JP7281315B2 - GLP-1 secretagogue - Google Patents

Description

The present invention relates to a material that exerts a GLP-1 secretagogue effect.
Regarding.

In recent years, antidiabetic agents targeting incretins (Intestine Secretion Insulin) have been developed. Incretin is a general term for hormones that are secreted from the gastrointestinal tract along with intake of nutrients and act on pancreatic β cells to promote insulin secretion. Two gastrointestinal hormones, GLP-1 (glucagon-like peptide-1) and GIP (glucose-dependent insulinotropic polypeptide), are known as hormones responsible for incretin action. GLP-1 is mainly secreted from L cells present in the lower small intestine and colon, and GIP is secreted from K cells abundantly present in the duodenum in the upper small intestine.

Incretins secreted by the influx of nutrients into the gastrointestinal tract and stimulation increase insulin secretion from pancreatic β-cells and suppress glucagon secretion from pancreatic α-cells as blood glucose levels rise, working to lower blood sugar. . On the other hand, when the blood sugar level is low, insulin secretion decreases and glucagon secretion increases, resulting in an increase in blood sugar level. Of the two incretins, GLP-1 has the effect of suppressing glucagon secretion from pancreatic α cells. It has also been shown in clinical trials that GLP-1 stimulates insulin secretion in type 2 diabetics, whereas GIP does not.
In addition, in tissues other than the pancreas, GLP-1 has physiological effects different from those of GIP. Reported extrapancreatic effects of GLP-1 include increased hepatic/muscular glucose uptake, improved insulin resistance, appetite suppression in the central nervous system, and delayed gastric excretion in the gastrointestinal tract (Non-Patent Documents 1-10). 3). From this, it is possible that increasing the effect of GLP-1 works favorably on blood sugar regulation and at the same time leads to improvement of obesity.

However, since GLP-1 is a polypeptide, it is inactivated by the in vivo enzyme, DPP-4 (dipeptidyl peptidase-4), even after administration, and its half-life is extremely short at several minutes. It is very difficult to keep constant. Therefore, in order to increase the GLP-1 concentration in vivo for a long period of time, it is desirable to promote endogenous GLP-1 secretion rather than exogenous administration.
In previous studies, as substances that promote the secretion of GLP-1, fatty acids such as palmitic acid and oleic acid (Non-Patent Documents 4-7), glucose (Non-Patent Documents 8-9), meat hydrolysates, gastrin Releasing peptide, carbachol, forskolin, ionomycin, phorbol myristate acetate, essential amino acids, leucine, isoleucine, skim milk, casein, leptin, muscarinic receptors M1 and M2, quillaja, lysophosphatidylinositol, marjoram, wormwood, ferulic acid, yucca , sugar ceramide, β-carotene, plant sterol, beet, chrysanthemum, paprika, orange, ginseng, loquat tea, etc. have been reported.

On the other hand, the applicant has found that chlorogenic acids have the effect of promoting the secretion of GLP-1 (Non-Patent Document 10).
However, no report has been made on the effect of combined use of chlorogenic acids and fatty acids on GLP-1.

Baggio, Laurie L., and Daniel J. Drucker., Gastroenterology 132.6, 2007, p.2131-2157 Seino, Yutaka, Mitsuo Fukushima, and Daisuke Yabe., Journal of diabetes investigation 1.1-2, 2010, p.8-23 Eur J Pharmacol, 440(2-3), 2002, p.269-279 Li Y, Kokrashvili Z, Mosinger B, Margolskee RF. Gustducin couples fatty acid receptors to GLP-1 release in colon. Am J Physiol Endocrinol Metab. 2013, 304(6), E651-60 Hirasawa, Akira, et al. "Free fatty acids regulate gut incretin glucagon-like peptide-1 secretion through GPR120." Nature medicine 11.1, 2005, p.90-94 Iakoubov, Roman, et al. "Protein kinase Cζ is required for oleic acid-induced secretion of glucagon-like peptide-1 by intestinal endocrine L cells." Endocrinology 148.3, 2007, p.1089-1098 Kaji, Izumi, Shin-ichiro Karaki, and Atsukazu Kuwahara. "Short-chain fatty acid receptor and its contribution to glucagon-like peptide-1 release." Digestion 89.1, 2014, p.31-36 Ritzel, U., et al. "Release of glucagon-like peptide-1 (GLP-1) by carbohydrates in the perfused rat ileum." Acta diabetologica 34.1, 1997, p.18-21 Kong, Marie-France, et al. "Effects of oral fructose and glucose on plasma GLP-1 and appetite in normal subjects." Peptides 20.5, 1999, p.545-551 Jokura, Hiroko, et al., Nutrition research 35(10), 2015, p.873-881

The present invention relates to providing a GLP-1 secretagogue that can be used in pharmaceuticals, foods, and the like. The present invention also relates to providing a composition capable of promoting GLP-1 secretion.

The present inventors investigated materials that promote the secretion of GLP-1, and found that the combination of chlorogenic acids and fatty acids increased the stimulation of the lower L cells of the small intestine and promoted the secretion of GLP-1 more than using them alone. It was found that the action was enhanced.

That is, the present invention relates to the following 1) to 9).
1) A GLP-1 secretagogue containing (A) chlorogenic acids and (B) fatty acid as active ingredients.
2) (A) GLP-1 secretagogue enhancers of fatty acids in the lower small intestine containing chlorogenic acids as active ingredients.
3) (B) GLP-1 secretagogue-enhancing agent of chlorogenic acids in the lower small intestine containing fatty acid as an active ingredient.
4) A food composition for promoting GLP-1 secretion comprising (A) chlorogenic acids and (B) fatty acid as active ingredients.
5) (A) A food composition for enhancing the GLP-1 secretion-promoting action of fatty acids in the lower small intestine, containing chlorogenic acids as an active ingredient.
6) (B) A food composition for enhancing the GLP-1 secretion promoting action of chlorogenic acids in the lower small intestine, containing a fatty acid as an active ingredient.
7) A method of enhancing the GLP-1 secretagogue effect of fatty acids in the lower small intestine with (A) chlorogenic acids (excluding medical practice for humans).
8) A method of enhancing the GLP-1 secretagogue effect of chlorogenic acids in the lower small intestine with (B) fatty acid (excluding medical practice for humans).
9) A composition containing (A) chlorogenic acids and (B) a fatty acid, which has a coating layer that coats at least (B) the fatty acid and dissolves in the lower small intestine.

INDUSTRIAL APPLICABILITY According to the present invention, it is possible to increase the stimulation of L cells in the lower small intestine and effectively promote the secretion of GLP-1.

GLP-1 secretion amount in the chlorogenic acids added group. GLP-1 secretion levels in the fatty acid-added group, the chlorogenic acids group, and the fatty acid-added group. GLP-1 secretion levels in the glucose-added group, the chlorogenic acids group, and the glucose-added group.

In one embodiment, the present invention provides a GLP-1 secretagogue and a GLP-1 secretagogue food composition comprising (A) chlorogenic acids and (B) fatty acid as active ingredients. In another embodiment, the present invention provides (A) an agent for enhancing the GLP-1 secretion-enhancing effect of fatty acids in the lower small intestine and a food composition for enhancing GLP-1 secretion-enhancing effect, containing chlorogenic acids as active ingredients. In another embodiment, the present invention provides (B) a GLP-1 secretagogue enhancer of chlorogenic acids in the lower small intestine containing a fatty acid as an active ingredient, and a food composition for enhancing GLP-1 secretagogue. . In another embodiment, the present invention provides a method for enhancing the GLP-1 secretagogue effect of fatty acids in the lower small intestine with (A) chlorogenic acids. Furthermore, in another embodiment, the present invention provides a method for enhancing the GLP-1 secretagogue effect of chlorogenic acids in the lower small intestine with (B) fatty acids.

As used herein, the term “chlorogenic acids” refers to monocaffeoylquinic acids such as 3-caffeoylquinic acid, 4-caffeoylquinic acid and 5-caffeoylquinic acid, 3-feruloylquinic acid and 4-ferroylquinic acid. It is a collective term for monoferuloylquinic acids such as ruquinic acid and 5-feruloylquinic acid. In the present invention, at least one of the above six types may be contained. Chlorogenic acids have stereoisomers, and in the present invention, pure stereoisomers thereof or mixtures of those stereoisomers can be used.
In the present specification, the content of (A) chlorogenic acids is defined based on the total amount of the above six types. In addition, when chlorogenic acids are salts or hydrates, the content of chlorogenic acids is a value converted into chlorogenic acids, which are free acids. Chlorogenic acids can be analyzed, for example, by an analysis method suitable for the situation of the sample to be measured among commonly known measurement methods. For example, it can be analyzed by the method described in Examples below.

(A) Chlorogenic acids may be in free form, salt or hydrate form. Salting can improve water solubility and increase physiological effectiveness. Salts of chlorogenic acids may be pharmaceutically acceptable salts. Examples of base substances for forming such salts include alkali metal hydroxides such as lithium hydroxide, sodium hydroxide and potassium hydroxide; alkaline earth metal hydroxides such as magnesium hydroxide and calcium hydroxide; inorganic bases such as ammonium hydroxide; basic amino acids such as arginine, lysine, histidine and ornithine; organic bases such as monoethanolamine, diethanolamine and triethanolamine. Among these, hydroxides of alkali metals or alkaline earth metals are preferred.

(A) Chlorogenic acids can be extracted from natural products containing them, particularly plants, and can also be industrially produced by chemical synthesis. In the present invention, a plant extract containing chlorogenic acids can also be used as (A) chlorogenic acids.
Examples of plants containing chlorogenic acids include coffee beans, cabbage, lettuce, artichokes, tomatoes, eggplants, potatoes, carrots, apples, pears, plums, peaches, apricots, cherries, sunflower seeds, mulukhiyah, kansho, nandin. leaves, blueberries, wheat and the like. Among them, coffee beans are preferable from the viewpoint of the content of chlorogenic acids. The bean species and production area of the coffee beans are not particularly limited.
The coffee beans may be green coffee beans, roasted coffee beans, or both. From the viewpoint of the content of chlorogenic acids, the roasted coffee beans are preferably lightly roasted coffee beans. The L value of the lightly roasted coffee beans is preferably 27 or more, more preferably 29 or more, still more preferably 35 or more, still more preferably 40 or more, from the viewpoint of chlorogenic acid content, etc., and less than 62 from the viewpoint of flavor. is preferred, 60 or less is more preferred, and 55 or less is even more preferred. Here, the "L value" as used herein means that black is L value 0 and white is L value 100, and the lightness of roasted coffee beans is measured by a color difference meter (for example, spectrophotometer SE2000, Co., Ltd.) after pulverization. ) manufactured by Nippon Denshoku Co., Ltd.).
The extraction method and extraction conditions can be selected as appropriate. For example, water, hot water, or a water-soluble organic solvent can be used by known methods such as batch extraction, drip extraction, and column extraction. Examples of water-soluble organic solvents include lower alcohols such as ethanol. For the extraction method, for example, the methods described in JP-A-58-138347, JP-A-59-51763, JP-A-62-111671, and JP-A-5-236918 can be employed. can. The coffee beans subjected to extraction may be unground or ground.
After the extraction, the obtained extract may be concentrated by known means such as normal pressure concentration, vacuum concentration, membrane concentration, etc. Alternatively, the extract or concentrate may be purified. Drying can be performed by known means such as spray drying and freeze drying.

(B) Fatty acid is preferably a linear or branched saturated fatty acid or unsaturated fatty acid having 2 to 22 carbon atoms. (B) Fatty acids include, for example, butanoic acid (butyric acid), pentanoic acid (valeric acid), hexanoic acid (caproic acid), heptanoic acid (enanthic acid), tetradecanoic acid (myristic acid), pentadecanoic acid (pentadecylic acid), hexadecanoic acid (palmitic acid), heptadecanoic acid (margaric acid), octadecanoic acid (stearic acid), 9-hexadecenoic acid (palmitoleic acid), cis-9-octadecenoic acid (oleic acid), 11-octadecenoic acid (vaccenic acid), cis,cis-9,12-octadecadienoic acid (linoleic acid), 9,12,15-octadecantrienoic acid (α-linolenic acid), 6,9,12-octadecatrienoic acid (γ-linolenic acid), 9,11,13-octadecatrienoic acid (eleostearic acid), 8,11-eicosadienoic acid, 5,8,11-eicosatrienoic acid (mead acid), 5,8,11-eicosatetraenoic acid (arachidonic acid), eicosapentaenoic acid, and docosahexaenoic acid. Two or more kinds of fatty acids may be combined. Among them, from the viewpoint of promoting GLP-1 secretion, those having 14 to 22 carbon atoms are preferable, and those having 16 to 18 carbon atoms are more preferable. Stereoisomers exist in fatty acids, and in the present invention, pure stereoisomers thereof or mixtures of those stereoisomers can be used.

(B) The fatty acid may be in free form or in salt form. Salting can improve water solubility and increase physiological effectiveness. Salts of fatty acids may be pharmaceutically acceptable salts. Examples of base substances for forming such salts include alkali metal hydroxides such as lithium hydroxide, sodium hydroxide and potassium hydroxide; alkaline earth metal hydroxides such as magnesium hydroxide and calcium hydroxide; Inorganic bases such as ammonium hydroxide; Basic amino acids such as arginine, lysine, histidine, ornithine; Organic bases such as monoethanolamine, diethanolamine, and triethanolamine. Metal hydroxides are preferred.

(B) Fatty acids may be derived from simple lipids and complex lipids.
Simple lipids are esters produced from fatty acids and alcohols, and include esters of fatty acids and lower alcohols such as methanol and ethanol, ester waxes of higher alcohols having 8 to 24 carbon atoms and fatty acids, and esters of glycerin and fatty acids. Glycerides (monoglycerides, diglycerides, triglycerides) and the like.
A complex lipid is a lipid containing phosphorus, nitrogen, etc. in addition to a simple lipid, and includes phospholipids, glycolipids, and the like.

(B) Fatty acids can be extracted from natural products containing them, particularly lipids, and can also be industrially produced by chemical synthesis.
Examples of lipids include natural oils such as soybean oil, corn oil, rapeseed oil, palm oil, palm kernel oil, olive oil, safflower oil, perilla oil, fish oil, beef tallow, lard, and butter, as well as many oils. You can get it from food. Examples of foods rich in fats and oils include seeds, nuts, fish meat, beef, milk, fresh cream, cheese, confectionery and the like.

As shown in Examples below, the combination of (A) chlorogenic acids and (B) fatty acids has the effect of increasing the amount of GLP-1 secreted from L cells. Moreover, the action is superior to the action of (A) chlorogenic acids and (B) fatty acids alone, and it can be said that the action is synergistic, exceeding the addition of the effects when they are added alone. On the other hand, (A) the combination of chlorogenic acids and glucose did not enhance the GLP-1 secretagogue effect.
Thus, even if a single component having an effect of increasing GLP-1 secretion is used in combination, the secretagogue effect is not always enhanced. 1 secretagogue and can be used to manufacture the same. In addition, (A) chlorogenic acids can be (B) a GLP-1 secretagogue enhancer of fatty acids for enhancing the GLP-1 secretagogue of fatty acids, and can be used for producing the same. . Furthermore, the (B) fatty acid can be (A) an agent for enhancing the GLP-1 secretagogue effect of chlorogenic acids for enhancing the GLP-1 secretagogue effect of chlorogenic acids, and can be used for producing the same. can.
In addition, when administered or ingested to animals including humans, (A) chlorogenic acids and (B) fatty acids can thereby promote GLP-1 secretion, and (A) chlorogenic acids can thereby promote fatty acid production. (B) Fatty acids can enhance the GLP-1 secretagogue of chlorogenic acids.
Furthermore, a combination of (A) chlorogenic acids and (B) fatty acids can be used to promote GLP-1 secretion, and (A) chlorogenic acids enhance the GLP-1 secretagogue effect of (B) fatty acids. (B) fatty acids can be used to enhance the GLP-1 secretagogue of (A) chlorogenic acids.
A combination of (A) chlorogenic acids and (B) fatty acids controls various conditions through the action of promoting GLP-1 secretion, for example, for the prevention and/or improvement of lifestyle-related diseases such as obesity and diabetes, It can be used to increase glucose uptake in the liver and muscles, to improve insulin resistance, to suppress appetite in the central nervous system, and to delay gastric emptying in the gastrointestinal tract.

As used herein, “GLP-1 secretion promotion” means promotion of GLP-1 secretion in vivo. The present invention is suitable for promoting GLP-1 secretion from L cells in the lower small intestine. "GLP-1 secretion promotion" refers to promotion of blood GLP-1 concentration increase accompanying GLP-1 secretion in vivo, maintenance of elevated GLP-1 concentration, or inhibition of reduction of elevated GLP-1 concentration, namely This is a concept that includes both stabilization of blood GLP-1 concentration.
"Use" means administration or ingestion to animals, including humans, and does not include medical procedures, i.e., methods of surgery, treatment or diagnosis of humans, more specifically physicians or physician's instructions. It is a concept that does not include the method by which a recipient performs surgery, therapy or diagnosis on humans.

In the present invention, either (A) chlorogenic acids or (B) fatty acid may be administered first or may be administered simultaneously. When the two drugs are not administered simultaneously, the administration interval between the two drugs can be appropriately selected as long as the effect of enhancing the GLP-1 secretagogue of (A) chlorogenic acids or (B) fatty acid is exhibited.
(A) chlorogenic acids and (B) fatty acids in combination, (A) chlorogenic acids and (B) fatty acids may be formulated into one dosage form as a compounding agent, or formulated separately. It may be a kit that can be used simultaneously or separately with an interval.

The GLP-1 secretagogue of the present invention, the fatty acid-based GLP-1 secretagogue-enhancing agent, and the chlorogenic acid-based GLP-1 secretagogue-enhancing agent (hereinafter also referred to as "GLP-1 secretagogue, etc.") are human Pharmaceuticals, quasi-drugs, foods or It can be used as a feed, and can also be a material or preparation used by being mixed with the drug, quasi-drug, food, or feed.

The dosage form of the pharmaceuticals (hereinafter also quasi-drugs) may be solid, semi-solid, or liquid, such as granules, powders, tablets, capsules, microcapsules, powders, lozenges, and liquids. , oral administration such as syrups; parenteral administration such as injections, suppositories, inhalants, percutaneous absorption agents, and external preparations. Oral administration and intra-intestinal administration are preferred. The size of the form can be arbitrarily adjusted according to the purpose of use.
Formulations of such various dosage forms may optionally contain pharmaceutically acceptable carriers such as excipients, binders, fillers, disintegrants, surfactants, lubricants, dispersants, Buffers, osmotic pressure adjusters, pH adjusters, emulsifiers, preservatives, stabilizers, preservatives, thickeners, fluidity improvers, flavoring agents, foaming agents, fragrances, coating agents, diluents, etc., and chlorogen It can be prepared by appropriately combining medicinal ingredients other than acids and fatty acids.

The above-mentioned foods include foods (foods for specified health uses, foods with function claims) which are claimed to promote GLP-1 secretion and are permitted to be labeled to that effect as necessary. Examples of indications are "improves insulin resistance", "improves HbA1C", and "helps lower fasting blood sugar". Foods for which function labeling is permitted can be distinguished from general foods.
The form of food may be solid, semi-solid or liquid, and various food compositions (breads, cakes, noodles, confectionery, jellies, frozen foods, ice creams, dairy products, shakes, beverages, etc.). Furthermore, nutritional supplement compositions in the same form as the oral formulations described above (solid formulations such as granules, powders, tablets, capsules, microcapsules, and lozenges) are also included. Beverages include soft drinks, tea drinks, coffee drinks, fruit juice drinks, carbonated drinks and the like. The size of the form can be arbitrarily adjusted according to the purpose of use.
Foods in various forms may optionally contain other food ingredients such as solvents, softeners, oils, emulsifiers, preservatives, acidulants, sweeteners, bittering agents, pH adjusters, stabilizers, colorants, Antioxidants, humectants, thickeners, fluidity improvers, foaming agents, fragrances, seasonings, etc., and active ingredients other than chlorogenic acids and fatty acids can be appropriately combined for preparation.

The form of the feed is preferably in the form of pellets, flakes, or mash. Examples include pet food for dogs, cats, small birds, and the like.
If necessary, the feed may contain other feed ingredients such as meat, protein, grains, rice bran, lees, sugars, vegetables, vitamins, minerals, gelling agents, shape retention agents, pH adjusters, Seasonings, preservatives, nutritional supplements, etc., and active ingredients other than chlorogenic acids and fatty acids can be appropriately combined and prepared by a conventional method.

In one embodiment, the present invention provides a composition containing (A) chlorogenic acids and (B) a fatty acid, which has a coating layer that coats at least (B) the fatty acid and dissolves in the lower small intestine. offer. The composition may be in the form of pharmaceuticals, quasi-drugs, or foods. Here, (A) chlorogenic acids and (B) fatty acids are the same as those described above as the forms of pharmaceuticals, quasi-drugs, and foods.
In general, fatty acids are absorbed in the upper part of the small intestine, so administration or ingestion through the upper part of the small intestine sufficiently promotes the secretion of GLP-1 by fatty acids in the lower part of the small intestine where L cells are highly localized. can't In contrast, the presence of a capsular layer in the composition of the present invention can result in the release of (B) fatty acids in the lower small intestine where L cells are highly localized and thus absorbed in the gastrointestinal tract in the lower small intestine. The coexistence of (A) chlorogenic acids and (B) fatty acids, which are difficult to digest, increases the stimulation of L cells by both, and is expected to enhance the effect of promoting GLP-1 secretion. Therefore, the composition of the present invention can be suitably used for promoting GLP-1 secretion.
The "coating layer that dissolves in the lower part of the small intestine" in the present invention may be a general enteric coating layer that does not dissolve until it reaches the lower part of the small intestine and dissolves in the lower part of the small intestine. From the viewpoint of targeted delivery, it is preferable to design the drug in consideration of the physical/physiological environment in the human gastrointestinal tract and the travel time in the gastrointestinal tract (for example, sustained release type, sustained release type, timed release type, etc.). Furthermore, it is preferred to design with a saliva- and/or gastric juice-resistant outer layer and an inner layer that controls the release of the active ingredient.
Examples of components constituting the coating layer include chitosan, hydroxypropylmethylcellulose acetate succinate, gelatin, starch, starch derivatives, cellulose, cellulose derivatives, waxes, hardened oils and the like. The coating layer can be provided with flavoring substances, flavoring agents, sweeteners, coloring agents, etc., as required.

The content of (A) chlorogenic acids in the GLP-1 secretagogue and the like of the present invention and the composition of the present invention is determined by the ease of administration or intake, saliva, gastric juice, pancreatic juice, and bile before reaching the small intestine and acting. , Considering that it is diluted with intestinal juice etc., it is preferably 0.00001 mass% or more, more preferably 0.00002 mass% or more, still more preferably 0.00005 mass% or more, still more preferably 0.0001 mass% % or more, preferably 1% by mass or less, more preferably 0.5% by mass or less, even more preferably 0.2% by mass or less, and even more preferably 0.1% by mass or less.

The content of (B) fatty acid in the GLP-1 secretagogue and the like of the present invention and the composition of the present invention depends on the ease of administration or intake, saliva, gastric juice, pancreatic juice, bile, Considering dilution with intestinal juice or the like, it is preferably 0.001% by mass or more, more preferably 0.002% by mass or more, still more preferably 0.005% by mass or more, and even more preferably 0.008% by mass. The content is preferably 1% by mass or less, more preferably 0.5% by mass or less, even more preferably 0.2% by mass or less, and even more preferably 0.1% by mass or less. When the fatty acid is a salt, the content of the fatty acid is a value converted to the fatty acid that is a free acid.

In the GLP-1 secretagogue and the like of the present invention and the composition of the present invention, the ratio of (A) chlorogenic acids and (B) fatty acid is the mass ratio [(A)/( B)] is preferably 0.001 or more, more preferably 0.005 or more, still more preferably 0.01 or more, and is preferably 10 or less, more preferably 5 or less, and still more preferably 1 or less. .

The dosage or intake of the GLP-1 secretagogue of the present invention, etc., and the dosage or intake of the composition of the present invention vary according to the body weight, sex, age, condition, or other factors of the subject of administration or intake. obtain. The dosage, route, interval of administration, and the amount and interval of intake can be appropriately determined by those skilled in the art. Considering that it is diluted by such as, usually per day per adult (60 kg), (A) as chlorogenic acids, preferably 10 mg or more, more preferably 50 mg or more, still more preferably 100 mg or more Also, it is preferably 30000 mg or less, more preferably 10000 mg or less, and still more preferably 5000 mg or less.
In addition, (B) fatty acid is preferably 10 mg or more, more preferably 50 mg or more, still more preferably 100 mg or more, and preferably 30000 mg or less, more preferably 1 adult (60 kg) per day. It is 10000 mg or less, more preferably 5000 mg or less.
In the present invention, it is preferred to administer or take such an amount once or several times a day, preferably once a day.

The GLP-1 secretagogue, etc. of the present invention and the composition of the present invention can be administered or taken according to any schedule.
The period of administration or intake is not particularly limited, but repeated and continuous administration or intake is preferable, and continuous administration or intake for 2 weeks or more, further 4 weeks or more, furthermore 8 weeks or more is preferable.

The GLP-1 secretagogue and the like of the present invention and the composition of the present invention are preferably administered or ingested at the time of eating/feeding, before eating/feeding, or after eating/feeding. More preferably, it is administered or taken within 10 minutes after eating/feeding. The contents of eating (feeding) are not particularly limited, and include meals or feeds containing proteins, carbohydrates and lipids, but preferred are meals or feeds containing carbohydrates and lipids.
Subjects for administration or intake are not particularly limited as long as they are humans or non-human animals that require or desire it. Preferred examples of subjects include simple obese people, diabetic patients, and those at risk.

(1) Preparation of test sample solution Lightly roasted coffee bean extracts were obtained by extracting ground lightly roasted coffee beans with hot water, treating in a column using activated carbon, and spray drying. The resulting lightly roasted coffee bean extract had a chlorogenic acid (6 types) content of 24.2% by mass and a caffeine content of 0.1% by mass.
HPLC (manufactured by Hitachi, Ltd.) was used to quantify chlorogenic acids and caffeine. In HPLC, 1 g of a sample was accurately weighed, diluted to 10 mL with an eluent, filtered through a membrane filter (GL Chromatodisc 25A, pore size 0.45 μm, GL Sciences Inc.), and subjected to analysis. Using 5-caffeoylquinic acid and caffeine as standard substances, 3-caffeoylquinic acid, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3-feruloylquinic acid, 4-feruloylquinic acid, 5 - Feruloylquinic acid and caffeine were quantified.

Krebs-Ringer Bicarbonate (KRB) Buffer (SIGMA-ALDRICH) to which CaCl ² (Fujifilm Wako Pure Chemical Industries, Ltd., final concentration 1 mM) solution and DPP IV Inhibitor (Merck Millipore, 2 mass% added) were added , dissolved the test sample. The test sample solution was prepared on the day of the test and kept at 37°C until just before the test.
The chlorogenic acid-containing test sample solution used in the experiment was adjusted to have a chlorogenic acid concentration of 3 μM and 300 μM, and was subjected to ultrasonic treatment for 30 minutes for dissolution. The fatty acid-containing test sample solution contains 0.4 mM oleic acid sodium salt (SIGMA-ALDRICH) and bovine serum albumin fraction v, Fatty acid free from bovine serum (SIGMA-ALDRICH) at a final concentration of 0.2%. A test sample solution was prepared containing 0.1% and sonicated for 30 minutes to dissolve. A glucose-containing test sample solution was prepared by adjusting D-(+)-Dextrose (Tokyo Chemical Industry Co., Ltd.) to a final concentration of 250 mM. As a control, a test sample solution containing no chlorogenic acids, fatty acids, or glucose was prepared.

(2) Cell Culture An experiment was performed using NCI-H716 cells (epithelial cells derived from human cecal adenocarcinoma, see Table 1) (American Type Culture Collection), which is an in vitro model of intestinal L cells. Cells were seeded in 75 cm ² flasks and cultured at 37° C. in the presence of 5% CO ² . RPMI1640 (containing 10% fetal bovine serum, high glucose) medium (Invitrogen) was used as the growth medium. DMEM (containing 10% fetal bovine serum, high glucose) medium (Invitrogen) was used to induce cell differentiation. A 48-well plate was coated with Matrigel (BD) at 60 μl/well, incubated at 37° C. for 30 minutes, and then seeded with cells at 2.5×10 ⁵ cells/500 μl/well. It was cultured and differentiated for days.

(3) GLP-1 secretion stimulation experiment The cells differentiated as described above were washed twice with 600 μL of KRB medium warmed to 37° C., replaced with 250 μL of the test sample solution, and treated at 37° C., 5% CO ^{2 .} Incubated in presence for 2 hours. The supernatant was collected in a microtube containing 10 μL of an aprotinin solution (Fuji Film Wako Pure Chemical Industries, Ltd., adjusted to 6500 KIU/mL) and centrifuged (5000 r/min, 5 minutes, 4° C.). The supernatant was recovered, and GLP-1 was quantified using GLUCAGON LIKE PEPTIDE-1 (ACTIVE) ELISA, 96-WELL PLATE (Merck Millipore). GLP-1 was measured according to the protocol of the kit, and the sample was diluted 10-fold with the attached Assay buffer.
For the test of significant difference, comparison between groups was evaluated by t-test. The significance level was all 5%, and statistical significance was determined when the P value fell below the significance level (*p<0.05, **p<0.01, vs. control). Microsoft Excel was used for the test of significance. The obtained numerical values are shown as mean±standard error.

(4) Results The quantification results of GLP-1 after adding the test sample solution are shown in FIGS. 1 to 3. FIG. Here, the amount of GLP-1 secreted is shown as a relative value to the amount of GLP-1 secreted when the control is added, with the amount of GLP-1 secreted when the control is added as "1".
The amount of GLP-1 secreted by adding 300 μM of chlorogenic acids was increased by about 15% compared to the control (Fig. 1). Fatty acid-induced GLP-1 secretion was increased by about 49% compared to the control (Fig. 2), and glucose-induced GLP-1 secretion was increased by about 65% compared to the control (Fig. 3). ). On the other hand, when chlorogenic acids and glucose were added together, the amount of GLP-1 secreted increased by about 51% compared to the control (Fig. 3). Therefore, no effect on GLP-1 secretion was observed by combining chlorogenic acids and glucose. On the other hand, when chlorogenic acids and fatty acids were added in combination, the amount of GLP-1 secretion increased by about 171% compared to the control. The effect on -1 secretion appeared to be synergistic.

Claims (9)

(A) chlorogenic acids that are 3-caffeoylquinic acid, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3-feruloylquinic acid, 4-feruloylquinic acid and 5-feruloylquinic acid and ( B) A GLP-1 secretagogue containing oleic acid as an active ingredient. (A) Effective chlorogenic acids that are 3-caffeoylquinic acid, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3-feruloylquinic acid, 4-feruloylquinic acid and 5-feruloylquinic acid GLP-1 secretagogue enhancer of oleic acid in the lower small intestine as a component. (B) 3-caffeoylquinic acid, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3-feruloylquinic acid, 4-feruloylquinic acid and 5-caffeoylquinic acid in the lower small intestine containing oleic acid as an active ingredient A GLP-1 secretagogue enhancer of chlorogenic acids , which are feruloylquinic acids . (A) chlorogenic acids that are 3-caffeoylquinic acid, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3-feruloylquinic acid, 4-feruloylquinic acid and 5-feruloylquinic acid and ( B) A food composition for promoting GLP-1 secretion containing oleic acid as an active ingredient. (A) Effective chlorogenic acids that are 3-caffeoylquinic acid, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3-feruloylquinic acid, 4-feruloylquinic acid and 5-feruloylquinic acid A food composition for enhancing the GLP-1 secretagogue effect of oleic acid in the lower small intestine as a component. (B) 3-caffeoylquinic acid, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3-feruloylquinic acid, 4-feruloylquinic acid and 5-caffeoylquinic acid in the lower small intestine containing oleic acid as an active ingredient A food composition for enhancing the GLP-1 secretagogue effect of chlorogenic acids , which are feruloylquinic acids . (A) 3-caffeoylquinic acid , 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3-feruloylquinic acid, 4-feruloylquinic acid and 5-feruloylquinic acid, chlorogenic acids (excluding medical procedures for humans). GLP of chlorogenic acids 3-caffeoylquinic acid, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3-feruloylquinic acid, 4-feruloylquinic acid and 5-feruloylquinic acid in the lower small intestine -1 A method of enhancing the secretagogue by (B) oleic acid (excluding medical treatment for humans). (A) chlorogenic acids that are 3-caffeoylquinic acid, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3-feruloylquinic acid, 4-feruloylquinic acid and 5-feruloylquinic acid and ( B) A composition containing oleic acid and having at least (B) a coating layer that coats the oleic acid and dissolves in the lower small intestine.

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