CN106974940B - Application of probiotics of scleritis in treating and preventing obesity and related diseases - Google Patents

Abstract

The invention discloses the application of a Firmicutes probiotic in the treatment and prevention of obesity-related diseases. Specifically, the Firmicutes probiotic of the present invention is used to prepare a composition or a For one or more uses selected from the group consisting of: (a) preventing and/or treating obesity; (b) lowering blood lipids; (c) preventing or treating cardiovascular disease; and/or (d) preventing and/or treating diabetes. The Firmicutes probiotics of the present invention can significantly reduce body weight, blood lipids and body fat ratio.

Description

Application of Firmicutes probiotics in the treatment and prevention of obesity and related diseases

technical field

The invention belongs to the field of microbiology, in particular, the invention relates to the application of Firmicutes probiotics in the treatment and prevention of obesity and related diseases, and also relates to compositions comprising Firmicutes probiotics and applications thereof.

Background technique

There are a large number of symbiotic microorganisms in the human body, most of which reside in the human gut, and the number exceeds 1000 trillion (10 ¹⁴ orders of magnitude), which is more than 10 times the total number of human cells. In the long evolutionary process, gut microbes have reached a good cooperation with humans and play a crucial role in human nutrition, metabolism and immunity. Many researchers regard the human gut microbial community more as a It is an "organ" of the human body, or the second genome of the human body, and the massive genetic information contained in it is closely related to human health. Through the study of nearly 10,000 samples of hundreds of diseases such as diabetes, coronary heart disease, obesity, colon cancer, etc., it is found that some specific species have a significant relationship with the disease. These results are used in the clinical assessment and diagnosis of the disease and later intervention. Therapy offers a whole new direction.

Obesity is a chronic disease, many factors can lead to obesity, and its origin has not been clearly studied. Obesity is also an inducing factor of a series of diseases, such as hypertension, diabetes, coronary heart disease, gallbladder disease, osteoarthritis, sleep apnea, breathing disorders, uterine tumors, prostate cancer, breast cancer and colon cancer. According to the NIH report, about 97 million Americans are currently overweight and obese, of which about 15.1 million are type 2 diabetes related to obesity, and about 200,000 people die each year from obesity-related diseases.

Obesity is usually an excess of body fat due to altered physiological or biochemical functions. Fats typically include neutral lipids, phospholipids and cholesterol. Fat gain is due to energy intake being greater than energy expenditure. In terms of pathogenesis, there are two types of obesity: (a) simple obesity and (b) second obesity. Simple obesity can be divided into idiopamic obesity and acquired obesity, and the number of patients with simple obesity can account for more than 95% of the total obesity. Congenital obesity is caused by a large number of fat cells and is common in childhood obesity. Acquired obesity is caused by larger size fat cells and is common in obesity in adulthood. Secondary obesity, also known as symptomatic obesity, is usually caused by endocrine or metabolic diseases.

There are currently five strategies for treating obesity: diet, exercise, behavioral therapy, drug therapy, and therapetltic operation. Which strategy is adopted depends mainly on the patient's health risk factors and the rate and effect of weight loss, and these strategies can be selected or combined in the treatment of obese patients. The speed and effectiveness of its weight loss is influenced by multiple factors such as age, height, family history, and risk factors. Diet-exercise therapy, i.e. eating low-calorie, low-fat foods and doing aerobic exercise, is generally considered unsuccessful for the general public and requires long-term and regular adherence; body fat removal surgery can achieve immediate results, but There are many limitations, such as surgical risks, unsustainable fat removal, and high cost.

Drug therapy is currently the main clinical method for treating obesity and obesity-related diseases such as diabetes. Mechanisms of drug therapy include appetite suppression, increased energy expenditure, stimulation of fat movement, decreased triglyceride synthesis, and inhibition of fat absorption. At present, the main drugs are: phenylpropanolamine (PPA), Roche (orlistat, XenicalⅢ) and sibutramine (ReductilTM). Hyperglycemia in some diabetics remains inadequately controlled by diet and/or exercise therapy or the use of the above therapeutic compounds. For these patients, exogenous insulin should be used. The use of exogenous insulin is an expensive and painful procedure for the patient, and it also brings a variety of complications to the patient. For example, miscalculation of insulin doses due to not eating or exercising regularly can lead to an insulin response (hypoglycemia). In addition, local or systemic hypersensitivity or immune resistance to the drug may also occur with the use of the drug.

At present, there is no effective method and drug for treating and preventing obesity and related diseases with few side effects in the art.

Therefore, there is an urgent need in the art to develop a new, non-toxic and side-effect drug for the treatment and prevention of obesity and related diseases.

SUMMARY OF THE INVENTION

An object of the present invention is to provide the use of Firmicutes probiotics in the treatment and prevention of obesity and related diseases.

Another object of the present invention is to provide an effective medicine, beverage, food composition, or animal feed composition for treating and preventing obesity and related diseases without toxic and side effects.

Another object of the present invention is to provide a method for reducing body weight and/or blood sugar and use thereof.

A first aspect of the present invention provides the use of a Firmicutes probiotic for the preparation of a composition or formulation for one or more uses selected from the group consisting of: (a) prevention and/or treating obesity; (b) lowering blood lipids; (c) preventing or treating cardiovascular disease; and/or (d) preventing and/or treating diabetes, wherein the Firmicutes probiotics are selected from the group consisting of: Coprococcus eutactus, Dialister invisus, or a combination thereof.

In another preferred embodiment, the Firmicutes probiotics include Coprococcuseutactus.

In another preferred embodiment, the Coprococcus eutactus is selected from the group consisting of Coprococcus eutactus ATCC 27759, Coprococcus eutactus ATCC 51897, Coprococcus eutactus L2-50, or a combination thereof.

In another preferred embodiment, the Firmicutes probiotics include Dialister invisus.

In another preferred embodiment, the Dialister invisus is selected from the group consisting of Dialister invisus DSM 15470, Dialister invisus E2.20, Dialister invisus E9.48, or a combination thereof.

In another preferred example, the Firmicutes probiotics include one or more selected from Table 3.

In another preferred example, the Firmicutes probiotics are selected from Table 3 and are from the same or different genera.

In another preferred example, the composition is selected from the following group: food composition, health care composition, pharmaceutical composition, beverage composition, feed composition, or a combination thereof.

In another preferred embodiment, the composition is an oral preparation.

In another preferred example, the composition is a liquid preparation, a solid preparation, or a semi-solid preparation.

In another preferred embodiment, the dosage form of the composition is selected from the group consisting of powder, powder, tablet, sugar coating, capsule, granule, suspension, solution, syrup, drop, and tongue Lower lozenge.

In another preferred embodiment, the food composition includes an emulsion product, a solution product, a powder product, or a suspension product.

In another preferred embodiment, the food composition includes dairy products, milk powder, or emulsion.

In another preferred embodiment, the liquid preparation is selected from the following group: a solution product or a suspension product.

A second aspect of the present invention provides the use of a Firmicutes probiotic for the preparation of a composition or formulation for one or more uses selected from the group consisting of: (i) reducing The level of monocyte chemoattractant protein-1 (MCP-1) in mammals; and/or (ii) improving leptin resistance and increasing the sensitivity to leptin in vivo, wherein the Firmicutes probiotics are selected from The lower group: Coprococcus eutactus, Dialister invisus, or a combination thereof.

In another preferred embodiment, the composition or preparation is also independently or additionally used for one or more purposes selected from the group consisting of:

(iii) inhibiting body weight gain in mammals;

(iv) reducing the body fat ratio (fat weight/body weight ratio) in mammals;

(v) reducing blood lipid levels in mammals;

(vi) increasing the level of high density lipoprotein (HDLC) in mammals;

(vii) reducing low density lipoprotein (LDLC) levels in a mammal.

In another preferred embodiment, the mammals include humans and rodents (eg, rats, mice).

In another preferred embodiment, the reducing the blood lipid level of the mammal comprises reducing the level of total cholesterol (TC) and/or the level of triglyceride.

A third aspect of the present invention provides a composition for treating and/or preventing obesity, the composition comprising: (i) a safe and effective amount of Firmicutes probiotics; and (ii) food or pharmacy An acceptable carrier; wherein the Firmicutes probiotic is selected from the group consisting of Coprococcus eutactus, Dialister invisus, or a combination thereof.

In another preferred embodiment, the Firmicutes probiotics include Coprococcuseutactus.

In another preferred embodiment, the Coprococcus eutactus is selected from the group consisting of Coprococcus eutactus ATCC 27759, Coprococcus eutactus ATCC 51897, Coprococcus eutactus L2-50, or a combination thereof.

In another preferred embodiment, the Firmicutes probiotics include Dialister invisus.

In another preferred embodiment, the Dialister invisus is selected from the group consisting of Dialister invisus DSM 15470, Dialister invisus E2.20, Dialister invisus E9.48, or a combination thereof.

In another preferred example, the composition is selected from the following group: food composition, health care composition, pharmaceutical composition, beverage composition, feed composition, or a combination thereof.

In another preferred example, the composition contains 1×10-1×10 ²⁰ cfu/mL or cfu/g Firmicutes probiotics, preferably 1×10 ⁴ -1×10 ¹⁵ cfu/mL or cfu/g of Firmicutes probiotics, based on the total volume or total weight of the composition.

In another preferred example, the composition contains 0.0001-99 wt %, preferably 0.1-90 wt % of the Firmicutes probiotics, based on the total weight of the composition.

In another preferred embodiment, the composition is in unit dosage form (one tablet, one capsule or one vial), and the mass of the composition in each unit dosage form is 0.05-5 g, preferably 0.1-1 g.

In another preferred example, the composition further contains other probiotics and/or prebiotics.

In another preferred example, the other probiotics are selected from the group consisting of lactic acid bacteria, bifidobacteria, lactobacillus acidophilus, or a combination thereof.

In another preferred embodiment, the prebiotics are selected from the group consisting of fructooligosaccharides (FOS), galactooligosaccharides (GOS), xylo-oligosaccharides (XOS), lactultooligosaccharides (LACT), soybean low Polysaccharide (SOS), inulin (Inulin), or a combination thereof.

The fourth aspect of the present invention provides a preparation method of the composition described in the third aspect of the present invention, comprising the steps:

(i) Firmicutes probiotics, and (ii) a food acceptable carrier or a pharmaceutically acceptable carrier are mixed to form the composition of the third aspect of the present invention.

In another preferred embodiment, the composition is an oral preparation.

The fifth aspect of the present invention provides a method for reducing body weight and/or blood lipids, by administering (i) Firmicutes probiotics or the composition of the third aspect of the present invention to the subject.

In another preferred embodiment, the administration comprises oral administration.

In another preferred embodiment, the administration dose is 0.01-5g/50kg body weight/day, preferably 0.1-2g/50kg body weight/day.

In another preferred embodiment, the subject includes mammals, such as humans.

It should be understood that within the scope of the present invention, the above-mentioned technical features of the present invention and the technical features specifically described in the following (eg, the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, it is not repeated here.

Description of drawings

Figure 1 shows the weight gain of mice in each group after gavage with faecalis concomitant compared with that before gavage.

Figure 2 shows the weight gain of mice in each group after intragastric administration of dialisteria turbidum compared with that before intragastric administration.

Figure 3 shows the weight gain of the mice in each group after gavage of the combined bacteria compared with that before gavage.

Figure 4 shows the body fat ratio of mice in each group after 9 weeks of gavage with Faecalicoccus congeners.

Figure 5 shows the body fat ratio of mice in each group after 9 weeks of intragastric administration of D. dialistica.

Figure 6 shows the body fat ratio of mice in each group after 9 weeks of gavage with the combined bacteria.

Figure 7 shows the effect of intragastric administration of Faecalicoccus concomitensis on blood lipids.

Figure 8 shows the effect of intragastric administration of dialisteria turbidum on blood lipids.

Figure 9 shows the effect of gavage combined bacteria on blood lipids.

Figure 10 shows the effect of gavage of Faecalicoccus congeners on monocyte chemoattractant protein-1 (MCP-1).

Figure 11 shows the effect of gavage of L. dialustris on monocyte chemoattractant protein-1 (MCP-1).

Figure 12 shows the effect of gavage combined bacteria on monocyte chemoattractant protein-1 (MCP-1).

Figure 13 shows the effect of gavage of Faecalicoccus congeners on leptin (Leptin, LEP).

Figure 14 shows the effect of gavage of L. dialustris on leptin (LEP).

Figure 15 shows the effect of gavage of the combination bacteria on leptin (Leptin, LEP).

Detailed ways

After extensive and in-depth research and experiments, the inventors unexpectedly found that Coprococcuseutactus and/or Dialister invisus can prevent and treat obesity and related diseases (such as cardiovascular disease). The active composition containing the above-mentioned Firmicutes probiotics was fed to the experimental subjects, and it was found that the composition could inhibit weight gain, reduce body fat ratio, lower blood lipids, and effectively alleviate cardiovascular and obesity diseases. The present invention has been completed on this basis.

As used herein, the term "comprising" means that the various ingredients can be used together in the mixture or composition of the present invention. Thus, the terms "consisting essentially of" and "consisting of" are encompassed by the term "comprising".

As used herein, the "body fat ratio" refers to the ratio of fat weight/body weight.

Firmicutes probiotics of the present invention and application thereof

As used herein, the "Firmicus probiotics of the present invention" refer to a group of bacteria of the phylum Firmicutes commensal in the intestinal tract, Gram-positive, rod-shaped, such as Clostridium XIVa cluster and IV cluster (Clostridium cluster XIVa and IV), including some species of Clostridium, Eubacterium, Ruminococcus and Anaerofilum. By interacting with other microbes in the gut, such probiotics play an important role in the balance of the gut microbiota, and at the same time, perform other specific or essential functions.

In the present invention, the "firmicute probiotics of the present invention" refers to a mixture of one or more bacterial genera of Coprococcuseutactus and Dialister invisus, or a One or more mixtures of bacteria in each genus.

Among them, Coprococcus eutactus is an obligate anaerobic bacterium that cannot move. When it shows a paired or chain-like paired state, it is Gram-positive, but it can quickly fade. In medium containing fermentable carbohydrates, it takes on a slightly elongated shape. But the bacteria are usually round, with a diameter of 0.7-1.3 μm.

Dialister invisus is an obligate anaerobic, immotile coccus (0.3-0.4 × 0.3-0.6 μm) that is Gram-negative. It can be in a single dispersed form, or in pairs, short chains, small clusters, etc.

The present invention provides the application of Firmicutes probiotics in the treatment and prevention of obesity and related diseases (such as cardiovascular diseases). The Firmicutes probiotics have the ability to (i) inhibit the subject's weight gain; (ii) reduce blood lipids; and (iii) reduce the body fat ratio in subjects consuming a high-fat diet. According to a preferred example of the present invention, C57BL fed with a high-fat food that can lead to obesity treated with the Firmicutes probiotics of the present invention (such as Faecalicoccus comorans, Dialisteria turbidum, or a combination thereof) /6J male mice, compared with untreated controls, showed slower weight gain and lower blood lipids, as well as various markers associated with obesity or cardiovascular disease, such as leptin (LEP) and monocytes Chemotactic protein-1 (MCP-1). Therefore, the Firmicutes-based probiotics of the present invention (eg, Faecalicoccus concomitans, Dialisteria turbidum, or a combination thereof) can be used to prevent and treat obesity and diseases caused by obesity, such as cardiovascular disease and the like.

Composition and its application

The present invention also provides a composition, preferably a pharmaceutical composition. The composition includes an effective amount of the Firmicutes probiotics of the present invention (such as Faecalicoccus concomitans, Dialisteria turbidum, or a combination thereof), and in a preferred embodiment, the composition further includes a probiotic selected from the group consisting of: Probiotics: Lactobacillus, Bifidobacterium, Lactobacillus acidophilus, or combinations thereof; and/or Prebiotics selected from the group consisting of fructooligosaccharides (FOS), galactooligosaccharides (GOS), xylo-oligosaccharides ( XOS), lactultooligosaccharides (LACT), soy oligosaccharides (SOS), inulin (Inulin), or a combination thereof.

In a preferred example, the composition is a liquid preparation, a solid preparation, or a semi-solid preparation.

In a preferred embodiment, the liquid preparation is selected from the following group: a solution product or a suspension product.

In a preferred embodiment, the dosage form of the composition is selected from the group consisting of powder, powder, tablet, sugar coating, capsule, granule, suspension, solution, syrup, drop, and sublingual lozenge.

The pharmaceutical composition of the present invention can be administered in any form of pharmaceutical tablet, injection or capsule, and the pharmaceutical preparation includes excipients, pharmaceutically acceptable vehicles and carriers, and these substances can be selected according to the route of administration. The pharmaceutical formulations of the present invention may further comprise auxiliary active ingredients.

Lactose, glucose, sucrose, sorbitol, mannose, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, fine crystalline cellulose, polyvinylpyrrolidone (PVP), cellulose, water, syrup, Methyl cellulose, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate or mineral oil etc. can be used as the carrier, excipient or diluent etc. of the pharmaceutical composition of the present invention.

In addition, the pharmaceutical composition of the present invention may further include lubricants, wetting agents, emulsifiers, suspension stabilizers, preservatives, sweeteners, flavors, and the like. The pharmaceutical composition of the present invention can be produced in enteric-coated formulations by various well-known methods so that the active ingredient of the pharmaceutical composition, ie, microorganisms, can pass smoothly through the stomach without being destroyed by gastric acid.

In addition, the microorganisms of the present invention can be used in the form of capsules prepared by conventional methods. For example, standard excipients and the freeze-dried microorganisms of the present invention are mixed to form pellets, which are then filled into gelatin capsules. In addition, the microorganisms and medicaments of the present invention allow the use of excipients such as liquid gum, cellulose, silicate or mineral oil, etc., to be mixed to make suspensions or dispersions, which can be filled into soft gelatin capsules middle.

The pharmaceutical composition of the present invention can be made into enteric-coated tablets for oral use. The term "enteric coating" in this application includes all conventional pharmaceutical acceptable coatings which are not degraded by gastric acid, but which are sufficiently decomposed in the small intestine to rapidly release the microorganisms of the present invention. The inventive casing can be maintained at 36-38°C for more than 2 hours in synthetic gastric acid such as pH=1 HCl solution, and preferably decompose within 1.0 hour in synthetic intestinal fluid such as pH=7.0 buffer.

The enteric coating of the present invention is coated at about 16-30 mg per tablet, preferably 16-25 mg, more preferably 16-20 mg. In the present invention, the thickness of the casing is 5-100 μm, and the ideal thickness is 20-80 μm. The enteric coating ingredients are selected from conventional polymers known to the public.

The preferred enteric coating of the present invention is composed of a cellulose acetate phthalate polymer or a trimellitate polymer and a copolymer of methacrylic acid (for example, containing more than 40% methacrylic acid and containing methyl cellulose phthalic acid) Copolymers of methacrylic acid of hydroxypropyl ester or its ester derivatives).

The viscosity of the cellulose acetate phthalate used in the enteric coating of the present invention is about 45-90 cp, the acetyl content is 17-26%, and the phthalic acid content is 30-40%. Cellulose acetate trimephthalate used in enteric coatings has a viscosity of about 5-21 cp and a content of 17-26% acetylphthalide. Cellulose acetate trimellitate is produced by Eastman Keda Company and can be used for the casing material in the present invention.

The hydroxypropyl methylcellulose phthalate used in the casing of the present invention generally has a molecular weight of 20,000-130,000 daltons, an ideal molecular weight of 80,000-100,000 daltons, and a hydroxypropyl content of 5-10%. The methoxy content is 18-24% and the phthaloyl content is 21-35%.

The hydroxypropyl methylcellulose phthalate used in the casing of the present invention is HP50, produced by Shin-EtsuChemidnl Co. Ltd., Japan. HP50 contains 6-10% hydroxypropyl, 20-24% methoxy, 21-27% propyl, and has a molecular weight of 84,000 Daltons. Another enteric coating material is HP55, HP55 contains 5-9% hydroxypropyl methylcellulose phthalate, 18-22% methoxy, 27-35% phthalic acid, and has a molecular weight of 78,000 Daltons.

The casings of the present invention are prepared by spraying the casing solution onto the core using conventional methods. All solvents in this enteric coating method are alcohols (eg, ethanol), ketones (eg, acetone), halogenated hydrocarbon compounds (eg, methylene chloride), or combinations thereof. Softeners such as di-n-butylphthalate and triacetin are added to the enteric coating solution in a ratio of 1 part coating to about 0.05 or about 0.3 parts softener. The spraying method is preferably carried out continuously, and the amount of material sprayed can be controlled according to the conditions used for coating. The spray pressure can be adjusted at will, in general, ideal results can be obtained at an average pressure of 1-1.5 Pa.

"Pharmaceutically effective amount" in the specification refers to an amount that can produce function or activity on humans and/or animals and can be accepted by humans/or animals. For example, in the present invention, it can be prepared to contain 1×10-1×10 ²⁰ cfu/ml or cfu/g (specially, it can contain 1×10 ⁴ to 1×10 ¹⁵ cfu/ml or cfu/g; more particularly Alternatively, formulations of Firmicutes probiotics of the present invention (eg, Faecalicoccus parafasciens, Dialisteria turbidum, or combinations thereof) may contain 1×10 ⁶ to 1×10 ¹¹ cfu/ml or cfu/g).

When used to prepare a pharmaceutical composition, the effective dose of the Firmicutes probiotics of the present invention (eg, Faecalicoccus concomitans, Dialisteria turbidum, or a combination thereof) used may vary with the mode of administration and the disease to be treated. vary in severity. Dosage form suitable for oral administration, comprising about 1×10-1×10 ²⁰ cfu/ml or cfu/g (specially, may contain 1×10 ⁴ -1× 10 ¹⁵ cfu/ml or cfu/g; more particularly, may contain 1 x 10 ⁶ -1 x 10 ¹¹ cfu/ml or cfu/g) of active Firmicutes probiotics (such as Faecalicoccus companis, Diali turbidum) Streptomyces or a combination thereof). This dosage regimen can be adjusted to provide optimal therapeutic response. For example, several divided doses may be administered daily, or the dose may be proportionally reduced, as dictated by the exigencies of the therapeutic situation.

The Firmicutes probiotics (such as Faecalicoccus companis, Dialisteria turbidum or a combination thereof) can be administered orally or the like. Solid carriers include: starch, lactose, dicalcium phosphate, microcrystalline cellulose, sucrose, and kaolin, while liquid carriers include: culture medium, polyethylene glycols, nonionic surfactants, and edible oils (eg, corn oil, peanut oil, etc.) and sesame oil), as long as it suits the characteristics of Firmicutes probiotics (eg, Faecalicoccus compartans, Dialisteria opacus, or a combination thereof) and the specific mode of administration required. Adjuvants commonly used in the preparation of pharmaceutical compositions may also advantageously be included, such as flavors, colors, preservatives and antioxidants such as vitamin E, vitamin C, BHT and BHA.

From the standpoint of ease of preparation and administration, the preferred pharmaceutical compositions are solid compositions, especially tablets and solid- or liquid-filled capsules. Oral administration is preferred.

The compositions of the present invention are administered to the individual one or more times per day. Dosage units for administration represent dosages which can be divided in form and suitable for use in humans or all other mammalian subjects. Each unit contains a pharmaceutically acceptable carrier and a therapeutically effective amount of the microorganism of the invention. The amount administered will vary with the weight and severity of obesity of the patient, the supplemental active ingredients included and the microorganism employed. In addition, if possible, the administration may be divided, and if necessary, the administration may be consecutive. Therefore, the said doses do not limit the present invention. In addition, the "composition" in the present invention means not only a medicine but also a functional food and a health supplement. In a preferred example, the composition includes: beverage, food, medicine, animal feed and the like.

In a preferred embodiment of the present invention, there is also provided a food composition, which contains an effective amount of Firmicutes probiotics (such as Faecalicoccus compartans, Dialisteria turbidum or a combination thereof), and the balance of A food acceptable carrier, the dosage form of the food composition is selected from solid, dairy, solution, powder, or suspension products.

In a preferred example, the formulation of the composition is as follows:

1 × 10-1 × 10 ²⁰ cfu/mL Firmicutes probiotics (such as Faecalicoccus compartans, Dialisteria turbidum, or a combination thereof); and a food or pharmaceutically acceptable carrier, and/or excipient Form.

In another preferred embodiment, the formula of the composition is as follows:

1 x 10 ⁶ -1 x 10 ¹¹ cfu/mL Firmicutes probiotics (such as Faecalicoccus comorbidis, Dialisteria opacus or a combination thereof); and a food or pharmaceutically acceptable carrier, and/or excipient.

Methods of reducing body weight and/or blood lipids

In another preferred example, the method comprises: ingesting the pharmaceutical composition, food composition, beverage composition, or combination thereof of the present invention. The experimental subjects are humans.

In another preferred example, the method comprises: ingesting the pharmaceutical composition, food composition, or animal feed of the present invention, or a combination thereof. The experimental object is an animal, preferably a mouse or a rabbit.

The main advantages of the present invention include:

(a) The Firmicutes probiotics of the present invention (eg, Faecalicoccus comorans, Dialisteria turbidum, or a combination thereof) can significantly reduce body weight, blood lipids, and body fat ratio.

(b) Firmicutes probiotics of the present invention (such as Faecalicoccus comorans, Dialisteria turbidum, or a combination thereof) can significantly reduce indicators (such as cholesterol and triglycerides).

(c) The Firmicutes probiotics of the present invention (eg, Faecalicoccus comorans, Dialisteria turbidum, or a combination thereof) can significantly reduce the levels of total cholesterol, triglycerides, and low-density lipoproteins.

(d) The Firmicutes probiotics of the present invention (eg, Faecalicoccus comorata, Dialisteria turbidum, or a combination thereof) can significantly increase the level of high-density lipoprotein.

(e) Firmicutes probiotics of the present invention (eg, Faecalicoccus comorans, Dialisteria opacus or a combination thereof) can improve insulin resistance, and can also reduce the risk of atherosclerosis and cardiovascular disease.

(f) The Firmicutes probiotics of the present invention (eg, Faecalicoccus parasites, Dialisteria turbidum, or a combination thereof) can significantly reduce the level of monocyte chemoattractant protein-1 (MCP-1).

(g) The Firmicutes probiotics of the present invention (eg, Faecalicoccus concomitans, Dialisteria turbidum, or a combination thereof) can effectively improve the leptin resistance associated with obesity, and increase the in vivo sensitivity to Leptin.

The present invention will be further described below in conjunction with specific embodiments. It should be understood that these examples are only used to illustrate the present invention and not to limit the scope of the present invention. The experimental method of unreceipted specific conditions in the following examples, usually according to conventional conditions such as people such as Sambrook, molecular cloning: conditions described in laboratory manual (New York: Cold Spring Harbor Laboratory Press, 1989), or according to "microorganism: Conditions described in the Laboratory Manual (edited by James Cappuccino and Natalie Sherman, Pearson Education Press), or as suggested by the manufacturer.

Example 1 Food composition containing bacterial components (such as faecalis commensal, dialisteria turbidum, or a combination thereof)

The raw materials are listed in Table 1.

Table 1 Food composition formula

raw material Mass percentage (%) bacterial components 0.5 milk 90.0 white sugar 9.5

The bacterial components in formulas 1-6 are single bacterial components, which respectively contain Coprococcus eutactus ATCC27759, Coprococcus eutactus ATCC 51897, Coprococcus eutactus L2-50, Dialisterinvisus DSM 15470, Dialister invisus E2.20, and Dialister invisus E9.48.

The bacterial component in formula 7 is a mixture of any two or more (preferably two or three) of the above-mentioned six kinds of bacteria (the weight ratio is 1:1 or 1:1:1).

Mix milk and sugar according to the above formula ratio, stir until completely mixed, preheat, homogenize under 20Mpa pressure, sterilize at about 90°C for 5-10 minutes, cool to 40-43°C, inoculate 1-100×10 ⁶ cfu/g bacteria Component, that is, a food composition made of a bacterial component (such as Faecalicoccus comorans, Dialisteria turbidum, or a combination thereof).

Example 2

Pharmaceutical compositions containing bacterial components (such as Faecalicoccus comorans, Dialisteria turbidum, or a combination thereof)

The raw material ratio is shown in Table 2.

Table 2 Pharmaceutical composition formula

raw material Mass percentage (%) bacterial components 1.0% lactose 2.0% yeast 2.0% Peptone 1.0% pure water 94.0%

The bacterial components in formulas 1-6 are single bacterial components, which respectively contain Coprococcus eutactus ATCC27759, Coprococcus eutactus ATCC 51897, Coprococcus eutactus L2-50, Dialisterinvisus DSM 15470, Dialister invisus E2.20, and Dialister invisus E9.48.

The bacterial component in formula 7 is a mixture of any two or more (preferably two or three) of the above-mentioned six kinds of bacteria (the weight ratio is 1:1 or 1:1:1). Mix lactose, yeast powder and peptone with pure water according to the proportion, preheat to 60-65℃, 20Mpa pressure to homogenize, sterilize at about 90℃ for 20-30 minutes, cool to 36-38℃, insert bacterial components ( 1-50×10 ⁶ cfu/mL), fermented at 36-38° C. to pH 6.0, centrifuged, and freeze-dried to a moisture content of less than 3% to prepare a freeze-dried product containing bacterial components. Weigh 0.5 g of the freeze-dried product containing bacterial components and mix them with maltodextrin in equal amounts and put them into capsules to make medicines containing bacterial components (such as Faecalicoccus faecalis, Dialisteria turbidum, or a combination thereof). combination.

Example 3 Therapeutic effect on obese model mice

Experimental Materials:

Mice: C57BL/6J male mice (purchased from Guangdong Provincial Medical Laboratory Animal Center) were purchased, which were normally raised mice, 6 weeks old. Mice were grown in the same environment and fed the same food.

6 strains of Firmicutes probiotics were obtained from the preservation institution and preserved in Shenzhen Huada Gene Research Institute. At the same time, Lactobacillus plantarum was selected, from the General Microorganism Center of the China Microorganism Culture Collection Management Committee (CGMCC), with the deposit number CGMCC No. 8198, as the control group (LP group), in the MRS medium, 37 ℃ cultured 24-48h.

Among them, the source information of the above six Firmicutes probiotics is shown in Table 3. All strains were identified by 16S rDNA sequencing and the experiment was started.

See Table 3 for strain information.

Table 3 Strain Information

Among them, Coprococcus eutactus L2-50 (Bacteria 3) was derived from the Rowett Research Institute (Barcenilla A, Pryde SE, Martin JC et al. Phylogenetic Relationships of Butyrate-Producing Bacteria from the Human Gut. Applied and Environmental Microbiology 2000) ;66:1654-1661.). Both Dialister invisus E2.20 (Bacteria 5) and Dialister invisus E9.48 (Bacteria 6) were obtained from the Department of Microbiology, Dental Institute, Guy's Tower, Guy's Hospital, King's College, London, UK. London, UK; J. Downes, M. Munson and W. G. Wade, Dialister invisus sp. nov., isolated from the human oral cavity, International Journal of Systematic and Evolutionary Microbiology (2003), 53, 1937–1940).

High-fat feed (HF): containing 78.8% basic feed, 1% cholesterol, 10% egg yolk powder, 10% lard and 0.2% bile salts, purchased from Nantong Trophy Feed Technology Co., Ltd.

Common maintenance feed: purchased from Guangdong Provincial Medical Laboratory Animal Center.

experimental method:

Normal feeding C57BL/6J adult male mice were selected and randomly divided into control group (CK), bacteria group (bacteria group 1, bacteria group 2, bacteria group 3, bacteria group 4, bacteria group 5, bacteria group 6 group) , Bacteria 1+Bacteria 5 groups, Bacteria 2+Bacteria 6 groups, Bacteria 3+Bacteria 4 groups), the control group (LP, Lactobacillus plantarum CGMCC No.8198) and the obesity model group (HF), 10 in each group, Eat and drink freely in an SPF (Specific Pathogen Free) environment. The LP group, HF group and bacterial group were fed with high-fat diet, and the CK group was fed with normal maintenance diet. After 4 weeks of feeding, the inoculum group and LP group were gavaged with the bacterial solution of the corresponding strain; the HF group and the CK group were gavaged with the same amount of medium for 9 weeks.

The amount of bacteria fed was 0.15mL/10g body weight, the bacterial concentration was 1×10 ⁷ cfu/mL, and the concentration after concentration was 1×10 ⁸ cfu/mL, and the frequency was once every other day. The bacterial liquid should be cultured in advance, activated every week to ensure freshness, and the concentration should be determined and adjusted to 1×10 ⁸ cfu/mL. For the single inoculum group, the corresponding bacterial solution was given by gavage at the above dose; for the mixed inoculum group, each single bacterial solution was mixed in equal proportions and then administered by gavage at the above dose.

During the experimental period, data such as body weight, state, and food intake of the mice were recorded every week. Glucose tolerance (OGTT) test was performed on mice in each group in the last week of the experiment. After the experiment, the mice were sacrificed, the weight of fat was recorded, and serum was collected to detect the content of blood lipids and protein factors with Elisa kit.

Experimental results:

(1) Effects of Faecalicoccus comorbidis, Dialisteria turbidum or their combination on the body weight of mice.

Table 4. The weight gain of mice in each group after gavage with faecalis coccus was compared with that before gavage (Fig. 1)

Note: The data in the table is the mean ± standard deviation. For any two groups of data in each column, no same letter after the number indicates significant difference (p<0.05). Table 5-15 is the same.

Table 5 The weight gain of mice in each group after gavage with dialisteria turbidum compared with that before gavage (Figure 2)

Table 6 The weight gain of the mice in each group after gavage with the combined bacteria compared with that before the gavage (Fig. 3)

The results are shown in Table 4-6 and Figure 1-3. The results showed that Faecalicoccus concomitans, Dialisteria opacus, or their combination could effectively slow down the weight gain of obese model mice (P<0.05).

(2) The effect of Faecalicoccus concomitans, Dialisteria turbidum or their combination on body fat ratio.

Table 7 Body fat ratio of mice in each group after 9 weeks of gavage with Faecalicoccus concomitans (Fig. 4)

grouping Fat weight/body weight x 100% CK 2.83±0.13e Bacteria 1 4.11±0.10c Bacteria 2 4.00±0.26cd Bacteria 3 3.85±0.19d LP 5.39±0.34b HF 7.48±0.46a

Table 8 Body fat ratio of mice in each group after 9 weeks of intragastric administration of dialisteria turbidum (Fig. 5)

grouping Fat weight/body weight x 100% CK 2.91±0.14e Bacteria 4 3.97±0.22cd Bacteria 5 4.16±0.23c Bacteria 6 3.76±0.20d LP 5.21±0.24b HF 746±064a

Table 9 Body fat ratio of mice in each group after 9 weeks of gavage with combined bacteria (Fig. 6)

grouping Fat weight/body weight x 100% CK 2.90±0.13d Bacteria 1+5 3.73±0.19c Bacteria 2+6 3.87±0.23c Bacteria 3+4 3.93±0.22c LP 5.42±0.17b HF 756±055a

The results are shown in Table 7-9 and Figure 4-6. The results showed that Faecalicoccus concomitans, Dialisteria turbidum or their combination could significantly reduce the body fat ratio of obese model mice (P<0.05).

(3) The effect of Faecalicoccus concomitans, Dialisteria turbidum or their combination on blood lipids.

Table 10 The blood lipid content of mice in each group after 9 weeks of gavage with Faecalicoccus concomitans (Figure 7)

grouping TC(mmol/L) TG(mmol/L) LDLC(mmol/L) HDLC(mmol/L) CK 3.811±0.144d 0.914±0.048d 1.260±0.057e 3.354±0.166a Bacteria 1 4.819±0.198c 1.041±0.080c 1.516±0.084c 3.529±0.123a Bacteria 2 4.790±0.226c 1.052±0.059c 1.479±0.083cd 3.510±0.160a Bacteria 3 4.823±0.251c 1.073±0.057c 1.457±0.102d 3.479±0.158a LP 5.385±0.253b 1.234±0.094b 1.872±0.124b 2.743±0.139b HF 6299±0257a 1303±0076a 2381±0157a 2193±0104c

Table 11 Blood lipid content of mice in each group after 9 weeks of intragastric administration of dialisteria turbidum (Fig. 8)

grouping TC(mmol/L) TG(mmol/L) LDLC(mmol/L) HDLC(mmol/L) CK 3.892±0.154d 0.983±0.047c 1.250±0.036d 3.376±0.202a Bacteria 4 4.896±0.230c 1.012±0.072c 1.395±0.082c 3.497±0.093a Bacteria 5 4.906±0.247c 1.014±0.054c 1.353±0.076c 3.439±0.160a Bacteria 6 4.917±0.201c 1.069±0.070c 1.364±0.071c 3.421±0.195a LP 5.258±0.302b 1.216±0.088b 1.831±0.131b 2.741±0.100b HF 6.390±0.290a 1.295±0.064a 2.357±0.126a 2.153±0.105c

Table 12 Blood lipid content of mice in each group after 9 weeks of gavage with combined bacteria (Fig. 9)

grouping TC(mmol/L) TG(mmol/L) LDLC(mmol/L) HDLC(mmol/L) CK 3.902±0.111d 0.964±0.046d 1.213±0.060d 3.287±0.239a Bacteria 1+5 4.822±0.139c 1.019±0.067c 1.369±0.077c 3.324±0.177a Bacteria 2+6 4.904±0.182c 1.017±0.061c 1.367±0.077c 3.249±0.215a Bacteria 3+4 4.834±0.192c 1.009±0.060c 1.381±0.082c 3.252±0.140a LP 5.498±0.280b 1.213±0.061b 1.742±0.105b 2.621±0.106b HF 6.511±0.252a 1.304±0.094a 2.357±0.166a 2.196±0.096c

The results are shown in Figure 7-9 and Table 10-12. The main components of blood lipids are cholesterol and triglycerides, and the elevated levels of cholesterol and triglycerides in plasma are related to the occurrence of atherosclerosis. The results showed that Faecalicoccus concomitensis, Dialisteria opacus or their combination could reduce blood lipids and reduce the related indicators of atherosclerosis-related diseases (such as cardiovascular diseases). In addition, the effect of Faecalicoccus concomitans, Dialisteria opacus or their combination to reduce total cholesterol (TC), total triglyceride (TG) and low density lipoprotein (LDLC), and increase high density lipoprotein (HDLC) Especially obvious (P<0.05).

(4) Effects of Faecalicoccus comorbidis, Dialisteria turbidum or their combination on leptin (Leptin, LEP) and monocyte chemoattractant protein-1 (MCP-1).

Table 13 Contents of leptin (Leptin, LEP) and monocyte chemoattractant protein-1 (MCP-1) in mice in each group after 9 weeks of gavage with Faecalicoccus congeners (Figure 10, Figure 13)

Table 14 Contents of leptin (Leptin, LEP) and monocyte chemoattractant protein-1 (MCP-1) in mice in each group after 9 weeks of intragastric administration of Alisteria turbidum (Figure 11, Figure 14)

grouping MCP-1 (pg/ml) LEP(pg/ml) CK 343.07±44.17c 1162.93±96.41c Bacteria 4 339.99±39.03c 1158.48±130.63c Bacteria 5 338.79±31.89c 1166.22±136.26c Bacteria 6 336.81±38.48c 1158.43±111.51c LP 365.95±48.92b 1264.09±158.29b HF 383.04±45.92a 1401.25±147.52a

Table 15 Contents of leptin (Leptin, LEP) and monocyte chemoattractant protein-1 (MCP-1) in mice in each group after gavage of the combined bacteria for 9 weeks (Figure 12, Figure 15)

grouping MCP-1 (pg/ml) LEP(pg/ml) CK 336.55±38.88c 1164.35±106.96c Bacteria 1+5 335.85±31.27c 1173.25±114.83c Bacteria 2+6 333.48±31.21c 1169.25±132.61c Bacteria 3+4 330.36±35.21c 1176.85±109.40c LP 366.47±24.75b 1259.06±144.03b HF 384.12±44.29a 1401.18±179.66a

The results are shown in Figure 10-15 and Table 13-15. The results showed that Faecalicoccus concomitensis, Dialisteria turbidum or their combination could significantly reduce the levels of leptin (LEP) and monocyte chemoattractant protein-1 (MCP-1) in the serum of obese model mice (P <0.05).

The results showed that Faecalicoccus concomitensis, Dialisteria opacus or their combination could improve leptin resistance and increase the sensitivity to leptin (LEP) in vivo; Serum MCP-1 level decreased after combined bacteria treatment, which was beneficial to improve insulin resistance and reduce the risk of atherosclerosis and cardiovascular disease.

All documents mentioned herein are incorporated by reference in this application as if each document were individually incorporated by reference. In addition, it should be understood that after reading the above teaching content of the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms also fall within the scope defined by the appended claims of the present application.

Claims (12)

1. the purposes of a Firmicutes probiotic, it is characterised in that, for the preparation of compositions or preparations, the compositions or preparations are used for one or more purposes selected from the group consisting of: (a) prevention and and/or (b) preventing or treating cardiovascular disease, wherein the Firmicutes probiotic is Dialister invisus, and the cardiovascular disease is selected from the group consisting of: Atherosclerosis and hyperlipidemia. 2. The use of claim 1, wherein the composition or formulation further comprises Coprococcus eutactus. 3. The use of claim 2, wherein the Coprococcus eutactus is selected from the group consisting of Coprococcus eutactus ATCC 27759, Coprococcus eutactus ATCC 51897, Coprococcus eutactus L2-50, or a combination thereof. 4. The use of claim 1, wherein the composition or formulation is also independently or additionally used for one or more uses selected from the group consisting of: (i) inhibiting body weight gain in mammals; (ii) reducing body fat ratio in mammals; (iii) reducing blood lipid levels in mammals. 5. The use of claim 4, wherein said reducing the blood lipid level of the mammal comprises: (i) increasing the level of high-density lipoprotein (HDLC) in mammals; (ii) reducing low density lipoprotein (LDLC) levels in a mammal. 6. A composition for treating and/or preventing obesity, the composition comprising: (i) a safe and effective amount of Firmicutes probiotics; and (ii) a pharmaceutically acceptable carrier; wherein the The Firmicutes probiotic is a combination of Coprococcus eutactus and Dialister invisus. 7. The composition of claim 6, wherein the composition contains 1×10-1×10 ²⁰ cfu/mL or cfu/g of Firmicutes probiotics, according to the composition Total volume or total weight. 8. The composition of claim 7, wherein the composition contains 1×10 ⁴ -1×10 ¹⁵ cfu/mL or cfu/g Firmicutes probiotics, according to the composition total volume or total weight. 9 . The composition of claim 6 , wherein the composition contains 0.0001-99 wt % of the Firmicutes probiotics, based on the total weight of the composition. 10 . 10 . The composition of claim 9 , wherein the composition contains 0.1-90 wt % of the Firmicutes probiotics, based on the total weight of the composition. 11 . 11. The composition of claim 6, further comprising other probiotics and/or prebiotics. 12. a preparation method of the described composition of claim 6, comprises the step: Mix (i) Firmicutes probiotics with (ii) a pharmaceutically acceptable carrier to form the composition of claim 6; wherein, the Firmicutes probiotics are Coprococcus eutactus) and Dialister invisus.

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