transferred to MRS liquid culture medium for bacterial enrichment and preserved in 30% glycerol to obtain the strain. The strain genome was extracted for 16S rDNA amplification and sequencing (performed by Suzhou Jinweizhi Biotechnology Co., Ltd.). The results were identified as Bifidobacterium adolescentis by NCBI sequence alignment and named Bifidobacterium adolescentis CCFM1386. It was deposited in Guangdong Provincial Microbiological Culture Collection on January 10, 2025, with the deposit number GDMCCNo: 65746.

[0089] 2. Preparation of postbiotics with Bifidobacterium adolescentis CCFM1386

[0090] (1) Resuscitating Bifidobacterium adolescentis CCFM1386 from a sterile tube by streaking, culturing in a modified MRS solid medium at 37°C in an anaerobic workstation for 48 h to obtain a single colony; picking a single colony and inoculating it into a modified MRS liquid medium, culturing at 37°C for 12-18 h to obtain culture medium 1;

[0091] The culture solution 1 was inoculated into the modified MRS liquid medium at an inoculum volume of 2% (v / v), and cultured anaerobically at 37°C for 12 h to obtain the seed solution;

[0092] The seed liquid was inoculated at 2% (v / v) in modified MRS liquid medium for expansion, and cultured anaerobically at 37°C for 18h, the viable cell count was recorded and the bacterial liquid a was obtained.

[0093] The bacterial liquid a was centrifuged at 8000r / min for 30min, and the supernatant and bacterial slurry were collected. The supernatant was heat-treated (65°C, 30min) and freeze-dried for standby, and the Bifidobacterium adolescentis CCFM1386 fermentation supernatant freeze-dried powder (denoted as CCFM1386-S) was prepared. The bacterial slurry was resuspended with 75% volume of double distilled water, and the resuspension was heat-treated (65°C, 30min) and then homogenized in a high-pressure homogenizer (1000MPa, 10P times). After homogenization, the supernatant was obtained by centrifugation at 8000r / min for 30min to obtain the bacterial lysate (denoted as CCFM1386-J).

[0094] The Bifidobacterium adolescentis CCFM1386 postbiotic (bacterial lysate CCFM1386-J and fermentation supernatant CCFM1386-S) was prepared by the above means.

[0095] Example 3: Effect of the postbiotic prepared from Bifidobacterium adolescentis CCFM1386 on the collagen content of HSF cells

[0096] (1) The logarithmic growth phase HSF cells were inoculated at 1×10 5 CFU / mL in a 6-well plate, and the cells were cultured overnight until the cells adhered. The old culture medium was discarded, and PBS was used to rinse 3 times. The blank group and the postbiotic treatment group were set up;

[0097] The blank group (control): containing cell culture medium and HSF cells, but not containing postbiotic;

[0098] The postbiotic treatment group: containing cell culture medium and HSF cells, and containing postbiotic.

[0099] The postbiotic was resuspended in cell culture medium (the amount of resuspended postbiotic was equivalent to the amount of postbiotic prepared from bacterial liquid with a concentration of 5.0×10 7 CFU / mL), and 100μL of the postbiotic prepared from Bifidobacterium adolescentis CCFM1386 was added.

[0100] (2) The above-mentioned hole plates were incubated at a temperature of 37°C for 24h, respectively.

[0101] (3) After incubation, the cell culture supernatant was collected and centrifuged at 1000r / min for 5min to obtain the supernatant. The COL I and COL III contents were detected by ELISA kit.

[0102] (4) After washing the cells in the plate twice with PBS, add 1 ml of trypsin containing 0.25% EDTA and digest at 37°C for 30 seconds. Observe the cells under a microscope until most of the cells become round. Then add 4 ml of DMEM complete medium to terminate the digestion. Gently pipette to completely detach the cells from the culture dish. Transfer the resulting cell suspension to a 15 ml centrifuge tube and centrifuge at 1000 rpm for 5 minutes. Remove the supernatant and resuspend the cells with a mixed protease inhibitor. Repeat freezing and thawing (quick freezing at -80°C and slow thawing at 4°C) three times to lyse the cells to obtain a cell lysate sample.

[0103] (5) Use the BCA protein concentration assay kit to detect the total protein concentration of the sample in (4) and calibrate the above data.

[0104] The results are as follows Figure 1 As shown in the results, compared with the control group, CCFM1386-S increased the content of COLⅠ secreted by HSF cells by approximately 20%, and CCFM1386-S significantly increased the content of COLⅢ secreted by HSF cells by approximately 18%. These results indicate that postbiotics prepared by CCFM1386 can increase the content of collagen secreted by HSF cells and have the potential to regulate collagen synthesis metabolism.

[0105] Example 4: Effect of postbiotics prepared from Bifidobacterium adolescentis CCFM1386 on collagen synthase activity in HSF cells

[0106] The preparation of cell lysate samples involved in the following examples is the same as that in Example 3. The enzymatic activity of lysine hydroxylase (LH1) in HSF cell lysate was detected by ELISA kit.

[0107] LH1 plays an important role in collagen synthesis and is one of the key enzymes in the post-modification process of collagen. LH1 is responsible for catalyzing the hydroxylation reaction of lysine residues to generate hydroxylysine, a process that is crucial for the stability and function of collagen. Hydroxylated lysine not only helps cross-link collagen fibers and enhances their mechanical strength, but also participates in the interaction between collagen and other extracellular matrix components, thus playing an important role in maintaining tissue structure and function. The results of LH1 are as follows Figure 2 As shown, compared with the control group (27.41 IU / g), the LH1 enzyme activity in HSF cells treated with CCFM1386-S was increased to 34.96 IU / g, and the LH1 enzyme activity in HSF cells treated with CCFM1386-J was increased to 41.72 IU / g. These results indicate that postbiotics prepared with CCFM1386 have the potential to promote collagen synthesis in the body.

[0108] Example 5: Effect of postbiotics prepared from Bifidobacterium adolescentis CCFM1386 on collagen content in the skin of aging mice

[0109] The preparation method of the postbiotics (CCFM1386-S and CCFM1386-J) of Bifidobacterium adolescentis CCFM1386 involved in the following examples is the same as that in Example 2, except that CCFM1386-J in this embodiment specifically refers to the bacterial solution a obtained in Example 2, which is centrifuged at 8000 r / min for 30 min to obtain the bacterial sludge, and the bacterial sludge is resuspended with double distilled water containing 75% of the volume of the original bacterial solution, and the resuspension is heat-treated at 65°C for 30 min without high-pressure homogenization.

[0110] (1) Twenty 8-week-old healthy male BALB / c mice were randomly divided into 4 cages with 5 mice in each cage. The 4 cages were: 1 cage for the blank group (control), 1 cage for the model group (codel), 1 cage for the CCFM1386-S group, and 1 cage for the CCFM1386-J group.

[0111] Blank group (control): physiological saline was used as a control;

[0112] Model group: normal saline was used as the control;

[0113] CCFM1386-S group: using Bifidobacterium adolescentis CCFM1386 metabolite (fermentation supernatant), the dose is: 500 mg / kg mouse body weight;

[0114] CCFM1386-J group: Bifidobacterium adolescentis CCFM1386 postbiotics (inactivated bacteria) were used at a dose of 500 mg / kg mouse body weight;

[0115] Among them, the inactivated bacteria or metabolites in the above groups were 1×10 9 Inactivated bacteria or metabolites prepared from a bacterial solution with an amount of CFU of viable bacteria.

[0116] The experiment lasted for 8 weeks: After the mice adapted for one week, all groups except the blank group were subcutaneously injected with D-galactose (500 mg / kg) at a dose of 0.2 mL / mouse / day. Starting from the second week, each intervention group used the corresponding strain-prepared postbiotic freeze-dried powder (inactivated bacteria or fermentation supernatant) dissolved in normal saline at a corresponding dose, and the mice were gavaged at a dose of 0.2 mL / mouse / day. The blank group and the model group were gavaged with the same amount of normal saline as a control until the end of the experiment. All groups had free access to water and food. The experimental process is as follows: Figure 3 shown.

[0117] After the experiment, the mice were killed and blood was collected from the eyeballs. After standing for 40 minutes, the blood was centrifuged at 3000 r / min for 20 minutes, and the blood supernatant was stored at -80°C. The back skin tissue was cut and ground into a homogenate at a weight-to-volume ratio of 1:9 with PBS. The homogenate was centrifuged at 3000 r / min for 20 minutes, and the skin supernatant was collected for detection using an ELISA kit.

[0118] The contents of type Ⅰ, Ⅲ, Ⅳ and Ⅶ collagen (COL Ⅰ, Ⅲ, Ⅳ, Ⅶ) in mouse skin were detected by ELISA kit. Figure 4 shown.

[0119] Compared with the blank group (8.18 μg / mg), the content of COLⅠ in the skin of the model group decreased significantly to 3.44 μg / mg. Oral administration of CCFM1386-S and CCFM1386-J groups significantly increased the content of COLⅠ compared with the model group, restoring the content of COLⅠ in the skin to 5.90 μg / mg and 5.23 μg / mg, respectively. Both groups can increase the content of COLⅠ.

[0120] The content of COLⅢ in the blank group was 1.94μg / mg, which was 2.4 times that of COLⅢ in the model group (0.81μg / mg). Oral administration of CCFM1386-S could significantly increase its content to 1.55μg / mg compared with the model group, and CCFM1386-J could also significantly increase its content to 1.41μg / mg compared with the model group. Both of them can increase the content of COLⅢ.

[0121] For COLⅣ, the content in the blank group was 15.29μg / mg, and the content in the model group was 6.21μg / mg. Oral administration of CCFM1386-S group can significantly increase the content of COLⅣ to 12.10μg / mg compared with the model group, and CCFM1386-J group can also significantly increase the content of COLⅣ to 11.40μg / mg compared with the model group.

[0122] For COLⅦ, the content in the blank group was 2.22 μg / mg, which was 2.6 times that of the model group (0.85 μg / mg). Only the oral CCFM1386-S group could significantly increase the content of COLⅦ compared with the model group, and this group restored the content to 1.98 μg / mg.

[0123] The above results show that the postbiotics (inactivated bacteria and supernatant) prepared from Bifidobacterium adolescentis CCFM1386 can significantly increase the collagen content in the skin of aging mice.

[0124] Example 6: Effects of Bifidobacterium adolescentis CCFM1386 and its prepared postbiotics on collagen synthase activity in aging mouse skin

[0125] The animal experiment design, gavage groups, and preparation of skin homogenate samples used in the ELISA kits involved in the following examples are the same as those in Example 5.

[0126] LH1 can catalyze the hydroxylation of lysine residues, thereby providing sites for glycosylation and affecting the assembly and cross-linking of collagen fibers; P4H can catalyze the hydroxylation of proline residues, promoting the correct folding and structural stability of collagen; LOX can catalyze the oxidative deamination reaction of lysine and hydroxylysine residues to form aldehyde derivatives, and condense with adjacent lysine amino groups or hydroxyl groups of hydroxylysine to form covalent cross-links, thereby stabilizing the structure of the fibrils. Both are key catalytic enzymes in the process of collagen synthesis. The enzymatic activities of LH1, P4H and LOX in mouse skin were detected by ELISA kits, and the results are as follows. Figure 5 shown.

[0127] Testing of LH1, P4H, and LOX enzyme activities revealed significant decreases in the model group compared to the blank group. Oral administration of CCFM1386-S significantly increased LH1 activity to 13.01 U / mg compared to 8.58 U / mg in the model group; P4H activity to 30.45 U / mg compared to 17.00 U / mg in the model group; and LOX activity to 13.43 U / mg, significantly higher than the model group's activity of 8.48 U / mg.

[0128] The above results show that the postbiotics (fermentation supernatant) prepared by Bifidobacterium adolescentis CCFM1386 can increase the enzyme activities of LH1, P4H and LOX in the skin of aging mice and promote the synthesis of collagen.

[0129] Example 7: Effects of Bifidobacterium adolescentis CCFM1386 and its prepared postbiotics on the antioxidant capacity of aging mouse skin

[0130] The animal experiment design, gavage groups, and preparation of skin homogenate samples used in the ELISA kits involved in the following examples are the same as those in Example 5.

[0131] The activities of superoxide dismutase (SOD) and catalase (CAT) in mouse skin were detected by ELISA kits. Figure 6 shown.

[0132] Regarding SOD enzyme activity, the SOD enzyme activity in the control group was 15.87 U / mg, while the SOD enzyme activity in the model group skin significantly decreased to 7.35 U / mg. Oral administration of CCFM1386-S increased it to 11.81 U / mg, and oral administration of CCFM1386-J increased it to 12.31 U / mg. Both were able to significantly increase SOD enzyme activity compared to the model group.

[0133] Compared with the control group (37.38 U / μg), the CAT activity in the model group decreased significantly to 16.32 U / μg. Oral administration of CCFM1386-S and CCFM1386-J increased the CAT activity in the skin to 23.62 U / μg and 25.10 U / μg, respectively, which was significantly higher than that in the model group.

[0134] Oxidative stress has an important impact on collagen synthesis and stability in skin tissue. SOD can catalyze the oxidation of superoxide anions (O2 - ) is converted into hydrogen peroxide (H2O2), and then CAT quickly decomposes H2O2 into water and oxygen, thereby effectively preventing ROS accumulation in cells, reducing the damage to collagen molecules caused by oxidative stress, and thus protecting the collagen structure.

[0135] In summary, the postbiotics (inactivated bacteria and supernatant) prepared from Bifidobacterium adolescentis CCFM1386 can increase the enzyme activities of CAT and SOD in the skin of aging mice, improve the antioxidant capacity of the skin, and increase the content and stability of collagen in the host skin.

[0136] Example 8: Effects of Bifidobacterium adolescentis CCFM1386 and its prepared postbiotics on inflammatory levels in aging mice

[0137] The animal experimental design, gavage groups and serum sample preparation involved in the following examples are the same as those in Example 5.

[0138] The levels of C-reactive protein (CRP) in mouse serum were detected by biochemical analyzer, and the levels of interleukin-6 (IL-6) and tumor necrosis factor α (TNF-α) in mouse skin were detected by ELISA kit. Figure 7 shown.

[0139] Elevated levels of CRP, an acute phase protein, typically reflect worsening systemic inflammation. The inflammatory environment can accelerate collagen degradation and inhibit collagen synthesis. The CRP level in the blank group was 4.58 mg / L, while that in the model group was 5.18 mg / L. Oral administration of CCFM1386-J significantly reduced serum CRP levels to 4.98 mg / L.

[0140] IL-6 is an important pro-inflammatory cytokine that can promote the expression of matrix metalloproteinases (MMPs) by activating signaling pathways, thereby destroying collagen fiber structure. Compared with the blank group (10.54 μg / mg), the IL-6 content in the skin of the model group increased significantly to 24.06 μg / mg. Oral administration of CCFM1386-J significantly reduced the IL-6 content in the skin to 16.89 μg / mg.

[0141] Tumor necrosis factor alpha (TNF-α), a potent pro-inflammatory cytokine, not only induces collagen degradation but also inhibits fibroblast function and reduces collagen regeneration. Compared to the blank control group (54.59 ng / mg), the TNF-α level in the model group's skin increased significantly to 112.29 μg / mg. Oral administration of CCFM1386-J significantly reduced skin TNF-α levels to 82.95 μg / mg.

[0142] In summary, oral administration of CCFM1386-J can alleviate the inflammation of aging mice to varying degrees and affect the synthesis and stability of collagen.

[0143] Example 9: Effects of Bifidobacterium adolescentis CCFM1386 and its postbiotics on the water content of the stratum corneum and skin elasticity of aging mice

[0144] The animal experiment design and gavage groups involved in the following examples are the same as those in Example 5. At the end of the experiment, the skin moisture tester (with stratum corneum moisture measurement probe Corneometer CM825) of German CK Company was used to detect the stratum corneum moisture content and the elasticity of the mouse back skin. The results are as follows: Figure 8 shown.

[0145] The moisture content of the stratum corneum is determined by Figure 8 It can be seen that compared with 78.83% in the blank group, the water content in the model group was significantly reduced to 52.87%. The water content in the stratum corneum of the oral CCFM1386-S group (71.99%) increased by 19.12% compared with the model group, and the water content in the CCFM1386-J group (68.43%) increased by 15.56% compared with the model group.

[0146] Skin elasticity R2 is determined by Figure 8 It can be seen that compared with the blank group (80.80%), the skin elasticity of the model group was significantly reduced to 54.17%, the skin elasticity of the CCFM1386-S group (79.53%) increased by 25.36% compared with the model group, and the skin elasticity of the CCFM1386-J group (75.20%) increased by 21.03% compared with the model group.

[0147] Collagen is the main component that maintains the skin's structure and elasticity. Its loss can weaken the skin's support and reduce its elasticity, leading to signs of aging such as wrinkles and sagging. At the same time, a decrease in collagen can also affect the skin's ability to retain water, causing it to lose moisture, become dry and rough, and further exacerbate skin aging.

[0148] From the above experimental results, it can be seen that the postbiotics (inactivated bacteria and supernatant) prepared from Bifidobacterium adolescentis CCFM1386 can significantly increase the moisture content and elasticity of aging skin.

[0149] Example 10: MASSON staining analysis of skin tissue sections of aging mice using Bifidobacterium adolescentis CCFM1386 and its prepared postbiotics

[0150] The animal experiment design and gavage groups involved in the following examples are the same as those in Example 5. At the end of the experiment, the mice were killed and the skin of the shaved back was removed for MASSON staining. The results are as follows: Figure 9 shown.

[0151] It can be clearly seen from the slice results that the collagen fibers in the dermis of the control group mice are tightly and orderly arranged, stained darker blue, and have a higher collagen content. The fiber bundle structure is complete and evenly distributed, with no obvious breakage or degeneration. In addition, the dermis and epidermis of the skin in the control group are tightly connected, showing good skin barrier function and elastic support. The collagen fibers in the dermis of the model group mice were stained significantly lighter, arranged loosely and disorderly, with fiber breakage and faults, and the collagen content was significantly reduced. Compared with the model group, the postbiotics of CCFM1386 significantly improved the collagen content and structure of mouse skin.

[0152] Although the present invention has been disclosed above in terms of preferred embodiments, it is not intended to limit the present invention. Anyone familiar with this technology can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be based on the definition of the claims.

Claims

1. A strain of Bifidobacterium adolescentis CCFM1386, characterized in that The Bifidobacterium adolescentis CCFM1386 was deposited in the Guangdong Provincial Microbiological Culture Collection Center on January 10, 2025, with the deposit number: GDMCC No: 65746.

2. A microbial preparation containing the Bifidobacterium adolescentis CCFM1386 according to claim 1.

3. A postbiotic, characterized in that: A postbiotic prepared using the Bifidobacterium adolescentis CCFM1386 of claim 1 or the microbial preparation of claim 2.

4. The postbiotic according to claim 3, characterized in that The postbiotics include bacterial lysates, inactivated or inactivated cells, fermentation supernatants, or powders prepared by drying any of the above.

5. The postbiotic according to claim 4, characterized in that The inactivated or inactivated cells are prepared as follows: after culturing the Bifidobacterium adolescentis CCFM1386 in a culture medium, collecting bacterial cells in the cell culture medium, and subjecting the culture medium to heat treatment to obtain inactivated bacterial cells; The preparation method of the bacterial lysate is as follows: after culturing the Bifidobacterium adolescentis CCFM1386 in a culture medium, collecting bacterial cells, homogenizing under high pressure, and obtaining the bacterial lysate from the supernatant of the centrifugation; The fermentation supernatant is the supernatant obtained by centrifuging the Bifidobacterium adolescentis CCFM1386 in a culture medium.

6. A product containing the Bifidobacterium adolescentis CCFM1386 according to claim 1 and / or the postbiotic according to any one of claims 3 to 5, characterized in that: The products include food, medicine, health products or daily chemical products.

7. Use of the Bifidobacterium adolescentis CCFM1386 according to claim 1, the microbial preparation according to claim 2, or the postbiotic according to any one of claims 3 to 5 in the preparation of a product that promotes collagen synthesis in skin fibroblasts, characterized in that: The product includes at least one of the following effects: (1) Increase the collagen content secreted by skin fibroblasts in vitro; (2) Increase the activity of collagen synthase in skin fibroblasts in vitro.

8. Use of the Bifidobacterium adolescentis CCFM1386 according to claim 1, the microbial preparation according to claim 2, or the postbiotic according to any one of claims 3 to 5 in the preparation of anti-aging drugs.

9. Use according to claim 8, characterized in that The application includes at least one of the following functions: (1) Increase the moisture and elasticity of skin tissue in aging individuals; (2) Increase the antioxidant enzyme activity in the skin tissue of aging individuals; (3) reduce inflammation levels in aging individuals; (4) Increase collagen content in skin tissue of aging individuals; (5) Increase the activity of collagen synthase in the skin tissue of aging individuals.

10. A method for promoting collagen synthesis by skin fibroblasts in vitro, characterized in that: The method comprises mixing the postbiotic according to any one of claims 3 to 5 with skin fibroblasts and incubating the mixture.

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