A strain of Lactobacillus paracasei and its application in anti-aging and anti-glycation

Abstract

The present invention belongs to the field of microbial technology, and specifically relates to a strain of Lactobacillus paracasei and its application in anti-aging and anti-glycation. The Lactobacillus paracasei provided by the present invention is Lactobacillus paracasei ( Lactobacillus paracasei ) BL-22, whose deposit number is CGMCC NO. 33475. Experiments in the present invention confirm that Lactobacillus paracasei BL-22 has significant effects on skin antioxidant protection, alleviating skin aging, and resisting glycation. At the same time, through animal experiments and research on the interaction of active ingredients, the interaction mechanism between Lactobacillus paracasei BL-22 and xylo-oligosaccharides was revealed, confirming that the combination of Lactobacillus paracasei BL-22 and xylo-oligosaccharides can exert a synergistic effect in alleviating skin aging and glycation. Therefore, the present invention can provide new research and development directions and technical support for the development of health products with anti-aging and anti-glycation effects.

Description

Technical Field

[0001] The invention belongs to the technical field of microorganisms, and particularly relates to a strain of Lactobacillus paracasei and application thereof in anti-aging and anti-glycation. Background Art

[0002] Skin aging is generally categorized as extrinsic and intrinsic. Extrinsic aging is primarily related to UV exposure, while intrinsic aging is closely linked to skin glycation. Glycation is primarily a non-enzymatic reaction between proteins (including collagen) and reducing sugars, generating irreversible, stable products such as advanced glycation end products (AGEs).

[0003] The extracellular matrix (ECM) is one of the primary targets of glycation reactions. Collagen reacts with sugars to form AGEs, which accumulate and cross-link with the ECM, leading to structural and functional damage to connective tissue. Furthermore, elastin and collagen, the primary structural molecules of the skin, can react with sugars through glycation, the covalent binding of sugars to proteins, and the subsequent destruction of AGEs, thereby exacerbating the aging process.

[0004] Currently, many active ingredients used for anti-aging and anti-glycation contain irritants such as retinol and acids. Long-term use can damage the skin barrier, causing skin allergies and inflammation. Therefore, developing more safe, natural active ingredients and related products that offer both excellent anti-aging and anti-glycation properties can provide a healthy and convenient new approach to addressing skin concerns. Summary of the Invention

[0005] In view of the deficiencies in the prior art, one of the objectives of the present invention is to provide a strain of Lactobacillus paracasei having anti-aging and anti-glycation effects.

[0006] The second object of the present invention is to provide the use of the above-mentioned Lactobacillus paracasei in the preparation of anti-aging and anti-glycation microbial preparations.

[0007] The third object of the present invention is to provide an anti-aging and anti-glycation microbial preparation.

[0008] A fourth object of the present invention is to provide an anti-aging and anti-glycation product.

[0009] To achieve the above object, the first aspect of the present invention provides a strain of Lactobacillus paracasei, which is named Lactobacillus paracasei BL-22 and is deposited in the General Microbiology Center of the China Culture Collection Administration of Microorganisms with a deposit number of CGMCC NO. 33475.

[0010] The second aspect of the present invention provides the use of the above-mentioned Lactobacillus paracasei in the preparation of anti-aging and anti-glycation microbial preparations.

[0011] The third aspect of the present invention provides an anti-aging and anti-glycation microbial preparation, wherein the microbial preparation comprises the Lactobacillus paracasei as described above.

[0012] In a preferred embodiment of the present invention, the microbial preparation further comprises xylooligosaccharides.

[0013] In a preferred embodiment of the present invention, the molecular weight of the xylooligosaccharide is 300-1050 Daltons.

[0014] In a preferred embodiment of the present invention, the ratio of the amount of Lactobacillus paracasei and xylooligosaccharide in the microbial preparation is (1×10 7 ~1×10 9 ) CFU: (100-500) mg. The present invention combines Lactobacillus paracasei BL-22 and xylooligosaccharides as an active ingredient, and the two can effectively enhance the anti-aging and anti-glycation effects of the skin through synergistic action.

[0015] In a further preferred embodiment of the present invention, the ratio of Lactobacillus paracasei to xylooligosaccharide in the microbial preparation is 1×10 8 CFU: 200mg.

[0016] The fourth aspect of the present invention provides an anti-aging and anti-glycation product, comprising an active component and excipients; the active component is the anti-aging and anti-glycation microbial preparation as described above.

[0017] In a preferred embodiment of the present invention, the product is one of a skin care product, a medicine, and a health food.

[0018] In a preferred embodiment of the present invention, the skin care product is one of skin care water, skin care lotion, essence, and face cream; the medicine is one of tablets, granules, oral liquid, and capsules; and the health food is one of pills, granules, powder, and oral liquid.

[0019] The present invention does not specifically limit the types of excipients. Technicians can select common excipient types based on the type of product preparation and preparation requirements. In a further preferred embodiment, when the product is a skin care product, the excipients used include one or more of a moisturizer, a pH adjuster, a thickener, and a preservative; moisturizers can be, for example, glycerin, hyaluronic acid, panthenol, propylene glycol, etc.; pH adjusters can be, for example, sodium lactate, arginine, aminomethyl propanol, etc.; thickeners can be, for example, xanthan gum, hydroxyethyl cellulose, carbomer, etc.; preservatives can be, for example, hydroxybenzoates, silicon dioxide, imidazolidinyl urea, etc. In a further preferred embodiment, when the product is a medicine or health food, the excipients used include one or more of a filler, a binder, a colorant, a disintegrant, and a preservative.

[0020] The technical solution of the present invention has the following advantages and beneficial effects:

[0021] The Lactobacillus paracasei BL-22 provided by the present invention is screened from traditional dairy products in Gannan Tibetan Autonomous Prefecture, Gansu Province. It has high biosafety. It has been verified that the strain not only has excellent DPPH free radical and hydroxyl free radical scavenging capabilities, exhibits outstanding antioxidant capacity, but also has excellent skin anti-aging and anti-glycation effects.

[0022] Furthermore, the present invention explored the effects of Lactobacillus paracasei BL-22 alone, oligoxylose alone, and the combined use of Lactobacillus paracasei BL-22 and oligoxylose on skin antioxidant, skin aging alleviation, and anti-glycation through animal experiments and studies on the interactions of active ingredients. The experiments showed that compared with the Lactobacillus paracasei BL-22 alone group and the oligoxylose alone group, the combined use of Lactobacillus paracasei BL-22 and oligoxylose in the present invention had the following outstanding effects: (1) significantly improved skin elasticity; (2) effectively increased the content of skin barrier-related proteins; (3) significantly reduced the level of inflammatory factor TNF-α; (4) exerted an effective anti-oxidative stress effect; (5) significantly reduced the content of AGEs and exerted an anti-glycation effect on the skin; (6) activated the TGF-β / Smad signaling pathway, promoted collagen synthesis, and effectively delayed skin aging.

[0023] Through comprehensive experimental research, the present invention reveals for the first time the interaction mechanism between Lactobacillus paracasei BL-22 and xylooligosaccharides, confirming that the combination of Lactobacillus paracasei BL-22 and xylooligosaccharides can exert a synergistic effect in alleviating skin aging and glycation. Therefore, the present invention can provide a new, healthy and convenient strategy for solving skin problems, and can also provide new research and development directions and technical support for the development of anti-aging and anti-glycation health products. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1The results of the comparison of the antioxidant capacity of different Lactobacillus paracasei in Experimental Example 1 of the present invention are shown in FIG. 1 , wherein (a) is the DPPH scavenging rate test result, and (b) is the hydroxyl radical scavenging ability test result;

[0025] Figure 2 The results of the skin elasticity test of mice in different groups in Experimental Example 2 of the present invention are as follows;

[0026] Figure 3 The test results of the expression levels of skin barrier-related proteins in different groups of mice in Experimental Example 2 of the present invention are as follows; (a) is the relative expression level of Loricrin protein, (b) is The relative expression of protein, (c) Relative expression of proteins;

[0027] Figure 4 These are the test results of the inflammatory factor TNF-α levels in the blood and skin of mice in different groups in Experimental Example 2 of the present invention; wherein (a) is the TNF-α content in the blood, and (b) is the TNF-α content in the skin;

[0028] Figure 5 The levels of SOD, GSH-Px, CAT, and MDA in the blood of mice in different groups in Experimental Example 2 of the present invention; wherein (a) is the SOD content, (b) is the GSH-Px content, (c) is the CAT content, and (d) is the MDA content;

[0029] Figure 6 The results of the AGE level test in the blood and skin of different groups of mice in Experimental Example 2 of the present invention are shown below; (a) is the AGE content in the blood, and (b) is the AGE content in the skin;

[0030] Figure 7 These are the SMAD2, SMAD3, TGF-β I and TGF-β II gene expression levels in the skin tissues of mice in different groups in Experimental Example 2 of the present invention; wherein, (a) is the SMAD2 gene expression level, (b) is the SMAD3 gene expression level; (c) is the TGF-β I gene expression level, and (b) is the TGF-β II gene expression level. DETAILED DESCRIPTION

[0031] In order to more clearly understand the above-mentioned purpose, features and advantages of the present invention, the present invention is further described in detail below in conjunction with specific embodiments. It should be noted that, in the case of no conflict, the embodiments of the present invention and the features in the embodiments can be combined with each other. In the following description, many specific details are set forth to facilitate a full understanding of the present invention, but the present invention can also be implemented in other ways different from those described herein. Therefore, the scope of protection of the present invention is not limited by the specific embodiments disclosed below. Under the premise of no conflict, the following technical features of the embodiments of the present invention can also be combined with each other.

[0032] In the present invention, the deposited information involved is as follows:

[0033] Deposit name: Lactobacillus paracasei BL-22, Latin name: Lactobacillus paracasei BL-22;

[0034] Deposit number: CGMCC NO. 33475;

[0035] Depository: General Microbiology Center, China Culture Collection Administration;

[0036] Storage address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing;

[0037] Date of preservation: January 20, 2025.

[0038] In the following embodiments of the present invention, oligoxylose has a molecular weight of 300-1050 Daltons and is a commercially available product of Henan Yichangqing Biotechnology Co., Ltd. Unless otherwise specified, other raw materials are commonly used in the art and are commercially available.

[0039] The separation, screening and identification processes of the Lactobacillus paracasei BL-22 and other comparative strains (Lactobacillus paracasei 1.0340, Lactobacillus paracasei 1.0384, Lactobacillus paracasei 3-1-6, Lactobacillus paracasei 3-1-8 and Lactobacillus paracasei 3-1-9) involved in the following examples or test examples are as follows:

[0040] Traditional dairy products from different families in Gannan Tibetan Autonomous Prefecture, Gansu Province were selected as test samples. After being numbered, they were transported to the laboratory under low temperature conditions. The samples showed a good consistency, with the unique flavor of naturally fermented milk and no odor. According to the method published by Lei Xueyan et al. (Lei Xueyan, Zhao Lixia, Li Yu, et al. Isolation and identification of lactic acid bacteria and screening of excellent strains in naturally fermented milk in Inner Mongolia [J]. Food and Fermentation Industries, 2025, 51(01): 11-17.), the samples were gradient diluted using a 10-fold gradient dilution method, and the dilution gradient was selected as 10. -4 , 10 -5, 10 -6 200 μL of the sample dilution was transferred to the MRS solid culture medium prepared in advance, spread evenly, and cultured at 37°C for 72 h.

[0041] After colonies formed, single colonies were isolated and purified by the plate streak method. Six purified Gram-positive strains were then inoculated into corresponding liquid culture media for 16S rRNA detection. BLAST homology comparison was performed using the NCBI (https: / / www.ncbi.nlm.nih.gov / ). Based on the homology results, the six isolated strains were identified as Lactobacillus paracasei and named Lactobacillus paracasei 1.0340, Lactobacillus paracasei 1.0384, Lactobacillus paracasei 3-1-6, Lactobacillus paracasei 3-1-8, Lactobacillus paracasei 3-1-9, and Lactobacillus paracasei BL-22 (the sequence homologies of the six strains were 100%, 100%, 100%, 99.93%, 100%, and 99.93%, respectively).

[0042] The 16S rRNA sequence of Lactobacillus paracasei BL-22 is specifically (as shown in SEQ ID NO.1):

[0043] GCACTTGTTCGACTTCCCCTAATCATTTGTCCCACCTTAGACGGCTCGCTCCCTAAAAGGGTTACGCCACCGGCTTCGGGTGTTACAAACTCTCATGGTGTGACGGGCGGTGTGTACAAGGCCCGGGAACGTATTCACCGCGGCGTGCTGATCCGCGATTACTAGCGATTCCGACTTCGTGTAGGCGAGTTGCAGCCTACAGTCCGAACTGAGAATGGCTTTAAGAGATTAGCTTGACCTCGCGGTCTCGCAACTCGTTGTACCATCCATTGTAGCACGTGTGTAGCCCAGGTCATAAGGGGCATGATGATTTGACGTCATCCCCACCTTCCTCCGGTTTGTCACCGGCAGTCTTACTAGAGTGCCCAACTAAATGCTGGCAACTAGTCATAAGGGTTGCGCTCGTTGCGGGACTTAACCCAACATCTCACGACACGAGCTGACGACAACCATGCACCACCTGTCATTTTGCCCCCGAAGGGGAAACCTGATCTCTCAGGTGATCAAAAGATGTCAAGACCTGGTAAGGTTCTTCGCGTTGCTTCGAATTAAACCACATGCTCCACCGCTTGTGCGGGCCCCCGTCAATTCCTTTGAGTTTCAACCTTGCGGTCGTACTCCCCAGGCGGAATGCTTAATGCGTTAGCTGCGGCACTGAAGGGCGGAAACCCTCCAACACCTAGCATTCATCGTTTACGGCATGGACTACCAGGGTATCTAATCCTGTTCGCTACCCATGCTTTCGAGCCTCAGCGTCAGTTACAGACCAGACAGCCGCCTTCGCCACTGGTGTTCTTCCATATATCTACGCATTTCACCGCTACACATGGAGTTCCACTGTCCTCTTCTGCACTCAAGTTTCCCAGTTTCCGATGCGCTTCCTCGGTTAAGCCGAGGGCTTTCACATCAGACTTAAAAAACCGCCTGCGCTCGCTTT。

[0044] Example 1

[0045] This example provides a strain of Lactobacillus paracasei, named Lactobacillus paracasei BL-22, which is deposited in the General Microbiology Center of the China Culture Collection Administration of Microorganisms with a deposit number of CGMCC NO. 33475.

[0046] This embodiment also provides an anti-aging and anti-glycation microbial preparation, comprising the above-mentioned Lactobacillus paracasei and oligoxylose. The dosage ratio of Lactobacillus paracasei BL-22 and oligoxylose is 1×10 8 CFU: 200mg. The molecular weight of xylooligosaccharide is 300~1050 Daltons.

[0047] Example 2

[0048] This example provides a strain of Lactobacillus paracasei, named Lactobacillus paracasei BL-22, which is deposited in the General Microbiology Center of the China Culture Collection Administration of Microorganisms with a deposit number of CGMCC NO. 33475.

[0049] This embodiment also provides an anti-aging and anti-glycation microbial preparation, comprising the above-mentioned Lactobacillus paracasei and oligoxylose. The dosage ratio of Lactobacillus paracasei BL-22 and oligoxylose is 1×10 7 CFU: 500mg. The molecular weight of xylo-oligosaccharide is 300~1050 Daltons.

[0050] Example 3

[0051] This example provides a strain of Lactobacillus paracasei, named Lactobacillus paracasei BL-22, which is deposited in the General Microbiology Center of the China Culture Collection Administration of Microorganisms with a deposit number of CGMCC NO. 33475.

[0052] This embodiment also provides an anti-aging and anti-glycation microbial preparation, comprising the above-mentioned Lactobacillus paracasei and oligoxylose. The dosage ratio of Lactobacillus paracasei BL-22 and oligoxylose is 1×10 9 CFU: 100mg. The molecular weight of xylo-oligosaccharide is 300~1050 Daltons.

[0053] Test Example 1: Screening of Lactobacillus paracasei

[0054] In this test example, the six strains of Lactobacillus paracasei obtained by the above screening (Lactobacillus paracasei 1.0340, Lactobacillus paracasei 1.0384, Lactobacillus paracasei 3-1-6, Lactobacillus paracasei 3-1-8, Lactobacillus paracasei 3-1-9, and Lactobacillus paracasei BL-22) were tested for their in vitro antioxidant capacity, including DPPH scavenging rate and hydroxyl radical scavenging capacity tests.

[0055] DPPH is commonly used to evaluate the antioxidant capacity of probiotics. Free radicals can cause oxidative damage to biomolecules, and the most harmful reactive oxygen species is the hydroxyl radical. Therefore, DPPH scavenging rate and hydroxyl radical scavenging ability are tested to screen Lactobacillus paracasei strains with the best application potential. The experimental steps are as follows:

[0056] (1) Strain activation and culture: Lactobacillus paracasei (1.0340, 1.0384, 3-1-6, 3-1-8, 3-1-9, BL-22) were inoculated into MRS sterile liquid culture medium at a 2% (v / v) inoculation volume, cultured for two generations at 37°C, and then subcultured for 18-24 h.

[0057] (2) Preparation of Lactobacillus paracasei cell suspension: The Lactobacillus paracasei cultured in the above step was centrifuged at 6000 rpm for 10 min at 4°C to obtain the cells, which were then washed three times with sterile saline and resuspended in saline to adjust the cell concentration to 10 9 CFU / mL, and used as cell suspension for subsequent experiments.

[0058] (3) DPPH free radical scavenging experiment: 1 mL of cell suspension or deionized water (as blank) was mixed with 1 mL of DPPH anhydrous ethanol solution (0.2 mM), and after thorough shaking, the mixture was incubated at 37 o C was placed in the dark for 30 minutes, centrifuged at 7000 r / min for 10 minutes, and the supernatant was collected. The absorbance was then measured at 517 nm, and the scavenging rate of DPPH free radicals was calculated according to the following formula:

[0059] ;

[0060] Among them, A 样 is the absorbance value of the sample; A 空 is the absorbance value of the blank group.

[0061] (4) Hydroxyl radical scavenging experiment: Take 1 mL of PBS, 1 mL of 1,10-phenanthroline (2.5 mM), 1 mL of ferrous sulfate (2.5 mM) and 1 mL of cell suspension and mix thoroughly. Then add 1 mL of hydrogen peroxide (20 mM) and let it stand at 37°C for 90 min. Replace the sample with an equal volume of deionized water as a blank, and replace hydrogen peroxide with an equal volume of deionized water as a control. The absorbance is measured at 536 nm. The scavenging ability of hydroxyl radicals is calculated as follows:

[0062] ;

[0063] Among them, A 样 is the absorbance value of the sample; A 空 is the absorbance value of the blank group, A 对 is the absorbance value of the control group.

[0064] The test results of DPPH scavenging rate and hydroxyl free radical scavenging ability are as follows Figure 1 shown.

[0065] Depend on Figure 1 As shown in Figure (a), among the six tested Lactobacillus paracasei strains, Lactobacillus paracasei 1.0340, Lactobacillus paracasei 3-1-6, Lactobacillus paracasei 3-1-9 and Lactobacillus paracasei BL-22 have higher DPPH scavenging abilities, among which Lactobacillus paracasei BL-22 shows the best DPPH scavenging rate, which can reach 52.7%.

[0066] Depend on Figure 1 As shown in Figure (b), different strains of Lactobacillus paracasei exhibited different hydroxyl radical scavenging abilities. 9 At CFU / mL, the hydroxyl radical scavenging rates of each bacterial suspension ranged from 14.3% to 50.3%. The descending order of scavenging ability was Lactobacillus paracasei BL-22 > Lactobacillus paracasei 3-1-6 > Lactobacillus paracasei 1.0384 > Lactobacillus paracasei 3-1-9 > Lactobacillus paracasei 1.0340 > Lactobacillus paracasei 3-1-8. Therefore, Lactobacillus paracasei BL-22 possessed the best hydroxyl radical scavenging ability.

[0067] This indicates that Lactobacillus paracasei BL-22 exhibits excellent DPPH and hydroxyl radical scavenging capabilities, achieving optimal antioxidant capacity. Therefore, Lactobacillus paracasei BL-22 was selected as the test strain for subsequent animal experiments.

[0068] Experimental Example 2: Effects on Skin Aging and Glycation Damage

[0069] This test example uses animal experiments to investigate the effects of different active ingredients on skin aging and glycation. Male BALB / c mice weighing 20-22g were selected as experimental animals. When constructing the experimental model, an aging mouse model was constructed by subcutaneous injection of D-galactose. The specific operation was as follows: after the mice were acclimated to normal feeding for one week, 0.1mL / mouse / day of D-galactose was subcutaneously injected into the remaining groups except the blank group. From the second week onwards, each intervention group was treated with Lactobacillus paracasei BL-22, oligoxylose, and a combination of Lactobacillus paracasei BL-22 and oligoxylose dissolved in PBS at corresponding doses, and the mice were gavaged at a rate of 0.1mL / mouse / day.

[0070] The blank group and the model group were gavaged with the same amount of normal saline every day as a control. The intervention group was gavaged with Lactobacillus paracasei BL-22 (1×10 9 CFU / kg mouse body weight), oligoxylose (500 mg / kg mouse body weight), and Lactobacillus paracasei BL-22 (1×10 8 CFU / kg mouse body weight) and xylooligosaccharides (200 mg / kg mouse body weight). Specific experimental groups and intervention measures are shown in Table 1.

[0071] Table 1. Experimental groups and intervention measures

[0072]

[0073] After six weeks of intervention, the mice were tested for various indicators, including skin elasticity, expression levels of skin barrier-related proteins, levels of inflammatory factors in the blood and skin, and expression levels of collagen-related genes. The results were statistically analyzed for differences. *p<0.05, **p<0.01, ***p<0.001, and ****p<0.0001 indicate significant differences between the groups.

[0074] 2.1 Effects on the elasticity of mouse back skin

[0075] Skin elasticity is usually related to skin 、 and natural Under natural conditions, with the increase of age, skin elasticity will gradually decrease. Therefore, the present invention uses a skin elasticity tester to measure the elasticity of the back skin of each group of mice. The results are as follows Figure 2 shown.

[0076] Depend on Figure 2It can be seen that compared with the blank group (skin elasticity 82%), the skin elasticity of the model group was significantly reduced to 48%. Compared with the model group, the skin elasticity of the xylo-oligosaccharide group increased by 11.1%, and the Lactobacillus paracasei BL-22 group increased by 15%. The Lactobacillus paracasei BL-22 + xylo-oligosaccharide combination group of the present invention had a more significant elasticity recovery effect (p < 0.0001), with skin elasticity restored to 78%, close to that of the blank group.

[0077] At the same time, the comparison of the dosage of active substances showed that the low dose of Lactobacillus paracasei BL-22 (1×10 8 CFU / kg) + oligoxylose (200 mg / kg) were used together to restore skin elasticity, which was significantly higher than that of high-dose oligoxylose (500 mg / kg, p < 0.0001) and high-dose BL-22 (1 × 10 9 CFU / kg, p<0.01) when used alone, it can be seen that the combined use of Lactobacillus paracasei BL-22 and xylooligosaccharides can play a synergistic role and significantly improve the recovery of skin elasticity on the basis of reducing the dosage of each component.

[0078] 2.2 Effects on the expression levels of skin barrier-related proteins on the back of mice

[0079] Loricrin is mainly expressed in the stratum corneum of the epidermis and is the main component of the cornified envelope of terminally differentiated keratinocytes. It is a characteristic marker of epithelial cell growth, differentiation and maturation, and plays an important role in epithelial cells. It can ensure the integrity of the physical and chemical properties of cells and the smooth progress of metabolic processes. In addition, It is mainly involved in the connection between skin and mucosal cells to maintain the integrity and stability of tissues. This experiment tested the expression levels of the above three proteins related to the skin barrier on the back of mice. Specifically, the skin on the back of the mice was cut immediately after the mice were killed and homogenized in liquid nitrogen. After that, lysis buffer was added for lysis, and the supernatant was collected after centrifugation at 12,000 rpm for 10 minutes at 4°C. The concentration of each protein was determined using a BCA kit, and GAPDH protein was used for normalization correction. Loricrin, 、 The protein expression results were as follows Figure 3 shown.

[0080] In this test, the present invention selected proteins (Loricrin, 、 ) expression level reflects the skin condition. Figure 3As shown in Figures (a), (b), and (c), compared with the blank group, the expression levels of Loricrin, Keratin 10, and Desmocular 1 proteins in the model group were significantly decreased (p < 0.0001), while oral administration of xylooligosaccharides and Lactobacillus paracasei BL-22 had a certain recovery effect on skin barrier-related proteins (p < 0.05, p < 0.01). Among them, the combination of Lactobacillus paracasei BL-22 + xylooligosaccharides used in the present invention showed the best recovery effect, significantly preventing the decrease in skin barrier-related proteins in mice. Specifically, in terms of restoring the relative expression levels of Loricrin and Keratin 10 proteins, the low-dose Lactobacillus paracasei BL-22 + xylooligosaccharide combination had no significant difference from the high-dose Lactobacillus paracasei BL-22, and the effect was significantly higher than that of xylooligosaccharides (p < 0.05). In terms of restoring the relative expression of Desmoglein 1 protein, the combination of Lactobacillus paracasei BL-22 and xylooligosaccharides was significantly higher than the high-dose Lactobacillus paracasei BL-22 and xylooligosaccharides alone (p < 0.05, p < 0.01). This shows that the combined use of Lactobacillus paracasei BL-22 and xylooligosaccharides in the present invention exhibits a synergistic effect and has a significant effect on restoring skin barrier-related proteins.

[0081] 2.3 Effects on TNF-α Levels in Mouse Blood and Skin

[0082] The experiment tested the content of inflammatory factor TNF-α in the blood and skin of mice. Specifically, after the intervention experiment, the mice were killed and blood was collected from the eyeballs. After standing for 40 minutes, the blood was centrifuged at 3000r / min for 20 minutes, and the blood supernatant was taken for ELISA detection; the back skin tissue was cut and ground into a homogenate at a weight-to-volume ratio of 1:10 with PBS, and the homogenate was centrifuged at 3000r / min for 20 minutes and the skin supernatant was taken. The TNF-α content in the serum and skin of mice was detected by ELISA kit, and the results are as follows: Figure 4 shown.

[0083] Depend on Figure 4As shown in Figures (a) and (b), compared with the blank group, the TNF-α levels in the serum and skin of the model group increased significantly, reaching 368.17 ng / L and 451.85 ng / L, respectively. After the model mice were gavaged with oligoxylose, Lactobacillus paracasei BL-22, and Lactobacillus paracasei BL-22 + oligoxylose, the TNF-α levels in the plasma and skin decreased to varying degrees. Among them, the combination of Lactobacillus paracasei BL-22 + oligoxylose showed the best ability to inhibit inflammatory factors in the skin and serum, reducing TNF-α levels to 238.98 ng / L and 370.85 ng / L, respectively, which was statistically significant compared with the model group (p < 0.0001). Moreover, compared with the oligoxylose and Lactobacillus paracasei BL-22 groups, the combination of Lactobacillus paracasei BL-22 + oligoxylose significantly reduced the TNF-α level in the blood (p < 0.001, p < 0.0001).

[0084] 2.4 Effects on SOD, GSH-Px, CAT, and MDA Levels in Mouse Blood

[0085] Elastin and collagen are the main structural molecules of the skin and can be destroyed by carbohydrates through glycation, covalent binding of sugars to proteins, and the subsequent generation of AGEs. These processes are closely related to the oxidation process. This experiment used ELISA kits to test the levels of SOD, GSH-Px, CAT, and MDA in the blood of mice to characterize the effects of different active components on the oxidation process. The results are as follows Figure 5 shown.

[0086] Depend on Figure 5 As shown in Figures (a), (b), (c), and (d), compared with the blank group, the activities of SOD, GSH-Px, and CAT in the serum of mice in the model group were significantly decreased (p < 0.0001), while the MDA content was significantly increased (p < 0.0001). Compared with the model group, after xylooligosaccharide treatment, MDA and CAT in the blood were significantly decreased (p < 0.05), but there were no significant differences in SOD and GSH-Px (p > 0.05). Compared with the model group, the activities of SOD, GSH-Px, and CAT were significantly increased in the Lactobacillus paracasei BL-22 + xylooligosaccharide treatment group, while the MDA content was significantly decreased, demonstrating that the combination of Lactobacillus paracasei BL-22 and xylooligosaccharide provided by the present invention can effectively resist oxidative stress.

[0087] 2.5 Effects on AGE Content in Mouse Blood and Skin

[0088] Endogenous aging is closely related to skin glycation. Glycation reaction is mainly a non-enzymatic reaction between proteins and reducing sugars. During this process, irreversible stable products AGEs are generated, and their production will damage skin fibroblasts. Therefore, reducing the production of AGEs is the core of anti-glycation and one of the effective ways to alleviate skin problems. This experiment tested the AGE content in the blood and skin of mice using an ELISA kit to evaluate the anti-glycation effect of different intervention groups. The results are as follows Figure 6 shown.

[0089] Combine Figure 6 As shown in Figures (a) and (b), after subcutaneous injection of D-galactose, the model group significantly increased blood and skin AGE levels compared to the blank group due to the oxidative effects of D-galactose (p < 0.0001). After intervention with xylooligosaccharides, Lactobacillus paracasei BL-22, and the combination of Lactobacillus paracasei BL-22 and xylooligosaccharides, blood and skin AGE levels decreased to varying degrees in each intervention group compared to the model group (p < 0.01, p < 0.001, and p < 0.0001, respectively). The Lactobacillus paracasei BL-22 and xylooligosaccharide combination showed the most significant reduction in AGE levels, demonstrating its synergistic effect and potential application in skin anti-glycation.

[0090] 2.6 Alleviating mouse aging through the TGF-β / Smad cell pathway

[0091] TGF-β is the main inducer of collagen synthesis in skin fibroblasts. The TGF-β / Smad signaling pathway can directly promote collagen synthesis. This experiment used β-actin for normalization correction and tested the expression levels of key genes involved in the TGF-β / Smad cell pathway (SMAD2, SMAD3, TGF-β I and TGF-β II). The results are as follows: Figure 7 shown.

[0092] Combine Figure 7As shown in Figures (a), (b), (c), and (d), the expression of SMAD2, SMAD3, TGF-β I, and TGF-β II genes in mice treated with D-galactose was significantly decreased compared to the blank control group (p < 0.0001). The expression levels of SMAD2, SMAD3, TGF-β I, and TGF-β II genes were significantly upregulated after administration of xylooligosaccharides, Lactobacillus paracasei BL-22, and Lactobacillus paracasei BL-22 + xylooligosaccharides (p < 0.05, p < 0.01, p < 0.001, and p < 0.0001). Furthermore, the effect of the Lactobacillus paracasei BL-22 + xylooligosaccharide group on restoring TGF-β gene expression was significantly greater than that of the xylooligosaccharide alone group and the Lactobacillus paracasei BL-22 alone group (p < 0.05, p < 0.01, and p < 0.0001). Given that the TGF-β / Smad pathway is an important signaling pathway for regulating collagen synthesis, this result also corresponds to the fact that the above-mentioned xylooligosaccharides, Lactobacillus paracasei BL-22, and Lactobacillus paracasei BL-22+xylooligosaccharides increase the expression of skin barrier-related proteins and skin elasticity in mice. This shows that the present invention uses a compound combination of Lactobacillus paracasei BL-22+xylooligosaccharides to promote collagen synthesis by activating the TGF-β / Smad signaling pathway, thereby effectively delaying skin aging.

[0093] In summary, the present invention has explored the effects of lactobacillus paracasei BL-22 alone, xylo-oligosaccharide alone and lactobacillus paracasei BL-22 and xylo-oligosaccharide coupling by animal experiment and active ingredient interaction, for the impact of skin anti-oxidation and alleviation of skin aging and anti-glycation. The test shows that the lactobacillus paracasei BL-22 alone group, xylo-oligosaccharide alone group and lactobacillus paracasei BL-22 and xylo-oligosaccharide coupling group all have the effect of anti-aging and anti-glycation. In particular, compared with the Lactobacillus paracasei BL-22 alone group and the Xylo-oligosaccharide alone group, the combined use of Lactobacillus paracasei BL-22 and Xylo-oligosaccharide has the following outstanding effects: (1) significantly improves skin elasticity; (2) effectively increases the content of skin barrier-related proteins; (3) significantly reduces the level of inflammatory factor TNF-α; (4) exerts an effective antioxidant stress effect; (5) significantly reduces the AGE content and exerts an anti-glycation effect on the skin; (6) activates the TGF-β / Smad signaling pathway, promotes collagen synthesis, and effectively delays skin aging.

[0094] Visible, the present invention has revealed the effect of Lactobacillus paracasei BL-22 in skin anti-aging and anti-glycation by comprehensive experimental exploration for the first time, and simultaneously by exploring the interaction mechanism between Lactobacillus paracasei BL-22 and xylo-oligosaccharides, confirmed that the coupling between Lactobacillus paracasei BL-22 and xylo-oligosaccharides can play a synergistic effect in alleviating skin aging and glycation. Therefore, the present invention can provide a kind of healthy and convenient new strategy for solving skin problems, and can also provide new research and development direction and technical support for the development of health products for anti-aging and anti-glycation.

[0095] Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or make equivalent replacements for some of the technical features therein; and these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. An anti-aging and anti-glycation microbial preparation, characterized in that: The microbial preparation comprises Lactobacillus paracasei and oligoxylose; the Lactobacillus paracasei is named Lactobacillus paracasei ( Lactobacillus paracasei ) BL-22, deposited in the General Microbiology Center of China Microorganism Culture Collection Administration Committee, with the deposit number CGMCC NO.33475; the molecular weight of the xylooligosaccharide is 300-1050 Daltons; in the microbial preparation, the dosage ratio of Lactobacillus paracasei to xylooligosaccharide is 1×10 8 CFU: 200mg.

2. An anti-aging and anti-glycation product, characterized in that: It comprises active components and excipients; the active component is the anti-aging and anti-glycation microbial preparation according to claim 1.

3. The anti-aging and anti-glycation product according to claim 2, characterized in that: The product described is a pharmaceutical product.

4. The anti-aging and anti-glycation product according to claim 3, characterized in that The medicine is one of tablets, granules, oral liquid and capsules.

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