KR20190068075A - Lactobacillus pentosus AO21-1 strain with Anti-Obesity Ability and Composition for treatment or improvement or preventing of obesity comprising the same - Google Patents

Abstract

A novel Pediococcus acidilactici AO21-1 strain of the present invention, a strain isolated from the human body has high stability, shows, in vitro testing, activity of inhibiting adipocyte differentiation, and is excellent in activity of ameliorating, preventing or treating obesity with weight loss observed in animal experiments. Therefore, the new strain of the present invention is unlikely to cause side effects, compared to the conventional diet foods or drugs that had a side effect problem, and thus can exhibit a weight loss effect without reducing food intake, thereby capable of being utilized as a pharmaceutical composition for treating or preventing obesity or a food composition for ameliorating or preventing obesity.

Description

Lactobacillus pentosus AO21-1 strain having anti-obesity activity and a composition for improving or treating obesity comprising the same,

The present invention relates to a novel Lactobacillus pentosus AO21-1 strain, a composition for preventing or treating obesity and a composition for improving obesity.

Obesity is becoming a serious chronic disease as humanity develops into a rich society. It is known that obesity is caused by various causes such as hypertrophy or hyperplasia of body fat cells morphologically, which is caused by imbalance of consumption and consumption of calories.

Obesity is not only psychologically disturbing individuals, but also a major cause of increased national medical expenditure, as it is a direct cause of increasing the risk of various adult diseases.

Specifically, obesity is directly associated with increased prevalence of various adult diseases such as type 2 diabetes, hypertension, hyperlipidemia, and cardiovascular disease, and is referred to as metabolic syndrome or insulin resistance syndrome by binding together diseases associated with obesity . They act as a cause of arteriosclerosis and cardiovascular disease.

Obesity is not only the most common malnutrition disorder in western society, but also in Korea, the frequency of obesity is rapidly increasing due to the improvement of diet and westernization of lifestyle by economic development, and the rate of Korean adult obesity is increased by 3% The problem is becoming more and more serious. Therefore, there is a demand for development of therapeutic agents or treatment methods for treating and preventing obesity.

Currently known herbal remedies include Xenical (Roche Pharmaceuticals, Switzerland), Reductil (Evothe, USA) and Exolise (Atopama, France), which are widely used as appetite suppressants, And most obesity drugs are appetite suppressants that suppress appetite by controlling the neurotransmitters associated with the hypothalamus. However, conventional therapeutic agents have side effects such as heart diseases, respiratory diseases, nervous system diseases and the like, and the persistence of their effects is also low, so that it is necessary to develop a more improved therapeutic agent for obesity. Also, currently developed products have satisfactory therapeutic effects There is little cure for cervical cancer, and development of a new treatment for obesity is required.

On the other hand, efforts have been made to reduce the blood cholesterol level by using lactic acid bacteria which have been regarded as safe microorganisms. Lactic acid bacteria are metabolites that use saccharides as an energy source and produce abundant lactic acid. They also produce various organic acids and bacteriocin and other antibacterial substances. However, Amine or ammonia are not produced, so it is a beneficial bacteria that prevents decay, inhibits harmful bacteria, and exhibits physiological activity that is helpful to humans. Among these, the Lactobacillus sp. Strain has been known for a long time as a key member of normal microbial communities in the intestinal tract of the human body and is important for maintaining healthy digestive organs and the vaginal environment, and the US Public Health Service According to the guidelines, all of the Lactobacillus strains deposited at the American Depositary Organization (ATCC) are now classified as "Bio-safty Level 1", which is considered to have no known potential risk of causing diseases in humans or animals have.

Korean Patent Registration No. 10-1471033 discloses a strain of Weissella sp. F22 (accession number: KACC 91867P) which is excellent in the ability to inhibit obesity. In Korean Patent Registration No. 10-0264361, a cholesterol- , And Lactobacillus plantarum PMO08 (KFCC-11028) strain having an activity to be defoamed against six kinds of conjugated bile acids is disclosed.

However, since no lactic acid bacteria-related technology superior in commercial anti-obesity effect has appeared, the inventors of the present invention have been studying probiotics which have no side effects in the body and are excellent in the effect of treating obesity, and newly discovered Lactobacillus pentosus AO21-1 The present inventors completed the present invention by confirming that the strain exhibits high survival and activity in the body of an animal and that the addition of the cells themselves as an effective ingredient of foods and medicines provides an excellent effect of preventing, .

Patent Document 1: Korean Patent Publication No. 10-1471033 Patent Document 2: Korean Patent Publication No. 10-0264361

DISCLOSURE Technical Problem The present invention has been made in order to solve the above problems, and it is an object of the present invention to provide a Lactobacillus pentosus AO21-1 strain (KCCM12144P) having an obesity-suppressing activity.

Another object of the present invention is to provide a probiotics comprising the Lactobacillus pentosus AO21-1 strain ( KCCM12144P ) or a culture thereof.

It is still another object of the present invention to provide a pharmaceutical composition for preventing or treating obesity comprising the Lactobacillus pentosus strain AO21-1 ( KCCM12144P ) or a culture thereof.

It is another object of the present invention to provide a food composition for preventing or ameliorating obesity and a food composition for improving inflammation caused by obesity, which comprises a culture of Lactobacillus pentosus strain AO21-1 ( KCCM12144P ) or a culture thereof .

In order to achieve the above object, the present invention provides Lactobacillus pentosus AO21-1 strain (KCCM12144P) having an obesity-suppressing activity.

The invention provides, probiotics (probiotics) comprising the Lactobacillus pento Versus (Lactobacillus pentosus) AO21-1 strain (KCCM12144P) or culture medium thereof to effect the different purposes.

The present invention to achieve the above another object, Lactobacillus pento Versus (Lactobacillus pentosus) provides AO21-1 strain (KCCM12144P) or pharmaceutical composition for preventing or treating obesity, comprising a culture medium thereof.

The Lactobacillus pentosus AO21-1 strain ( KCCM12144P ) or a culture thereof may be contained in an amount of 0.01 to 50% by weight based on the total weight of the composition.

The composition may be one that inhibits adipocyte differentiation.

The composition may be one which reduces body weight.

The present invention provides a food composition for preventing or ameliorating obesity comprising Lactobacillus pentosus AO21-1 strain ( KCCM12144P ) or a culture thereof to achieve the above-mentioned further object.

In another aspect, the present invention provides a Lactobacillus pento Versus (Lactobacillus pentosus) AO21-1 strain (KCCM12144P) or food composition for improving inflammation caused by obesity comprising a culture medium thereof to effect the further object.

The Lactobacillus pentosus strain AO21-1 according to the present invention is a strain isolated from a human body and has high stability and exhibits an ability to inhibit adipocyte differentiation in a test tube test, Improvement, prevention, or therapeutic activity. Therefore, the novel strain of the present invention has a low possibility of side effects, so that it can show a diet effect without adjusting the amount of food to be consumed unlike the conventional dietary functional foods or medicines, which have side effects. Therefore, a pharmaceutical composition for obesity treatment or prevention, Or a food composition for prevention.

Therefore, the novel strain according to the present invention can be used as a pharmaceutical material useful for preventing, improving and treating obesity.

1 is a diagram showing a series of analysis procedures for selecting strains having an obesity-suppressing activity according to Experimental Example 1 of the present invention.
FIG. 2 is a graph showing lipid accumulation by Oil red O staining of each of 36 strains measured according to Experimental Example 1 of the present invention. FIG.
FIG. 3 is a diagram illustrating a procedure of setting an experimental animal and an experimental group according to Experimental Example 2 of the present invention.
4 is a graph showing a change in body weight in each experimental group according to Experimental Example 2 of the present invention.
FIG. 5 is a graph showing the weight change of each experimental group according to Experimental Example 4 of the present invention measured weekly. FIG.
FIG. 6 is a graph showing an analysis of body weight gain in each experimental group according to Experimental Example 4 of the present invention. The feed intake (g / day), the amount of water (g / day) and the calorie intake (kcal / day) Respectively.

Hereinafter, various aspects and various embodiments of the present invention will be described in more detail.

One aspect of the present invention relates to a Lactobacillus pentosus AO21-1 strain (KCCM12144P) having an obesity-suppressing activity.

Lactobacillus pentosus strain AO21-1 according to the present invention is a strain isolated from a human body and deposited on November 2, 2017 at the Korean Microorganism Conservation Center (KCCM).

The Lactobacillus pentosus AO21-1 strain according to the present invention is a 16S rDNA comprising the nucleotide sequence shown in SEQ ID NO: 1, and the Lactobacillus pentosus AO21-1 strain is purple when stained with Gram- Gram-positive bacilli, and glucose is used as a carbon source, galactose, lactose, maltose, xylose, trehalose, sucrose, raffinose, arabinose and the like.

Also, the Lactobacillus bacteria pento Versus (Lactobacillus pentosus) AO21-1 strains may represent more active in addition to obesity, inflammation suppressing activity by the obesity. Lactobacillus pentosus strain AO21-1 , when ingested via oral administration, inhibits adipocyte differentiation and reduces host body weight by increasing dietary fat excretion while reducing host body dietary fat absorption .

The Lactobacillus pentosus strain AO21-1 of the present invention is excellent in survival ability against gastric acid or bile, and exhibits a high survival ability in the animal body, so that the obesity-suppressing activity can be maintained in the body for a long time.

Another aspect of the present invention relates to probiotics comprising Lactobacillus pentosus strain AO21-1 ( KCCM 12144P ) or a culture thereof.

The Lactobacillus pentosus AO21-1 strain according to the present invention can be used in the form of a dried powder or in a cultured state as it is.

In the present invention, the culture broth is prepared by adding Lactobacillus pentosus strain AO21-1 to a known liquid medium or solid medium capable of supplying nutrients so that the strain AO21-1 can grow and survive in vitro, and the Lactobacillus pentosus strain ) Means a whole medium including a cultured strain obtained by culturing the AO21-1 strain for a certain period of time, its metabolites, extra nutrients, etc., and also includes a culture solution obtained by removing the strain after culturing the strain.

The culture broth may be centrifuged or filtered or concentrated, and this step may be carried out according to the needs of a person skilled in the art.

In the present invention, probiotics refers to live microorganisms, that is, live fungi, which provide beneficial health to the intestinal flora, that is, the health of the host. Generally, probiotics are consumed as part of fermented foods such as yogurt or as dietary supplements.

The Lactobacillus pentosus strain AO21-1 of the present invention is a strain of the genus Lactobacillus known as probiotics and has excellent survival ability against gastric acid and bile and has an obesity activity effect on its own. Therefore, the probiotic bacteria such as probiotics, Can be used.

The Lactobacillus pentosus AO21-1 strain ( KCCM12144P ) or a culture thereof is preferably contained in an amount of 0.01 to 50% by weight based on the total weight of the composition.

The number of viable cells of the Lactobacillus pentosus strain AO21-1 in the composition may be adjusted so as to be contained at a concentration of less than 5 x 10 < ⁷ > CFU, which should be selected according to the patient and the situation, The scope of the present invention is not limited by the number of living cells.

Another aspect of the present invention relates to a pharmaceutical composition for prevention or treatment of obesity comprising a Lactobacillus pentosus strain AO21-1 ( KCCM12144P ) or a culture thereof, and a food composition for preventing or improving obesity.

The Lactobacillus pentosus AO21-1 strain according to the present invention can be used in the form of a dried powder or in a cultured state as it is.

In the present invention, the culture broth is prepared by adding Lactobacillus pentosus strain AO21-1 to a known liquid medium or solid medium capable of supplying nutrients so that the strain AO21-1 can grow and survive in vitro, and the Lactobacillus pentosus strain ) Means a whole medium including a cultured strain obtained by culturing the AO21-1 strain for a certain period of time, its metabolites, extra nutrients, etc., and also includes a culture solution obtained by removing the strain after culturing the strain. The culture medium from which the strain has been removed may be sterilized to include dead cells, filtered or centrifuged to remove cells or centrifuged supernatant.

The culture broth may be centrifuged or filtered or concentrated, and this step may be carried out according to the needs of a person skilled in the art.

The Lactobacillus pento Versus (Lactobacillus pentosus) AO21-1 strain (KCCM12144P) or culture medium thereof has a composition containing 0.01 to 50% by weight, based on the total weight, it may be used immediately, it is concentrated to be used, or concentrated and dried It can be diluted.

Increasing the dietary fat emissions while improving, the treatment or prevention of obesity of the present invention, while proliferation is Lactobacillus pento Versus (Lactobacillus pentosus) AO21-1 strain to inhibit the differentiation of mast cells, and reduces the absorption of dietary fats by the host Is expected to reduce the body weight of the host, and the composition may be used as the liquid itself, or it may be dried and powdered.

Means that the Lactobacillus pentosus strain AO21-1 or the culture thereof contains an amount sufficient to achieve an obesity improvement, treatment or prophylactic efficacy or activity.

Refers to all the actions of inhibiting or delaying the development, spread and recurrence of obesity by administration of the composition, and the treatment means all the actions of improving or improving the symptoms of obesity by administration of the composition.

The food composition may be formulated in the form of a capsule, tablet, powder, granule, liquid, ring, flake, paste, syrup, gel, jelly or bar form, Gum, confectionery, etc., and it is produced in the form of general food. If it is taken, it means bringing about a certain effect on health. However, unlike general medicine, it may occur when taking food for a long time There is no side effect.

The food composition is very useful because it can be ingested routinely. The amount of the Lactobacillus pentosus strain AO21-1 or the culture thereof added in such a food composition can not be uniformly determined depending on the type of the target food. However, the amount of the Lactobacillus pentosus AO21-1 added in the range that does not impair the original taste of the food And is usually in the range of 0.01 to 50% by weight, preferably 0.1 to 20% by weight based on the target food. In the case of food compositions in the form of capsules, tablets, powders, granules, liquids, rings, flakes, pastes, syrups, gels, jellies or bar, the content is usually 0.1 to 100% by weight, preferably 0.5 to 80% Of the total weight of the composition.

The food composition may contain, as an active ingredient, a Lactobacillus pentosus AO21-1 strain or a culture thereof, as well as a component that is ordinarily added in the course of food production, for example, a protein, a carbohydrate, a fat, Nutrients, flavoring agents, and flavoring agents. Examples of the above-mentioned carbohydrates are monosaccharides such as glucose, fructose, and the like; Disaccharides such as maltose, sucrose, oligosaccharides and the like; And polysaccharides such as dextrin, cyclodextrin and the like, and sugar alcohols such as xylitol, sorbitol and erythritol.

As a flavoring agent, a natural flavoring agent [tau Martin, stevia extract (e.g., rebaudioside A, glycyrrhizin, etc.) and synthetic flavor (saccharin, aspartame, etc.) can be used.

For example, when the food composition of the present invention is prepared from a drink and a beverage, it may be added to the Lactobacillus pentosus AO21-1 strain of the present invention or a culture thereof in addition to citric acid, liquid fructose, sugar, glucose, acetic acid, And various plant extracts may be further included.

Also, in a pharmaceutical composition for preventing or treating obesity, which comprises Lactobacillus pentosus strain AO21-1 or a culture thereof as an active ingredient, the number of viable cells of Lactobacillus pentosus strain AO21-1 is 5 x 10 < Less than ⁷ CFU, preferably less than 5 x 10 < ³ > 5 x 10 < ⁷ > CFU, but this should be selected depending on the patient and the circumstances, and the scope of the present invention is not limited by the number of viable bacteria in the composition.

In addition, in the above composition, the culture medium may be, for example, 0.001 mg / kg or more, preferably 0.1 mg / kg or more, more preferably 10 mg / kg or more, still more preferably 100 mg / Preferably 250 mg / kg or more, and most preferably 0.1 g / kg or more. Lactobacillus pento Versus (Lactobacillus pentosus) AO21-1 strain even if it is administered in an excess amount of viable cells in the strain isolated from the human body because there is no side effect to the human body of Lactobacillus pento Versus (Lactobacillus pentosus) AO21-1 strains contained in the composition of the present invention The quantitative upper limit can be selected by a person skilled in the art within a suitable range.

The pharmaceutical composition may be prepared by using pharmaceutically acceptable and physiologically acceptable adjuvants in addition to the above-mentioned active ingredients. Examples of the adjuvants include excipients, disintegrants, sweeteners, binders, coating agents, swelling agents, lubricants, A flavoring agent and the like can be used.

The pharmaceutical composition may be formulated to contain at least one pharmaceutically acceptable carrier in addition to the above-described effective ingredients for administration.

The pharmaceutical form of the pharmaceutical composition may be a granule, a powder, a tablet, a coated tablet, a capsule, a suppository, a liquid, a syrup, a juice, a suspension, an emulsion, a drip agent or an injectable liquid agent. For example, for formulation into tablets or capsules, the active ingredient may be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like. Also, if desired or necessary, suitable binders, lubricants, disintegrants and coloring agents may also be included as a mixture. Suitable binders include, but are not limited to, natural sugars such as starch, gelatin, glucose or beta-lactose, natural and synthetic gums such as corn sweeteners, acacia, tracker candles or sodium oleate, sodium stearate, magnesium stearate, sodium Benzoate, sodium acetate, sodium chloride, and the like. Disintegrants include, but are not limited to, starch, methyl cellulose, agar, bentonite, xanthan gum and the like.

Acceptable pharmaceutical carriers for compositions that are formulated into a liquid solution include sterile water and sterile water suitable for the living body such as saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, And other conventional additives such as an antioxidant, a buffer, and a bacteriostatic agent may be added as needed. In addition, diluents, dispersants, surfactants, binders, and lubricants may be additionally added to formulate into injectable solutions, pills, capsules, granules or tablets such as aqueous solutions, suspensions, emulsions and the like.

Further, it can be suitably formulated according to each disease or ingredient, using the method disclosed in Remington's Pharmaceutical Science, Mack Publishing Company, Easton PA as an appropriate method in the field.

The pharmaceutical composition may be administered orally or parenterally. In the case of parenteral administration, the pharmaceutical composition may be administered by intravenous injection, subcutaneous injection, muscle injection, intraperitoneal injection, transdermal administration or the like, preferably oral administration.

The appropriate dosage of the pharmaceutical composition will vary depending on factors such as the formulation method, the manner of administration, the age, weight, sex, pathological condition, food, administration time, route of administration, excretion rate and responsiveness of the patient, The physician can easily determine and prescribe dosages effective for the desired treatment or prophylaxis. According to a preferred embodiment, the daily dosage of the pharmaceutical composition is 0.001-10 g / kg.

The pharmaceutical composition may be prepared in a unit dose form by formulating it using a pharmaceutically acceptable carrier and / or excipient according to a method which can be easily carried out by a person having ordinary skill in the art to which the present invention belongs. Into a capacity container. The formulations may be in the form of solutions, suspensions or emulsions in oils or aqueous media, or in the form of excipients, powders, granules, tablets or capsules, and may additionally contain dispersing or stabilizing agents.

Hereinafter, the present invention will be described in more detail with reference to Examples and the like, but the scope and content of the present invention can not be construed to be limited or limited by the following Examples. It will be apparent to those skilled in the art that various modifications and variations can be made in the present invention without departing from the spirit and scope of the present invention as set forth in the following claims. It is natural that it belongs to the claims.

In addition, the experimental results presented below only show representative experimental results of the embodiments and the comparative examples, and the respective effects of various embodiments of the present invention which are not explicitly described below will be specifically described in the corresponding part.

Example  1. 36 strains

In order to select candidates for probiotics having an activity of inhibiting obesity, 36 strains derived from human body (Table 1) were collected, and each strain was isolated and identified and shown in Table 1. Each strain was incubated with MRS medium (Difco, 288110) at 37 ° C for 18-24 hours.

number Strain name One Lactobacillus sakei subsp. Sakei 2 Streptococcus infantarius subsp. Coli 3 Bacillus licheniformis 4 Bacillus siamensis 5 Lactobacillus sakei subsp. Carnosus 6 Enterococcus faecalis 7 Enterococcus hirae 8 Carnobacterium maltaromaticum BA 9 Lactobacillus reuteri 10 Enterococcus faecium 11 Enterococcus faecium 12 Enterococcus faecium 13 Lactobacillus fermentum 14 Lactobacillus mucosae 15 Lactobacillus pentosus AO21-1 16 Lactobacillus acidilactici 17 Lactobacillus zeae 18 Enterococcus faecium 19 Lactobacillus reuteri 20 Lactobacillus johnsonii 21 Lactobacillus acidophilus 22 Lactobacillus gasseri 23 Lactobacillus ruminis 24 Lactobacillus mesenteroides subsp. Mesenteroides 25 Obesumbacterium proteus 26 D29 27 I01-7 28 I02-4 29 M40a 30 Lactobacillus acidophilus 31 Lactobacillus rhamnosus 32 pediococcus pentosaceus 33 Bifidobacterium lactis 34 Bifidobacterium breve 35 Lactobacillus acidophilus 36 Lactobacillus intestinalis

Experimental Example  1. Selection of strains with activity to inhibit adipocyte formation

In order to evaluate the anti-obesity efficacy of the above candidate candidates, the ability to inhibit the differentiation of pre-adipocytes (3T3-L1 cells) into adipocytes was analyzed and compared (Example 1).

(1) Experimental method

The accumulation of lipids in fat cells is the most representative feature of obesity. Adipocytes are formed by the differentiation of pre-adipocytes derived from stem cells. During the differentiation of 3T3-L1 cells (adipocytes) into adipocytes, morphological, biochemical Through the change, the intracellular fat sphere is formed, and as the differentiation progresses, the size of the fat sphere becomes larger. Since most lipids are composed of proteins such as triglyceride and perilipin A, the level of lipid differentiation can be confirmed by measuring the intracellular triglyceride content.

Therefore, 3T3-L1 cells, which are representative preadipocytes, were treated with high-grade candidate strains, and the activity of inhibiting adipogenesis was evaluated by measuring the triglyceride (TG) content by inducing differentiation.

Cell culture was maintained for at least 8 days for the above TG content measurement experiment. In order to exclude the influence of the higher candidate strains on the growth of 3T3-L1 cells, the upper candidate strains were treated in the form of dead cells.

Prior to the TG experiment, the cytotoxicity of WST (water soluble tetrazolium) was examined in the upper candidate strains. As a result, the cells did not show cytotoxicity at 1 × 10 ⁷ to 1 × 10 ⁶ CFU, In order to measure the inhibitory activity, the concentration of dead cells of the upper candidate strains to be treated was fixed at 1 × 10 ⁷ to 1 × 10 ⁶ CFU / well.

Specifically, Oil Red O staining was carried out according to the method of Negrel and Dani in order to investigate the effect of the candidate strains on the degree of fat accumulation in the lipid differentiation process. FBS + PS + Insulin + Dexamethasone + IBMX + 1 + 10 + CFU / mL were treated with 3 × 10 ⁷ CFU / mL of each of the above 36 candidate strains of the above candidate strains in 100% confluent 3T3- Insulin). After 10 days of differentiation, the culture medium was removed, washed with PBS, and fixed with 10% formaldehyde for 10 minutes. 10% formaldehyde was removed and the cells were fixed for more than 1 hour by adding saturated formaldehyde again. After that, 60% isopropanol was added to the flask. The flask was washed with distilled water, stained with oil red O staining solution for 30 minutes, and washed with distilled water. The intracellular lipid accumulation of stained lipid was observed with a microscope and the effect of 36 kinds of strains on fat accumulation during lipid differentiation was observed visually. To confirm the accumulation of fat, isopropanol was added to the dried oil to elute the oil red O dye, and absorbance was measured at 490 nm using a multi-plate reader. At this time, isopropanol was used as a blank.

(2) Conclusion

FIG. 1 is a diagram showing a series of analysis procedures for selecting strains having an obesity-suppressing activity according to Experimental Example 1 of the present invention. Fig. 3 is a graph showing lipid accumulation using oil red O staining method. Fig.

2, 3, 5, 6, 7, 8, 12, 13, 14, 15, 16, and 16 of the 36 strains of the respective strains, 24, and 29 strains were found to have decreased TG content by more than 10%. Among them, 4 strains (strain number: 3, 6, 13, 15) were selected.

Among the four strains selected, in order to secondly select strains exhibiting excellent activity of inhibiting obesity in vivo, the in vivo inhibitory activity in vivo was compared with that of the in vivo test described below.

Experimental Example  2. Identification of effects of obesity inhibitory activity on selected strains by animal experiment

end. Experimental animals and Experimental group

C57BL / 6 6 week old males were purchased from Central Experimental Animals, and they were raised at a temperature of 20 ± 2 ℃ and humidity of 55 ± 5% for 12 hours. Experimental animals were purified for 2 weeks after purchase, and 5 mice per group were placed in the experimental group as shown in Table 2.

division
(N = 5) Experimental group Dietary regulation Normal diet Normal control group (ND, 10% fat) fed general diet, PBS 5 times a week, 100 μl / head orally for 8 weeks high fat diet A negative control diet (HFD, 45% fat), which consumed high fat diets for induction of obesity, was administered 5 times per week for 8 weeks with 100 μl / head orally A Consume high fat diets for obesity-induced, and strain number 3 (Bacillus licheniformis) to 0 and then orally administered five times a week 10 ⁸ CFU / head for 4 weeks, 5-8 weeks five times a week 5X10 ⁸ CFU / head during Oral administration B Consume high fat diets for obesity-induced, and strain No. 6 (Enterococcus faecalis) 0 to after oral administration of five times a week 10 ⁸ CFU / head for 4 weeks, 5-8 weeks five times a week 5X10 ⁸ CFU / head during Oral administration C Consume high fat diets for obesity-induced, and strain No. 13 (Lactobacillus fermentum) 0 to after oral administration of five times a week 10 ⁸ CFU / head for 4 weeks, 5-8 weeks five times a week 5X10 ⁸ CFU / head during Oral administration D Ingesting a high-fat diet-induced obesity and for the strain No. 15 (Lactobacillus pentosus AO21-1) for 0-4 weeks after oral administration of 5 times 10 ⁸ CFU / head for a week, 5-8 weeks five times a week for ⁸ 5X10 Oral administration of CFU / head

In all experimental groups except for normal diet and negative control (Hihg fat diet), each strain was mixed with feed, followed by oral administration of feed and strain. FIG. 3 shows a procedure of setting the experimental animal and experimental group according to Experimental Example 2 of the present invention.

I. Weight, feed intake and Quantity of water

All experimental groups were examined for changes in body weight (g), feed intake (g), and water volume (㎖) from the time of feeding the feed to sacrificing the experimental animals. The diet and water intake were measured three times a week. The cages were changed twice a week. Body weight was measured once a week and compared.

FIG. 4 is a graph showing a change in body weight in each experimental group. According to this, the decrease in weight gain was clearly confirmed in Lactobacillus pentosus strain AO21-1 , strain 15.

Experimental Example  3. Identification of selected strains

16S rDNA  analysis

The final selected strains were analyzed for 16S rRNA gene (SEQ ID NO: 1), which is a nucleotide conservation sequence of bacteria, using a universal bacterial primer (518F, 800R) for PCR reaction. The results were interpreted using NCBI blast (http://www.ncbi.nlm.nih.gov/).

The present inventors named Lactobacillus pentosus AO21-1 as " Lactobacillus pentosus AO21-1 (Accession No .: KCCM12144P)" and deposited it on November 2, 2017 at the Korean Microorganism Conservation Center.

Name of depository: Korea Microorganism Conservation Center (overseas)

Accession number: KCCM12144P

Checked on: 20171102

Experimental Example  4. Selected final AO21 -1 in the Animal Experiment of Obesity Sho sign

end. Culture of the strain

The Lactobacillus pentosus AO21-1 strain was inoculated intact into a solid medium (Lactobacilli MRS Agar; BD Difco Co, USA) to produce a final selected Lactobacillus pentosus AO21-1 strain , And cultured at 37 ° C for 24 hours. After thoroughly checking whether the cells were contaminated or formed a single colony during the culturing process, the following experiment was conducted. After the cultivation was completed, a colony was taken from each solid medium and inoculated into 5 mL of a liquid medium (Lactobacilli MRS broth; BD Difco Co, USA) and incubated at 37 ° C and 140 rpm for 24 hours. Was inoculated in 200 mL of another liquid medium (Lactobacilli MRS broth; BD Difco Co, USA) in a 1% (v / v) inoculation stepwise. The activated Lactobacillus pentosus AO21-1 strain was cultured at 37 ° C for 24 hours at 37 ° C and centrifuged at 6000 rpm at 4 ° C for 15 minutes (Avanti JE, Beckman Coulter, USA ) To recover the cells. The recovered cells were washed twice with sterilized 0.85% physiological saline, and lyophilized (FDCF-12003, OPERON, Korea), powdered and stored at -80 ° C until use.

I. Experimental animals and Experimental group

C57BL / 6 male mice, 6 weeks old, were purchased from a central experimental animal and maintained at a temperature of 20 ± 2 ° C and a humidity of 55 ± 5% for 12 hours. Experimental animals were purified for 2 weeks after purchase, and 10 rats per group were randomly assigned to a complete block design.

During the refinement period, the litter was mixed at intervals of 2 to 3 days to homogenize intestinal microflora in the animal model. After that, 10 rats were placed in each experimental group. Normal diet (ND) group was fed normal diet and the other group was fed high diet (Rodent diet with 45% kcal Fat, Research Diets, USA). The Lactobacillus pentosus strain AO21-1 was weighed to a concentration of 5 × 10 ⁷ CFU / head, suspended in sterilized PBS, and administered orally over a period of 5 times per week for 10 weeks.

At the end of the experiment period, the animals were fasted for 12 hours or more. Blood was collected from orbital blood, left at room temperature for 30 minutes or more, and centrifuged at 1,690 × g for 10 minutes. . Liver fat, epididymal fat, retroperitoneal fat, saliva fat and shabbat liver brown fat were extracted, washed with saline to remove water and white fat removed from the brown fat tissue of the scabbard liver was removed and the weight of each was measured.

division
(N = 10) Experimental group ND Oral administration of 100 μl / head for 10 weeks, 5 times a week, a normal control group (ND, 10% fat) HFD The subjects were fed a high-fat diets (Rodent diet with 45% kcal Fat, Research Diets, USA), PBS for 5 weeks, 100 μl / head for 10 weeks Lactobacillus pentosus AO21-1 Fat diet for induction of obesity, and the final selected Lactobacillus pentosus AO21-1 strain was orally administered 5 x 10 < ⁷ > CFU / head five times a week for 10 weeks

All. Weight, feed intake and Quantity of water

All experimental groups were examined for body weight change (g), feed intake (g / day), and water volume (g / day) of the experimental animals from the feeding time to the experimental animal sacrifice. The diet and water intake were measured three times a week. The cages were changed twice a week. Body weight was measured once a week and compared.

FIG. 5 is a graph showing the change in body weight in each experimental group by week. FIG. 6 is a graph showing an analysis of body weight gain in each experimental group. FIG. 5 is a graph showing the relationship between feed intake (g / day) The intake (kcal / day) is shown in Table 4.

ND HFD Lactobacillus pentosus AO21-1 P value Average of daily food intake
(g / day) 3.56 ± 0.05 2.64 ± 0.04 2.57 + 0.04 <0.001 Average of daily water intake
(g / day) 3.81 ± 0.08 2.96 ± 0.09 2.95 + - 0.10 <0.001 Average of daily calorie intake
(kcal / day) 12.83 + - 0.17 13.36 ± 0.21 13.00 + 0.04 N.S.

According to Figures 5, 6 and Table 4, Lactobacillus pentosus strain AO21-1 was administered to mice fed with a high fat diet for 10 weeks. As a result, compared to the HFD group fed only with high fat diets, As shown in FIG.

In other words, since check that the gain less weight than the HFD group consumed the high fat diet, Lactobacillus pento Versus (Lactobacillus pentosus) AO21-1 strain was confirmed that even though the intake of high-fat diet, and the weight gain inhibition compared to the HFD group bar, Lactobacillus pento Versus (Lactobacillus pentosus) AO21-1 strains it can be seen that has a weight gain suppressing effect.

The calorie intake was calculated from the amount of feed consumed. Mean daily dietary intake and water intake were significantly different between ND and HFD groups. It is expected that this is due to differences in the constituents and properties of feed, and ND group consumes more feed and water intake than HFD group.

HFD group and Lactobacillus pentosus AO21 -1 groups are different from each food, water, bar hayeotneun confirmed that no difference in calorie intake, feed weight gain inhibitory activity by a Lactobacillus pento Versus (Lactobacillus pentosus) AO21-1 strain evidence by experiments prior through which or It is not influenced by the decrease of caloric intake.

la. Liver, epididymal fat, Posterior peritoneum  Fat, Cry  Fat, Shoulder bag  Local analysis

At the end of the experimental period, liver, epididymal fat, retroperitoneal fat, follicular fat and shabu - shabu brown fat were extracted, washed with saline to remove water, and white fat attached to brown fat tissue After removal, weights were measured relative to body weight and are shown in Table 5 below. In Table 5, Liver is liver fat, EFT is epididymis fat, RFT is retroperitoneal fat, IFT is fat, and iBAT is interscapular brown adipose tissue.

Tissue weight
(mg) ND HFD Lactobacillus pentosus AO21-1 P value Liver 834.6 ± 15.48 916.16 + 28.37 842.60 ± 15.88 N.S. (0.055) EFT 270.49 + - 19.51 ^a 1741.68 ± 188.37 ^b 1550.53 + - 134.21 ^b <0.001 RFT 82.00 ± 22.96 ^a 642.58 + - 57.26 ^b 579.81 + - 45.60 ^b <0.001 IFT 159.19 ± 14.20 ^a 873.74 + 84.50 ^b 758.48 + 49.47 ^b <0.001 iBAT 54.27 ± 2.59 ^a 88.10 ± 5.91 ^c 72.98 + - 6.94 ^b 0.003

According to Table 5, the HFD group consuming high fat diet showed slightly increased liver weight compared to ND group and Lactobacillus pentosus AO21-1.

In addition, the weight of the epididymal white fat tissue (EFT), the retroperitoneal white fat tissue (RFT), the white fat tissue (IFT), and the brown fat tissue of the scabbit liver (iBAT) On the contrary, when the Lactobacillus pentosus AO21-1 strain was administered, the fat weight in each tissue was significantly decreased.

Is increased in the HFD group consumed a high fat, Lactobacillus pento Versus (Lactobacillus pentosus) when administered to AO21-1 strain, it is understood that it has significantly decreased.

<110> KOREA FOOD RESEARCH INSTITUTE <120> Lactobacillus pentosus AO21-1 strain with Anti-Obesity Ability          and Composition for treatment or prevention          obesity comprising the same <130> HPC-7704 <160> 1 <170> KoPatentin 3.0 <210> 1 <211> 1454 <212> DNA <213> Artificial Sequence <220> <223> 16s rDNA of Lactobacillus pentosus AO21-1 <400> 1 gctggcggcg tgcctaatac atgcaagtcg aacgaactct ggtattgatt ggtgcttgca 60 tcatgattta catttgagtg agtggcgaac tggtgagtaa cacgtgggaa acctgcccag 120 aagcggggga taacacctgg aaacagatgc taataccgca taacaacttg gaccgcatgg 180 tccgagtttg aaagatggct tcggctatca cttttggatg gtcccgcggc gtattagcta 240 gatggtgggg taacggctca ccatggcaat gatacgtagc cgacctgaga gggtaatcgg 300 ccacattggg actgagacac ggcccaaact cctacgggag gcagcagtag ggaatcttcc 360 acaatggacg aaagtctgat ggagcaacgc cgcgtgagtg aagaagggtt tcggctcgta 420 aaactctgtt gttaaagaag aacatatctg agagtaactg ttcaggtatt gacggtattt 480 aaccagaaag ccacggctaa ctacgtgcca gcagccgcgg taatacgtag gtggcaagcg 540 ttgtccggat ttattgggcg taaagcgagc gcaggcggtt ttttaagtct gatgtgaaag 600 ccttcggctc aaccgaagaa gtgcatcgga aactgggaaa cttgagtgca gaagaggaca 660 gtggaactcc atgtgtagcg gtgaaatgcg tagatatatg gaagaacacc agtggcgaag 720 gcggctgtct ggtctgtaac tgacgctgag gctcgaaagt atgggtagca aacaggatta 780 gataccctgg tagtccatac cgtaaacgat gaatgctaag tgttggaggg tttccgccct 840 tcagtgctgc agctaacgca ttaagcattc cgcctgggga gtacggccgc aaggctgaaa 900 ctcaaaggaa ttgacggggg cccgcacaag cggtggagca tgtggtttaa ttcgaagcta 960 cgcgaagaac cttaccaggt cttgacatac tatgcaaatc taagagatta gacgttccct 1020 tcggggacat ggatacaggt ggtgcatggt tgtcgtcagc tcgtgtcgtg agatgttggg 1080 ttaagtcccg caacgagcgc aacccttatt atcagttgcc agcattaagt tgggcactct 1140 ggtgagactg ccggtgacaa accggaggaa ggtggggatg acgtcaaatc atcatgcccc 1200 ttatgacctg ggctacacac gtgctacaat ggatggtaca acgagttgcg aactcgcgag 1260 agtaagctaa tctcttaaag ccattctcag ttcggattgt aggctgcaac tcgcctacat 1320 gaagtcggaa tcgctagtaa tcgcggatca gcatgccgcg gtgaatacgt tcccgggcct 1380 tgtacacacc gcccgtcaca ccatgagagt ttgtaacacc caaagtcggt ggggtaacct 1440 tttaggaacc agcc 1454

Claims (8)

Lactobacillus pentosus AO21-1 strain ( KCCM12144P ) having obesity inhibitory activity . A probiotics comprising the Lactobacillus pentosus strain AO21-1 ( KCCM 12144P ) or a culture thereof of claim 1. A pharmaceutical composition for preventing or treating obesity comprising the Lactobacillus pentosus strain AO21-1 ( KCCM12144P ) or a culture thereof according to claim 1. The pharmaceutical composition for preventing or treating obesity according to claim 3, wherein the Lactobacillus pentosus AO21-1 strain ( KCCM 12144P ) or a culture thereof is contained in an amount of 0.01 to 50% by weight based on the total weight of the composition Composition. The pharmaceutical composition according to claim 3, wherein the composition inhibits mast cell differentiation. The pharmaceutical composition for preventing or treating obesity according to claim 3, wherein the composition reduces weight. A composition for prevention or improvement of obesity comprising Lactobacillus pentosus strain AO21-1 ( KCCM12144P ) or a culture thereof according to claim 1. A composition for improving inflammation caused by obesity comprising the Lactobacillus pentosus AO21-1 strain ( KCCM12144P ) of claim 1 or a culture thereof.

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