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WO2016049883A1 - 拟杆菌在治疗或预防肥胖相关疾病中的用途 - Google Patents

拟杆菌在治疗或预防肥胖相关疾病中的用途 Download PDF

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Publication number
WO2016049883A1
WO2016049883A1 PCT/CN2014/087994 CN2014087994W WO2016049883A1 WO 2016049883 A1 WO2016049883 A1 WO 2016049883A1 CN 2014087994 W CN2014087994 W CN 2014087994W WO 2016049883 A1 WO2016049883 A1 WO 2016049883A1
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WIPO (PCT)
Prior art keywords
bacteroides
pharmaceutical composition
obesity
cfu
food
Prior art date
Application number
PCT/CN2014/087994
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English (en)
French (fr)
Inventor
宁光
洪洁
王卫庆
刘瑞欣
王计秋
Original Assignee
上海交通大学医学院附属瑞金医院
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Application filed by 上海交通大学医学院附属瑞金医院 filed Critical 上海交通大学医学院附属瑞金医院
Priority to US15/515,343 priority Critical patent/US10350248B2/en
Priority to PCT/CN2014/087994 priority patent/WO2016049883A1/zh
Priority to EP14903005.8A priority patent/EP3202891B1/en
Priority to CN201480082378.9A priority patent/CN107002023B/zh
Publication of WO2016049883A1 publication Critical patent/WO2016049883A1/zh

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/66—Microorganisms or materials therefrom
    • A61K35/74—Bacteria
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K20/00—Accessory food factors for animal feeding-stuffs
    • A23K20/10—Organic substances
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/66—Microorganisms or materials therefrom
    • A61K35/74—Bacteria
    • A61K35/741—Probiotics
    • A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
    • A61K35/747—Lactobacilli, e.g. L. acidophilus or L. brevis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0053—Mouth and digestive tract, i.e. intraoral and peroral administration
    • A61K9/0056—Mouth soluble or dispersible forms; Suckable, eatable, chewable coherent forms; Forms rapidly disintegrating in the mouth; Lozenges; Lollipops; Bite capsules; Baked products; Baits or other oral forms for animals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/04—Anorexiants; Antiobesity agents
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales

Definitions

  • the present invention relates to the field of microbiology, in particular to the use of Bacteroides for the preparation of medicaments, and to pharmaceutical compositions, medicaments, foods comprising Bacteroides and their use in the treatment or prevention of obesity-related diseases.
  • Obesity is a chronic disease, and many factors can lead to obesity. The origin of the disease has not been studied so far. Obesity is also an inducing factor for a range of diseases such as hypertension, diabetes, coronary heart disease, gallbladder disease, osteoarthritis, respiratory apnea, respiratory disorders, uterine tumors, prostate cancer, breast cancer and colon cancer. According to NHI, about 97 million Americans are currently overweight and obese, with about 15.1 million people with type 2 diabetes associated with obesity, and about 200,000 people die each year from obesity-related diseases.
  • Obesity is usually a body fat excess due to changes in physiological or biochemical functions. Fat usually includes neutral lipids, phospholipids and cholesterol. The increase in fat is due to the energy intake being greater than the energy consumed.
  • simple obesity can be divided into idiopamic obesity and acquired obesity, and the number of simple obesity patients can account for more than 95% of 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 adulthood obesity.
  • Secondary obesity also known as symptomatic obesity, is usually caused by endocrine or metabolic diseases.
  • Medication is currently the main method of clinical treatment of obesity and its obesity-related diseases such as diabetes.
  • the mechanisms of drug treatment include appetite suppression, increased energy expenditure, stimulation of fat movement, reduction of triglyceride synthesis, and inhibition of fat absorption.
  • the main drugs are: phenylpropanolamine (PPA), orlistat (Xenical III) and sibutramine (ReductilTM).
  • PPA phenylpropanolamine
  • orlistat Xenical III
  • ReductilTM sibutramine
  • Hyperglycemia in certain diabetics is still not properly controlled by diet and/or exercise therapy or using the above therapeutic compounds.
  • exogenous insulin should be used.
  • the use of exogenous insulin is a very expensive and painful method and can cause multiple complications to the patient. For example, an incorrect insulin dose calculation can result in an insulin response (hypoglycemia) due to lack of eating or abnormal exercise.
  • local or systemic allergy or immune resistance to the drug may occur with the drug.
  • an object of the present invention is to propose the use of Bacteroides in the treatment or prevention of obesity-related diseases.
  • the invention provides the use of Bacteroides in the preparation of a medicament for the treatment or prevention of obesity-related diseases.
  • the inventors have found that by providing the drug to an animal, it is possible to effectively reduce the body weight and blood sugar of the animal, and thus can be used for treating or preventing obesity and diseases caused by obesity, such as diabetes.
  • the Bacteroides is Bacteroides intestinalis.
  • the animal's body weight and blood sugar can be effectively reduced, and it can be effectively used for treating or preventing obesity and diseases caused by obesity.
  • the Bacteroides is at least one selected from the group consisting of Bacteroides intestinalis DSM 17393, Bacteroides intestinalis JCM 13266, and Bacteroides intestinalis AM-1.
  • the effect of reducing animal body weight and blood sugar is remarkable.
  • the obesity-related disease is selected from the group consisting of obesity or diabetes.
  • the medicament is for reducing body weight or blood sugar of an animal.
  • the animal is a mammal, preferably a human, a murine and a rabbit.
  • the invention provides a pharmaceutical composition.
  • the pharmaceutical composition comprises: Bacteroides and/or a metabolite thereof; and a pharmaceutically acceptable excipient.
  • the inventors have surprisingly found that by providing the animal with the pharmaceutical composition of the present invention, it is possible to effectively reduce the body weight and blood sugar of the animal, thereby effectively treating or preventing obesity and diseases caused by obesity.
  • the Bacteroides is Bacteroides intestinalis.
  • the animal's body weight and blood sugar can be effectively reduced, and it can be effectively used for treating or preventing obesity and diseases caused by obesity.
  • the Bacteroides is at least one selected from the group consisting of Bacteroides intestinalis DSM 17393, Bacteroides intestinalis JCM 13266, and Bacteroides intestinalis AM-1.
  • the effect of reducing animal body weight and blood sugar is remarkable.
  • the pharmaceutical composition is in a solid state, and the pharmaceutical composition comprises 1 ⁇ 10-1 ⁇ 10 20 cfu/g of the Bacteroides and/or a metabolite thereof, or the drug
  • the composition is in a liquid state, and the pharmaceutical composition comprises 1 x 10-1 x 10 20 cfu/mL of the Bacteroides and/or its metabolites. Therefore, the effect of reducing animal body weight and blood sugar is ideal. If the content of Bacteroides is too low, the effect of reducing body weight and blood sugar is not satisfactory. If the content of Bacteroides is too high, the effect of reducing body weight and blood sugar is not significantly improved, resulting in waste. .
  • 1 ⁇ 10-1 ⁇ 10 20 cfu/g or 1 ⁇ 10-1 ⁇ 10 20 cfu/mL of the Bacteroides and/or its metabolites used herein means at a concentration of 1 ⁇ 10-1 ⁇ 10 20 cfu / g or 1 ⁇ 10-1 ⁇ 10 20 cfu / mL bacteria and / or concentration of the metabolite produced by bacteria.
  • the pharmaceutical composition is solid and the pharmaceutical composition comprises not less than 1 ⁇ 10 6 cfu / g of the Bacteroides and / or metabolites thereof, preferably 1 ⁇ 10 11 cfu g of the above and / Bacteroides and/or its metabolites, or the pharmaceutical composition is in a liquid state, and the pharmaceutical composition comprises not less than 1 ⁇ 10 6 cfu/mL of the Bacteroides and/or its metabolites, preferably 1 ⁇ 10 11 cfu/mL and above of the Bacteroides and/or its metabolites.
  • the effect of reducing animal body weight and blood sugar is better.
  • the pharmaceutically acceptable excipient is selected from the group consisting of a carrier, an excipient, a diluent, a lubricant, a wetting agent, an emulsifier, a suspension stabilizer, a preservative, a sweetener, and a fragrance. At least one of them.
  • the pharmaceutically acceptable adjuvant is selected from the group consisting of lactose, glucose, sucrose, sorbitol, mannose, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, fine At least one of crystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, and mineral oil.
  • the pharmaceutical composition further comprises: at least one of Lactobacillus bergii, Lactobacillus rhamnosus GM-020, and Lactobacillus rhamnosus PL60.
  • the invention provides a medicament.
  • the medicament comprises: the pharmaceutical composition described above.
  • the medicament of the present invention can effectively reduce the body weight and blood sugar of an animal, thereby being capable of treating or preventing obesity and obesity-related diseases such as diabetes and the like.
  • the medicament is in the form of at least one preparation selected from the group consisting of granules, capsules, tablets, powders, oral solutions, suspensions and emulsions.
  • administration is easy.
  • the invention provides a food product.
  • the food product comprises: Bacteroides and/or a metabolite thereof; and a food acceptable acceptable adjuvant. The inventors have found that by taking the food of the present invention, the animal's body weight and blood sugar are significantly lowered, thereby being able to treat or prevent obesity and obesity-related diseases.
  • the Bacteroides is Bacteroides intestinalis.
  • the animal's body weight and blood sugar can be effectively reduced, and it can be effectively used for treating or preventing obesity and diseases caused by obesity.
  • the Bacteroides is at least one selected from the group consisting of Bacteroides intestinalis DSM 17393, Bacteroides intestinalis JCM 13266, and Bacteroides intestinalis AM-1.
  • the effect of reducing animal body weight and blood sugar is remarkable.
  • the food product is in a solid state, and the food product comprises 1 ⁇ 10-1 ⁇ 10 20 cfu/g of the Bacteroides and/or a metabolite thereof, or the food product is in a liquid state, and The food product comprises 1 x 10-1 x 10 20 cfu/mL of the Bacteroides and/or its metabolites. Therefore, the effect of reducing animal body weight and blood sugar is ideal. If the content of Bacteroides is too low, the effect of reducing body weight and blood sugar is not satisfactory. If the content of Bacteroides is too high, the effect of reducing body weight and blood sugar is not significantly improved, resulting in waste. .
  • the food product is in a solid state, and the food product comprises not less than 1 ⁇ 10 6 cfu / g of the Bacteroides and / or its metabolites, preferably 1 ⁇ 10 11 cfu / g and above
  • the Bacteroides and/or its metabolite, or the food product is in a liquid state, and the food product comprises not less than 1 ⁇ 10 6 cfu/mL of the Bacteroides and/or its metabolites, preferably 1 ⁇ 10 11% fufu/mL and above of the Bacteroides and/or its metabolites.
  • the effect of reducing animal body weight and blood sugar is better.
  • the food acceptable excipient is selected from the group consisting of a carrier, an excipient, a diluent, a lubricant, a wetting agent, an emulsifier, a suspension stabilizer, a preservative, a sweetener, and At least one of the spices.
  • the food acceptable excipient is selected from the group consisting of lactose, glucose, sucrose, sorbitol, mannose, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, At least one of fine crystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, and mineral oil.
  • the food product further comprises: at least one of Lactobacillus bergii, Lactobacillus rhamnosus GM-020, and Lactobacillus rhamnosus PL60.
  • the food product is in the form of at least one preparation selected from the group consisting of solids, dairy products, solution preparations, powder preparations and suspension preparations. Therefore, it is convenient to take.
  • the invention provides a feed.
  • the feed comprises Bacteroides and/or its metabolites. Providing the animal with the feed of the present invention can effectively reduce the body weight and blood sugar of the animal.
  • the Bacteroides is Bacteroides intestinalis.
  • the animal's body weight and blood sugar can be effectively reduced, and it can be effectively used for treating or preventing obesity and diseases caused by obesity.
  • the Bacteroides is at least one selected from the group consisting of Bacteroides intestinalis DSM 17393, Bacteroides intestinalis JCM 13266, and Bacteroides intestinalis AM-1.
  • the effect of reducing animal body weight and blood sugar is remarkable.
  • the feed is solid, and the feed comprises 1 ⁇ 10-1 ⁇ 10 20 cfu/g of the Bacteroides and/or a metabolite thereof, or the feed is in a liquid state, and The feed comprises 1 x 10-1 x 10 20 cfu/mL of the Bacteroides and/or its metabolites. Therefore, the effect of reducing animal body weight and blood sugar is ideal. If the content of Bacteroides is too low, the effect of reducing body weight and blood sugar is not satisfactory. If the content of Bacteroides is too high, the effect of reducing body weight and blood sugar is not significantly improved, resulting in waste. .
  • the feed is solid, and the feed comprises not less than 1 ⁇ 10 6 cfu / g of the Bacteroides and / or its metabolites, preferably 1 ⁇ 10 11 cfu / g and above
  • the Bacteroides and/or its metabolite, or the feed is in a liquid state, and the feed comprises not less than 1 ⁇ 10 6 cfu/mL of the Bacteroides and/or its metabolites, preferably 1 ⁇ 10 11% fufu/mL and above of the Bacteroides and/or its metabolites.
  • the effect of reducing animal body weight and blood sugar is better.
  • the present invention provides the use of the aforementioned pharmaceutical composition, the aforementioned medicament, and the aforementioned foodstuff for treating or preventing obesity-related diseases.
  • the inventors have found that by providing the animal with the pharmaceutical composition described above, the aforementioned drug, and the aforementioned food, it is possible to effectively reduce the body weight and blood sugar of the animal, thereby enabling the treatment or prevention of obesity and obesity-related diseases. .
  • the obesity-related disease is obesity or diabetes.
  • the invention provides a method of treating or preventing a disease associated with obesity.
  • the method comprises: providing the animal with the pharmaceutical composition described above, the preceding drug or the aforementioned food product.
  • Providing the animal with the pharmaceutical composition described above, the aforementioned drug, and the aforementioned food can effectively reduce the body weight and blood sugar of the animal, thereby effectively treating or preventing the effects of obesity and obesity-related diseases.
  • FIG. 1 shows experimental mouse grouping information and processing steps in accordance with one embodiment of the present invention
  • Figure 2 shows the trend of weight gain of three groups of mice fed within 10 weeks according to one embodiment of the present invention
  • Figure 3 shows a comparison of the P value of the difference in body weight gain of three groups of mice fed within 10 weeks according to one embodiment of the present invention
  • Figure 4 is a graph showing the change in blood glucose of a mouse at the 10th week of feeding according to one embodiment of the present invention
  • FIG. 5 shows blood glucose tolerance in mice fed week 10 (comparison of blood glucose values 120 minutes after feeding glucose solution) in accordance with one embodiment of the present invention.
  • Bacteroides has the following physiological characteristics: rod-shaped, the size of the cells is about 0.8 ⁇ m ⁇ 1 ⁇ 5.0 ⁇ m, no spores, no movement, scattered. It is capable of fermenting several carbohydrates to produce acid, including glucose, lactose, sucrose, maltose, xylose, arabinose, cellobiose, mannose, raffinose and rhamnose.
  • Culture conditions Modified PYG medium (DSMZ Medium 104), anaerobic culture at 37 ° C for 1-2 days.
  • Bacteroides has an effect of treating or preventing obesity-related diseases, and the active composition containing Bacteroides is fed to a test subject and found to be capable of inhibiting body weight. Increase, lower blood sugar, and effectively reduce diabetes and obesity.
  • the invention provides the use of Bacteroides for the preparation of a medicament for the treatment or prevention of obesity-related diseases.
  • the inventors have found that by providing the drug to an animal, it is possible to effectively reduce the body weight and blood sugar of the animal, and thus can be used for treating or preventing obesity and diseases caused by obesity, such as diabetes.
  • the Bacteroides is Bacteroides intestinalis.
  • the animal's body weight and blood sugar can be effectively reduced, and it can be effectively used for treating or preventing obesity and diseases caused by obesity.
  • the Bacteroides is at least one selected from the group consisting of Bacteroides intestinalis DSM 17393 (DSM 17393), Bacteroides intestinalis JCM 13266 (JCM 13266), and Bacteroides intestinalis AM-1 (AM-1).
  • DSM 17393 Bacteroides intestinalis DSM 17393
  • JCM 13266 Bacteroides intestinalis JCM 13266
  • AM-1 Bacteroides intestinalis AM-1
  • the obesity-related disease is selected from the group consisting of obesity or diabetes.
  • the Bacteroides is used to reduce animal body weight or blood glucose.
  • the animal is a mammal, preferably a human, a murine and a rabbit.
  • the subject ingests a high fat food, and strains DSM 17393, JCM 13266, and AM-1 each have the ability to inhibit the subject's weight gain.
  • strains DSM 17393, JCM 13266, and AM-1 each have the ability to inhibit the subject's weight gain.
  • CF57BL/6J mice fed with high fat foods which can cause obesity which were treated with strains DSM 17393, JCM 13266 and AM-1, respectively, compared with the untreated control group, It maintains no significant increase in body weight and contributes to a drop in blood sugar.
  • the strain can be used to treat or prevent obesity and diseases caused by obesity, such as diabetes.
  • the invention provides a pharmaceutical composition.
  • the pharmaceutical composition comprises: Bacteroides and/or a metabolite thereof; and a pharmaceutically acceptable excipient.
  • the inventors have surprisingly found that by providing the animal with the pharmaceutical composition of the present invention, it is possible to effectively reduce the body weight and blood sugar of the animal, thereby effectively treating or preventing obesity and diseases caused by obesity.
  • the Bacteroides is Bacteroides intestinalis.
  • the animal's body weight and blood sugar can be effectively reduced, and it can be effectively used for treating or preventing obesity and diseases caused by obesity.
  • the Bacteroides is at least one selected from the group consisting of Bacteroides intestinalis DSM 17393, Bacteroides intestinalis JCM 13266, and Bacteroides intestinalis AM-1.
  • the effect of reducing animal body weight and blood sugar is remarkable.
  • the pharmaceutical composition is in a solid state, and the pharmaceutical composition comprises 1 ⁇ 10 - 1 ⁇ 10 20 cfu / g of the Bacteroides and / or a metabolite thereof, or the drug combination
  • the substance is in a liquid state, and the pharmaceutical composition comprises 1 x 10-1 x 10 20 cfu/mL of the Bacteroides and/or its metabolite. Therefore, the effect of reducing animal body weight and blood sugar is ideal. If the content of Bacteroides is too low, the effect of reducing body weight and blood sugar is not satisfactory. If the content of Bacteroides is too high, the effect of reducing body weight and blood sugar is not significantly improved, resulting in waste. .
  • the pharmaceutical composition is in a solid state, and the pharmaceutical composition comprises not less than 1 ⁇ 10 6 cfu / g of the Bacteroides and / or its metabolite, preferably 1 ⁇ 10 11 cfu / g and above of the Bacteroides and / or its metabolites, or the pharmaceutical composition is in a liquid state, and the pharmaceutical composition comprises not less than 1 ⁇ 10 6 cfu / mL of the Bacteroides and / or The metabolite thereof is preferably 1 ⁇ 10 11 cfu/mL or more of the Bacteroides and/or its metabolite.
  • the effect of reducing animal body weight and blood sugar is better.
  • the pharmaceutically acceptable excipient is selected from the group consisting of a carrier, an excipient, a diluent, a lubricant, a wetting agent, an emulsifier, a suspension stabilizer, a preservative, a sweetener, and a fragrance. At least one of them.
  • the pharmaceutically acceptable adjuvant is selected from the group consisting of lactose, glucose, sucrose, sorbitol, mannose, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, fine At least one of crystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, and mineral oil.
  • the pharmaceutical composition further comprises: at least one of Lactobacillus bergii, Lactobacillus rhamnosus GM-020, and Lactobacillus rhamnosus PL60.
  • the invention provides a medicament.
  • the medicament comprises: the pharmaceutical composition described above.
  • the medicament of the present invention can effectively reduce the body weight and blood sugar of an animal, thereby being capable of treating or preventing obesity and obesity-related diseases such as diabetes and the like.
  • the medicament is in the form of at least one preparation selected from the group consisting of granules, capsules, tablets, powders, oral solutions, suspensions and emulsions.
  • administration is easy.
  • the medicament of the present invention may also be administered in the form of any one of a pharmaceutical tablet, an injection or a capsule, and the medicament may include an excipient, a pharmaceutically acceptable medium, and a carrier, which may be administered according to The drug route is chosen.
  • the medicament of the present invention can be produced in a casing formulation by various known methods, so that the active ingredient of the drug, that is, the microorganism (Bacteroides) can smoothly pass through the stomach without being destroyed by gastric acid.
  • the active ingredient of the drug that is, the microorganism (Bacteroides)
  • the microorganism of the invention may be used in the form of a capsule prepared by conventional methods.
  • a standard excipient is mixed with the cold-dried microorganism of the present invention to prepare a pellet, and then the pellet is filled into a gelatin capsule.
  • the microorganism of the present invention may be mixed with an excipient which is allowed to be used by a drug such as liquid glue, cellulose, silicate or mineral oil to prepare a suspension or dispersion, and the suspension or dispersion may be filled into soft gelatin.
  • a drug such as liquid glue, cellulose, silicate or mineral oil
  • the medicament of the invention may be formulated into enteric coated tablets for oral use.
  • Terms in this article "Casing” includes all coatings which are permitted for use with conventional drugs. These coatings are not degraded by gastric acid, but are sufficiently decomposed in the small intestine and rapidly release the microorganism of the present invention.
  • the casing of the present invention is coated at about 16-30 mg per tablet, preferably about 16-25 mg per tablet, more preferably about 16-20 mg per tablet.
  • the thickness of the casing is 5 to 100 ⁇ m, and the thickness is preferably 20 to 80 ⁇ m.
  • the casing component is selected from conventional polymers known to the public.
  • Preferred casings of the present invention are cellulose acetate phthalate polymers or trimellitate polymers and copolymers of methacrylic acid (for example, containing 40% or more of methacrylic acid and containing methylcellulose phthalic acid) Preparation of a copolymer of hydroxypropyl acrylate or its ester derivative of methacrylic acid).
  • the cellulose acetate phthalate used in the casing of the present invention has a viscosity of about 45 to 90 cp, an acetyl content of 17 to 26%, and a phthalic acid content of 30 to 40%.
  • the cellulose acetate terephthalate used in the casing has a viscosity of about 5-21 cs and an acetamidine content of 17-26%.
  • Cellulose acetate trimellitate is produced by Eastman Kodak Company and can be used as the casing material in the present invention.
  • the hydroxypropyl methylcellulose phthalate used in the casing of the present invention has a molecular weight of generally 20,000 to 130,000 Daltons, an ideal molecular weight of 80,000 to 100,000 Daltons, and a hydroxypropyl content of 5 to 10%.
  • the methoxy group content is 18-24%, and the phthaloyl group content is 21-35%.
  • HP50 Hydroxypropyl methylcellulose phthalic acid used in the casing of the present invention
  • HP50 contains 6-10% hydroxypropyl, 20-24% methoxy, 21-27% propyl and has a molecular weight of 84,000 Daltons.
  • Another casing material is HP55, HP55 contains 5-9% hydroxypropyl methylcellulose phthalate, 18-22% methoxy, 27-35% phthalic acid, and its molecular weight is 78,000 Daltons.
  • the casing of the present invention can be prepared by spraying the casing solution onto the core using a conventional method.
  • All solvents in the enteric coating process are alcohols (e.g., ethanol), ketones (e.g., acetone), halogenated hydrocarbon compounds (e.g., dichloromethane), or combinations thereof.
  • a softening agent such as di-n-butyl phthalate and triacetin is added to the casing solution in a ratio of about 1 part of the laundry plus about 0.05 part or about 0.3 part of the softening agent.
  • the spraying method is preferably carried out continuously, and the amount of the sprayed material can be controlled according to the conditions employed for the coating.
  • the spray pressure can be adjusted at will, and in general, the desired result can be obtained at an average pressure of 1-1.5 bar.
  • the expression "pharmaceutically effective amount” as used herein refers to an amount which can produce function or activity to a human and/or animal and which can be accepted by humans and/or animals.
  • it may be prepared to contain 1 ⁇ 10 -1 ⁇ 10 20 cfu / ml or 1 ⁇ 10 - 1 ⁇ 10 20 cfu / g (particularly, may contain 1 ⁇ 10 4 - 1 ⁇ 10 15 cfu / mL or 1 ⁇ 10 4 -1 ⁇ 10 15 cfu / g; more specifically, may contain 1 ⁇ 10 6 - 1 ⁇ 10 11 cfu / ml or 1 ⁇ 10 6 - 1 ⁇ 10 11 cfu / g) of Bacteroides And/or a pharmaceutical preparation of its metabolite.
  • the effective dose of Bacteroides or its metabolites used may vary depending on the mode of administration and the severity of the condition to be treated.
  • a dosage form suitable for internal administration comprising about 1 x 10-1 x 10 20 cfu/g or 1 x 10-1 x 10 20 cfu/ml in intimate admixture with a solid or liquid pharmaceutically acceptable carrier (particularly Containing 1 ⁇ 10 4 - 1 ⁇ 10 15 cfu / g or 1 ⁇ 10 4 - 1 ⁇ 10 15 cfu / mL; more specifically, may contain 1 ⁇ 10 6 - 1 ⁇ 10 11 cfu / g or 1 ⁇ 10 6 -1 x 10 11 cfu/ml) of active Bacteroides or the active ingredient produced by fermentation.
  • This dosage regimen can be adjusted as needed to provide an optimal therapeutic response. For example, several separate doses may be administered per day, or the dose may be proportionally reduced, as is critical to the condition of the treatment.
  • the Bacteroides or its metabolites of the present invention can be administered by oral or the like.
  • Solid carriers include: starch, lactose, dicalcium phosphate, microcrystalline cellulose, sucrose and kaolin, while liquid carriers include: medium, polyethylene glycol, nonionic surfactants and edible oils (such as corn oil, peanut oil). And sesame oil), as long as it is suitable for the characteristics of Bacteroides or its metabolites and the specific mode of administration required.
  • Adjuvants commonly used in the preparation of medicaments may also be advantageously included, such as flavoring agents, coloring agents, preservatives, and antioxidants such as vitamin E, vitamin C, BHT, and BHA.
  • preferred drugs are in solid form, especially tablets and solid filled or liquid filled capsules.
  • Oral administration is the preferred mode of administration.
  • Dosage unit of administration means a dose that is formally separable and suitable for use in humans or all other mammalian individuals.
  • Each unit contains a pharmaceutically acceptable carrier and a therapeutically effective amount of a microorganism of the invention.
  • the amount administered varies depending on the weight of the patient and the severity of obesity, the supplementary active ingredient included, and the microorganism used. Further, if possible, it can be administered separately and can be administered continuously if necessary. Therefore, the amount administered does not limit the invention.
  • the invention provides a food product.
  • the food product comprises: Bacteroides and/or a metabolite thereof; and a food acceptable acceptable adjuvant. The inventors have found that by taking the food of the present invention, the animal's body weight and blood sugar are significantly lowered, thereby being able to treat or prevent obesity and obesity-related diseases.
  • the Bacteroides is Bacteroides intestinalis.
  • the animal's body weight and blood sugar can be effectively reduced, and it can be effectively used for treating or preventing obesity and diseases caused by obesity.
  • the Bacteroides is at least one selected from the group consisting of Bacteroides intestinalis DSM 17393, Bacteroides intestinalis JCM 13266, and Bacteroides intestinalis AM-1.
  • the effect of reducing animal body weight and blood sugar is remarkable.
  • the food product is in a solid state, and the food product comprises 1 ⁇ 10-1 ⁇ 10 20 cfu/g of the Bacteroides and/or a metabolite thereof, or the food product is in a liquid state, and The food product comprises 1 x 10-1 x 10 20 cfu/mL of the Bacteroides and/or its metabolites. Therefore, the effect of reducing animal body weight and blood sugar is ideal. If the content of Bacteroides is too low, the effect of reducing body weight and blood sugar is not satisfactory. If the content of Bacteroides is too high, the effect of reducing body weight and blood sugar is not significantly improved, resulting in waste. .
  • the food product is in a solid state, and the food product comprises not less than 1 ⁇ 10 6 cfu / g of the Bacteroides and / or its metabolites, preferably 1 ⁇ 10 11 cfu / g and above
  • the Bacteroides and/or its metabolite, or the food product is in a liquid state, and the food product comprises not less than 1 ⁇ 10 6 cfu/mL of the Bacteroides and/or its metabolites, preferably 1 ⁇ 10 11% fufu/mL and above of the Bacteroides and/or its metabolites.
  • the effect of reducing animal body weight and blood sugar is better.
  • the food acceptable excipient is selected from the group consisting of a carrier, an excipient, a diluent, a lubricant, a wetting agent, an emulsifier, a suspension stabilizer, a preservative, a sweetener, and At least one of the spices.
  • the food acceptable excipient is selected from the group consisting of lactose, glucose, sucrose, sorbitol, mannose, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, At least one of fine crystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, and mineral oil.
  • the food product further comprises: at least one of Lactobacillus bergii, Lactobacillus rhamnosus GM-020, and Lactobacillus rhamnosus PL60.
  • the food product is in the form of at least one preparation selected from the group consisting of solids, dairy products, solution preparations, powder preparations and suspension preparations. Therefore, it is convenient to take.
  • the invention provides a feed.
  • the feed comprises Bacteroides and/or its metabolites. Providing the animal with the feed of the present invention can effectively reduce the body weight and blood sugar of the animal.
  • the Bacteroides is Bacteroides intestinalis.
  • the animal's body weight and blood sugar can be effectively reduced, and it can be effectively used for treating or preventing obesity and diseases caused by obesity.
  • the Bacteroides is at least one selected from the group consisting of Bacteroides intestinalis DSM 17393, Bacteroides intestinalis JCM 13266, and Bacteroides intestinalis AM-1.
  • the effect of reducing animal body weight and blood sugar is remarkable.
  • the feed is solid, and the feed comprises 1 ⁇ 10-1 ⁇ 10 20 cfu/g of the Bacteroides and/or a metabolite thereof, or the feed is in a liquid state, and The feed comprises 1 x 10-1 x 10 20 cfu/mL of the Bacteroides and/or its metabolites. Therefore, the effect of reducing animal body weight and blood sugar is ideal. If the content of Bacteroides is too low, the effect of reducing body weight and blood sugar is not satisfactory. If the content of Bacteroides is too high, the effect of reducing body weight and blood sugar is not significantly improved, resulting in waste. .
  • the feed is solid, and the feed comprises not less than 1 ⁇ 10 6 cfu / g of the Bacteroides and / or its metabolites, preferably 1 ⁇ 10 11 cfu / g and above
  • the Bacteroides and/or its metabolite, or the feed is in a liquid state, and the feed comprises not less than 1 ⁇ 10 6 cfu/mL of the Bacteroides and/or its metabolites, preferably 1 ⁇ 10 11% fufu/mL and above of the Bacteroides and/or its metabolites.
  • the effect of reducing animal body weight and blood sugar is better.
  • the present invention provides the use of the aforementioned pharmaceutical composition, the aforementioned medicament, and the aforementioned foodstuff for treating or preventing obesity-related diseases.
  • the inventors have found that by providing the animal with the pharmaceutical composition described above, the aforementioned drug, and the aforementioned food, it is possible to effectively reduce the body weight and blood sugar of the animal, thereby enabling the treatment or prevention of obesity and obesity-related diseases. .
  • the obesity-related disease is obesity or diabetes.
  • the invention provides a method of treating or preventing a disease associated with obesity.
  • the method comprises: providing the animal with the pharmaceutical composition described above, the preceding drug or the aforementioned food product.
  • Providing the animal with the pharmaceutical composition described above, the aforementioned drug, and the aforementioned food can effectively reduce the body weight and blood sugar of the animal, thereby effectively treating or preventing the effects of obesity and obesity-related diseases.
  • mice Mouse C57BL/6J (purchased from the Experimental Animal Center of Southern Medical University), which is a normal-fed mouse, 6 weeks old, a total of 25 mice, the growth process of the mice is in the same environment, and the same food is fed.
  • mice were randomly divided into 5 groups of 5 each.
  • mice Six weeks old C57BL/6J mice, which were normally fed (without sterility), were selected for the experiment.
  • the mice were tested for insulin tolerance by the glucose tolerance (OGGT) test by introducing the Bacteroides DSM 17393, JCM 13266 and AM-1 to be tested in the intestine of the mice by feeding method.
  • OGGT glucose tolerance
  • the specific experimental grouping information and processing steps are shown in Figure 1. The experimental steps are as follows:
  • mice Twenty-five mice were housed in the same environment to 6 weeks of age (food without disinfection), the amount of feed was recorded and the body weight changes and physical condition of the mice were monitored regularly.
  • mice Twenty-five mice were divided into five experimental groups (5 replicates per experimental group), which were placed in the ultra-clean workbench and officially entered the experimental period. The first feeding change treatment was performed according to the experimental design.
  • Feeding method adopts gavage method, feeding timing quantitative, unified for all mice standard.
  • the amount of the cultured bacteria was set to 0.15 mL/10 g 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, and the activation should be fresh every week, and the concentration should be determined separately.
  • mice During the experimental period, the body weight, body length, and food intake of the mice were recorded weekly.
  • the blood glucose level of the mice was measured by a blood glucose meter, and the symptoms and time points of diabetes in each group were recorded.
  • Method for measuring blood glucose tolerance Fasting blood glucose was measured by starvation for 10 hours, and postprandial blood glucose was measured 60 minutes and 120 minutes after feeding the glucose solution.
  • feeding the test bacteria can significantly reduce the blood sugar of obese mice. From the results of Fig. 5, feeding test bacteria can significantly improve the blood glucose tolerance of obese mice. Thus, it was shown that Bacteroides can be effectively used to reduce body weight and blood sugar.
  • a pharmaceutical composition containing Bacteroides intestinalis DSM 17393, Bacteroides intestinalis JCM 13266 or Bacteroides intestinalis AM-1, respectively, was prepared as follows:
  • the raw material ratio of the pharmaceutical composition is shown in Table 4.
  • lactose, yeast powder and peptone were mixed uniformly with purified water, preheated to 60-65 ° C, homogenized under pressure of 20 Mpa, then sterilized at 90 ° C for 20-30 minutes, then cooled to 36.
  • 0.5 g of the lyophilized product was weighed and mixed with maltodextrin in an equal amount, and then placed in a capsule to prepare a capsule containing Bacteroides intestinalis DSM 17393 or Bacteroides intestinalis JCM 13266 or Bacteroides intestinalis AM-1.
  • Seed tank (GYS-500C);
  • the live bacterial dry powder preparations of Bacteroides intestinalis DSM 17393, Bacteroides intestinalis JCM 13266, and Bacteroides intestinalis AM-1 are prepared separately, and the specific steps are as follows:
  • the above-mentioned activated strains were inoculated into fresh modified PYG medium at a ratio of 3%, and expanded culture was carried out for 24 hours at 37 ° C, 160 r / min shaker to obtain expanded cultured strains.
  • the above-mentioned expanded culture strain is inoculated into each fermenting dish according to the inoculation amount of 1-5%, and the strain and the solid fermentation medium are thoroughly stirred and mixed, naturally flattened, and then fermented at 37 ° C. Hours in order to obtain fermented wheat bran.
  • the fermented wheat bran obtained above is placed in a dry box and dried at a constant temperature of 60 ° C for 12-16 hours. Then, the dried fermented wheat bran is pulverized by a 100 mesh sieve to obtain a dry powder preparation of live bacteria, which is stored in a bag. Among them, some samples were taken for live bacterial count, and the live bacterial powder preparations with a live bacterial count of 1 billion to 20 billion were selected for use.
  • a dry powder preparation of live bacteria obtained by obtaining Bacteroides intestinalis DSM 17393, Bacteroides intestinalis JCM 13266, and Bacteroides intestinalis AM-1 was separately prepared to obtain a feed additive.
  • the above-mentioned feed additives were added to the feed ingredients of the animal feed in an amount of 0.1%, and then subjected to extrusion granulation by a granulator to obtain Bacteroides intestinalis DSM 17393, Bacteroides intestinalis JCM 13266, and Bacteroides intestinalis AM-, respectively. 1 feed additive feed.

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Abstract

提供了拟杆菌(Bacteroides)在制备药物中的用途，所述药物用于治疗或预防肥胖相关疾病，还提供了包含拟杆菌的药物组合物、药物、食品、饲料及其在治疗或预防肥胖相关疾病中的用途，以及治疗或预防肥胖相关疾病的方法。

Description

拟杆菌在治疗或预防肥胖相关疾病中的用途 技术领域
本发明涉及微生物学领域，具体地，涉及拟杆菌(Bacteroides)在制备药物中的用途，也涉及包含拟杆菌的药物组合物、药物、食品及其在治疗或预防肥胖相关疾病中的用途。
背景技术
肥胖症是一种慢性疾病，许多因素都会导致肥胖，其发病起源至今未研究清楚。肥胖同时也是一系列疾病的诱导因子，如高血压、糖尿病、冠心病、胆囊病、骨关节炎、睡眠中呼吸窒息、呼吸失调、子宫瘤、前列腺癌、乳腺癌和结肠癌等。根据NHI的报告，目前约有9700万美国人超重和过度肥胖，其中与过度肥胖相关的II型糖尿病人数达到约1510万人，每年约有20万人死于与肥胖症相关的疾病。
肥胖通常是由于生理或生物化学功能改变而导致的体脂肪过剩。脂肪通常包括中性脂、磷脂和胆固醇。脂肪的增加是由于能量摄入大于能量的消耗。从发病原理上讲，肥胖有两种类型：(a)单纯性肥胖(simple obesity)和(b)继发性肥胖(second obesity)。单纯性肥胖可划分为先天性肥胖(idiopamic obesity)和后天性肥胖(acquired obesity)，单纯性肥胖患者数量可占95％以上的总肥胖数目。先天性肥胖是由大量的脂肪细胞所致，且常见于儿童期肥胖。后天性肥胖是由更大尺寸的脂肪细胞所致，且常见于成年期肥胖。继发性肥胖又被称为症状性肥胖，其通常是由内分泌或新陈代谢疾病所致。
目前有五种治疗肥胖的策略：节食、锻炼、行为治疗、药物治疗和康复手术(therapetltic operation)。采取哪种策略主要视病人的健康危险因子和体重减轻的速度和效果而定，可选择或组合这些策略对肥胖病人进行治疗。其体重减轻的速度和效果受诸如年龄、身高、家族史和危险因子等多个因素的影响。饮食—锻炼疗法，即吃低热量、低脂肪食物和进行有氧锻炼，但这种方法一般认为对普通大众不成功，且需要长期经常性地坚持；去除体内脂肪手术能达到立竿见影的效果，但存在诸多限制，如手术风险、除脂效果难以持久和花费过于昂贵等。
药物治疗是目前主要的临床治疗肥胖及其肥胖相关疾病(如糖尿病)的方法。药物治疗的机制包括抑制食欲、增加能量消耗、刺激脂肪移动、降低甘油三酯合成和抑制脂肪吸收。目前主要的药物为：苯丙醇胺(phenylpropanolamine，PPA)、罗氏鲜(orlistat，XenicalⅢ)和诺美婷(sibutramine，ReductilTM)。某些糖尿病人的高血糖通过饮食和/或锻炼疗法或使用上述治疗化合物仍然不能得到适当控制。对于这些病人，应使用外源胰岛素。对病人来说， 使用外源胰岛素是很昂贵和痛苦的方法，还会给病人带来多种并发症。例如，由于没有吃饭或不正常锻炼，胰岛素剂量的计算错误会导致胰岛素响应(低血糖)。此外，使用药物还可能发生对药物的局部或全身过敏或免疫抗性。
目前本领域还没有一种有效的、副作用小的治疗或预防肥胖及其相关疾病的方法和药物，因此本领域迫切需要开发一种的新的，无毒副作用的，用于治疗或预防肥胖及其相关疾病的药物。
发明内容
本发明旨在至少在一定程度上解决相关技术中的技术问题之一。为此，本发明的一个目的在于提出拟杆菌在治疗或预防肥胖相关疾病中的用途。
在本发明的第一方面，本发明提供了拟杆菌在制备药物中的用途，所属药物用于治疗或预防肥胖相关疾病。发明人发现，通过给动物提供该药物，能够有效降低动物的体重和血糖，进而能够用于治疗或预防肥胖及由肥胖引起的疾病，如糖尿病等。
根据本发明的实施例，所述拟杆菌为Bacteroides intestinalis。由此，能够有效降低动物的体重和血糖，进而能够有效用于治疗或预防肥胖及由肥胖引起的疾病。
根据本发明的实施例，所述拟杆菌为选自Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266和Bacteroides intestinalis AM-1中的至少一种。由此，降低动物体重和血糖的效果显著。
根据本发明的实施例，所述肥胖相关疾病为选自肥胖症或糖尿病。
根据本发明的实施例，所述药物用于降低动物体重或血糖。
根据本发明的实施例，所述动物为哺乳动物，优选人、鼠类和兔类。
在本发明的第二方面，本发明提供了一种药物组合物。根据本发明的实施例，该药物组合物包括：拟杆菌和/或其代谢产物；以及药学上可接受的辅料。发明人惊喜地发现，通过给动物提供本发明的药物组合物，能够有效降低动物的体重和血糖，进而能够有效治疗或预防肥胖及由肥胖引起的疾病。
根据本发明的实施例，所述拟杆菌为Bacteroides intestinalis。由此，能够有效降低动物的体重和血糖，进而能够有效用于治疗或预防肥胖及由肥胖引起的疾病。
根据本发明的实施例，所述拟杆菌为选自Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266和Bacteroides intestinalis AM-1中的至少一种。由此，降低动物体重和血糖的效果显著。
根据本发明的实施例，所述药物组合物呈固态，并且所述药物组合物包括1×10-1×1020cfu/g的所述拟杆菌和/或其代谢产物，或者，所述药物组合物呈液态，并且所述 药物组合物包括1×10-1×1020cfu/mL的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果理想，如果拟杆菌含量过低，降低动物体重和血糖的效果不理想，如果拟杆菌含量过高，则降低动物体重和血糖的效果无明显提高，造成浪费。
需要说明的是，本文中所使用的表达方式“1×10-1×1020cfu/g或1×10-1×1020cfu/mL的所述拟杆菌和/或其代谢产物”是指浓度为1×10-1×1020cfu/g或1×10-1×1020cfu/mL的细菌和/或该浓度的细菌所产生的代谢产物。
所述药物组合物呈固态，并且所述药物组合物包括不低于1×106cfu/g的所述拟杆菌和/或其代谢产物，优选1×1011cfu/g及以上的所述拟杆菌和/或其代谢产物，或者所述药物组合物呈液态，并且所述药物组合物包括不低于1×106cfu/mL的所述拟杆菌和/或其代谢产物，优选1×1011cfu/mL及以上的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果较佳。
根据本发明的实施例，所述药学上可接受的辅料为选自载体、赋形剂、稀释剂、润滑剂、润湿剂、乳化剂、悬浮液稳定剂、防腐剂、甜味剂以及香料中的至少一种。
根据本发明的实施例，所述药学上可接受的辅料为选自乳糖、葡萄糖、蔗糖、山梨糖醇、甘露糖、淀粉、阿拉伯胶、磷酸钙、藻酸盐、明胶、硅酸钙、细结晶纤维素、聚乙烯吡咯烷酮、纤维素、水、糖浆、甲基纤维素、羟基苯甲酸甲酯、羟基苯甲酸丙酯、滑石、硬脂酸镁和矿物油中的至少一种。
根据本发明的实施例，所述药物组合物进一步包括：加氏乳杆菌，鼠李糖乳杆菌GM-020和鼠李糖乳杆菌PL60中的至少一种。由此，有利于提高降低动物体重和血糖的效果。
在本发明的第三方面，本发明提供了一种药物。根据本发明的实施例，所述药物包括：前面所述的药物组合物。本发明的药物能够有效降低动物的体重和血糖，进而能够治疗或预防肥胖和肥胖相关疾病，如糖尿病等。
根据本发明的实施例，所述药物呈选自颗粒剂、胶囊、片剂、粉末剂、口服液、混悬液和乳剂中的至少一种制剂形式。由此，易于给药。
在本发明的第四方面，本发明提供了一种食品。根据本发明的实施例，所述食品包括：拟杆菌和/或其代谢产物；以及食品学上可接受的辅料。发明人发现，服用本发明的该食品，动物的体重和血糖明显降低，进而能够治疗或预防肥胖和肥胖相关疾病。
根据本发明的实施例，所述拟杆菌为Bacteroides intestinalis。由此，能够有效降低动物的体重和血糖，进而能够有效用于治疗或预防肥胖及由肥胖引起的疾病。
根据本发明的实施例，所述拟杆菌为选自Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266和Bacteroides intestinalis AM-1中的至少一种。由此，降低动物体重和血糖的效果显著。
根据本发明的实施例，所述食品呈固态，并且所述食品包括1×10-1×1020cfu/g的所述拟杆菌和/或其代谢产物，或者所述食品呈液态，并且所述食品包括1×10-1×1020cfu/mL的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果理想，如果拟杆菌含量过低，降低动物体重和血糖的效果不理想，如果拟杆菌含量过高，则降低动物体重和血糖的效果无明显提高，造成浪费。
根据本发明的实施例，所述食品呈固态，并且所述食品包括不低于1×106cfu/g的所述拟杆菌和/或其代谢产物，优选1×1011cfu/g及以上的所述拟杆菌和/或其代谢产物，或者所述食品呈液态，并且所述食品包括不低于1×106cfu/mL的所述拟杆菌和/或其代谢产物，优选1×1011cfu/mL及以上的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果较佳。
根据本发明的实施例，所述食品学上可接受的辅料为选自载体、赋形剂、稀释剂、润滑剂、润湿剂、乳化剂、悬浮液稳定剂、防腐剂、甜味剂以及香料中的至少一种。
根据本发明的实施例，所述食品学上可接受的辅料为选自乳糖、葡萄糖、蔗糖、山梨糖醇、甘露糖、淀粉、阿拉伯胶、磷酸钙、藻酸盐、明胶、硅酸钙、细结晶纤维素、聚乙烯吡咯烷酮、纤维素、水、糖浆、甲基纤维素、羟基苯甲酸甲酯、羟基苯甲酸丙酯、滑石、硬脂酸镁和矿物油中的至少一种。
根据本发明的实施例，所述食品进一步包括：加氏乳杆菌，鼠李糖乳杆菌GM-020和鼠李糖乳杆菌PL60中的至少一种。由此，有利于提高降低动物体重和血糖的效果。
根据本发明的实施例，所述食品呈选自固体、乳品、溶液制品、粉末制品和悬浮液制品中的至少一种制剂形式。由此，服用方便。
在本发明的第五方面，本发明提供了一种饲料。根据本发明的实施例，该饲料包括拟杆菌和/或其代谢产物。对动物提供本发明的饲料，可以有效降低动物的体重和血糖。
根据本发明的实施例，所述拟杆菌为Bacteroides intestinalis。由此，能够有效降低动物的体重和血糖，进而能够有效用于治疗或预防肥胖及由肥胖引起的疾病。
根据本发明的实施例，所述拟杆菌为选自Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266和Bacteroides intestinalis AM-1中的至少一种。由此，降低动物体重和血糖的效果显著。
根据本发明的实施例，所述饲料呈固态，并且所述饲料包括1×10-1×1020cfu/g的所述拟杆菌和/或其代谢产物，或者所述饲料呈液态，并且所述饲料包括1×10-1×1020cfu/mL的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果理想，如果拟杆菌含量过低，降低动物体重和血糖的效果不理想，如果拟杆菌含量过高，则降低动物体重和血糖的效果无明显提高，造成浪费。
根据本发明的实施例，所述饲料呈固态，并且所述饲料包括不低于1×106cfu/g的所述拟杆菌和/或其代谢产物，优选1×1011cfu/g及以上的所述拟杆菌和/或其代谢产物，或者所述饲料呈液态，并且所述饲料包括不低于1×106cfu/mL的所述拟杆菌和/或其代谢产物，优选1×1011cfu/mL及以上的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果较佳。
在本发明的第六方面，本发明提供了前面所述的药物组合物、前面所述的药物、前面所述的食品在治疗或预防肥胖相关疾病中的用途。发明人发现，通过对动物提供前面所述的药物组合物、前面所述的药物、前面所述的食品，能够有效降低动物的体重和血糖，进而能够发挥治疗或预防肥胖和肥胖相关疾病的功效。
根据本发明的实施例，所述肥胖相关疾病为肥胖症或糖尿病。
在本发明的第七方面，本发明提供了一种治疗或预防肥胖相关疾病的方法。根据本发明的实施例，该方法包括：对动物提供前面所述的药物组合物、前面的药物或前面所述的食品。对动物提供前面所述的药物组合物、前面所述的药物、前面所述的食品，能够有效降低动物的体重和血糖，进而能够有效治疗或预防肥胖和肥胖相关疾病的功效。
应理解，在本发明范围内中，本发明的上述各技术特征和在下文(如实施例)中具体描述的各技术特征之间都可以互相组合，从而构成新的或优选的技术方案。限于篇幅，在此不再一一累述。
附图说明
图1显示了根据本发明一个实施例，实验小鼠分组信息和处理步骤；
图2显示了根据本发明一个实施例，饲喂10周内3组小鼠体重增加趋势；
图3显示了根据本发明一个实施例，饲喂10周内3组小鼠体重增长差异P值比较结果；
图4显示了根据本发明一个实施例，饲喂第10周小鼠血糖变化曲线；以及
图5显示了根据本发明一个实施例，饲喂第10周鼠血糖耐受情况(喂食葡萄糖溶液后120分钟血糖值比较)。
具体实施方式
下面结合具体实施例，进一步阐述本发明。应理解，这些实施例仅用于说明本发明而不用于限制本发明的范围。下列实施例中未注明具体条件的实验方法，通常按照常规条件如Sambrook等人，分子克隆：实验室手册(New York：Cold Spring Harbor Laboratory Press，1989)中所述的条件，或按照《微生物：实验手册》(James Cappuccino和Natalie Sherman编，Pearson Education出版社)中所述的条件，或按照制造厂商所建议的条件。
拟杆菌具有如下生理特性：杆状，菌体大小约0.8μm×1～5.0μm，不产芽孢，不运动，分散存在。能够发酵几种碳水化合物产酸，包括葡萄糖、乳糖、蔗糖、麦芽糖、木糖、阿拉伯糖、纤维二糖、甘露糖、棉子糖和鼠李糖。培养条件：改良PYG培养基中(DSMZ Medium104)，37℃厌氧培养1-2天。发明人经过广泛而深入的研究和实验，意外地发现，拟杆菌(Bacteroides)具有治疗或预防肥胖相关疾病的作用，将含有拟杆菌的活性组合物饲喂食实验对象，发现该组合物能够抑制体重增加，降低血糖，有效减轻糖尿病及肥胖等病症。
因而，在本发明的第一方面，本发明提供了拟杆菌在制备药物中的用途，所述药物用于治疗或预防肥胖相关疾病。发明人发现，通过给动物提供该药物，能够有效降低动物的体重和血糖，进而能够用于治疗或预防肥胖及由肥胖引起的疾病，如糖尿病等。
根据本发明的实施例，所述拟杆菌为Bacteroides intestinalis。由此，能够有效降低动物的体重和血糖，进而能够有效用于治疗或预防肥胖及由肥胖引起的疾病。
根据本发明的实施例，所述拟杆菌为选自Bacteroides intestinalis DSM 17393(DSM 17393)、Bacteroides intestinalis JCM 13266(JCM 13266)和Bacteroides intestinalis AM-1(AM-1)中的至少一种。由此，降低动物体重和血糖的效果显著。
根据本发明的实施例，所述肥胖相关疾病为选自肥胖症或糖尿病。
根据本发明的实施例，所述拟杆菌用于降低动物体重或血糖。
根据本发明的实施例，所述动物为哺乳动物，优选人、鼠类和兔类。
根据本发明的实施例，受试者摄入高脂肪的食物，菌株DSM 17393、JCM 13266和AM-1均具有抑制该受验者体重增加的能力。根据本发明的实施例，分别经菌株DSM 17393、JCM 13266和AM-1来治疗的被喂以可导致肥胖的高脂肪的食物的CF57BL/6J小鼠，与未接受治疗的对照组相比，其可保持体重无明显增加，且有助于血糖下降。由此，表明所述菌株能够用以治疗或预防肥胖及由肥胖引起的疾病，如糖尿病等。
在本发明的第二方面，本发明提供了一种药物组合物。根据本发明的实施例，该药物组合物包括：拟杆菌和/或其代谢产物；以及药学上可接受的辅料。发明人惊喜地发现，通过给动物提供本发明的药物组合物，能够有效降低动物的体重和血糖，进而能够有效治疗或预防肥胖及由肥胖引起的疾病。
根据本发明的实施例，所述拟杆菌为Bacteroides intestinalis。由此，能够有效降低动物的体重和血糖，进而能够有效用于治疗或预防肥胖及由肥胖引起的疾病。
根据本发明的实施例，所述拟杆菌为选自Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266和Bacteroides intestinalis AM-1中的至少一种。由此，降低动物体重和血糖的效果显著。
根据本发明的实施例，所述药物组合物呈固态，并且所述药物组合物包括 1×10-1×1020cfu/g的所述拟杆菌和/或其代谢产物，或者所述药物组合物呈液态，并且所述药物组合物包括1×10-1×1020cfu/mL的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果理想，如果拟杆菌含量过低，降低动物体重和血糖的效果不理想，如果拟杆菌含量过高，则降低动物体重和血糖的效果无明显提高，造成浪费。
根据本发明的实施例，所述药物组合物呈固态，并且所述药物组合物包括不低于1×106cfu/g的所述拟杆菌和/或其代谢产物，优选1×1011cfu/g及以上的所述拟杆菌和/或其代谢产物，或者所述药物组合物呈液态，并且所述药物组合物包括不低于1×106cfu/mL的所述拟杆菌和/或其代谢产物，优选1×1011cfu/mL及以上的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果较佳。
根据本发明的实施例，所述药学上可接受的辅料为选自载体、赋形剂、稀释剂、润滑剂、润湿剂、乳化剂、悬浮液稳定剂、防腐剂、甜味剂以及香料中的至少一种。
根据本发明的实施例，所述药学上可接受的辅料为选自乳糖、葡萄糖、蔗糖、山梨糖醇、甘露糖、淀粉、阿拉伯胶、磷酸钙、藻酸盐、明胶、硅酸钙、细结晶纤维素、聚乙烯吡咯烷酮、纤维素、水、糖浆、甲基纤维素、羟基苯甲酸甲酯、羟基苯甲酸丙酯、滑石、硬脂酸镁和矿物油中的至少一种。
根据本发明的实施例，所述药物组合物进一步包括：加氏乳杆菌，鼠李糖乳杆菌GM-020和鼠李糖乳杆菌PL60中的至少一种。由此，有利于提高降低动物体重和血糖的效果。
在本发明的第三方面，本发明提供了一种药物。根据本发明的实施例，所述药物包括：前面所述的药物组合物。本发明的药物能够有效降低动物的体重和血糖，进而能够治疗或预防肥胖和肥胖相关疾病，如糖尿病等。
根据本发明的实施例，所述药物呈选自颗粒剂、胶囊、片剂、粉末剂、口服液、混悬液和乳剂中的至少一种制剂形式。由此，易于给药。
根据本发明的实施例，本发明的药物也可以以药物片剂，针剂或胶囊的任一种形式给药，所述药物可以包括赋形剂、药物允许的媒介和载体，这些物质可根据给药途径进行选择。
根据本发明的一些实施例，本发明的药物可通过多种公知的方法以肠衣制剂生产，以便于药物的活性成分即微生物(拟杆菌)能顺利通过胃而不被胃酸所破坏。
根据本发明的另一些实施例，本发明的微生物可以常规方法制备的胶囊形式使用。例如，标准赋形剂和本发明的冷干微生物混合制成小球药丸，然后将药丸装填入明胶胶囊中。此外，本发明的微生物可以和药物允许使用的赋形剂如液体胶、纤维素、硅酸盐或矿物油等混合制作悬浮液或分散液，这种悬浮液或分散液可装入软的明胶胶囊中。
根据本发明的另一些实施例，本发明的药物可制成肠衣片供口服使用。本文中的术语 “肠衣”，包括所有常规药物允许使用的包衣，这些包衣不被胃酸降解，但在小肠中能充分分解并快速释放出本发明的微生物。木发明的肠衣能在合成胃酸如pH＝1的HCl溶液中在36-38℃条件下维持2小时以上，并优选在合成肠液如pH＝7.0的缓冲液中在1.0小时内分解。
本发明的肠衣为以每片约16-30mg进行包衣，较佳地每片约16-25mg，更佳地每片约16-20mg进行包衣。本发明中肠衣厚度为5-100μm，理想的厚度为20-80μm。肠衣成分选自己公开知晓的常规聚合物。本发明优选的肠衣由纤维素乙酸邻苯二甲酸酯聚合物或偏苯三酸酯聚合物以及异丁烯酸的共聚物(例如，含有40％以上异丁烯酸和含有甲基纤维素邻苯二甲酸羟丙酯或其酯类衍生物的异丁烯酸的共聚物)制备。
本发明中肠衣所使用的纤维素乙酸邻苯二甲酸酯的粘度为约45-90cp，乙酰含量17-26％，邻苯二甲酸含量30-40％。用于肠衣中的纤维素乙酸偏苯二酸酯粘度为约5-21cs，乙酞含量17-26％。纤维素乙酸偏苯三酸酯由Eastman科达公司生产，可用作本发明中的肠衣材料。
用于本发明肠衣中的羟丙基甲基纤维素邻苯二甲酸酷，分子量一般为20,000-130,000道尔顿，理想分子量为80,000-100,000道尔顿，羟丙基含量为5-10％，甲氧基含量为18-24％，邻苯二甲酰基含量为21-35％。
用于本发明肠衣中的羟丙基甲基纤维素邻苯二甲酸酷为HP50，由日本Shin-Etsu Chemidnl Co.Ltd.生产。HP50含有6-10％羟丙基，20-24％甲氧基，21-27％的丙基，其分子量为84,000道尔顿。另一种肠衣物质为HP55，HP55含有5-9％的羟丙基甲基纤维素邻苯二甲酸酯，18-22％甲氧基，27-35％的邻苯二甲酸，其分子量为78,000道尔顿。
本发明肠衣可以通过如下步骤进行制备：使用常规方法将肠衣溶液喷雾到核心上。该肠包衣方法中所有溶剂为醇类(如乙醇)、酮类(如丙酮)、卤代烃化合物(如二氯甲烷)、或其组合物。将软化剂如二-正丁基邻苯二甲酸酯和三乙酸甘油酯加入到肠衣溶液中，其比例为1份包衣物加约0.05份或约0.3份软化剂。喷雾方法优选连续执行，所喷雾的料量可根据包衣所采用的条件进行控制。喷雾压力可随意调节，一般而言，能在平均1-1.5巴压力下获得理想的结果。
需要说明的是，本文中所使用的表达方式“药物有效量”是指可对人和/或动物产生功能或活性的且可被人和/或动物所接受的量。比如，在本发明中，可制备含有1×10-1×1020cfu/ml或1×10-1×1020cfu/g(特别的，可含有1×104-1×1015cfu/mL或1×104-1×1015cfu/g；更特别地，可含有1×106-1×1011cfu/ml或1×106-1×1011cfu/g)的拟杆菌和/或其代谢产物的药物制剂。
当用于制备药物时，所用的拟杆菌或其代谢产物的有效剂量可随施用的模式和待治疗的疾病的严重程度而变化。适用于内服的剂量形式，包含与固态或液态药学上可接受的载体密切混合的约1×10-1×1020cfu/g或1×10-1×1020cfu/ml(特别的，可含有1×104-1×1015cfu/g 或1×104-1×1015cfu/mL；更特别地，可含有1×106-1×1011cfu/g或1×106-1×1011cfu/ml)的活性拟杆菌或发酵产生的活性成分。根据需要，可调节此剂量方案以提供最佳治疗应答。例如，由治疗状况的迫切要求，可每天给予若干次分开的剂量，或将剂量按比例地减少。
本发明所述的拟杆菌或其代谢产物可通过口服等途径给予。固态载体包括：淀粉、乳糖、磷酸二钙、微晶纤维素、蔗糖和白陶土，而液态载体包括：培养基、聚乙二醇、非离子型表面活性剂和食用油(如玉米油、花生油和芝麻油)，只要适合拟杆菌或其代谢产物特性和所需的特定给药方式。在制备药物中通常使用的佐剂也可有利地被包括，例如调味剂、色素、防腐剂和抗氧化剂如维生素E、维生素C、BHT和BHA。
从易于制备和给药的立场看，优选的药物呈固态形式，尤其是片剂和固体填充或液体填充的胶囊。口服给药是优选的给药方式。
将本发明的药物施用给所述个体，可以每天给药1次或多次。给药剂量单位表示其形式上能分开且适用于人类或其他所有哺乳动物个体的剂量。每一单位含有药物允许的载体和有效治疗量的本发明微生物。给药量随病人的体重和肥胖严重程度、所包括的补充活性组份和所使用的微生物而变化。此外如可能，可分开给药，并且如需要可连续给药。因此，所述给药量不会对本发明造成限制。
在本发明的第四方面，本发明提供了一种食品。根据本发明的实施例，所述食品包括：拟杆菌和/或其代谢产物；以及食品学上可接受的辅料。发明人发现，服用本发明的该食品，动物的体重和血糖明显降低，进而能够治疗或预防肥胖和肥胖相关疾病。
根据本发明的实施例，所述拟杆菌为Bacteroides intestinalis。由此，能够有效降低动物的体重和血糖，进而能够有效用于治疗或预防肥胖及由肥胖引起的疾病。
根据本发明的实施例，所述拟杆菌为选自Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266和Bacteroides intestinalis AM-1中的至少一种。由此，降低动物体重和血糖的效果显著。
根据本发明的实施例，所述食品呈固态，并且所述食品包括1×10-1×1020cfu/g的所述拟杆菌和/或其代谢产物，或者所述食品呈液态，并且所述食品包括1×10-1×1020cfu/mL的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果理想，如果拟杆菌含量过低，降低动物体重和血糖的效果不理想，如果拟杆菌含量过高，则降低动物体重和血糖的效果无明显提高，造成浪费。
根据本发明的实施例，所述食品呈固态，并且所述食品包括不低于1×106cfu/g的所述拟杆菌和/或其代谢产物，优选1×1011cfu/g及以上的所述拟杆菌和/或其代谢产物，或者所述食品呈液态，并且所述食品包括不低于1×106cfu/mL的所述拟杆菌和/或其代谢产物，优选1×1011cfu/mL及以上的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果 较佳。
根据本发明的实施例，所述食品学上可接受的辅料为选自载体、赋形剂、稀释剂、润滑剂、润湿剂、乳化剂、悬浮液稳定剂、防腐剂、甜味剂以及香料中的至少一种。
根据本发明的实施例，所述食品学上可接受的辅料为选自乳糖、葡萄糖、蔗糖、山梨糖醇、甘露糖、淀粉、阿拉伯胶、磷酸钙、藻酸盐、明胶、硅酸钙、细结晶纤维素、聚乙烯吡咯烷酮、纤维素、水、糖浆、甲基纤维素、羟基苯甲酸甲酯、羟基苯甲酸丙酯、滑石、硬脂酸镁和矿物油中的至少一种。
根据本发明的实施例，所述食品进一步包括：加氏乳杆菌，鼠李糖乳杆菌GM-020和鼠李糖乳杆菌PL60中的至少一种。由此，有利于提高降低动物体重和血糖的效果。
根据本发明的实施例，所述食品呈选自固体、乳品、溶液制品、粉末制品和悬浮液制品中的至少一种制剂形式。由此，服用方便。
在本发明的第五方面，本发明提供了一种饲料。根据本发明的实施例，该饲料包括拟杆菌和/或其代谢产物。对动物提供本发明的饲料，可以有效降低动物的体重和血糖。
根据本发明的实施例，所述拟杆菌为Bacteroides intestinalis。由此，能够有效降低动物的体重和血糖，进而能够有效用于治疗或预防肥胖及由肥胖引起的疾病。
根据本发明的实施例，所述拟杆菌为选自Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266和Bacteroides intestinalis AM-1中的至少一种。由此，降低动物体重和血糖的效果显著。
根据本发明的实施例，所述饲料呈固态，并且所述饲料包括1×10-1×1020cfu/g的所述拟杆菌和/或其代谢产物，或者所述饲料呈液态，并且所述饲料包括1×10-1×1020cfu/mL的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果理想，如果拟杆菌含量过低，降低动物体重和血糖的效果不理想，如果拟杆菌含量过高，则降低动物体重和血糖的效果无明显提高，造成浪费。
根据本发明的实施例，所述饲料呈固态，并且所述饲料包括不低于1×106cfu/g的所述拟杆菌和/或其代谢产物，优选1×1011cfu/g及以上的所述拟杆菌和/或其代谢产物，或者所述饲料呈液态，并且所述饲料包括不低于1×106cfu/mL的所述拟杆菌和/或其代谢产物，优选1×1011cfu/mL及以上的所述拟杆菌和/或其代谢产物。由此，降低动物体重和血糖的效果较佳。
在本发明的第六方面，本发明提供了前面所述的药物组合物、前面所述的药物、前面所述的食品在治疗或预防肥胖相关疾病中的用途。发明人发现，通过对动物提供前面所述的药物组合物、前面所述的药物、前面所述的食品，能够有效降低动物的体重和血糖，进而能够发挥治疗或预防肥胖和肥胖相关疾病的功效。
根据本发明的实施例，所述肥胖相关疾病为肥胖症或糖尿病。
在本发明的第七方面，本发明提供了一种治疗或预防肥胖相关疾病的方法。根据本发明的实施例，该方法包括：对动物提供前面所述的药物组合物、前面的药物或前面所述的食品。对动物提供前面所述的药物组合物、前面所述的药物、前面所述的食品，能够有效降低动物的体重和血糖，进而能够有效治疗或预防肥胖和肥胖相关疾病的功效。
实施例1
1、实验材料
小鼠：小鼠C57BL/6J(购自南方医科大学实验动物中心)，为正常饲养小鼠，6周龄，一共25只，小鼠生长过程处于同一环境，且喂养同样的食物。
菌株：Bacteroides intestinalis DSM 17393，购自DSMZ(Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH)；Bacteroides intestinalis JCM 13266，购自JCM(Japan Collection of Microorganisms)；Bacteroides intestinalis AM-1，来自日本北海道大学农业学院微生物生理实验室(Laboratory of Microbial Physiology,Research Faculty of Agriculture,Hokkaido University)。
高脂肪食物(HF)：含58％猪油、25.6％的碳水化合物、16.4％的蛋白，购自北京华阜康生物科技股份有限公司。
常规老鼠食物：购自南方医科大学动物实验中心。
2、小鼠分组和诱导处理
本实施例共设置5个实验组，将小鼠随机分为5组，每组各5只。
选取正常喂养(无需无菌)的6周龄C57BL/6J小鼠进行实验。通过喂食法为小鼠肠道分别引入待检验的拟杆菌DSM 17393、JCM 13266和AM-1，通过葡萄糖耐量(OGGT)测试，鉴定小鼠的胰岛素耐受能力。具体实验分组信息和处理步骤见图1。实验步骤具体如下：
将25只小鼠置于同一环境正常饲养至6周龄(食物无需消毒)，记录喂食量并定期监控小鼠体重变化与身体状况。
25只小鼠均分为5个实验组(每实验组5只重复)，分别置于超净工作台中，正式进入实验期。根据实验设计进行第一次喂食改变处理。
在实验期中小鼠必须生活在对应的培养箱中，不可对换任意培养箱中的小鼠。保持培养箱中的清洁等问题。喂食方法采取灌喂(gavage)法，喂食定时定量，对所有小鼠统一 标准。
灌喂菌量定为0.15mL/10g体重，菌浓度为1×107cfu/mL，浓缩后浓度为1×108cfu/mL，频率为隔天一次。菌液需提前培养，每周活化保证新鲜，并分别测定浓度。
实验期中，每周记录小鼠体重、体长、进食量等数据，用血糖仪检测小鼠血糖含量，记录各组小鼠出现糖尿病症状及时间点。血糖耐受能力测定方法：先饥饿10小时测定空腹血糖，再于喂食葡萄糖溶液后60分钟和120分钟测定餐后血糖。
3、实验结果
(1)饲喂10周，各组小鼠体重增长结果见表1和图2。
表1
Figure PCTCN2014087994-appb-000001
(2)饲喂10周，各组小鼠体重增长差异显著性分析结果见图3。
由表1和图2、图3的结果可知，饲喂10周后，饲喂检验菌能显著降低肥胖鼠体重。
(3)饲喂第10周，各组小鼠血糖含量测定结果见表2、图4和图5。
表2
Figure PCTCN2014087994-appb-000002
由表2和图4的结果可知，饲喂检验菌可以显著降低肥胖鼠的血糖。由图5的结果可知，饲喂检验菌可以显著改善肥胖鼠的血糖耐受能力。由此，表明拟杆菌能够有效用于降低体重和血糖。
实施例2：食品的制备
在本实施例中，制备分别含有拟杆菌Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266和Bacteroides intestinalis AM-1的食品，具体如下：
1、食品的原料配比如表3所示。
表3
Figure PCTCN2014087994-appb-000003
按照表3所示的配方和比例，将牛奶和白糖混合，然后搅拌至完全混匀，预热后，于20Mpa压力下均质，接着于90℃左右杀菌5-10分钟，随后冷却至40-43℃，然后接种拟杆菌，接种数量为：每1g上述混合物接种Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266或Bacteroides intestinalis AM-1菌1-100×106cfu，即得。
实施例3：药物的制备
在本实施例中，制备分别含有拟杆菌Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266或Bacteroides intestinalis AM-1的药物组合物，具体如下：
药物组合物的原料配比见表4。
表4
Figure PCTCN2014087994-appb-000004
按照表4所示的比例将乳糖、酵母粉、蛋白胨以纯净水混合均匀，预热到60-65℃，于 20Mpa压力下均质，然后于90℃左右杀菌20-30分钟，随后冷却至36-38℃，然后接入Bacteroides intestinalis DSM 17393或Bacteroides intestinalis JCM 13266或Bacteroides intestinalis AM-1活菌接种量为(1-50×106cfu/mL)，接着于36-38℃发酵至pH值为6.0，离心后，冷冻干燥至水份含量小于3％，即得Bacteroides intestinalis DSM 17393或Bacteroides intestinalis JCM 13266或Bacteroides intestinalis AM-1菌冷冻干燥物。然后，称取0.5克冷冻干燥物与麦芽糊精等量混合后装入胶囊中，即制成含Bacteroides intestinalis DSM 17393或Bacteroides intestinalis JCM 13266或Bacteroides intestinalis AM-1菌的胶囊。
实施例4：饲料的制备
设备：
1、种子罐(GYS-500C)；
2、发酵罐(GYS-1500C)；
3、发酵培养箱(SPX-1500)；
4、搅拌粉粹机(SWFL82D)；
5、卧式压力蒸汽灭菌器(ZY.01.001)；
6、高速离心喷雾干燥机(LPG1000)；
7、热收缩包装机(SM-4525L)；
8、定量粉剂灌装机(DXDCF2000A)；
9、造粒机
首先，按照固体发酵方法，分别制备Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266、Bacteroides intestinalis AM-1的活菌干粉制剂，具体步骤如下：
1、菌种准备
(1)菌种活化
室温放置菌种甘油冻藏管，待该冻藏管内液体全部融化，在火焰旁边将其打开，并用200微升的移液器将菌种甘油冻藏管内液体吹吸均匀。然后，用移液器从已吹吸均匀的冻藏关内吸取200微升液体，接种至50ml体积的新鲜的改良PYG液体培养基中，并于37℃、160r/min摇床上进行活化培养24h，以便获得经过活化的菌种。
(2)扩大培养
将上述经过活化的菌种按照3％的比例接种到新鲜的改良PYG培养基内，并于37℃、160r/min摇床上进行扩大培养24h，以便获得经过扩大培养的菌种。
2、固体发酵
(1)固体发酵培养基配制及分装
称取适量麦麸，并按照1：1-2的质量比加入水将其搅拌均匀，然后将混合物于115℃ 高温下灭菌15-20分钟，以便获得固体发酵培养基。接着，于操作台上，将不低于70℃的固体发酵培养基分装于经过灭菌的干净发酵盘中，并将固体发酵培养基中聚集成团的麦麸捏散。
(2)接种
将上述经过扩大培养的菌种按照1-5％的接种量，接种到各发酵盘中，并将菌种与固体发酵培养基充分搅拌混匀，自然铺平，然后于37℃下进行发酵48小时，以便获得发酵麦麸。
3、发酵麦麸粉碎
将上述获得的发酵麦麸置于干燥箱中，60℃恒温烘干12-16h，然后，利用100目筛将经过烘干的发酵麦麸进行粉碎，以便获得活菌干粉制剂，装袋保存。其中，留取部分样品进行活菌计数，选取每克活菌数为10亿-200亿的活菌干粉制剂备用。
由此，分别制备获得Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266、Bacteroides intestinalis AM-1的活菌干粉制剂，得到饲料添加剂。最后，将上述饲料添加剂分别以0.1％添加量加入到动物饲料的饲料原料中，再通过造粒机进行挤压造粒，分别制得含Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266、Bacteroides intestinalis AM-1饲料添加剂的饲料。
在本说明书的描述中，参考术语“一个实施例”、“一些实施例”、“示例”、“具体示例”、或“一些示例”等的描述意指结合该实施例或示例描述的具体特征、结构、材料或者特点包含于本发明的至少一个实施例或示例中。在本说明书中，对上述术语的示意性表述不必须针对的是相同的实施例或示例。而且，描述的具体特征、结构、材料或者特点可以在任一个或多个实施例或示例中以合适的方式结合。此外，在不相互矛盾的情况下，本领域的技术人员可以将本说明书中描述的不同实施例或示例以及不同实施例或示例的特征进行结合和组合。
尽管上面已经示出和描述了本发明的实施例，可以理解的是，上述实施例是示例性的，不能理解为对本发明的限制，本领域的普通技术人员在本发明的范围内可以对上述实施例进行变化、修改、替换和变型。

Claims (29)

  1. 拟杆菌在制备药物中的用途，所述药物用于治疗或预防肥胖相关疾病。
  2. 根据权利要求1所述的用途，其特征在于，所述拟杆菌为Bacteroides intestinalis。
  3. 根据权利要求2所述的用途，其特征在于，所述拟杆菌为选自Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266和Bacteroides intestinalis AM-1中的至少一种。
  4. 根据权利要求1所述的用途，其特征在于，所述肥胖相关疾病为肥胖症或糖尿病。
  5. 根据权利要求1所述的用途，其特征在于，所述药物用于降低动物体重或血糖。
  6. 根据权利要求1所述的用途，其特征在于，所述动物为哺乳动物，优选人、鼠类和兔类。
  7. 一种药物组合物，其特征在于，包括：
    拟杆菌和/或其代谢产物；以及
    药学上可接受的辅料。
  8. 根据权利要求7所述的药物组合物，其特征在于，所述拟杆菌为Bacteroides intestinalis。
  9. 根据权利要求8所述的药物组合物，其特征在于，所述拟杆菌为选自Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266和Bacteroides intestinalis AM-1中的至少一种。
  10. 根据权利要求7所述的药物组合物，其特征在于，所述药物组合物呈固态，并且所述药物组合物包括1×10-1×1020cfu/g的所述拟杆菌和/或其代谢产物，或者
    所述药物组合物呈液态，并且所述药物组合物包括1×10-1×1020cfu/mL的所述拟杆菌和/或其代谢产物。
  11. 根据权利要求10所述的药物组合物，其特征在于，所述药物组合物呈固态，并且所述药物组合物包括不低于1×106cfu/g的所述拟杆菌和/或其代谢产物，优选1×1011cfu/g及以上的所述拟杆菌和/或其代谢产物，或者
    所述药物组合物呈液态，并且所述药物组合物包括不低于1×106cfu/mL的所述拟杆菌和/或其代谢产物，优选1×1011cfu/mL及以上的所述拟杆菌和/或其代谢产物。
  12. 根据权利要求7所述的药物组合物，其特征在于，所述药学上可接受的辅料为选自载体、赋形剂、稀释剂、润滑剂、润湿剂、乳化剂、悬浮液稳定剂、防腐剂、甜味剂以及香料中的至少一种。
  13. 根据权利要求7所述的药物组合物，其特征在于，所述药学上可接受的辅料为选自乳糖、葡萄糖、蔗糖、山梨糖醇、甘露糖、淀粉、阿拉伯胶、磷酸钙、藻酸盐、明胶、 硅酸钙、细结晶纤维素、聚乙烯吡咯烷酮、纤维素、水、糖浆、甲基纤维素、羟基苯甲酸甲酯、羟基苯甲酸丙酯、滑石、硬脂酸镁和矿物油中的至少一种。
  14. 根据权利要求7所述的药物组合物，其特征在于，进一步包括：加氏乳杆菌，鼠李糖乳杆菌GM-020和鼠李糖乳杆菌PL60中的至少一种。
  15. 一种药物，其特征在于，包括：
    权利要求7-14中任一项所述的药物组合物。
  16. 根据权利要求15所述的药物，其特征在于，所述药物呈选自颗粒剂、胶囊、片剂、粉末剂、口服液、混悬液和乳剂中的至少一种制剂形式。
  17. 一种食品，其特征在于，包括：
    拟杆菌和/或其代谢产物；以及
    食品学上可接受的辅料。
  18. 根据权利要求17所述的食品，其特征在于，所述拟杆菌为Bacteroides intestinalis。
  19. 根据权利要求18所述的食品，其特征在于，所述拟杆菌为选自Bacteroides intestinalis DSM 17393、Bacteroides intestinalis JCM 13266和Bacteroides intestinalis AM-1中的至少一种。
  20. 根据权利要求17所述的食品，其特征在于，所述食品呈固态，并且所述食品包括1×10-1×1020cfu/g的所述拟杆菌和/或其代谢产物，或者
    所述食品呈液态，并且所述食品包括1×10-1×1020cfu/mL的所述拟杆菌和/或其代谢产物。
  21. 根据权利要求20所述的食品，其特征在于，所述食品呈固态，并且所述食品包括不低于1×106cfu/g的所述拟杆菌和/或其代谢产物，优选1×1011cfu/g及以上的所述拟杆菌和/或其代谢产物，或者
    所述食品呈液态，并且所述食品包括不低于1×106cfu/mL的所述拟杆菌和/或其代谢产物，优选1×1011cfu/mL及以上的所述拟杆菌和/或其代谢产物。
  22. 根据权利要求17所述的食品，其特征在于，所述食品学上可接受的辅料为选自载体、赋形剂、稀释剂、润滑剂、润湿剂、乳化剂、悬浮液稳定剂、防腐剂、甜味剂以及香料中的至少一种。
  23. 根据权利要求17所述的药物组合物，其特征在于，所述食品学上可接受的辅料为选自乳糖、葡萄糖、蔗糖、山梨糖醇、甘露糖、淀粉、阿拉伯胶、磷酸钙、藻酸盐、明胶、硅酸钙、细结晶纤维素、聚乙烯吡咯烷酮、纤维素、水、糖浆、甲基纤维素、羟基苯甲酸甲酯、羟基苯甲酸丙酯、滑石、硬脂酸镁和矿物油中的至少一种。
  24. 根据权利要求17所述的食品，其特征在于，进一步包括：加氏乳杆菌，鼠李糖乳 杆菌GM-020和鼠李糖乳杆菌PL60中的至少一种。
  25. 根据权利要求17所述的食品，其特征在于，所述食品呈选自固体、乳品、溶液制品、粉末制品和悬浮液制品中的至少一种制剂形式。
  26. 一种饲料，其特征在于，包括：拟杆菌和/或其代谢产物。
  27. 权利要求7-14中任一项所述的药物组合物、权利要求15或16所述的药物、权利要求17-25中任一项所述的食品或权利要求26所述的饲料在治疗或预防肥胖相关疾病中的用途。
  28. 根据权利要求27所述的用途，其特征在于，所述肥胖相关疾病为选自肥胖症或糖尿病。
  29. 一种治疗或预防肥胖相关疾病的方法，其特征在于，对动物提供权利要求7-14中任一项所述的药物组合物、权利要求15或16所述的药物、权利要求17-25中任一项所述的食品或权利要求26所述的饲料。
PCT/CN2014/087994 2014-09-30 2014-09-30 拟杆菌在治疗或预防肥胖相关疾病中的用途 WO2016049883A1 (zh)

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EP14903005.8A EP3202891B1 (en) 2014-09-30 2014-09-30 Uses of bacteroides in treatment or prevention of obesity or diabetes
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