JP6866354B2 - Tie2 activation composition - Google Patents

Description

The present invention relates to a Tie2 activation composition and the like. More specifically, the present invention contains, as an active ingredient, one or more components selected from the group consisting of novel iridoid compounds (Compound A and Compound B), oleanoside A, and kaempferol. The present invention relates to a Tie2 activation composition, a vascular permeability suppressing composition, a blood vessel maturation composition, a blood vessel normalization composition, a blood vessel stabilization composition, and a lymphatic vessel stabilization composition. The present invention also relates to novel iridoid compounds (Compound A and Compound B) and compositions containing the compounds.

Olive (Olea europaea) is a plant belonging to the genus Olive of the Oleaceae family, and is widely cultivated in the Mediterranean region and its fruits, and its fruits are widely used worldwide for olive oil extraction and edible use.

Olive fruits are rich in polyphenols such as hydroxytyrosol, acteoside, and oleuropein (Non-Patent Documents 1 and 2), and olive extracts and their polyphenol components have an arteriosclerosis inhibitory effect (Non-Patent Document 3). ), High blood pressure suppressing action (Patent Document 1), bone weight loss suppressing action (Non-Patent Document 4) and the like.

In recent years, it has been found that aging and functional deterioration of blood vessels affect skin properties such as wrinkles and rough skin, and it is known that the normality of blood vessels is effective as an index for determining skin properties. These blood vessels not only send oxygen and nutrients to organs throughout the body through blood, but also carry waste products from the tissues and excrete them out of the body, which is an important tissue for the human body to maintain vital activities. .. In general, aging of blood vessels and disruption of function have a great effect on human health such as arteriosclerosis, and also lead to external findings such as wrinkles and swelling. In recent years, attention has been paid to the activation of the receptor tyrosine kinase Tie2 (Tyrosine kinase with Ig and ECF homology domain 2) for the aging of blood vessels. It is known that when Tie2 is activated by angiopoietin-1 from the parietal cells of blood vessels, adhesion between cells of vascular endothelium is induced, which contributes to stabilization of vascular endothelial cells. In recent years, ingredients and foods having this Tie2 activating effect have been found, and are used in supplements, beverages, etc., for the purpose of cosmetology, prevention of swelling, prevention of poor circulation, etc. (Patent Documents 2 to 4).

Japanese Unexamined Patent Publication No. 2005-334022 Japanese Unexamined Patent Publication No. 2014-09977 Japanese Unexamined Patent Publication No. 2013-241356 Japanese Unexamined Patent Publication No. 2011-102272

J. Agric. Food Chem., Vol.52, pp479-484, 2004 J. Agric. Food Chem., Vol.53, pp8963-8969, 2005, Atherosclerosis, vol.188, No.1, pp35-42, 2006 Clin. Nutr., vol.25, No.5, 859-868, 2006

An object of the present invention is to provide a composition having a Tie2 activating effect, a vascular permeability suppressing effect, a blood vessel maturation effect, a blood vessel normalizing effect, a blood vessel stabilizing effect, a lymphatic vessel stabilizing effect, and the like. The purpose is.

In order to solve the above problems, the present inventors conducted diligent research, and as a result, among the components isolated from the olive fruit extract, oleanoside A and kenferol have a Tie2 activating effect, a vascular permeability suppressing effect, and a blood vessel. It was newly found that it has a maturation action, a blood vessel normalization action, a blood vessel stabilizing action, a lymphatic vessel stabilizing action, and the like. Furthermore, novel iridoid compounds (Compound A and Compound B) were found from the olive fruit extract, and the novel compounds also had a Tie2 activating effect, a vascular permeability suppressing effect, a blood vessel maturation effect, a blood vessel normalizing effect, and a blood vessel. It has been clarified that it has a stabilizing action, a lymphatic vessel stabilizing action, and the like, and the present invention has been completed.

（２）前記（Ｉ）で表される化合物、前記（ＩＩ）で表される化合物、前記（ＩＩＩ）で表される化合物（Oleanoside A）、前記（ＩＶ）で表される化合物（Kaempferol）、及びそれらの塩からなる群から選択される１以上の成分を含有する、血管透過性抑制用組成物。
（３）前記（Ｉ）で表される化合物、前記（ＩＩ）で表される化合物、前記（ＩＩＩ）で表される化合物（Oleanoside A）、前記（ＩＶ）で表される化合物（Kaempferol）、及びそれらの塩からなる群から選択される１以上の成分を含有する、血管の成熟化用組成物。
（４）前記（Ｉ）で表される化合物、前記（ＩＩ）で表される化合物、前記（ＩＩＩ）で表される化合物（Oleanoside A）、前記（ＩＶ）で表される化合物（Kaempferol）、及びそれらの塩からなる群から選択される１以上の成分を含有する、血管の正常化用組成物。
（５）前記（Ｉ）で表される化合物、前記（ＩＩ）で表される化合物、前記（ＩＩＩ）で表される化合物（Oleanoside A）、前記（ＩＶ）で表される化合物（Kaempferol）、及びそれらの塩からなる群から選択される１以上の成分を含有する、血管の安定化用組成物。
（６）前記（Ｉ）で表される化合物、前記（ＩＩ）で表される化合物、前記（ＩＩＩ）で表される化合物（Oleanoside A）、前記（ＩＶ）で表される化合物（Kaempferol）、及びそれらの塩からなる群から選択される１以上の成分を含有する、リンパ管の安定化用組成物。
（７）血管透過性抑制作用、血管の成熟化作用、血管の正常化作用、血管の安定化作用、又はリンパ管の安定化作用が、Ｔｉｅ２活性化作用に起因するものである、（２）〜（６）のいずれかに記載の組成物。
（８）食品組成物である、（１）〜（７）のいずれかに記載の組成物。
（９）飲料組成物である、（１）〜（７）のいずれかに記載の組成物。
（１０）Ｔｉｅ２を活性化することにより発揮される機能、血管透過性抑制により発揮される機能、血管の成熟化により発揮される機能、血管の正常化により発揮される機能、血管の安定化により発揮される機能、及びリンパ管安定化により発揮される機能からなる群から選択される１以上の機能の表示を付した、（１）〜（９）のいずれかに記載の組成物。
（１１）機能表示が、「血管透過性亢進を抑制する」、「血管内因子の血管外への漏出を抑制する」、「血管を正常な状態に改善する」、「血管を正常な状態に維持する」、「血管内皮細胞同士の乖離を抑制する」、「血管内皮細胞の細胞死を抑制する」、及び「組織間液の速やかな回収機能を保持する」からなる群から選択される１以上である、（１０）に記載の組成物。
（１２）Ｔｉｅ２活性化のための、前記（Ｉ）で表される化合物、前記（ＩＩ）で表される化合物、前記（ＩＩＩ）で表される化合物（Oleanoside A）、前記（ＩＶ）で表される化合物（Kaempferol）、及びそれらの塩からなる群から選択される１以上の使用。
（１３）前記（Ｉ）で表される化合物、前記（ＩＩ）で表される化合物、前記（ＩＩＩ）で表される化合物（Oleanoside A）、前記（ＩＶ）で表される化合物（Kaempferol）、及びそれらの塩からなる群から選択される１以上を有効成分として使用する、Ｔｉｅ２を活性化する方法。
（１４）下記式（Ｉ）

で表される化合物。
（１５）下記式（ＩＩ）

で表される化合物。
（１６）（１４）又は（１５）に記載の化合物を含有する組成物。
（１７）（１４）及び（１５）に記載の化合物を含有する組成物。
（１８）食品組成物である、（１６）又は（１７）に記載の組成物。
（１９）飲料組成物である、（１６）又は（１７）に記載の組成物。That is, the aspects of the present invention include, but are not limited to, the following inventions.
(1) The compound represented by the following (I), the compound represented by the following (II), the compound represented by the following (III) (Oleanoside A), the compound represented by the following (IV) (Kaempferol), A Tie2 activating composition containing one or more components selected from the group consisting of and salts thereof:

(2) The compound represented by (I), the compound represented by (II), the compound represented by (III) (Oleanoside A), and the compound represented by (IV) (Kaempferol). A composition for suppressing vascular permeability, which comprises one or more components selected from the group consisting of and salts thereof.
(3) The compound represented by the above (I), the compound represented by the above (II), the compound represented by the above (III) (Oleanoside A), the compound represented by the above (IV) (Kaempferol), A composition for vascular maturation, which comprises one or more components selected from the group consisting of and salts thereof.
(4) The compound represented by (I), the compound represented by (II), the compound represented by (III) (Oleanoside A), and the compound represented by (IV) (Kaempferol). A composition for normalizing blood vessels, which comprises one or more components selected from the group consisting of and salts thereof.
(5) The compound represented by the above (I), the compound represented by the above (II), the compound represented by the above (III) (Oleanoside A), the compound represented by the above (IV) (Kaempferol), A composition for stabilizing blood vessels, which comprises one or more components selected from the group consisting of and salts thereof.
(6) The compound represented by the above (I), the compound represented by the above (II), the compound represented by the above (III) (Oleanoside A), the compound represented by the above (IV) (Kaempferol), A composition for stabilizing lymphatic vessels, which comprises one or more components selected from the group consisting of and salts thereof.
(7) The vascular permeability suppressing action, the blood vessel maturation action, the blood vessel normalizing action, the blood vessel stabilizing action, or the lymphatic vessel stabilizing action are caused by the Tie2 activating action, (2). The composition according to any one of (6).
(8) The composition according to any one of (1) to (7), which is a food composition.
(9) The composition according to any one of (1) to (7), which is a beverage composition.
(10) Function exerted by activating Tie2, function exerted by suppressing vascular permeability, function exerted by maturation of blood vessels, function exerted by normalization of blood vessels, by stabilization of blood vessels The composition according to any one of (1) to (9), which is labeled with one or more functions selected from the group consisting of the functions exerted and the functions exerted by lymphatic vessel stabilization.
(11) Function indications are "suppress vascular hyperpermeability", "suppress leakage of intravascular factors to the outside of blood vessels", "improve blood vessels to normal state", and "reduce blood vessels to normal state". Selected from the group consisting of "maintaining", "suppressing divergence between vascular endothelial cells", "suppressing cell death of vascular endothelial cells", and "maintaining the function of rapidly collecting intertissue fluid" 1 The composition according to (10), which is the above.
(12) The compound represented by (I), the compound represented by (II), the compound represented by (III) (Oleanoside A), and the compound represented by (IV) for activating Tie2. One or more uses selected from the group consisting of compounds to be (Kaempferol), and salts thereof.
(13) The compound represented by the above (I), the compound represented by the above (II), the compound represented by the above (III) (Oleanoside A), the compound represented by the above (IV) (Kaempferol), And a method of activating Tie2 using one or more selected from the group consisting of salts thereof as an active ingredient.
(14) The following formula (I)

The compound represented by.
(15) The following formula (II)

The compound represented by.
(16) A composition containing the compound according to (14) or (15).
(17) A composition containing the compounds according to (14) and (15).
(18) The composition according to (16) or (17), which is a food composition.
(19) The composition according to (16) or (17), which is a beverage composition.

According to the present invention, a composition containing one or more components selected from the group consisting of novel iridoid compounds (Compound A and Compound B), oleanoside A, kenpherol, and salts thereof as an active ingredient has a Tie2 activating action. , Vascular permeability inhibitory action, blood vessel maturation action, blood vessel normalizing action, blood vessel stabilizing action, and lymphatic vessel stabilizing action can be exerted. In addition, by strengthening the connection between vascular endothelial cells by normalizing blood vessels according to the present invention, nutrition is distributed to the skin through blood vessels, and cosmetic effects such as improvement of skin firmness and texture and prevention of wrinkles are obtained. Be done. In addition, blood flow is improved by closing the gaps between endothelial cells of blood vessels and lymph vessels, and water and waste products are collected from the tissue, which is effective in improving various symptoms such as coldness, stiff shoulders, swelling, and bears. And preventive effect.

FIG. 1 shows the chemical structures of novel iridoid compounds (Compound A and Compound B), oleanoside A, and kaempferol. FIG. 2 shows the purification flow of the olive fruit extract. ^{FIG. 3 shows 1} H-NMR and ¹³ C-NMR signals of a novel iridoid compound (Compound A) and megalithactonol. FIG. 4 shows the results of two-dimensional NMR analysis of a novel iridoid compound (Compound A). Specifically, as shift correlation spectrum analysis, ¹ H- ¹ H shift correlation 2D NMR ( ¹ H- ¹ H correlation spectroscopy (COSY)) analysis and ¹ H- ¹³ C long range shift correlation 2D NMR ( ¹ H) -Detected heteronuclear multiple bond connectivity (HMBC)) Shows the results of the analysis. ^{FIG. 5 shows 1} H-NMR and ¹³ C-NMR signals of the novel iridoid compound (Compound B) and oleanoside A. FIG. 6 shows the results of two-dimensional NMR analysis of the novel iridoid compound (Compound B). Specifically, as a shift correlation spectrum analysis, the results of ¹ H- ¹³ C long range shift correlation two-dimensional NMR ( ¹ H-detected heteronuclear multiple bond connectivity (HMBC)) analysis are shown. FIG. 7 shows the results of comparing the Tie2 activating effects of novel iridoid compounds (Compound A and Compound B), oleanoside A, kaempferol, hydroxytyrosol, and tyrosol with those of the control group (DMSO). FIG. 8 shows the results of evaluating the inhibitory effect of novel iridoid compounds (Compound A and Compound B) on the increase in vascular permeability caused by vascular endothelial growth factor (VEGF), which is a vascular permeability factor. Shown.

1. 1. Composition
1-1. Composition One aspect of the present invention is a composition containing one or more components selected from the group consisting of Compound A, Compound B, oleanoside A, kaempferol, and salts thereof as an active ingredient. The active ingredient contained in the composition of the present invention is a compound represented by the following formulas (I) to (IV).

The compounds represented by the formulas (I) and (II) are novel iridoid compounds. The compound of the formula (I) is 2- (3,4-dihydroxyphenyl) ethyl (E) -2- (5-ethylidene-2-oxotetrahydropyran-4-yl) acetate (2- (3) , 4-dihydroxyphenyl) ethyl (E) -2- (5-ethylidene-2-oxotetrahydropyran-4 -yl) acetate), and may be referred to as "Compound A" in the present specification. The compound of the formula (II) is 2- (3,4-dihydro-3-hydroxymethyl-5-methoxycarbonyl-2-methyl-2H-pyran-4-yl) acetic acid (2-(3,4-dihydro-3-hydroxymethyl-5-methoxycarbonyl-2-methyl-2H-pyran-4-yl) acetic acid under the IUPAC name. 3,4-dihydro-3-hydroxymethyl-5-methoxycarbonyl-2-methyl-2H-pyran-4-yl) acetic acid), and may be referred to as "Compound B" in the present specification. The compound represented by the formula (III) is Oleanoside A, and the compound represented by the formula (IV) is kaempferol.

The compound in the present invention may be in the form of any pharmacologically acceptable salt (including inorganic and organic salts). As used herein, the term "salt" refers to any pharmacologically acceptable salt (including inorganic and organic salts), for example, sodium salt, potassium salt, calcium salt, magnesium salt, ammonium salt, hydrochloric acid. Salts, sulfates, nitrates, phosphates, organic acid salts (acetate, citrate, maleate, malate, oxalate, lactate, succinate, fumarate, propionate, formate , Butts, picrates, benzenesulfonates, trifluoroacetates, etc.), but are not limited thereto. Salts of the compounds of the invention can be readily prepared by one of ordinary skill in the art by any method known in the art.

1-2. Isolation / Synthesis of Active Ingredients in the Composition of the Present Invention The active ingredients (Compound A, Compound B, Oleanoside A, Kenferol, and / or salts thereof) contained in the composition of the present invention are derived from natural products. It may be produced by extraction / isolation or by a chemical synthesis method, and is not particularly limited. When the active ingredient contained in the composition of the present invention is produced from a natural product, the natural product used as a raw material is not particularly limited, but it is preferable to use olive, for example. Olive is a plant of the family Oleaceae, and olive fruits can be used as the main raw material, but leaves, seeds, stems and the like may be mixed. When olive fruit is used as a raw material, it may be used as it is, or it may be dried by freeze-drying or the like. In addition, the residue after squeezing the oil from the olive fruit can be used as it is or in a dried state. It is preferable to use only olive juice, which is an aqueous layer obtained by separating the liquid phase obtained from olive fruits into an oil layer and an aqueous layer, as a raw material.

The olive varieties are not particularly limited, but for example, varieties such as Manzanillo, Lucca, Nevadillo Blanco, Mission, Picual, Albequina, Ohibranka, Cornicabra, Gordar, Moroiolo, Frantio, Colatina, and Leccino can be preferably used. it can.

Here, in the present invention, the "olive fruit juice" refers to an extract obtained from an aqueous layer obtained by separating the liquid phase obtained from the olive fruit extract into an oil layer and an aqueous layer. The olive fruit extract can also be extracted from olives, for example, using an aqueous solvent. Examples of the aqueous solvent include water and alcohols (for example, ethanol). As the olive fruit extract, olive juice or an aqueous solvent extract may be used as it is, or an extract obtained by filtering these and then concentrating polyphenols on an adsorption column and spray-drying may be used. The olive fruit extract can be obtained through a step of separating and purifying olive fruits. In the olive fruit separation step, the raw material olive fruit is squeezed and separated into oil and fruit juice by centrifugation to obtain olive juice. The squeezing / centrifugation is performed by a method known to those skilled in the art. Can be done. In the purification step, the extract obtained by filtering and concentrating the olive fruit juice obtained in the separation step can be used as the olive fruit extract in the present invention. Further, the filtrate obtained by filtering olive juice is added to a column filled with an adsorbent resin to remove components that are not adsorbed to the adsorbent resin such as sugars, minerals and organic acids, and then an aqueous solvent (for example, water, ethanol) is used. , Or a mixture thereof) may be used as an eluate eluted as an olive fruit extract. Those skilled in the art can appropriately set the conditions for the type of adsorbent resin and the elution solvent of the column according to the type and form of olives used.

Extraction of the active ingredient contained in the composition of the present invention from a raw material derived from a natural product can be carried out using a solvent, but the present invention is not limited thereto. Any solvent may be used as long as the active ingredient contained in the composition of the present invention can be extracted, and water, an aqueous solution such as a buffer solution, an organic solvent, or a combination thereof can be used. The organic solvent may be any generally used one, and for example, alcohols and ketones can be used. Examples of alcohols include lower alcohols (methanol, ethanol, propanol, isopropanol, butanol, isobutanol, etc.) and liquid polyhydric alcohols (1,3-butylene alcohol, propylene glycol, glycerin, etc.). Preferably, hydrous lower alcohols or lower alcohols are used. Examples of ketones include acetone, butanone and the like. The amount of the solvent used can be appropriately set according to the amount of the raw material to be treated, the content of the target compound in the raw material, and the like. For example, 1.0 to 1000 mL, preferably 2. You may use 0 to 500 mL, more preferably 5.0 to 100 mL of solvent.

The active ingredient contained in the composition of the present invention can be further purified to increase the purity after extraction from the raw material, if necessary. Purification may be carried out by any method, for example, by chromatography. Examples of chromatography include, but are not limited to, ion exchange chromatography, hydrophobic chromatography, hydrophilic chromatography, gel filtration chromatography and the like. The purity of the sample can be confirmed by high performance liquid chromatography or LC-MS which is a combination of high performance liquid chromatography and a mass spectrometer, but is not limited thereto. Further, the chemical structure of the active ingredient contained in the composition of the present invention can be analyzed by nuclear magnetic resonance (NMR), mass spectrum, or the like. The composition of the present invention does not include naturally occurring animals and plants that have not been processed at all.

When the active ingredient contained in the composition of the present invention is chemically synthesized, the compound of the present invention is produced by appropriately selecting a generally known organic chemistry method using an available compound as a starting material. can do.

1-3. Composition for activating Tie2, etc. One aspect of the present invention is activating Tie2 containing one or more components selected from the group consisting of Compound A, Compound B, oleanoside A, kaempferol, and salts thereof as an active ingredient. Composition for use.

In the present invention, the "Tie2 activation composition" refers to a composition having the ability to convert Tie2 into an active substance (phosphorylated Tie2) by phosphorylating Tie2. Activation of a receptor called Tie2 present in vascular endothelial cells plays an important role in stabilizing the structure of capillaries. This Tie2 is shown to be present not only in vascular endothelial cells but also in lymphatic endothelial cells, and is activated by angiopoietin-1 released from parietal cells, strengthening the adhesion between endothelial cells and the parietal cells, and the vascular structure. Involved in stabilization. It has also been reported that the activity of Tie2 decreases with age, for example, and it has been clarified that blood vessels and lymph vessels also age accordingly. Therefore, the Tie2 activating composition in the present invention can prevent and improve the aging of blood vessels and lymph vessels by the Tie2 activating action.

In addition, when the vascular structure becomes unstable due to aging of blood vessels, nutritional components such as plasma components leak from the blood vessels, and the necessary components cannot be carried to the necessary places. As a result, various troubles such as deterioration of skin condition may occur. Therefore, the Tie2 activating composition in the present invention can stabilize vascular endothelial cells by the Tie2 activating action and exert a vascular permeation inhibitory action. By firmly binding the endothelial cells of blood vessels, nourishment is distributed to the skin, and cosmetic effects such as improvement of skin firmness and texture and prevention of wrinkles can be obtained. In addition, blood flow is improved by closing the gaps between endothelial cells in blood vessels and lymph vessels, and water and waste products are collected from the tissues to improve various symptoms such as coldness, stiff shoulders, swelling, and bears. , The effect of prevention is obtained. Therefore, the present invention is a composition for activating Tie2 containing one or more components selected from the group consisting of Compound A, Compound B, oleanoside A, kaempferol, and salts thereof as an active ingredient, and is a blood vessel. The composition may have a permeability inhibitory action, a blood vessel maturation action, a blood vessel normalizing action, a blood vessel stabilizing action, or a lymphatic vessel stabilizing action.

As another aspect of the present invention, a composition for suppressing vascular permeability, a composition for maturating blood vessels, a composition for normalizing blood vessels, a composition for stabilizing blood vessels, a composition for stabilizing lymphatic vessels, and the like can be mentioned. Be done. These compositions preferably contain one or more ingredients selected from the group consisting of Compound A, Compound B, oleanoside A, kaempferol, and salts thereof as an active ingredient.

In the present invention, "inhibition of vascular permeability" means suppressing the increase in vascular permeability caused by VEGF (Vascular Endothelial Growth Factor) or the like. "Maturation of blood vessels" is to induce adhesion between vascular endothelial cells and vascular parietal cells to form focal adhesions between vascular endothelial cells so that intravascular environmental factors do not easily leak out of the blood vessels. To say. "Normalization of blood vessels" means to enhance the adhesion between vascular endothelial cells and promote the lining of blood vessel wall cells to vascular endothelial cells, thereby leading to the disordered growth of blood vessels and blood vessels with vascular permeability disruption. It refers to the normalization of abnormal blood vessels. "Stabilization of blood vessels" means damage to existing blood vessels, divergence between vascular endothelial cells, action of suppressing divergence between vascular endothelial cells and vascular parietal cells, and suppression of cell death of vascular endothelial cells. .. "Lymphatic vessel stabilization" refers to stabilizing lymphatic vessels so that lymphatic endothelial cells are properly arranged and lined, and retains the function of promptly collecting interstitial fluid.

1-4. Content of Active Ingredient in Composition The content of Compound A, Compound B, oleanoside A, kaempferol, and salts thereof in the composition of the present invention is the content of the present invention in consideration of its administration form, administration method, and the like. The amount may be any amount as long as the desired effect can be obtained, and is not particularly limited. For example, when Compound A is used as the active ingredient, the content of Compound A in the composition of the present invention is 0.01% by weight to 99% by weight, preferably 0.1% by weight or more, based on the total amount of the composition. It is 50% by weight, more preferably 1% by weight to 30% by weight, and even more preferably 10% by weight to 30% by weight. When Compound B is used as the active ingredient, the content of Compound B in the composition of the present invention is 0.01% by weight to 99% by weight, preferably 0.1% by weight to 50% by weight, based on the total amount of the composition. %, More preferably 1% by weight to 30% by weight, even more preferably 10% by weight to 30% by weight. When oleanoside A is used as the active ingredient, the content of oleanoside A in the composition of the present invention is 0.01% by weight to 99% by weight, preferably 0.1% by weight to 50% by weight, based on the total amount of the composition. %, More preferably 1% by weight to 30% by weight, even more preferably 10% by weight to 30% by weight. When kenferol is used as the active ingredient, the content of kenferol in the composition of the present invention is 0.001% by weight to 99% by weight, preferably 0.01% by weight to 50% by weight, based on the total amount of the composition. %, More preferably 0.1% by weight to 30% by weight, even more preferably 1% by weight to 30% by weight. As used herein, "% by weight" means% by weight of weight / weight (w / w) unless otherwise noted. When various active ingredients are in the form of salts, hydrates, etc., the above contents shall be calculated after converting them into free substances.

The contents of Compound A, Compound B, oleanoside A, kaempferol, and salts thereof can be measured according to methods known to those skilled in the art. For example, LC-MS / MS, HPLC, or the like can be used for measurement by setting appropriate conditions according to the type of active ingredient and the like.

1-5. Other Ingredients The composition of the present invention may contain any additive or ingredient in addition to the active ingredient, depending on its form. These additives and / or ingredients are not particularly limited, but are, for example, additives such as excipients, binders, disintegrants, abundant agents, antioxidants, preservatives, flavoring agents, odorants, coloring agents, and fragrances. Can be used. For example, L-ascorbic acid, catechin, dextrin, cyclodextrin, calcium carbonate, trehalose and the like can be used. Further, the composition may contain a known component, or may contain a component known to be effective for preventing aging of blood vessels and cosmetology. For example, Kiraya, Huangli, Ginkgo, Oyster, Ukon, Chrysanthemum, Natsume, Kuco, Chamomile, Butcher Bloom, Sanzashi, Star Fruit, Getto, Hass, Louis Boss, Indian Dates, Karin, Sijuum, which have been reported to activate Tie2. Guava, Hihatsu, Siberian carrot, Mango ginger, Koryo carrot, Akigumi, Okahijiki, Harigiri, Ryobu, Yabukanzo, Hasuimo, Mitsubautsugi, Kusagi, Mube, Sanshosou, Konara, Kunugi, Akinonogeshi, Suishogaki Extracts of Nikkei, Ginger, Ousei, Gyokuchiku, Caronin, Vulture, grape seeds and pine bark containing procyanidin, which are effective in improving vascular endothelium, extracts of peanut seed coat, lychee seed coat, L-citrulin, L-arginine, etc. Amino acids can be used.

1-6. Uses The compositions of the present invention can exhibit various physiological actions as described above. For example, by applying the composition of the present invention, vascular permeability suppressing effect, blood vessel maturation effect, blood vessel normalizing effect, blood vessel stabilizing effect, lymphatic vessel stabilizing effect, Tie2 activating effect and the like are exhibited. As a result, it is possible to obtain beauty effects such as improvement of skin firmness and texture, prevention of wrinkles, and improvement / prevention effects such as coldness, stiff shoulders, swelling, and bears.

The composition of the present invention can be applied to either therapeutic use (medical use) or non-therapeutic use (non-medical use). Examples of such compositions include pharmaceuticals (pharmaceutical compositions), foods and drinks (including foods, beverages, food and drink compositions, food compositions, beverage compositions), cosmetics (cosmetic compositions), and the like. Not limited to.

The composition of the present invention is a solid agent such as a tablet (including a coated tablet), a granule, a powder, a powder, or a capsule, or an oral solution, a suspension, an emulsion, a syrup, or a syrup, according to a known method. It can be provided as an agent formulated in a liquid agent such as a drink agent. These agents can be taken as they are or with water or the like. Further, after preparation in a form that can be easily blended (for example, powder form or granule form), it can be used as a raw material for, for example, pharmaceuticals and foods and drinks, but is not limited to this form. The dosage form of the composition of the present invention can be arbitrarily selected depending on the intended purpose as a cosmetic (cosmetic composition), for example, agents such as creams, ointments, emulsions, solutions, gels, powders and granules. It can be in the form of shapes, packs, lotions, powders, sticks and the like. The formulation forms include a wide range of forms such as aqueous solution type, solubilization type, emulsification type, powder type, oil liquid type, gel type, ointment type, aerosol type, water-oil two-layer system and water-oil-powder three-layer system. can do. That is, for basic cosmetics, facial cleansers, lotions, lotions, milky lotions, creams, gels, essences (essences), facial masks, massage agents, foundations, lipsticks, bathing agents, shampoos, hair conditioners, hair tonics. , Ointments, pastes and sheet products (absorbent articles such as sanitary products, wipes and wet tissues, etc.) can be widely used. The dosage form and form of the present invention are not limited to this. In addition, various oil agents, surfactants, gelling agents, preservatives, antioxidants, solvents, alcohols, water, chelating agents, thickeners, ultraviolet absorbers, emulsion stabilizers, pH Adjusters, pigments, fragrances and the like can be appropriately blended.

Specific examples of the beverage of the present invention include Karyu tea beverage, black tea beverage, green tea beverage, fruit juice beverage, vegetable juice, sports drink, isotonic beverage, enhanced water, mineral water, near water, nutritional drink, beauty drink, and various types. Alcoholic beverages and the like can be mentioned.

In addition, the beverage of the present invention is made into a packaged beverage after undergoing steps such as sterilization as necessary. For example, a sterilized packaged beverage can be produced by a method of filling a container with a beverage and then performing heat sterilization or the like, or a method of sterilizing the beverage and then filling the container in a sterile environment.

The type of container used for packaged beverages is not particularly limited. For example, resin containers such as PET bottles, paper containers such as paper packs, glass containers such as glass bottles, metal containers such as aluminum cans and steel cans, and aluminum pouches. Any container that is normally used for beverages can be used.

The food or drink of the present invention is not particularly limited, and is, for example, a health food, a functional food (including a food for specified health use, a food for specified health use with conditions, a food with nutritional function, a food with functional claim), a food for special use, or a food for special use. Examples include health supplements. The content of the active ingredient in the food and drink of the present invention is as described above for the composition.

For example, when preparing a known food or drink, the food or drink of the present invention may be prepared by mixing a predetermined amount of the active ingredient with the raw material thereof according to a known method for producing the food or drink, or a ready-made food or drink. It may be prepared by adding the active ingredient to a known food or drink in the predetermined amount. The known food and drink may originally contain the active ingredient, and if the active ingredient is in the predetermined amount, it can be appropriately blended and prepared.

The composition of the present invention can be applied by an appropriate method according to its form. The application method is not particularly limited as long as the active ingredient according to the present invention can be transferred into the circulating blood. For example, it may be in the form of an oral solid preparation, an oral liquid preparation such as an oral liquid or a syrup, or a parenteral preparation such as an injection, an external preparation, a suppository or a transdermal absorbent. Not limited to. In addition, in this specification, "application" is used as including all aspects such as ingestion, ingestion, drinking, and percutaneous absorption. The applied amount of the composition of the present invention is timely set according to its form, administration method, purpose of use and age, body weight, and symptom of the patient or animal to be administered, and is not constant. The effective human intake of the composition of the present invention is also not constant and is not particularly limited. The effective human intake of the composition of the present invention is, for example, 0.1 mg to 400 mg (0.002 mg / kg to 8 mg / kg), preferably 1 mg to 300 mg (0.02 mg / kg) per day for a person weighing 50 kg. ~ 6 mg / kg), more preferably 10 mg to 200 mg (0.2 mg / kg to 4 mg / kg). The composition of the present invention may be applied once or divided into several times within a day within a desired intake range. The application period is also arbitrary. The effective human intake of the composition of the present invention is the intake of the composition of the present invention showing an effective effect in humans, and the type of the active ingredient contained in the composition is not particularly limited.

The application target of the composition of the present invention is preferably human, but domestic animals such as cows, horses and goats, pet animals such as dogs, cats and rabbits, or experimental animals such as mice, rats, guinea pigs and monkeys. It may be. When applied to animals other than humans, the daily application amount per mouse (about 20 g) is a condition such as the content of the active ingredient in the composition, the condition of the target person, body weight, sex and age. It depends on, for example, 0.002 mg / kg to 8 mg / kg, preferably 0.02 mg / kg to 6 mg / kg, and more preferably 0.2 mg / kg to 4 mg / kg. In another aspect, the present invention provides an indication of a function exerted by vascular permeability suppression, blood vessel maturation, blood vessel normalization, blood vessel stabilization, lymphatic vessel stabilization, or Tie2 activation. The present invention relates to a composition containing the agent of the present invention. Such display or function display is not particularly limited, but for example, "suppresses vascular hyperpermeability", "suppresses leakage of intravascular factors to the outside of blood vessels", and "improves blood vessels to a normal state". , "Maintaining blood vessels in a normal state", "Suppressing divergence between vascular endothelial cells", "Suppressing cell death of vascular endothelial cells", or "Maintaining the function of prompt recovery of interstitial fluid" Etc., or a display or a function display that can be equated with these. In the present specification, indications such as the indication and the function indication may be attached to the composition itself, or may be attached to the container or packaging of the composition.

2. Use for activating Tie2 One aspect of the present invention is one or more selected from the group consisting of Compound A, Compound B, oleanoside A, kaempferol, and salts thereof, which suppress vascular permeability, vascular. It is used for maturation, vascular normalization, vascular stabilization, lymphatic vessel stabilization, or Tie2 activation, preferably for Tie2 activation.

Further, by using the present invention, Tie2 is activated to obtain a vascular permeability suppressing effect, a blood vessel maturation effect, a blood vessel normalizing effect, a blood vessel stabilizing effect, a lymphatic vessel stabilizing effect, and the like. You can also. The use is in humans or non-human animals and may be therapeutic or non-therapeutic. Here, "non-therapeutic" is a concept that does not include medical treatment, that is, treatment of the human body by treatment.

3. 3. Method for activating Tie2 One aspect of the present invention is a method for activating Tie2, which uses one or more selected from the group consisting of Compound A, Compound B, oleanoside A, kaempferol, and salts thereof as an active ingredient. is there. Specifically, one aspect of the present invention provides a method of activating Tie2, which comprises administering a therapeutically effective amount of the active ingredient to a subject requiring Tie2 activation. The target that requires Tie2 activation is as described above for the composition.

Further, one aspect of the present invention is to suppress vascular permeability and mature blood vessels by using one or more selected from the group consisting of Compound A, Compound B, oleanoside A, kaempferol, and salts thereof as an active ingredient. It is also a method for normalizing blood vessels, stabilizing blood vessels, or stabilizing lymphatic vessels.

Further, in the present specification, the therapeutically effective amount is taken when one or more selected from the group consisting of Compound A, Compound B, oleanoside A, kaempferol, and salts thereof is ingested by the above-mentioned subject. It is the amount at which Tie2 activation is obtained as compared to a non-subject. The specific effective amount is set in a timely manner depending on the administration form, administration method, purpose of use, age, body weight, symptomatology, etc. of the subject, and is not constant.

In the method of the present invention, the active ingredient may be administered as it is or as a composition containing the active ingredient so as to obtain the therapeutically effective amount.

4. Novel Iridoid Compound One aspect of the present invention is a novel iridoid compound represented by the following formula (I) or (II).

The compound of the formula (I) is 2- (3,4-dihydroxyphenyl) ethyl (E) -2- (5-ethylidene-2-oxotetrahydropyran-4-yl) acetate (2-yl) under the IUPAC name. It is expressed as (3,4-dihydroxyphenyl) ethyl (E) -2- (5-ethylidene-2-oxotetrahydropyran-4 -yl) acetate), and may be described as "Compound A" in this specification. The compound of the formula (II) is 2- (3,4-dihydro-3-hydroxymethyl-5-methoxycarbonyl-2-methyl-2H-pyran-4-yl) acetic acid (2-(3,4-dihydro-3-hydroxymethyl-5-methoxycarbonyl-2-methyl-2H-pyran-4-yl) acetic acid under the IUPAC name. 3,4-dihydro-3-hydroxymethyl-5-methoxycarbonyl-2-methyl-2H-pyran-4-yl) acetic acid), and may be referred to as "Compound B" in the present specification.

The compounds in the present invention may be in the form of any pharmacologically acceptable salt (including inorganic and organic salts). As described above, the term "salt" as used herein is not particularly limited as long as it is a pharmacologically acceptable salt (including an inorganic salt and an organic salt), and is, for example, a sodium salt, a potassium salt, or calcium. Salts, magnesium salts, ammonium salts, hydrochlorides, sulfates, nitrates, phosphates, organic acid salts (acetate, citrate, maleate, malate, oxalate, lactate, succinate, fumal Examples thereof include acid salts, propionates, acidates, benzoates, picphosphates, benzenesulfonates, trifluoroacetates, etc.). Salts of the compounds of the invention can be readily prepared by one of ordinary skill in the art by any method known in the art.

4-1. Method for Producing New Iridoid Compound The novel iridoid compound (Compound A, Compound B) of the present invention may be produced by extracting / isolating it from a natural product or by a chemical synthesis method, and is not particularly limited. When produced from a natural product, the natural product used as a raw material is not particularly limited, but it is particularly preferable to use olive. Olive is a plant of the family Oleaceae, and olive fruits can be used as the main raw material, but leaves, seeds, stems and the like may be mixed. When olive fruit is used as a raw material, it may be used as it is, or it may be dried by freeze-drying or the like. In addition, the residue after squeezing the oil from the olive fruit can be used as it is or in a dried state. It is preferable to use only olive juice, which is an aqueous layer obtained by separating the liquid phase obtained from olive fruits into an oil layer and an aqueous layer, as a raw material. "Olive" and "olive juice" are as described above with respect to the composition.

Extraction of the novel iridoid compound of the present invention from a raw material derived from a natural product (for example, olive fruit extract) can be carried out using a solvent, but is not limited thereto. Any solvent may be used as long as the active ingredient contained in the composition of the present invention can be extracted, and water, an aqueous solution such as a buffer solution, an organic solvent, or a combination thereof can be used. The organic solvent may be any generally used one, and for example, alcohols or acetone can be used. Examples of alcohols include lower alcohols (methanol, ethanol, propanol, isopropanol, butanol, isobutanol, etc.) and liquid polyhydric alcohols (1,3-butylene alcohol, propylene glycol, glycerin, etc.). Preferably, hydrous lower alcohols or lower alcohols are used. The amount of the solvent used can be appropriately set according to the amount of the raw material to be treated, the content of the target compound in the raw material, and the like. For example, 1.0 to 1000 mL, preferably 2. You may use 0 to 500 mL, more preferably 5.0 to 100 mL of solvent.

The novel iridoid compound of the present invention can be further purified to increase its purity after extraction from the raw material, if necessary. Purification may be carried out by any method, for example, by chromatography. Examples of chromatography include, but are not limited to, ion exchange chromatography, hydrophobic chromatography, hydrophilic chromatography, gel filtration chromatography and the like. The purity of the sample can be confirmed by high performance liquid chromatography or LC-MS which is a combination of high performance liquid chromatography and a mass spectrometer, but is not limited thereto. Further, the chemical structure of the novel iridoid compound of the present invention can be analyzed by nuclear magnetic resonance (NMR), mass spectrum, or the like.

When the novel iridoid compound of the present invention is chemically synthesized, a desired compound can be produced by appropriately selecting a generally known organic chemistry method using an available compound as a starting material.

4-2. Composition One aspect of the present invention is a composition containing Compound A and / or Compound B. As described above, the novel iridoid compounds of the present invention, Compound A and Compound B, have various physiological actions, for example, Tie2 activating action, vascular permeability suppressing action, blood vessel maturation action, and blood vessel normalizing action. , Blood vessel stabilizing action, lymphatic vessel stabilizing action, etc. are shown. Therefore, in some embodiments, the composition of the present invention has a Tie2 activating effect, a vascular permeability inhibitory effect, a blood vessel maturation effect, a blood vessel normalizing effect, a blood vessel stabilizing effect, or a lymphatic vessel stabilizing effect. Have. Examples of the composition include pharmaceuticals (pharmaceutical compositions), foods and drinks (including foods, beverages, food and drink compositions, food compositions, beverage compositions), cosmetics (cosmetic compositions), and the like. , Not limited to these.

As described above, Compound A and / or Compound B contained in the composition of the present invention may be produced by extracting and isolating from a natural product, or may be produced by a chemical synthesis method, and is not particularly limited. When the active ingredient contained in the composition of the present invention is produced from a natural product, the natural product used as a raw material is not particularly limited, but it is preferable to use olive, for example. The method for isolating and synthesizing Compound A and / or Compound B is also as described above. The composition of the present invention does not include naturally occurring animals and plants that have not been processed at all, or extracts themselves that do not contain other components.

The content of Compound A and / or Compound B in the composition of the present invention is not particularly limited. For example, the content of Compound A in the composition of the present invention is 0.01% by weight based on the total amount of the composition. ~ 99% by weight, preferably 0.1% by weight to 50% by weight, more preferably 1% by weight to 30% by weight, even more preferably 10% by weight to 30% by weight, and Compound B in the composition of the present invention. The content of is 0.01% by weight to 99% by weight, preferably 0.1% by weight to 50% by weight, more preferably 1% by weight to 30% by weight, still more preferably, based on the total amount of the composition. It is 10% by weight to 30% by weight. Other components and the like that can be blended in the composition are as described above.

Hereinafter, the present invention will be described in more detail based on examples. The present invention is not limited to these examples.

Example 1: Structure determination of Compound A and Compound B
1-1. Preparation of Olive Fruit Extract Olive fruits are squeezed, centrifuged and separated into oil and juice. After filtering the olive juice obtained here, components that are not adsorbed to the adsorbed resin such as sugars, minerals, and organic acids are removed by an adsorption column, the hydroalcohol-eluting part is concentrated, and the olive fruit is spray-dried. Used as an extract. Subsequently, this fruit extract (500 g) was subjected to column chromatography (resin amount; 10 L, elution solvent; ethanol: water (0: 100 → 70: 30)) using Diaion HP20 to fractionate the olive fruit extract. , Each fraction was obtained as follows (Fr.1: 179 g, Fr.2: 120 g, Fr.3: 85 g, Fr.4: 71 g, Fr.5: 26 g). Of the fractions in which Tie2 activity was observed in Fr.1 to Fr.5, Fr.4 (50% ethanol elution part, 20 g) was subjected to column chromatography using Toyopearl HW-40C (resin amount; 500 mL, elution). Solvent; methanol: water (10:90 → 70:30) → acetone: water (70:30)) was carried out, and Fr.4 was fractionated into 19 fractions (Fr.4a to Fr.4s). Fraction Fr.4k in which Tie2 activity was observed in Fr.4a to Fr.4s was subjected to preparative HPLC (Preparative HPLC) under the following analysis condition 1, and further divided into 6 fractions (Fr.4ka to 4kf). Among the fractions in which the highest Tie2 activity was observed in Fr.4ka to Fr.4kf, Fr.4kb and Fr.4kd were purified using preparative HPLC under the following analytical conditions 2 and 3. , Compound A (46.2 mg), Compound B (19.5 mg) and Oleanoside A (98.1 mg) were isolated. For Fr.4r in which Tie2 activity was observed, Kaempferol (3.7 mg) was isolated by performing preparative HPLC (Preparative HPLC) under analytical condition 4. In addition, FIG. 2 shows the purification flow of the olive fruit extract.

<Analysis condition 1>
Detection: 220 nm
Flow velocity: 15 mL / min
Column: Capcell Pack C18 TYPE AQ φ3.0 × 25.0 cm (SHISEIDO)
Mobile phase: CH ₃ CN: H ₂ O (32:68)
Column temperature: Room temperature 25 ° C
<Analysis condition 2>
Detection: 220 nm
Flow velocity: 10 mL / min
Column: TSK-Gel Amide-80 φ2.15 × 30.0 cm (TOSOH)
Mobile phase: CH ₃ CN: H ₂ O (0 min, 100: 0 → 15 min, 100: 0 → 20 min, 80:20 → 35 min, 80:20)
Column temperature: Room temperature 25 ° C
<Analysis condition 3>
Detection: 220 nm
Flow velocity: 10 mL / min
Column: COSMOSIL Cholester φ2.0 × 25.0 cm (nacalai tesque)
Mobile phase: CH ₃ CN: H ₂ O (20:80)
Column temperature: Room temperature 25 ° C
<Analysis condition 4>
Detection: 280 nm
Flow velocity: 15 mL / min
Column: Capcell Pack C18 TYPE AQ φ3.0 × 25.0 cm (SHISEIDO)
Mobile phase: CH ₃ CN: H ₂ O (27:73)
Column temperature: Room temperature 25 ° C

1-2. Structural analysis of Compound A
Compound A was obtained as a colorless oily substance, and the results of high-resolution electrospray-ionization mass spectrometry (HRESI-MS) revealed that it had a _{molecular formula of C 17} H ₂₀ O _6. Moreover, from the UV spectrum pattern (λmax 282), it was inferred that this compound has a benzene ring in the molecule. Next, in ¹ H-nuclear magnetic resonance ( ¹ H-NMR), a set of ABX signals [δ _H : 6.62 (1H, dd, J = 2.0, 8.0 Hz), 6.74 (1H, d, J = 2.0 Hz) ), 6.82 (1H, d, J = 8.0 Hz)] and the eight peak patterns observed in ¹³ _{C-NMR (δ c} : 34.6, 66.0, 115.5, 116.4, 121.3, 130.7, 143.5, 143.6) suggested the presence of hydroxytyrosol. Furthermore, in ¹ H-NMR, one methyl-derived signal, three methylene-derived signals, one methine-derived signal, and one olefin proton were confirmed. ^When units other than hydroxytyrosol were confirmed by 13 C-NMR, one methyl-derived signal, three methylene-derived signals, two methine-derived signals, and three quaternary carbon-derived signals were confirmed. Subsequently, for the purpose of clarifying these bonding modes, ¹ H- ¹ H correlation spectroscopy ( ¹ H- ¹ H COSY), ¹ H-detected multiple quantum coherence (HMQC), and ¹ H-detected multiple-bond connectivity (1 H-detected multiple-bond connectivity (HMQC)). HMBC) was measured and analyzed in detail, and it was clarified that the structure of this unit is a compound characterized by iridoid. In the iridoid unit whose structure was clarified this time by detailed analysis of spectral data centered on two-dimensional NMR with reference to an example of isolation of olive-derived iridoids, the hydroxyl group at the 3-position of megalithactol is a carboxyl group. It was clarified that it was a converted structure.

Subsequently, when HMBC was analyzed to examine the binding mode of each unit, a correlation was confirmed between the proton at the 8'position of the hydroxytyrosol unit and the carbonyl carbon at the 3-position of megalitholactol. It was revealed that the 8'position of hydroxytyrosol and the 3-position of megalitholactol are ester-bonded. From the above spectral data, the structure of Compound A was determined. ¹ H-NMR and ¹³ C-NMR signals of Compound A and megalitholactonol are shown in FIG. The results of two-dimensional NMR analysis of Compound A ( ¹ H- ¹ H correlation spectroscopy (COSY) analysis and ¹ H-detected heteronuclear multiple bond connectivity (HMBC) analysis) are shown in FIG.

1-3. Structural analysis of Compound B
Compound B was obtained as a colorless oily substance. ^{In 1} H-NMR, one methyl group-derived signal, two methylene group-derived signals, three methine group-derived signals, one methoxy group-derived signal, and one olefin-derived signal were confirmed. ^{13 In} C-NMR, there are two methyl groups (δ _c : 19.4, 51.5), two methylene groups (δ _c : 38.2, 62.3), and four methine groups (δ _c : 29.6, 45.0, 74.1, 156.2). ), Three quaternary carbons (δ _c : 108.5, 169.3, 174.3), a total of 11 carbon signals were confirmed. From the above ¹ H-NMR and ¹³ C-NMR results and UV absorption could not be confirmed, this compound was presumed to be an iridoid. ^{Next, two-dimensional NMR (1} H- ¹ H COSY, HMQC, HMBC) was analyzed for the purpose of confirming these binding modes, and the structure of this compound was clarified. The structure clarified by the above analysis is similar to that of Oleanosides reported as one of the components of olive fruits, and when their spectral data are compared, the signals at positions 6, 7, and 8 are compared. It was confirmed that there was a good correlation except for. In addition to the above results, the structure of Compound B was determined based on the fact that the HMBC correlation from methylene at position 8 to carbonyl carbon at position 7 could not be confirmed with this compound and the results of the molecular weight obtained by MS. .. ¹ H-NMR and ¹³ C-NMR signals of Compound B and oleanoside A are shown in FIG. In addition, the results of two-dimensional NMR analysis of Compound B ( ¹ H- ¹ H correlation spectroscopy (COSY) analysis and ¹ H detected heteronuclear multiple bond connectivity (HMBC) analysis) are shown in FIG. Oleanoside A and Kaempferol were identified by comparison with the literature values described in the paper.

Example 2: Evaluation of Tie2 activating action
Compound A and Compound B were dissolved in a serum vascular endothelial cell growth medium (Humedia-EG2, manufactured by Kurabo Industries Ltd.) so that the final concentration was 10 μg / mL, and a test sample was prepared. In addition, a test sample in which angiopoietin-1 was dissolved in Humedia-EG2 to a concentration of 500 ng / mL was prepared and used as a positive control.

Next, normal human umbilical vein endothelial cells (HUVEC) cultured to confluence ^{were seeded on a 96-well plate at 2.0 × 10 4} cells / 0.1 mL / well, and cultured overnight using Human-EG2. After overnight culturing, HUVEC was replaced with 0.1 mL of vascular endothelial cell basal medium (Humedia-EB2, manufactured by Kurabo Industries Ltd.) 3 hours before cell stimulation (addition of test sample), and culturing was performed again. Then, 0.1 mL of the test sample was added into the well, and the incubation was carried out for 20 minutes. After incubation, intracellular phosphorylated Tie2 levels were measured using an immunoassay kit (R & D Systems, Human Phospho-Tie2 (Y992) Immunoassay) according to the protocol. As a negative control, phosphorylated Tie2 was also measured for dimethyl sulfoxide (DMSO) used for dissolving the test sample. Then, the Tie2 activation rate was calculated according to the following formula, and the phosphorylation effect was evaluated.

Tie2 activation rate (%) = measured value of phosphorylated Tie2 when the test sample was added / measured value of phosphorylated Tie2 with negative control) × 100

The results are shown in Table 1 and FIG. As shown in Table 1 and FIG. 7, Tie2 activating action was confirmed in Compound A, Compound B, oleanoside A and kaempferol.

Example 3: Inhibitory action of Compound A and Compound B on vascular permeability Using the DMSO solution (25 μM) of Compound A or B prepared in Example 1 as a test sample, vascular endothelial growth factor (vascular), which is a vascular permeability factor, was used. The inhibitory effect on increased vascular permeability caused by endothelial growth factor (VEGF) was evaluated using the Miles assay method. This animal experiment was conducted based on a plan approved by the head of the institution after being examined by the in-house animal experiment committee in compliance with the Act on Welfare and Management of Animals and other related laws and regulations.

3-1. Test method
After pre-rearing 8-week-old Balb / c female mice for 1 week or longer, 8 dosing plots were set for each mouse (4 on each side of the back median) and evaluated at n = 6. did.

15 minutes after intravenous administration of 100 μL of 1% Evans blue dye prepared in physiological saline under anesthesia, equal amounts of VEGF (final concentration 300 ng / mL) and each test sample were mixed, and 15 μL of that 15 μL was used for the back skin. Was administered within. Forty minutes after intradermal administration, the dye leaked into the skin was extracted with 500 μL formamide, and the amount of dye was quantified by absorbance at 620 nm. In addition, a section in which neither VEGF nor the test sample was administered and a section in which only VEGF was administered were provided at the same time. Dunnett's multiple comparison test was used for the significance test.

3-2. As a result, the amount of the leaked Evans blue pigment is shown in FIG. The vascular permeability suppression rate of the test sample was 89% for Compound A and 139% for Compound B. From the above, it was confirmed that both Compound A and Compound B have the effect of suppressing the permeability from blood vessels.

The present invention provides a composition having a Tie2 activating effect, a vascular permeability suppressing effect, a blood vessel maturation effect, a blood vessel normalizing effect, a blood vessel stabilizing effect, a lymphatic vessel stabilizing effect, and the like. Is. As described above, the present invention provides a new composition or the like having preferable physiological activity, and can be used for foods and drinks and the like.

Claims (10)

Tie2 activation composition containing the compound (Oleanoside A) represented by the following (III) and / or a salt thereof as an active ingredient:

A composition for suppressing vascular permeability containing the compound (Oleanoside A) represented by (III) and / or a salt thereof as an active ingredient, wherein the vascular permeability suppressing action is caused by the Tie2 activating action. The composition. A composition for vascular maturation containing the compound (Oleanoside A) represented by (III) and / or a salt thereof as an active ingredient, wherein the vascular maturation action is caused by the Tie2 activating action. The composition. A composition for normalizing blood vessels containing the compound (Oleanoside A) represented by (III) and / or a salt thereof as an active ingredient, wherein the normalizing action of blood vessels is caused by the activating action of Tie2. The composition. A composition for stabilizing blood vessels containing the compound (Oleanoside A) represented by (III) and / or a salt thereof as an active ingredient, wherein the stabilizing action of blood vessels is caused by the activating action of Tie2. The composition. A composition for stabilizing lymphatic vessels containing the compound (Oleanoside A) represented by (III) and / or a salt thereof as an active ingredient, and the lymphatic vessel stabilizing action is caused by the Tie2 activating action. The composition which is to be used. The composition according to any one of claims 1 to 6, which is a food composition. The composition according to any one of claims 1 to 6, which is a beverage composition. Functions exerted by activating Tie2, functions exerted by suppressing vascular permeability, functions exerted by maturation of blood vessels, functions exerted by normalization of blood vessels, and functions exerted by stabilization of blood vessels. The composition according to any one of claims 1 to 8, which is labeled with one or more functions selected from the group consisting of functions and functions exerted by lymphatic vessel stabilization. Function indications are "suppress vascular hyperpermeability", "suppress leakage of intravascular factors out of blood vessels", "improve blood vessels to normal state", "maintain blood vessels in normal state" , "Suppresses dissociation between vascular endothelial cells", "Suppresses cell death of vascular endothelial cells", and "Maintains rapid recovery function of intertissue fluid". , The composition according to claim 9.

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