Flavonoid-containing composition

Abstract

The inventors have found that quercetin and genistein, individually, suppress the expression of some aging-related genes while promoting the expression of other aging-related genes. Therefore, the main objective was to maintain the repressive effect on some aging-related genes that quercetin, genistein, and their glycosides may have, while suppressing the effect of promoting the expression of other aging-related genes. [Solution] The above problem can be solved by combining two or three components selected from the group consisting of component (i): quercetin and / or its glycosides, component (ii): genistein and / or its glycosides, and component (iii): nobiletin and / or its glycosides.

Description

[Technical Field]

[0001] This disclosure relates to compositions, etc., containing two or three components selected from the group consisting of component (i): quercetin and / or its glycosides, component (ii): genistein and / or its glycosides, and component (iii): nobiletin and / or its glycosides. [Background technology]

[0002] As we age, the structure and function of blood vessels deteriorate, such as arteriosclerosis and decreased endothelial function, which is known to increase the risk of various diseases. For example, stroke, vascular dementia, myocardial infarction, hypertension, diabetes, and renal failure have been reported to be closely related to vascular aging (Non-Patent Literature 1). Since these diseases are factors that cause a decline in quality of life and an increase in mortality, preventing and / or improving vascular aging is important from the perspective of extending healthy life expectancy.

[0003] In recent years, research has focused particularly on vascular endothelial cells within blood vessels. For example, Non-Patent Literature 2 reports that in transgenic mice (E-DNIκB mice) in which functional NF-κB signaling in vascular endothelial cells was inhibited, inflammation and oxidative stress in vascular endothelial cells were reduced, the progression of vascular aging was suppressed, and lifespan was extended. Furthermore, Non-Patent Literature 3 shows that in control mice fed a high-calorie diet, p53 expression in vascular endothelial cells was significantly increased, leading to metabolic abnormalities, while in vascular endothelial cell-specific p53-deficient mice (EC-p53KO), insulin sensitivity was improved and fat accumulation decreased. It should be noted that p53 is a factor known to be involved in aging.

[0004] These studies suggest that inhibiting vascular aging may contribute to reducing disease risk and extending healthy life expectancy. Approaches to prevent or delay vascular aging include the use of antioxidants and anti-inflammatory drugs, and lifestyle improvements (moderate exercise, balanced diet, smoking cessation, etc.). In recent years, it has also been reported that certain flavonoids and polyphenols may have anti-vascular aging effects. [Prior art documents] [Non-patent literature]

[0005] [Non-Patent Document 1] Han Y, Kim SY. Endothelial senescence in vascular diseases: current understanding and future opportunities in senotherapeutics. Exp Mol Med. 2023;55(1):1-12. doi:10.1038 / s12276-022-00906-w [Non-Patent Document 2] Hasegawa Y, Saito T, Ogihara T, et al. Blockade of the nuclear factor-κB pathway in the endothelium prevents insulin resistance and prolongs life spans. Circulation. 2012;125(9):1122-1133. doi:10.1161 / CIRCULATIONAHA.111.054346 [Non-Patent Document 3] Yokoyama M, Okada S, Nakagomi A, et al. Inhibition of endothelial p53 improves metabolic abnormalities related to dietary obesity. Cell Rep. 2014;7(5):1691-1703. doi:10.1016 / j.celrep.2014.04.046 [Overview of the Initiative] [Problems that the invention aims to solve]

[0006] The inventors have found that quercetin and genistein, individually, suppress the expression of some aging-related genes while promoting the expression of other aging-related genes. Therefore, the main objective was to maintain the repressive effect on some aging-related genes that quercetin, genistein, and their glycosides may have, while suppressing the effect of promoting the expression of other aging-related genes. [Means for solving the problem]

[0007] The inventors have found that by combining two or three components selected from the group consisting of component (i): quercetin and / or its glycosides, component (ii): genistein and / or its glycosides, and component (iii): nobiletin and / or its glycosides, it is possible to suppress the expression-promoting effect on other aging-related genes while maintaining the expression-suppressing effect on some aging-related genes that quercetin and genistein, and their glycosides, may have. Further improvements have been made to complete this disclosure.

[0008] This disclosure includes, for example, the following subjects: Section 1. Ingredient (i): Quercetin and / or its glycosides, Component (ii): Genistein and / or its glycosides, Component (iii): Nobiletin and / or its glycosides An anti-aging composition containing two or three components selected from the group consisting of the following. Section 2. Ingredient (i): Quercetin and / or its glycosides, Component (ii): Genistein and / or its glycosides, Component (iii): A composition for anti-vascular endothelial aging, comprising two or three components selected from the group consisting of nobiletin and / or its glycosides. Section 3. The composition according to item 1 or 2, used for the suppression of cell cycle arrest factors and / or for the suppression of inflammation. Section 4. It contains component (i) and component (ii), and the molar ratio (G / Q) of the content of component (ii) (G) to the content of component (i) (Q) is 1 / 10 to 10. It contains component (i) and component (iii), and the molar ratio (N / Q) of the content of component (iii) to the content of component (i) (Q) is 1 / 10 to 10, or A composition according to any one of claims 1 to 3, comprising component (ii) and component (iii), wherein the molar ratio (N / G) of the content of component (iii) to the content of component (ii) (G) is 1 / 10 to 10. Section 5. It contains component (i) and component (ii), and the molar ratio (G / Q) of the content of component (ii) (G) to the content of component (i) (Q) is 1 / 5 to 5. It contains component (i) and component (iii), and the molar ratio (N / Q) of the content of component (iii) to the content of component (i) (Q) is 1 / 5 to 5, or A composition according to any one of claims 1 to 3, comprising component (ii) and component (iii), wherein the molar ratio (N / G) of the content of component (iii) to the content of component (ii) (G) is 1 / 5 to 5. Section 6. Component (i): Quercetin and / or its glycosides are incorporated into the composition, Component (ii): Genistein and / or its glycosides, Component (iii): Nobiletin and / or its glycosides A method for producing a composition in which the CXCL1 and / or IL-8 expression promoting effect of component (i) is suppressed, characterized by incorporating at least one component selected from the group consisting of the above into the composition. Section 7. Component (ii): Genistein and / or its glycosides are incorporated into the composition, Component (i): Quercetin and / or its glycosides, and Component (iii): Nobiletin and / or its glycoside A method for producing a composition in which the promoting effect of CXCL1 and / or IL-8 expression of component (ii) is suppressed, characterized by blending at least one component selected from the group consisting of Item 8. Component (i): Quercetin and / or its glycoside Component (ii): Genistein and / or its glycoside, and Component (iii): Nobiletin and / or its glycoside A method for suppressing the promoting effect of CXCL1 and / or IL-8 expression of component (i), characterized by using in combination at least one component selected from the group consisting of Item 9. Component (ii): Genistein and / or its glycoside Component (i): Quercetin and / or its glycoside, and Component (iii): Nobiletin and / or its glycoside A method for suppressing the promoting effect of CXCL1 and / or IL-8 expression of component (ii), characterized by using in combination at least one component selected from the group consisting of

Advantages of the Invention

[0009] According to the present disclosure, while maintaining the expression-suppressing effect on some aging-related genes that quercetin, genistein, and their glycosides may have, the expression-promoting effect on other aging-related genes can be suppressed.

Brief Description of the Drawings

[0010] [Figure 1] Test 1: Fluorescence microscope images (left in the figure) and fluorescence intensities (relative values with the fluorescence intensity at p.4 set to 1, right in the figure) when detecting SA-β-gal positive cells using the β-galactosidase detection fluorescent reagent SPiDER-βGal. In the figure, the upper end of the bar graph indicates the average value, and the error bar indicates the standard deviation (the same applies hereinafter). **p<0.01, n = 6. [Figure 2]Experiment 1: Results of analysis of SA-β-gal-positive cells by flow cytometry are shown. **p<0.01, n=5. [Figure 3] Experiment 1: Results of analyzing cells that highly express SA-β-gal by flow cytometry are shown. **p<0.01, n=4. [Figure 4] Experiment 1: The results of cell cycle analysis of p.4 and p.21 cells by flow cytometry are shown. The vertical axis of the graph shows the percentage of total cells (% cell). n=2. [Figure 5] Test 1: Shows gene expression levels for IL-8, CXCL1, ICAM-1, p16, and p21. **p<0.01, n=6. [Figure 6] Test 1: Shows gene expression levels of SIRT1, NRF1, and TFAM. **p<0.01, *p<0.05, n=6. [Figure 7] Test 1: Shows gene expression levels of IL-1β, CCL2, VCAM-1, and E-selectin. **p<0.01 vs p.4. n=6. [Figure 8] Experiment 2: Results when genistein was added are shown. In the figure, C represents the control (no flavonoid added) (the same applies hereafter). n=2. [Figure 9] Test 2: Results when quercetin was added are shown. n=2. [Figure 10] Test 2: Results when nobiletin was added are shown. n=2. [Figure 11] Study 3: Shows the expression levels of CXCL1 and IL-8 when genistein and quercetin are combined. *p<0.05 vs quercetin 20 μM. n=3 [Figure 12] Test 3: Shows p21 expression levels when genistein and nobiletin are combined. *p<0.05 vs genistein 20 μM. n=3 [Figure 13] Experiment 3: Shows the gene expression levels when quercetin and nobiletin are combined. **p<0.01 vs quercetin 20uM, *p<0.05 vs quercetin 20μM. n=4. [Figure 14] (Continued from Figure 13) The gene expression levels when quercetin and nobiletin are combined are shown. **p<0.01 vs quercetin 20μM, *p<0.05 vs quercetin 20μM. n=4. [Figure 15] Experiment 3: Shows the expression levels of each gene when genistein and quercetin are combined. **p<0.01 vs quercetin 20μM, *p<0.05 vs quercetin 20μM. n=4. [Figure 16] (Continued from Figure 15) The expression levels of each gene when genistein and quercetin are combined are shown. **p<0.01 vs quercetin 20μM, *p<0.05 vs quercetin 20μM. n=4. [Figure 17] Experiment 3: Shows the expression levels of each gene when genistein and quercetin are combined. **p<0.01 vs genistein 20μM, *p<0.05 vs genistein 20μM. n=4. [Figure 18] (Continued from Figure 17) This shows the expression levels of each gene when genistein and quercetin are combined. **p<0.01 vs genistein 20μM, *p<0.05 vs genistein 20μM. n=4. [Figure 19] Test 3: Shows the expression levels of each gene when genistein and nobiletin are combined. **p<0.01 vs genistein 20μM, *p<0.05 vs genistein 20μM. n=4. [Figure 20] (Continued from Figure 19) The expression levels of each gene when genistein and nobiletin are combined are shown. **p<0.01 vs genistein 20μM, *p<0.05 vs genistein 20μM. n=4. [Modes for carrying out the invention]

[0011] The embodiments included in this disclosure will be described in more detail below. This disclosure preferably includes, but is not limited to, anti-aging compositions, anti-vascular endothelial aging compositions, etc., and encompasses everything disclosed herein and recognizable to those skilled in the art.

[0012] I. Compositions of the Disclosure The compositions included in this disclosure contain two or three components selected from the group consisting of component (i): quercetin and / or its glycosides, component (ii): genistein and / or its glycosides, and component (iii): nobiletin and / or its glycosides. Hereinafter, the compositions included in this disclosure may be referred to as "the compositions of this disclosure."

[0013] Quercetin is a type of flavonoid and a polyphenol compound that can be found mainly in plants such as fruits, vegetables, and tea leaves. The quercetin used in the technology of this disclosure may be of plant origin or chemically synthesized. The molar mass of quercetin is 302.236 g / mol. The structure of quercetin is shown below. [ka]

[0014] Genistein is a type of isoflavone that can be found in soybeans and other leguminous plants. The genistein used in the art of this disclosure may be of plant origin or may be chemically synthesized. The molar mass of genistein is 270.24 g / mol. The structure of genistein is shown below. [ka]

[0015] Nobiletin is a type of polymethoxyflavonoid that can be found mainly in the peels of citrus fruits. The nobiletin used in the art of this disclosure may be of plant origin or chemically synthesized. The molar mass of nobiletin is 402.39 g / mol. The structure of nobiletin is shown below. [ka]

[0016] In the technologies of this disclosure, quercetin, genistein, and nobiletin may optionally take the form of glycosides bound to sugars. The sugar constituting the glycoside is not particularly limited and may be a monosaccharide, disaccharide, or trisaccharide composed of one or more monosaccharides selected from the group consisting of glucose, galactose, mannose, ribose, deoxyribose, arabinose, xylose, rhamnose, fucose, and fructose. The aforementioned monosaccharide may be the L-form, the D-form, or a mixture of the L-form and D-form, and it is preferable that it be the enantiomer which is more abundant in nature. Therefore, it is preferable that the sugar constituting the glycoside is a monosaccharide, disaccharide, or trisaccharide composed of one or more monosaccharides selected from the group consisting of D-glucose, D-galactose, D-mannose, D-ribose, D-deoxyribose, L-arabinose, D-xylose, L-rhamnose, L-fucose, and D-fructose. Furthermore, the monosaccharides mentioned above may be in the α-form, the β-form, or a mixture of both, but it is preferable that they be the anomer that is more abundant in nature.

[0017] More specifically, disaccharides include maltose, sucrose, lactose, rutinose, cellobiose, trehalose, lactulose, cellobiose, isomaltose, isotrehalose, neotrehalose, turanose, palatinose, and mannobiose. More specifically, trisaccharides include maltotriose, melegitose, raffinose, nigerotriose, and kestose.

[0018] Examples of quercetin glycosides include rutin, quercitrin, and isoquercitrin. Rutin has a structure in which rutinose, composed of D-glucose and L-rhamnose, is O-glycosidically bonded to the hydroxyl group at position 3 of quercetin. Rutin is the 3-O-rutinoside of quercetin. Quercitrin has a structure in which L-rhamnose is O-glycosidically bonded to the hydroxyl group at position 3 of quercetin. Quercitrin is the 3-O-rhamnoside of quercetin. Isoquercitrin has a structure in which D-glucose is O-glycosidically bonded to the hydroxyl group at position 3 of quercetin. Isoquercitrin is the 3-O-glucoside of quercetin.

[0019] Genistin is an example of a glycoside of genistein. Genistin has a structure in which D-glucose is O-glycosidically bonded to the hydroxyl group at position 7 of genistein. Genistin is the 7-O-glucoside of genistein.

[0020] The compositions of this disclosure contain two or three components selected from the group consisting of component (i): quercetin and / or its glycosides, component (ii): genistein and / or its glycosides, and component (iii): nobiletin and / or its glycosides, as described above. In other words, they contain any combination of two components (i) and (ii), two components (i) and (iii), two components (ii) and (iii), or three components (i), (ii), and (iii).

[0021] If the composition of the present disclosure contains both quercetin and a glycoside of quercetin as component (i), the sum of the moles of quercetin and the moles of the glycoside of quercetin is the number of moles of component (i). Similarly, if the composition of the present disclosure contains both genistein and a glycoside of genistein as component (ii), the sum of the moles of genistein and the moles of the glycoside of genistein is the number of moles of component (ii). If the composition of the present disclosure contains both nobiletin and a glycoside of nobiletin as component (iii), the sum of the moles of nobiletin and the moles of the glycoside of nobiletin is the number of moles of component (iii).

[0022] If the composition of this disclosure contains component (i) and component (ii), the molar ratio (G / Q) of the content of component (ii) (G) to the content of component (i) (Q) is not particularly limited, but may be, for example, 1 / 2000 to 2000. The upper or lower limit of the range may be 1 / 2000, 1 / 1500, 1 / 1000, 1 / 800, 1 / 600, 1 / 400, 1 / 200, 1 / 100, 1 / 90, 1 / 80, 1 / 70, 1 / 60, 1 / 50, 1 / 40, 1 / 30, 1 / 20, 1 / 10, 1 / 9, 1 / 8, 1 / 7, 1 / 6, 1 / 5, 1 / 4, 1 / 3, 1 / 2, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 100, 200, 400, 600, 800, 1000, 1500, or 2000. G / Q is preferably 1 / 1000 to 1000 or 1 / 500 to 500, more preferably 1 / 200 to 200 or 1 / 100 to 100, even more preferably 1 / 50 to 50 or 1 / 10 to 10, and particularly preferably 1 / 5 to 5 or 1 / 3 to 3.

[0023] If the composition of this disclosure contains component (i) and component (iii), the molar ratio (N / Q) of the content of component (iii) to the content of component (i) (Q) is not particularly limited, but may be, for example, 1 / 2000 to 2000. The upper or lower limit of the range may be 1 / 2000, 1 / 1500, 1 / 1000, 1 / 800, 1 / 600, 1 / 400, 1 / 200, 1 / 100, 1 / 90, 1 / 80, 1 / 70, 1 / 60, 1 / 50, 1 / 40, 1 / 30, 1 / 20, 1 / 10, 1 / 9, 1 / 8, 1 / 7, 1 / 6, 1 / 5, 1 / 4, 1 / 3, 1 / 2, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 100, 200, 400, 600, 800, 1000, 1500, or 2000. N / Q is preferably 1 / 1000 to 1000 or 1 / 500 to 500, more preferably 1 / 200 to 200 or 1 / 100 to 100, even more preferably 1 / 50 to 50 or 1 / 10 to 10, and particularly preferably 1 / 5 to 5 or 1 / 3 to 3.

[0024] If the composition of the present disclosure contains component (ii) and component (iii), the molar ratio (N / G) of the content of component (iii) to the content (G) of component (ii) is not particularly limited, but may be, for example, 1 / 2000 to 2000. The upper or lower limit of the range may be 1 / 2000, 1 / 1500, 1 / 1000, 1 / 800, 1 / 600, 1 / 400, 1 / 200, 1 / 100, 1 / 90, 1 / 80, 1 / 70, 1 / 60, 1 / 50, 1 / 40, 1 / 30, 1 / 20, 1 / 10, 1 / 9, 1 / 8, 1 / 7, 1 / 6, 1 / 5, 1 / 4, 1 / 3, 1 / 2, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 100, 200, 400, 600, 800, 1000, 1500, or 2000. The N / G ratio is preferably 1 / 1000 to 1000 or 1 / 500 to 500, more preferably 1 / 200 to 200 or 1 / 100 to 100, even more preferably 1 / 50 to 50 or 1 / 10 to 10, and particularly preferably 1 / 5 to 5 or 1 / 3 to 3.

[0025] If the composition of the present disclosure contains both quercetin and a glycoside of quercetin as component (i), the sum of the content of quercetin and the content of a glycoside of quercetin is the content of component (i). Similarly, if the composition of the present disclosure contains both genistein and a glycoside of genistein as component (ii), the sum of the content of genistein and the content of a glycoside of genistein is the content of component (ii). If the composition of the present disclosure contains both nobiletin and a glycoside of nobiletin as component (iii), the sum of the content of nobiletin and the content of a glycoside of nobiletin is the content of component (iii).

[0026] If the composition of this disclosure contains component (i), its content is not particularly limited, but may be, for example, 0.000001 to 1% by mass of the whole composition. The upper or lower limits of the range are 0.000001, 0.000005, 0.00001, 0.00005, 0.0001, 0.0005, 0.001, 0.002, 0.005, 0.008, 0.01, 0.012, 0.014, 0.016, 0.018, 0.02, 0.022, 0.024, 0.026, 0.028, 0.03, 0.032, 0.034, 0.036, 0.038, 0.04, 0.042, 0.044, 0 The amount may be 0.046, 0.048, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.2, 0.22, 0.24, 0.26, 0.28, 0.3, 0.32, 0.34, 0.36, 0.38, 0.4, 0.42, 0.44, 0.46, 0.48, 0.5, 0.6, 0.7, 0.8, 0.9, or 1% by mass. The content of component (i) is preferably 0.00001 to 0.5% by mass, more preferably 0.0001 to 0.2% by mass, and even more preferably 0.001 to 0.1% by mass, relative to the total composition.

[0027] If the composition of this disclosure contains component (ii), its content is not particularly limited, but may be, for example, 0.000001 to 1% by mass relative to the whole composition. The upper or lower limits of the range are 0.000001, 0.000005, 0.00001, 0.00005, 0.0001, 0.0005, 0.001, 0.002, 0.005, 0.008, 0.01, 0.012, 0.014, 0.016, 0.018, 0.02, 0.022, 0.024, 0.026, 0.028, 0.03, 0.032, 0.034, 0.036, 0.038, 0.04, 0.042, 0.044, 0 The amount may be 0.046, 0.048, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.2, 0.22, 0.24, 0.26, 0.28, 0.3, 0.32, 0.34, 0.36, 0.38, 0.4, 0.42, 0.44, 0.46, 0.48, 0.5, 0.6, 0.7, 0.8, 0.9, or 1% by mass. The content of component (ii) is preferably 0.00001 to 0.5% by mass, more preferably 0.0001 to 0.2% by mass, and even more preferably 0.001 to 0.1% by mass, relative to the total composition.

[0028] If the composition of this disclosure contains component (iii), its content is not particularly limited, but may be, for example, 0.000001 to 1% by mass relative to the whole composition. The upper or lower limits of the range are 0.000001, 0.000005, 0.00001, 0.00005, 0.0001, 0.0005, 0.001, 0.002, 0.005, 0.008, 0.01, 0.012, 0.014, 0.016, 0.018, 0.02, 0.022, 0.024, 0.026, 0.028, 0.03, 0.032, 0.034, 0.036, 0.038, 0.04, 0.042, 0.044, 0 The amount may be 0.046, 0.048, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.2, 0.22, 0.24, 0.26, 0.28, 0.3, 0.32, 0.34, 0.36, 0.38, 0.4, 0.42, 0.44, 0.46, 0.48, 0.5, 0.6, 0.7, 0.8, 0.9, or 1% by mass. The content of component (iii) is preferably 0.00001 to 0.5% by mass, more preferably 0.0001 to 0.2% by mass, and even more preferably 0.001 to 0.1% by mass, relative to the total composition.

[0029] The compositions of this disclosure contain two or three components selected from the group consisting of component (i): quercetin and / or its glycosides, component (ii): genistein and / or its glycosides, and component (iii): nobiletin and / or its glycosides, and may further contain other components insofar as the desired effect is obtained. Examples of such other components include pharmacologically or food-hygiene-acceptable bases, carriers, solvents, dispersants, emulsifiers, buffers, stabilizers, excipients, binders, disintegrants, lubricants, thickeners, antioxidants, preservatives, coatings, colorants, etc. Flavonoids and flavonoid glycosides other than components (i) to (iii) may also be included. These components can be used individually or in combination of two or more.

[0030] The compositions of this disclosure are not particularly limited, but may include, for example, oral compositions. The compositions of this disclosure may also include pharmaceutical compositions or food and beverage compositions. If they are food and beverage compositions, they may include, for example, processed foods, beverages, health foods (nutrient function foods, foods with functional claims, foods for specified health uses, etc.), supplements, and foods for the sick (hospital meals, sick person meals, or nursing care meals, etc.).

[0031] The compositions of this disclosure can be prepared by conventional methods by combining two or three components selected from the group consisting of component (i): quercetin and / or its glycosides, component (ii): genistein and / or its glycosides, and component (iii): nobiletin and / or its glycosides, with other components as needed.

[0032] II. Application As described above, the inventors have found that quercetin and genistein, individually, suppress the expression of some aging-related genes while promoting the expression of other aging-related genes. On the other hand, according to the technology of this disclosure, the repressive effect on some aging-related genes that quercetin and genistein, and their glycosides, may have can be maintained while the pro-expression effect on other aging-related genes can be suppressed. More specifically, when the composition of this disclosure contains component (i): quercetin and / or its glycoside, the repressive effect on the expression of the aging-related genes ICAM-1 (Intercellular adhesion molecule-1) and / or p16 that component (i) may have can be maintained while the pro-expression effect on CXCL1 (CXC motif chemokine ligand 1) and IL-8 (Interleukin-8) can be suppressed. When the composition of this disclosure contains component (ii): genistein and / or its glycosides, the inhibitory effect on the expression of at least one gene selected from the group consisting of CXCL1, IL-8, ICAM-1, and p16 that component (ii) may have can be maintained while the p21 expression promoting effect can be suppressed. For this reason, a superior anti-aging effect can be achieved compared to when component (i) or component (ii) is used alone. Note that IL-8, CXCL1, and ICAM-1 are inflammation-related factors, and p16 and p21 are cell cycle regulatory (arrest) factors.

[0033] From the above effects, it is understood that the composition of this disclosure has at least one effect selected from the group consisting of anti-aging effect, anti-vascular endothelial aging effect, cell cycle arrest factor inhibitory effect, and anti-inflammatory effect. Therefore, the composition of this disclosure can be used, for example, in at least one application selected from the group consisting of anti-aging, anti-vascular endothelial aging, cell cycle arrest factor inhibitory effect, and anti-inflammatory effect.

[0034] The anti-aging effect of the compositions disclosed can be confirmed, for example, by adding the compositions to an evaluation system containing senescent cells, which results in a decrease in the expression level of at least one aging-related factor compared to when the compositions are not added. Specific examples of aging-related factors include inflammation-related factors such as IL-8, CXCL1, and ICAM-1, and cell cycle arrest factors such as p16 and p21. The anti-vascular endothelial aging effect of the compositions disclosed can be confirmed, for example, by adding the compositions to an evaluation system containing senescent-induced vascular endothelial cells, which results in a decrease in the expression level of at least one aging-related factor compared to when the compositions are not added. The inhibitory effect of the compositions disclosed on cell cycle arrest factors can be confirmed, for example, by adding the compositions to an evaluation system containing senescent cells (preferably senescent-induced vascular endothelial cells), which results in a decrease in the expression level of at least one cell cycle arrest factor compared to when the compositions are not added. The anti-inflammatory effect of the compositions disclosed herein can be confirmed, for example, by adding the compositions to an evaluation system including senescent cells (preferably senescence-induced vascular endothelial cells), which results in a decrease in the expression level of at least one inflammation-related factor compared to the case where the compositions are not added. In this disclosure, the term "factor" is used to include both genes and proteins.

[0035] The method of administering the compositions of this disclosure is not particularly limited and may be administered, for example, orally or parenterally. Parenteral administration may include, for example, intravenous, arterial, intramuscular, subcutaneous, abdominal, rectal, and topical administration, as well as transdermal administration. The compositions of this disclosure are preferably administered orally.

[0036] The dosage (intake) of the composition disclosed herein is not particularly limited and can be appropriately selected depending on the form and method of administration of the composition disclosed herein. For example, when administered orally to a human (adult), the total intake of two or three components selected from the group consisting of component (i): quercetin and / or its glycosides, component (ii): genistein and / or its glycosides, and component (iii): nobiletin and / or its glycosides may be 0.01 to 10,000 mg per day, preferably 0.1 to 50,000 mg, more preferably 0.3 to 2,000 mg, even more preferably 1 to 1,000 mg, and particularly preferably 1 to 500 mg. The above amount may be administered or ingested to the subject in, for example, 1 to 3 divided doses per day.

[0037] The frequency of administration (ingestion) of the compositions disclosed herein is not particularly limited. For example, they may be administered or ingested once or more times a day, once or more times a week, or once or more times a month. Furthermore, the duration of application of the technology disclosed herein is not particularly limited.

[0038] The subjects to whom the compositions of this disclosure are administered or ingested are not particularly limited, and may be, for example, humans or other mammals. Examples of other mammals include rats, mice, rabbits, cattle, pigs, dogs, cattle, sheep, and monkeys.

[0039] While not particularly limited, the compositions of this disclosure are preferably applied to persons aged 40 years or older, more preferably to persons aged 45 years or older, even more preferably to persons aged 50 years or older or 55 years or older, and particularly preferably to persons aged 60 years or older. The lower limit of the range may be 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, or 70 years.

[0040] Preferably, the compositions disclosed herein are intended for use by people who want to prevent aging, people who want to suppress aging, people who want to improve aging, people who want to prevent aging of blood vessels (especially vascular endothelium), people who want to suppress aging of blood vessels (especially vascular endothelium), people who want to improve aging of blood vessels (especially vascular endothelium), people who want to maintain and / or improve vascular function, people who want to maintain and / or improve the flexibility of blood vessels that decreases with age, people who want to prevent inflammation (especially inflammation in the vascular endothelium), people who want to suppress inflammation (especially inflammation in the vascular endothelium), and people who want to improve inflammation (especially inflammation in the vascular endothelium).

[0041] The disclosure also includes a method for producing a composition in which the CXCL1 and / or IL-8 expression-promoting effect of component (i) is suppressed, characterized by incorporating component (i): quercetin and / or its glycoside into the composition, and incorporating at least one component selected from the group consisting of component (ii): genistein and / or its glycoside, and component (iii): nobiletin and / or its glycoside into the composition; and a method for producing a composition in which the CXCL1 and / or IL-8 expression-promoting effect of component (ii) is suppressed, characterized by incorporating component (ii): genistein and / or its glycoside into the composition, and incorporating at least one component selected from the group consisting of component (i): quercetin and / or its glycoside, and component (iii): nobiletin and / or its glycoside into the composition. Furthermore, this disclosure also includes a method for suppressing the CXCL1 and / or IL-8 expression-promoting effect of component (i), characterized by using component (i): quercetin and / or its glycosides in combination with at least one component selected from the group consisting of component (ii): genistein and / or its glycosides, and component (iii): nobiletin and / or its glycosides; and a method for suppressing the CXCL1 and / or IL-8 expression-promoting effect of component (ii), characterized by using component (ii): genistein and / or its glycosides in combination with at least one component selected from the group consisting of component (i): quercetin and / or its glycosides, and component (iii): nobiletin and / or its glycosides. The contents described with respect to the compositions of this disclosure may be appropriately incorporated into the respective manufacturing methods and methods described above.

[0042] In this specification, the term “comprising” includes not only “containing” but also “essentially consisting of” and “consisting of.” Furthermore, this disclosure encompasses all combinations of the constituent elements described herein.

[0043] Furthermore, the various characteristics (properties, structure, function, etc.) described in each embodiment of this disclosure above may be combined in any way to identify the subject matter covered by this disclosure. In other words, this disclosure covers all subject matter consisting of any combination of the combinable characteristics described herein. [Examples]

[0044] The embodiments of this disclosure will be described in more detail below with examples, but the embodiments of this disclosure are not limited to the examples below. In these embodiments, quercetin may be referred to as "Q", genistein as "G", and nobiletin as "N". The density of the culture medium used for cell culture was approximately 1.0 g / mL in all cases.

[0045] Experiment 1. Induction of aging in human vascular endothelial cells 1-1. Method 1-1-1. Evaluation of SA-β-gal-positive cells (Staining - Microscope) Human umbilical vein endothelial cells (HUVEC, Lonza) were subcultured until they reached passage 4 (Passage 4, hereafter p.4) or passage 21 (Passage 21, hereafter p.21). Subsequently, p.4 cells measured 2.0 × 10⁶ cells. 4 At a density of cells / 1 mL / well, P.21 is 2.5 × 10⁻⁶ 4Cells were seeded at a density of 1 mL / well into 12-well plates and cultured for 3 days in growth medium (MCDB 131 medium (Thermo Fisher Scientific) containing 10% FBS (Fetal Bovine Serum, Biowest), 1% Antibiotics (Gibco), 2 mM L-Glutamine (Fujifilm Wako Pure Chemical Corporation), and 10 ng / mL FGF (Kaken Pharmaceutical Co., Ltd)). After washing once with PBS(-), Bafilomycin A1 (Focus Biomolecules), diluted to 100 nM in evaluation medium (MCDB 131 medium (Thermo Fisher Scientific) containing 10% FBS (Fetal Bovine Serum, Biowest), 1% Antibiotics (Gibco), and 2 mM L-Glutamine (Fujifilm Wako Pure Chemical Corporation)), was added and incubated for 1 hour.

[0046] After 1 hour, the cells were washed twice with PBS(-) and fixed for 3 minutes using 4% paraformaldehyde-phosphate buffer (FUJIFILM Wako Pure Chemical). After washing three times with PBS(-), SPiDER-βGal (Dojindo), dissolved in DMSO and diluted with McIlvaine buffer (pH 6.0), was added and incubated for 30 minutes. Then, Hoechst 33342 (Thermo Scientific), diluted with PBS(-) containing 0.3% Triton X-100 (Sigma-Aldrich), was added and incubated for 15 minutes. After washing three times with PBS(-), fluorescence images were acquired using the EVOS M7000 Imaging System (Thermo Scientific). Fluorescence intensity was quantified by processing the fluorescence images using Celleste 5 Image Analysis Software (Thermo Scientific).

[0047] 1-1-2. Evaluation of SA-β-gal-positive cells (Staining-Flow cytometry) Human umbilical vein endothelial cells (HUVEC, Lonza) were subcultured until they reached passage 4 (Passage 4, hereafter p.4) or passage 21 (Passage 21, hereafter p.21). Subsequently, p.4 cells measured 2.0 × 10⁶ cells. 4 At a density of cells / 1 mL / well, P.21 is 2.5 × 10⁻⁶ 4 Cells were seeded at a density of 1 mL / well into 12-well plates and cultured for 3 days in growth medium (MCDB 131 medium (Thermo Fisher Scientific) containing 10% FBS (Fetal Bovine Serum, Biowest), 1% Antibiotics (Gibco), 2 mM L-Glutamine (Fujifilm Wako Pure Chemical Corporation), and 10 ng / mL FGF (Kaken Pharmaceutical Co., Ltd)). After washing once with HBSS, Bafilomycin A1 (Focus Biomolecules), diluted to 100 nM in evaluation medium (MCDB 131 medium (Thermo Fisher Scientific) containing 10% FBS (Fetal Bovine Serum, Biowest), 1% Antibiotics (Gibco), and 2 mM L-Glutamine (Fujifilm Wako Pure Chemical Corporation)), was added and incubated for 1 hour.

[0048] After 1 hour, SPiDER-βGal (Dojindo), dissolved in DMSO and diluted in evaluation medium, was added and incubated for 30 minutes. After washing once with HBSS, the cells were detached from the plate using trypsin and collected. The collected cells were centrifuged at 4°C (1,000 rpm, 5 minutes), and the supernatant was removed. The cells were washed again with HBSS, centrifuged at 4°C (1,000 rpm, 5 minutes), and the supernatant was aspirated and removed. Subsequently, the precipitated cells were dispersed in HBSS, and then dispersed individually using a cell strainer (40 μm, Corning). The dispersed cells were analyzed by fluorescence measurement using a BD Accuri C6 Plus Flow Cytometer (BD Biosciences).

[0049] 1-1-3. Cell cycle evaluation (Staining-flow cytometry) Human umbilical vein endothelial cells (HUVEC, Lonza) were subcultured until they reached passage 4 (Passage 4, hereafter p.4) or passage 21 (Passage 21, hereafter p.21). Subsequently, p.4 cells measured 2.0 × 10⁶ cells. 4 At a density of cells / 1 mL / well, P.21 is 2.5 × 10⁻⁶ 4Cells were seeded at a density of 1 mL / well into 12-well plates and cultured for 3 days in growth medium (MCDB 131 medium (Thermo Fisher Scientific) containing 10% FBS (Fetal Bovine Serum, Biowest), 1% Antibiotics (Gibco), 2 mM L-Glutamine (Fujifilm Wako Pure Chemical Corporation), and 10 ng / mL FGF (Kaken Pharmaceutical Co., Ltd)). After reaching confluence, the cells were washed once with PBS(-), detached from the plate using trypsin, and collected. The collected cells were centrifuged at 20°C (300 g, 5 min), the supernatant was removed, and the cells were dispersed in PBS(-). A predetermined number of cells were mixed with Cell Cycle Assay Solution Deep Red (Dojindo) and incubated at 37°C under light-shielded conditions for 15 minutes. After dispersing the cells individually using a cell strainer (40 μm, Corning), fluorescence measurements were performed and analyzed using a BD Accuri C6 Plus Flow Cytometer (BD Biosciences).

[0050] 1-1-4. Evaluation of the expression of inflammation-related genes, cell cycle regulatory genes, and mitochondrial function-related genes (qPCR) Human umbilical vein endothelial cells (HUVEC, Lonza) were subcultured until they reached passage 4 (Passage 4, hereafter p.4) or passage 21 (Passage 21, hereafter p.21). Subsequently, p.4 cells measured 2.0 × 10⁶ cells. 4 At a density of cells / 1 mL / well, P.21 is 2.5 × 10⁻⁶ 4Cells were seeded at a density of 1 mL / well into 12-well plates and cultured for 3 days in growth medium (MCDB 131 medium (Thermo Fisher Scientific) containing 10% FBS (Fetal Bovine Serum, Biowest), 1% Antibiotics (Gibco), 2 mM L-Glutamine (Fujifilm Wako Pure Chemical Corporation), and 10 ng / mL FGF (Kaken Pharmaceutical Co., Ltd)). After reaching confluence, the cells were switched to evaluation medium (MCDB 131 medium (Thermo Fisher Scientific) containing 10% FBS (Fetal Bovine Serum, Biowest), 1% Antibiotics (Gibco), and 2 mM L-Glutamine (Fujifilm Wako Pure Chemical Corporation)) and cultured for 30 hours.

[0051] Total RNA was extracted from cultured cells using the RNeasy Mini Kit (Qiagen). cDNA was synthesized from the extracted total RNA using the PrimeScript RT reagent Kit (Takara Bio). Using the synthesized cDNA as a template, gene expression was quantified using QuantStudio 5 Real-Time PCR (Thermo Fisher Scientific) with specific primers for each gene and an intercalator method using TB Green Premix Ex Taq II FAST qPCR (Takara Bio). The expression levels of each gene were corrected to the RPS18 expression levels, and the relative values ​​at p.21 were calculated, with p.4 expression set as 1. The primer sets used for quantifying each gene expression are shown in the table below. These primer sets were also used in the tests described later.

[0052] [Table 1]

[0053] 1-2. Results Senescent cells are known to be positive for SA-β-gal (senescence-associated beta galactosidase). The results regarding SA-β-gal positive cells are shown in Figures 1-3. Figure 1 shows fluorescence microscope images (left) and fluorescence intensity (relative value with fluorescence intensity at p.4 set to 1, right) when SA-β-gal positive cells were detected using the β-galactosidase detection fluorescent reagent SPiDER-βGal. Figure 2 shows the results of flow cytometry analysis of SA-β-gal positive cells. Figure 3 shows the results of flow cytometry analysis of cells that highly express SA-β-gal. As shown in Figures 1-3, the expression of SA-β-gal, a phenotype of senescence, was shown to be upregulated at p.21. These results suggest that senescence is induced in HUVEC cells that have undergone repeated passage.

[0054] Figure 4 shows the results of flow cytometry analysis of the cell cycles of cells p.4 and p.21. As shown in Figure 4, p.21 showed a decrease in G0 / G1 and S phase cells, and an increase in G2 phase cells.

[0055] The expression levels of each gene are shown in Figures 5-7. Note that all the genes evaluated are those reported to be associated with aging (aging-related genes). The genes shown in Figure 5 all showed significantly increased expression at p.21. This result is consistent with previous findings that the expression of these genes is induced in senescent cells. The genes shown in Figure 6 are all mitochondrial function-related genes, and their expression was significantly decreased. This result is consistent with previous findings that mitochondrial function is impaired in senescent cells. Contrary to previous findings regarding senescent cells, the genes shown in Figure 7 either showed significantly decreased expression at p.21 or there was no significant difference in expression levels between p.4 and p.21.

[0056] In subsequent tests, the inventors used the expression levels of IL-8, CXCL1, ICAM-1, p16, and p21 shown in FIG. 5 as indicators of cell senescence. Note that IL-8, CXCL1, and ICAM-1 are inflammation-related factors, and p16 and p21 are cell cycle control (arrest) factors.

[0057] Experiment 2. Effects of flavonoids on aging vascular endothelial cells (1) - Effects of single flavonoids 2-1. Method Genistein (Tokyo Chemical Industry), quercetin (Tokyo Chemical Industry), and nobiletin (Tokyo Chemical Industry) were each dissolved using DMSO (Dimethyl sulfoxide, FUJIFILM Wako Pure Chemical Corporation), and these were used as flavonoid samples for subsequent cell tests (the same applies in Test 3).

[0058] Human umbilical vein endothelial cells (HUVEC, Lonza) of p.21 were 2.0 × 10 4Cells were seeded at a density of 1 mL / well in 12-well plates and cultured for 3 days in growth medium (MCDB 131 medium (Thermo Fisher Scientific) containing 10% FBS (Fetal Bovine Serum, Biowest), 1% Antibiotics (Gibco), 2 mM L-Glutamine (Fujifilm Wako Pure Chemical Corporation), and 10 ng / mL FGF (Kaken Pharmaceutical Co., Ltd)). After reaching confluence, the medium was changed to evaluation medium (MCDB 131 medium (Thermo Fisher Scientific) containing 10% FBS (Fetal Bovine Serum, Biowest), 1% Antibiotics (Gibco), and 2 mM L-Glutamine (Fujifilm Wako Pure Chemical Corporation)). Genistein, quercetin, and nobiletin flavonoids, diluted to final concentrations of 5, 10, 20, 40, and 80 μM in the evaluation medium, were added individually and incubated for 6 hours.

[0059] Total RNA was extracted from cells using the RNeasy Mini Kit (Qiagen). cDNA was synthesized from the extracted total RNA using the PrimeScript RT reagent Kit (Takara Bio). Using the synthesized cDNA as a template, gene expression was quantified using QuantStudio 5 Real-Time PCR (Thermo Fisher Scientific) with primers specific to each gene and an intercalator method using TB Green Premix Ex Taq II FAST qPCR (Takara Bio). The expression levels of each gene were corrected for the expression level of RPS18, and the relative values ​​were calculated with the gene expression level when p.21 was cultured in evaluation medium without flavonoids set to 1.

[0060] 2-2. Results Figure 8 shows the results when genistein was added. In the figure, C represents the control (no flavonoid addition) (the same applies hereafter). As shown in Figure 8, genistein addition resulted in a decrease in the expression levels of CXCL1, IL-8, ICAM-1, and p16 compared to the control, but an increase in the expression level of p21. In other words, it was suggested that genistein alone suppresses the expression of aging-related genes CXCL1, IL-8, ICAM-1, and p16 in senescent endothelial cells, while increasing the expression of p21.

[0061] Figure 9 shows the results when quercetin was added. As shown in Figure 9, the expression levels of ICAM-1 and p16 decreased compared to the control group with the addition of quercetin, but the expression levels of CXCL1 and IL-8 increased. This suggests that quercetin alone suppresses the expression of aging-related genes ICAM-1 and p16 in senescent endothelial cells, while increasing the expression of CXCL1 and IL-8.

[0062] The results when nobiletin was added are shown in Figure 10. As shown in Figure 10, no genes showed a significant increase in expression.

[0063] 2-3. Summary From Experiment 2, it was suggested that genistein and quercetin have the effect of suppressing the expression of some aging-related genes (i.e., a desirable effect), while also increasing the expression of other aging-related genes (i.e., an undesirable effect). Therefore, the inventors then attempted to suppress the undesirable effects of genistein and quercetin while maintaining the desirable effects by combining them with flavonoids.

[0064] Experiment 3. Effects of flavonoids on aging vascular endothelial cells (2) - Effects of flavonoid combinations 3-1. Method Human umbilical vein endothelial cells (HUVEC, Lonza) on page 21 were measured in 2.0 × 10⁻¹⁶ units. 4Cells were seeded at a density of 1 mL / well in 12-well plates and cultured for 3 days in growth medium (MCDB 131 medium (Thermo Fisher Scientific) containing 10% FBS (Fetal Bovine Serum, Biowest), 1% Antibiotics (Gibco), 2 mM L-Glutamine (Fujifilm Wako Pure Chemical Corporation), and 10 ng / mL FGF (Kaken Pharmaceutical Co., Ltd)). After reaching confluence, the medium was changed to MCDB 131 medium (Thermo Fisher Scientific) containing 10% FBS (Fetal Bovine Serum, Biowest), 1% Antibiotics (Gibco), and 2 mM L-Glutamine (Fujifilm Wako Pure Chemical Corporation). Three single flavonoids—genistein, quercetin, and nobiletin—as well as combinations of two flavonoids selected from the three, were added so that the final concentrations of each flavonoid reached the concentrations shown in Figures 8-20, and incubated for 6 hours.

[0065] Total RNA was extracted from cells using the RNeasy Mini Kit (Qiagen). cDNA was synthesized from the extracted total RNA using the PrimeScript RT reagent Kit (Takara Bio). Using the synthesized cDNA as a template, gene expression was quantified using QuantStudio 5 Real-Time PCR (Thermo Fisher Scientific) with primers specific to each gene and an intercalator method using TB Green Premix Ex Taq II FAST qPCR (Takara Bio). The expression levels of each gene were corrected for the expression level of RPS18, and the relative values ​​were calculated with the gene expression level when p.21 was cultured in evaluation medium without flavonoids set to 1.

[0066] 3-2. Results As described above, quercetin alone can increase the expression of CXCL1 and IL-8 in senescent endothelial cells. Furthermore, genistein alone can increase the expression of p21 in senescent endothelial cells, which presents a challenge. Therefore, the inventors evaluated the expression levels of CXCL1 and IL-8 when quercetin and genistein were combined, and the expression level of p21 when genistein and nobiletin were combined. The results are shown in Figures 11 and 12.

[0067] As shown in Figure 11, when combined with genistein, the effect of quercetin in increasing CXCL1 and IL-8 expression, which was present when added alone, was significantly suppressed. Furthermore, as shown in Figure 12, when combined with nobiletin, the effect of genistein in increasing p21 expression, which was present when added alone, was significantly suppressed.

[0068] The inventors evaluated the expression levels of each gene in the same manner as in the above test, by changing the ratio of each flavonoid combination. The expression levels of each gene when quercetin and nobiletin are combined are shown in Figures 13 and 14. In the figures, "p.21" represents the control without the addition of flavonoids (the same applies hereafter). As shown in Figure 13, the CXCL1 and IL-8 expression-promoting effect of quercetin alone was suppressed when combined with nobiletin. Furthermore, the ICAM-1, p.16, and p.21 expression-suppressing effects that quercetin and nobiletin each possessed individually were maintained even when quercetin and nobiletin were combined.

[0069] Figures 15-18 show the expression levels of each gene when genistein and quercetin are combined. Note that Figures 15 and 17, and Figures 16 and 18, are graphs showing the same data, but differ in that Figures 15 and 16 perform significance tests on the data for 20 μM quercetin (Q 20 μM), while Figures 17 and 18 perform significance tests on the data for 20 μM genistein (G 20 μM). As shown in Figures 15-18, the CXCL1 and IL-8 expression-promoting effects of quercetin alone were suppressed when combined with genistein. The p.21 expression-promoting effect of genistein alone was suppressed when combined with quercetin. Furthermore, the ICAM-1 and p16 expression-promoting effects of genistein alone, and those of quercetin alone, were maintained when genistein and quercetin were combined.

[0070] Figures 19 and 20 show the expression levels of each gene when genistein and nobiletin are combined. As shown in Figures 19 and 20, the p.21 expression-promoting effect of genistein alone was suppressed when combined with nobiletin. Furthermore, the ICAM-1 expression-suppressing effect that genistein alone possessed was maintained even when combined with quercetin and nobiletin.

Claims

1. Component (i): Quercetin and / or its glycosides, Component (ii): Genistein and / or its glycosides, Ingredients (iii): Nobiletin and / or its glycosides An anti-aging composition containing two or three components selected from the group consisting of the following.

2. Component (i): Quercetin and / or its glycosides, Component (ii): Genistein and / or its glycosides, Ingredient (iii): An anti-vascular endothelial aging composition containing two or three ingredients selected from the group consisting of nobiletin and / or its glycosides.

3. The composition according to claim 1 or 2, used for suppressing cell cycle arrest factors and / or inflammation.

4. It contains component (i) and component (ii), and the molar ratio (G / Q) of the content of component (ii) (G) to the content of component (i) (Q) is 1 / 10 to 10. It contains component (i) and component (iii), and the molar ratio (N / Q) of the content of component (iii) to the content of component (i) (Q) is 1 / 10 to 10, or The composition according to claim 1 or 2, comprising component (ii) and component (iii), wherein the molar ratio (N / G) of the content of component (iii) to the content of component (ii) (G) is 1 / 10 to 10.

5. It contains component (i) and component (ii), and the molar ratio (G / Q) of the content of component (ii) (G) to the content of component (i) (Q) is 1 / 5 to 5. It contains component (i) and component (iii), and the molar ratio (N / Q) of the content of component (iii) to the content of component (i) (Q) is 1 / 5 to 5, or The composition according to claim 1 or 2, comprising component (ii) and component (iii), wherein the molar ratio (N / G) of the content of component (iii) to the content of component (ii) (G) is 1 / 5 to 5.

6. Component (i): A composition comprising quercetin and / or its glycosides, Component (ii): Genistein and / or its glycosides, Ingredients (iii): Nobiletin and / or its glycosides A method for producing a composition in which the CXCL1 and / or IL-8 expression promoting effect of component (i) is suppressed, characterized by incorporating at least one component selected from the group consisting of the above into the composition.

7. Ingredient (ii): Genistein and / or its glycosides are incorporated into the composition, Component (i): Quercetin and / or its glycosides, Ingredients (iii): Nobiletin and / or its glycosides A method for producing a composition in which the CXCL1 and / or IL-8 expression promoting effect of component (ii) is suppressed, characterized by incorporating at least one component selected from the group consisting of the above into the composition.

8. Ingredient (i): Quercetin and / or its glycosides, Component (ii): Genistein and / or its glycosides, Ingredients (iii): Nobiletin and / or its glycosides A method for suppressing the CXCL1 and / or IL-8 expression-promoting effect of component (i), characterized by using it in combination with at least one component selected from the group consisting of the following.

9. Ingredient (ii): Genistein and / or its glycosides, Component (i): Quercetin and / or its glycosides, Ingredients (iii): Nobiletin and / or its glycosides A method for suppressing the CXCL1 and / or IL-8 expression-promoting effect of component (ii), characterized by using it in combination with at least one component selected from the group consisting of the following.

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