A postbiotic and its use in modulating the direction of lipid metabolism

Abstract

The present application relates to a probiotic and its application in regulating the direction of lipid metabolism. Specifically, the present application relates to the use of the probiotic in the preparation of a medicine or food. The present application also relates to a method for regulating the body weight of a subject, and a method for inhibiting or reducing the absorption of lipids in the gastrointestinal tract of a subject.

Description

Technical Field

[0001] This application relates to an epigenetic and its application in regulating lipid metabolism. Specifically, this application relates to the use of the epigenetic in the preparation of pharmaceuticals. This application also relates to a method for regulating the weight of a subject, and a method for inhibiting or reducing lipid absorption in the gastrointestinal tract of a subject. Background Technology

[0002] In modern society, improved living standards, high-calorie diets, and sedentary lifestyles have led to a gradual increase in obesity rates and a series of metabolic abnormalities. For example, excessive lipid deposition can cause hyperlipidemia and non-alcoholic fatty liver disease. Furthermore, fat cell dysfunction can cause systemic inflammation and arteriosclerosis, resulting in hypertension and cardiovascular diseases. With the increasing number of overweight and obese individuals, the incidence of these diseases is rising rapidly, becoming a major health concern for urban populations today, and the obese and overweight population is trending towards younger ages.

[0003] Currently, the treatment of obesity, overweight, or lipid metabolism disorders mainly relies on lifestyle interventions, including a low-saturated fatty acid diet and moderate-to-vigorous physical activity. In addition, medication may be used, such as the lipase inhibitor orlistat; or medications for treating hypertension or dyslipidemia, as well as cholesterol-lowering drugs. These medications have some effect in treating obesity or lipid metabolism disorders, but long-term use can cause serious side effects on liver and kidney function.

[0004] Studies have found that probiotics play an important role in human physiological metabolism by producing metabolites and regulating gut microbiota. According to the definition of probiotics given by British scientist Roy Fuller in 1989: a live microbial supplement that, after ingestion, exerts beneficial effects on the host by improving the balance of the gut microbiota. The China Nutrition and Health Food Association has released the group standard T / CNHFA 006-2022, "Grading Specification for Live Bacteria Rate in Probiotic Foods," which grades the live bacteria rate of probiotic foods, emphasizing that ensuring a sufficient number of live probiotics reach the intestines is one of the fundamental conditions for probiotics to exert their efficacy.

[0005] Recent scientific research has shown that certain inactivated probiotics also possess unique biological activities, termed "metasecobiotics." The International Association for the Study of Probiotics and Prebiotics defines "metasecobiotics" as: preparations of non-living microorganisms and / or their related components that provide health benefits to the host. The efficacy of metabiotics derives from the microbial cells themselves and their growth metabolites. Metabiotic preparations are generally considered to include: inactivated or dead microbial cells; macromolecules secreted by microorganisms or bound to the cell surface, such as proteins, lipids, and carbohydrates; microbial metabolites such as short-chain fatty acids (SCFAs) and organic acids; and cell wall components such as lipoteichoic acid and peptidoglycan. As scientific research continues, the definition and scope of metabiotics will be further refined. Compared to probiotics, inactivated probiotics have natural advantages for commercial production and product application.

[0006] Therefore, developing an inactivated probiotic strain that can regulate or improve abnormal lipid metabolism has high market value and broad applications. Summary of the Invention

[0007] The patent applicant initially screened a probiotic strain—Bifidobacterium breve 207-1, with the microbial preservation number GDMCC No. 60962. During subsequent research, it was unexpectedly discovered that the metabiotic prepared from this strain after fermentation and inactivation exhibits outstanding efficacy in promoting lipolysis and / or inhibiting fat absorption. Therefore, the metabiotic of this application and compositions containing it have great potential in the preparation of remedies for diseases and / or symptoms related to fat accumulation.

[0008] Therefore, in a first aspect, this application provides the use of an epigenetic, or a composition containing said epigenetic, in the preparation of a medicament for the prevention and / or improvement of diseases and / or symptoms related to fat gain in a subject;

[0009] The metabiotic is prepared by fermenting and then inactivating Bifidobacterium breve, which is deposited at the Guangdong Provincial Center for Microbial Culture Collection with accession number GDMCC No. 60962.

[0010] Preparation of post-biotics

[0011] The postgenetic agent of the present invention can be prepared using various methods known in the art.

[0012] In some embodiments, the metabiotic is prepared by inactivating Bifidobacterium breve. In some embodiments, the metabiotic comprises Bifidobacterium breve in the form of dead bacteria.

[0013] In some embodiments, the metabiotic is prepared by fermenting and then inactivating Bifidobacterium breve. In some embodiments, the metabiotic comprises Bifidobacterium breve in dead form, as well as primary and / or secondary metabolites generated during in vitro fermentation.

[0014] In this article, metageners may include many different components.

[0015] In some embodiments, the metabiotic includes cell components of *Bifidobacterium breve*. In some embodiments, the metabiotic includes cell lysates of *Bifidobacterium breve* cells. In some embodiments, the metabiotic includes peptidoglycan, lipoteichoic acid, cell wall peptides, cell wall polysaccharides, fimbriae, etc.

[0016] In some embodiments, metabiotics include primary and / or secondary metabolites of Bifidobacterium breve. In some embodiments, metabiotics include short-chain fatty acids (SCFAs, such as acetic acid, propionic acid, and butyric acid), extracellular polysaccharides, functional proteins, vitamins (such as biotin, cobalamin, folic acid, niacin, pantothenic acid, pyridoxine, riboflavin, and thiamine), etc.

[0017] In such embodiments, those skilled in the art can choose appropriate methods to prepare various inactivated formulations of the Bifidobacterium breve contained in the metabiotic. Therefore, the dosage forms of the metabiotic include, but are not limited to, pills, powders, capsules, tablets (e.g., effervescent tablets), film-coated tablets, orally soluble granules, and liquids.

[0018] In some implementations, the metagener is a powder.

[0019] In some embodiments, the metabiotic is a bacterial powder.

[0020] In some embodiments, the bacterial powder is prepared by culturing the Bifidobacterium breve, inactivating it, collecting the precipitate, and drying it.

[0021] In some embodiments, the bacterial powder is prepared by fermenting and then inactivating the Bifidobacterium breve, collecting the precipitate by centrifugation, and then freeze-drying it under vacuum.

[0022] In some implementations, the Bifidobacterium breve is fermented using MRS medium.

[0023] In some embodiments, the *Bifidobacterium breve* is heat-inactivated at 70–95°C.

[0024] In some embodiments, the drug is used to prevent and / or improve diseases and / or symptoms caused by fat accumulation in a subject.

[0025] In some implementations, the disease and / or symptoms are selected from weight gain, obesity, fatty liver, fat accumulation (e.g., visceral fat accumulation, subcutaneous fat accumulation), lipid metabolism disorders, or any combination thereof.

[0026] In some embodiments, the subcutaneous fat deposits are selected from abdominal fat deposits, arm fat deposits, leg fat deposits, or any combination thereof.

[0027] In some embodiments, the visceral fat accumulation is selected from periintestinal fat accumulation, perirenal fat accumulation, perigonadal fat accumulation, or any combination thereof.

[0028] In some embodiments, the disease caused by the lipid metabolism abnormality is selected from: hyperlipidemia, non-alcoholic fatty liver disease, hypertension, cardiovascular disease, or any combination thereof.

[0029] In some implementations, the obese subject has a BMI greater than 23.9 kg / m² (e.g., BMI greater than 25 kg / m², greater than 26 kg / m², greater than 27 kg / m², greater than 28 kg / m², greater than 29 kg / m², greater than 30 kg / m²).

[0030] In some embodiments, the drug can promote lipolysis and / or inhibit fat absorption.

[0031] In some implementations, the drug is able to maintain the subject's weight and / or BMI.

[0032] In some implementations, the drug is able to reduce the subject's weight and / or BMI.

[0033] In some embodiments, the drug enables the subject to have a healthy BMI (e.g., 18.5-23.9 kg / m²).

[0034] In some embodiments, the drug is administered to subjects with a healthy BMI to maintain the subject's weight and / or BMI. In some embodiments, the drug is administered to subjects with an overweight BMI to reduce the subject's weight and / or BMI, or to bring the subject's weight and / or BMI toward a healthy weight and / or BMI (e.g., 18.5-23.9 kg / m²).

[0035] In some implementations, administering the drug to a subject can increase the subject's feeling of fullness.

[0036] In some implementations, administering the drug to a subject can reduce the subject's food intake.

[0037] In some embodiments, the drug may also contain additional active ingredients (e.g., compounds).

[0038] In some embodiments, the additional active ingredient can promote lipolysis and / or inhibit fat absorption; for example, L-carnitine.

[0039] In some implementations, the additional active ingredient can accelerate metabolism; for example, tea polyphenols, caffeine.

[0040] In some embodiments, the additional active ingredient is a lipase inhibitor; for example, orlistat.

[0041] In this document, the term "medicine" encompasses both medicines for human use and medicines for animal use (i.e., veterinary applications). In some embodiments, the medicine is used for human use.

[0042] In some embodiments, the pharmaceutical composition comprises a formulation of an epigenetic agent.

[0043] In some embodiments, the pharmaceutical composition comprises a pharmaceutically acceptable carrier.

[0044] In some embodiments, the pharmaceutical composition is formulated for oral administration.

[0045] In some embodiments, the drug is a drug that targets gastrointestinal release or a drug that is released in a controlled manner in the gastrointestinal tract.

[0046] In some embodiments, the drug is in the form of pills, powders, capsules, tablets (e.g., effervescent tablets), film-coated tablets, orally soluble granules, liquids, suppositories, or enemas.

[0047] In some embodiments, the subject is a mammal. In some embodiments, the mammal is selected from rats, pigs, rabbits, monkeys, sheep, and humans.

[0048] In some embodiments, the amount of metabiotic added to the drug is 0.001g-0.1g.

[0049] In some embodiments, the amount of metabiotic added to the drug is 0.001-0.005g, 0.005-0.01g, 0.01-0.05g, or 0.05-0.1g.

[0050] It is understood that those skilled in the art are capable of administering an effective amount of postbiotic to a subject based on the subject's specific circumstances.

[0051] In some embodiments, since the metabiotic contains inactivated Bifidobacterium breve, the number of Bifidobacterium breve cells is used as the unit of measurement for Bifidobacterium breve. In some embodiments, the number of Bifidobacterium breve cells in the metabiotic is 10-1. 8 -10 14 / g (e.g., 10) 8 -10 10 / g, 10 10 -10 12 / g, 10 12 -10 14 / g).

[0052] Therefore, when the amount of metabiotic added to the drug is 0.001g, the number of metabiotic cells is 10. 5 -10 11 When the amount of metabiotic added to the drug is 0.1g, the number of metabiotic cells is 10. 7 -10 13 .

[0053] In some embodiments, the composition comprises the postbiotic and microorganisms selected from bacteria, fungi, or any combination thereof.

[0054] In some implementations, the microorganism is a probiotic.

[0055] In some implementations, the microorganism is yeast.

[0056] In some embodiments, the yeast is selected from Saccharomyces cerevisiae, Saccharomyces boulardii, Kluyveromyces marxianus, or any combination thereof.

[0057] In some embodiments, the bacteria are selected from the genera *Lactobacillus*, *Bifidobacterium*, *Bacillus*, *Propionibacterium*, *Streptococcus*, *Lactococcus*, *Pediococcus*, *Enterococcus*, *Staphylococcus*, or any combination thereof.

[0058] In some embodiments, the bacteria of the genus *Lactobacillus* are selected from: *Lactobacillus paracasei*, *Lactobacillus acidophilus*, *Lactobacillus brevis*, *Lactobacillus jensenii*, *Lactobacillus iners*, *Lactobacillus casei*, *Lactobacillus crispatus*, *Lactobacillus curvatus*, *Lactobacillus delbrueckii*, *Lactobacillus fermentum*, *Lactobacillus gasseri*, *Lactobacillus helveticus*, *Lactobacillus johnsonii*, *Lactobacillus plantarum*, and *Lactobacillus reuteri*. Lactobacillus reuteri, Lactobacillus rhamnosus, Lactobacillus sakei, Lactobacillus salivarius, or any combination thereof.

[0059] In some embodiments, the bacteria of the genus Bifidobacterium are selected from: Bifidobacterium animalis, Bifidobacterium bifidum, Bifidobacterium breve, Bifidobacterium infantis, Bifidobacterium longum, Bifidobacterium adolescentis, or any combination thereof.

[0060] In some embodiments, the bacteria of the genus Bacillus are selected from Bacillus subtilis, Bacillus coagulans, or any combination thereof.

[0061] In some embodiments, the bacteria of the genus Propionibacterium are selected from: Propionibacterium shermanii, Propionibacterium freudenreichii, Propionibacterium acidipropionici, or any combination thereof.

[0062] In some embodiments, the Streptococcus bacteria are selected from Streptococcus thermophilus, Streptococcus salivarius, or any combination thereof.

[0063] In some embodiments, the bacteria of the genus *Lactococcus* is *Lactococcus lactis*.

[0064] In some embodiments, the Enterococcus species are selected from Enterococcus faecalis, Enterococcus faecium, Enterococcus mundtii, or any combination thereof.

[0065] In a second aspect, this application provides a method for regulating the weight of a subject, the method comprising: administering an effective amount of postbiotic to the subject, wherein the postbiotic is prepared by fermenting and then inactivating Bifidobacterium breve, the Bifidobacterium breve being deposited at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC No. 60962.

[0066] In some embodiments, the effective amount of postgenetic agent is 0.001g-0.1g; for example, 0.001-0.005g, 0.005-0.01g, 0.01-0.05g, 0.05-0.1g.

[0067] In some implementations, the obese subject has a BMI greater than 23.9 kg / m².

[0068] In some implementations, the method is able to maintain the subject's weight.

[0069] In some implementations, the method is able to reduce the subject's weight.

[0070] In some implementations, the method enables the subject to have a healthy BMI (e.g., 18.5-23.9 kg / m²).

[0071] In some embodiments, the subject is a mammal. In some embodiments, the mammal is selected from rats, pigs, rabbits, monkeys, sheep, and humans.

[0072] In some implementations, the method is a method for adjusting the weight of a subject for non-therapeutic purposes.

[0073] In such implementations, the subject does not have or has been diagnosed with any disease (e.g., obesity).

[0074] In such implementations, the subject has a healthy BMI.

[0075] In such implementations, the subject gains weight due to an unhealthy diet (e.g., a high-fat diet, a high-sugar diet, a high-cholesterol diet) or environmental factors.

[0076] In such embodiments, an effective amount of the post-gene or a composition containing the post-gene is given to the subject to maintain the subject's weight and / or BMI, or to reduce the subject's weight and / or BMI.

[0077] In such implementations, the frequency and manner of administering the postbiotic or a composition containing the postbiotic to the subject can be adjusted based on the subject's characteristics (e.g., age, sex, race, weight, height, BMI, body fat percentage, and / or medical history).

[0078] On the other hand, this application provides the use of an epigenetic or a composition containing said epigenetic in the preparation of a medicament for regulating the weight of a subject; wherein the epigenetic is prepared by fermentation and inactivation of Bifidobacterium breve, which is deposited at the Guangdong Provincial Center for Microbial Culture Collection with accession number GDMCC No. 60962.

[0079] In some embodiments, the amount of the post-genetic agent added is 0.001g-0.1g; for example, 0.001-0.005g, 0.005-0.01g, 0.01-0.05g, 0.05-0.1g.

[0080] In some embodiments, the subject is a mammal. In some embodiments, the mammal is selected from rats, pigs, rabbits, monkeys, sheep, and humans.

[0081] In some embodiments, the Bifidobacterium breve is present in the metabiotic as a dead bacterium.

[0082] In such embodiments, those skilled in the art can choose suitable methods to prepare various inactivated formulations of Bifidobacterium breve. Inactivated formulations include, but are not limited to, pills, powders, capsules, tablets (e.g., effervescent tablets), film-coated tablets, orally soluble granules, and liquids.

[0083] In some embodiments, the Bifidobacterium breve is a powder.

[0084] In some embodiments, the Bifidobacterium breve is a bacterial powder.

[0085] In some embodiments, the bacterial powder is prepared by culturing the Bifidobacterium breve, inactivating it, collecting the precipitate, and drying it.

[0086] In some embodiments, the bacterial powder is prepared by fermenting and then inactivating the Bifidobacterium breve, collecting the precipitate by centrifugation, and then freeze-drying it under vacuum.

[0087] In some implementations, the Bifidobacterium breve is fermented using MRS medium.

[0088] In some embodiments, the *Bifidobacterium breve* is heat-inactivated at 70–95°C.

[0089] In a third aspect, this application provides a method for inhibiting or reducing lipid absorption in the gastrointestinal tract of a subject, the method comprising: administering an effective amount of an epigenetic to the subject, wherein the epigenetic is prepared by fermenting and then inactivating Bifidobacterium breve, the Bifidobacterium breve being deposited at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC No. 60962.

[0090] In some embodiments, the effective amount of postgenetic agent is 0.001g-0.1g; for example, 0.001-0.005g, 0.005-0.01g, 0.01-0.05g, 0.05-0.1g.

[0091] In some implementations, the method is able to maintain the subject's weight.

[0092] In some implementations, the method is able to reduce the subject's weight.

[0093] In some implementations, the method is able to maintain the subject's BMI (e.g., 18.5-23.9 kg / m²).

[0094] In some embodiments, the subject is a mammal. In some embodiments, the mammal is selected from rats, pigs, rabbits, monkeys, sheep, and humans.

[0095] In some implementations, the method is a method for inhibiting or reducing lipid absorption in the gastrointestinal tract of a subject for non-therapeutic purposes.

[0096] In such implementations, the subject does not have or has been diagnosed with any disease (e.g., obesity).

[0097] In such implementations, the subject has a healthy BMI.

[0098] In such implementations, the subject gains weight due to an unhealthy diet (e.g., a high-fat diet, a high-sugar diet, a high-cholesterol diet) or environmental factors.

[0099] In such embodiments, an effective amount of the post-biotic composition comprising the post-biotic is given to a subject to maintain the subject's weight and / or BMI, or to reduce the subject's weight and / or BMI.

[0100] In such implementations, the frequency and manner of administering the postbiotic or a composition containing the postbiotic to the subject can be adjusted based on the subject's characteristics (e.g., age, sex, race, weight, height, BMI, body fat percentage, and / or medical history).

[0101] On the other hand, this application provides the use of an epigenetic or a composition containing said epigenetic in the preparation of a pharmaceutical composition for inhibiting or reducing lipid absorption in the gastrointestinal tract of a subject; wherein the epigenetic is prepared by fermentation and inactivation of Bifidobacterium breve, which is deposited at the Guangdong Provincial Center for Microbial Culture Collection with accession number GDMCC No. 60962.

[0102] In some embodiments, the amount of the post-biotic added to the drug is 0.001g-0.1g; for example, 0.001-0.005g, 0.005-0.01g, 0.01-0.05g, 0.05-0.1g.

[0103] In some embodiments, the subject is a mammal. In some embodiments, the mammal is selected from rats, pigs, rabbits, monkeys, sheep, and humans.

[0104] In some embodiments, the Bifidobacterium breve is present in the metabiotic as a dead bacterium.

[0105] In such embodiments, those skilled in the art can choose suitable methods to prepare various inactivated formulations of Bifidobacterium breve. Inactivated formulations include, but are not limited to, pills, powders, capsules, tablets (e.g., effervescent tablets), film-coated tablets, orally soluble granules, and liquids.

[0106] In some embodiments, the Bifidobacterium breve is a powder.

[0107] In some embodiments, the Bifidobacterium breve is a bacterial powder.

[0108] In some embodiments, the bacterial powder is prepared by culturing the Bifidobacterium breve, inactivating it, collecting the precipitate, and drying it.

[0109] In some embodiments, the bacterial powder is prepared by fermenting and then inactivating the Bifidobacterium breve, collecting the precipitate by centrifugation, and then freeze-drying it under vacuum.

[0110] In some implementations, the Bifidobacterium breve is fermented using MRS medium.

[0111] In some embodiments, the *Bifidobacterium breve* is heat-inactivated at 70–95°C.

[0112] Terminology Definition

[0113] In this invention, unless otherwise stated, the scientific and technical terms used herein have the meanings commonly understood by those skilled in the art. Furthermore, to better understand this invention, definitions and explanations of relevant terms are provided below.

[0114] As used herein, the term "lipid metabolism" is a biochemical reaction, specifically referring to the process by which lipids in an organism are synthesized, broken down, digested, absorbed, and transported under the action of various related enzymes. The main lipids in the blood include cholesterol, triglycerides (TAG), phospholipids (PL), and free fatty acids. In some embodiments, lipid metabolism processes fats into substances needed by the body to ensure the operation of normal physiological functions.

[0115] As used in this article, the term "dyslipidemia" refers to abnormalities in the synthesis, breakdown, digestion, absorption, and transport of lipids in the body, resulting in an excess or deficiency of lipids in tissues. Long-term high-cholesterol, high-saturated-fat, and high-calorie diets, genetic factors, apolipoprotein abnormalities, mental exertion, lack of exercise, and stress can all lead to dyslipidemia. Dyslipidemia may result in hyperlipidemia, non-alcoholic fatty liver disease, hypertension, and cardiovascular disease.

[0116] As used herein, the term "pharmaceuticalally acceptable carrier" means a carrier that is pharmacologically and / or physiologically compatible with the subject and the active ingredient, which is well known in the art (see, for example, Remington's Pharmaceutical Sciences. Edited by Gennaro AR, 19th ed. Pennsylvania: Mack Publishing Company, 1995), and includes, but is not limited to: pH adjusters, surfactants, adjuvants, and ionic strength enhancers. For example, pH adjusters include, but are not limited to, phosphate buffers; surfactants include, but are not limited to, cationic, anionic, or nonionic surfactants, such as Tween-80; and ionic strength enhancers include, but are not limited to, sodium chloride.

[0117] As used herein, the term "dietary supplement" refers to an edible product that provides a beneficial effect to a consumer (e.g., nutritional, preventative, therapeutic, or other beneficial effect). In this document, dietary supplements encompass products such as health products, nutritional supplements, and tonics.

[0118] As used herein, the term "drug" encompasses drugs used in human medicine and veterinary medicine for both human and animal use, as well as drugs intended for inclusion in animal feed (e.g., livestock feed and / or pet food). Furthermore, as used herein, the term "drug" refers to any substance that provides therapeutic, preventative, and / or beneficial effects. The term "drug" as used herein is not necessarily limited to substances requiring marketing approval, but includes substances that can be used in cosmetics, health products, foods (including, for example, animal feed and beverages), probiotic cultures, and dietary supplements.

[0119] Beneficial effects of the invention

[0120] The patent applicant initially screened a probiotic strain—Bifidobacterium breve 207-1, with the microbial preservation number GDMCC No. 60962. During subsequent research, it was unexpectedly discovered that this strain, after inactivation, exhibits outstanding efficacy in promoting lipolysis and / or inhibiting fat absorption. Therefore, the strain and compositions containing it in this application have great potential in the preparation of medicines for diseases and / or symptoms related to fat accumulation, such as those for weight gain, obesity, fatty liver, and fat accumulation.

[0121] The embodiments of the present invention will now be described in detail with reference to the accompanying drawings and examples. However, those skilled in the art will understand that the following drawings and examples are for illustrative purposes only and are not intended to limit the scope of the invention. Various objects and advantages of the present invention will become apparent to those skilled in the art from the following detailed description of the drawings and preferred embodiments. Attached Figure Description

[0122] Figure 1 The fluorescence intensity diagrams showing the effects of different sample treatments on zebrafish to promote lipolysis are shown.

[0123] Figure 2 The study shows the staining of blood vessels in the intestines and tail of zebrafish after different sample treatments.

[0124] Figure 3 The graphs showing the changes in body weight of mice in different treatment groups at different time points are shown. Compared with the model group, #p < 0.05.

[0125] Figure 4 The total weight gain of mice in different treatment groups before and after treatment is shown. Compared with the model group, #p < 0.05.

[0126] Figure 5 HE staining results of liver tissue from mice in different treatment groups are shown.

[0127] Instructions on the Preservation of Biological Materials

[0128] Bifidobacterium breve 207-1 has been deposited at the Guangdong Microbial Culture Collection Center (GDMCC), located on the 5th floor of Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC No. 60962, and the deposit date is January 15, 2020. Detailed Implementation

[0129] The invention will now be described with reference to the following embodiments, which are intended to illustrate the invention (and not limit it).

[0130] Unless otherwise specified, the experiments and methods described in the embodiments are performed in accordance with conventional methods well known in the art and described in various references. For example, conventional techniques such as immunology, biochemistry, chemistry, molecular biology, microbiology, cell biology, genomics, and recombinant DNA used in this invention can be found in Sambrook, Fritsch, and Maniatis, *Molecular Cloning: A Laboratory Manual*, 2nd edition (1989); *Current Protocols in Molecular Biology* (edited by FM. Ausubel et al., (1987)); the *Methods in Enzymology* series (academic publishing company): *PCR 2: A PRACTICAL APPROACH* (edited by MJ. MacPherson, BD. Hames, and GR. Taylor, (1995)); and *Animal Cell Culture*. CELLCULTURE (edited by R.R. Freshney (1987)).

[0131] Furthermore, unless specific conditions are specified in the examples, conventional conditions or conditions recommended by the manufacturer should be followed. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products. Those skilled in the art will understand that the examples are described by way of illustration and are not intended to limit the scope of protection claimed by the invention. All disclosures and other references mentioned herein are incorporated herein by reference in their entirety.

[0132] Example 1. Test strain

[0133] Preparation method of post-genetic agents

[0134] Live *Bifidobacterium breve* 207-1 bacteria were fermented on MRS medium at 35-37℃ for 24-72 hours, followed by heat inactivation at 70-95℃. The inactivated bacteria were centrifuged to obtain a precipitate, which was then freeze-dried under vacuum. The dried bacteria were sieved to obtain the final metabiotic. The metabiotic passed various tests, including sensory requirements, net content, total lactic acid bacteria count, bacterial count, and coliform count, meeting the requirements of the State Administration for Quality Supervision, Inspection and Quarantine Order No. 75, "Regulations on the Supervision and Management of Quantitatively Packaged Commodities." The test showed that the number of *Bifidobacterium breve* 207-1 bacteria in the metabiotic was 3 × 10⁻⁶.11 / g.

[0135] Source of experimental strains

[0136] The *Bifidobacterium breve* strains used in this experiment were all derived from the proprietary strain library of By-Health. These strains were isolated from stool samples of normal full-term newborns born at West China Hospital of Obstetrics and Gynecology, Sichuan University. Specifically, fresh stool samples were collected from infants aged 1-4 months using sterile collection tubes. Immediately after sampling, the samples were temporarily stored at 4°C and then transported to the laboratory by sampling personnel for dilution and culture. If immediate operation was not possible, the samples were anaerobically stored at 4°C and cultured on the same day. Subsequently, single strains were isolated and purified using the plate method. The specific species of the isolated strains were identified using bioMérieux's API 50 CH and 16S rDNA sequencing, numbered, and deposited in By-Health's proprietary strain library. Among them, *Bifidobacterium breve* 207-1 was previously deposited and patented due to its acid and bile salt resistance characteristics.

[0137] Example 2. Evaluation of the efficacy of metabiotics in promoting lipolysis in zebrafish

[0138] 2.1 Laboratory Animals

[0139] The melanin allele mutant translucent Albino strain zebrafish were raised in aquarium water at 28℃ (water quality: 200 mg of instant sea salt added per 1 L of reverse osmosis water, conductivity 450~550 μS / cm; pH 6.5~8.5; hardness 50~100 mg / L CaCO3), bred and provided by our company's aquarium. The laboratory animal use license number is: SYXK (Zhejiang) 2022-0004, and the husbandry and management meet the requirements of international AAALAC certification (certification number: 001458).

[0140] Zebrafish were bred through natural pair mating. Zebrafish aged 2 days post-fertilization (2 dpf) were used to determine the maximum detectable concentration (MTC) of the samples for promoting lipolysis and to evaluate the efficacy.

[0141] 2.2 MTC Measurement

[0142] Zebrafish of the Albino strain with a 2-dpf melanin allele mutation were randomly selected and treated in 6-well plates, with 30 zebrafish treated in each well (experimental group). Samples were administered via water-soluble solution, and a normal control group was also included. The volume per well was 3 mL. After treatment at 28℃ for 2 days, the MTC of the samples compared to normal zebrafish was measured. Under the experimental conditions, the MTC of the metagenerogenin promoting lipolysis was 2000 μg / mL.

[0143] 2.3 Evaluation of efficacy in promoting fat breakdown (phenotype)

[0144] Zebrafish of the Albino strain with a 2 dpf melanin allele mutant were randomly selected and placed in 6-well plates, with 30 zebrafish treated in each well (experimental group). Samples were administered via water-soluble solution, with resveratrol (11.4 μg / mL, Shanghai Aladdin Biochemical Technology Co., Ltd.) as the positive control. A normal control group was also included, with a volume of 3 mL per well. After treatment at 28℃ for 1 day, Nile red dye was administered via water-soluble solution to each experimental group. After another day of treatment at 28℃, 10 zebrafish from each experimental group were randomly selected and photographed under a fluorescence microscope. Data were analyzed and collected using NIS-Elements D 3.20 advanced image processing software. The fluorescence intensity of the yolk sac fat in zebrafish was analyzed, and the statistical analysis results of this index were used to evaluate the efficacy of the sample in promoting fat decomposition. Statistical results are expressed as mean ± SE. Statistical analysis was performed using SPSS 26.0 software. p < 0.05 indicated statistical significance. The experimental results are shown in Table 1 and [Table data would be inserted here]. Figure 1 .

[0145] Table 1. Experimental results evaluating the efficacy of the samples in promoting fat breakdown (n = 10)

[0146]

[0147] Note: Compared with the normal control group, *p < 0.05, ***p < 0.001

[0148] 2.4 Evaluation of the efficacy of promoting fat breakdown (genes)

[0149] Uncoupling protein 1 (ucp1) (Gene ID: 83908) is a specific protein on the inner mitochondrial membrane. When activated, ucp1 uncouples the oxidative phosphorylation of the mitochondrial respiratory chain, thereby inhibiting ATP synthesis and releasing energy as heat, increasing energy expenditure. Studies have shown that overexpression of ucp1 in white adipose tissue reduces body weight in obese mice; ucp1 activation increases energy output and reduces fatty acid synthesis.

[0150] Total RNA was extracted from each group of zebrafish in step 1.3 using the Universal RNA Extraction TL Kit C (Foshan Aowei Biotechnology Co., Ltd.). The concentration and purity of the total RNA were measured using a UV-Vis spectrophotometer, and all samples were found to be of acceptable quality. 2.00 μg of total RNA from the zebrafish sample was taken, and 20.0 μL of cDNA was synthesized according to the instructions of the cDNA first-strand synthesis kit. The ucp1 primer information is shown in Table 2.

[0151] Table 2. Primer sequence information for β-actin and ucp1 genes

[0152]

[0153] The expression of β-actin and ucp1 genes was detected by q-PCR. β-actin was used as an internal control for gene expression, and the relative RNA expression level of the ucp1 gene was calculated. Statistical results are expressed as mean ± SE. Statistical analysis was performed using SPSS 26.0 software. p < 0.05 was considered statistically significant. The results are shown in Table 3.

[0154] Table 3. Evaluation results of the efficacy of the samples in promoting lipolysis (ucp1 gene) (n = 3)

[0155]

[0156] Note: Compared with the normal control group,

[0157] The results indicate that metabiotics can promote lipolysis in zebrafish. Specifically, phenotypically, the fluorescence intensity of yolk sac fat was significantly reduced compared to the control group, and the lipolysis effect was best at a concentration of 2000 μg / mL.

[0158] At the gene level, compared with the control group, treatment with inactivated metabiotics at 1000 μg / mL and 2000 μg / mL significantly upregulated the relative expression level of UCP1. This result suggests that metabiotics may reduce the body weight of obese mice by upregulating UCP1 expression.

[0159] Example 3. Evaluation of the effect of metabiotics on inhibiting fat absorption in zebrafish

[0160] 3.1 Laboratory Animals

[0161] Wild-type AB strain zebrafish were raised in aquarium water at 28℃ (water quality: 200mg of instant sea salt added per 1L of reverse osmosis water, conductivity 450~550 μS / cm; pH 6.5~8.5; hardness 50~100 mg / L CaCO3), bred and provided by our company's aquarium. The laboratory animal use license number is: SYXK (Zhejiang) 2022-0004, and the husbandry and management meet the requirements of international AAALAC certification (certification number: 001458).

[0162] Zebrafish reproduce through natural pair mating. Zebrafish aged 5 dpf were used to determine the maximum detectable concentration (MTC) of their fat-inhibiting efficacy and to evaluate its effectiveness.

[0163] 3.2 MTC Measurement

[0164] Five dpf wild-type AB strain zebrafish were randomly selected and placed in beakers, with 30 zebrafish treated in each beaker (experimental group). Samples were administered in water solution, and a normal control group and a model control group were also set up. Each beaker had a volume of 20 mL. After treatment at 28℃ for 1 hour, except for the normal control group, all other concentration groups were fed pure egg yolk powder in water solution to establish a dietary fat absorption model. After continuing treatment at 28℃ for another day, the MTC of the sample on the model zebrafish was measured. Under the experimental conditions, the MTC of metabiotic inhibition of fat absorption was 2000 μg / mL.

[0165] 3.3 Evaluation of efficacy in inhibiting fat absorption (phenotype)

[0166] Five dpf wild-type AB strain zebrafish were randomly selected and placed in beakers, with 30 zebrafish treated in each beaker (experimental group). Samples were administered in water, with orlistat (Shandong New Era Pharmaceutical Co., Ltd.) at a concentration of 15.0 μg / mL as a positive control. A normal control group and a model control group were also included. Each beaker had a volume of 20 mL. After treatment at 28℃ for 1 h, except for the normal control group, all other concentration groups were fed pure egg yolk powder in water to establish a dietary fat absorption model. After another day of treatment at 28℃, whole-body fat was stained with Oil Red O. After destaining and bleaching, 10 zebrafish were randomly selected from each experimental group and photographed under a dissecting microscope. Data were collected using NIS-Elements D 3.20 advanced image processing software to analyze the staining intensity of intestinal and tail blood vessel fat. The statistical analysis results of this index were used to evaluate the efficacy of the samples in inhibiting fat absorption. Statistical results are expressed as mean ± SE. Statistical analysis was performed using SPSS 26.0 software; p < 0.05 indicated statistical significance. The results are shown in Table 4. Fat staining of zebrafish intestinal and tail blood vessels is shown in... Figure 2 .

[0167] Table 4. Evaluation results of the phenotypic effect of the samples on inhibiting fat absorption (n = 10)

[0168]

[0169] Compared with the model control group, ***p < 0.001

[0170] 3.4 Evaluation of the efficacy of inhibiting fat absorption (genes)

[0171] adipor2 (Gene ID: 560140) encodes the adiponectin receptor AdipoR2. Adiponectin is a hormone secreted by adipocytes that increases fatty acid burning and energy expenditure. Adiponectin activates the AMPK and PPARα pathways, thereby stimulating fatty acid oxidation, increasing fatty acid burning, and reducing tissue cholesterol levels in the liver. AdipoR2 is the receptor for the full-length adiponectin, mediating increased AMPK and PPARα ligand activity, as well as adiponectin's ability to oxidize fatty acids and take up glucose. Obesity reduces adiponectin levels and also reduces adipor2 expression, leading to decreased adiponectin sensitivity and reduced fatty acid burning capacity, creating a vicious cycle. The lepa gene (Gene ID: 100150233) encodes leptin, a protein hormone secreted by adipocytes. Leptin plays a major role in the regulation of energy homeostasis. Circulating leptin binds to leptin receptors in the brain, activating downstream signaling pathways that inhibit eating and promote energy expenditure. In obese individuals, leptin levels increase, thus inhibiting food intake, while weight loss leads to decreased leptin levels, thus increasing food intake.

[0172] Total RNA was extracted from each group of zebrafish in step 2.3 using the Universal RNA Extraction TL Kit C. The concentration and purity of total RNA were determined using a UV-Vis spectrophotometer. 2.00 μg of total RNA from the zebrafish sample was taken, and 20.0 μL of cDNA was synthesized according to the instructions of the cDNA first-strand synthesis kit. Primer information is shown in Table 5.

[0173] Table 5. Primer sequence information for β-actin, adipor2, and lepa genes

[0174]

[0175] The expression of β-actin, adipor2, and lepa genes was detected by q-PCR. β-actin was used as an internal control for gene expression, and the relative RNA expression levels of adipor2 and lepa genes were calculated. Statistical results are expressed as mean ± SE. Statistical analysis was performed using SPSS 26.0 software, and p < 0.05 was considered statistically significant. The results are shown in Table 6.

[0176] Table 6. Results of the experiment evaluating the efficacy of the samples in inhibiting fat absorption (gene) (n = 3)

[0177]

[0178] Compared with the model control group, *p < 0.05, **p < 0.01, ***p < 0.001

[0179] The results indicate that metabiotics have the effect of inhibiting fat absorption in zebrafish. Specifically, phenotypically, compared with the model control group, the staining intensity of fat in the intestinal and tail blood vessels was significantly reduced. Compared with the positive control drug, the fluorescence intensity at different concentrations was lower than that of orlistat, showing a better effect of inhibiting fat absorption.

[0180] At the gene level, compared with the model control group, metabiotic treatment significantly upregulated the relative expression level of adipor2 and downregulated the relative expression level of lepa. This result suggests that metabiotics may inhibit fat absorption by regulating the relative expression levels of these two genes.

[0181] Example 4. Study on the effect of post-biotics in improving obesity in mice

[0182] Eight-week-old male C57BL / 6J mice were housed at an ambient temperature of 21±2℃ and humidity of 30-70%, with 12-hour light-dark cycles, and free access to water and food. After 7 days of acclimatization, they were randomly divided into three groups of 16 mice each. The control group (CON) was fed a normal diet and administered saline by gavage; the model group (HFD) was fed a high-fat, high-cholesterol, high-fructose diet (HFHCD) and administered saline by gavage; and the inactivated bacteria group (HK207-1) was fed a high-fat, high-cholesterol, high-fructose diet and administered postbiotics (0.01g of inactivated bacteria powder dissolved in 200ul of saline by gavage). Food intake and body weight were recorded weekly. Mice were sacrificed at week 5, and blood and organs were collected.

[0183] After blood collection from the eyeballs, the blood was allowed to stand for at least 2 hours, then centrifuged at 2000×g at 4°C for 20 min to collect the supernatant. The supernatant was then collected and centrifuged again at 2000×g at 4°C for 5 min before serum separation. Liver, visceral fat (periglenoid, perirenal, and perigonadal fat), and inguinal subcutaneous fat were collected and weighed. Liver tissue was subjected to H&E staining; transcriptomics analysis was performed to detect the mRNA expression levels of the adipogenic gene SCD1 (Gene ID: 20249), the lipolysis gene HSL (Gene ID: 16890), and the fatty acid β-oxidation gene ACOX3 (Gene ID: 80911).

[0184] The body weight of mice in the model group was significantly higher than that in the control group two weeks after modeling. Figure 3 The total weight gain in the model group was significantly higher than that in the control group, indicating successful modeling. However, the body weight of mice fed with postbiotics was significantly lower than that of the model group after 5 weeks. Figure 4 Furthermore, the total weight gain in the group was significantly lower than that in the model group, indicating that post-biotics can improve obesity.

[0185] Mice fed a high-fat diet showed significantly higher levels of liver, subcutaneous, and visceral fat accumulation compared to the control group. Metabiotic intervention significantly reduced liver, periintestinal, and perirenal fat accumulation in mice, demonstrating potential in improving subcutaneous fat accumulation. This suggests that metabiotics can improve visceral fat accumulation and abdominal obesity. Liver transcriptome data showed that, compared to the model group, metabiotic intervention significantly downregulated the lipid synthesis gene SCD1 and significantly upregulated the lipid oxidation gene ACOX3. These results indicate that metabiotic intervention can regulate lipid metabolism by promoting lipolysis and lipid oxidation.

[0186] From liver staining results ( Figure 5 It can be seen that the high-fat diet model group caused ballooning lesions in liver cells, resulting in a certain degree of fatty liver. The intervention of metabiotics can alleviate this lesion, indicating that metabiotics have the effect of improving fatty liver.

[0187] Table 7. Fat weight of mouse organs

[0188]

[0189] Compared with the control group, *p < 0.05; compared with the model group, #p < 0.05

[0190] Table 8. Expression levels of genes involved in lipid metabolism in the liver

[0191]

[0192] Compared with the control group, *p < 0.05; compared with the model group, #p < 0.05

[0193] Although specific embodiments of the invention have been described in detail, those skilled in the art will understand that various modifications and variations can be made to the details based on all the published teachings, and all such changes are within the scope of protection of the invention. The entire scope of the invention is given by the appended claims and any equivalents thereof.

Claims

1. Use of a postbiotic or a composition comprising the postbiotic in the preparation of a medicament for preventing and / or ameliorating perirenal or perirenal fat accumulation in a subject; wherein the postbiotic is prepared by fermenting Bifidobacterium breve at 35-37 °C for 24-72 h, followed by heat inactivation at 70-95 °C, centrifuging the inactivated bacteria to obtain a precipitate, and vacuum freeze-drying the precipitate to obtain the postbiotic; wherein the Bifidobacterium breve is deposited with the Guangdong Microbial Culture Collection Center, and has the accession number GDMCC No. 60962.

2. The use of claim 1, wherein the subject has a BMI greater than 23.9 kg / m2 2 .

3. The use of claim 1, wherein, The Bifidobacterium breve is in the form of dead bacteria in the medicament.

4. The use of claim 1, wherein the medicament further comprises an additional active ingredient.

5. The use of claim 4, wherein the additional active ingredient is capable of promoting fat decomposition and / or inhibiting fat absorption.

6. The use of claim 4, wherein the additional active ingredient is L-carnitine.

7. The use of claim 4, wherein the additional active ingredient is capable of accelerating metabolism.

8. The use of claim 4, wherein the additional active ingredient is selected from tea polyphenols, caffeine, or any combination thereof.

9. The use of claim 8, wherein the additional active ingredient is a lipase inhibitor.

10. The use of claim 9, wherein the lipase inhibitor is orlistat.

11. The use of claim 1, wherein, The medicament has one or more characteristics selected from the following: (1) the medicament is a targeted gastrointestinal release medicament, or a controlled release medicament in the gastrointestinal tract; (2) the medicament further comprises a pharmaceutically acceptable carrier; (3) the medicament is in the form of a pill, powder, capsule, tablet, film-coated tablet, orally dissolving granules, liquid, suppository, or enema.

12. The use of claim 1, having one or more characteristics selected from the following: (1) the subject is a mammal; (2) the amount of postbiotic added to the medicament is 0.001 g-0.1 g; (3) the amount of postbiotic added to the medicament is 0.001-0.005 g, 0.005-0.01 g, 0.01-0.05 g, or 0.05-0.1 g.

13. The use of claim 12, wherein the mammal is selected from a mouse, a pig, a rabbit, a monkey, a sheep, a human, or any combination thereof.

14. The use of any one of claims 1-13, wherein, The composition comprises the postbiotic, and a microorganism selected from a bacterium, a fungus, or any combination thereof.

15. The use of claim 14, wherein the microorganism is a probiotic.

16. The use of claim 14, wherein the microorganism is a yeast.

17. The use of claim 16, wherein the yeast is selected from Saccharomyces cerevisiae, Saccharomyces boulardii, Kluyveromyces marxianus, or any combination thereof.

18. The use according to claim 14, wherein the bacteria are selected from the genera *Lactobacillus* spp., *Bifidobacterium* spp., *Bacillus* spp., *Propionibacterium* spp., *Streptococcus* spp., *Lactococcus* spp., *Pediococcus* spp., *Enterococcus* spp., *Staphylococcus* spp., or any combination thereof.

19. The use of claim 18, wherein, The intended use has one or more of the following characteristics: (1) The bacteria of the genus *Lactobacillus* mentioned are selected from: *Lactobacillus paracasei*, *Lactobacillus acidophilus*, *Lactobacillus brevis*, *Lactobacillus jensenii*, *Lactobacillus iners*, *Lactobacillus casei*, *Lactobacillus crispatus*, *Lactobacillus curvatus*, *Lactobacillus delbrueckii*, *Lactobacillus fermentum*, *Lactobacillus gasseri*, *Lactobacillus helveticus*, *Lactobacillus johnsonii*, *Lactobacillus plantarum*, and *Lactobacillus reuteri*. Lactobacillus reuteri, Lactobacillus rhamnosus, Lactobacillus sakei, Lactobacillus salivarius, or any combination thereof; (2) the bacteria of the genus Bifidobacterium are selected from the group consisting of Bifidobacterium animalis, Bifidobacterium bifidum, Bifidobacterium breve, Bifidobacterium infantis, Bifidobacterium longum, Bifidobacterium adolescentis, or any combination thereof; (3) the bacteria of the genus Bacillus are selected from the group consisting of Bacillus subtilis, Bacillus coagulans, or any combination thereof; (4) the bacteria of the genus Propionibacterium are selected from the group consisting of Propionibacterium shermanii, Propionibacterium freudenreichii, Propionibacterium acidipropionici, or any combination thereof; (5) the bacteria of the genus Streptococcus are selected from the group consisting of Streptococcus thermophilus, Streptococcus salivarius, or any combination thereof; (6) the bacteria of the genus Lactococcus is Lactococcus lactis; (7) the bacteria of the genus Enterococcus are selected from the group consisting of Enterococcus faecalis, Enterococcus faecium, Enterococcus mundtii, or any combination thereof.

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