KR20170017176A - Composition for increasing lactic acid bacteria in intestine and method for producing lactic acid bacteria using the same - Google Patents

Abstract

The present invention relates to a composition for increasing intestinal lactobacilli, containing a compound represented by chemical formula 1, a derivative thereof, or a pharmaceutically or sitologically acceptable salt as an active ingredient. According to the present invention, the composition for increasing intestinal lactobacilli induces proliferation of intestinal lactobacilli while inhibiting proliferation of bad intestinal bacteria, thereby relieving diarrhea, enteritis, and gastrointestinal disturbance.

Description

TECHNICAL FIELD The present invention relates to a composition for increasing lactic acid bacteria in the intestine and a method for producing lactic acid bacteria using the same.

The present specification describes a composition for increasing lactic acid bacteria in the intestines and a method for producing lactic acid bacteria using the same.

Within the intestines of the human body, various microbial populations are clustered and affect health in various aspects depending on the composition of the microbial community. If the balance of these microbial communities is destroyed by stress, aging, diet, infection of harmful bacteria, etc., it may lead to rapid growth of harmful microorganisms, which can lead to metabolic diseases, inflammatory diseases and the like and exacerbate health. In this case, antibiotics are used for therapeutic purposes, and the use of antibiotics can cause side effects that cause the resistance of harmful microorganisms.

As an alternative to this, it is possible to use a probiotic which directly feeds beneficial microorganisms to the body to increase the beneficial bacteria in the intestines. However, in order to commercialize them, it is necessary to produce such beneficial microorganisms with high production yield. However, microorganisms which are beneficial to human body are very difficult to grow in vitro, and thus are difficult to produce. In order to exhibit the effect as a probiotics, the microorganisms must survive the lyophilization or pelletization process during the commercialization process, and must survive the strong acid and bile acid to settle in the intestines. However, since probiotics with all of these conditions are rare, the probiotics that we ingest actually have very little effect compared to the efficacy of the original microorganism.

As a method for overcoming this problem, a prebiotics for improving the health by selectively activating the growth of beneficial microorganisms present in the human body has been proposed and many researches and developments have been made.

Korean Patent Publication No. 10-1395875

In one aspect, the present invention provides a composition capable of inducing proliferation of a lactic acid bacterium in a field which is difficult to mass-propagate in vitro, thereby improving diarrhea, enteritis, gastrointestinal disorder, etc. of a subject.

In another aspect, the present invention provides a composition for inducing the proliferation of lactic acid bacteria in the intestines and inhibiting the proliferation of noxious bacteria in the intestines to thereby improve diarrhea, enteritis, gastrointestinal disorders and the like.

In another aspect, the present invention provides a method for producing a lactic acid bacterium having a high production yield by propagating lactic acid bacteria which are difficult to mass-propagate in vitro in mammalian intestines.

In one aspect of the present invention, there is provided a composition for intestinal lactobacillus growth comprising a compound of the following formula (1), a derivative thereof, or a pharmaceutically or pharmacologically acceptable salt thereof as an active ingredient.

≪ Formula 1 >

Wherein each of R ₁ , R ₂ , R ₃ and R ₄ is independently a linear or branched C ₁ to C ₁₈ alkyl group, a C ₁ to C ₁₈ alkenyl group, a C ₁ to C ₁₈ alkynyl group, C ₃ to C ₆ cyclic alkyl group.

Another aspect of the present invention provides a method for producing a lactic acid bacterium comprising orally administering the composition for intracoronary lactic acid bacteria growth to a mammal other than a human and extracting the intestinal lactic acid bacteria from the excrement of the mammal.

In one aspect, the present invention provides a composition capable of inducing proliferation of a lactic acid bacterium in a field which is difficult to mass-propagate in vitro, thereby improving diarrhea, enteritis and gastrointestinal disorder of a subject.

In another aspect, the present invention can provide a composition for inducing the proliferation of lactic acid bacteria in the intestines and inhibiting the proliferation of noxious bacteria in the intestines to thereby improve diarrhea, enteritis, gastrointestinal disorders and the like.

In another aspect, the present invention provides a method of producing a lactic acid bacterium having a high production yield by propagating lactic acid bacteria which are difficult to mass-propagate in vitro in mammalian intestines.

FIG. 1 is a graph showing the concentration of Sirengarolol according to the fraction of ginseng fruit extract. FIG.
FIG. 2 is a graph showing the frequency of enterobacteriaceae in the intestines analyzed from the side separated from the large intestine of the rat administered with the test substance.
FIG. 3 is a graph showing the frequency of intestinal noxious organisms analyzed in the side separated from the large intestine of a mouse to which an experimental substance was orally administered.
FIG. 4 shows the effect of Sirling's lycinol and its extract on the proliferation of Lactobacillus luteri.
FIG. 5 is a graph showing the improvement effect of ulcerative colitis induced by dextran sulfate sodium (DSS) in mice administered with orally administered ginseng fruit extract containing Sirling's resinoid.

Embodiments of the present application will now be described in more detail with reference to the accompanying drawings. However, the techniques disclosed in the present application are not limited to the embodiments described herein but may be embodied in other forms. It should be understood, however, that the embodiments disclosed herein are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the invention to those skilled in the art. 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 invention.

As used herein, " derivative " means any compound that is substituted with the same or different substituents at substitutable positions of the compounds. Examples of such substituents include, but are not limited to, for example, a hydroxyl group, a phenoxy group, a thienyl group, a furyl group, a pyridyl group, a cyclohexyl group, an alkyl alcohol group, an alkyldialcohol group, a substituted or unsubstituted phenyl group, An unsubstituted C1 to C20 bicyclic hydrocarbon group; A C3 to C30 cyclic hydrocarbon group substituted or unsubstituted with a hydroxyl group, a hydroxymethyl group, a methyl group, or an amino group; Or < / RTI > sugar moieties.

As used herein, the term " sugar residue "refers to a group in which one hydrogen atom has been removed from the polysaccharide molecule, and thus may refer to a residue derived, for example, from a monosaccharide or oligosaccharide.

As used herein, " substituted " means that at least one hydrogen atom in the functional group is replaced by a halogen (F, Cl, Br or I), a hydroxy group, a nitro group, a cyano group, (Wherein R is a C1 to C10 alkyl group), an amino group (-NH2, -NH (R '), -N (R ") (R"'), R ', R " C30 alkyl group, a C3 to C30 heteroaryl group, a C3 to C30 heteroaryl group, or a C2 to C30 heteroaryl group, C30 < / RTI > heterocycloalkyl group.

As used herein, "hetero" means that a carbon atom is substituted with any one atom selected from the group consisting of N, O, S and P.

As used herein, the term " pharmaceutically acceptable "refers to the approval of a government or equivalent regulatory agency that can be used on animals, and more specifically on humans, by avoiding significant toxic effects when used in conventional medicinal dosage Received, approved, or listed in a pharmacopoeia or other general pharmacopoeia.

As used herein, "pharmaceutically acceptable salt" means a salt according to one aspect of the present invention which is pharmaceutically acceptable and possesses the desired pharmacological activity of the parent compound. The salt may be formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and the like; (3-hydroxybenzoyl) benzoic acid, or an acetic acid, propionic acid, hexanoic acid, cyclopentenepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, Benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethane-disulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, 4- chlorobenzenesulfonic acid, 2- 4-methylbicyclo [2,2,2] -oct-2-en-1-carboxylic acid, glucoheptonic acid, 3-phenylpropionic acid, trimethylacetic acid, tert Acid addition salts formed with organic acids such as butylacetic acid, laurylsulfuric acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, muconic acid; Or salts formed when the acidic proton present in the parent compound is substituted. In addition, the compounds according to the present specification may include not only pharmaceutically acceptable salts, but also all salts, hydrates and solvates which can be prepared by conventional methods.

The composition for intestinal lactobacillus growth according to one embodiment of the present invention may include a compound of the following formula 1, a derivative thereof, or a pharmaceutically or pharmacologically acceptable salt thereof as an active ingredient.

≪ Formula 1 >

Wherein each of R ₁ , R ₂ , R ₃ and R ₄ is independently a linear or branched C ₁ to C ₁₈ alkyl group, a C ₁ to C ₁₈ alkenyl group, a C ₁ to C ₁₈ alkynyl group, C ₃ to C ₆ cyclic alkyl group.

The compound of Formula 1 is a lignan compound, including, but not limited to, syringaresinol, eudesmin, yangambin, or sesamin. In an embodiment, the lignan compound comprises the compound of Formula 1 or an isomer thereof. (+) - sialing regresin, (-) - seringgrecinol, (+) - diethylenepentamine, and the like are more preferred. Desmin, (-) - yudesmin, (+) - persimmon, (-) - persimmon, (+) - sesamin or (-) - sesamin.

(2)

(3)

Wherein R ₁ , R ₂ , R ₃ , and R ₄ are as defined in Formula 1.

In an embodiment, the lignan compound may be obtained through chemical synthesis or may be obtained from one or more of flaxseed, white, ocher, sesame and ginseng.

In an embodiment, the compound may be obtained by purifying an extract derived from one or more of flaxseed, yellow, white, oak, sesame and ginseng.

The above flaxseed, yellowtail, oak, and sesame include all parts of the plant such as leaf, stem, root, fruit or seed of each plant, and the ginseng fruit may include the seed part of the ginseng fruit, have.

The above-mentioned extract is a broad concept including all the substances obtained by extracting the components of natural products, regardless of the extraction method, the extraction solvent, the extracted component or the form of the extract.

For example, the extract may be obtained by extracting at least one selected from the group consisting of flaxseed, yellow white, ocher, sesame and ginseng in water, an organic solvent, and a mixture of water and an organic solvent. The organic solvent includes, but is not limited to, at least one selected from the group consisting of alcohol, acetone, ether, ethyl acetate, diethyl ether, ethyl methyl ketone and chloroform. Wherein the alcohol comprises a C1 to C5 lower alcohol and the C1 to C5 lower alcohol includes at least one member selected from the group consisting of methanol, ethanol, isopropyl alcohol, n-propyl alcohol, n-butanol and isobutanol , But is not limited thereto.

In an embodiment, the extract may be an extract of ginseng fruit. Ginseng fruit extract, alcohol extract of ginseng fruit pulp; Eluting the alcohol extract with a solvent containing at least one of water and alcohol to obtain a fraction; And fractionating the obtained fractions by chromatography using, for example, thin film chromatography (TLC) using an organic solvent as a developing solvent. The organic solvent may include at least one selected from the group consisting of alcohol, acetone, ether, ethyl acetate, diethyl ether, ethyl methyl ketone and chloroform, and the alcohol may include C1 to C5 alcohols.

For example, the compound of Formula 1 may be syringaresinol of Formula 4 below.

≪ Formula 4 >

(-) - sialing regreon of formula (5) or (8R, 8'R) - (-) - +) -Single ring may be a recurrence, for example (8R, 8'R) - (+) - Syringal resorcin.

≪ Formula 5 >

(6)

The lactic acid bacteria may include one or more of Lactobacillus sp. Strain and Bifidobacterium sp. Strain. For example, the lactic acid bacteria may be one or more of Lactobacillus reuteri and Bifidobacterium adolescentis.

Lactobacillus lutea is a typical intestinal bacterium, and it is the most important probiotic in the lactic acid bacteria industry. Lactobacillus luteriens ferment glycerol and produce antibiotics rutherin, which has a strong antimicrobial activity. Therefore, it shows excellent efficacy in preventing and treating common diarrhea, enteritis, allergies and candidiasis in infants and young children (Urbanska et al., Eur. J. Pediatr 2014). Am J Physiol Gastrointest Liver Physiol. 2010). In addition, it is possible to improve insulin resistance by controlling the secretion of inflammatory cytokines in the intestinal mucosa, thereby reducing inflammatory bowel disease, irritable bowel syndrome, and gastrointestinal disorders (Liu et al. (Poutahidis et al., Plos One 2014)), but also in the development of cognitive function in the brain (Hsieh et al., Nutrition & Metabolism 2013), inhibiting the reduction of testosterone by aging (PCT / EP2012 / 074023).

Such Lactobacillus luteri is a strain that is difficult to produce in large quantities by culturing because the growth conditions in vitro are very difficult.

Accordingly, the composition according to one embodiment of the present invention is orally administered to a subject to activate the proliferation of Lactobacillus lurii present in the intestine of a subject, thereby increasing the lactobacillus lutei in the intestine and thereby preventing diarrhea, enteritis, gastrointestinal disorder, , Candidiasis, and the like can be exhibited as prebiotics that show improvement, treatment and prevention effects.

In an embodiment, the compound of Formula 1, which is an active ingredient, may be included in an amount of 0.001 wt% to 80 wt%, specifically 0.005 wt% to 60 wt%, more specifically 0.01 wt% to 30 wt% based on the total weight of the composition . Can be effectively exhibited within the above range to exhibit an intended effect, can satisfy both the stability of the composition and the stability of the formulation, and can be excellent in terms of cost-effectiveness. It is possible to exhibit sufficient stomach and intestinal disease ameliorating effect within the above range and to prevent the safety such as toxicity and the formulation stability problem when the active ingredient is contained in an excessive amount.

In another embodiment of the present invention, the composition for intestinal lactobacillus growth may comprise the compound of formula 1 as an extract of at least one of flaxseed, yellowtail, oak, sesame and ginseng fruit. The composition for enhancing intestinal lactobacillus according to one embodiment of the present invention, except that it is contained as an extract of at least one of flaxseed, yellow white, ocher, sesame and ginseng, So the description is omitted.

In the present embodiment, the flaxseed, yellowtail, oak, and sesame include all the parts of the plant, such as leaves, stems, roots, fruits, or seeds of each plant. The ginseng fruit may include seeds, peel or flesh of ginseng fruit, for example, a ginseng fruit concentrate obtained by removing solid content from the total juice except seeds from the ginseng fruit.

The above-mentioned extract is a broad concept including all the substances obtained by extracting the components of natural products, regardless of the extraction method, the extraction solvent, the extracted component or the form of the extract.

For example, the extract may be obtained by extracting at least one selected from the group consisting of flaxseed, yellow white, ocher, sesame and ginseng in water, an organic solvent, and a mixture of water and an organic solvent. The organic solvent includes, but is not limited to, at least one selected from the group consisting of alcohol, acetone, ether, ethyl acetate, diethyl ether, ethyl methyl ketone and chloroform. Wherein the alcohol comprises a C1 to C5 lower alcohol and the C1 to C5 lower alcohol includes at least one member selected from the group consisting of methanol, ethanol, isopropyl alcohol, n-propyl alcohol, n-butanol and isobutanol , But is not limited thereto.

In an embodiment, the extract may be an extract of ginseng fruit. Ginseng fruit extract, alcohol extract of ginseng fruit pulp; Eluting the alcohol extract with a solvent containing at least one of water and alcohol to obtain a fraction; And fractionating the obtained fractions by chromatography using, for example, thin film chromatography (TLC) using an organic solvent as a developing solvent. The organic solvent may include at least one selected from the group consisting of alcohol, acetone, ether, ethyl acetate, diethyl ether, ethyl methyl ketone and chloroform, and the alcohol may include C1 to C5 alcohols.

The extract may contain 0.01% to 40% by weight, for example 0.04% to 24% by weight, of the compound of Formula 1 based on the total weight of the extract.

In another embodiment of the present invention, the composition for intramuscular increase in lactic acid bacteria as a prebiotics may be a composition to be administered simultaneously with a probiotic Lactobacillus reuteri .

The composition according to the embodiments of the present invention may be provided as various types of food additives or functional foods containing the active ingredient. Cheese, yogurt, juice, probiotic agent, and health food containing the above-mentioned active ingredient, and may be used in various other food additives.

The composition according to embodiments of the present invention may be a health food composition. The health food composition may be formulated into a pill, a capsule, a tablet, a granule, a caramel or a drink. In other embodiments, it may be processed in the form of a liquid, powder, granule, tablet or tea bag.

The composition may be administered by various methods such as simple drinking, injection administration, spraying or squeezing.

The composition may contain other components or the like which can give a synergistic effect to the main effect within a range that does not impair the main effect of the present invention. For example, additives such as perfume, coloring agent, bactericide, antioxidant, preservative, moisturizing agent, thickening agent, inorganic salt, emulsifier and synthetic polymer substance may be further added for improvement of physical properties. In addition, it may further contain auxiliary components such as water-soluble vitamins, oil-soluble vitamins, polymer peptides, polymeric polysaccharides and seaweed extract. These ingredients can be selected by a person skilled in the art depending on the formulation or purpose of use, and the amount thereof can be selected within a range that does not impair the purpose and effect of the present invention. For example, the amount of addition of the above components may be from 0.01% by weight to 5% by weight, for example from 0.01% by weight to 3% by weight, based on the total weight of the composition.

The composition according to embodiments of the present invention may be a pharmaceutical composition.

In an embodiment, the pharmaceutical composition further comprises an inorganic or inorganic carrier which is commonly used according to a conventional method, and may be formulated into various forms such as solid, semi-solid or liquid, orally or parenterally.

The oral dosage form may be, for example, tablets, pills, granules, soft capsules, powders, granules, powders, emulsions, syrups, pellets and the like. The parenteral dosage forms may be, for example, Lotions, sprays, suspensions, emulsions, suppositories, and the like. The pharmaceutical composition may further contain a preservative, a stabilizer, a wetting or emulsifying accelerator, a pharmaceutical adjuvant such as a salt or buffer for controlling osmotic pressure, and other therapeutically useful substances.

The pharmaceutical composition may be administered orally, parenterally, topically, transdermally, subcutaneously, rectally, intravenously, intramuscularly, intraperitoneally, and the like.

It should be understood that the actual dosage of the active ingredient should be determined in light of various relevant factors such as the severity of the symptoms, the route of administration selected, the age, sex, weight and health status of the subject. In general, the dosage of the active ingredient is from 0.001 mg / kg / day to 2000 mg / kg / day, for example from 0.5 mg / kg / day to 1500 mg / kg / day.

In one embodiment of the present invention, a kit comprising the composition for intracoronary lactic acid bacteria growth and the Lactobacillus reuteri according to the embodiments of the present invention can be provided.

The method for producing a lactic acid bacterium according to an embodiment of the present invention may include orally administering the composition of the present invention to mammals other than human, and extracting intestinal lactic acid bacteria from the excreta of the mammal. According to the production method according to the present embodiment, it is possible to economically mass-produce a lactic acid bacterium which is difficult to mass-produce because of difficult growth and proliferation conditions in vitro, and thus produced lactic acid bacteria can be provided as probiotics.

The mammal is not limited as long as it is a mammal other than a human. For example, mammals such as rats such as Mus or Rattus can be used in terms of production efficiency and economy.

The lactic acid bacteria may include one or more of Lactobacillus sp. Strain and Bifidobacterium sp. Strain. For example, the lactic acid bacteria may be at least one of Lactobacillus reuteri and Bifidobacterium adolescentis, for example, Lactobacillus reuteri.

Hereinafter, the present invention will be described in detail with reference to Examples, Comparative Examples and Test Examples. It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not to be construed as limiting the scope of the present invention. It will be self-evident.

[Preparation Example 1] Preparation of ginseng fruit extract

100 kg of fruit from 4 years old or more ginseng was washed and 3 ~ 10 times of purified water of ginseng fruit and ginseng fruit volume washed in seed separator was added to the seeds of ginseng fruit to the other parts (flesh, . Approximately 75 kg of ginseng seeds were collected, and juice containing the perilla, flesh, and juice of the ginseng fruit was obtained.

Decanter was used to separate ginseng fruit, pectin and other solids from the juice as food processing equipment and concentrated under reduced pressure to prepare a first concentrate.

To the first concentrate is added a 15-fold volume of ethanol to the ginseng fruit concentrate and subjected to reflux extraction at about 85 占 폚. The resultant was filtered and concentrated under reduced pressure to prepare ginseng fruit extract.

The resulting ginseng fruit extract solids were purified by reversed-phase (ODS) column chromatography. The elution solvents used were 100% / 0%, 90% / 10%, 80% / 20% %, 70% / 30%, 60% / 40%, 50% / 40%, 40% / 60%, 30% / 70%, 20% / 80%, 10% / 90%, 0% / 100% Elution with solvents resulted in fractions of GB-1 to GB-10. The content of (+) - Syringal resinol in each fraction is shown in FIG.

The extract of GB-1 fraction shown in Fig. 1 was used as a ginseng fruit extract in the following test examples.

[Preparation Example 2] Preparation of (+) - Syringal Resinol

(1) Preparation of 4- (benzyloxy) -3,5-dimethoxybenzaldehyde (Compound 8) 4- (benzyloxy) -3,5-dimethoxybenzaldehyde

Syringaldehyde (Compound 7) (3 g, 16.47 mmol) and potassium carbonate (6.83 g, 49.4 mmol) were dissolved in DMF (16 mL) in a nitrogen atmosphere, benzyl bromide (2.94 mL, 24.7 mmol), and the mixture was stirred at room temperature overnight. After confirming the termination of the reaction by TLC, the reaction solution is extracted with water and ethyl acetate. The extracted organic layer is washed three times with water, dehydrated with anhydrous MgSO ₄ , and then filtered with a filter. The filtrate was concentrated and purified by column chromatography (5: 1 hexanes / EtOAc) to give compound 8 as a white solid (4.45 g, 99.2%). NMR data of Compound 8 is as follows.

¹ H NMR (300 MHz, CDCl ₃ )? 3.90 (s, 6H), 5.13 (s, 2H), 7.11 (s, 2H), 7.29-7.48 (m, 5H), 9.86 ^{13 C NMR (75 MHz, CDCl} 3) δ 56.2, 75.0, 106.7, 128.1, 128.2, 128.4, 128.6, 131.9, 137.2, 191.1.

(2) Synthesis of 1- (4- (benzyloxy) -3,5-dimethoxyphenyl) prop-2 -en-1-ol < / RTI > (Compound 9)

In a nitrogen atmosphere, Compound 8 (4.04 g, 14.8 mmol) obtained above was dissolved in anhydrous THF (30 mL), and the temperature was lowered to -78 ° C. Vinylmagnesium bromide (17.8 mL of 1M THF solution) was added, stirred at -78 ° C for 10 minutes, then cooled to 0 ° C and stirred for another 2 hours. After completion of the reaction was confirmed by TLC, the reaction solution was quenched with a saturated aqueous NH ₄ Cl solution and extracted with ethyl acetate. The extracted organic layer is washed three times with water, dehydrated with anhydrous MgSO ₄ , and then filtered with a filter. The filtrate was concentrated and purified by column chromatography (4: 1 hexane / EtOAc) to give Compound 9 (3.97 g, 89.1%) as a pale yellow solid. NMR data of Compound 9 is as follows.

¹ H NMR (300 MHz, CDCl ₃ )? 3.83 (s, 6H), 4.99 (s, 2H), 5.14 (m, 1H), 5.22 (ddd, J = 10.5,1.2,1.2 Hz, 1H) (ddd, J = 16.8, 1.8, 1.8 Hz, 1H), 6.05 (ddd, J = 16.5, 10.5, 6.0 Hz, 1H), 6.60 (s, 2H), 7.28-7.51 (m, 5H); ¹³ C NMR (75 MHz, CDCl ₃ )? 56.1, 75.0, 75.4, 102.5, 103.4, 115.3, 127.8, 128.1, 128.4, 137.9, 138.3, 140.0, 153.6.

(3) Synthesis of (R) - (4- (benzyloxy) -3,5-dimethoxyphenyl) ((R) 5-dimethoxyphenyl) ((R) -oxiran-2-yl) methanol) (Compound 10)

In an atmosphere of nitrogen, add dried anhydrous methylene chloride (45 mL) to dried powdered 4 Å molecular sieve (10.5 g) and lower the temperature to -20 ° C. Titanium isopropoxide (1.48 mL, 4.99 mmol) is slowly added to the solution, and the mixture is stirred for 30 min. After the addition of (-) - DIPT (diisopropyltryptamine) (1.18 mL, 5.99 mmol) After 30 minutes cumene hydroxide (1.85 mL, 80%, 5.99 mmol) is added and stirred for 30 minutes. The compound 21 (3 g, 9.99 mmol) dissolved in anhydrous methylene chloride (10 mL) is then slowly added dropwise. After stirring at -20 < 0 > C for 5 hours, the temperature is raised to 0 < 0 > C and stirred overnight. After confirming the reaction by TLC, 10% NaOH solution (30 mL, in saturated NaCl solution) is added, stirred for 3 hours, and then filtered with a filter. The filtrate is extracted with methylene chloride and washed with brine solution. The mixture was dehydrated with anhydrous Na ₂ SO ₄ , filtered through a filter, and then the filtrate was concentrated and purified by column chromatography (3: 1 hexane / EtOAc) to obtain Compound 10. NMR data of Compound 10 are as follows.

^{_{[α] D -47.5 o (c}} 1.0, CHCl 3); ^{1 H NMR (300 MHz, CDCl} 3) δ 2.32 (d, J = 2.1 Hz, 1H), 2.79 (dd, J = 4.8, 3.9 Hz, 1H), 2.94 (dd, J = 4.8, 2.7 Hz, 1H) , 3.22 (q, J = 3.3 Hz, 1H), 3.84 (s, 6H), 4.82 (t, J = 2.1 Hz, 1H), 5.00 (s, 2H), 6.61 (s, 2H), 7.29-7.51 m, 5H); ^{13 C NMR (75 MHz, CDCl} 3) δ 43.73, 54.98, 56.14, 71.11, 74.99, 76.57, 77.00, 77.42, 103.39, 127.78, 128.11, 128.41, 135.20, 136.84, 137.78, 153.69.

(4) Synthesis of 2- (4 - ((R) - ((E) -3- (4- (benzyloxy) -3,5- dimethoxyphenyl) allyloxy) Yl) methyl) -2,6-dimethoxyphenoxy) tetrahydro-2H-pyran (2- (4- ((R) -dimethoxyphenyl) allyl) oxy) ((R) -oxiran-2-yl) methyl) -2,6-dimethoxyphenoxy) tetrahydro-2H- pyran

Add dried THF / DMSO (10: 1) (20 mL) to NaH (0.633 g, 60% dispersion, 15.82 mmol) in a nitrogen atmosphere and lower the temperature to 0 ° C. Compound 23 (0.5 g, 1.58 mmol) dissolved in THF (10 mL) was added and stirred. Then, cinnamyl bromide (Compound 11) (1.15 g, 3.16 mmol) dissolved in THF (10 mL) was added and stirred at 0 ° C for 30 minutes, then the temperature was raised to room temperature and stirred overnight. The reaction solution is quenched with water and extracted with ethyl acetate. The extracted organic layer is washed three times with water, dehydrated with anhydrous MgSO ₄ , and then filtered with a filter. The filtrate was concentrated and purified by column chromatography (3: 1 hexane / EtOAc) to give compound 12 (760 mg, 80.27%) as a pale yellow oil. NMR data of Compound 12 are as follows.

^{1 H NMR (300 MHz, CDCl} 3) δ 2.82 (m, 2H), 3.19 (dt, J = 2.7, 3.9 Hz, 1H), 3.83 (s, 6H), 3.84 (s, 6H), 4.12 (m, 2H), 4.32 (d, J = 5.1 Hz, 1H), 5.01 (s, 2H), 5.02 (s, 2H), 6.18 (dt, J = 15.9, 6.0 Hz, 1H), 6.47 (d, J = 17.1 Hz, 1 H) 6.59 (d, J = 4.8 Hz, 4H), 7.28-7.52 (m, 10H); ^{13 C NMR (75 MHz, CDCl} 3) δ 45.3, 54.4, 56.1, 56.2, 69.7, 75.0, 75.1, 80.1, 103.7, 104.4, 125.1, 127.8, 128.1, 128.4, 128.5, 132.3, 132.8, 134.0, 137.9, 153.6 , 156.7.

(5) Preparation of Compound 13

Cp ₂ TiCl ₂ (0.182 g, 1.463 mmol) dissolved in THF (20 mL) was added to a flask containing activated zinc dust (0.146 g, 4.44 mmol) in a nitrogen atmosphere and stirred for 1 hour to obtain Cp ₂ TiCl (green solution). In another flask, Compound 25 (0.19 g, 0.635 mmol) was dissolved in THF (16 mL), adjusted to 60 ° C, and the Cp ₂ TiCl solution prepared above was added for 20 minutes using a cannula. After stirring for 20 minutes, I ₂ (0.105 g, 0.825 mmol) dissolved in THF (4 mL) was added and the reaction solution was stirred for 1 hour. After completion of the reaction is confirmed by TLC, saturated NH ₄ Cl aqueous solution is added to the reaction solution to terminate the reaction and extracted with diethyl ether. The extracted organic layer is washed three times with 10% Na ₂ S ₂ O ₃ solution and brine solution, dehydrated with anhydrous Na ₂ SO ₄ and filtered. The filtrate was concentrated and purified by column chromatography (3: 1 hexane / EtOAc) to give compound 13 as a pale red oil. NMR data of Compound 13 is as follows.

^{1 H NMR (300 MHz, CDCl} 3) δ 3.11 (m, 2H), 3.84 (s, 12H), 3.93 (dd, J = 9.4, 3.6 Hz, 2H), 4.31 (dd, J = 9.3, 7.2 Hz, 2H), 4.75 (d, J = 3.9 Hz, 2H), 4.99 (s, 4H), 6.57 (s, 4H), 7.28-7.51 (m, 10H); ¹³ C NMR (75 MHz, CDCl ₃ )? 54.3, 56.2, 72.0, 75.0, 86.0, 103.0, 127.8, 128.1, 128.4, 136.8, 137.8, 153.7.

(6) Preparation of (+) - Syringal Resinol (Formula 6)

Compound 13 is dissolved in ethanol / THF (1: 1) (9 mL) and the temperature is adjusted to 65 [deg.] C. Raney nickel (0.15 g) was added thereto, and the mixture was stirred at 65 ° C for 1 hour. After completion of the reaction was confirmed by TLC, the reaction solution was filtered using acetone and celite. The filtrate was concentrated and purified by column chromatography (1: 1 hexane / EtOAc) to obtain a pale yellow solid Respectively. The obtained compound is (8R, 8'R) - (+) - Syringalisinol can be confirmed from the following NMR data and [?] _D value of +40.9 ^o (c 0.1, CHCl ₃ ). In the following Test Examples, the (+) - Syringal resynol prepared above was used.

^{_{[α] D +40.9 o (c}} 0.1, CHCl 3); ^{1 H NMR (400 MHz, CDCl} 3) δ 3.10 (m, 2H), 3.90 (s, 12H), 3.90 (m, 2H), 4.28 (dd, J = 8.8, 6.8 Hz, 2H), 4.73 (d, J = 4.4 Hz, 2H), 5.48 (s, 2H), 6.59 (s, 2H); ^{13 C NMR (100 MHz, CDCl} 3) δ 54.4, 56.4, 71.8, 86.1, 102.7, 132.1, 134.3, 147.1.

[Preparation Example 3] Separation of (+) - Syringal resinol

The extract fraction GB-1 of Preparation Example 1 was concentrated and subjected to Sephadex LH-20 column choromatography using 50% water-soluble methanol. A fraction having a high content of SIRT1 activity and Sirling's resinol was fractionated and concentrated. The residue was subjected to preparative silica gel TLC using chloroform: methanol (10: 1) as a developing solvent. As a result, Rf The active fraction having a value of 0.67 was purified.

NMR spectroscopic analysis and database search were performed to identify the active compound isolated and purified as syringaresinol. Mass spectrometry was performed to confirm this, and ESI-mass was measured in a positive mode. As a result, [M + Na] + was observed at m / z 440.9 and [2M + na] + was observed at m / z 858.9, . As a result of NMR spectroscopic analysis, the same results as in Chemical Formula 6 were obtained. Thus, it was confirmed that the purified and purified compound was syringaresinol.

[Experimental Example 1] Effect of siringa ginsenoside and ginseng fruit extract on intestinal microorganism proliferation

1. Experimental animals

C57BL / 6 male mice at 28 weeks of age were purchased from a central laboratory animal and then were adapted for two months in the Pacific rim animal room environment and were tested at 36 weeks of age. The animals were fed a sterile diet (D12450B, Research Diets, New Brunswick, NJ, USA) and water ad libitum in a sterilized fiter-top cage ℃ and humidity of 50 ± 10% were induced, and illumination was automatically adjusted day and night at intervals of 12 hours. Animal management was treated according to the constraint IACUC (IACUC14-014).

2. Dietary siregarecein and ginseng fruit extract

(10 mg / kg), (+) - seringgreinol high concentration group (50 mg / kg), ginseng fruit low concentration group (100 mg / kg) and high concentration of ginseng fruit (500 mg / kg). The concentration unit is mg of sample per kg of rat weight. (+) - singalgreinol was dissolved in Methylcellulose and each sample was orally administered to the above concentrations once a day for 10 weeks once a day.

3. Analysis of the fungus in the feces of the large intestine

At week 46, sacrificed by cervical dislocation, DNA was separated from the feces by ChunLab analysis in dry state, and the 16 rRNA gene was amplified by PCR, followed by pyrosequencing, followed by EzTaxon Each microorganism was identified by BALSTN search using -e database and pairwiase global sequence alignment. The results are shown in Figs.

In the results shown in Fig. 2, in the case of Sirling's high concentration group and the high concentration group of ginseng fruit, the Lactobacillus sp. Strains were 17 times and 7 times, and the Bifidobacterium sp. In the high concentration group, 5 times more proliferation was increased.

 3, the strains of Staphylococcus spp., Desulfovibrio sp., Clostridium spp., And U. obtellium spp., Which are harmful bacteria, were significantly higher in the (+) - seringgreinol high concentration group and the high concentration group of ginseng fruit It can be confirmed that it is statistically decreased.

From the above results, it was confirmed that the oral administration of (+) - Syringalisinol and ginseng fruit extract significantly increased the lactic acid bacterial proliferation in Lactobacillus genus and Bifidobacterium genus in the intestines and remarkably inhibited the proliferation of harmful bacteria .

[Experimental Example 2] Effect of siring ginsenoside and ginseng fruit on the proliferation of Lactobacillus lutei

1. Strain used

Lactobacillus lutein KCTC 3594 was purchased from the Korea Biological Resources Center (KCTC).

2. Lactobacillus lutei proliferation assay

The number of generated colonies was counted, and the average number of colonies was multiplied by a dilution factor to calculate the number of viable cells per ml of the culture. (+) - singaleceinol or ginseng fruit extract was added to 5 ml of MRS broth (Difco, USA) at a concentration of 0 to 200 μg / ml and sterilized. Lactobacillus lutea was inoculated into sterile 5 ml MRS broth medium by adjusting the bacterial concentration so that the absorbance value at 660 nm was 0.3, and then 100 μl was inoculated. After incubation at 37 ° C for 6 hours and 24 hours, the number of bacteria was measured.

From the results shown in Fig. 4, the sering ginsenoside and ginseng fruit extract promoted the proliferation of Lactobacillus lutei in a concentration-dependent manner.

[Test Example 3] Improvement of ulcerative colitis in (+) - singeringisinol and ginseng fruit

1. Experimental animals

C57BL / 6 male mice at 28 weeks of age were purchased from a central laboratory animal and then adapted for 2 months in a pathogen-free facility animal room environment of Pacific Pharmaceuticals. The animals were fed a sterile diet (D12450B, Research Diets, New Brunswick, NJ, USA) and water ad libitum in a sterilized fiter-top cage ℃ and humidity of 50 ± 10% were induced, and illumination was automatically adjusted day and night at intervals of 12 hours. Animal management was treated according to the constraint IACUC (IACUC14-014).

2. The induction of ulcerative colitis with DSS and the extraction of syringalisinol and ginseng fruit extract

3% (W / V) DSS is fed for 8 days to induce ulcerative colitis. At this time, 10 rats per group were separated and administered orally for 8 days each in the DSS group (saline administration), seringgaresinol 50 mg / kg, and ginseng fruit 500 mg / kg once a day.

3. Measurement of disease activity index (DAI)

DAI was measured by lesions such as stool and diarrhea.

As can be seen from FIG. 5, ulcerative colitis was improved by about 50% in rats fed high (+) - seringgarisinol concentration compared to DSS. The ginseng fruit extract also showed that ulcerative colitis was improved by 20% in rats fed high concentration.

[Experimental Example 4] A method of isolating Lactobacillus luterium from the intestine of a rat fed with (+) - sialing reginol

In order to isolate the strain according to the invention, the sirens of recipient mice were inoculated into an MRS medium containing 0.02% sodium azide and incubated at 37 ° C for 24 hours. After culturing, the culture solution was taken with 10 μl of platinum, and the mixture was plated on a medium containing 0.02% sodium azide and cultured at 37 ° C. for 48 hours to isolate colloid, followed by bacterial pyrosequencing And confirmed that it was Lactobacillus lutea.

[Formulation Example 1] Tablets

(+) -Singalgreinol 100 mg, lactose 400 mg, corn starch 400 mg, and magnesium stearate 2 mg were mixed and tableted according to a conventional preparation method.

 [Formulation Example 2]

(+) - singalgreinol 100 mg, lactose 400 mg, corn starch 400 mg and magnesium stearate 2 mg were mixed and filled in gelatin capsules according to the conventional preparation method of capsules to prepare capsules.

[Formulation Example 3]

(+) - Syringalisinol (50 mg), anhydrous crystalline glucose (250 mg) and starch (550 mg) were mixed and granulated into granules using a fluidized bed granulator.

[Formulation Example 4] Drinking agent

50 mg of ginseng fruit extract containing (+) - seringgreinol, 10 g of glucose, 0.6 g of citric acid, and 25 g of liquid oligosaccharide are mixed and 300 ml of purified water is added to each bottle. And the mixture was sterilized at 130 DEG C for 4 to 5 seconds to prepare a drink.

[Formulation Example 5] Caramel formulation

(+) - Syringal lecinol 50 mg, corn syrup 1.8 g, defatted milk 0.5 g, soybean lecithin 0.5 g, butter 0.6 g, vegetable hardened oil 0.4 g, sugar 1.4 g, margarine 0.58 g, And then caramelized.

[Formulation Example 6] Health food

ingredient content (+) - Syringal Resinol 1000 mg Vitamin mixture Vitamin A acetate 70 [mu] g Vitamin E 1.0 mg Vitamin B1 0.13 mg Vitamin B2 0.15 mg Vitamin B6 0.5 mg Vitamin B12 0.2 [mu] g Vitamin C 10 mg Biotin 10 [mu] g Nicotinic acid amide 1.7 mg Folic acid 50 [mu] g Calcium pantothenate 0.5 mg Mineral mixture Ferrous sulfate 1.75 mg Zinc oxide 0.82 mg Magnesium carbonate 25.3 mg Potassium monophosphate 15 mg Dicalcium phosphate 55 mg Potassium citrate 90 mg Calcium carbonate 100 mg Magnesium chloride 24.8 mg

Although the composition ratio of the vitamin and the mineral mixture is relatively mixed with the ingredient suitable for health food, the compounding ratio of the vitamin and mineral mixture may be arbitrarily changed. The ingredients may be mixed according to a conventional method of manufacturing health food, And can be used in the manufacture of a health food composition according to a conventional method.

[Formulation Example 7] Health drinks

ingredient content (+) - Syringal Resinol 1000 mg Citric acid 1000 mg oligosaccharide 100 g Plum concentrate 2 g Taurine 1 g Purified water Balance Total volume 900 ml

As shown in Table 2, the remaining amount of purified water was added to a total volume of 900 ml, and the above components were mixed according to a conventional health drink manufacturing method. After stirring for about 1 hour at 85 캜, the solution was filtered Sterilized in a 2-liter container, sterilized by sealing, and stored in a refrigerator to prepare a health beverage composition.

Claims (13)

A composition for intestinal lactobacillus growth comprising a compound of the following formula (1), a derivative thereof or a pharmaceutically or pharmacologically acceptable salt thereof as an active ingredient:
≪ Formula 1 >

Wherein each of R ₁ , R ₂ , R ₃ and R ₄ is independently a linear or branched C ₁ to C ₁₈ alkyl group, a C ₁ to C ₁₈ alkenyl group, a C ₁ to C ₁₈ alkynyl group, C ₃ to C ₆ cyclic alkyl group. The method according to claim 1,
Wherein the lactic acid bacterium comprises at least one of a Lactobacillus sp. Strain and a Bifidobacterium sp. Strain. The method according to claim 1,
The lactic acid bacteria may be selected from the group consisting of Lactobacillus reuteri and Bifidobacterium ( Bifidobacterium < RTI ID = 0.0 > adolescentis ). < / RTI > The method according to claim 1,
Wherein the compound of Formula 1 is (+) - Syringaresinol. 5. The method of claim 4,
Wherein the (+) - Syringalisinol is a form contained in one or more extracts selected from the group consisting of flaxseed, yellowtail, ocher, sesame and ginseng. The method according to claim 1,
Wherein the composition is for improving diarrhea, enteritis or gastrointestinal disorders. The composition for intestinal lactobacillus growth according to any one of claims 1 to 6, which is administered simultaneously with Lactobacillus reuteri . The composition according to any one of claims 1 to 6, wherein the composition is a health food composition. The composition according to any one of claims 1 to 6, wherein the composition is a pharmaceutical composition. A kit comprising the composition for enhancing intestinal lactic acid bacteria according to any one of claims 1 to 6 and Lactobacillus reuteri . A method for producing a lactic acid bacterium comprising orally administering a composition according to any one of claims 1 to 6 to a mammal other than a human and extracting the intestinal lactic acid bacteria from the excrement of the mammal. 12. The method of claim 11,
Wherein said non-human mammal is a mouse. 12. The method of claim 11,
Wherein the lactic acid bacterium is Lactobacillus reuteri .

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