Probiotic metagen preparation for improving skin barrier and skin micro-ecological environment as well as preparation method and application of probiotic metagen preparation

Abstract

The invention discloses a probiotic post-generation preparation capable of improving a skin barrier and a skin micro-ecological environment as well as a preparation method and application of the probiotic post-generation preparation. The bacterial strains in the probiotic post-generation preparation comprise lactobacillus plantarum MH95 of which the preservation number is CGMCC (China General Microbiological Culture Collection Center) No.31053 and lactobacillus paracasei FB97 of which the preservation number is CGMCC No.30464. The invention further discloses a preparation method of the probiotic post-generation preparation capable of improving the skin barrier and the skin micro-ecological environment and a preparation method and application of the probiotic post-generation preparation capable of improving the skin barrier and the skin micro-ecological environment of the probiotic post-generation preparation capable of improving the skin barrier and the skin micro-ecological environment of the probiotic post-generation preparation capable of improving the skin barrier and the skin micro-ecological environment. MH95 and FB97 are compounded to prepare a related metagen preparation, potential interaction exists between MH95 and FB97, and MH95 and FB97 can be mutually matched and synergistically interacted, specifically, expression of fibronectin (FLG) and LOR in a damaged skin barrier is improved, expression of related inflammatory factors is inhibited, and the skin micro-ecological environment is regulated and controlled.

Description

Technical Field

[0001] The present invention belongs to the field of microbial technology, and in particular relates to a probiotic postbiotic preparation for improving skin barrier and skin microecological environment, and a preparation method and application thereof. Background Art

[0002] The skin barrier is a structural barrier formed by lipids between epidermal cells and keratin in the stratum corneum. The skin barrier can prevent the body from releasing too much water and prevent harmful substances such as chemicals or microorganisms from entering our body. Damage to the skin barrier will cause dry skin, skin aging, skin sensitivity such as photodermatitis, irritant dermatitis, hormone-dependent dermatitis and other diseases. Atopic dermatitis (AD) is a chronic inflammatory skin disease caused by impaired skin barrier function, which manifests as dry skin, itching, erythema and inflammation. The filaggrin gene (FLG) is a key protein in maintaining the skin barrier function. Its mutation leads to serious defects in the skin barrier function, increasing skin water loss and penetration of external irritants. Loricrin (LOR) is the most abundant protein in the keratin envelope, accounting for about 70% to 80% of the total protein, and is an important protein in maintaining the skin barrier. When the skin barrier function is damaged, the response of the immune system will also be affected. When the skin barrier is damaged, the chance of exposure to external irritants and microorganisms increases, stimulating the immune cells in the skin (such as keratinocytes, Langerhans cells, etc.) to excessively secrete inflammatory factors, triggering a series of inflammatory reactions.

[0003] The skin lesions of patients with atopic dermatitis are inhabited by a large number of Staphylococcus aureus, whose metabolites and toxins further damage the skin barrier through various mechanisms (such as superantigen action, toxin action, biofilm formation, etc.), aggravate the inflammatory response, increase the risk of infection, and affect the treatment effect. Therefore, improving the skin barrier and eliminating Staphylococcus aureus have become key links in the treatment of patients with atopic dermatitis.

[0004] At present, the technical means for repairing the skin barrier and treating dermatitis mainly include microneedle technology, laser treatment technology, electroporation technology, skin transplantation surgery and antibiotics and other drug treatments. Although microneedles, lasers and skin transplants have significant effects, they are expensive and have a long recovery period, which is not conducive to their use and popularization in daily life. Although antibiotics such as mupirocin and neomycin can quickly relieve symptoms of inflammation and infection, long-term use may cause skin atrophy, pigmentation and other problems. Probiotic products applied on the skin can balance the flora of the skin microecological environment and promote skin barrier repair. Therefore, it is of great significance to develop a probiotic preparation product that can effectively improve the skin barrier and skin microecological environment. Summary of the invention

[0005] In view of the deficiencies in the prior art, the object of the present invention is to provide a probiotic postbiotic preparation for improving the skin barrier and skin microecological environment, and a preparation method and application thereof.

[0006] In order to achieve the purpose of the invention, the present invention adopts the following technical solutions:

[0007] In a first aspect, the present invention provides a probiotic postbiotic preparation for improving skin barrier and skin microecological environment, wherein the strains in the probiotic postbiotic preparation include Lactobacillus plantarum MH95 with a preservation number of CGMCC No.31053 and Lactobacillus paracasei FB97 with a preservation number of CGMCC No.30464.

[0008] The present invention develops a new probiotic compounding method and a new strategy for improving the skin barrier and skin microecological environment, that is, Lactobacillus plantarum MH95 and Lactobacillus paracasei FB97 are compounded to prepare relevant postbiotic preparations, and it is found that there is a potential interaction between the two, which can cooperate with each other and synergize. When the amount of bacteria is consistent, it has a more significant and excellent effect than a single postbiotic or other postbiotic compounding method. It is specifically manifested in improving the expression of filaggrin (FLG) and loricrin (LOR) in damaged skin barriers, inhibiting the expression of related inflammatory factors, and regulating the skin microecological environment.

[0009] Preferably, the ratio of the bacterial counts of Lactobacillus plantarum MH95 and Lactobacillus paracasei FB97 is 1:10-10:1, for example, 1:10, 1:9, 1:8, 1:7, 1:6, 1:5, 1:4, 1:3, 1:2, 1:1, 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, etc. Other specific point values ​​within this numerical range can be selected, and they will not be described one by one here.

[0010] Preferably, the total concentration of the strain in the probiotic postbiotic preparation is not less than 1×10 6 / mL or 1×10 6 Pieces / g, for example 1×10 6 pieces / mL(pieces / g), 2×10 6 pieces / mL(pieces / g), 5×10 6 pieces / mL(pieces / g), 8×10 6 pieces / mL(pieces / g), 1×10 7pieces / mL(pieces / g), 5×10 7 pieces / mL(pieces / g), 1×10 8 Pieces / mL (pieces / g), etc. Other specific point values ​​within this numerical range can be selected and will not be described here one by one.

[0011] Preferably, the dosage form of the probiotic postbiotic preparation includes liquid, powder, capsule, tablet or granule.

[0012] Preferably, the probiotic postbiotic preparation further comprises a protective agent.

[0013] Preferably, the protective agent comprises any one of skim milk, sucrose, lactose, trehalose, dextran, gelatin, dextrin, gum arabic, sodium alginate, polyvinyl pyrrolidone, sorbitol or xylitol, or a combination of at least two thereof.

[0014] In a second aspect, the present invention provides a method for preparing a probiotic postbiotic preparation according to the first aspect, wherein the probiotic postbiotic preparation is a solution preparation method comprising: inoculating activated Lactobacillus plantarum MH95 and activated Lactobacillus paracasei FB97 into a culture medium, respectively, and culturing at 35-38° C. for 18-30 hours to obtain a bacterial solution, mixing the two bacterial solutions according to a bacterial count ratio, adding a protective agent, and sterilizing to obtain the probiotic postbiotic preparation;

[0015] The above-mentioned "35-38°C" may be, for example, 35°C, 35.2°C, 35.5°C, 35.8°C, 36°C, 36.2°C, 36.5°C, 36.8°C, 37°C, 37.2°C, 37.5°C, 37.8°C, 38°C, etc.;

[0016] The above-mentioned "18-30h" can be, for example, 18h, 19h, 20h, 21h, 22h, 23h, 24h, 25h, 26h, 27h, 28h, 29h, 30h, etc. Other specific point values ​​within the numerical range can be selected, and they will not be described one by one here.

[0017] Preferably, the preparation method of the probiotic postbiotic preparation in the form of a powder comprises: inoculating activated Lactobacillus plantarum MH95 and activated Lactobacillus paracasei FB97 into a culture medium respectively, culturing at 35-38° C. for 18-30 hours to obtain a bacterial solution, mixing the two bacterial solutions according to a bacterial count ratio, adding a protective agent, and then sterilizing and drying to obtain the probiotic postbiotic preparation;

[0018] The above-mentioned "35-38°C" may be, for example, 35°C, 35.2°C, 35.5°C, 35.8°C, 36°C, 36.2°C, 36.5°C, 36.8°C, 37°C, 37.2°C, 37.5°C, 37.8°C, 38°C, etc.;

[0019] The above-mentioned "18-30h" can be, for example, 18h, 19h, 20h, 21h, 22h, 23h, 24h, 25h, 26h, 27h, 28h, 29h, 30h, etc. Other specific point values ​​within the numerical range can be selected, and they will not be described one by one here.

[0020] Preferably, the sterilization temperature is 70-130°C, for example, it can be 70°C, 75°C, 80°C, 85°C, 90°C, 95°C, 100°C, 105°C, 110°C, 115°C, 120°C, 125°C, 130°C, etc.; the time is 15-40min, for example, it can be 15min, 16min, 18min, 20min, 22min, 25min, 28min, 30min, 32min, 35min, 38min, 40min, etc., and other specific point values ​​within the numerical range can be selected, which will not be elaborated here.

[0021] In a third aspect, the present invention provides a use of the probiotic postbiotic preparation according to the first aspect in preparing a product having the function of improving the skin barrier and / or improving the skin microecological environment;

[0022] The products include medicines, foods, health products, daily chemical products or household textiles.

[0023] Compared with the prior art, the present invention has the following beneficial effects:

[0024] The present invention develops a new probiotic compounding method and a new strategy for improving the skin barrier and skin microecological environment, that is, Lactobacillus plantarum MH95 and Lactobacillus paracasei FB97 are compounded to prepare relevant postbiotic preparations, and it is found that there is a potential interaction between the two, which can cooperate with each other and synergize. When the amount of bacteria is consistent, it has a more significant and excellent effect than a single postbiotic or other postbiotic compounding method. It is specifically manifested in improving the expression of filaggrin (FLG) and loricrin (LOR) in damaged skin barriers, inhibiting the expression of related inflammatory factors, and regulating the skin microecological environment.

[0025] The classification name of MH95 involved in the present invention is Lactiplantibacillus plantarum, the preservation unit is the General Microbiology Center of China Microbiological Culture Collection Administration, the preservation time is June 24, 2024, the preservation number is CGMCC No.31053, and the address is: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

[0026] The FB97 involved in the present invention is classified and named as Lactobacillus paracasei, and the deposit unit is the General Microbiology Center of China Microorganism Culture Collection Administration, the deposit time is April 28, 2024, the deposit number is CGMCC No.30464, and the address is: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing. DETAILED DESCRIPTION

[0027] In order to further explain the technical means and effects adopted by the present invention, the technical solution of the present invention is further described below in conjunction with the preferred embodiments of the present invention, but the present invention is not limited to the scope of the embodiments.

[0028] The classification name of MH95 mentioned below is Lactiplantibacillus plantarum, and the deposit number is CGMCC No.31053;

[0029] The FB97 referred to below is named as Lacticaseibacillus paracasei, and its deposit number is CGMCC No.30464;

[0030] The ATCC 8014 bacteria referred to below is Lactobacillus plantarum ATCC 8014.

[0031] The following lipopolysaccharides (LPS, Escherichiacoli055:B5) were purchased from Beijing Solebow Technology Co., Ltd.

[0032] RAW264.7 mouse macrophages were purchased from the cell bank of the Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences;

[0033] DMEM medium was purchased from Gibco Advanced DMEM from Thermo Fisher Scientific China Co., Ltd.

[0034] The components of the MRS medium involved below are: yeast powder 5.0 g / L, beef extract 10.0 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 trihydrate 2.6 g / L, manganese sulfate monohydrate 0.25 g / L, magnesium sulfate heptahydrate 0.5 g / L and Tween-80 1 mL / L, pH 6.2-6.4.

[0035] The preparation method of the activated bacterial suspension involved below is to inoculate Lactobacillus plantarum MH95, Lactobacillus paracasei FB97 and ATCC 8014 strains into MRS culture medium at a concentration of 2%, and then culture them at 37° C. with shaking for 24 hours to obtain activated MH95 bacterial solution, activated FB97 bacterial solution and activated ATCC 8014 bacterial solution.

[0036] The cell culture medium mentioned below is fibroblast culture medium, purchased from Shenzhen Zike Biotechnology Co., Ltd., model number GMS12049.1;

[0037] The in vitro reconstructed epidermis model and supporting culture medium involved in the following examples were purchased from Guangdong Boxi Biotechnology Co., Ltd., and the model is 3D epidermis model EpiKutis.

[0038] Example 1

[0039] The present embodiment provides a probiotic postbiotic preparation, the preparation method of which is to mix the activated MH95 bacterial solution and the activated FB97 bacterial solution in a bacterial count ratio of 3:1 to obtain a mixed bacterial solution. Use NaOH to adjust the pH of the mixed bacterial solution to 7.0, centrifuge at 5000rpm for 5 minutes, discard the supernatant, and collect the centrifugal precipitate (bacterial mud). Add a protective agent to the bacterial mud in proportion (bacterial mud: 30% trehalose solution = 1:1, W / W), stir continuously at 80°C and 200rpm, maintain for 30 minutes to complete inactivation, and obtain an inactivated emulsion; spray dry the inactivated emulsion and crush it to obtain a total bacterial concentration of 1×10 7 / g of inactivated bacterial powder to obtain the probiotic postbiotic preparation.

[0040] Example 2

[0041] The present embodiment provides a probiotic postbiotic preparation, the preparation method of which is to mix the activated MH95 bacterial solution and the FB97 bacterial solution in a bacterial count ratio of 1:1 to obtain a mixed bacterial solution. Use NaOH to adjust the pH of the mixed bacterial solution to 7.0, centrifuge at 5000rpm for 5 minutes, discard the supernatant, and collect the centrifugal precipitate (bacterial mud). Add a protective agent to the bacterial mud in proportion (bacterial mud: 30% sucrose solution = 1:1, W / W), stir continuously at 100°C and 250rpm, maintain for 20 minutes to complete inactivation, and obtain an inactivated emulsion. Spray dry the inactivated emulsion and crush it to obtain a total bacterial concentration of 5×10 6 / g of inactivated bacterial powder to obtain the probiotic postbiotic preparation.

[0042] Example 3

[0043] The present embodiment provides a probiotic postbiotic preparation, the preparation method of which is to mix the activated MH95 bacterial solution and the FB97 bacterial solution in a bacterial count ratio of 1:6 to obtain a mixed bacterial solution. Use NaOH to adjust the pH of the mixed bacterial solution to 7.0, centrifuge at 5000rpm for 5 minutes, discard the supernatant, and collect the centrifugal precipitate (bacterial mud). Add a protective agent to the bacterial mud in proportion (bacterial mud: 30% lactose solution = 1:1, W / W), stir continuously at 120°C and 180rpm, maintain for 18 minutes to complete inactivation, and obtain an inactivated emulsion. Spray dry the inactivated emulsion and crush it to obtain a total bacterial concentration of 2×10 6 / g of inactivated bacterial powder to obtain the probiotic postbiotic preparation.

[0044] Comparative Example 1

[0045] This comparative example provides a probiotic postbiotic preparation, the preparation method of which is different from that of Example 1 in that the activated MH95 bacterial solution is replaced with an ATCC 8014 bacterial solution of equal bacterial count, and subsequent treatment is performed to obtain a total bacterial concentration of 1×10 7 / g of inactivated bacterial powder, that is, probiotic postbiotic preparation.

[0046] Comparative Example 2

[0047] This comparative example provides a probiotic postbiotic preparation, the preparation method of which is different from that of Example 1 in that FB97 bacterial solution is not added and the reduced bacterial count is proportionally distributed to MH95 bacterial solution, and subsequent treatment is performed to obtain a total bacterial concentration of 1×10 7 / g of inactivated bacterial powder, that is, probiotic postbiotic preparation.

[0048] Comparative Example 3

[0049] This comparative example provides a probiotic postbiotic preparation, the preparation method of which is different from that of Example 1 in that MH95 bacterial solution is not added and the reduced bacterial count is proportionally distributed to FB97 bacterial solution, and subsequent treatment is performed to obtain a total bacterial concentration of 1×10 7 / g of inactivated bacterial powder, that is, probiotic postbiotic preparation.

[0050] Comparative Example 4

[0051] This comparative example provides a probiotic postbiotic preparation, the preparation method of which is different from that of Example 1 in that the activated MH95 bacterial solution and the activated FB97 bacterial solution are respectively replaced with ATCC 8014 bacterial solution with the same bacterial count, and the total bacterial concentration is obtained by subsequent treatment. 7 / g of inactivated bacterial powder, that is, probiotic postbiotic preparation.

[0052] Test Example 1

[0053] Anti-inflammatory experiments

[0054] Sample treatment: The probiotic postbiotic preparations prepared in Examples 1-3 and Comparative Examples 1-4 were mixed with deionized water to prepare a 50 μg / mL postbiotic preparation solution for testing.

[0055] Test method:

[0056] (1) RAW264.7 mouse macrophages in the logarithmic growth phase with good morphology were selected and inoculated in DMEM culture medium containing 10% FBS fetal bovine serum and incubated for 24 h. 5 The cells were seeded at a density of cells / well in a 24-well plate and incubated in an incubator at 37°C and 5% CO2 for 2 hours. The cells were stimulated with 1 μg / mL lipopolysaccharide for 24 hours to establish a cellular inflammation model.

[0057] (2) 100 μL of the above-treated probiotic postbiotic preparation sample solution was added to each group of cells. After incubation at 37°C for 24 h, the cell supernatant was collected and tested according to the instructions of the ELISA kit and the nitric oxide detection kit. The contents of α-tumor necrosis factor (TNF-α), nitric oxide (NO), interleukin-1β (IL-1β) and interleukin-6 (IL-6) in the collected cell supernatant were detected respectively. The experimental results were analyzed by data processing software. The results are shown in Table 1.

[0058] Table 1

[0059]

[0060]

[0061] Macrophages are important effector cells of the natural immune response. Bacterial lipopolysaccharide is an effective substance that stimulates macrophages to release inflammatory mediators. Bacterial lipopolysaccharide binds to macrophage surface antigen recognition receptors, inducing macrophages to secrete proinflammatory cytokines (TNF-α, NO, IL-1β, IL-6, etc.) and inflammatory mediators (carbon monoxide, etc.) and related enzymes, further triggering cascade reactions, leading to tissue injury. Therefore, by constructing an inflammatory model of related macrophages, using a probiotic postbiotic preparation to treat the relevant model, the content changes of related proinflammatory cytokines are detected to verify the anti-inflammatory effect of the relevant products. It can be seen from the data in the table above that the samples in each experimental group have different degrees of inhibitory effects on proinflammatory factors, and the MH95 strain and the FB97 strain have significant inhibitory effects on the four proinflammatory mediators after being used in combination, proving that the probiotic postbiotic preparation provided by the present invention has excellent proinflammatory effects.

[0062] Test Example 2

[0063] Skin barrier improvement experiment

[0064] Test method:

[0065] (1) The in vitro reconstructed epidermal model was revived and cultured normally in a CO2 incubator (37°C, 5% CO2) using the matching culture medium according to the method in the instruction manual.

[0066] (2) Sodium dodecyl sulfate powder was mixed with epidermal model liquid culture medium to prepare a sodium dodecyl sulfate solution with a mass concentration of 0.1%. 25 μL of the sodium dodecyl sulfate solution was evenly applied on the surface of the resuscitated epidermal model to establish a 3D epidermal damage model (control group).

[0067] (3) Sodium dodecyl sulfate powder and postbiotic preparation powder (Example 1-3 and Comparative Example 1-4) were mixed with the epidermal model liquid culture medium, and the final system had a sodium dodecyl sulfate mass concentration of 0.1% and a postbiotic preparation mass concentration of 5%. 25 μL of the above system was evenly applied on the surface of the resuscitated epidermal model (Example 1-3 and Comparative Example 1-4).

[0068] (4) The control group, Examples 1-3 and Comparative Examples 1-4 were placed in a CO2 incubator (37°C, 5% CO2) and incubated for 24 hours. The models were then washed with PBS solution and fixed with 4% paraformaldehyde. After fixation for 24 hours, the models were removed by circumcision and the expression of filaggrin (FLG) and loricrin (LOR) was detected by immunofluorescence method. The increase rate of protein expression level in each test group compared with that in the control group was calculated. The results are shown in Table 2.

[0069] Table 2

[0070] FLG expression level increase rate Increase rate of LOR expression level Example 1 52.69％ 63.04％ Example 2 50.86％ 48.89％ Example 3 47.73％ 50.9％ Comparative Example 1 28.11％ 10.59％ Comparative Example 2 38.61％ 40.89％ Comparative Example 3 22.44％ 9.43％ Comparative Example 4 20.29％ 11.94％

[0071] Some highly irritating test substances or ultraviolet rays can cause clinical acute damage to the skin barrier, leading to dry skin and erythema. The anionic surfactant sodium dodecyl sulfate has amphiphilic (hydrophilic and lipophilic) characteristics. It can damage the skin barrier after contacting the skin at a specific concentration, and obtain a related epidermal damage model, which is used to test the repair and improvement effects of related samples on damaged skin barriers. Filaggrin and loricrin are important keratinized envelope structural proteins, which play an important role in resisting external stimuli of the skin and protecting the skin barrier. The efficacy of related products is evaluated by the changes in barrier-related protein indicators.

[0072] From the data in the table, we can see that the samples in each experimental group can increase the expression of epidermal filaggrin and loricrin to varying degrees, thereby improving and repairing the skin barrier; and when the MH95 strain is used in combination with the FB97 strain, it has a significant improvement on the relevant skin barrier, with the best effect.

[0073] Test Example 3

[0074] Skin microecological environment flora regulation test

[0075] Preparation of test bacterial solution:

[0076] ① Staphylococcus aureus (ATCC 6538): After thawing the freeze-dried bacterial powder at 25℃, evenly inoculate it on the surface of MRS solid culture medium. After constant temperature culture at 37℃ for 24h, take an appropriate amount of bacterial cells and transfer them to MRS liquid culture medium and continue to culture at 37℃ for 24h. Adjust the concentration of bacterial suspension to 10 6 CFU / mL.

[0077] ② Staphylococcus epidermidis (ATCC 29213): After thawing the freeze-dried bacterial powder at 25℃, evenly inoculate it on the surface of MRS solid culture medium. After constant temperature culture at 37℃ for 24h, take an appropriate amount of bacterial cells and transfer them to MRS liquid culture medium and continue to culture at 37℃ for 24h. Adjust the concentration of bacterial suspension to 10 6 CFU / mL.

[0078] Preparation of probiotic postbiotic preparation culture medium:

[0079] When preparing the MRS liquid culture medium, the probiotic postbiotic preparations prepared in Examples 1-3 and Comparative Examples 1-4 were added to the culture medium at a concentration of 1 mg / mL, and the culture medium was sterilized at 121° C. for 20 min before use.

[0080] Test method: The prepared Staphylococcus aureus suspension and Staphylococcus epidermidis suspension were inoculated into MRS liquid culture medium containing probiotic postbiotic preparations at a volume fraction of 7% as the experimental group, and the same inoculation amount was inoculated into MRS culture medium without probiotic postbiotic preparations as the control group. All groups were cultured at 37°C at a rotation speed of 120 rpm for 18 hours, diluted for plate counting, and the bacterial inhibition / promotion rate was calculated according to the following formula.

[0081] X(%)=[(C0-C1) / C0]×100%

[0082] Wherein, X is the bacterial inhibition / promotion rate. When X is a positive number, it means that this group of samples has an inhibitory effect on the relevant test bacteria; when X is a negative number, it means that this group of samples has a proliferation-promoting effect on the relevant test bacteria; C0 is the test bacteria concentration of the control group, and C1 is the test bacteria concentration of the experimental group.

[0083] The results are shown in Table 3

[0084] Table 3

[0085] Staphylococcus aureus inhibition rate Staphylococcus epidermidis growth promotion rate Example 1 90％ -342.11％ Example 2 81.2％ -184.21％ Example 3 80.4％ -154.14％ Comparative Example 1 56.78％ -25.44％ Comparative Example 2 70.25％ -125.56％ Comparative Example 3 59.37％ -41.45％ Comparative Example 4 55.67％ -4.14％

[0086] The metabolites and toxins produced by Staphylococcus aureus can damage the skin barrier, aggravate inflammatory responses, and disrupt the balance of the skin's microecological environment; while Staphylococcus epidermidis and Staphylococcus aureus can compete with each other, and increasing the number of Staphylococcus epidermidis can inhibit the proliferation of Staphylococcus aureus, thereby maintaining skin health by regulating the balance of the skin's microecological environment.

[0087] From the data in the table, it can be seen that the samples in each experimental group can inhibit Staphylococcus aureus to varying degrees while promoting the proliferation of Staphylococcus epidermidis. Among them, the combination of MH95 strain and FB97 strain has a better effect, which can significantly increase the number of Staphylococcus epidermidis and inhibit the proliferation of Staphylococcus aureus, thereby maintaining skin health by regulating the balance of skin microecology.

[0088] Test Example 4

[0089] Product safety testing

[0090] According to the subject inclusion criteria, 15 women and 15 men were selected to participate in the experiment and randomly divided into 3 groups (5 men and 5 women in each group), and the probiotic postbiotic preparation products of Examples 1-3 (the postbiotic preparation was mixed with pure water to adjust the mass concentration to 5% and then tested) were respectively accepted for trial. Use a syringe to take 0.025mL of sample and place it in the medicine chamber of the spot test tape. Immediately apply the spot test tape with the sample to the normal skin of the subject's left forearm, and gently press the tape with the palm of your hand to apply it evenly to the skin. The test site is marked for observation, and the patch is applied every 24 hours. Half an hour after the patch is removed, it is inspected according to the terminology of the CTFA guidelines. A total of 6 patch tests were conducted. On the sixth day after the patch was completed, an additional inspection was conducted to observe delayed reactions. The data were compared using the results of the last inspection, and the results are shown in Table 4.

[0091] Table 4

[0092]

[0093] It can be seen from the results in the table that the probiotic postbiotic preparation product prepared by the present invention has no irritation to the skin and no positive reaction, which proves that the product is highly safe, less irritating and relatively mild.

[0094] In summary, the present invention has developed a new probiotic compounding method and a new strategy for improving skin barrier and skin microecological environment, that is, Lactobacillus plantarum MH95 and Lactobacillus paracasei FB97 are compounded and prepared into relevant postbiotic preparations, and it is found that there is a potential interaction between the two, which can cooperate with each other and synergize. When the amount of bacteria is consistent, it has a more significant and excellent effect than a single postbiotic or other postbiotic compounding method. It is specifically manifested in improving the expression of filaggrin (FLG) and loricrin (LOR) in damaged skin barriers, inhibiting the expression of related inflammatory factors, and regulating the skin microecological environment.

[0095] The applicant declares that the present invention illustrates the technical solution of the present invention through the above-mentioned embodiments, but the present invention is not limited to the above-mentioned embodiments, that is, it does not mean that the present invention must rely on the above-mentioned embodiments to be implemented. Those skilled in the art should understand that any improvement of the present invention, equivalent replacement of various raw materials of the product of the present invention, addition of auxiliary components, selection of specific methods, etc., all fall within the protection scope and disclosure scope of the present invention.

[0096] The preferred embodiments of the present invention are described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the technical concept of the present invention, a variety of simple modifications can be made to the technical solution of the present invention, and these simple modifications all belong to the protection scope of the present invention.

[0097] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present invention will not further describe various possible combinations.

Claims

1. A probiotic postbiotic preparation for improving skin barrier and skin microecological environment, characterized in that: The strains in the probiotic postbiotic preparation include Lactobacillus plantarum MH95 with a preservation number of CGMCC No.31053 and Lactobacillus paracasei FB97 with a preservation number of CGMCC No.30464.

2. The probiotic postbiotic preparation according to claim 1, characterized in that The bacterial count ratio of Lactobacillus plantarum MH95 and Lactobacillus paracasei FB97 is 1:10-10:

1.

3. The probiotic postbiotic preparation according to claim 1 or 2, characterized in that: The total concentration of the strain in the probiotic postbiotic preparation is not less than 1×10 6 / mL or 1×10 6 Pieces / g.

4. The probiotic postbiotic preparation according to any one of claims 1 to 3, characterized in that The dosage form of the probiotic postbiotic preparation includes liquid, powder, capsule, tablet or granule.

5. The probiotic postbiotic preparation according to any one of claims 1 to 4, characterized in that The probiotic postbiotic preparation also includes a protective agent.

6. The probiotic postbiotic preparation according to claim 5, characterized in that The protective agent includes any one of skim milk, sucrose, lactose, trehalose, dextran, gelatin, dextrin, gum arabic, sodium alginate, polyvinyl pyrrolidone, sorbitol or xylitol, or a combination of at least two thereof.

7. A method for preparing a probiotic postbiotic preparation according to any one of claims 1 to 6, characterized in that: The preparation method of the probiotic postbiotic preparation in the form of a solution comprises: inoculating activated Lactobacillus plantarum MH95 and activated Lactobacillus paracasei FB97 into a culture medium respectively, culturing at 35-38° C. for 18-30 hours to obtain a bacterial solution, mixing the two bacterial solutions according to a bacterial count ratio, adding a protective agent, and sterilizing to obtain the probiotic postbiotic preparation.

8. The method for preparing the probiotic postbiotic preparation according to any one of claims 1 to 6, characterized in that: The preparation method of the probiotic postbiotic preparation in the form of powder comprises: inoculating activated Lactobacillus plantarum MH95 and activated Lactobacillus paracasei FB97 into a culture medium respectively, culturing at 35-38° C. for 18-30 hours to obtain bacterial liquid, mixing the two bacterial liquids according to the bacterial count ratio, adding a protective agent, sterilizing and drying to obtain the probiotic postbiotic preparation.

9. The preparation method according to claim 7 or 8, characterized in that: The sterilization temperature is 70-130° C. and the time is 15-40 minutes.

10. Use of the probiotic postbiotic preparation according to any one of claims 1 to 6 in the preparation of a product having the function of improving the skin barrier and / or improving the skin microecological environment; The products include medicines, foods, health products, daily chemical products or household textiles.

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