Lactobacillus paracasei and application thereof in anti-aging and anti-saccharification

Abstract

The invention belongs to the technical field of microorganisms, and particularly relates to lactobacillus paracasei and application thereof in anti-aging and anti-saccharification. The lactobacillus paracasei provided by the invention is lactobacillus paracasei BL-22, and the preservation number of the lactobacillus paracasei BL-22 is CGMCC (China General Microbiological Culture Collection Center) NO. 33475. The lactobacillus paracasei provided by the invention is a lactobacillus paracasei strain. Tests prove that the lactobacillus paracasei BL-22 has remarkable effects in the aspects of skin oxidation resistance, skin aging relieving and saccharification resistance, and meanwhile, through animal tests and research on interaction of active components, an interaction mechanism between the lactobacillus paracasei BL-22 and xylooligosaccharide is disclosed. It is proved that the lactobacillus paracasei BL-22 and the xylooligosaccharide are combined to play a synergistic effect in the aspects of relieving skin aging and saccharification. Therefore, the invention can provide a new research and development direction and technical support for the development of anti-aging and anti-saccharification health products.

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 divided into exogenous and intrinsic aging. Exogenous aging is mainly related to ultraviolet radiation, while intrinsic aging is closely related to skin glycation. Glycation reaction is mainly a non-enzymatic reaction between proteins (including collagen, etc.) and reducing sugars, generating irreversible stable products such as advanced glycation end products (AGEs).

[0003] The extracellular matrix (ECM) is one of the main targets of glycation reactions. Collagen reacts with sugar to produce AGEs. The accumulated AGEs cross-link with the ECM, resulting in structural and functional damage to the connective tissue. In addition, elastin and collagen are the main structural molecules of the skin, which can react with carbohydrates through glycation, covalent binding of sugars to proteins, and subsequent destruction of the generated AGEs, thereby exacerbating the skin aging process.

[0004] At present, many active ingredients used for anti-aging and anti-glycation contain irritating ingredients such as retinol and acids, which can easily damage the skin barrier and cause skin sensitivity and inflammatory reactions if used for a long time. Therefore, developing more safe, natural, and excellent anti-aging and anti-glycation active ingredients and related products can provide a healthy and convenient new way to solve skin problems. Summary of the invention

[0005] In view of the deficiencies in the prior art, one of the purposes of the present invention is to provide a strain of Lactobacillus paracasei which has 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 Microbiological Center of China National Microbiological Culture Collection Committee 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, in the microbial preparation, the dosage ratio of Lactobacillus paracasei and xylo-oligosaccharide is (1×10 7 ~1×10 9 ) CFU: (100-500) mg. The present invention uses Lactobacillus paracasei BL-22 and xylooligosaccharide as active ingredients, 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, in the microbial preparation, the dosage ratio of Lactobacillus paracasei to xylo-oligosaccharide 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 an auxiliary material; 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; the health food is one of pills, granules, powder, and oral liquid.

[0019] The present invention does not specifically limit the types of auxiliary materials, and technicians can select common types of auxiliary materials according to the types of product preparation and preparation requirements. In a further preferred embodiment, when the product is a skin care product, the auxiliary materials used include one or more of a moisturizer, a pH adjuster, a thickener, and a preservative; moisturizers can be, for example, glycerol, 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 a health food, the auxiliary materials used include one or more of a filler, an adhesive, 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 invention is screened from traditional dairy products in Gannan Tibetan Autonomous Prefecture of Gansu Province, has high biological safety, and is verified to have not only excellent DPPH free radical and hydroxyl free radical scavenging capabilities, but also outstanding antioxidant capacity, and excellent skin anti-aging and anti-glycation effects.

[0022] Furthermore, the present invention explores the effects of Lactobacillus paracasei BL-22 alone, xylooligosaccharides alone, and the combined use of Lactobacillus paracasei BL-22 and xylooligosaccharides on skin antioxidant, skin aging relief, and anti-glycation through animal experiments and studies on the interaction of active components. The experiment shows that compared with the Lactobacillus paracasei BL-22 alone group and the xylooligosaccharides alone group, the present invention uses Lactobacillus paracasei BL-22 and xylooligosaccharides in combination, which 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 anti-oxidative stress effect; (5) significantly reduces the AGE content and exerts skin anti-glycation effect; (6) activates the TGF-β / Smad signaling pathway, promotes the synthesis of collagen, and effectively delays skin aging.

[0023] The present invention, through comprehensive experimental exploration, reveals for the first time the interaction mechanism between Lactobacillus paracasei BL-22 and xylo-oligosaccharides, and confirms that the combination of 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 healthy and convenient new 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 are a comparison of the antioxidant capacity of different Lactobacillus paracasei in Experimental Example 1 of the present invention; wherein (a) is the DPPH clearance 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, and (b) is The relative expression of proteins, (c) Relative expression of proteins;

[0027] Figure 4 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 test results of AGE levels in the blood and skin of mice in different groups in Experimental Example 2 of the present invention are shown in FIG. 1 , wherein (a) is the AGE content in the blood, and (b) is the AGE content in the skin;

[0030] Figure 7 The expression levels of SMAD2, SMAD3, TGF-β I and TGF-β II genes in skin tissues of mice in different groups in Experimental Example 2 of the present invention; wherein, (a) is the expression level of SMAD2 gene, and (b) is the expression level of SMAD3 gene; (c) is the expression level of TGF-β I gene, and (b) is the expression level of TGF-β II gene. 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 absence of 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 protection scope of the present invention is not limited by the specific embodiments disclosed below. Under the premise of not conflicting, 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 Microbiological 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, the molecular weight of xylo-oligosaccharide is 300-1050 Daltons, and it is a commercial product of Henan Yichangqing Biotechnology Co., Ltd. Unless otherwise specified, other raw materials are all materials commonly used in the art and can be obtained through commercial channels.

[0039] The separation, screening and identification process 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, Lactobacillus paracasei 3-1-9) involved in the following embodiments 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 thick 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 gradiently diluted using the 10-fold gradient dilution method, and the dilution gradient was selected as 10 -4 , 10 -5, 10 -6 Take 200 μL of the sample dilution and transfer it to the MRS solid culture medium prepared in advance. Spread it evenly and culture it at 37℃ for 72h.

[0041] After colonies were formed, single colonies were isolated and purified by the plate streak method, and then the six purified Gram-positive strains were inoculated into the corresponding liquid culture medium for 16S rRNA detection. Finally, BLAST homology comparison was performed in NCBI (https: / / www.ncbi.nlm.nih.gov / ). According to the homology results, the six isolated strains were determined to be Lactobacillus paracasei, and they were 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, respectively (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 embodiment provides a strain of Lactobacillus paracasei, named Lactobacillus paracasei BL-22, which is deposited in the General Microbiological Center of China National Microbiological Culture Collection Administration 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 xylo-oligosaccharide. The dosage ratio of Lactobacillus paracasei BL-22 and xylo-oligosaccharide is 1×10 8 CFU: 200mg. The molecular weight of xylooligosaccharide is 300~1050 Daltons.

[0047] Example 2

[0048] This embodiment provides a strain of Lactobacillus paracasei, named Lactobacillus paracasei BL-22, which is deposited in the General Microbiological Center of China National Microbiological Culture Collection Administration 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 xylo-oligosaccharide. The dosage ratio of Lactobacillus paracasei BL-22 and xylo-oligosaccharide is 1×10 7 CFU: 500mg. The molecular weight of xylooligosaccharide is 300~1050 Daltons.

[0050] Example 3

[0051] This embodiment provides a strain of Lactobacillus paracasei, named Lactobacillus paracasei BL-22, which is deposited in the General Microbiological Center of China National Microbiological Culture Collection Administration 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 xylo-oligosaccharide. The dosage ratio of Lactobacillus paracasei BL-22 and xylo-oligosaccharide is 1×10 9 CFU: 100mg. The molecular weight of xylooligosaccharide 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 usually used to evaluate the antioxidant capacity of probiotics. Free radicals can cause oxidative damage to biological molecules, among which the most harmful reactive oxygen free radical is hydroxyl free radical. Therefore, DPPH scavenging rate and hydroxyl free radical scavenging ability tests are used to screen Lactobacillus paracasei with the best application potential. The test 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 rate, cultured for two generations at 37°C, and then subcultured for 18-24 h.

[0057] (2) Preparation of Lactobacillus paracasei cell suspension: The cultured Lactobacillus paracasei cell suspension 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 sufficient shaking, the mixture was incubated at 37 o C was placed away from light for 30 minutes, centrifuged at 7000r / min for 10 minutes, and the supernatant was taken. Then the absorbance was measured at 517nm, 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 place 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. Measure the absorbance 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 strains of Lactobacillus paracasei tested, 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 showed different hydroxyl radical scavenging abilities. 9 CFU / mL, the scavenging rate of each bacterial suspension on hydroxyl free radicals ranged from 14.3% to 50.3%, and the scavenging ability was from high to low in the order of 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. It can be seen that the tested strain of Lactobacillus paracasei BL-22 has the best scavenging ability on hydroxyl free radicals.

[0067] It can be seen that Lactobacillus paracasei BL-22 showed excellent DPPH free radical and hydroxyl free radical scavenging ability and could achieve the best antioxidant capacity. Therefore, Lactobacillus paracasei BL-22 was selected as the test strain for subsequent animal experiments.

[0068] Test 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 adapted to normal feeding for one week, 0.1mL / mouse / day of D-galactose was subcutaneously injected in the remaining groups except the blank group. From the second week, each intervention group was respectively dissolved in PBS with a corresponding dose of Lactobacillus paracasei BL-22, xylo-oligosaccharides, and a composition of Lactobacillus paracasei BL-22+xylo-oligosaccharides, and the mice were gavaged at 0.1mL / mouse / day.

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

[0071] Table 1. Experimental groups and intervention measures

[0072] After six weeks of intervention, various indicators of mice were tested, including mouse skin elasticity, expression levels of skin barrier-related proteins, levels of inflammatory factors in mouse blood and skin, and expression levels of collagen-related genes, and the experimental results were statistically analyzed for differences. Among them, *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001, represent significant differences between the groups.

[0073] 2.1 Effects on the elasticity of the back skin of mice

[0074] 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.

[0075] 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 elastic recovery effect (p < 0.0001), and the skin elasticity was restored to 78%, which was close to the blank group.

[0076] At the same time, the comparison of the dosage of active substances showed that low dose of Lactobacillus paracasei BL-22 (1×10 8 CFU / kg) + oligoxylose (200 mg / kg) had a significantly higher effect on restoring skin elasticity than high-dose oligoxylose (500 mg / kg, p < 0.0001) and high-dose BL-22 (1×10 9 CFU / kg, p<0.01) alone, it can be seen that the combined use of Lactobacillus paracasei BL-22 and oligoxylose can play a synergistic role and significantly improve the recovery of skin elasticity on the basis of reducing the dosage of each component.

[0077] 2.2 Effects on the expression levels of skin barrier-related proteins on the back of mice

[0078] 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, ensuring the integrity of the physical and chemical properties of cells and the smooth progress of metabolic processes. It is mainly involved in the connection between skin and mucosal cells to maintain tissue integrity and stability. This experiment tested the expression levels of the above three proteins related to the skin barrier on the back of mice. Specifically, the back skin 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 are as follows Figure 3 shown.

[0079] In this experiment, 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, Keratin10, and Desmocular1 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 present invention adopts a combination of Lactobacillus paracasei BL-22 + xylooligosaccharides, which shows the best recovery effect and significantly prevents the decrease of mouse skin barrier-related proteins. Specifically, in terms of restoring the relative expression of Loricrin and Keratin 10 proteins, there is no significant difference between the combination of low-dose Lactobacillus paracasei BL-22 + xylooligosaccharides and high-dose Lactobacillus paracasei BL-22, and the effect is significantly higher than xylooligosaccharides (p < 0.05). In terms of restoring the relative expression of Desmoglein 1 protein, the combination of Lactobacillus paracasei BL-22 + oligoxylose was significantly higher than the single application of high-dose Lactobacillus paracasei BL-22 and oligoxylose (p < 0.05, p < 0.01). It can be seen that the combined use of Lactobacillus paracasei BL-22 and oligoxylose in the present invention shows a synergistic effect and has a significant restorative effect on skin barrier-related proteins.

[0080] 2.3 Effects on the levels of inflammatory factor TNF-α in the blood and skin of mice

[0081] The experiment tested the content of inflammatory factor TNF-α in mouse blood and skin. Specifically, after the intervention experiment, mice were killed to collect eyeball blood, which was allowed to stand for 40 minutes and then centrifuged at 3000r / min for 20 minutes. 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, centrifuged at 3000r / min for 20 minutes, and the skin supernatant was taken. The ELISA kit was used to detect the content of TNF-α in mouse serum and skin. The results are as follows: Figure 4 shown.

[0082] Depend on Figure 4As shown in Figures (a) and (b), compared with the blank group, the TNF-α in the serum and skin of the model group increased significantly, rising to 368.17ng / L and 451.85ng / L, respectively. After the modeling mice were gavaged with oligoxylose, Lactobacillus paracasei BL-22, and Lactobacillus paracasei BL-22 + oligoxylose, the TNF-α content in plasma and skin decreased to different levels. Among them, the combination of Lactobacillus paracasei BL-22 + oligoxylose showed the best ability to inhibit inflammatory factors in the skin and serum, reducing the TNF-α level to 238.98ng / L and 370.85ng / 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).

[0083] 2.4 Effects on the levels of SOD, GSH-Px, CAT and MDA in the blood of mice

[0084] Elastin and collagen are the main structural molecules of the skin and can be destroyed by carbohydrates through glycosylation, 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 content 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.

[0085] 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 the model group mice were significantly decreased (p < 0.0001), and the content of MDA was significantly increased (p < 0.0001). Compared with the model group, after the intervention of oligoxylose, MDA and CAT in the blood were reduced, with significant differences (p < 0.05), but there was no significant difference in SOD and GSH-Px (p > 0.05). Compared with the model group, the activities of SOD, GSH-Px, and CAT in the combined group of Lactobacillus paracasei BL-22 + oligoxylose were significantly increased, and the content of MDA was significantly decreased, which proved that the combination of Lactobacillus paracasei BL-22 + oligoxylose provided by the present invention can play an effective anti-oxidative stress role.

[0086] 2.5 Effects on AGE Content in Blood and Skin of Mice

[0087] Endogenous aging is closely related to skin glycation. Glycation reaction is mainly a non-enzymatic reaction between protein and reducing sugar. In this process, irreversible stable products AGEs are generated, and their production will damage skin fibroblasts. Therefore, reducing the production of AGE 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.

[0088] Combination Figure 6 As shown in Figures (a) and (b), after subcutaneous injection of D-galactose, the AGE content in the blood and skin of the model group increased significantly compared with the blank group due to the oxidation of D-galactose (p < 0.0001). After intervention with oligoxylose, Lactobacillus paracasei BL-22, and Lactobacillus paracasei BL-22 + oligoxylose, the AGE content in the blood and skin of each intervention group decreased to varying degrees compared with the model group (p < 0.01, p < 0.001, p < 0.0001). Among them, the combination of Lactobacillus paracasei BL-22 + oligoxylose was the most significant in reducing the AGE content, showing its synergistic effect and application prospects in skin anti-glycation.

[0089] 2.6 Alleviating mouse aging through the TGF-β / Smad cell pathway

[0090] TGF-β is the main inducer of collagen synthesis in skin fibroblasts, and 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.

[0091] Combination Figure 7As shown in Figures (a), (b), (c), and (d), after D-galactose modeling, the SMAD2, SMAD3, TGF-β I, and TGF-β II genes of mice were significantly decreased compared with the blank group (p < 0.0001). After the administration of xylo-oligosaccharides, Lactobacillus paracasei BL-22, and Lactobacillus paracasei BL-22 + xylo-oligosaccharides, the expression levels of SMAD2, SMAD3, TGF-β I, and TGF-β II genes were significantly upregulated (p < 0.05, p < 0.01, p < 0.001, p < 0.0001). In addition, in terms of restoring the expression level of TGF-β gene, the effect of Lactobacillus paracasei BL-22 + xylo-oligosaccharides group was significantly higher than that of the xylo-oligosaccharides group and the Lactobacillus paracasei BL-22 group (p < 0.05, p < 0.01, p < 0.0001). In view of the fact 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 xylo-oligosaccharides, Lactobacillus paracasei BL-22, and Lactobacillus paracasei BL-22+xylo-oligosaccharides increase the expression of skin barrier-related proteins and skin elasticity in mice. This shows that the present invention adopts Lactobacillus paracasei BL-22+xylo-oligosaccharides for compound combination, which can promote the synthesis of collagen by activating the TGF-β / Smad signaling pathway, thereby effectively delaying skin aging.

[0092] In summary, the present invention has explored the effects of single use of Lactobacillus paracasei BL-22, single use of xylo-oligosaccharide and combined use of Lactobacillus paracasei BL-22 and xylo-oligosaccharide on skin anti-oxidation and alleviation of skin aging and anti-glycosylation through animal experiments and the study of active ingredient interactions. The test shows that the single use group of Lactobacillus paracasei BL-22, single use group of xylo-oligosaccharide and combined use group of Lactobacillus paracasei BL-22 and xylo-oligosaccharide all have the effects of anti-aging and anti-glycosylation. 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 improving skin elasticity; (2) effectively increasing the content of skin barrier-related proteins; (3) significantly reducing the level of inflammatory factor TNF-α; (4) exerting an effective antioxidant effect; (5) significantly reducing AGE content and exerting skin anti-glycation effect; (6) activating the TGF-β / Smad signaling pathway, promoting collagen synthesis, and effectively delaying skin aging.

[0093] It can be seen that the present invention, through comprehensive experimental exploration, has revealed the effect of Lactobacillus paracasei BL-22 in skin anti-aging and anti-glycation for the first time, and by exploring the interaction mechanism between Lactobacillus paracasei BL-22 and xylo-oligosaccharides, it has been confirmed that the combination 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 new healthy and convenient 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.

[0094] 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 substitutions for some of the technical features therein; and these modifications or substitutions do not make the essence of the corresponding technical solutions deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A strain of Lactobacillus paracasei, characterized in that: The paracasei lactobacillus is named as paracasei lactobacillus ( Lactobacillus paracasei )BL-22, deposited in the General Microbiology Center of China Microorganism Culture Collection Administration, with the deposit number CGMCC NO. 33475.

2. An application of Lactobacillus paracasei as claimed in claim 1, characterized in that, Application in the preparation of anti-aging and anti-glycation microbial preparations.

3. An anti-aging and anti-glycation microbial preparation, characterized in that: The microbial preparation comprises the Lactobacillus paracasei as claimed in claim 1.

4. The anti-aging and anti-glycation microbial preparation according to claim 3, characterized in that: The microbial preparation also includes xylooligosaccharides.

5. The anti-aging and anti-glycation microbial preparation according to claim 4, characterized in that: The molecular weight of the xylooligosaccharide is 300-1050 Daltons.

6. The anti-aging and anti-glycation microbial preparation according to claim 4 or 5, characterized in that: In the microbial preparation, the dosage ratio of Lactobacillus paracasei and xylo-oligosaccharide is (1×10 7 ~1×10 9 )CFU∶(100~500)mg.

7. The anti-aging and anti-glycation microbial preparation according to claim 6, characterized in that: In the microbial preparation, the dosage ratio of Lactobacillus paracasei to xylo-oligosaccharide is 1×10 8 CFU: 200mg.

8. An anti-aging and anti-glycation product, characterized in that: It comprises active components and auxiliary materials; the active component is the anti-aging and anti-glycation microbial preparation according to any one of claims 3 to 7.

9. The anti-aging and anti-glycation product according to claim 8, characterized in that: The product is one of skin care products, medicines and health foods.

10. The anti-aging and anti-glycation product according to claim 9, characterized in that: The skin care product is one of skin care water, skin care milk, essence, and face cream; the medicine is one of tablets, granules, oral liquid, and capsules; the health food is one of pills, granules, powder, and oral liquid.

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