Application of animal bifidobacterium subspecies F1-7 in preparation of medicine for preventing or treating sarcopenia

Abstract

The invention discloses application of a bifidobacterium animalis subspecies F1-7 in preparation of a medicine for preventing or treating sarcopenia, belongs to the technical field of microorganism application, and provides application of the bifidobacterium animalis subspecies F1-7 in preparation of the medicine for preventing or treating sarcopenia. The invention relates to an application of bifidobacterium animalis subspecies F1-7 in preparation of drugs for improving bile acid metabolic function decline caused by sarcopenia. The invention relates to an application of bifidobacterium animalis subspecies F1-7 in preparation of a medicine for improving intestinal flora imbalance caused by sarcopenia. The invention finds that the bifidobacterium animalis subspecies F1-7 can effectively improve the skeletal muscle quality of mice with sarcopenia, improve the condition of tension reduction, regulate intestinal flora, promote bile acid metabolism and further relieve muscular atrophy, and enriches a probiotic library for improving sarcopenia.

Description

Technical Field

[0001] The present invention belongs to the technical field of microbial applications, and particularly relates to the application of Bifidobacterium animalis subsp. F1-7 in the preparation of drugs for preventing or treating sarcopenia. Background Art

[0002] Sarcopenia, abbreviated as muscle loss, is a progressive and widespread syndrome of skeletal muscle mass and strength decline, specifically manifested as a decrease in the number and cross-sectional area of muscle fibers and the disruption of the net protein degradation balance. Sarcopenia may increase the risks of falls, fractures, and low quality of life, further leading to frailty, disability, and high mortality. Currently, there is no specific treatment plan or effective drug for sarcopenia, and the main recommendations for preventing and treating sarcopenia are moderate exercise and a high-quality diet. Therefore, dietary intervention is recognized as a more scientific and reasonable strategy for improving sarcopenia.

[0003] The gut microbiota may regulate the homeostasis of skeletal muscle, that is, the "gut-muscle axis" hypothesis has become a research hotspot in recent years. A large number of studies have shown that gut microbiota dysbiosis and reduced bile acid metabolism are associated with muscle atrophy. Sarcopenia is characterized by a decrease in beneficial gut bacteria and an increase in harmful bacteria, and the disorder of the gut microbiota, which will block the bile acid metabolism dominated by gut microbiota, weaken the muscle metabolism signaling pathway affected by bile acids in the gut-muscle axis, and finally exacerbate muscle atrophy. In summary, abnormal gut microbiota and blocked bile acid metabolism will further affect the development of sarcopenia. Therefore, restoring gut microbiota homeostasis and promoting bile acid metabolism may be a strategy for improving sarcopenia. In this process, probiotics or their probiotic products that are safe and have no toxic side effects become a relatively potential object. However, the role and mechanism of Bifidobacterium animalis in sarcopenia are not yet clear, and it is necessary to deeply explore the strains that can improve sarcopenia, which will be of great significance for seeking prevention and treatment strategies for sarcopenia. Summary of the Invention

[0004] To solve the above technical problems, the present invention proposes the application of Bifidobacterium animalis subsp. F1-7 in the preparation of drugs for preventing or treating sarcopenia. Bifidobacterium animalis subsp. F1-7 can effectively improve the skeletal muscle mass of sarcopenia mice, improve the decline in tensile strength, and can also regulate the gut microbiota to promote bile acid metabolism, thereby alleviating muscle atrophy, enriching the probiotic library for improving sarcopenia.

[0005] To achieve the above object, the present invention provides the application of Bifidobacterium animalis subsp. F1-7 in the preparation of drugs for preventing or treating sarcopenia.

[0006] Preferably, the 16S rDNA nucleotide sequence of the Bifidobacterium animalis subsp. F1-7 is as shown in SEQ ID NO.1; the Bifidobacterium animalis subsp. F1-7 was deposited at the China Center for Type Culture Collection on December 2, 2020. The deposit address is Wuhan University, Wuhan, China, and the deposit number is: CCTCC NO: M2020833.

[0007] Preferably, the Bifidobacterium animalis subsp. F1-7 can relieve the muscle mass decline caused by sarcopenia, improve the decline in motor ability, improve the decline in bile acid metabolism, and correct the intestinal flora imbalance caused by sarcopenia, so as to achieve the effect of preventing or treating sarcopenia.

[0008] Preferably, the improvement of the muscle mass decline caused by sarcopenia is to improve the reduction of the farnesoid X receptor protein level in the intestine, and then increase the level of fibroblast growth factor 15 in the body through the farnesoid X receptor / fibroblast growth factor 15 pathway, so as to relieve the muscle mass decline caused by sarcopenia.

[0009] Preferably, the improvement of the decline in bile acid metabolism is to increase the levels of chenodeoxycholic acid and deoxycholic acid, and reduce the levels of T-β-MCA and 7-ketodeoxycholic acid, thereby improving bile acid metabolism.

[0010] Preferably, the correction of the intestinal flora imbalance caused by sarcopenia is to increase the relative abundances of Enterococcus and Bifidobacterium, while reducing the relative abundances of Dubosiella and Desulfovibrio, thereby correcting the intestinal flora imbalance caused by sarcopenia.

[0011] The present invention also provides the use of Bifidobacterium animalis subsp. F1-7 in the preparation of a drug for improving the decline in bile acid metabolism function caused by sarcopenia. The 16S rDNA nucleotide sequence of the Bifidobacterium animalis subsp. F1-7 is as shown in SEQ ID NO.1; the Bifidobacterium animalis subsp. F1-7 was deposited at the China Center for Type Culture Collection on December 2, 2020. The deposit address is Wuhan University, Wuhan, China, and the deposit number is: CCTCC NO: M2020833.

[0012] The present invention also provides the use of Bifidobacterium animalis subsp. F1-7 in the preparation of a drug for improving the intestinal flora imbalance caused by sarcopenia. The 16S rDNA nucleotide sequence of the Bifidobacterium animalis subsp. F1-7 is as shown in SEQ ID NO.1; the Bifidobacterium animalis subsp. F1-7 was deposited at the China Center for Type Culture Collection on December 2, 2020. The deposit address is Wuhan University, Wuhan, China, and the deposit number is: CCTCC NO: M2020833.

[0013] The present invention also provides a method for preparing fermented milk using Bifidobacterium animalis subsp. F1-7, comprising the following steps: heating pure milk to 55 °C, adding granulated sugar with a volume fraction of 6%, sterilizing at 95 °C for 10 min, and cooling to obtain treated milk; inoculating Bifidobacterium animalis subsp. F1-7 and a commercial starter of type 2 into the treated milk at a volume ratio of 2:1, fermenting at a constant temperature of 42 °C until the pH reaches 4.5, terminating the fermentation, and performing after-ripening at 4 °C for 24 h to obtain fermented milk;

[0014] The 16S rDNA nucleotide sequence of the Bifidobacterium animalis subsp. F1-7 is as shown in SEQ ID NO.1; the Bifidobacterium animalis subsp. F1-7 was deposited at the China Center for Type Culture Collection on December 2, 2020, with the deposit address being Wuhan University, Wuhan, China, and the deposit number being: CCTCC NO:M2020833.

[0015] The present invention also provides the application of the fermented milk in the preparation of a functional food for improving intestinal flora imbalance.

[0016] Compared with the prior art, the present invention has the following advantages and technical effects:

[0017] The present invention discovers that Bifidobacterium animalis subsp. F1-7 significantly improves muscle protein metabolism in sarcopenia mice and alleviates muscle atrophy in mice by upregulating the gene expression of farnesoid X receptor and fibroblast growth factor 15 in the intestine, increasing the level of fibroblast growth factor 15 in the body, and downregulating the gene expression of Atrogin-1 and MuRF1 in muscle; Bifidobacterium animalis subsp. F1-7 can correct the intestinal flora imbalance caused by sarcopenia, increase the relative abundances of Enterococcus and Bifidobacterium, and simultaneously reduce the relative abundances of Dubosiella and Desulfovibrio; Bifidobacterium animalis subsp. F1-7 can improve the decreased bile acid metabolism in sarcopenia, increase the levels of farnesoid X receptor-activated bile acids chenodeoxycholic acid and deoxycholic acid in the intestine, and reduce the levels of farnesoid X receptor-inhibiting bile acids T-β-MCA and 7-ketodeoxycholic acid. In the present invention, Bifidobacterium animalis subsp. F1-7 can be used as an auxiliary starter to prepare fermented milk. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for use in the embodiments. Obviously, the drawings in the following description are only some embodiments of the present invention, and those of ordinary skill in the art can also obtain other drawings based on these drawings without creative efforts.

[0019] Figure 1 Figure for measuring the effects of Bifidobacterium animalis subsp. F1-7 intervention on skeletal muscle mass and motor ability in sarcopenia mice. Among them, (a) shows the weight measurement of extensor digitorum longus (EDL), tibialis anterior (TA), gastrocnemius (GA), and soleus (SOL); (b) shows the muscle mass index measurement; (c) shows the grip strength measurement. Different letters a, b, and c in the figure represent statistical differences. HM represents the normal mouse group, SM represents the sarcopenia mouse model group, and F1-7 represents the Bifidobacterium animalis subsp. F1-7 intervention group;

[0020] Figure 2 Figure for measuring the effects of Bifidobacterium animalis subsp. F1-7 intervention on muscle pathological damage in sarcopenia mice. Among them, (a) shows the normal mouse group, (b) shows the sarcopenia mouse model group, (c) shows the Bifidobacterium animalis subsp. F1-7 intervention group, and the scale bar is 100 μm;

[0021] Figure 3 Figure for measuring the effects of Bifidobacterium animalis subsp. F1-7 intervention on the gene expression levels of muscle atrophy markers MuRF-1 and Atrogin-1. Among them, (a) shows the gene expression level of MuRF-1, (b) shows the gene expression level of Atrogin-1. Different letters a, b, and c in the figure represent statistical differences. HM represents the normal mouse group, SM represents the sarcopenia mouse model group, and F1-7 represents the Bifidobacterium animalis subsp. F1-7 intervention group;

[0022] Figure 4 Figure for analyzing the protein expression of ileal FXR / FGF15 by immunohistochemical staining and Elisa detection after Bifidobacterium animalis subsp. F1-7 intervention. Among them, (a) shows the representative immunohistochemical picture of ileal FXR, (b) shows the area analysis of ileal FXR immunohistochemistry, (c) shows the level of FGF15, (d) shows the relative mRNA expression level of FXR, (e) shows the relative mRNA expression level of FGF15, (f) shows the relative mRNA expression level of FGFR4, (g) shows the relative mRNA expression level of KLB. Different letters a, b, and c in the figure represent statistical differences. HM represents the normal mouse group, SM represents the sarcopenia mouse model group, and F1-7 represents the Bifidobacterium animalis subsp. F1-7 intervention group;

[0023] Figure 5This is a diagram showing the effects of the intervention of Bifidobacterium animalis subsp. F1-7 at the genus level on the gut microbiota. In the figure, HM represents the normal mouse group, SM represents the sarcopenia mouse model group, F1-7 represents the Bifidobacterium animalis subsp. F1-7 intervention group, Bifidobacterium represents the genus Bifidobacterium, Lachnoclostridium represents the genus Lachnoclostridium, Enterococcus represents the genus Enterococcus, Dubosiella represents the genus Dubosiella, and desulfovibrio represents the genus Desulfovibrio;

[0024] Figure 6 This is the determination of the effects of the intervention of Bifidobacterium animalis subsp. F1-7 on bile acid metabolism. Among them, (a) is the level of chenodeoxycholic acid, (b) is the level of deoxycholic acid, (c) is the level of T-β-MCA, and (d) is the level of 7-ketodeoxycholic acid. In the figure, different letters for a, b, and c indicate statistical differences. HM represents the normal mouse group, SM represents the sarcopenia mouse model group, and F1-7 represents the Bifidobacterium animalis subsp. F1-7 intervention group;

[0025] Figure 7 This is the sensory evaluation results of the fermented milk of Bifidobacterium animalis subsp. F1-7 and the control group fermented milk. Among them, (a) is the texture evaluation, (b) is the flavor evaluation, and (c) is the overall acceptance evaluation. In the figure, F1-7 represents the fermented milk of Bifidobacterium animalis subsp. F1-7, and the control group represents the control group fermented milk. Detailed implementation manners

[0026] Now, various exemplary implementation manners of the present invention will be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, characteristics, and implementation schemes of the present invention.

[0027] It should be understood that the terms described in the present invention are only for describing specific implementation manners and are not used to limit the present invention. Additionally, for the numerical ranges in the present invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Each intermediate value within any stated value or stated range, as well as each smaller range between any other stated value or intermediate value within the stated range, is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded from the range.

[0028] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials related to the documents. In case of conflict with any incorporated document, the content of this specification shall prevail.

[0029] Without departing from the scope or spirit of the present invention, various modifications and variations can be made to the specific embodiments of the description of the present invention, which are obvious to those skilled in the art. Other embodiments obtained from the description of the present invention are obvious to those skilled in the art. The description and examples of the present invention are merely exemplary.

[0030] As used herein, the terms "comprising", "including", "having", "containing", etc. are open-ended terms, meaning including but not limited to.

[0031] Source of the materials used in the present invention: Bifidobacterium animalis subsp F1-7 is derived from the feces of healthy infants. The Bifidobacterium animalis subsp F1-7 was deposited at the China Center for Type Culture Collection on December 2, 2020. The deposit address is Wuhan University, Wuhan, China, and the deposit number is: CCTCC NO: M2020833.

[0032] The nucleotide sequence of the 16S rDNA of Bifidobacterium animalis subsp. F1-7 is shown in SEQ ID NO. 1, SEQ ID NO.1: GGGTGGGGGGCGTTCTTACACATGCAGTCGAACGGGATCCCTGGCAGCTTGCTGTCG GGGTGAGAGTGGCGAACGGGTGAGTAATGCGTGACCAACCTGCCCTGTGCACCGGAATAGCTCCTGGAAACGGGTGGTAATACCGGATGCTCCGCTCCATCGCATGGTGGGGTGGGAAATGCTTTTGCGGCATGGGATGGGGTCGCGTCCTATCAGCTTGTTGGCGGGGTGATGGCCCACCAAGGCGTTGACGGGTAGCCGGCCTGAGAGGGTGACCGGCCACATTGGGACTGAGATACGGCCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGCACAATGGGCGCAAGCCTGATGCAGCGACGCCGCGTGCGGGATGGAGGCCTTCGGGTTGTAAACCGCTTTTGTTCAAGGGCAAGGCACGGTTTCGGCCGTGTTGAGTGGATTGTTCGAATAAGCACCGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGGTGCGAGCGTTATCCGGATTTATTGGGCGTAAAGGGCTCGTAGGCGGTTCGTCGCGTCCGGTGTGAAAGTCCATCGCCTAACGGTGGATCTGCGCCGGGTACGGGCGGGCTGGAGTGCGGTAGGGGAGACTGGAATTCCCGGTGTAACGGTGGAATGTGTAGATATCGGGAAGAACACCAATGGCGAAGGCAGGTCTCTGGGCCGTCACTGACGCTGAGGAGCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGGTGGATGCTGGATGTGGGGCCCTTTCCACGGGTCCCGTGTCGGAGCCAACGCGTTAAGCATCCCGCCTGGGGAGTACGGCCGCAAGGCTAAAACTCAAAGAAATTGACGGGGGCCCGCACAAGCGGCGGAGCATGCGGATTAATTCGATGCAACGCGAAGAACCTTACCTGGGCTTGAC。.

[0033] Example 1

[0034] I. Effects of Bifidobacterium animalis subsp. F1-7 on improving sarcopenia in mice

[0035] 1. Experimental animals: Male C57BL / 6J mice at 8 weeks old were purchased from Jinan Pengyue Laboratory Animal Breeding Co., Ltd. During the feeding period, they were exposed to 12-hour day-night cycle lighting, with free access to water and facilities. All mice were housed in the breeding experimental room of the Marine Biological Resources Development Center of Ocean University of China. The experiment was approved by the Laboratory Animal Ethics Committee of the College of Food Science and Engineering, Ocean University of China, and the ethical review number was SPXY2024032601.

[0036] 2. Experimental method: Dexamethasone was used to construct a sarcopenia model in mice for 10 days and the model was verified. During the modeling period, mice in the sarcopenia mouse model group (SM group) and the Bifidobacterium animalis subsp. F1-7 intervention group (F1-7 group) were intraperitoneally injected with 10 mg / kg / d dexamethasone injection every day. Mice in the normal mouse group (HM group) were only intraperitoneally injected with normal saline corresponding to their body weight. The subcutaneous administration volume was 0.1 mL / 10 g for 10 consecutive days. After successful construction, the mice were intervened by gavage. Each mouse in the F1-7 group was gavaged with 0.1 mL of Bifidobacterium animalis subsp. F1-7 bacterial suspension with a concentration of 1×10 9 CFU / mL. The HM group and the SM group were gavaged with an equal amount of PBS (purchased from Beijing Solarbio Science & Technology Co., Ltd.). Gavage continued for 5 weeks. The body weight of the mice was recorded weekly, and the motor ability was tested. After the intervention, all mice were fasted but allowed water for 12 hours, weighed, and then anesthetized and sacrificed by intraperitoneal injection of 2% pentobarbital sodium (0.25 mL / 100 g). Blood was collected from the orbital venous plexus, and the serum was separated for biochemical index detection; muscle tissues were separated, part of which was fixed in 4% paraformaldehyde for pathological analysis, and part was used to prepare muscle homogenates; ileum, colon tissues and colon contents were separated, and the above samples were all stored in a -80°C refrigerator for subsequent detection.

[0037] (1) After the mice were sacrificed, the tibialis anterior muscle (TA), extensor digitorum longus (EDL), gastrocnemius muscle (GA), and soleus muscle (SOL) were quickly dissected and weighed, and the skeletal muscle index was calculated (skeletal muscle index = muscle wet weight / fasting body weight × 100%). The motor ability of the mice was analyzed based on the tensile monitoring data during the experimental process.

[0038] (2) Histological observation of muscle tissues was performed by preparing H&E-stained sections of the tibialis anterior muscle and extensor digitorum longus of the mice.

[0039] (3) The gene expression levels of skeletal muscle atrophy markers ubiquitin ligase Atrogin-1 and MuRF-1 in mouse muscle tissues were analyzed by qRT-PCR. Total muscle RNA was extracted using Trizol reagent. cDNA was synthesized using a reverse transcription kit (purchased from Shanghai Abm Biotechnology Co., Ltd.) according to the instructions. The gene levels of Atrogin-1 and MuRF-1 expressed in muscle were detected using a fluorescence quantitative PCR instrument. The relative amounts of mRNA transcripts were calculated by the 2-ΔΔCT method and normalized by the control β-actin. The primers used are shown in Table 1.

[0040] Table 1 Primers for qRT-PCR

[0041]

[0042] 3. Test results:

[0043] The improvement effect of mouse muscle mass was obtained by weighing the mouse muscle mass and calculating the skeletal muscle index, as Figure 1 shown in (a), Figure 1 shown in (b), and Figure 1 shown in (c). It can be seen that compared with the HM group, the F1-7 group, that is, the treatment with Bifidobacterium animalis subsp. F1-7, significantly reduced the muscle mass loss caused by sarcopenia; by analyzing the change in mouse pulling force to analyze the change in motor ability, it was found that after the intervention of the F1-7 group, that is, Bifidobacterium animalis subsp. F1-7, the decline in motor ability of sarcopenic mice was significantly improved, and the muscle strength was significantly improved.

[0044] Muscle histology analysis was performed by observing the morphology of the muscles of each group of mice under an optical microscope. As Figure 2 shown in (a), Figure 2 shown in (b), and Figure 2 shown in (c), it can be seen that Figure 2 in (a), the muscle fiber structure of the mice in the HM group was intact, and the muscle fibers were arranged neatly and tightly; in contrast, Figure 2 in (b) of the SM group, the skeletal muscle fibers of the mice became thinner, the gaps between the muscle fibers increased, and the muscle fibers were not arranged neatly. Figure 2 In (c) of the F1-7 group, that is, after the intervention of Bifidobacterium animalis subsp. F1-7, compared with the SM group, the muscle fibers were denser, the gaps decreased, and the muscle mass was improved.

[0045] By detecting the mRNA expression levels of Atrogin-1 and MuRF-1 in the muscle, the improvement effect of Bifidobacterium animalis subsp. F1-7 on sarcopenia can be effectively reflected. As Figure 3 shown in (a) and Figure 3As shown in (b), sarcopenia can cause the upregulation of the expression of muscle atrophy markers, while the F1-7 group, i.e., treatment with animal Bifidobacterium subspecies F1-7, can significantly reduce the expression levels of MuRF-1 and Atrogin-1, thereby improving muscle atrophy.

[0046] 2. Mechanism of Bifidobacterium animalis subspecies F1-7 in improving sarcopenia in mice

[0047] 1. Experimental animals and methods: Based on the above experimental animals, intestinal tissue fixed in paraformaldehyde was sent for immunohistochemical staining to analyze the protein expression of intestinal farnesoid X receptor (FXR). The primary antibody was anti-FXR1 rabbit monoclonal antibody (purchased from Absin Shanghai Biotechnology Co., Ltd.), and the sections were stained with 3,3'-diaminobenzidine (DAB). The images were captured using a microscope at 40×, and the mean fluorescence intensity was quantitatively analyzed using Image J software. The level of mouse fibroblast growth factor 15 (FGF15) was measured using a kit (purchased from Shanghai Jianglai Biotechnology Co., Ltd.).

[0048] qRT-PCR was used to analyze the gene expression of intestinal signaling pathway factors FXR and FGF5, as well as FGF15 receptor fibroblast growth factor receptor 4 (FGFR4) and Klotho-β protein (KLB) in muscle. Total RNA from ileum and muscle was extracted using Trizol reagent. cDNA was synthesized using a reverse transcription kit (purchased from Shanghai Aibimeng Biotechnology Co., Ltd.) according to the instructions, and the gene levels of FXR and FGF5 expressed in intestinal epithelial cells and FGFR4 and KLB expressed in muscle tissue were detected using a fluorescent quantitative PCR instrument. The relative amount of mRNA transcripts was calculated by the 2-ΔΔCT method and normalized by the control β-actin. The primers used are detailed in Table 2.

[0049] Table 2 Primers for qRT-PCR

[0050]

[0051] 2. Test results:

[0052] Immunohistochemical staining and Elisa were used to analyze the protein expression of FXR / FGF15 in the ileum. Figure 4 Middle (a), Figure 4 Middle (b), Figure 4 Middle (c), Figure 4 Middle (d), Figure 4 Middle (e), Figure 4 Middle (f) and Figure 4 The results in (g) show that the intervention of F1-7 group, namely Bifidobacterium animalis subspecies F1-7, can significantly improve the decrease of FXR protein level in the intestine of mice (p<0.05,Figure 4 In (d) of Figure 6, and further increase the level of FGF15 in the body through the FXR / FGF15 pathway; the qRT-PCR results further confirmed that the intervention of Bifidobacterium animalis subsp. F1-7 in the F1-7 group could effectively increase the expression of FXR and FGF15 genes in the intestine of sarcopenia mice ( Figure 4 in (d) of Figure 6 and Figure 4 in (e) of Figure 6); after the intervention of Bifidobacterium animalis subsp. F1-7 in the F1-7 group, the expression levels of FGFR4 and KLB, the receptor genes of FGF15 in muscle, were also increased ( Figure 4 in (f) of Figure 6 and Figure 4 in (g) of Figure 6), further verifying the realization of the FXR / FGF15 pathway.

[0053] III. Regulatory effect of Bifidobacterium animalis subsp. F1-7 on intestinal flora

[0054] 1. Experimental animals and methods: Based on the above experimental animals, fecal samples were collected at the end of the experiment, immediately placed in liquid nitrogen and then transferred to -80 °C for storage. The changes in intestinal flora were analyzed by 16S rRNA microbiota gene sequencing.

[0055] 2. Experimental results: The results of 16S rRNA microbiota gene sequencing analysis of fecal samples are as Figure 5 shown. The intervention of Bifidobacterium animalis subsp. F1-7 in the F1-7 group increased the relative abundances of Enterococcus and Bifidobacterium while decreasing the relative abundances of Dubosiella and Desulfovibrio.

[0056] IV. Regulatory effect of Bifidobacterium animalis subsp. F1-7 on bile acid metabolism

[0057] 1. Experimental animals and methods: Based on the above experimental animals, fecal samples were collected at the end of the experiment, immediately placed in liquid nitrogen and then transferred to -80 °C for storage. The changes in bile acids were analyzed by targeted metabolomics.

[0058] 2. Experimental results: Through analysis, the metabolic levels of the FXR-activated bile acids chenodeoxycholic acid (CDCA) and deoxycholic acid (DCA) in the SM group were generally lower than those in the HM group. As Figure 6 shown in (a) of Figure 7, Figure 6 in (b) of Figure 7, Figure 6 in (c) of Figure 7, Figure 6 and in (d) of Figure 7, the intervention of Bifidobacterium animalis subsp. F1-7 in the F1-7 group significantly increased the levels of CDCA and DCA, significantly decreased the levels of T-β-MCA and 7-ketodeoxycholic acid, improved the bile acid metabolism in sarcopenia, and promoted the activation of FXR.

[0059] Example 2

[0060] I. Application of Bifidobacterium animalis subsp. F1-7 as an adjunct culture in the preparation of fermented milk

[0061] 1. Test method: The strain of Bifidobacterium animalis subsp. F1-7 needs to be cultured in MRS liquid medium (purchased from Qingdao Haibo Biotechnology Co., Ltd.) at 37 °C for 48 h and passaged twice. The specific steps of subculture are to inoculate the activated bacterial liquid cultured for 48 h into fresh MRS medium at a ratio of 2% by volume, place it in an upright culture bag, and anaerobically culture it at 37 °C for 48 h. The bacterial liquid of the third generation is used for subsequent experiments to ensure the vitality and adaptability of the strain. Subsequently, pure milk is heated to 55 °C, 6% by volume of granulated sugar is added, stirred and homogenized, then sterilized at 95 °C for 10 min, and cooled to obtain the treated milk. The common inoculation ratio of Bifidobacterium in yogurt is 1×10 7 CFU / mL. At the stage of inoculating the strains, Bifidobacterium animalis subsp. F1-7 and a commercial starter culture of 2 strains (purchased from Angel Biotechnology Co., Ltd.) are inoculated into the treated milk at a volume ratio of 2:1, mixed evenly, and fermented at a constant temperature of 42 °C until the pH reaches 4.5, then the fermentation is terminated. Subsequently, it is placed in a refrigerator at 4 °C for 24 h of after-ripening to improve the flavor and texture, obtaining fermented milk, and stored at 4 °C to ensure the stability of the product and the viable bacteria survival rate. The fermented milk without adding Bifidobacterium animalis subsp. F1-7 (only adding the commercial starter culture, and other steps are the same as the experimental group) is used as the control group for index detection and analysis.

[0062] 2. Test results:

[0063] Table 2 Results of texture analysis of fermented milk

[0064]

[0065]

[0066] In terms of acidity, the pH value of the F1-7 group was stable at about 4.5 after fermentation, slightly lower than that of the control group, indicating that Bifidobacterium animalis subsp. F1-7 can slightly promote lactic acid production. The detection of viable bacteria count showed that the viable bacteria count of the fermented milk in the F1-7 group reached more than 107 CFU / mL, meeting the national standard and having good bacterial survival rate and probiotic characteristics. As shown in Table 2, in terms of texture, the hardness, adhesiveness and elasticity of the fermented milk in the F1-7 group were all improved compared with the control group, and the coagulation state was more stable, indicating that Bifidobacterium animalis subsp. F1-7 can affect the protein network structure and improve the texture characteristics of yogurt. As Figure 7 in (a), Figure 7 in (b) and Figure 7As shown in the sensory evaluation results in (c), the fermented milk of the F1-7 group showed good performance in terms of consistency, taste, and overall acceptance, but was slightly inferior to the control group. Overall, Bifidobacterium animalis subsp. F1-7 can be used as an adjunct starter in the production of fermented milk.

[0067] Bifidobacterium animalis subsp. F1-7 of the present invention can be used as an adjunct starter to prepare functional fermented milk. However, it is not limited thereto, and also includes other forms of dairy products.

[0068] The present invention provides the application of Bifidobacterium animalis subsp. F1-7 in the preparation of drugs for improving sarcopenia. Bifidobacterium animalis subsp. F1-7 can effectively regulate the intestinal flora, improve bile acid metabolism, and thus relieve sarcopenia, and can enrich the probiotic library for improving sarcopenia.

[0069] The above-described embodiments are only descriptions of the preferred embodiments of the present invention, and do not limit the scope of the present invention. Without departing from the design spirit of the present invention, various deformations and improvements made by those of ordinary skill in the art to the technical solutions of the present invention shall fall within the protection scope determined by the claims of the present invention.

Claims

1. Use of Bifidobacterium animalis subsp. F1-7 in the preparation of a medicament for preventing or treating sarcopenia.

2. The application according to claim 1, wherein The 16S rDNA nucleotide sequence of Bifidobacterium animalis subsp. F1-7 is shown as SEQ ID NO.1; Bifidobacterium animalis subsp. F1-7 was deposited at the China Center for Type Culture Collection on December 2, 2020. The deposit address is Wuhan University, Wuhan, China, and the deposit number is: CCTCC NO: M2020833.

3. The application according to claim 1, wherein By alleviating the decrease in muscle mass caused by sarcopenia, improving the decline in motor ability, improving the decline in bile acid metabolism, and correcting the intestinal flora dysbiosis caused by sarcopenia, Bifidobacterium animalis subsp. F1-7 achieves the effect of preventing or treating sarcopenia.

4. The application according to claim 3, wherein Alleviating the decrease in muscle mass caused by sarcopenia means improving the decrease in the level of farnesoid X receptor protein in the intestine, and then increasing the level of fibroblast growth factor 15 in the body through the farnesoid X receptor / fibroblast growth factor 15 pathway, so as to alleviate the decrease in muscle mass caused by sarcopenia.

5. The application according to claim 3, characterized in that, Improving the decline in bile acid metabolism means increasing the levels of chenodeoxycholic acid and deoxycholic acid, and decreasing the levels of T-β-MCA and 7-ketodeoxycholic acid, thereby improving bile acid metabolism.

6. The application according to claim 3, wherein Correcting the intestinal flora dysbiosis caused by sarcopenia means increasing the relative abundances of Enterococcus and Bifidobacterium, while decreasing the relative abundances of Dubosiella and Desulfovibrio, thereby correcting the intestinal flora dysbiosis caused by sarcopenia.

7. Use of Bifidobacterium animalis subsp. F1-7 in the preparation of a drug for improving the decline of bile acid metabolism function caused by sarcopenia, characterized in that, The 16S rDNA nucleotide sequence of Bifidobacterium animalis subsp. F1-7 is shown as SEQ ID NO.1; Bifidobacterium animalis subsp. F1-7 was deposited at the China Center for Type Culture Collection on December 2, 2020. The deposit address is Wuhan University, Wuhan, China, and the deposit number is: CCTCC NO: M2020833.

8. Use of Bifidobacterium animalis subsp. F1-7 in the preparation of a drug for improving gut microbiota dysbiosis caused by sarcopenia, characterized in that, The 16S rDNA nucleotide sequence of Bifidobacterium animalis subsp. F1-7 is shown as SEQ ID NO.1; Bifidobacterium animalis subsp. F1-7 was deposited at the China Center for Type Culture Collection on December 2, 2020. The deposit address is Wuhan University, Wuhan, China, and the deposit number is: CCTCC NO: M2020833.

9. A method for preparing fermented milk using Bifidobacterium animalis subsp. F1-7, characterized in that, It includes the following steps: Heat pure milk to 55°C, add white granulated sugar with a volume fraction of 6%, sterilize at 95°C for 10 min, and cool to obtain treated milk; inoculate Bifidobacterium animalis subsp. F1-7 and a commercial starter of type 2 into the treated milk at a volume ratio of 2:1, ferment at a constant temperature of 42°C until the pH reaches 4.5, terminate fermentation, and ripen at 4°C for 24 h to obtain fermented milk. The 16S rDNA nucleotide sequence of Bifidobacterium animalis subsp. F1-7 is shown as SEQ ID NO.1; Bifidobacterium animalis subsp. F1-7 was deposited at the China Center for Type Culture Collection on December 2, 2020. The deposit address is Wuhan University, Wuhan, China, and the deposit number is: CCTCC NO: M2020833.

10. Use of the fermented milk according to claim 9 in the preparation of a functional food for improving intestinal flora imbalance.

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