Use of ligilactobacillus salivarius in preparing drug for treating obesity and metabolism-related diseases

Abstract

The present application relates to use of Ligilactobacillus salivarius in preparing a drug for preventing and / or treating diseases associated with overweight and / or obesity . The strain can inhibit the weight growth of a high-fat diet-induced obesity model mouse, reduce the blood lipid level of the high-fat diet-induced obesity model mouse, improve the liver injury index of the high-fat diet-induced obesity model mouse, reduce the area of white adipose cells of the high-fat diet-induced obesity model mouse, improve the colon GLP-1 level of the high-fat diet-induced obesity model mouse, improve the intestinal flora imbalance of the high-fat diet-induced obesity model mouse, and improve the liver glycolipid metabolism of the high-fat diet-induced obesity model mouse; and reduce the waist circumference, hip circumference, body fat mass, BMI, and weight of overweight and overweight and / or obesity volunteers, and maintain the muscle content, promoting overweight and overweight and / or obesity people to carry out safe, effective, and sustainable weight management.

Description

Use of lactobacillus salivarius in preparation of medicine for treating obesity and metabolic related diseases

[0001] Cross-reference to Related Applications

[0002] This application claims priority to Chinese Patent Application No. 202410542178.2, filed on April 30, 2024, and titled “Use of Lactobacillus salivarius in Preparation of Medicine for Treating Obesity,” the entire contents of which are incorporated herein by reference. This application also claims priority to Chinese Patent Application No. 202411842434.6, filed on December 13, 2024, and titled “Use of Lactobacillus salivarius in Preparation of Medicine for Treating Metabolic Related Diseases,” the entire contents of which are incorporated herein by reference. TECHNICAL FIELD

[0003] The present application relates to the use of Lactobacillus salivarius in the preparation of medicine for treating obesity, the use of Lactobacillus salivarius in the preparation of medicine for treating metabolic related diseases, and belongs to the field of biological medicine. BACKGROUND

[0004] Obesity is a chronic metabolic disease caused by the interaction of genetic factors, environmental factors and other factors. The World Health Organization (WHO) defines overweight and obesity for adults as follows: a body mass index (BMI) greater than or equal to 25 is overweight; a body mass index greater than or equal to 30 is obese. According to the data of the World Health Organization (WHO) in 2022, there are 2.5 billion adults aged 18 and above worldwide who are overweight, of which more than 890 million adults suffer from obesity. The main features of obesity are excessive weight gain and lipid accumulation. Studies have reported that obesity has become an independent risk factor for various metabolic syndromes such as non-alcoholic fatty liver disease, as well as hyperlipidemia, hyperglycemia, hypertension, and various cardiovascular diseases. Adipose tissue plays an important regulatory role in metabolic disorders, inflammation activation and other aspects caused by obesity. In obesity, adipose tissue not only stores excessive lipids, but also has endocrine function, and the adipokines produced can mediate the occurrence of insulin resistance and other mechanisms, causing metabolic disorders in the body. Therefore, prevention and treatment of obesity is very important for maintaining human health.

[0005] For obesity, there are currently some weight loss drugs on the market, however, these weight loss drugs all have obvious adverse reactions and cannot be taken long-term. For example, orlistat targeting pancreatic lipase often has mild to moderate gastrointestinal adverse reactions such as fatty stool, diarrhea and abdominal pain after taking, and occasionally has serious liver adverse reactions such as cholelithiasis, cholestatic hepatitis and subacute liver failure; and the side effects of GLP-1 receptor agonists represented by semaglutide targeting GLP-1 receptor include nausea, diarrhea, vomiting, constipation, abdominal pain, headache, fatigue, indigestion, dizziness, gastroenteritis, gastroesophageal reflux disease and hypoglycemia in type II diabetes patients. Therefore, it is necessary to find weight loss drugs with good effect and safe and harmless long-term use.

[0006] Ligilactobacillus salivarius is a probiotic existing in the oral cavity and digestive tract of the human body, which has multiple health benefits such as improving intestinal health, enhancing intestinal barrier, reducing inflammation and strengthening immune function. In 2003, Ligilactobacillus salivarius was officially approved by the Ministry of Health as an edible strain and entered the "List of Strains for Food", which has industrial applications in probiotic food and health products. Based on the characteristics of long-term safe and harmless use of probiotics, there are currently related researches trying to use Ligilactobacillus salivarius for the prevention and treatment of obesity, but the effects of Ligilactobacillus salivarius provided by these researches in inhibiting obesity and obesity-related diseases are not ideal. For example, in the patent application with publication number CN113337440A, Ligilactobacillus salivarius MG-587 can reduce weight, but its effect in weight loss is not significant (at the end of the experiment, the body weight of the model group and the drug administration group was compared, the p value was 0.0678, with no statistical difference). Therefore, it is urgent to find Ligilactobacillus salivarius with better weight loss effect to develop weight loss drugs with good effect and safe and harmless long-term use.

[0007] Metabolic diseases are diseases caused by metabolic problems, mainly including type 2 diabetes (T2DM), obesity and non-alcoholic fatty liver disease (NAFLD) and the like. The occurrence of metabolic diseases is usually related to metabolic factors such as insulin resistance, abnormal blood glucose control and lipid metabolism disorder, and these metabolic factors are also closely related to each other. Metabolic diseases have a significant impact on human health, and need to be treated in time to control their development, but the existing drugs for treating metabolic diseases still have significant defects, which seriously limit the clinical use of metabolic disease treatment drugs, and need to be overcome.

[0008] For example, semaglutide, as a GLP-1 analogue, can help the pancreas release insulin, control blood sugar, reduce appetite, and reduce weight by simulating the physiological effects of GLP-1, thereby treating type 2 diabetes and obesity. However, semaglutide has obvious side effects, which can cause nausea, diarrhea, vomiting, constipation, abdominal pain, abdominal distension, and indigestion, and can cause hair loss, bone loss, anemia, and other problems caused by rapid weight loss. In addition, semaglutide can cause aggravation of diabetic retinopathy and diseases such as cholecystitis and cholestasis.

[0009] For example, semaglutide, as a GLP-1 analogue, can help the pancreas release insulin, control blood sugar, reduce appetite, and reduce weight by simulating the physiological effects of GLP-1, thereby treating type 2 diabetes and obesity. However, semaglutide has obvious side effects, which can cause nausea, diarrhea, vomiting, constipation, abdominal pain, abdominal distension, and indigestion, and can cause hair loss, bone loss, anemia, and other problems caused by rapid weight loss. In addition, semaglutide can cause aggravation of diabetic retinopathy and diseases such as cholecystitis and cholestasis.

[0010] Ligilactobacillus salivarius is a probiotic bacteria, and it was officially approved by the Ministry of Health in 2003 as an edible strain and entered the list of strains that can be used in food. It has industrial applications in probiotic food and health products. Based on the safety and harmlessness of long-term probiotic consumption, if Ligilactobacillus salivarius that can improve the level of GLP-1 in the body can be found, it will be very helpful for the clinical treatment of metabolic diseases such as type 2 diabetes. However, at present, there is no report of Ligilactobacillus salivarius that can effectively improve the level of GLP-1 in the body through in vivo experiments. SUMMARY

[0011] To solve the above problems, the present application provides the use of Ligilactobacillus salivarius XA-1416 in the preparation of a medicament, wherein the medicament has any one of the following functions:

[0012] (a) preventing and / or treating obesity;

[0013] (b) preventing and / or treating obesity-related diseases;

[0014] (c) regulating intestinal flora;

[0015] The Lactobacillus salivarius XA-1416 is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 40307, and the preservation date is September 5, 2022.

[0016] In an embodiment of the present application, the obesity-related disease includes obesity-induced metabolic syndrome, obesity-induced cardiovascular disease, and / or obesity-induced intestinal flora imbalance.

[0017] In an embodiment of the present application, the obesity-induced metabolic syndrome includes non-alcoholic fatty liver; the obesity-induced cardiovascular disease includes hyperlipidemia (high blood lipids) and / or hyperglycemia.

[0018] In an embodiment of the present application, the prevention and / or treatment of obesity includes inhibiting body weight gain, reducing white adipocyte area, inhibiting preadipocyte differentiation into adipocytes, reducing liver cell lipid accumulation, improving blood lipid levels, improving liver damage indicators, improving bile acid metabolism, increasing intestinal farnesoid X receptor (FXR) antagonist levels, and / or improving intestinal flora.

[0019] In an embodiment of the present application, the prevention and / or treatment of obesity-related diseases includes inhibiting body weight gain, reducing white adipocyte area, inhibiting preadipocyte differentiation into adipocytes, reducing liver cell lipid accumulation, improving blood lipid levels, improving liver damage indicators, improving bile acid metabolism, increasing intestinal farnesoid X receptor (FXR) antagonist levels, and / or improving intestinal flora.

[0020] In an embodiment of the present application, the improvement of blood lipid levels includes reducing serum total cholesterol (TC) and / or low-density lipoprotein cholesterol (LDL-C).

[0021] In an embodiment of the present application, the improvement of liver damage indicators includes reducing serum alanine aminotransferase (ALT) levels.

[0022] In an embodiment of the present application, the improvement of bile acid metabolism includes promoting the conversion of intestinal conjugated bile acids to free bile acids.

[0023] In an embodiment of the present application, the increase of intestinal farnesoid X receptor antagonist levels includes increasing the levels of ursodeoxycholic acid (UDCA), glyco-ursodeoxycholic acid (GUDCA), and / or tauroursodeoxycholic acid (TUDCA) in the intestine.

[0024] In an embodiment of the present application, the improving the intestinal flora comprises reducing the abundance of obesity-related bacteria in the intestine, increasing the abundance of beneficial bacteria in the intestine, improving the ratio of obesity-related bacteria in the intestine, and / or improving the intestinal microbiota functional pathway; the obesity-related bacteria refers to bacteria enriched in the intestine of obese individuals.

[0025] In an embodiment of the present application, the obesity-related bacteria comprises Lachnospiraceae, Acetatifactor, and / or Lachnoclostridium.

[0026] In an embodiment of the present application, the beneficial bacteria comprises Lactobacillus and / or Bifidobacterium.

[0027] In an embodiment of the present application, the ratio of obesity-related bacteria comprises Firmicutes / Bacteroidetes.

[0028] In an embodiment of the present application, the improving the intestinal microbiota functional pathway comprises promoting the function of energy metabolism and lipid metabolism related pathways in the intestine.

[0029] In an embodiment of the present application, the energy metabolism and lipid metabolism related pathways comprise pentose phosphate pathway, 4-deoxy-L-threo-hex-4-enopyranuronate degradation pathway, and / or Mevalonate pathway.

[0030] In an embodiment of the present application, the obesity-related disease is hyperlipidemia; the preventing and / or treating hyperlipidemia comprises improving blood lipid levels, improving bile acid metabolism, and / or increasing the level of intestinal farnesoid X receptor (FXR) antagonists.

[0031] In an embodiment of the present application, the obesity-related disease is non-alcoholic fatty liver; the preventing and / or treating non-alcoholic fatty liver comprises reducing liver cell lipid accumulation, improving blood lipid levels, improving liver damage indicators, improving bile acid metabolism, and / or increasing the level of intestinal farnesoid X receptor (FXR) antagonists.

[0032] In an embodiment of the present application, the obesity-related disease is obesity-induced intestinal flora imbalance; the preventing and / or treating obesity-induced intestinal flora imbalance comprises reducing the abundance of obesity-related bacteria in the intestine, increasing the abundance of beneficial bacteria in the intestine, improving the ratio of obesity-related bacteria in the intestine, and / or improving the intestinal microbiota functional pathway.

[0033] In an embodiment of the present application, the adjusting the intestinal flora comprises increasing the abundance of intestinal probiotics and / or reducing the abundance of intestinal harmful bacteria.

[0034] In an embodiment of the present application, the intestinal probiotics comprise Bifidobacterium and / or Lactobacillus; and the intestinal harmful bacteria comprise Clostridium perfringens, Enterococcus and / or Enterobacter.

[0035] In an embodiment of the present application, the obesity comprises high-fat diet-induced obesity, chemical drug-induced obesity and / or genetically modified obesity.

[0036] In an embodiment of the present application, the obesity-related disease comprises high-fat diet-induced obesity-related disease, chemical drug-induced obesity-related disease and / or genetically modified obesity.

[0037] In an embodiment of the present application, the medicament further comprises a pharmaceutical carrier and / or a pharmaceutical excipient.

[0038] In an embodiment of the present application, the pharmaceutical carrier comprises microcapsules, microspheres, nanoparticles and / or liposomes.

[0039] In an embodiment of the present application, the pharmaceutical excipient comprises solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavorings, preservatives, suspending agents, coating materials, fragrances, antiadherents, integrating agents, penetration enhancers, pH regulators, buffers, plasticizers, surfactants, antifoaming agents, thickeners, inclusion agents, humectants, absorbents, diluents, flocculants and deflocculants, filter aids and / or release retarders.

[0040] In an embodiment of the present application, the dosage form of the medicament is powder, granules, capsules, tablets, pills or oral liquid.

[0041] In an embodiment of the present application, the number of viable bacteria of the Salivarius Ligilactobacillus XA-1416 in the medicament is not less than 1×10 6 CFU / mL or 1×10 6 CFU / g.

[0042] The technical solution of the present application has the following advantages:

[0043] The present application provides the application of Salivarius Ligilactobacillus XA-1416 in the preparation of a medicament for preventing and / or treating obesity, preventing and / or treating obesity-related diseases, or adjusting intestinal flora.

[0044] (1) Lactobacillus salivarius XA-1416 can effectively inhibit the weight gain of high-fat diet-induced obese model mice caused by high-fat diet;

[0045] (2) Lactobacillus salivarius XA-1416 can significantly reduce the levels of TC (total cholesterol) and LDL-C (low-density lipoprotein cholesterol) in the serum of high-fat diet-induced obese model mice;

[0046] (3) Lactobacillus salivarius XA-1416 can significantly reduce the level of ALT (alanine aminotransferase) in the serum of high-fat diet-induced obese model mice;

[0047] (4) Lactobacillus salivarius XA-1416 can significantly reduce the average area of white adipocytes in the tissues (testicular adipose tissue and liver tissue) of high-fat diet-induced obese model mice;

[0048] (5) Lactobacillus salivarius XA-1416 can significantly increase the proportion of free and conjugated bile acids in the ileal contents of high-fat diet-induced obese model mice, promoting the conversion of conjugated bile acids to free bile acids in the intestines of high-fat diet-induced obese model mice;

[0049] (6) Lactobacillus salivarius XA-1416 can significantly increase the level of FXR antagonists (UDCA, GUDCA and TUDCA) in the intestines of high-fat diet-induced obese model mice;

[0050] (7) Lactobacillus salivarius XA-1416 can significantly improve the intestinal flora imbalance (reduce the abundance of obesity-related bacteria in the intestine, increase the abundance of beneficial bacteria in the intestine, improve the ratio of obesity-related bacteria in the intestine, and improve the functional pathways of intestinal microbiota) of high-fat diet-induced obese model mice caused by high-fat diet;

[0051] (8) Lactobacillus salivarius XA-1416 can significantly inhibit fat accumulation in the Hep G2 cell model;

[0052] (9) Lactobacillus salivarius XA-1416 can significantly inhibit the differentiation of preadipocytes into adipocytes in the 3T3-L1 cell model;

[0053] (10) Lactobacillus salivarius XA-1416 can regulate the intestinal flora (increase the abundance of beneficial bacteria in the intestine and reduce the abundance of harmful bacteria in the intestine).

[0054] Therefore, Lactobacillus salivarius XA-1416 has great application prospects in the preparation of drugs for preventing and / or treating obesity, preventing and / or treating obesity-related diseases (including obesity-induced hyperlipidemia, obesity-induced non-alcoholic fatty liver and / or obesity-induced intestinal flora imbalance, etc.), or regulating the intestinal flora.

[0055] In addition, the Lactobacillus salivarius XA-1416 belongs to intestinal probiotics and is in the "List of Bacterial Strains for Food", which has the advantage of long-term safe and harmless use.

[0056] To solve the above problems, the application provides the application of Ligilactobacillus Salivarius in the preparation of a drug for promoting the secretion of glucagon-like peptide-1, wherein the Ligilactobacillus Salivarius is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 40307.

[0057] In an embodiment of the application, the promotion of the secretion of glucagon-like peptide-1 includes the regulation of bile acid metabolism and / or short-chain fatty acid metabolism.

[0058] In an embodiment of the application, the promotion of the secretion of glucagon-like peptide-1 by regulating bile acid metabolism includes the promotion of the level of hyodeoxycholic acid, the inhibition of the activity of famesoid X receptor while activating the transmembrane receptor G protein-coupled bile acid receptor 5, thereby promoting the secretion of glucagon-like peptide 1.

[0059] In an embodiment of the application, the promotion of the secretion of glucagon-like peptide-1 by regulating short-chain fatty acid metabolism includes the activation of intestinal endocrine cells by promoting the level of short-chain fatty acids, thereby promoting the secretion of glucagon-like peptide 1.

[0060] In an embodiment of the application, the drug further contains a pharmaceutically acceptable excipient.

[0061] In an embodiment of the application, the pharmaceutically acceptable excipient comprises a solvent, a propellant, a solubilizer, a cosolvent, an emulsifier, a coloring agent, a binding agent, a disintegrating agent, a filling agent, a lubricant, a wetting agent, an osmotic pressure regulator, a stabilizer, a glidant, a flavoring agent, a preservative, a suspending agent, a coating material, an aromatic agent, an anti-adhesion agent, an integrating agent, a penetration enhancer, a pH regulator, a buffer, a plasticizer, a surfactant, a foaming agent, an antifoaming agent, a thickening agent, a clathrate agent, a humectant, an absorbent, a diluent, a flocculating agent and a deflocculating agent, a filter aid, and / or a release retardant.

[0062] In an embodiment of the application, the dosage form of the drug is a powder, a granule, a capsule, a tablet, a pill, or an oral liquid.

[0063] In an embodiment of the application, in the drug, the viable bacterial count of the Lactobacillus salivarius XA-1416 is not less than 1×10 6 CFU / mL or 1×10 6 CFU / g.

[0064] The application also provides a use of Ligilactobacillus Salivarius in the preparation of a drug for preventing and / or treating diseases related to lipid metabolism and / or sugar metabolism, wherein the Ligilactobacillus Salivarius is preserved in the China General Microbiological Culture Collection Center on September 5, 2022, and the preservation number is CGMCC No. 40307.

[0065] In an embodiment of the application, the diseases related to lipid metabolism include fatty liver disease and / or obesity; the diseases related to sugar metabolism include diabetes.

[0066] In an embodiment of the application, the fatty liver disease includes non-alcoholic fatty liver disease (NAFLD); the diabetes includes type 1 diabetes (T1DM), type 2 diabetes (T2DM), and / or gestational diabetes.

[0067] In an embodiment of the application, the prevention and / or treatment of diseases related to lipid metabolism and / or sugar metabolism includes increasing the level of glucagon-like peptide 1 (GLP-1) in the intestine, increasing the level of pancreatic polypeptide (PP) in the intestine, increasing the level of Leptin in the intestine, improving lipid metabolism in the liver, and / or improving sugar metabolism in the liver.

[0068] In an embodiment of the application, the increase in the level of glucagon-like peptide 1 (GLP-1) in the intestine includes increasing the level of glucagon-like peptide 1 in the intestine by regulating bile acid metabolism and / or short-chain fatty acid metabolism.

[0069] In an embodiment of the application, the increase in the level of glucagon-like peptide 1 (GLP-1) in the intestine by regulating bile acid metabolism includes increasing the level of hyocholic acid (HCA) in the intestine, inhibiting the activity of farnesoid X receptor (FXR) while activating transmembrane receptor G protein-coupled bile acid receptor 5 (TGR5), thereby promoting the secretion of glucagon-like peptide 1, and thereby increasing the level of glucagon-like peptide 1 in the intestine.

[0070] The increase in the level of glucagon-like peptide 1 (GLP-1) in the intestine by regulating short-chain fatty acid metabolism includes increasing the level of short-chain fatty acid in the intestine, activating intestinal endocrine cells, and thereby increasing the level of glucagon-like peptide 1 in the intestine.

[0071] In an embodiment of the application, the increase in the level of pancreatic polypeptide (PP) in the intestine includes increasing the level of pancreatic polypeptide in the intestine by regulating short-chain fatty acid metabolism.

[0072] In an embodiment of the present application, the increasing the level of pancreatic polypeptide in the intestine by modulating short-chain fatty acid metabolism comprises activating the enteroendocrine cells in the intestine by increasing the level of short-chain fatty acids in the intestine, thereby increasing the level of pancreatic polypeptide in the intestine.

[0073] In an embodiment of the present application, the increasing the level of leptin in the intestine by modulating short-chain fatty acid metabolism comprises activating the enteroendocrine cells in the intestine by increasing the level of short-chain fatty acids in the intestine, thereby increasing the level of leptin in the intestine.

[0074] In an embodiment of the present application, the increasing the level of leptin in the intestine by modulating short-chain fatty acid metabolism comprises activating the enteroendocrine cells in the intestine by increasing the level of short-chain fatty acids in the intestine, thereby increasing the level of leptin in the intestine.

[0075] In an embodiment of the present application, the short-chain fatty acids comprise acetic acid, propionic acid, butyric acid, valeric acid and / or isovaleric acid.

[0076] In an embodiment of the present application, the improving the lipid metabolism in the liver comprises improving the lipid metabolism in the liver by activating the AMPK signaling pathway, the FXR-SHP-SREBP-1c signaling pathway and / or the FXR-PPARa signaling pathway.

[0077] In an embodiment of the present application, the improving the lipid metabolism in the liver by activating the AMPK signaling pathway comprises activating the AMPK signaling pathway by up-regulating the expression of the AMPK gene in the liver, thereby improving the lipid metabolism in the liver.

[0078] the improving the lipid metabolism in the liver by the FXR-SHP-SREBP-1c signaling pathway comprises activating the FXR-SHP-SREBP-1c signaling pathway by up-regulating the expression of the FXR gene in the liver, thereby inhibiting the liver fat synthesis, and further improving the lipid metabolism in the liver.

[0079] the improving the lipid metabolism in the liver by the FXR-PPARa signaling pathway comprises activating the FXR-PPARa signaling pathway by up-regulating the expression of the FXR gene in the liver, thereby promoting the fatty acid oxidation, and further improving the lipid metabolism in the liver.

[0080] In an embodiment of the present application, the improving the glucose metabolism in the liver comprises improving the glucose metabolism in the liver by the AMPK signaling pathway and / or the FXR-SHP signaling pathway.

[0081] In an embodiment of the present application, the improving the glucose metabolism in the liver by the AMPK signaling pathway comprises activating the AMPK signaling pathway by up-regulating the expression of the AMPK gene in the liver, thereby improving the glucose metabolism in the liver.

[0082] The improvement of the sugar metabolism of the liver through the FXR-SHP signal pathway comprises activating the FXR-SHP signal pathway by up-regulating the expression of the FXR gene in the liver, thereby inhibiting hepatic gluconeogenesis, and further improving the sugar metabolism of the liver.

[0083] In an embodiment of the present application, the medicine further comprises a pharmaceutically acceptable excipient.

[0084] In an embodiment of the present application, the pharmaceutically acceptable excipient comprises a solvent, a propellant, a solubilizer, a co-solvent, an emulsifier, a coloring agent, a binder, a disintegrant, a filler, a lubricant, a wetting agent, an osmotic pressure regulator, a stabilizer, a glidant, a flavoring agent, a preservative, a suspending agent, a coating material, an aromatic agent, an anti-adhesion agent, an integrating agent, a penetration enhancer, a pH regulator, a buffer, a plasticizer, a surfactant, a foaming agent, an antifoaming agent, a thickening agent, an inclusion agent, a humectant, an absorbent, a diluent, a flocculating agent and a deflocculating agent, a filter aid, and / or a release retardant.

[0085] In an embodiment of the present application, the dosage form of the medicine is a powder, a granule, a capsule, a tablet, a pill, or an oral liquid.

[0086] In an embodiment of the present application, in the medicine, the viable cell count of the Ligilactobacillus Salivarius XA-1416 is not less than 1 x 10 6 CFU / mL or 1 x 10 6 CFU / g.

[0087] The present application also provides a pharmaceutical composition, wherein the components of the pharmaceutical composition comprise Ligilactobacillus Salivarius; the Ligilactobacillus Salivarius is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 40307; and the pharmaceutical composition has the functions shown in any one of the following:

[0088] (a) promoting the secretion of glucagon-like peptide-1;

[0089] and / or, (b) preventing and / or treating diseases related to lipid metabolism and / or sugar metabolism.

[0090] In an embodiment of the present application, the promotion of the secretion of glucagon-like peptide-1 comprises regulating bile acid metabolism and / or short-chain fatty acid metabolism.

[0091] In an embodiment of the present application, the promotion of the secretion of glucagon-like peptide-1 through the regulation of bile acid metabolism comprises promoting the level of hyocholic acid, thereby inhibiting the activity of famesoid X receptor while activating the transmembrane receptor G protein-coupled bile acid receptor 5, so as to promote the secretion of glucagon-like peptide 1.

[0092] In an embodiment of the present application, the promoting glucagon-like peptide-1 secretion by regulating short-chain fatty acid metabolism comprises activating enteroendocrine cells by promoting the level of short-chain fatty acids, thereby promoting the secretion of glucagon-like peptide 1.

[0093] In an embodiment of the present application, the disease related to lipid metabolism comprises fatty liver disease and / or obesity; the disease related to sugar metabolism comprises diabetes.

[0094] In an embodiment of the present application, the fatty liver disease comprises non-alcoholic fatty liver disease (NAFLD); the diabetes comprises type 1 diabetes (T1DM), type 2 diabetes (T2DM) and / or gestational diabetes.

[0095] In an embodiment of the present application, the preventing and / or treating diseases related to lipid metabolism and / or sugar metabolism comprises increasing the level of glucagon-like peptide 1 (GLP-1) in the intestine, increasing the level of pancreatic polypeptide (PP) in the intestine, increasing the level of Leptin in the intestine, improving lipid metabolism in the liver and / or improving sugar metabolism in the liver.

[0096] In an embodiment of the present application, the increasing the level of glucagon-like peptide 1 (GLP-1) in the intestine comprises increasing the level of glucagon-like peptide 1 in the intestine by regulating bile acid metabolism and / or short-chain fatty acid metabolism.

[0097] In an embodiment of the present application, the increasing the level of glucagon-like peptide 1 in the intestine by regulating bile acid metabolism comprises increasing the level of hyocholic acid (HCA) in the intestine, inhibiting the activity of famesoid X receptor (FXR) while activating transmembrane receptor G protein-coupled bile acid receptor 5 (TGR5), thereby promoting the secretion of glucagon-like peptide 1, and further increasing the level of glucagon-like peptide 1 in the intestine.

[0098] The increasing the level of glucagon-like peptide 1 in the intestine by regulating short-chain fatty acid metabolism comprises activating enteroendocrine cells by increasing the level of short-chain fatty acids in the intestine, thereby increasing the level of glucagon-like peptide 1 in the intestine.

[0099] In an embodiment of the present application, the increasing the level of pancreatic polypeptide (PP) in the intestine comprises increasing the level of pancreatic polypeptide in the intestine by regulating short-chain fatty acid metabolism.

[0100] In an embodiment of the present application, the increasing the level of pancreatic polypeptide in the intestine by regulating short-chain fatty acid metabolism comprises activating enteroendocrine cells by increasing the level of short-chain fatty acids in the intestine, thereby increasing the level of pancreatic polypeptide in the intestine.

[0101] In an embodiment of the present application, the increasing the level of Leptin in the intestine comprises increasing the level of Leptin in the intestine by modulating short chain fatty acid metabolism.

[0102] In an embodiment of the present application, the increasing the level of Leptin in the intestine by modulating short chain fatty acid metabolism comprises increasing the level of Leptin in the intestine by activating intestinal endocrine cells through increasing the level of short chain fatty acids in the intestine.

[0103] In an embodiment of the present application, the short chain fatty acids comprise acetic acid, propionic acid, butyric acid, valeric acid and / or isovaleric acid.

[0104] In an embodiment of the present application, the improving lipid metabolism in the liver comprises improving lipid metabolism in the liver by activating AMPK signaling pathway, FXR-SHP-SREBP-1c signaling pathway and / or FXR-PPARa signaling pathway.

[0105] In an embodiment of the present application, the improving lipid metabolism in the liver by activating AMPK signaling pathway comprises improving lipid metabolism in the liver by activating AMPK signaling pathway through up-regulating the expression of AMPK gene in the liver;

[0106] the improving lipid metabolism in the liver by FXR-SHP-SREBP-1c signaling pathway comprises improving lipid metabolism in the liver by activating FXR-SHP-SREBP-1c signaling pathway through up-regulating the expression of FXR gene in the liver, thereby inhibiting hepatic lipogenesis, and further improving lipid metabolism in the liver;

[0107] the improving lipid metabolism in the liver by FXR-PPARa signaling pathway comprises improving lipid metabolism in the liver by activating FXR-PPARa signaling pathway through up-regulating the expression of FXR gene in the liver, thereby promoting fatty acid oxidation, and further improving lipid metabolism in the liver.

[0108] In an embodiment of the present application, the improving glucose metabolism in the liver comprises improving glucose metabolism in the liver by AMPK signaling pathway and / or FXR-SHP signaling pathway.

[0109] In an embodiment of the present application, the improving glucose metabolism in the liver by AMPK signaling pathway comprises improving glucose metabolism in the liver by activating AMPK signaling pathway through up-regulating the expression of AMPK gene in the liver;

[0110] the improving glucose metabolism in the liver by FXR-SHP signaling pathway comprises improving glucose metabolism in the liver by activating FXR-SHP signaling pathway through up-regulating the expression of FXR gene in the liver, thereby inhibiting hepatic gluconeogenesis, and further improving glucose metabolism in the liver.

[0111] In an embodiment of the present application, the pharmaceutical composition contains a first active ingredient and a second active ingredient; the first active ingredient is Ligilactobacillus Salivarius; and the second active ingredient comprises glucagon-like peptide-1, glucagon-like peptide-1 analogs and / or glucagon-like peptide-1 receptor agonists.

[0112] In an embodiment of the present application, the pharmaceutical composition further contains pharmaceutically acceptable excipients.

[0113] In an embodiment of the present application, the pharmaceutically acceptable excipients include solvents, propellants, solubilizers, co-solvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavorings, preservatives, suspending agents, coating materials, fragrances, anti-adhesion agents, integrating agents, penetration enhancers, pH regulators, buffers, plasticizers, surfactants, foaming agents, antifoaming agents, thickening agents, inclusion agents, humectants, absorbents, diluents, flocculants and deflocculants, filter aids and / or release retardants.

[0114] In an embodiment of the present application, the dosage form of the pharmaceutical composition is powder, granules, capsules, tablets, pills or oral liquid.

[0115] In an embodiment of the present application, the number of viable bacteria of Ligilactobacillus Salivarius XA-1416 in the pharmaceutical composition is not less than 1×10 6 CFU / mL or 1×10 6 CFU / g.

[0116] The technical solution of the present application has the following advantages:

[0117] The present application provides the use of Ligilactobacillus Salivarius XA-1416 in the preparation of a drug for promoting the secretion of glucagon-like peptide-1, or a drug for preventing and / or treating diseases related to lipid metabolism and / or glucose metabolism. Animal experiments show that:

[0118] (1) Ligilactobacillus Salivarius XA-1416 can increase the level of Hyocholic acid (HCA) in the intestinal tract, activate the transmembrane receptor G protein-coupled bile acid receptor 5 (TGR5) and inhibit the activity of farnesoid X receptor (FXR), thereby promoting the secretion of glucagon-like peptide 1 in the intestinal tract, and effectively controlling blood glucose levels;

[0119] (2) L. salivarius XA-1416 can activate intestinal endocrine cells by increasing the level of short-chain fatty acids (including acetic acid, propionic acid, butyric acid, valeric acid, and isovaleric acid, etc.) in the intestine, thereby increasing the levels of glucagon-like peptide 1, pancreatic polypeptide, and leptin in the intestine;

[0120] (3) L. salivarius XA-1416 can activate the AMPK signaling pathway by up-regulating the expression of the AMPK gene in the liver, thereby improving lipid metabolism and glucose metabolism in the liver;

[0121] (4) L. salivarius XA-1416 can activate the FXR-SHP-SREBP-1c signaling pathway by up-regulating the expression of the FXR gene in the liver, thereby inhibiting liver fat synthesis and further improving liver lipid metabolism;

[0122] (5) L. salivarius XA-1416 can activate the FXR-PPARa signaling pathway by up-regulating the expression of the FXR gene in the liver, thereby promoting fatty acid oxidation and further improving liver lipid metabolism;

[0123] (6) L. salivarius XA-1416 can activate the FXR-SHP signaling pathway by up-regulating the expression of the FXR gene in the liver, thereby inhibiting hepatic gluconeogenesis and further improving liver glucose metabolism,

[0124] Therefore, L. salivarius XA-1416 has great application prospects in the preparation of drugs for promoting the secretion of glucagon-like peptide-1, or drugs for preventing and / or treating diseases related to lipid metabolism and / or glucose metabolism (mainly including type 2 diabetes, fatty liver disease, and obesity, etc.).

[0125] The present application also provides the use of L. salivarius in the preparation of a drug for promoting the secretion of glucagon-like peptide-1, characterized in that the L. salivarius is preserved in the China General Microbiological Culture Collection Center with a preservation number of CGMCC No. 40307.

[0126] In an embodiment of the present application, the promotion of the secretion of glucagon-like peptide-1 includes the regulation of bile acid metabolism and / or short-chain fatty acid metabolism.

[0127] In an embodiment of the present application, the promotion of the secretion of glucagon-like peptide-1 by regulating bile acid metabolism includes the promotion of the level of hyocholic acid to inhibit the activity of famesoid X receptor while activating the transmembrane receptor G protein-coupled bile acid receptor 5, thereby promoting the secretion of glucagon-like peptide 1.

[0128] The promoting glucagon-like peptide-1 secretion by regulating short-chain fatty acid metabolism comprises activating intestinal endocrine cells by promoting the level of short-chain fatty acids, thereby promoting the secretion of glucagon-like peptide 1.

[0129] The application also provides a use of Ligilactobacillus Salivarius in the preparation of a medicament for preventing and / or treating diseases related to lipid metabolism and / or sugar metabolism, wherein the Ligilactobacillus Salivarius is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 40307.

[0130] In an embodiment of the application, the diseases related to lipid metabolism comprise fatty liver disease and / or obesity; and the diseases related to sugar metabolism comprise diabetes.

[0131] In an embodiment of the application, the preventing and / or treating diseases related to lipid metabolism and / or sugar metabolism comprises increasing the level of glucagon-like peptide 1 in the intestine, increasing the level of pancreatic polypeptide in the intestine, increasing the level of leptin in the intestine, improving lipid metabolism in the liver, and / or improving sugar metabolism in the liver.

[0132] In an embodiment of the application, the increasing the level of glucagon-like peptide 1 in the intestine comprises increasing the level of glucagon-like peptide 1 in the intestine by regulating bile acid metabolism and / or short-chain fatty acid metabolism.

[0133] In an embodiment of the application, the improving lipid metabolism in the liver comprises improving lipid metabolism in the liver by activating the AMPK signaling pathway, the FXR-SHP-SREBP-1c signaling pathway, and / or the FXR-PPARα signaling pathway.

[0134] In an embodiment of the application, the improving sugar metabolism in the liver comprises improving sugar metabolism in the liver by the AMPK signaling pathway and / or the FXR-SHP signaling pathway.

[0135] The application also provides a pharmaceutical composition, wherein the components of the pharmaceutical composition comprise Ligilactobacillus Salivarius; the Ligilactobacillus Salivarius is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 40307; and the pharmaceutical composition has the functions shown in any one of the following:

[0136] (a) promoting glucagon-like peptide-1 secretion;

[0137] and / or (b) preventing and / or treating diseases related to lipid metabolism and / or sugar metabolism.

[0138] In an embodiment of the present application, the drug has any one of the following functions of preventing and / or treating obesity: increasing BAT / weight ratio and colonic GLP-1 level, improving body shape index, reducing visceral fat, reducing body fat, reducing body weight and BMI, avoiding muscle loss during weight loss, reducing inflammation level, anti-oxidation, anti-aging

[0139] The improved body shape index includes reducing waist circumference, hip circumference and waist-hip ratio of overweight / obese people;

[0140] The reduced inflammation level includes reducing IL-6 gene expression level and increasing IL-10 gene expression level;

[0141] The anti-oxidation includes reducing T-AOC level and MDA level in serum;

[0142] The anti-aging includes increasing LCA level.

[0143] In an embodiment of the present application, the Ligilactobacillus salivarius XA-1416 has high activity of Bile Salt Hydrolase (BSH).

[0144] In an embodiment of the present application, the Ligilactobacillus salivarius XA-1416 is used in combination with a GLP-1 drug,

[0145] The GLP-1 drug is Semaglutide.

[0146] In an embodiment of the present application, the Ligilactobacillus salivarius XA-1416 reduces the increase of pancreatic organ index caused by the GLP-1 drug.

[0147] The present application also provides a method for preventing and / or treating obesity and related diseases, comprising administering Ligilactobacillus salivarius to overweight / obese subjects, the Ligilactobacillus salivarius being deposited with the China General Microbiological Culture Collection Center under the accession number CGMCC No. 40307.

[0148] In an embodiment of the present application, the obesity-related diseases include obesity-induced metabolic syndrome, obesity-induced cardiovascular disease and / or obesity-induced intestinal flora imbalance.

[0149] In an embodiment of the present application, the obesity-induced metabolic syndrome includes non-alcoholic fatty liver disease;

[0150] The obesity-induced cardiovascular disease includes hyperlipidemia and / or hyperglycemia.

[0151] In an embodiment of the present application, the prevention and / or treatment of obesity comprises inhibiting body weight gain, reducing white adipocyte area, inhibiting differentiation of preadipocytes into adipocytes, reducing hepatic lipid accumulation, improving blood lipid level, improving liver injury indicators, improving bile acid metabolism, increasing intestinal farnesoid X receptor antagonist level, and / or improving intestinal flora.

[0152] In an embodiment of the present application, the improvement of blood lipid level comprises reducing serum total cholesterol and / or low-density lipoprotein cholesterol;

[0153] The improvement of liver injury indicators comprises reducing serum alanine aminotransferase level;

[0154] The improvement of bile acid metabolism comprises promoting the conversion of intestinal conjugated bile acids into free bile acids;

[0155] The increase of intestinal farnesoid X receptor antagonist level comprises increasing the levels of ursodeoxycholic acid, glyco-ursodeoxycholic acid, and / or tauroursodeoxycholic acid in the intestine;

[0156] The improvement of intestinal flora comprises reducing the abundance of intestinal obesity-related bacteria, increasing the abundance of intestinal beneficial bacteria, improving the ratio of intestinal obesity-related bacteria, and / or improving intestinal microbiota functional pathways; the obesity-related bacteria refers to bacteria enriched in the intestine of obese individuals.

[0157] In an embodiment of the present application, the obesity-related bacteria comprises Lachnospiraceae, Acetatifactor, and / or Lachnoclostridium;

[0158] The beneficial bacteria comprises Lactobacillus and / or Bifidobacterium;

[0159] The ratio of obesity-related bacteria comprises Firmicutes / Bacteroidetes;

[0160] The improvement of intestinal microbiota functional pathways comprises promoting the function of intestinal energy metabolism and lipid metabolism-related pathways.

[0161] In an embodiment of the present application, the energy metabolism and lipid metabolism-related pathways comprise pentose phosphate pathway, 4-deoxy-L-threo-hex-4-enopyranuronate degradation pathway, and / or mevalonate pathway.

[0162] In an embodiment of the present application, the obesity-related disease is hyperlipidemia; the prevention and / or treatment of hyperlipidemia comprises improving blood lipid levels, improving bile acid metabolism, and / or increasing the level of intestinal farnesoid X receptor antagonists;

[0163] The obesity-related disease is non-alcoholic fatty liver; the prevention and / or treatment of non-alcoholic fatty liver comprises reducing liver cell lipid accumulation, improving blood lipid levels, improving liver damage indicators, improving bile acid metabolism, and / or increasing the level of intestinal farnesoid X receptor antagonists;

[0164] The obesity-related disease is obesity-induced intestinal flora imbalance; the prevention and / or treatment of obesity-induced intestinal flora imbalance comprises reducing the abundance of intestinal obesity-related bacteria, increasing the abundance of intestinal beneficial bacteria, improving the ratio of intestinal obesity-related bacteria, and / or improving intestinal microbiota functional pathways.

[0165] In an embodiment of the present application, the obesity comprises high-fat diet-induced obesity, chemical drug-induced obesity, and / or genetically modified obesity;

[0166] The obesity-related disease comprises high-fat diet-induced obesity-related disease, chemical drug-induced obesity-related disease, and / or genetically modified obesity.

[0167] In an embodiment of the present application, the prevention and / or treatment of obesity comprises improving the ratio of BAT to body weight and the level of colonic GLP-1, improving body shape indicators, reducing visceral fat, reducing body fat, reducing body weight and BMI, avoiding muscle loss during weight loss, reducing inflammation levels, antioxidant, anti-aging

[0168] The improvement of body shape indicators comprises reducing waist circumference, hip circumference, and waist-to-hip ratio in overweight / obese populations;

[0169] The reduction of inflammation levels comprises reducing IL-6 gene expression levels and increasing IL-10 gene expression levels;

[0170] The antioxidant comprises reducing the levels of T-AOC and MDA in serum;

[0171] The anti-aging comprises increasing the level of LCA.

[0172] The present application also provides a method for regulating intestinal flora, comprising administering Ligilactobacillus Salivarius to overweight / obese subjects, the Ligilactobacillus Salivarius is deposited with the China General Microbiological Culture Collection Center, and the deposit number is CGMCC No. 40307.

[0173] In an embodiment of the present application, the viable count of the Ligilactobacillus salivarius XA-1416 is not less than 1x10 6 CFU / mL or 1x10 6 CFU / g.

[0174] The present application also provides a method for preventing and / or treating diseases related to lipid metabolism and / or sugar metabolism, comprising administering Ligilactobacillus salivarius to an overweight / obese subject, wherein the Ligilactobacillus salivarius is preserved in the China General Microbiological Culture Collection Center with a preservation number of CGMCC No. 40307.

[0175] In an embodiment of the present application, the diseases related to lipid metabolism include fatty liver disease and / or obesity; and the diseases related to sugar metabolism include diabetes.

[0176] In an embodiment of the present application, the prevention and / or treatment of diseases related to lipid metabolism and / or sugar metabolism includes increasing the level of glucagon-like peptide 1 in the intestine, increasing the level of pancreatic polypeptide in the intestine, increasing the level of leptin in the intestine, improving lipid metabolism in the liver, and / or improving sugar metabolism in the liver.

[0177] In an embodiment of the present application, the increase in the level of glucagon-like peptide 1 in the intestine includes increasing the level of glucagon-like peptide 1 in the intestine by regulating bile acid metabolism and / or short-chain fatty acid metabolism.

[0178] In an embodiment of the present application, the improvement in lipid metabolism in the liver includes improving lipid metabolism in the liver by activating the AMPK signaling pathway, the FXR-SHP-SREBP-1c signaling pathway, and / or the FXR-PPARα signaling pathway.

[0179] In an embodiment of the present application, the improvement in sugar metabolism in the liver includes improving sugar metabolism in the liver by the AMPK signaling pathway and / or the FXR-SHP signaling pathway.

[0180] In an embodiment of the present application, the viable count of the Ligilactobacillus salivarius XA-1416 is not less than 1x10 6 CFU / mL or 1x10 6 CFU / g.

[0181] The present application also provides the use of Ligilactobacillus salivarius in the preparation of a medicament, characterized in that the medicament has any one of the following functions:

[0182] (a) preventing and / or treating overweight;

[0183] (b) preventing and / or treating overweight-related diseases;

[0184] (c) inhibiting weight gain.

[0185] In an embodiment of the present application, the Ligilactobacillus salivarius XA-1416 is combined with a GLP-1 drug, and the GLP-1 drug is Semaglutide.

[0186] In an embodiment of the present application, the Ligilactobacillus salivarius XA-1416 helps alleviate the adverse effects of GLP-1 drugs on the pancreas;

[0187] The adverse effects are decreased pancreatic organ index and pancreatic tissue damage.

[0188] In an embodiment of the present application, the obesity-related diseases include obesity and / or overweight-induced metabolic syndrome, obesity and / or overweight-induced cardiovascular disease, and obesity and / or overweight-induced intestinal flora imbalance.

[0189] Biological material preservation

[0190] A strain of Ligilactobacillus salivarius XA-1416, taxonomically named Ligilactobacillus salivarius, was preserved in the China General Microbiological Culture Collection Center on September 5, 2022, with the preservation number CGMCC No. 40307 and the preservation address being No. 3, Beichen West Road, Chaoyang District, Beijing. BRIEF DESCRIPTION OF DRAWINGS

[0191] Figure 1: Utilization of carbon sources salicin (a) and rhamnose (b) by Ligilactobacillus salivarius XA-1416.

[0192] Figure 2: Phylogenetic analysis of Ligilactobacillus salivarius XA-1416 with reference genomes (a), and ANI value distribution of genomes with 100% sequence similarity to the 16S rRNA gene sequence of XA-1416 (b).

[0193] Figure 3: Flowchart of the experimental design of the high-fat diet-induced obesity model.

[0194] Figure 4: HE staining of epididymal adipose tissue of mice in each group (x200, scale = 50 pm).

[0195] Figure 5: Average area of epididymal white adipocytes of mice in each group.

[0196] Figure 6: HE staining of liver tissues of mice in each group (x200, scale bar = 50 pm).

[0197] Figure 7: Ratio of free bile acids to conjugated bile acids in ileal contents of mice in two groups.

[0198] Figure 8: Levels of FXR antagonists UDCA, GUDCA and TUDCA in ileal contents of mice in two groups. In Figure 8, A is the level of UDCA; B is the level of GUDCA; C is the level of TUDCA.

[0199] Figure 9: Beta diversity of intestinal microbiota of mice in each group at the ASVs level.

[0200] Figure 10: Beta diversity of intestinal microbiota of mice in each group at the Genus level.

[0201] Figure 11: Beta diversity of intestinal microbiota of mice in model group and XA-1416 group at the ASVs level.

[0202] Figure 12: Beta diversity of intestinal microbiota of mice in model group and XA-1416 group at the Genus level.

[0203] Figure 13: Differences in abundance of intestinal microbiota of mice in three groups on obesity-related bacteria.

[0204] Figure 14: Differences in abundance of intestinal microbiota of mice in three groups on probiotic Lactobacillus and Bifidobacterium.

[0205] Figure 15: Differences in abundance of intestinal microbiota of mice in three groups on specific functions.

[0206] Figure 16: Differences in intestinal microbiota of mice in three groups on the ratio of Firmicutes / Bacteroidetes.

[0207] Figure 17: Effect of saliva combined with Lactobacillus XA-1416 on lipid accumulation of Hep G2 cells.

[0208] Figure 18: Effect of saliva combined with Lactobacillus XA-1416 on adipogenic differentiation of 3T3-L1 cells (preadipocytes).

[0209] In Figures 1-18, *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001.

[0210] Figure 19: Effect of saliva combined with Lactobacillus XA-1416 on colon GLP-1 level of model mice.

[0211] Figure 20: Effect of Lactobacillus salivarius XA-1416 on the level of porcine cholic acid (HCA) in the ileum content of model mice.

[0212] Figure 21: Effect of Lactobacillus salivarius XA-1416 on the level of PP in the colon of model mice.

[0213] Figure 22: Effect of Lactobacillus salivarius XA-1416 on the level of Leptin in the colon of model mice.

[0214] Figure 23: Effect of Lactobacillus salivarius XA-1416 on the expression level of AMPK gene in the liver tissue of model mice.

[0215] Figure 24: Effect of Lactobacillus salivarius XA-1416 on the expression level of FXR gene in the liver tissue of model mice.

[0216] Figure 25: Effect of Lactobacillus salivarius XA-1416 on the expression level of SHP gene in the liver tissue of model mice.

[0217] Figure 26: Effect of Lactobacillus salivarius XA-1416 on the expression level of SREBP-1c gene in the liver tissue of model mice.

[0218] Figure 27: Effect of Lactobacillus salivarius XA-1416 on the expression level of PPAR-α gene in the liver tissue of model mice.

[0219] Figure 28: Effect of Lactobacillus salivarius XA-1416 on the level of Butyric acid in the cecal content of model mice.

[0220] Figure 29: Effect of Lactobacillus salivarius XA-1416 on the level of Acetic acid in the cecal content of model mice.

[0221] Figure 30: Effect of Lactobacillus salivarius XA-1416 on the level of Propionic acid in the cecal content of model mice.

[0222] Figure 31: Effect of Lactobacillus salivarius XA-1416 on the level of Valeric acid in the cecal content of model mice.

[0223] Figure 32: Effect of Lactobacillus salivarius XA-1416 on the level of Isovaleric acid in the cecal content of model mice.

[0224] In Figures 19-32, *, p<0.05; **, p<0.01; ***, p<0.001; ****, p<0.0001.

[0225] Figure 33: High-fat diet-induced preventive obesity mouse model experiment (a) and therapeutic obesity mouse model experiment (b) design flowchart.

[0226] Figure 34: Concentration of conjugated bile acids (a) and activity of bile salt hydrolase (BSH) (b) at each time point.

[0227] Figure 35: Total antioxidant capacity (T-AOC) of serum of each group of mice in the preventive obesity mouse model.

[0228] Figure 36: Malondialdehyde (MDA) level of serum of each group of mice in the preventive obesity mouse model.

[0229] Figure 37: Blood glucose level of each group of mice during OGTT (Oral glucose tolerance test) in the preventive obesity mouse model.

[0230] Figure 38: Area under the curve (AUC) of OGTT of each group in the preventive obesity mouse model.

[0231] Figure 39: HE staining of colon tissue of each group of mice (x200, scale = 100 pm) in the preventive obesity mouse model.

[0232] Figure 40: PAS staining of colon tissue of each group of mice (x200, scale = 100 pm) in the preventive obesity mouse model.

[0233] Figure 41: Fluorescent double staining of colon tissue of each group of mice (x200, scale = 100 pm) in the preventive obesity mouse model.

[0234] Figure 42: LCA level in ileal contents of each group of mice in the preventive obesity mouse model.

[0235] Figure 43: Effect of Lactobacillus salivarius XA-1416 on butyric acid level in cecal contents of obese mice in the preventive obesity mouse model.

[0236] Figure 44: Effect of Lactobacillus salivarius XA-1416 on acetic acid level in cecal contents of obese mice in the preventive obesity mouse model.

[0237] Figure 45: Effect of Lactobacillus salivarius XA-1416 on propionic acid level in cecal contents of obese mice in the preventive obesity mouse model.

[0238] Figure 46: Effect of Salivaria conjuncta Lactobacillus XA-1416 on Valeric acid level in cecal content of obese mice in a preventive model of obesity.

[0239] Figure 47: Effect of Salivaria conjuncta Lactobacillus XA-1416 on Isovaleric acid level in cecal content of obese mice in a preventive model of obesity.

[0240] Figure 48: Effect of Salivaria conjuncta Lactobacillus XA-1416 on GLP-1 level in colon of obese mice in a preventive model of obesity.

[0241] Figure 49: Effect of Salivaria conjuncta Lactobacillus XA-1416 on Hyocholic acid (HCA) level in ileal content of obese mice in a preventive model of obesity.

[0242] Figure 50: Effect of Salivaria conjuncta Lactobacillus XA-1416 on PP level in colon of obese mice in a preventive model of obesity.

[0243] Figure 51: Effect of Salivaria conjuncta Lactobacillus XA-1416 on Leptin level in colon of obese mice in a preventive model of obesity.

[0244] Figure 52: Effect of Salivaria conjuncta Lactobacillus XA-1416 on AMPK gene expression in liver tissue of obese mice in a preventive model of obesity.

[0245] Figure 53: Effect of Salivaria conjuncta Lactobacillus XA-1416 on FXR gene expression in liver tissue of obese mice in a preventive model of obesity.

[0246] Figure 54: Effect of Salivaria conjuncta Lactobacillus XA-1416 on SHP gene expression in liver tissue of obese mice in a preventive model of obesity.

[0247] Figure 55: Effect of Salivaria conjuncta Lactobacillus XA-1416 on SREBP-1c gene expression in liver tissue of obese mice in a preventive model of obesity.

[0248] Figure 56: Effect of Salivaria conjuncta Lactobacillus XA-1416 on PPAR-a gene expression in liver tissue of obese mice in a preventive model of obesity.

[0249] Figure 57: Effect of Salivaria conjuncta Lactobacillus XA-1416 on IL-6 gene expression in liver tissue of obese mice in a preventive model of obesity.

[0250] Figure 58: Effect of Salivaria conjuncta Lactobacillus XA-1416 on IL-10 gene expression in liver tissue of obese mice in a preventive model of obesity.

[0251] Figure 59: Effect of saliva combined with Lactobacillus XA-1416 on the expression of TNF-a gene in liver tissue of obese mice in a preventive obesity mouse model.

[0252] Figure 60: Effect of saliva combined with Lactobacillus XA-1416 on serum TG level of obese mice in a therapeutic obesity mouse model.

[0253] Figure 61: Effect of saliva combined with Lactobacillus XA-1416 on serum TC level of obese mice in a therapeutic obesity mouse model.

[0254] Figure 62: Effect of saliva combined with Lactobacillus XA-1416 on serum LDL-C level of obese mice in a therapeutic obesity mouse model.

[0255] Figure 63: Effect of saliva combined with Lactobacillus XA-1416 on serum HDL-C level of obese mice in a therapeutic obesity mouse model.

[0256] Figure 64: Effect of saliva combined with Lactobacillus XA-1416 on serum ALT level of obese mice in a therapeutic obesity mouse model.

[0257] Figure 65: Effect of saliva combined with Lactobacillus XA-1416 on serum AST level of obese mice in a therapeutic obesity mouse model.

[0258] Figure 66: Effect of saliva combined with Lactobacillus XA-1416 on the ratio of brown adipose tissue (BAT) to body weight of obese mice in a therapeutic obesity mouse model.

[0259] Figure 67: Effect of saliva combined with Lactobacillus XA-1416 on colon GLP-1 level of obese mice in a therapeutic obesity mouse model.

[0260] Figure 68: Effect of saliva combined with Lactobacillus XA-1416 and Semaglutide on body weight (a), epididymal fat (eWAT) weight (b) and food intake (c) of obese mice in a therapeutic obesity mouse model.

[0261] Figure 69: Effect of saliva combined with Lactobacillus XA-1416 and Semaglutide on pancreas organ index of obese mice in a therapeutic obesity mouse model.

[0262] Figure 70: HE staining of pancreas tissue of mice in each group (x200, scale = 50 pm) in a therapeutic obesity mouse model.

[0263] Figure 71: Flowchart of the trial eating process of human trials.

[0264] Figure 72: Effect of saliva Lactobacillus consortium XA-1416 on waist circumference of volunteers in human trial.

[0265] Figure 73: Distribution of the reduction in waist circumference and the percentage of the relevant number of volunteers after taking saliva Lactobacillus consortium XA-1416 in human trial.

[0266] Figure 74: Effect of saliva Lactobacillus consortium XA-1416 on hip circumference of volunteers in human trial.

[0267] Figure 75: Distribution of the reduction in hip circumference and the percentage of the relevant number of volunteers after taking saliva Lactobacillus consortium XA-1416 in human trial.

[0268] Figure 76: Effect of saliva Lactobacillus consortium XA-1416 on waist-to-hip ratio of volunteers in human trial.

[0269] Figure 77: Distribution of the reduction in waist-to-hip ratio and the percentage of the relevant number of volunteers after taking saliva Lactobacillus consortium XA-1416 in human trial.

[0270] Figure 78: Effect of saliva Lactobacillus consortium XA-1416 on subcutaneous fat of volunteers in human trial.

[0271] Figure 79: Effect of saliva Lactobacillus consortium XA-1416 on visceral fat of volunteers in human trial.

[0272] Figure 80: Effect of saliva Lactobacillus consortium XA-1416 on body fat of volunteers in human trial.

[0273] Figure 81: Effect of saliva Lactobacillus consortium XA-1416 on BMI of volunteers in human trial.

[0274] Figure 82: Effect of saliva Lactobacillus consortium XA-1416 on body weight of volunteers in human trial.

[0275] Figure 83: Effect of saliva Lactobacillus consortium XA-1416 on Lean body mass of volunteers in human trial.

[0276] Figure 84: Effect of saliva Lactobacillus consortium XA-1416 on skeletal muscle mass of volunteers in human trial.

[0277] Figure 85: Effect of saliva Lactobacillus consortium XA-1416 on muscle mass of volunteers in human trial.

[0278] In Figures 33-85, *, p<0.05; **, p<0.01; ***, p<0.001; ****, p<0.0001. DETAILED DESCRIPTION

[0279] The following examples are provided to better enable those skilled in the art to further understand the application, and are not intended to limit the content and scope of the application. The application is not limited to the best mode contemplated by the inventors, and is intended to encompass any and all products which fall within the scope of the application along with future products developed by the inventors or others based on the present application, or the like. Any and all such products are intended to fall within the scope of the application.

[0280] The specific experimental procedures or conditions are not specified in the following experimental examples, and can be performed according to the conventional experimental procedures described in the literature or the conditions. The reagents or instruments used are not specified by the manufacturer, and are conventional reagent products that can be obtained commercially. The animal bifidobacterium XA-768 involved in the following examples is described in the patent application text with publication number CN115778987A.

[0281] The culture medium involved in the following examples is as follows:

[0282] MRS solid medium: proteose peptone 10.0 g / L, beef infusion powder 5.0 g / L, glucose 20.0 g / L, sodium acetate 5.0 g / L, yeast extract powder 4.0 g / L, ammonium citrate 2.0 g / L, potassium phosphate dibasic 2.0 g / L, magnesium sulfate 0.2 g / L, Tween 80.0 1 mL / L, agar 15.0 g / L, manganese sulfate 0.05 g / L, pH 6.2.

[0283] MRS liquid medium: proteose peptone 10.0 g / L, beef infusion powder 5.0 g / L, glucose 20.0 g / L, sodium acetate 5.0 g / L, yeast extract powder 4.0 g / L, ammonium citrate 2.0 g / L, potassium phosphate dibasic 2.0 g / L, magnesium sulfate 0.2 g / L, Tween 80.0 1 mL / L, manganese sulfate 0.05 g / L, pH 6.2.

[0284] YCFA solid medium: Tryptone 10.0 g / L, Yeast extract 2.5 g / L, Sodium bicarbonate 4.0 g / L, Glucose 2.0 g / L, Maltose 2.0 g / L, Cellobiose 2.0 g / L, Cysteine hydrochloride 1.0 g / L, Dipotassium hydrogen phosphate 0.25 g / L, Monopotassium phosphate 0.45 g / L, Ammonium sulfate 0.9 g / L, Sodium chloride 0.9 g / L, Magnesium sulfate heptahydrate 0.09 g / L, Calcium chloride dihydrate 0.09 g / L, Resazurin 1.0 mL / L, Hematinic chloride 0.01 g / L, VFA mix 6.2 mL / L (VFA mix: Acetic acid 17.0 mL, Propionic acid 6.0 mL, n-Valeric acid 1.0 mL, Isovaleric acid 1.0 mL, Isobutyric acid 1.0 mL), vitamin solution I 1.0 mL / L (vitamin solution I: Biotin 5.0 mg, Vitamin B12 5.0 mg, p-Aminobenzoic acid 15.0 mg, VB Folic acid 25.0 mg, Vitamin B6 hydrochloride 75 mg), Agar 15.0 g / L, Distilled water to 1 L, pH 7.0.

[0285] BHI liquid medium: Tryptone 10.0 g / L, Sodium Chloride 5.0 g / L, Disodium Hydrogen Phosphate 2.5 g / L, Dextrose 2.0 g / L, Heart Extract Powder 9.8 g / L, Brains Extract Powder 7.7 g / L, pH 7.4.

[0286] Modified Gifu Anaerobic Medium (mGAM): Tryptone 5.0 g / L, Peptone 5.0 g / L, Soy peptone 3.0 g / L, Yeast extract 2.5 g / L, Beef powder 2.2 g / L, Digestive serum powder 10.0 g / L, Beef liver extract powder 1.2 g / L, Glucose 0.5 g / L, Monopotassium phosphate 2.5 g / L, Sodium chloride 3.0 g / L, Soluble starch 5.0 g / L, L-Cysteine 0.3 g / L, L-Arginine 1.0 g / L, L-Tryptophan 0.2 g / L, Sodium thioglycolate 0.3 g / L, pH 7.3.

[0287] ​API medium (oligotrophic medium): Trypticase 5.0 g / L, Yeast extract 5.0 g / L, Sodium Chloride 2.5 g / L, L-tryptophane 0.2 g / L, L-cystine 0.4 g / L, Hemin 0.005 g / L, Vitamin K1 0.01 g / L, Sodium sulfite, pH 7.3.

[0288] The digestive juice involved in the following examples is as follows:

[0289] 0.25% trypsin digestive juice: first, 2.5 g of porcine-derived trypsin (purchased from Gibco Company) and 0.2 g of EDTA were dissolved in PBS buffer (purchased from Solarbio, product number P1020), then the pH was adjusted to 7.4 with HCl, and finally PBS buffer was added to make up to 1 L, thus obtaining 0.25% trypsin digestive juice.

[0290] The preparation method of the bacterial agent and the culture supernatant involved in the following examples is as follows:

[0291] The saliva joint lactobacillus bacterial liquid was inoculated into MRS liquid medium at an inoculation amount of 4% (v / v), and incubated in a 37°C constant temperature incubator for 3 days to obtain a culture liquid. The culture liquid was centrifuged at 8000g for 10 min to obtain saliva joint lactobacillus culture supernatant and saliva joint lactobacillus cells. The saliva joint lactobacillus cells were washed with normal saline and resuspended in a drug solvent (purchased from Solarbio, product number P1020) to obtain a saliva joint lactobacillus bacterial agent, which was stored at -80°C for later use.

[0292] Experimental Example 1: Subspecies determination of saliva joint lactobacillus XA-1416

[0293] In this experimental example, the subspecies determination experiment of saliva joint lactobacillus XA-1416 was carried out, and the specific process was as follows:

[0294] Phenotype Microarray 1 and 2A (purchased from BIOLOG, item number 12111 and 12112) were used to detect the degree of utilization of carbon sources Salicin and L-Rhamnose by Lactobacillus salivarius XA-1416. All Lactobacillus salivarius XA-1416 monoclonal colonies on the anaerobic blood agar medium (purchased from Becton Dickinson) were collected with a throat swab, and a cotton swab was inserted into a sterile saline test tube that had been deoxygenated, the cotton swab was agitated and rubbed against the wall of the test tube to uniformly dissolve all the bacteria on the cotton swab into sterile saline to prepare bacterial suspension A. After centrifugation (20°C, 7000g, 5min) of bacterial suspension A, the supernatant was discarded, and an equal volume of sterile saline was used to resuspend it, and after centrifugation and discarding the supernatant, API medium (oligoculture medium) was added to prepare bacterial suspension B. Bacterial suspension B was added to the Phenotype Microarray 1 and Phenotype Microarray 2A plates, the 96-well plate was sealed with a sealing film, and placed on an enzyme marker, and the growth curve was detected on the machine.

[0295] The results are shown in Figure 1, Lactobacillus salivarius XA-1416 can utilize L-Rhamnose as a carbon source, but not Salicin as a carbon source for growth. According to the classification criteria mentioned in the literature “Morrison Rogosa et al., Species differentiation of oral lactobacilli from man including description of Lactobacillus salivarius nov spec and Lactobacillus cellobiosus nov spec, J. Bacteriol. 65:681-699, 1953.” Lactobacillus salivarius XA-1416 can be clearly classified as Lactobacillus salivarius subsp. salivarius.

[0296] Experimental Example 2: Genome analysis of Lactobacillus salivarius XA-1416

[0297] This experimental example obtains the genome sequence by genome sequencing of Lactobacillus salivarius XA-1416, analyzes the differences between XA-1416 strain and literature reported strains from the genome perspective to show its uniqueness, and predicts the BSH activity potential of XA-1416 strain by predicting the BSH gene in the genome sequence. The specific process is as follows:

[0298] The DNA of Lactobacillus gasseri XA-1416 strain was extracted by SDS method, the quality of the extracted DNA was detected by agarose gel electrophoresis, and the concentration of the extracted DNA was quantified by 2.0. 2.0. The sequencing of the bacterial genome was performed by Novogene Co., Ltd. The 1D library was constructed using the Nanopore platform, and the three-generation sequencing of the bacterial genome was completed; the 350bp small fragment library was constructed using the Illumina platform, and the two-generation sequencing of the bacterial genome was completed. After quality control, the sequences obtained by three-generation and two-generation sequencing were spliced and assembled using Unicycler software, the chromosomal and plasmid sequences were screened, and the chromosomal sequence was assembled into a circular genome (if it is a linear genome, it is a linear genome sequence), i.e. the final 0gap completed graph sequence.

[0299] The reference genome of Lactobacillus gasseri was downloaded to perform core genome-based phylogenetic analysis to determine the species information of strain XA-1416, and to calculate the ANI similarity of the strain and the reference genome, and to analyze the similarity of strain XA-1416 and the reference genome. Among them, the comparative genome analysis and the core genome construction use orthorfinder software, and the phylogenetic analysis uses the best calculation model evaluated by iqtree software to calculate the phylogeny. The average nucleotide similarity (ANI) analysis is calculated using fastANI software.

[0300] The assembled Lactobacillus gasseri XA-1416 genome was predicted using the Prodigal software. The Bile salt hydrolase (BSH) gene sequence downloaded from the UniProt database was used to construct a database, and the potential BSH gene in the genome was found by comparison and search, and the BSH activity potential of the strain was speculated.

[0301] The genome of Lactobacillus gasseri XA-1416 consists of 1 circular genome and 1 plasmid, with a total length of 1,942,197bp and a GC content of 33.04%. The genome encodes 1,831 genes, with a total length of 1.71Mbp, an average gene length of 932bp, and a total length of the coding region accounting for 87.87% of the whole genome. The core genome consisting of 1401 common homologous genes of Lactobacillus gasseri was subjected to phylogenetic analysis, and the results are shown in Figure 2a. The results show that strain XA-1416 clusters with all reference genomes of Lactobacillus gasseri, and therefore XA-1416 belongs to Lactobacillus gasseri.

[0302] Due to the high similarity of 16S rRNA gene sequences of each species under Lactobacillus, using 16S rRNA gene as a standard molecular marker for differential analysis has great limitations (see literature: Kim E, Yang SM, Lim B, Park SH, Rackerby B, Kim HY. Design of PCR assays to specifically detect and identify 37 Lactobacillus species in a single 96well plate. BMC Microbiol. 2020 Apr 15;20(1):96. doi: 10.1186 / s12866-020-01781-z). This includes saliva associated lactobacillus. In this experimental example, the 16S rRNA gene sequence of saliva associated lactobacillus XA-1416 was compared with the NCBI Refseq genome database, and 37 reference genomes with 100% similarity to the 16S rRNA gene sequence of XA-1416 were found. ANI analysis of these 38 genomes found that the ANI similarity of these reference genomes to strain XA-1416 was between 96.84-98.02% (b in Figure 2). These results show that even though XA-1416 has high similarity in 16S rRNA gene sequence with some reference strains, at the genome level, strain XA-1416 has higher uniqueness compared to the reference genomes that have been reported so far.

[0303] The BSH gene alignment search results of saliva associated lactobacillus XA-1416 are shown in Table 1, which shows that the genome of saliva associated lactobacillus strain XA-1416 has two genes with high homology to the reference BSH gene sequence, one of which is located on the circular chromosome of the strain, with a similarity of 53.846% to the reference gene; one is located on the plasmid of the strain, with a similarity of 97.531% to the reference gene. Therefore, it is speculated that saliva associated lactobacillus XA-1416 has BSH activity.

[0304] Table 1 BSH alignment results of saliva associated lactobacillus XA-1416 genome

[0305] Experimental Example 3: Effect of saliva associated lactobacillus XA-1416 on high-fat diet-induced obese model mice

[0306] This experimental example provides an experiment of the effect of saliva associated lactobacillus XA-1416 on high-fat diet-induced obese model mice, and the experimental process is as follows:

[0307] C57BL6 mice (purchased from Guangdong Vantoll Life Experimental Animal Technology Co., Ltd.) were taken, male, 5 weeks (mice age at the beginning of the experiment), 50, after quarantine and adaptive feeding, the high-fat diet feeding group was changed to high-fat diet. After changing to high-fat diet for 1 week, mice were ranked by weight gain except for the normal control group, and the last 20% of mice were eliminated, and randomly divided into groups, 8 in each group. This experiment designed 5 groups, of which Group 1 was the normal control group (Normal diet group), and AIN93M was provided during the experiment, and Groups 2-5 were high-fat diet feeding groups, and obesity models were established by feeding high-fat diet (Research Diets, D12492) for 12 weeks. Group 2 is a high-fat diet-induced obese mouse model group (Model group), Group 3 is a positive control group (Olistat group), and the positive drug Orlistat is administered intragastrically, Group 4 is a probiotic treatment group (XA-1416 group), and the saliva combined with lactobacillus XA-1416 agent is administered intragastrically, and Group 5 is a probiotic treatment group (XA-768 group), and the animal bifidobacterium XA-768 agent is administered intragastrically. The administration period is 11 weeks, and the test drug is administered orally, the positive drug dose is 10 mg / kg, the agent dose is 2 x 10 8 CFU / mouse, the drug volume is 200 μL, the drug solvent is 0.1% (w / v, g / 100 mL) L-cysteine hydrochloride PBS buffer, and the specific experimental design is shown in Figure 3 and Table 2. During the experiment, the mice were observed weekly, and the body weight and food intake of the mice were recorded. At the end of the experiment, fresh mouse feces were taken in a sterile EP tube, frozen in liquid nitrogen, and stored at -80°C, and the mouse was dissected to collect tissues.

[0308] The mouse testicular adipose tissue and liver tissue were fixed with 4% (w / v, g / 100 mL) paraformaldehyde. The fixed tissue was then embedded, and then a paraffin section was prepared using a paraffin section machine, with a thickness of 3 microns. After sectioning, hematoxylin and eosin staining (H&E staining) was performed, and the stained sections needed to be mounted. After mounting, an optical microscope was used for microscopic examination to observe the morphological structure of the tissue and possible pathological changes.

[0309] Take 25 mg of mouse ileum contents and add to a 2 mL thick centrifuge tube, add 2 small steel beads and 400 μL of pre-cooled (pre-cooled to 4°C) precipitant (acetonitrile:methanol = 7:3, v / v) to each tube, and put into a tissue grinder for grinding (grinding conditions are set to power 50 Hz, time 300 s) to obtain a ground liquid sample. Remove the steel beads, and after the ground liquid sample is placed in a -20°C refrigerator for 120 min, centrifuge at 25000 g, 4°C for 15 min, take the supernatant to a 1.5 mL EP tube to obtain supernatant sample A. Add 50 μL of supernatant sample A, standard sample and blank to the corresponding hole sites, and then add 150 μL of internal standard-containing precipitant, seal the aluminum film, shake for 2 min first, and then centrifuge at 4000 rpm, 4°C for 30 min to obtain supernatant sample B. Add 80 μL of pure water to a new V-shaped 96-well plate, take 80 μL of supernatant sample B, seal the aluminum film, shake for 2 min, and then centrifuge at 4000 rpm, 4°C for 2 min to obtain the pretreated sample. The pretreated sample is quantitatively detected by high performance liquid chromatography-tandem mass spectrometry (LC-MS / MS) for 15 kinds of bile acids (cholic acid, lithocholic acid, glycocholic acid, glycochenodeoxycholic acid, glycodeoxycholic acid, glycolithocholic acid, glycoursocholic acid, taurochenodeoxycholic acid, taurodeoxycholic acid, taurocholylithocholic acid, tauroursodeoxycholic acid, ursodeoxycholic acid, chenodeoxycholic acid, deoxycholic acid and taurocholic acid, respectively).

[0310] Take mouse feces and use a DNA extraction kit (purchased from QIAGEN, item number 47016) to extract DNA. PCR amplify the V4 region of the 16S rRNA gene; the PCR amplification reaction is carried out under the condition of a volume of 50 μL, containing 25 μL of 2x Premix Taq, 1 μL of each primer (10 μM) and 3 μL of DNA (20 ng / μL) template; amplify by thermal cycling: 94°C initialization for 5 minutes; 30 cycles of 94°C denaturation for 30 seconds, 52°C annealing for 30 seconds, and 72°C extension for 30 seconds; finally 72°C for 10 minutes of final extension. The length and concentration of the PCR product are detected by 1% (w / v, g / 100 mL) agarose gel electrophoresis. The PCR product is purified, and then Ultra TM IIDNA Library Prep Kit for Sequencing library was generated. Library quality was assessed using Qubit 2.0 Fluorometer. Finally, sequencing was performed on Illumina Nova6000 platform to generate 250bp paired-end short sequences. For the data off the machine, sequences such as adapters, primers, poly-A tails were filtered using cutadapt (V3.4), then the data was annotated for species by Dada2 (V1.22) and silva138 database. Picrust2 (V2.4.1) was used to perform functional prediction analysis for the samples. ASVs (Amplicon sequencing variants) with average abundance lower than 0.1% were filtered out. On R (v3.6.3), various Packages were used to perform species difference analysis (MaAsLin2), Beta diversity analysis (phyloseq), PERMANOVA analysis (vegan v2.6.4), and boxplot drawing for strain and functional abundance (ggplot2) for the samples. For the calculation of the difference between groups, first, the data was log transformed, then ANOVA was used to analyze the difference between the three groups, and finally, Tukey's HSD test (honestly significant difference) was used for multiple comparisons to see the significance between each pair of groups.

[0311] As shown in Tables 3-7, the body weight of the mice in the XA-1416 group was lower than that in the Model group from the 7th week after administration, and was significantly lower than that in the Model group at the end of the experiment, and was lower than that in the Olistat group, while the body weight of the mice in the XA-768 group had no significant difference compared with the Model group. It is shown that saliva combined with Lactobacillus XA-1416 is more effective than Olistat in inhibiting the weight gain of mice, thereby treating obesity, while animal Bifidobacterium XA-768 has no such effect.

[0312] As shown in Tables 8-11, compared with the Normal diet group, the levels of total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and alanine aminotransferase (ALT) in the serum of the Model group mice were significantly increased, and the level of high-density lipoprotein cholesterol (HDL-C) was significantly decreased. As shown in Table 7, compared with the Model group, the levels of total cholesterol (TC) in the serum of the Olistat group and the XA-1416 group mice were decreased, and the level of total cholesterol (TC) in the serum of the XA-1416 group mice was significantly lower than that of the Model group. It is indicated that the saliva combined with Lactobacillus XA-1416 can effectively reduce the level of total cholesterol (TC) in the serum of the high-fat diet-induced obese model mice, thereby treating hyperlipidemia (i.e., hyperlipidemia). In recent years, low-density lipoprotein cholesterol (LDL-C) has replaced total cholesterol (TC) in clinical practice as a key blood lipid measurement index for predicting cardiovascular risk. As shown in Tables 9-10, compared with the Model group, the level of low-density lipoprotein cholesterol (LDL-C) in the serum of the XA-1416 group mice was significantly decreased, which was significantly higher than that of the positive control drug Olistat group. At the same time, the level of high-density lipoprotein cholesterol (HDL-C) in the serum of the XA-1416 group mice did not significantly decrease compared with the Model group. It is indicated that the saliva combined with Lactobacillus XA-1416 can effectively reduce the level of low-density lipoprotein cholesterol (LDL-C) in the serum of the high-fat diet-induced obese model mice, thereby reducing the risk of cardiovascular disease. In addition, alanine aminotransferase (ALT) is a biochemical index widely used in clinical practice to assess the health status of the liver. As shown in Table 10, compared with the Model group, the level of alanine aminotransferase (ALT) in the serum of the XA-1416 group mice was significantly decreased, and the degree of decrease was similar to that of the Olistat group. It is indicated that the saliva combined with Lactobacillus XA-1416 can effectively reduce the level of alanine aminotransferase (ALT) in the serum of the high-fat diet-induced obese model mice, thereby treating non-alcoholic fatty liver.

[0313] As shown in Figures 4-5, compared with the Normal diet group, the fat cell area of the epididymal adipose tissue (i.e., the adipose tissue attached to the epididymis) of the Model group mice increased, and the cell wall was thin. Compared with the Model group, the fat cell area of the epididymal adipose tissue of the Olistat group and the XA-1416 group mice was significantly smaller (the average area of the epididymal adipose tissue of the Normal diet group mice was 0.002394 mm 2 , the average area of the epididymal adipose tissue of the Model group mice was 0.006445 mm 2 , and the average area of the epididymal adipose tissue of the XA-1416 group mice was 0.005129 mm 2). As can be seen from Figure 6, compared with the Normal diet group, the fat cells in the liver tissue of the Model group mice obviously increased, and the white vacuoles became larger. However, the number of fat cells in the liver tissue of the Olistat group and the XA-1416 group was less than that of the Model group, and the liver fat infiltration was obviously smaller than that of the Model group. It is shown that saliva combined with Lactobacillus XA-1416 can effectively reduce the average area of fat cells in the tissues (testicular adipose tissue and liver tissue) of high-fat diet-induced obese model mice, thereby treating obesity and non-alcoholic fatty liver.

[0314] Intestinal microbiota can have a significant impact on the host's metabolic homeostasis by altering bile acid levels. On the one hand, the intestinal microbiota uses bile salt hydrolase (BSH) to decouple conjugated bile acids into free bile acids, which increases the excretion of lipids in feces and reduces the efficiency of lipid digestion and absorption in the intestine, which has a positive impact on lipid metabolism and energy balance (see literature: Yu Wang et al. 2022); on the other hand, the intestinal microbiota plays a key role in the occurrence and development of obesity through the intestinal flora-bile acid-farnesoid X receptor (FXR) signaling pathway. FXR is an important transcription factor mainly present in the liver and ileum, and inhibiting intestinal FXR transcriptional activity can regulate diet-induced lipid and cholesterol metabolism disorders. Studies have shown that FXR dysfunction helps reduce body weight and fat mass in diet-induced obese mice, and the absence of intestinal FXR can effectively prevent weight gain, and FXR antagonists can inhibit adipogenesis by inhibiting the intestinal FXR signaling pathway (see literature: Fei Li et al. 2013). In addition, studies have confirmed that FXR antagonists have the potential to lower blood lipids, and conjugated bile acids in the ileum, such as UDCA (ursodeoxycholic acid), GUDCA (glyco-ursodeoxycholic acid), and TUDCA (tauro-ursodeoxycholic acid), as FXR antagonists, can inhibit the intestinal FXR-FGF15 / 19-FGFR4 pathway, activate bile acid synthase in the liver, promote the conversion of cholesterol to bile acids, and thus reduce cholesterol levels in the blood (see literature: Fengjie Huang et al. 2019), which helps to improve obesity-related metabolic dysfunction in mice (see literature: Lulu Sun et al. 2018). In view of this, promoting the conversion of conjugated bile acids to free bile acids or targeting intestinal FXR strategies may provide a safer and more effective treatment for obesity and obesity-related diseases. As can be seen from Figure 7, the ratio of free bile acids to conjugated bile acids in the ileal contents of mice in the XA-1416 group was significantly higher than that in the Model group (the ratio of free bile acids to conjugated bile acids in the ileal contents of mice in the Model group was 0.5967, and the ratio of free bile acids to conjugated bile acids in the ileal contents of mice in the XA-1416 group was 1.387), indicating that XA-1416 has a significant regulatory effect on bile acid metabolism, converting conjugated bile acids to free bile acids.As can be seen from Figure 8, compared with the Model group, the levels of UDCA (ursodeoxycholic acid), GUDCA (glyco-ursodeoxycholic acid) and TUDCA (tauro-ursodeoxycholic acid) in the XA-1416 group all showed an upward trend, especially the contents of UDCA (ursodeoxycholic acid) and GUDCA (glyco-ursodeoxycholic acid) were significantly up-regulated (the average UDCA level in the ileal contents of mice in the Model group was 42.57 ng / mg, the average UDCA level in the ileal contents of mice in the XA-1416 group was 121.3 ng / mg; the average GUDCA level in the ileal contents of mice in the Model group was 1.449 ng / mg, the average GUDCA level in the ileal contents of mice in the XA-1416 group was 3.324 ng / mg; the average TUDCA level in the ileal contents of mice in the Model group was 373.4 ng / mg, the average TUDCA level in the ileal contents of mice in the XA-1416 group was 495.7 ng / mg), indicating that XA-1416 has a promoting effect on the level of intestinal FXR antagonists. This result confirms that saliva combined with lactobacillus XA-1416 can treat obesity and hyperlipidemia (i.e. hyperlipidemia) through the bile acid metabolic pathway and the FXR signaling pathway at the same time.

[0315] Based on the composition of gut microbiota, beta diversity analysis was performed and PCoA plot was drawn to observe the distribution among groups. As shown in FIGS. 9-10, there were significant differences in species composition among the Normal diet group, the Model group and the XA-1416 group at both ASV and Genus levels (PERMANOVA P=0.001). As shown in FIGS. 11-12, there were also significant differences between the Model group and the XA-1416 group (PERMANOVA P=0.016), indicating that high-fat diet had a significant impact on the composition of intestinal microbiota of mice, and XA-1416 could significantly change the intestinal flora disorder caused by high-fat diet. Further analysis found 3 species significantly differentially enriched in the intestines of obese individuals (FIG. 13), 2 significantly changed probiotics (FIG. 14) and 4 significantly different functional pathways (FIG. 15). As shown in FIGS. 13-14, compared with the Normal diet group, the Model group significantly increased the abundance of Lachnospiraceae family and Acetatifactor and Lachnoclostridium (see documents: Ruixin Liu et al. 2017, Vanessa Palmas et al. 2021, Li Sun et al. 2021, Guoqiang Yao et al. 2021 and Jiali chen et al. 2021), while the XA-1416 group significantly reduced the abundance of the above bacteria compared with the Model group, so that the abundance of these obesity-related genera tended to approach the level of the Normal diet group. As shown in FIG. 15, in the Model group, the pentose phosphate pathway related to energy metabolism and the 4-deoxy-L-threo-hex-4-enopyranuronate degradation pathway and the Mevalonate metabolic pathway related to lipid metabolism were significantly decreased, while in the XA-1416 group, they showed a trend of recovery. In the intestinal microbiota of obese individuals, the increase of the Firmicutes / Bacteroidetes ratio is an important feature (see documents: Yong Fan et al. 2020). As shown in FIG. 16, the Firmicutes / Bacteroidetes ratio of the Model group increased compared with the Normal diet group, while the ratio decreased in the XA-1416 group and the Model group, and was closer to the normal level.These results show that saliva combined with Lactobacillus XA-1416 can significantly reduce the abundance of intestinal bacteria enriched in obese individuals (i.e., obesity-associated bacteria), significantly increase the abundance of beneficial intestinal bacteria, significantly improve the proportion of intestinal bacteria enriched in obese individuals, and significantly improve the intestinal microbial community functional pathways, thereby treating high-fat diet-induced intestinal dysbiosis.

[0316] Table 2 Experimental grouping and administration design

[0317] Table 3 Body weight (g) of the Normal diet group

[0318] Table 4 Body weight (g) of the Model group

[0319] Table 5 Body weight (g) of the Olistat group

[0320] Table 6 Body weight (g) of the XA-1416 group

[0321] Table 7 Body weight (g) of the XA-768 group

[0322] Table 8 Serum cholesterol levels (mmol / L)

[0323] Table 9 Serum low-density lipoprotein levels (mmol / L)

[0324] Table 10 Serum high-density lipoprotein levels (mmol / L)

[0325] Table 11 Serum alanine aminotransferase levels (U / L)

[0326] Experimental Example 4: Regulation of intestinal flora by saliva combined with Lactobacillus XA-1416

[0327] According to the determination criteria for regulating intestinal flora in the Technical Guidelines for Health Food Function Inspection and Evaluation (2023 Edition), the microbiome analysis of the feces of the mice in Experimental Example 3 was performed to evaluate the effect of saliva combined with Lactobacillus XA-1416 on the abundance of beneficial bacteria Bifidobacterium and Lactobacillus in the intestine, as well as potential harmful bacteria Clostridium perfringens, Enterococcus, and Enterobacter. The experimental methods and analysis methods are described in detail in Experimental Example 3.

[0328] As can be seen from FIG. 14, compared with the Normal diet group, the abundance of Bifidobacterium and Lactobacillus in the Model group was significantly decreased, however, after treatment with saliva combined with Lactobacillus XA-1416, the abundance of Bifidobacterium and Lactobacillus was significantly increased compared with the Model group, and Clostridium perfringens, Enterococcus and Enterobacter were not detected before and after treatment with saliva combined with Lactobacillus XA-1416. Therefore, saliva combined with Lactobacillus XA-1416 can regulate intestinal flora.

[0329] Experimental Example 5: Effect of saliva combined with Lactobacillus XA-1416 on lipid accumulation of Hep G2 liver cells

[0330] This experimental example provides an experiment of the effect of saliva combined with Lactobacillus XA-1416 on lipid accumulation of Hep G2 liver cells, and the experimental process is as follows:

[0331] The Hep G2 cells (purchased from the Chinese Academy of Sciences Cell Bank) cultured in a T75 bottle were centrifuged at 250 x g for 5 minutes, the supernatant was discarded, 5 mL of PBS buffer preheated to 37°C was sucked into the culture bottle with a pipette, the cells were gently washed, then 3 mL of 0.25% trypsin digestion solution preheated to 37°C was added to the culture bottle to cover the cells, and the culture bottle was transferred to a cell incubator for digestion. After 2 min of digestion, under a microscope, gaps appeared between the cells, and the cells became round, then the digestion was terminated. 6 mL of complete culture medium (purchased from Gibco) was transferred to the culture bottle, and the digested cells were blown evenly with a pipette gun and then transferred to a 15 mL centrifuge tube for centrifugation at 250 x g for 5 minutes, the supernatant was discarded, and the cell precipitate was obtained. The cell precipitate was resuspended with 1 mL of complete culture medium to obtain cell suspension A. 10 μL of cell suspension A was mixed with 10 μL of trypan blue (purchased from Biyun Tian) to determine the cell count in parallel twice. According to the counting results, the Hep G2 cells were resuspended in fresh complete culture medium preheated to 37°C at a cell concentration of 1.5 x 10 5 The cell suspension B was inoculated in a 96-well black multi-well plate at a seeding amount of 100 μL per well, and then the 96-well plate was transferred to a cell incubator at 37°C, 5% (v / v) CO2 for culture.

[0332] After 24h of culture, the 96-well plates were centrifuged at 250xg for 5min, the supernatant was discarded, and the wells of the 96-well plates were grouped and dosed. For the negative control group, 200μL of complete culture medium was added to the wells; for the model group, 200μL of complete culture medium containing 0.4mM free fatty acids (Palmitic Acid and Oleic Acid at a final concentration ratio of 3:1) was added to the wells; for the positive control group, 200μL of complete culture medium containing 50μM caffeine and 0.4mM free fatty acids (Palmitic Acid and Oleic Acid at a final concentration ratio of 3:1) was added to the wells; for the XA-1416 intervention group, 200μL of complete culture medium containing 5% (here 5% refers to 5% of the total volume of complete culture medium) XA-1416 culture supernatant and 0.4mM free fatty acids (Palmitic Acid and Oleic Acid at a final concentration ratio of 3:1) was added to the wells; for the probiotic culture medium control group, 200μL of complete culture medium containing 5% (here 5% refers to 5% of the total volume of complete culture medium) mGAM culture medium (purchased from Qingdao Haibo Biology) and 0.4mM free fatty acids (Palmitic Acid and Oleic Acid at a final concentration ratio of 3:1) was added to the wells. See Table 12 for details.

[0333] After dosing was complete, the 96-well plates were centrifuged at 250xg for 3min, and then the cells were incubated at 37℃ in a 5% (v / v) cell incubator for 36h. After 36h of incubation, the cells were washed once with PBS buffer, the supernatant was discarded, and 100μL of lipid droplet staining solution (purchased from Biyun Tian, item number C2051M) was added to each well of the 96-well plate, which was incubated at room temperature (25℃) for 20min in the dark. After incubation, the cells were washed twice with PBS buffer. The fluorescence intensity of the cells at Ex / Em = 485 / 535nm was detected after Nile Red staining, and statistical analysis of the data was performed to compare the effects of different interaction components on the accumulation of cell lipids. The results of the detection are shown in Figure 17.

[0334] As shown in Figure 17, compared with the negative control group, the fluorescence signal of the model group increased significantly, indicating that the accumulation of cell lipids in the model group increased after treatment with the inducer. The fluorescence intensity of the positive drug 50μM caffeine decreased significantly compared with the model group. Without affecting the accumulation of cell lipids, 5% XA-1416 culture supernatant showed a significant effect on inhibiting the accumulation of cell lipids compared with the model group (the average fluorescence intensity of the negative control group was 0.08903, the average fluorescence intensity of the model group was 0.09247, and the average fluorescence intensity of the XA-1416 group was 0.08822). In summary, saliva combined with Lactobacillus XA-1416 can inhibit the accumulation of lipids in liver cells.

[0335] Table 12 Grouping and cell treatment method

[0336] Experimental Example 6: Effect of Lactobacillus salivarius XA-1416 on the differentiation of preadipocytes into adipocytes in a 3T3-L1 model

[0337] This experimental example provides an experiment on the effect of Lactobacillus salivarius XA-1416 on the differentiation of preadipocytes into adipocytes in a 3T3-L1 model, and the experimental process is as follows:

[0338] The 3T3-L1 cells (purchased from the Cell Bank of the Chinese Academy of Sciences) cultured in T75 bottles were centrifuged at 250 x g for 5 minutes, the supernatant was discarded, 5 mL of preheated PBS buffer at 37°C was sucked into the culture bottle with a pipette, the cells were gently rinsed, then 3 mL of preheated 0.25% trypsin digestion solution at 37°C was added to the culture bottle to cover the cells and the culture bottle was transferred to the cell incubator for digestion. After 2 minutes of digestion, the intercellular gaps were observed under a microscope, and the digestion was terminated when the cells became round. 6 mL of complete culture medium (purchased from Gibco) was transferred to the culture bottle, and the digested cells were evenly blown with a pipette gun and then transferred to a 15 mL centrifuge tube for centrifugation at 250 x g for 5 minutes. The supernatant was discarded, and the cell pellet was obtained. The cell pellet was resuspended with 1 mL of complete culture medium to obtain cell suspension A. 10 μL of cell suspension A was mixed with 10 μL of trypan blue (purchased from Biyun Tian) to determine the cell count in parallel twice. According to the counting results, 3T3-L1 cells were resuspended in fresh preheated complete culture medium at 37°C to a cell concentration of 3 x 10 4 individuals / mL to obtain cell suspension B. The cell suspension B was inoculated in a 96-well black multi-well plate at a seeding amount of 100 μL per well, and then the 96-well plate was transferred to a cell incubator at 37°C and 5% (v / v) CO2 for culture.

[0339] After 24 hours of culture, the 96-well plate was centrifuged at 250 x g for 5 minutes, the supernatant was discarded, and the wells of the 96-well plate were grouped and sampled. 100 μL of complete culture medium was added to the wells of the negative control group, 200 μL of complete culture medium containing 10 μg / mL of insulin was added to the wells of the model group, 200 μL of complete culture medium containing 1 μM of retinoic acid (RA) and 10 μg / mL of insulin was added to the wells of the positive control group, 200 μL of complete culture medium containing 5% (here 5% refers to 5% of the total volume of the complete culture medium) of XA-1416 culture supernatant and 10 μg / mL of insulin was added to the wells of the XA-1416 intervention group, and 200 μL of complete culture medium containing 5% (here 5% refers to 5% of the total volume of the complete culture medium) of mGAM culture medium (purchased from Qingdao Haibo Biology) and 10 μg / mL of insulin was added to the wells of the probiotic culture medium control group, as shown in Table 13.

[0340] After the addition was completed, the 96-well plate was centrifuged at 250xg for 3 minutes, and then the cells were placed in a 37°C, 5% (v / v) cell incubator for 36h. After 36h of culture, 1 wash was performed using PBS buffer, the supernatant was discarded, 100μL of lipid droplet staining solution (purchased from Biyun Tian, item number C2051M) was added to each well of the 96-well plate, and incubation was performed at room temperature (25°C) for 20 minutes in the dark. After incubation, two washes were performed using PBS buffer. The fluorescence intensity of the cells at Ex / Em = 485 / 535nm was detected after Nile red staining, and statistical analysis of the data was performed to compare the effects of different interaction components on the differentiation of preadipocytes into adipocytes. The results of the detection are shown in Figure 18.

[0341] As shown in Figure 18, compared with the negative control group, the differentiation of preadipocytes into adipocytes in the model group was significantly increased after induction. Compared with the model group, the positive control group (1μM retinoic acid) showed inhibition of the differentiation of preadipocytes into adipocytes, but there was no significant difference. 5% mGAM had no effect on the differentiation of preadipocytes into adipocytes, and compared with the model group, 5% XA-1416 culture supernatant significantly inhibited the differentiation of preadipocytes into adipocytes, and the inhibitory effect was better than that of the positive control drug (the average fluorescence intensity of the negative control group was 0.1506, the average fluorescence intensity of the model group was 0.1746, and the average fluorescence intensity of the XA-1416 group was 0.1313). In summary, Lactobacillus salivarius XA-1416 can inhibit the differentiation of preadipocytes into adipocytes.

[0342] Table 13 Grouping and cell treatment method

[0343] Experimental Example 7: Effect of Lactobacillus salivarius XA-1416 on high-fat diet-induced mice

[0344] This experimental example provides an experiment on the effect of Lactobacillus salivarius XA-1416 on high-fat diet-induced mice, and the experimental process is as follows:

[0345] 1. Experimental method

[0346] C57BL6 mice (male, 5 weeks old, purchased from Guangdong VitoLihua Experimental Animal Technology Co., Ltd.) were taken, and after quarantine and adaptive feeding, the experiment was carried out, and the whole experimental period was 12 weeks. After the start of the experiment, the mice were randomly divided into three groups, 8 in each group, among which, Group 1 is the normal control group (Normal diet group), AIN93M feed is provided during the experiment for 12 weeks, Group 2 is the high-fat diet induced obese mouse model group (Model group), high-fat feed (Research Diets, D12492) is fed during the experiment for 12 weeks to establish an obesity model, and Group 3 is the probiotic treatment group (XA-1416 group), high-fat feed (Research Diets, D12492) is fed during the experiment for 12 weeks to establish an obesity model, and at the same time, salivary combined with lactobacillus XA-1416 agent is administered by gavage, the gavage period is 11 weeks, and the high-fat feed is fed for 1 week before gavage. The dose of the agent is 2x10 8 CFU / mouse, the drug volume is 200μL, the drug solvent is PBS buffer containing 0.1% (w / v, g / 100mL) L-cysteine hydrochloride, and the specific experimental design is shown in Table 14. At the end of the experiment, the colon tissue, cecal contents and ileal contents of the mice were collected for further study.

[0347] The mouse colon tissue and PBS buffer were mixed at a ratio of 1:9 (W / V, 1g:9mL) and then homogenized. The supernatant was collected by centrifugation at 4°C, 12,000 rpm for 10 min. The protein concentration in the supernatant was detected using a nanodrop oneC S2201957 (USA) ultramicro spectrophotometer. The GLP-1 level, Leptin and PP level were determined using an enzyme-linked immunosorbent assay kit (Jiangsu Meilan Industrial Co., Ltd., China).

[0348] The mouse ileum contents were weighed 25 mg into a 2 mL thick centrifuge tube, 2 steel beads and 400 μL of pre-cooled (pre-cooled to 4°C) precipitant (acetonitrile:methanol = 7:3, v / v) were added to each tube, and the tissue grinder was put into the tissue grinder for grinding (the grinding condition was set to power 50 Hz, time 300 s) to obtain the grinding liquid sample. The steel beads were removed, and the grinding liquid sample was placed in a -20°C refrigerator for 120 min, then centrifuged at 25000 g and 4°C for 15 min, and the supernatant was taken into a 1.5 mL EP tube. 20 μL of the sample and the standard curve were taken, 60 μL of pre-cooled 50% methanol was added, shaken for 5 min, precipitated at -20°C for 4 h, centrifuged at 20000 g and 4°C for 15 min, and 20 μL of the supernatant was taken into an EP tube. Next, the derivatization step was performed. 20 μL of 200 mM 3-NPH and 20 μL of 120 mM EDC-6% pyridine mixed solution were added to the EP tube containing 20 μL of the supernatant, shaken and incubated at 25°C for 30 min, centrifuged, and the supernatant was taken for machine analysis. The porcine cholic acid in the pretreated sample was detected by high performance liquid chromatography-tandem mass spectrometry (LC-MS / MS).

[0349] The mouse liver tissue was used to extract total RNA using Trizol reagent (Beijing Ploygen Technologies Inc., Beijing, China), and reverse transcribed into cDNA by high capacity cDNA kit (Beijing Ploygen Technologies Inc., Beijing, China). Real-time PCR analysis was performed using Hiper SYBR Premix reagent to detect the mRNA levels of AMPK, FXR, small heterodimer protein (SHP), sterol regulatory element binding protein (SREBP-1c), and peroxisome proliferator-activated receptor alpha (PPAR-α) in liver tissue, and GAPDH was used as an internal reference for standardization. The primers used for PCR are shown in Table 15.

[0350] 0.1 g of mouse cecal contents was mixed with 1.2 mL of 0.5% (w / v, g / 100 mL) aqueous phosphoric acid solution, and then centrifuged to separate the supernatant. Next, the supernatant was filtered through a 0.22 micron filter, and then subjected to gas chromatography-mass spectrometry (GC-MS) detection (the analysis and measurement work was performed by a professional team of Shanghai Applied Protein Technology Co., Ltd.). The sample was mixed with formic acid and centrifuged to collect the supernatant. Then, ethyl acetate was added to extract, and the upper ethyl acetate phase was obtained. The ethyl acetate phase was mixed with 4-methylvaleric acid, and filtered through an organic phase filter membrane, to detect short-chain fatty acids (the content of short-chain fatty acids was evaluated according to the relative peak area with the standard and 4-methylvaleric acid internal standard).

[0351] 2. Experimental results

[0352] Glucagon-like peptide-1 (GLP-1) is a kind of insulinotropic gut hormone secreted by intestinal L cells. GLP-1 can promote insulin secretion to reduce blood glucose level, and can also slow gastric emptying, reduce appetite, and help control body weight. As can be seen from FIG. 19, compared with mice fed with normal diet, the GLP-1 level in the colon of the model group mice fed with high-fat diet decreased by 51%, and after treatment with XA-1416, the GLP-1 level increased by 49% compared with the model group. As can be seen from the results of FIG. 19, XA-1416 has a significant restoring effect on GLP-1 level.

[0353] Farnesoid X receptor (FXR) is a kind of nuclear receptor protein mainly expressed in tissues such as liver and intestine, and plays a key role in regulating bile acid, lipid and glucose metabolism. Transmembrane receptor G protein-coupled bile acid receptor 5 (TGR5) is a kind of G protein-coupled receptor, and activation of the receptor can induce GLP-1 release in the intestine. After primary bile acid is converted into secondary bile acid by intestinal microbiota in the intestine, it regulates the secretion of GLP-1 in L cells by binding to TGR5 and nuclear receptor FXR. After FXR in the intestine is activated, the secretion of GLP-1 is reduced. Hyocholic acid (HCA) in bile acid is both an activator of TGR5 and an antagonist of FXR. As can be seen from FIG. 20, the level of HCA in mice treated with XA-1416 increased by 110%, which is significantly higher than that in the model group. The results of FIGS. 19-20 show that XA-1416 promotes the secretion of GLP-1 by increasing the level of HCA, activating TGR5 receptor and inhibiting FXR activity at the same time.

[0354] Based on the results of FIGS. 19-20, it can be inferred that Lactobacillus salivarius XA-1416 can promote the secretion of GLP-1 in the intestine by increasing the level of hyocholic acid (HCA) in the intestine, activating transmembrane receptor G protein-coupled bile acid receptor 5 (TGR5) and inhibiting the activity of farnesoid X receptor (FXR) at the same time, thereby effectively controlling blood glucose level and having great application prospect in the treatment of type 2 diabetes.

[0355] Pancreatic polypeptide (PP) is an endogenous polypeptide secreted by PP cells in endocrine pancreas, which can regulate pancreatic secretion, affect liver glycogen storage and gastrointestinal peristalsis and secretion, inhibit appetite, and delay postprandial blood glucose and insulin rise. As can be seen from FIG. 21, compared with the normal diet group, the colon PP level of the model group also decreased by 46%, however, after treatment with XA-1416, the PP level of the XA-1416 group increased by 35% compared with the model group, which is close to the normal level. As can be seen from the results of FIG. 21, XA-1416 also has a positive regulating effect on PP level, and can effectively control blood glucose level by promoting the secretion of PP in the intestine.

[0356] Leptin is a protein secreted by adipocytes, which is closely related to the regulation of food intake, energy metabolism, and fat storage. As can be seen from FIG. 22, compared with the normal diet group, the colon Leptin level of the model group was also significantly reduced by 5%, however, after XA-1416 treatment, the Leptin level of the XA-1416 group was significantly improved, and 100% restored to the normal level. As can be seen from the results of FIG. 22, XA-1416 also has a positive regulation effect on the Leptin level, which can effectively improve lipid metabolism by promoting the secretion of Leptin in the intestinal tract.

[0357] Adenosine monophosphate-activated protein kinase (AMPK) is a key energy sensing molecule, which plays an important role in regulating cell energy balance and promoting metabolic health. As can be seen from FIG. 23, compared with the normal diet group, the expression of AMPK gene in the liver of the model group was significantly reduced, however, after XA-1416 treatment, the expression of AMPK gene in the liver of the XA-1416 group was significantly increased compared with the model group. The results of FIG. 23 show that XA-1416 has a significant up-regulation effect on the expression of liver AMPK gene, which can improve lipid metabolism and glucose metabolism in the liver by activating the AMPK signaling pathway.

[0358] After FXR is activated in the liver, the expression of SREBP-1c is down-regulated in a SHP-dependent manner, thereby inhibiting the generation of liver fat. FXR can also up-regulate PPAR-a to increase fatty acid oxidation metabolism. In addition, FXR reduces hepatic gluconeogenesis through SHP to improve glucose metabolism. As can be seen from FIG. 24, the expression of FXR gene in the liver of the model group was significantly reduced, however, after XA-1416 treatment, the expression level of FXR was significantly increased, indicating that XA-1416 can significantly increase the expression level of FXR. As can be seen from FIGS. 25-27, compared with the normal diet group, the expression of SHP gene in the liver of the model group was significantly decreased, the expression of SREBP-1c gene was significantly increased, and the expression of PPARa gene was significantly decreased, indicating that the metabolic regulation of the liver of the model group was disordered, however, after XA-1416 treatment, the gene expression of SHP, SREBP-1c and PPARa in the liver was significantly restored, close to the normal diet group. In combination with the results of FIGS. 24-27, XA-1416 activates the FXR-SHP-SREBP-1c signaling pathway by increasing the expression level of FXR in the liver, effectively inhibits the synthesis of liver fat, and promotes fatty acid oxidation through the FXR-PPARa signaling pathway. In addition, XA-1416 also reduces hepatic gluconeogenesis through the FXR-SHP pathway, thereby improving glucose metabolism.

[0359] In combination with the results of FIGS. 23-27, it is inferred that Lactobacillus salivarius XA-1416 can effectively improve lipid metabolism and glucose metabolism in the liver by activating the AMPK signaling pathway and the FXR-SHP-SREBP-1c and FXR-SHP signaling pathways, and thus has great application prospects in the treatment of fatty liver disease.

[0360] Short-chain fatty acids increase satiety signals such as GLP-1, Leptin, and PP by binding to specific receptors in the intestinal tract. As can be seen from FIGS. 28-32, the levels of short-chain fatty acids (acetic acid, propionic acid, butyric acid, valeric acid, and isovaleric acid) in the cecal contents of the model group mice decreased significantly compared with the normal diet group. This change may be related to the imbalance of the intestinal microbial community structure in the obese state, thereby affecting the production of short-chain fatty acids. However, after treatment with XA-1416, the levels of short-chain fatty acids such as acetic acid, propionic acid, butyric acid, valeric acid, and isovaleric acid increased significantly, with a 40% increase in butyric acid, and a significant increase of 84%, 43%, and 42% in valeric acid, propionic acid, and acetic acid, respectively. The results of FIGS. 28-32 show that XA-1416 can promote the production of five kinds of short-chain fatty acids, and the increase in the level of short-chain fatty acids has a positive activation effect on intestinal endocrine cells, thereby increasing the secretion of GLP-1, Leptin, and PP.

[0361] In combination with the results of FIGS. 19, 21-22, and 28-32, it is inferred that Lactobacillus salivarius XA-1416 can increase the levels of short-chain fatty acids (including acetic acid, propionic acid, butyric acid, valeric acid, and isovaleric acid) in the intestinal tract, activate intestinal endocrine cells, thereby increasing the levels of glucagon-like peptide 1, pancreatic polypeptide, and leptin in the intestinal tract, and thus effectively control blood glucose levels and improve lipid metabolism, and thus has great application prospects in the treatment of type 2 diabetes and obesity.

[0362] Table 14: Experimental grouping and administration design

[0363] Table 15: Specific primer sequences

[0364] The establishment of a prophylactic experimental model was as follows:

[0365] The present experimental example provides an experiment on the effect of Lactobacillus salivarius XA-1416 on high-fat diet-induced obese model mice, and the experimental process is as follows:

[0366] C57BL6 mice (purchased from Guangdong Vantoll Life Experimental Animal Technology Co., Ltd.) were selected, male, 5 weeks (mice age at the beginning of the experiment), 50, after quarantine and adaptive feeding, the high-fat diet feeding group was changed to high-fat diet. After changing to high-fat diet for 1 week, mice were ranked by weight gain except for the normal control group, and the last 20% of mice were eliminated and randomly divided into groups, 8 mice per group. This experiment designed 5 groups, of which Group 1 was the normal control group (Normal diet group), and AIN93M was provided during the experiment, and Groups 2-5 were high-fat diet feeding groups, and obesity models were established with high-fat diet (Research Diets, D12492) for 12 weeks. Group 2 is a high-fat diet-induced obese mouse model group (Model group), Group 3 is a positive control group of orlistat (Orlistat group), which is administered by gavage with the positive drug orlistat, Group 4 is a probiotic treatment group (XA-1416 group), which is administered by gavage with saliva combined with lactobacillus XA-1416 agent, and Group 5 is a probiotic treatment group (XA-768 group), which is administered by gavage with animal bifidobacterium XA-768 agent. The administration period is 11 weeks, and the test drug is administered by oral gavage, the positive drug dose is 10 mg / kg, the agent dose is 2x108CFU / mouse, the drug volume is 200 μL, and the drug solvent is PBS buffer containing 0.1% (w / v, g / 100 mL) L-cysteine hydrochloride. The specific experimental design is shown in Figure 33(a) and Table 16. During the experiment, the mice were observed and the body weight and food intake of the mice were recorded every week. At the end of the experiment, fresh feces of the mice were collected in sterile EP tubes, frozen in liquid nitrogen, and stored at 80°C, and the tissues were collected by dissection.

[0367] Table 16 Experimental grouping and administration design

[0368] The establishment of the treatment type obese mouse model, the experimental process is as follows:

[0369] The entire experiment is divided into a modeling phase and an intervention phase. 35 male C57BL6 mice (purchased from Guangdong Vantoll Life Experimental Animal Technology Co., Ltd.) aged 8 weeks were selected. During the entire experiment, the normal control group was provided with maintenance feed AIN93M, and the remaining mice were provided with high-fat feed (purchased from Research Diets, catalog number D12492).

[0370] The modeling phase lasted for 8 weeks. At the end of the modeling, the body weight of the mice receiving high-fat feed needed to be at least 20% higher than that of the normal control group mice, which was used as the standard for the successful establishment of the obesity model. After the end of the modeling phase, in addition to the normal control group, the other mice were randomly divided into 4 groups according to the body weight, 7 mice per group. The day of grouping was defined as day 0.

[0371] In the 6-week intervention phase, a total of 5 groups were designed, with Group 1 being the normal control group, Group 2 being the high-fat diet-induced obese mouse model group (HFD group), Group 3 being the positive control group (Semaglutide group), subcutaneously injected with the positive drug Semaglutide twice a week at a dose of 10 nmol / kg, Group 4 being the probiotic treatment group (XA-1416 group), gavaged with XA-1416 bacterial agent daily at a dose of 2 x 10 9 CFU / each, with a dosing volume of 200 μL, and Group 5 being the combination group (Semaglutide + XA-1416 group), subcutaneously injected with the positive drug Semaglutide twice a week at a dose of 10 nmol / kg and gavaged with XA-1416 bacterial agent daily at a dose of 2 x 10 9 CFU / each). During the experiment, the body weight and food intake of the mice were monitored and recorded every week. At the end of the experiment, the mice were dissected to collect tissue samples. The specific experimental design is shown in Figure 33(b) and Table 17.

[0372] Table 17. Experimental grouping and dosing design

[0373] Experimental Example 8: Acquisition of Salivaria lactis XA-1416

[0374] This experimental example provides the acquisition process of Salivaria lactis XA-1416, which is as follows:

[0375] In an anaerobic workstation, 29 frozen fecal samples from healthy people in Shenzhen were mixed by equal volume to obtain a mixed sample. After 0.5 mL of the mixed sample was taken into an anaerobic blood culture bottle and incubated at 37°C in a constant temperature incubator for 3 days, 0.1 mL of the 3-day cultured mixed sample was first taken into 0.9 mL of PBS buffer containing 1 g / L L-cysteine hydrochloride (purchased from Solarbio, catalog number P1020) to obtain a 10 -1 dilution, and then 0.1 mL of the 10 -2 dilution was taken into 0.9 mL of PBS buffer containing 1 g / L L-cysteine hydrochloride to obtain a 10 -3 dilution, and so on to obtain 10 -4 , 10 -5 , 10 -6, 10-7 dilution; 10-5, 10-6 and 10-7 dilutions were coated on the oxygen-removed YCFA solid medium at a coating amount of 100 μL / plate (3 oxygen-removed YCFA solid media for each gradient dilution), and after 3 days of static culture in a 37°C constant temperature incubator, single colonies were picked and inoculated into oxygen-removed BHI liquid medium, and cultured in a 37°C constant temperature incubator for 3 days to obtain bacterial liquid; the strains corresponding to each bacterial liquid were numbered, and the steps recorded in the textbook "Microbiology" (Shen Ping, Chen Xiangdong, ed.) were referred to for Gram staining, strain identification, physiological and biochemical experiments, and genomic identification analysis, and strains with typical characteristics of L. salivarius were selected to obtain strain XA-1416;

[0376] The strain identification process is as follows:

[0377] The XA-1416 bacterial body was taken, and the bacterial genome extraction kit was used to extract the genome of XA-1416. The 27F / 1492R primer pair with sequences shown in SEQ ID NO. 14 and SEQ ID NO. 15 (Table 18) was used to amplify the extracted genome of XA-1416 as a template to obtain the 16S rRNA of XA-1416 (the 16S rDNA sequence of XA-1416 is shown in Table 18 SEQ ID NO. 13); the 16S rDNA of XA-1416 was subjected to nucleic acid sequence alignment in the Blastn program of NCBI, and the results showed that the strain was L. salivarius, which was named L. salivarius XA-1416.

[0378] Table 18 XA-1416 16s and primer sequences

[0379] Experimental Example 9: Functional gene prediction of L. salivarius XA-1416

[0380] Prodigal software was used to predict the coding genes of the assembled L. salivarius XA-1416 genome. The bile salt hydrolase (BSH) gene sequence downloaded from the UniProt database was used to construct the database, and the potential BSH gene in the genome was found by alignment, and the BSH activity potential of the strain was predicted.

[0381] In addition, the BSH gene was further screened using the PLMSearch tool. First, the predicted protein sequence was aligned with its built-in database using PLMSearch to identify proteins highly homologous to the BSH protein. Then, the homologous protein pairs screened by PLMSearch were further precisely aligned and scored using the PLMAlign tool to ensure the accuracy of the results.

[0382] As shown in Table 19, the genome of Lactobacillus salivarius XA-1416 has two genes highly homologous to the reference BSH gene sequence, one of which is located on the circular chromosome of the strain, with a similarity of 97.846% to the reference gene; one is located on the plasmid of the strain, with a similarity of 98.457% to the reference gene, and moreover, the proteins encoded by the two genes have a similarity of more than 99% to the BSH protein. It is therefore inferred that Lactobacillus salivarius XA-1416 has high BSH activity.

[0383] Table 19 BSH alignment results of the genome of Lactobacillus salivarius XA-1416

[0384] Experimental Example 10: In vitro study of BSH activity of Lactobacillus salivarius XA-1416

[0385] This experimental example evaluates the BSH activity of Lactobacillus salivarius XA-1416 in vitro, and the experimental process is as follows:

[0386] The substrate metabolized by the strain is 6 common conjugated bile acids (BS): glycochenodeoxycholic acid, taurodeoxycholic acid, taurochenodeoxycholic acid, glycochenodeoxycholic acid, taurocholic acid, and glycocholic acid, which are divided into high-concentration substrate group (HBS) and low-concentration substrate group (LBS) (Table 20). The initial substrate concentrations of the two groups are the same, and after the same time of culture, the conjugated bile acids and their metabolic rates are determined.

[0387] When 45 mL of bacterial solution reaches the growth plateau, the supernatant is removed by centrifugation, and an equal volume of high-concentration bile salt medium (H) and low-concentration bile salt medium (L) is added, both of which are prepared from M9 medium, so that the initial total bile salt concentrations of the two groups are 6 mM and 1.2 mM, respectively. After 3 hours of culture, samples are taken, centrifuged, and the supernatant is taken for LC-MS detection. Liquid chromatography-mass spectrometry (LC-MS) is used for detection. The metabolic rate of conjugated bile acids is defined as the rate of decrease in the concentration of conjugated bile acids in the strain sample within 3 hours.

[0388] As shown in FIG. 34(a), the concentration of conjugated bile acids decreased with time. Lactobacillus salivarius XA-1416 showed a significant bile salt degrading ability regardless of the initial concentration of conjugated bile acids. As shown in FIG. 34(b), the BSH activity of Lactobacillus salivarius XA-1416 in high and low concentrations of bile salts was 97,200 μg / L / h and 42,500 μg / L / h, respectively. This indicates that Lactobacillus salivarius XA-1416 has high BSH activity.

[0389] M9 medium: glucose 40 g / L, disodium hydrogen phosphate 6 g / L, potassium dihydrogen phosphate 3 g / L, sodium chloride 0.5 g / L, ammonium chloride 1 g / L, calcium chloride 0.01 g / L, magnesium sulfate 0.24 g / L

[0390] Table 20 Preparation method of high concentration bile salt medium (HM) and low concentration bile salt medium (LM)

[0391] Experimental Example 11: Effect of Lactobacillus salivarius XA-1416 on a prophylactic obesity model mouse

[0392] Before the animals were sacrificed, the mice were subjected to an oral glucose tolerance test (OGTT). Before the test, the mice were fasted for 12 hours, and then a glucose solution at a dose of 1.5 g / kg was administered by gavage. After the administration of glucose, blood samples were collected by tail clipping at 0, 15, 30, 60, 90, and 120 minutes, and the blood glucose level was measured. According to the curve of the blood glucose level over time, the area under the curve (AUC) was calculated to evaluate the glucose tolerance of the animals in each group.

[0393] The mice were taken blood from the eyeball, and the blood samples were allowed to clot at room temperature. After the blood samples were centrifuged at 3000 rpm for 10 minutes, the serum was separated. The serum samples were subjected to TG, TC, HDL-C, LDL-C, and ALT detection according to the kit (North China Biotech Co., Ltd.) instructions, and the biochemical indicators were detected using a Mindray BS-420 automatic biochemical analyzer.

[0394] The colon tissues of the mice were subjected to PAS (periodic acid-Schiff) staining. The colon tissues were fixed using 4% (w / v, g / 100 mL) paraformaldehyde, and then dehydrated, paraffin-embedded, and cut into 5 μm-thick sections. The deparaffinized sections were placed in a high iodine solution for 10 minutes, and then transferred to a Schiff reagent for incubation for 15 minutes, followed by staining with Mayer's hematoxylin and differentiation and blueing treatment. The stained tissue sections were then observed for morphology using a 200-fold optical microscope.

[0395] Mouse colon tissues were fixed with 4% paraformaldehyde, dehydrated, embedded, and sectioned (4 pm). After deparaffinization and rehydration, the sections were incubated with primary antibody (ZO-1 and Occludin, dilution ratio of 1:500) at 4°C overnight. Subsequently, the tissue sections were incubated with CY3-labeled secondary antibody and stained with DAPI for counterstaining of the nucleus. These sections were digitally scanned at 400x magnification using a 3DHISTECH Pannoramic MIDI digital slide scanner and analyzed by CaseViewer software.

[0396] Total Antioxidant Capacity (T-AOC) is a general term for the antioxidant capacity of the body's antioxidant system, which can reflect the antioxidant capacity of the body. As can be seen from FIG. 35, compared with the Normal diet group, the total antioxidant capacity (T-AOC) in the serum of the Model group mice was significantly reduced. However, the T-AOC level in the serum of the Orlistat group and the XA-1416 group mice was significantly improved compared with the Model group. Malondialdehyde (MDA) is one of the end products of lipid peroxidation reaction, and its content can indirectly estimate the degree of lipid peroxidation, which is an index reflecting the oxidative damage of the body. As can be seen from FIG. 36, compared with the Normal diet group, the malondialdehyde (MDA) level in the serum of the Model group mice was significantly increased, and the MDA level in the serum of the Orlistat group and the XA-1416 group mice was significantly lower than that of the Model group. The results showed that XA-1416 had significant antioxidant capacity.

[0397] As can be seen from FIG. 37, compared with the Normal diet group, the blood glucose level of the Model group mice rapidly increased after oral glucose, and the decline rate was relatively slow, indicating that the Model group mice had impaired glucose tolerance. However, the blood glucose level of the Orlistat group and the XA-1416 group mice increased significantly lower than that of the Model group, and the blood glucose level returned to normal level after 30 min, indicating that Orlistat and XA-1416 can effectively improve the rapid increase of blood glucose induced by high-fat diet. As can be seen from FIG. 38, the AUC of the Model group was significantly higher than that of the Normal diet group, and the AUC of the Orlistat group and the XA-1416 group was significantly lower than that of the Model group, further confirming that the Orlistat group and the XA-1416 group can effectively alleviate glucose intolerance.

[0398] As shown in FIG. 39, the intestinal glands of the Normal diet group were clear, the crypts were deep and complete, indicating that the colon structure was healthy. Compared with the Normal diet group, the crypts of the Model group were shallow, the glands were irregularly arranged, and the colon was obviously inflamed. The colon tissue structure of the Orlistat group and the XA-1416 group was relatively complete, and the crypts and glands were relatively normal, showing a certain degree of protective effect, and the protective effect of the XA-1416 group was the best.

[0399] As shown in FIG. 40, compared with the Normal diet group, the Model group had reduced colonic mucus secretion and significantly reduced PAS staining, indicating that the colon was damaged and caused dysfunction. The number of goblet cells and the colonic mucus secretion function of the Orlistat group and the XA-1416 group were restored compared with the Model group, and the PAS staining was similar to that of the Normal diet group, indicating that Orlistat and XA-1416 had a certain protective effect on the colon, and the protective effect of XA-1416 was the best.

[0400] As shown in FIG. 41, Occludin and ZO-1 are important tight junction proteins in colon tissue. Compared with the Normal diet group, the expression of Occludin and ZO-1 in the Model group was significantly reduced, indicating that the intestinal barrier function was damaged and the intestinal permeability was increased. The expression of Occludin and ZO-1 in the Orlistat group and the XA-1416 group was improved compared with the Model group, indicating that Orlistat and XA-1416 could effectively improve the symptoms of intestinal barrier damage and increased intestinal permeability caused by high fat, and the effect of XA-1416 was the best.

[0401] LCA (lithocholic acid) activates Sirtuins by binding to TULP3 protein, and then deacetylates and inhibits the key subunit V1E1 of lysosomal proton pump v-ATPase. This process activates AMPK through the lysosomal glucose sensing pathway, thereby simulating the metabolic and anti-aging benefits brought by caloric restriction.

[0402] As shown in FIG. 42, compared with the Model group, the level of LCA in the XA-1416 group was significantly improved, increasing by 295%. The results showed that XA-1416 could significantly improve the level of LCA and had the potential of anti-aging.

[0403] Experimental Example 12: Effect of saliva combined with Lactobacillus XA-1416 on the intestines and liver of a prophylactic obesity model mouse

[0404] Take 0.1 g of mouse cecal contents and mix thoroughly with 1.2 mL of 0.5% (w / v, g / 100 mL) aqueous phosphoric acid solution, then centrifuge to separate the supernatant. Next, the supernatant is filtered through a 0.22 micron filter, and then subjected to gas chromatography-mass spectrometry (GC-MS) detection (the analysis and measurement work is performed by a professional team of Shanghai Applied Protein Technology Co., Ltd.). The sample is mixed with formic acid and centrifuged to collect the supernatant. Then, ethyl acetate is added to extract the upper ethyl acetate phase. The ethyl acetate phase is mixed with 4-methylvaleric acid and filtered through an organic phase filter membrane to detect short-chain fatty acids (the short-chain fatty acid content is evaluated according to the relative peak area with the standard and 4-methylvaleric acid internal standard).

[0405] Mix the mouse colon tissue and PBS buffer at a ratio of 1:9 (W / V, 1 g:9 mL) and homogenize. The supernatant is collected by centrifugation at 4°C, 12,000 rpm for 10 min. The protein concentration in the supernatant is detected using a nanodrop oneC S2201957 (USA) spectrophotometer. The GLP-1 level, Leptin and PP level are determined using an enzyme-linked immunosorbent assay kit (Jiangsu Meilan Industrial Co., Ltd., China).

[0406] Weigh 25 mg of mouse ileal contents into a 2 mL thick centrifuge tube, add 2 small steel balls and 400 μL of pre-cooled (pre-cooled to 4°C) precipitant (acetonitrile:methanol=7:3, v / v) to each tube, and put it into a tissue grinder for grinding (grinding conditions are set to power 50 Hz, time 300 s) to obtain a grinding liquid sample. Remove the steel balls, and after the grinding liquid sample is placed in a -20°C refrigerator for 120 min, centrifuge at 25000g, 4°C for 15 min, and take the supernatant to a 1.5 mL EP tube. Take 20 μL of the sample and the standard curve, add 60 μL of pre-cooled 50% methanol, shake for 5 min, precipitate at -20°C for 4 h, centrifuge at 20000g, 4°C for 15 min, and take 20 μL of the supernatant to an EP tube. Next, the derivatization step is performed. Add 20 μL of 200 mM 3-NPH and 20 μL of 120 mM EDC-6% pyridine mixture to the EP tube containing 20 μL of the supernatant, respectively, shake and incubate at 25°C for 30 min, centrifuge, take the supernatant, and load it onto the machine. The porcine cholic acid in the pretreated sample is detected by high performance liquid chromatography-tandem mass spectrometry (LC-MS / MS for short).

[0407] Total RNA was extracted from mouse liver tissues using Trizol reagent (Beijing Pishow Biotechnology Co., Ltd., Beijing, China) and reverse transcribed into cDNA by High Capacity cDNA Reverse Transcription Kit (Beijing Pishow Biotechnology Co., Ltd., Beijing, China). Real-time PCR was performed using Hiper SYBR Premix reagent to detect the mRNA levels of AMPK, FXR, small heterodimer partner (SHP), sterol regulatory element binding protein 1c (SREBP-1c), and peroxisome proliferator-activated receptor alpha (PPAR-α) in liver tissues, with GAPDH as an internal reference for standardization. The primers used for PCR are shown in Table 21.

[0408] Short-chain fatty acids increase GLP-1, Leptin, and PP, etc. satiety signals by binding to specific receptors in the gut. As can be seen from FIGS. 43-47, the levels of short-chain fatty acids (acetic acid, propionic acid, butyric acid, valeric acid, and isovaleric acid) in the cecal contents of model mice decreased significantly compared with the normal diet group. This change may be related to the imbalance of the intestinal microbial community structure in the obese state, thereby affecting the production of short-chain fatty acids. However, after treatment with XA-1416, the levels of short-chain fatty acids such as acetic acid, propionic acid, butyric acid, valeric acid, and isovaleric acid increased significantly, especially the level of butyric acid increased by 40%, and the levels of valeric acid, propionic acid, and acetic acid increased by 84%, 43%, and 42%, respectively.

[0409] Glucagon-like peptide-1 (GLP-1) is a pro-insulin intestinal hormone secreted by intestinal L cells. GLP-1 can promote insulin secretion to reduce blood glucose levels, and can also slow gastric emptying, reduce appetite, and help control weight. As can be seen from FIG. 48, compared with mice fed with a normal diet, the GLP-1 level in the colon of model mice fed with a high-fat diet decreased by 51%, and after treatment with XA-1416, the GLP-1 level increased by 49% compared with the model group. As can be seen from the results of FIG. 48, XA-1416 has a significant restoring effect on the GLP-1 level.

[0410] Farnesoid X receptor (FXR) is a nuclear receptor protein mainly expressed in tissues such as liver and intestine, which plays a key role in regulating bile acid, lipid and glucose metabolism. Transmembrane receptor G protein-coupled bile acid receptor 5 (TGR5) is a G protein-coupled receptor, and activation of the receptor can induce the release of GLP-1 in the intestine. After primary bile acids are converted into secondary bile acids by intestinal microbiota in the intestine, the secretion of L cell GLP-1 is regulated by binding to TGR5 and nuclear receptor FXR. After FXR in the intestine is activated, the secretion of GLP-1 is reduced. Hyocholic acid (HCA) in bile acid is both an activator of TGR5 and an antagonist of FXR. As can be seen from FIG. 49, the level of HCA in mice treated with XA-1416 increased by 110%, which was significantly higher than that in the model group. The results of FIGS. 48-49 show that XA-1416 promotes the secretion of GLP-1 by increasing the level of HCA, activating TGR5 receptor while inhibiting FXR activity.

[0411] Based on the results of FIGS. 48-49, it is inferred that Lactobacillus salivarius XA-1416 can promote the secretion of glucagon-like peptide 1 in the intestine by increasing the level of hyocholic acid (HCA) in the intestine, activating transmembrane receptor G protein-coupled bile acid receptor 5 (TGR5) while inhibiting the activity of farnesoid X receptor (FXR), thereby effectively controlling blood glucose levels, and having great application prospects in the treatment of type 2 diabetes.

[0412] Pancreatic polypeptide (PP) is an endogenous polypeptide secreted by PP cells in endocrine pancreas, which can regulate pancreatic secretion, affect liver glycogen storage and gastrointestinal peristalsis and secretion, suppress appetite, and delay the increase of postprandial blood glucose and insulin. As can be seen from FIG. 50, the level of PP in the colon of the model group was also significantly reduced by 46% compared with the normal diet group, however, after treatment with XA-1416, the level of PP in the XA-1416 group was significantly increased by 35% compared with the model group, close to the normal level. As can be seen from the results of FIG. 50, XA-1416 also has a positive regulatory effect on the level of PP, which can effectively control blood glucose levels by promoting the secretion of PP in the intestine.

[0413] Leptin is a protein secreted by adipocytes, which is closely related to the regulation of food intake, energy metabolism, and fat storage in the body. As can be seen from FIG. 51, the level of Leptin in the colon of the model group was also significantly reduced by 5% compared with the normal diet group, however, after treatment with XA-1416, the level of Leptin in the XA-1416 group was significantly increased, and 100% restored to the normal level. As can be seen from the results of FIG. 51, XA-1416 also has a positive regulatory effect on the level of Leptin, which can effectively improve lipid metabolism by promoting the secretion of Leptin in the intestine.

[0414] Based on the results of FIGS. 43-48 and 50-51 and the results of FIGS. 52-56, it is inferred that Lactobacillus salivarius XA-1416 can effectively control blood glucose levels and improve lipid metabolism by increasing the levels of short-chain fatty acids (including acetic acid, propionic acid, butyric acid, valeric acid, and isovaleric acid, etc.) in the intestine, activating intestinal endocrine cells, thereby increasing the levels of glucagon-like peptide 1, pancreatic polypeptide, and leptin in the intestine, and thus has great application prospects in the treatment of type 2 diabetes and overweight and / or obesity.

[0415] Adenosine monophosphate-activated protein kinase (AMPK) is a key energy-sensing molecule that plays an important role in regulating cellular energy balance and promoting metabolic health. As can be seen from FIG. 52, the expression level of the AMPK gene in the liver of the model group was significantly lower than that of the normal diet group, whereas the expression level of the AMPK gene in the liver of the XA-1416 group was significantly higher than that of the model group after treatment with XA-1416. The results of FIG. 52 show that XA-1416 has a significant up-regulating effect on the expression of the AMPK gene in the liver, and can improve lipid metabolism and glucose metabolism in the liver by activating the AMPK signaling pathway.

[0416] After FXR is activated in the liver, the expression of SREBP-1c is down-regulated in a SHP-dependent manner, thereby inhibiting the generation of liver fat. FXR can also up-regulate PPAR-a to increase fatty acid oxidation metabolism. In addition, FXR reduces hepatic gluconeogenesis through SHP to improve glucose metabolism. As can be seen from FIG. 53, the expression level of the FXR gene in the liver of the model group was significantly lower than that of the normal diet group, whereas the expression level of FXR was significantly increased after treatment with XA-1416, indicating that XA-1416 can significantly increase the expression level of FXR. As can be seen from FIGS. 54-56, the expression level of the SHP gene in the liver of the model group was significantly lower than that of the normal diet group, the expression level of the SREBP-1c gene was significantly higher, and the expression level of the PPARa gene was significantly lower, indicating that the metabolic regulation of the liver of the model group was disordered, whereas the expression levels of the SHP, SREBP-1c, and PPARa genes in the liver were significantly restored after treatment with XA-1416, approaching those of the normal diet group. Based on the results of FIGS. 53-56, it can be seen that XA-1416 activates the FXR-SHP-SREBP-1c signaling pathway by increasing the expression level of FXR in the liver, effectively inhibits liver fat synthesis, and promotes fatty acid oxidation through the FXR-PPARa signaling pathway. In addition, XA-1416 also reduces hepatic gluconeogenesis through the FXR-SHP pathway, thereby improving glucose metabolism.

[0417] In combination with the results of FIGS. 53-56, it is inferred that Lactobacillus salivarius XA-1416 can effectively improve lipid metabolism and glucose metabolism in the liver by activating the AMPK signaling pathway and the FXR-SHP-SREBP-1c and FXR-SHP signaling pathways, and thus has great application prospects in the treatment of fatty liver disease.

[0418] As shown in FIG. 57, compared with the normal diet group, the expression level of IL-6 gene in the liver of the model group mice was significantly increased. However, after treatment with XA-1416, the expression level of IL-6 gene in the liver was significantly decreased, close to the level of the normal diet group. As shown in FIG. 58, compared with the normal diet group, the expression level of IL-10 gene in the liver of the model group mice was significantly decreased. However, after treatment with XA-1416, the expression level of IL-10 gene in the liver was significantly increased. As shown in FIG. 59, compared with the normal diet group, the expression level of TNF-a gene in the liver of the model group mice was significantly increased. However, after treatment with XA-1416, the expression level of TNF-a gene in the liver was significantly decreased. The results show that XA-1416 can effectively regulate the expression of inflammation-related genes in the liver of obese mice, and can reduce the inflammatory response.

[0419] Table 21 Specific primer sequences

[0420] Experimental Example 13: Effect of Lactobacillus salivarius XA-1416 on treatment of obese model mice

[0421] The eyeballs of the mice were taken out to obtain blood, which was left to stand at room temperature. After the blood sample was coagulated, it was centrifuged at 3000 rpm for 10 min to separate the serum. The serum sample was detected for TG, TC, HDL-C, LDL-C, ALT and AST according to the instructions of the kit (Zhongsheng Beikong Biotechnology Co., Ltd.), and the biochemical indicators were detected using Mindray BS-420 automatic biochemical analyzer.

[0422] The brown adipose tissue of the mice was taken out, and the tissues were weighed using a balance, and the weight of each tissue sample was recorded.

[0423] The colon tissue of the mice was mixed with PBS buffer at a ratio of 1:9 (W / V, 1 g:9 mL) to homogenate. The supernatant was collected by centrifugation at 4°C and 12,000 rpm for 10 min. The protein concentration in the supernatant was detected using a nanodrop oneC S2201957 (USA) spectrophotometer. The GLP-1 level was determined using an enzyme-linked immunosorbent assay kit (Jiangsu Meilan Industrial Co., Ltd., China).

[0424] As can be seen from FIGS. 60-65, compared with the ND group, the TG, TC, LDL-C, ALT and AST levels in the serum of the HFD group mice were significantly increased, and the HDL-C level was significantly decreased, indicating that the obesity model was successfully constructed, and the HFD group mice showed characteristics of dyslipidemia and liver function damage. Moreover, compared with the HFD group, the serum TG, TC, LDL-C and ALT levels of the Semaglutide group and the XA-1416 group mice were significantly reduced, while the HDL-C level was significantly increased. In addition, the XA-1416 group showed a significant effect in reducing the serum AST level, and was superior to the Semaglutide group. These results show that XA-1416 can effectively improve the blood lipid metabolism disorder of high-fat diet-induced obese model mice, significantly regulate the levels of TG, TC, LDL-C and HDL-C in serum, and restore blood lipid balance. At the same time, XA-1416 can also significantly reduce the levels of ALT and AST in the serum of obese mice, showing its significant protective effect on the liver.

[0425] Brown adipose tissue (BAT) is a special adipose tissue, which contains abundant mitochondria and a large number of small lipid droplets in the cells, and is mainly distributed in the interscapular region, axillary fossa and posterior neck, etc. BAT releases energy in the form of heat through mitochondrial respiration uncoupling mediated by uncoupling protein 1 (UCP1), thereby increasing energy consumption. As can be seen from FIG. 66, compared with the HFD group, the BAT / weight ratio of the Semaglutide group decreased significantly by 12%, while the BAT / weight ratio of the XA-1416 group increased significantly by 15%.

[0426] As can be seen from FIG. 67, compared with the HFD group, the colon GLP-1 level of the Semaglutide group increased significantly by 42%, while the colon GLP-1 level of the XA-1416 group increased more significantly by 49%, which was 7% higher than that of the Semaglutide group.

[0427] The results show that XA-1416 has a significant effect on improving the BAT / weight ratio and colon GLP-1 level, and the effect is superior to that of Semaglutide.

[0428] Experimental Example 14: Effect of saliva combined with Lactobacillus XA-1416 and Semaglutide on treatment of obese model mice

[0429] The epididymal fat tissue and pancreas tissue of the mice were taken and weighed using a balance, and the weight of each tissue sample was recorded.

[0430] The mouse pancreas tissue was fixed with 4% (w / v, g / 100 mL) paraformaldehyde. The fixed tissue was then embedded, and then a section was prepared using a paraffin microtome, and the thickness of the section was 3 pm. The tissue section was deparaffmized with xylene I and xylene II, dehydrated with gradient ethanol, and washed with tap water, followed by nuclear staining, washing, differentiation, and finally dehydration, transparency, and mounting. After mounting, the morphology of the tissue structure and possible pathological changes were observed under a light microscope.

[0431] As can be seen from FIG. 68(a), the body weight of the Semaglutide group was significantly reduced compared with the HFD group, however, the body weight of the Semaglutide+XA1416 group was reduced more significantly, indicating that the combined treatment is superior to Semaglutide alone in terms of weight loss effect. As can be seen from FIG. 68(b), the epididymal fat weight showed a similar trend. Compared with the HFD group, the epididymal fat weight of the Semaglutide group did not show a significant decrease, however, the epididymal fat weight of the Semaglutide+XA1416 group was significantly reduced, indicating that the combined treatment may also be more effective in reducing white adipose tissue. As can be seen from FIG. 68(c), compared with the HFD group, the food intake of the Semaglutide group was reduced by 9%, however, in the Semaglutide+XA1416 group, the food intake was reduced by 16%, indicating that the combined treatment showed a more significant effect in suppressing appetite.

[0432] The pancreas organ index is the ratio of pancreas weight (mg) to mouse body weight (g), which is used to evaluate the health status of the pancreas. As can be seen from FIG. 69, compared with the HFD group, the pancreas organ index was significantly increased when Semaglutide was used alone, indicating that Semaglutide caused damage to the pancreas. However, the pancreas organ index of the Semaglutide+XA1416 group was lower than that of the Semaglutide group, indicating that XA-1416 has the effect of alleviating pancreatic damage.

[0433] In addition, as can be seen from FIG. 70, the pancreas of the HFD group appears edematous, and the islet cells are sparsely distributed, but there is no serious damage to the tissue structure. However, the use of Semaglutide alone causes the pancreas to be significantly swollen, the cytoplasm to be slightly vacuolated, the cells to be loosely arranged and disordered, and the inflammatory cells to increase. However, when Semaglutide is used in combination with XA-1416, the damage to the pancreas is significantly improved, and the pancreas is only slightly swollen, and the cells are arranged in an orderly manner. This further indicates that XA-1416 helps to reduce the adverse effects of Semaglutide on the pancreas.

[0434] The human trial process is as follows:

[0435] The purpose of this study is to evaluate the effectiveness of the XA-1416 probiotic product in weight loss and fat reduction, as well as its potential impact on the volunteers' lives. Forty adults aged 18 to 65 years old with a BMI between 22 and 35 kg / m2 are recruited as volunteers. After voluntarily signing the informed consent form, the volunteers enter the screening period, provide personal information, and undergo BMI review. Volunteers who meet the inclusion criteria will be further reviewed for eligibility based on inclusion and exclusion criteria.

[0436] The inclusion criteria include: voluntarily participating and signing the informed consent form; age 18 to 65 years old, regardless of gender; BMI between 22 and 35 kg / m2; being able to take the probiotic daily for three months and cooperate with the completion of body composition testing; receiving dietary and lifestyle advice. The exclusion criteria include: oral antibiotics within one month before enrollment; having diabetes, cardiovascular disease, thyroid abnormalities, anemia, or digestive diseases; pregnant or lactating women; alcohol abuse or long-term drinking; using weight loss or fat reduction related health nutrition products or internal medicine for disease treatment (except for those who agree to stop); allergic to probiotics; participating in a weight loss program in the past month (except for those who have rebounded); and other conditions deemed unsuitable for participation by the research staff.

[0437] This project is a single-arm, single-center study, with a plan to enroll 40 volunteers. Volunteers will take 1 bag of XA-1416 probiotic products (each bag contains at least 20 billion CFU of live bacteria) daily for 90 days, and it is recommended to take it 1 hour after meals. Baseline data (without taking probiotics) will be collected on day -2, and BMI and body composition data (using ZHAIKER body composition tester) will be measured at 30 days (±5 days), 60 days (±7 days), and 90 days (±7 days) after taking, respectively. Volunteers need to complete a questionnaire every week within 90 days to report their feelings and health status. After completing the 90-day follow-up, all data are analyzed and summarized, and the project is completed.

[0438] After all volunteers complete the 1-month visit, a mid-term analysis is performed, including drawing the weight, body fat, and waist circumference change curve, comparing the changes in indicators before and after the intervention, and analyzing the change distribution under different grouping conditions. After completing the 3-month visit, a final analysis is performed, which is the same as the mid-term analysis, and an additional comparative analysis after 1 month, 2 months, and 3 months of intervention is added. The specific experimental design is shown in FIG. 71.

[0439] The digestive juice involved in the following experimental examples is as follows:

[0440] 0.25% trypsin digestive juice: first weigh 2.5 g of porcine-derived trypsin (purchased from Gibco Company), 0.2 g of EDTA is dissolved in PBS buffer (purchased from Solarbio, product number P1020), then adjust the pH to 7.4 with HCl, and finally add PBS buffer to 1 L to obtain 0.25% trypsin digestive juice.

[0441] The preparation method of the bacterial agent and culture supernatant involved in the following experimental examples is as follows:

[0442] Take the Lactobacillus salivarius bacterial liquid and inoculate it in the MRS liquid medium at an inoculation amount of 4% (v / v), and place it in a 37°C constant temperature incubator for static culture for 3 days to obtain a culture liquid; centrifuge the culture liquid at 8000 g for 10 min to obtain Lactobacillus salivarius culture supernatant and Lactobacillus salivarius cells; resuspend the Lactobacillus salivarius cells in a physiological saline after washing with physiological saline to obtain a Lactobacillus salivarius bacterial agent, and store the Lactobacillus salivarius bacterial agent at -80°C for later use.

[0443] Experimental Example 15: Effect of Lactobacillus salivarius XA-1416 on the body type of overweight and / or obese people

[0444] After the volunteers took Lactobacillus salivarius XA-1416 for 90 days, the body type indicators showed significant improvement, as follows:

[0445] As shown in Table 22 and FIGS. 72-73, the waist circumference of the volunteers significantly decreased month by month, with an average waist circumference reduction of 3.12 cm and a maximum reduction of 12 cm. Among the 33 volunteers, 85% (28 people) had a reduced waist circumference, of which 36.4% (12 people) had a waist circumference reduction of 4 cm or more.

[0446] As shown in Table 23 and FIGS. 74-75, after taking for 90 days, 64% (21 people) of the volunteers achieved a reduction in hip circumference, with an average reduction of 1.33 cm and a maximum reduction of 9 cm. Among them, 33.3% (11 people) had a hip circumference reduction of 2 cm or more, with a specific distribution of 18.2% of the volunteers having a reduction of 2-4 cm and 15.2% of the volunteers having a reduction of 4 cm or more.

[0447] As shown in Table 24 and FIGS. 76-77, the waist-to-hip ratio (WHR) of the volunteers significantly decreased month by month during the 3-month period of taking XA-1416. 79% (26 people) of the volunteers achieved a reduction in waist-to-hip ratio, with an average reduction of 0.019 and a maximum reduction of 0.07. Among them, 51.5% (17 people) of the volunteers achieved a waist-to-hip ratio reduction value of 0.02 or more.

[0448] The results show that saliva combined with Lactobacillus XA-1416 has a significant effect on improving the body shape of overweight and / or obese people. By reducing waist circumference, hip circumference and waist-to-hip ratio, XA-1416 not only helps volunteers to shape a healthier body shape, but also reduces the risk of metabolic diseases associated with overweight and / or obesity.

[0449] Table 22: Waist circumference (cm) records

[0450] Table 23: Hip circumference (cm) records

[0451] Table 24: Waist-to-hip ratio

[0452] Experimental Example 16: Effect of saliva combined with Lactobacillus XA-1416 on body fat of overweight and / or obese people

[0453] Body fat mass refers to the total weight of all adipose tissue in the human body, mainly including subcutaneous fat and visceral fat, and is an important indicator for assessing body composition and health risks. On the one hand, subcutaneous fat is adipose tissue located below the skin and distributed throughout the body (such as the abdomen, thighs, hips, etc.), which is directly stored between the skin and muscles and is the main form of fat storage in the human body. On the other hand, visceral fat is the fat located in the abdominal cavity and surrounding internal organs (such as the liver, pancreas, and intestinal tract), and is more closely related to the increase in various health risks than subcutaneous fat.

[0454] As shown in FIGS. 78-80, after the subjects took saliva combined with Lactobacillus XA-1416 for three consecutive months, the body fat composition showed systematic improvement: the subcutaneous fat decreased by 1.2% compared to the initial value, and the total body fat decreased by 1.3% (1.5 kg), and most notably, the visceral fat index showed a significant decrease (P<0.05) in the second month of intervention. In summary, XA-1416 not only has a systemic fat-reducing effect, but also exhibits a specific targeting effect on visceral fat, which has a higher metabolic risk.

[0455] Experimental Example 17: Effect of Lactobacillus salivarius XA-1416 on weight loss in overweight and / or obese population

[0456] Body weight is the most intuitive indicator of obesity, but due to differences in height and body composition among individuals, the simple body weight indicator is difficult to fully reflect the obesity situation. Body Mass Index (BMI, unit: kg / m2) is a commonly used clinical standard for assessing overall obesity, has good correlation with body fat ratio, and can effectively reflect the incidence risk of obesity-related diseases.

[0457] As can be seen from Figures 81-82, compared with the initial value, the BMI of the subjects decreased by 0.37 kg / m2 and the body weight decreased by 0.98 kg after taking XA-1416.

[0458] These results show that XA-1416 has a significant effect on weight loss and reduction of body fat, and further confirms its improvement effect on overweight and / or obesity in the population through the decrease of BMI.

[0459] Experimental Example 18: Effect of Lactobacillus salivarius XA-1416 on muscle in overweight and / or obese population

[0460] As mentioned in the literature "Clinical effectiveness of semaglutide on weight loss, body composition, and muscle strength in Chinese adults Eur Rev Med Pharmacol Sci. 2023 Oct; 27(20): 9908-9915.", GLP-1 receptor agonists such as semaglutide can effectively reduce body weight, but at the same time, it will cause a large amount of muscle loss. After 24 weeks of intervention, the amount of muscle loss can account for 4.8% of the total amount of body weight loss (p<0.001). The reduction of muscle tissue will cause a variety of health problems such as immune function decline, which is one of the main side effects of GLP-1 receptor agonist drugs.

[0461] As can be seen from Figures 83-85, after 3 months of human trial, the body fat of the subjects decreased significantly, however, the lean body mass, skeletal muscle mass and total muscle mass of the subjects remained unchanged.

[0462] The results show that XA-1416 can effectively protect muscle tissue and avoid muscle loss caused by weight loss while reducing fat, thereby maintaining the health status of the body while reducing body weight.

[0463] As can be seen from the results of the above open-label trial, saliva combined with Lactobacillus XA-1416 showed significant effects in reducing body fat, body weight and BMI, especially having a unique advantage in reducing waist circumference and visceral fat. In addition, XA-1416 can effectively protect muscle tissue during weight loss and avoid muscle loss caused by weight loss, providing a safe, effective and sustainable weight management solution for overweight and / or obese people.

[0464] Obviously, the above embodiments are only examples for the purpose of clarity, and are not limiting to the embodiments. Based on the above description, other different forms of changes or variations can also be made by those of ordinary skill in the art. All embodiments do not need to be exhausted here. The obvious changes or variations derived therefrom are still within the protection scope of the present application.

Claims

1. Use of Ligilactobacillus salivarius in the manufacture of a medicament, characterized in that, The medicament has any one of the following functions: (a) preventing and / or treating obesity; (b) preventing and / or treating obesity-related diseases; (c) regulating intestinal flora; The saliva combined with Lactobacillus is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 40307.

2. Use according to claim 1, wherein The obesity-related diseases include obesity-induced metabolic syndrome, obesity-induced cardiovascular diseases and / or obesity-induced intestinal flora imbalance.

3. Use according to claim 2, wherein the compound is ###0002### The obesity-induced metabolic syndrome includes non-alcoholic fatty liver; the obesity-induced cardiovascular diseases include hyperlipidemia and / or hyperglycemia.

4. Use according to any one of claims 1 to 3, characterized in that, The prevention and / or treatment of obesity includes inhibiting body weight gain, reducing white adipocyte area, inhibiting preadipocyte differentiation into adipocytes, reducing liver cell lipid accumulation, improving blood lipid levels, improving liver damage indicators, improving bile acid metabolism, increasing intestinal farnesoid X receptor antagonist levels and / or improving intestinal flora.

5. Use according to any one of claims 1 to 3, wherein The prevention and / or treatment of obesity-related diseases includes inhibiting body weight gain, reducing white adipocyte area, inhibiting preadipocyte differentiation into adipocytes, reducing liver cell lipid accumulation, improving blood lipid levels, improving liver damage indicators, improving bile acid metabolism, increasing intestinal farnesoid X receptor antagonist levels and / or improving intestinal flora.

6. Use according to claim 4 or 5, wherein the compound is ###0002### The improvement of blood lipid levels includes reducing serum total cholesterol and / or low-density lipoprotein cholesterol; The improvement of liver damage indicators includes reducing serum alanine aminotransferase levels; The improvement of bile acid metabolism includes promoting the conversion of intestinal conjugated bile acids to free bile acids; The increase of intestinal farnesoid X receptor antagonist levels includes increasing the levels of ursodeoxycholic acid, glyco-ursodeoxycholic acid and / or tauroursodeoxycholic acid in the intestine; The improvement of intestinal flora includes reducing the abundance of obesity-related bacteria in the intestine, increasing the abundance of beneficial bacteria in the intestine, improving the ratio of obesity-related bacteria in the intestine and / or improving the functional pathways of intestinal microbiota; the obesity-related bacteria refer to bacteria enriched in the intestine of obese individuals.

7. Use according to claim 6, wherein the compound is ###0002### The obesity-related bacteria include Lachnospiraceae, Acetatifactor and / or Lachnoclostridium; The beneficial bacteria include Lactobacillus and / or Bifidobacterium; The ratio of obesity-related bacteria includes Firmicutes / Bacteroidetes; The improvement of intestinal microbiota functional pathways includes promoting the function of energy metabolism and lipid metabolism related pathways in the intestine.

8. Use according to claim 7, wherein the compound is ###0002### The energy metabolism and lipid metabolism related pathways include pentose phosphate pathway, 4-deoxy-L-threo-hex-4-enopyranuronate degradation pathway and / or mevalonate pathway.

9. Use according to any one of claims 1 to 8, characterized in that, The obesity-related diseases are hyperlipidemia; the prevention and / or treatment of hyperlipidemia includes improving blood lipid levels, improving bile acid metabolism and / or increasing intestinal farnesoid X receptor antagonist levels; The obesity-related disease is non-alcoholic fatty liver disease; the prevention and / or treatment of non-alcoholic fatty liver disease includes reducing liver cell lipid accumulation, improving blood lipid levels, improving liver damage indicators, improving bile acid metabolism, and / or increasing intestinal farnesoid X receptor antagonist levels. The obesity-related disease is obesity-induced intestinal flora imbalance; the prevention and / or treatment of obesity-induced intestinal flora imbalance includes reducing the abundance of intestinal obesity-related bacteria, increasing the abundance of intestinal beneficial bacteria, improving the ratio of intestinal obesity-related bacteria, and / or improving intestinal microbiota functional pathways.

10. Use according to any one of claims 1 to 9, characterized in that, The obesity includes high-fat diet-induced obesity, chemical drug-induced obesity, and / or genetically modified obesity. The obesity-related disease includes high-fat diet-induced obesity-related disease, chemical drug-induced obesity-related disease, and / or genetically modified obesity.

11. Use of Ligilactobacillus salivarius in the preparation of a medicament for promoting the secretion of glucagon-like peptide-1, characterized in that, The saliva joint lactobacillus is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 40307.

12. The use according to claim 11, wherein the compound is ###00010### or a pharmaceutically acceptable salt thereof. The promotion of glucagon-like peptide-1 secretion includes the promotion of bile acid metabolism and / or short-chain fatty acid metabolism.

13. The use according to claim 12, wherein the compound is ###00010### or a pharmaceutically acceptable salt thereof. The promotion of glucagon-like peptide-1 secretion by regulating bile acid metabolism includes the promotion of swine cholalic acid levels, the inhibition of farnesoid X receptor activity while activating the transmembrane receptor G protein-coupled bile acid receptor 5, thereby promoting the secretion of glucagon-like peptide 1. The promotion of glucagon-like peptide-1 secretion by regulating short-chain fatty acid metabolism includes the activation of intestinal endocrine cells by promoting the levels of short-chain fatty acids, thereby promoting the secretion of glucagon-like peptide 1.

14. Use of Ligilactobacillus salivarius in the preparation of a medicament for the prevention and / or treatment of a disease associated with lipid metabolism and / or sugar metabolism, characterized in that, The saliva joint lactobacillus is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 40307.

15. The use according to claim 14, wherein the compound is ###0006### The disease related to lipid metabolism includes fatty liver disease and / or obesity; the disease related to sugar metabolism includes diabetes.

16. Use according to claim 14 or 15, wherein The prevention and / or treatment of diseases related to lipid metabolism and / or sugar metabolism includes increasing the level of glucagon-like peptide 1 in the intestine, increasing the level of pancreatic polypeptide in the intestine, increasing the level of leptin in the intestine, improving the lipid metabolism of the liver, and / or improving the sugar metabolism of the liver.

17. The use according to claim 16, wherein The increase in the level of glucagon-like peptide 1 in the intestine includes the increase in the level of glucagon-like peptide 1 in the intestine by regulating bile acid metabolism and / or short-chain fatty acid metabolism.

18. The use of claim 16, wherein, The improvement of the lipid metabolism of the liver includes the improvement of the lipid metabolism of the liver by activating the AMPK signaling pathway, the FXR-SHP-SREBP-1c signaling pathway, and / or the FXR-PPARα signaling pathway.

19. The use of claim 16, wherein, The improvement of the sugar metabolism of the liver includes the improvement of the sugar metabolism of the liver by the AMPK signaling pathway and / or the FXR-SHP signaling pathway.

20. A pharmaceutical composition comprising, The components of the pharmaceutical composition include Ligilactobacillus Salivarius; the Ligilactobacillus Salivarius is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 40307; the pharmaceutical composition has the functions shown in any one of the following: (a) promoting glucagon-like peptide-1 secretion; and / or, (b) preventing and / or treating diseases related to lipid metabolism and / or glucose metabolism.

21. The use of any one of claims 1-3, wherein the medicament has any one of the following functions of preventing and / or treating obesity: increasing BAT / body weight ratio and colonic GLP-1 level, improving body shape index, reducing visceral fat, reducing body fat, reducing body weight and BMI, avoiding muscle loss during weight loss, reducing inflammation level, anti-oxidation, anti-aging The improvement of the body shape index includes reducing the waist circumference, hip circumference and waist-hip ratio of the overweight / obese population; The reduction of the inflammation level includes reducing the IL-6 gene expression level and increasing the IL-10 gene expression level; The anti-oxidation includes reducing the T-AOC level and MDA level in serum; The anti-aging includes increasing the LCA level.

22. The use according to claim 4 or 5, wherein The Ligilactobacillus salivarius XA-1416 has high activity of Bile Salt Hydrolase (BSH).

23. The use of any one of claims 1-3, wherein the Ligilactobacillus salivarius XA-1416 is combined with a GLP-1 medicament, The GLP-1 medicament is Semaglutide.

24. The use of any one of claims 1-3, wherein the Ligilactobacillus salivarius XA-1416 reduces the increase of pancreatic organ index caused by the GLP-1 medicament.

25. A method for preventing and / or treating obesity and its related diseases, comprising administering Ligilactobacillus salivarius to an overweight / obese subject, the Ligilactobacillus salivarius is deposited with the China General Microbiological Culture Collection Center under the accession number CGMCC No. 40307.

26. The method of claim 25, wherein the obesity-related diseases include obesity-induced metabolic syndrome, obesity-induced cardiovascular disease and / or obesity-induced intestinal flora imbalance.

27. The method of claim 26, wherein the obesity-induced metabolic syndrome includes non-alcoholic fatty liver disease; The obesity-induced cardiovascular disease includes hyperlipidemia and / or hyperglycemia.

28. The method of claim 25, wherein the prevention and / or treatment of obesity includes inhibiting body weight gain, reducing white adipocyte area, inhibiting the differentiation of preadipocytes into adipocytes, reducing liver cell lipid accumulation, improving blood lipid level, improving liver damage index, improving bile acid metabolism, increasing intestinal farnesoid X receptor antagonist level and / or improving intestinal flora.

29. The method of claim 28, wherein the improvement of blood lipid level includes reducing serum total cholesterol and / or low-density lipoprotein cholesterol; The improvement of liver damage index includes reducing serum alanine aminotransferase level; The improvement of bile acid metabolism includes promoting the conversion of intestinal conjugated bile acids to free bile acids; The increasing intestinal farnesoid X receptor antagonist level includes increasing the level of ursodeoxycholic acid, glyco-ursodeoxycholic acid and / or tauroursodeoxycholic acid in the intestine; The improving intestinal flora includes reducing the abundance of obesity-related bacteria in the intestine, increasing the abundance of beneficial bacteria in the intestine, improving the ratio of obesity-related bacteria in the intestine and / or improving the intestinal microbiota functional pathway; the obesity-related bacteria refers to the bacteria enriched in the intestine of obese individuals.

30. The method of claim 29, wherein the obesity-related bacteria comprises Lachnospiraceae, Acetatifactor and / or Lachnoclostridium. The beneficial bacteria comprises Lactobacillus and / or Bifidobacterium; The ratio of obesity-related bacteria comprises Firmicutes / Bacteroidetes; The improving intestinal microbiota functional pathway comprises promoting the function of energy metabolism and lipid metabolism related pathways in the intestine.

31. The method of claim 30, wherein the energy metabolism and lipid metabolism related pathways comprise pentose phosphate pathway, 4-deoxy-L-threo-hex-4-enopyranuronate degradation pathway and / or mevalonate pathway.

32. The method of claim 25, wherein the obesity-related disease is hyperlipidemia; the preventing and / or treating hyperlipidemia comprises improving blood lipid levels, improving bile acid metabolism and / or increasing intestinal farnesoid X receptor antagonist levels; The obesity-related disease is non-alcoholic fatty liver; the preventing and / or treating non-alcoholic fatty liver comprises reducing liver cell lipid accumulation, improving blood lipid levels, improving liver damage indicators, improving bile acid metabolism and / or increasing intestinal farnesoid X receptor antagonist levels; The obesity-related disease is obesity-induced intestinal flora imbalance; the preventing and / or treating obesity-induced intestinal flora imbalance comprises reducing the abundance of obesity-related bacteria in the intestine, increasing the abundance of beneficial bacteria in the intestine, improving the ratio of obesity-related bacteria in the intestine and / or improving the intestinal microbiota functional pathway.

33. The method of claim 25, wherein the obesity comprises high-fat diet-induced obesity, chemical drug-induced obesity and / or genetically modified obesity; The obesity-related disease comprises high-fat diet-induced obesity-related disease, chemical drug-induced obesity-related disease and / or genetically modified obesity.

34. The method of claim 25, wherein the preventing and / or treating obesity comprises increasing the ratio of BAT to body weight and the level of colonic GLP-1, improving body shape indicators, reducing visceral fat, reducing body fat, reducing body weight and BMI, avoiding muscle loss during weight loss, reducing inflammation levels, antioxidant, anti-aging The improving body shape indicators comprises reducing waist circumference, hip circumference and waist-to-hip ratio in overweight / obese individuals; The reducing inflammation levels comprises reducing IL-6 gene expression levels, increasing IL-10 gene expression levels; The antioxidation includes reducing the level of T-AOC and MDA in serum; The anti-aging includes increasing the level of LCA.

35. A method for regulating intestinal flora, comprising administering Ligilactobacillus Salivarius to an overweight / obese subject, the Ligilactobacillus Salivarius is deposited with the China General Microbiological Culture Collection Center under the accession number CGMCC No. 40307.

36. The method of claim 25 or 35, wherein the viable cell number of Lactobacillus salivarius XA-1416 is not less than 1 x 10 6 CFU / mL or 1 x 10 6 CFU / g.

37. A method for preventing and / or treating diseases related to lipid metabolism and / or sugar metabolism, comprising administering Ligilactobacillus Salivarius to an overweight / obese subject, the Ligilactobacillus Salivarius is deposited with the China General Microbiological Culture Collection Center under the accession number CGMCC No. 40307.

38. The method of claim 37, wherein the diseases related to lipid metabolism include fatty liver disease and / or obesity; the diseases related to sugar metabolism include diabetes.

39. The method of claim 38, wherein the preventing and / or treating diseases related to lipid metabolism and / or sugar metabolism comprises increasing the level of glucagon-like peptide 1 in the intestine, increasing the level of pancreatic polypeptide in the intestine, increasing the level of leptin in the intestine, improving lipid metabolism in the liver, and / or improving sugar metabolism in the liver.

40. The method of claim 39, wherein the increasing the level of glucagon-like peptide 1 in the intestine comprises increasing the level of glucagon-like peptide 1 in the intestine by regulating bile acid metabolism and / or short-chain fatty acid metabolism.

41. The method of claim 39, wherein the improving lipid metabolism in the liver comprises improving lipid metabolism in the liver by activating the AMPK signaling pathway, the FXR-SHP-SREBP-1c signaling pathway, and / or the FXR-PPARa signaling pathway.

42. The method of claim 39, wherein the improving sugar metabolism in the liver comprises improving sugar metabolism in the liver by the AMPK signaling pathway and / or the FXR-SHP signaling pathway.

43. The method of claim 37, wherein the viable cell count of Lactobacillus salivarius XA-1416 is not less than 1 x 10 6 CFU / mL or 1 x 10 6 CFU / g.

44. Use of Ligilactobacillus salivarius in the manufacture of a medicament, characterized in that, The drug has any one of the following functions: (a) preventing and / or treating overweight; (b) preventing and / or treating overweight-related diseases; (c) inhibiting weight gain.

45. The use of any one of claims 1-3, wherein the Ligilactobacillus Salivarius XA-1416 is used in combination with a GLP-1 drug, and the GLP-1 drug is Semaglutide.

46. The use of any one of claims 1-3, wherein the Ligilactobacillus Salivarius XA-1416 helps alleviate the adverse effects of GLP-1 drug administration on the pancreas. The adverse effects are decreased pancreatic organ index and pancreatic tissue damage.

47. The method of claim 25, wherein the obesity-related diseases include obesity and / or overweight-induced metabolic syndrome, obesity and / or overweight-induced cardiovascular disease, and obesity and / or overweight-induced intestinal flora imbalance.

Images