Adzuki bean-derived food material and production method of the same

Abstract

To provide an adzuki bean-derived food material including a bile acid micelle dissolubility inhibiting action and a bile acid binding insolubilized material forming action derived from adzuki beans, and to provide a production method of the same.SOLUTION: An adzuki bean-derived food material as a food material derived from Vigna angularis contains (+)-catechin 7-O-β-D-glucopyranoside, (+)-epicatechin 7-O-β-D-glucopyranoside and (+)-catechin, and includes a bile acid micelle dissolubility inhibiting action and a bile acid binding insolubilized matter forming action. A production method of the adzuki bean-derived food material includes the steps of: an adzuki bean immersion step of immersing adzuki beans in ethanol-containing water; a liquid matter collecting step of removing adzuki beans from the immersion water after the adzuki bean immersion step, and collecting a liquid matter; and a drying step of drying the liquid matter obtained in the liquid matter collecting step so as to obtain a dried matter.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a food material derived from adzuki beans, which has the effect of inhibiting bile acid micelle solubility and the effect of forming bile acid-bound insolubilized products, and a method for producing the food material. [Background technology]

[0002] Azuki beans (Vigna angularis) have been commonly consumed since ancient times, primarily through cooking (cooking) to soften them and used as an ingredient in dishes such as red rice, or with added sugar to be made into confections such as "ogura bean paste." The pericarp (skin) of azuki beans is known to contain polyphenols, saponins, and even glycosides bound to sugar molecules. Analysis of these components from the boiling water of azuki beans, i.e., hot water extracts, has revealed that they have antiallergic activity, hypoglycemic activity, inhibition of cell adhesion, prevention of hyperlipidemia, and regulation of bone metabolism.

[0003] Among natural substances, including those contained in adzuki beans, research is being conducted on the components and effects related to glucose metabolism. Abnormal glucose metabolism in particular makes it difficult to control blood sugar levels, which is a major cause of diabetes. When diabetes worsens, it can cause blindness and peripheral necrosis. Diabetes is a type of lifestyle-related disease, and there are concerns that its onset may be due to an accumulation of irregular eating habits and lifestyle habits. Therefore, it may be possible to prevent the onset of the disease by taking measures based on daily eating habits and lifestyle habits.

[0004] Taking this into consideration, the applicant of the present application has clarified the relationship between the catechins contained in adzuki bean seeds, (+)-catechin 7-O-β-D-glucopyranoside and (+)-epicatechin 7-O-β-D-glucopyranoside, and sugar metabolism enzymes, and has provided sugar metabolism enzyme inhibitors based on the catechins in adzuki beans (Patent Document 1). [Prior art documents] [Patent documents]

[0005] [Patent Document 1] Patent Publication No. 2022-30599 Summary of the Invention [Problem to be solved by the invention]

[0006] The present inventors have investigated further the ecological effects of adzuki bean components, particularly (+)-catechin 7-O-β-D-glucopyranoside, (+)-epicatechin 7-O-β-D-glucopyranoside, and (+)-catechin. To investigate the cholesterol-lowering effects in vitro, we used cholesterol micelles, bile acids, and Caco-2 cells as an intestinal model. (+)-Catechin 7-O-β-D-glucopyranoside, (+)-epicatechin 7-O-β-D-glucopyranoside, and (+)-catechin each inhibited micelle solubility in a dose-dependent manner (IC50 values: C7G: 0.23 ± 0.03 mg / ml; E7G: 0.22 ± 0.02 mg / ml). Furthermore, (+)-Catechin 7-O-β-D-glucopyranoside and (+)-epicatechin 7-O-β-D-glucopyranoside exhibited binding activity with bile acids, rendering them insoluble. When Caco-2 cells were treated with C7G or E7G, the amount of cholesterol taken up was significantly reduced compared to control cells, and no cytotoxicity was observed under the experimental conditions used. As a result, the inventors of the present invention have discovered a food material having the effect of inhibiting the uptake of cholesterol in the body and a method for producing the same.

[0007] An object of the present invention is to provide a food material derived from adzuki beans that has the effect of inhibiting the micelle solubility of bile acids derived from adzuki beans and the effect of forming bile acid-bound insolubilized products, and a method for producing the same. [Means for solving the problem]

[0008] The above objectives are achieved by the following: (1) A food material derived from adzuki beans, which contains (+)-catechin 7-O-β-D-glucopyranoside, (+)-epicatechin 7-O-β-D-glucopyranoside, and (+)-catechin, and which has the effect of inhibiting bile acid micelle solubility and forming bile acid-bound insoluble products.

[0009] (2) The food material derived from adzuki beans according to (1) above, which has an inhibitory effect on the uptake of cholesterol in the body. (3) The food material derived from adzuki beans according to (1) or (2) above, wherein the food material is derived from alcohol-containing soaking water for adzuki beans. (4) The food material derived from adzuki beans according to any one of (1) to (3) above, wherein the food material derived from adzuki beans is a freeze-dried product.

[0010] The above objectives are also achieved by the following: (5) A method for producing a food material derived from adzuki beans, comprising the steps of: soaking adzuki beans in ethanol-containing water; removing the adzuki beans from the soaking water after the soaking; collecting a liquid; and drying the liquid obtained in the liquid collecting step to obtain a dried product, thereby obtaining a food material containing (+)-catechin 7-O-β-D-glucopyranoside or (+)-catechin 7-O-β-D-glucopyranoside and (+)-epicatechin 7-O-β-D-glucopyranoside, and having the ability to inhibit bile acid micelle solubility and form bile acid-bound insoluble products.

[0011] (6) The method for producing a food material derived from adzuki beans according to (5) above, wherein the food material derived from adzuki beans contains (+)-catechin. (7) The method for producing a food material derived from adzuki beans according to (5) or (6) above, wherein the adzuki bean soaking step uses ethanol-containing water having an ethanol concentration of 30 to 50%. (8) The method for producing a food material derived from adzuki beans according to any one of (5) to (7) above, wherein the adzuki bean soaking step comprises soaking adzuki beans in the ethanol-containing water for 12 hours or more. (9) The method for producing a food material derived from adzuki beans according to any one of (5) to (8) above, wherein the drying step is a heat-drying step or a vacuum freeze-drying step. (10) The method for producing a food material derived from adzuki beans according to any one of (5) to (9) above, wherein the soaking time in the adzuki bean soaking step is 12 to 48 hours. (11) A method for producing a food material derived from adzuki beans according to any one of (5) to (10) above, wherein the amount of ethanol-containing water used for soaking the adzuki beans in the adzuki bean soaking step is from the same amount to twice the weight of the adzuki beans. (12) A method for producing a food material derived from adzuki beans, comprising the steps of: soaking dried adzuki beans with their outer skins in ethanol-containing water having an ethanol concentration of 30 to 50% for 12 to 48 hours; removing the adzuki beans from the soaking water after the adzuki bean soaking step to collect a liquid; and drying the liquid obtained in the liquid collection step to obtain a dried product, thereby obtaining a food material containing (+)-catechin 7-O-β-D-glucopyranoside or (+)-catechin 7-O-β-D-glucopyranoside, (+)-epicatechin 7-O-β-D-glucopyranoside, and (+)-catechin, and having the ability to inhibit bile acid micelle solubility and form bile acid-bound insoluble products. [Effects of the Invention]

[0012] The adzuki bean-derived food material of the present invention contains (+)-catechin 7-O-β-D-glucopyranoside, (+)-epicatechin 7-O-β-D-glucopyranoside, and (+)-catechin, and has the effect of inhibiting bile acid micelle solubility and forming bile acid-bound insoluble products. Therefore, it has the effect of inhibiting the micelle solubility of bile acids and inhibiting the uptake of cholesterol by forming bile acid-bound insolubilized products, and is derived from adzuki beans, so it is safe to ingest.

[0013] The method for producing a food material derived from adzuki beans of the present invention includes an adzuki bean soaking step in which adzuki beans are soaked in ethanol-containing water, a liquid collection step in which the adzuki beans are removed from the soaking water after the adzuki bean soaking step and a liquid collection step in which the liquid collection step is dried to obtain a dried product, thereby obtaining a food material containing (+)-catechin 7-O-β-D-glucopyranoside or (+)-catechin 7-O-β-D-glucopyranoside and (+)-epicatechin 7-O-β-D-glucopyranoside and having the ability to inhibit bile acid micelle solubility and form bile acid-bound insolubilized products. This makes it possible to easily produce a food material derived from adzuki beans that has the effect of inhibiting bile acid micelle solubility and inhibiting cholesterol uptake in the body by forming bile acid-bound insolubilized products, and is derived from adzuki beans and is safe to ingest. [Brief explanation of the drawings]

[0014] [Figure 1] FIG. 1 is a graph showing the test results for examples of the food material derived from adzuki beans of the present invention. [Figure 2] FIG. 2 is a graph showing the test results for examples of the food material derived from adzuki beans of the present invention. DETAILED DESCRIPTION OF THE INVENTION

[0015] The food material derived from adzuki beans of the present invention will now be described. The adzuki bean-derived food material of the present invention is derived from adzuki beans. The adzuki bean-derived food material contains (+)-catechin 7-O-β-D-glucopyranoside (hereinafter referred to as "C7G"), (+)-epicatechin 7-O-β-D-glucopyranoside (hereinafter referred to as "E7G"), and (+)-catechin, and has the effects of inhibiting bile acid micelle solubility and forming bile acid-bound insolubilized products.

[0016] Furthermore, the azuki bean-derived food material of the present invention contains (+)-catechin 7-O-β-D-glucopyranoside, (+)-epicatechin 7-O-β-D-glucopyranoside, and (+)-catechin, and has an inhibitory effect on the bioinsulation of cholesterol (cholesterol absorption inhibitory effect) due to its inhibitory effect on bile acid micelle solubility and its effect on the formation of bile acid-bound insoluble compounds.

[0017] The azuki bean-derived food material of the present invention is preferably derived from alcohol-containing soaking water for azuki beans. The azuki bean-derived food material is preferably a dried product. The dried product is preferably a freeze-dried product or a heat-dried product.

[0018] To investigate the cholesterol-lowering activity of C7G and E7G, we used cholesterol micelles, bile acids, and Caco-2 cells as an intestinal model. We confirmed that C7G and E7G each inhibited micelle solubility in a dose-dependent manner, with inhibitory activity comparable to that of (+)-catechin (IC50 values: C7G, 0.23 ± 0.03 mg / ml; E7G, 0.22 ± 0.02 mg / ml; (+)-catechin, 0.26 ± 0.11 mg / ml).

[0019] It has been confirmed that the food material of the present invention, specifically a food material derived from alcohol-containing soaking water for adzuki beans, contains a large amount of (+)-catechin 7-O-β-D-glucopyranoside (C7G, "Chemical Formula 1" below), as well as (+)-epicatechin 7-O-β-D-glucopyranoside (E7G, "Chemical Formula 2" below) and (+)-catechin ("Chemical Formula 3" below, molecular weight 290).

[0020] [ka]

[0021] [ka]

[0022] [ka]

[0023] The development of metabolic syndrome is associated with several risk factors, such as high blood cholesterol, high blood pressure, and obesity. Patients with high blood cholesterol are said to be at risk of developing serious heart and circulatory diseases.

[0024] In the body, cholesterol is ingested through bile and daily diet. To be utilized, cholesterol undergoes intestinal absorption through two key steps. The first step is the micellization of cholesterol, which facilitates its efficient translocation across the brush border membrane (BBM) of enterocytes. This step requires bile acids, which are produced from cholesterol and secreted from the liver. Bile acids are functional components of cholesterol micelles that surround cholesterol. Because cholesterol has low solubility and is unlikely to cross the BBM without micellization, micelle formation is necessary for cholesterol absorption. The second step is transport across the BBM by Niemann-Pick C1 Like-1 (NPC1L1), a transporter expressed on the plasma membrane.

[0025] NPC1L1 plays an important role in cholesterol uptake. After this two-step process, excess cholesterol taken up from the intestine accumulates in blood vessels and blocks blood flow, resulting in the risk of the diseases mentioned above. To reduce these risks, it is effective to lower blood cholesterol levels by inhibiting the formation of cholesterol micelles or suppressing cholesterol absorption through the intake of certain medications and foods.

[0026] Cholesterol can pass through the unstirred aqueous layer in the intestinal tract by dissolving in bile acid micelles. Therefore, the food material of the present invention contains (+)-catechin 7-O-β-D-glucopyranoside, (+)-epicatechin 7-O-β-D-glucopyranoside, and (+)-catechin, and thereby has a bile acid micelle solubility inhibitory effect that inhibits the solubility of bile acid micelles in which cholesterol is dissolved, and a bile acid-bound insolubilized product forming effect that inhibits the formation of bile acid micelles and forms insolubilized products by binding with bile acids. This inhibits the passage of cholesterol through the unstirred aqueous layer in the intestinal tract, thereby providing an inhibitory effect on the biouptake of cholesterol.

[0027] Furthermore, the food material of the present invention has the effect of inhibiting the absorption of cholesterol in small intestinal epithelial cells after passing through the unstirred water layer in the intestinal tract (cholesterol absorption inhibitory effect in small intestinal epithelial cells). As a result, it has a good inhibitory effect on the biouptake of cholesterol, and because it is derived from adzuki beans, it is safe for the body to ingest.

[0028] Furthermore, it was confirmed that the food material of the present invention exhibits binding activity with bile acids and converts them into insoluble forms. Furthermore, when Caco-2 cells were treated with C7G or E7G, the amount of cholesterol taken up was significantly reduced compared to control cells, and no cytotoxicity was observed under the experimental conditions used.

[0029] The food material of the present invention contains C7G, E7G, and (+)-catechin, and is therefore believed to have the ability to inhibit micelle solubility, bind to bile acids, and suppress cholesterol absorption. The food material of the present invention is useful as a medicinal supplement for suppressing elevated cholesterol levels. In the dried food material of the present invention, the content of (+)-catechin 7-O-β-D-glucopyranoside in the total amount of (+)-catechin 7-O-β-D-glucopyranoside, (+)-epicatechin 7-O-β-D-glucopyranoside, and (+)-catechin is preferably 10 to 40% by weight, and particularly preferably 20 to 30% by weight; the content of (+)-epicatechin 7-O-β-D-glucopyranoside is preferably 0.1 to 5% by weight, and particularly preferably 0.5 to 3% by weight; and the content of (+)-catechin is preferably 0.5 to 10% by weight, and particularly preferably 1 to 5% by weight.

[0030] Next, a method for producing the adzuki bean-derived food material of the present invention having the effect of inhibiting bile acid micelle solubility and the effect of forming bile acid-bound insolubilized products will be described.

[0031] The method for producing a food material derived from adzuki beans of the present invention includes an adzuki bean soaking step in which adzuki beans are soaked in ethanol-containing water, a liquid collection step in which the adzuki beans are removed from the soaking water after the adzuki bean soaking step and a liquid is collected, and a drying step in which the liquid obtained in the liquid collection step is dried to obtain a dried product, thereby producing a food material derived from adzuki beans that contains (+)-catechin 7-O-β-D-glucopyranoside or (+)-catechin 7-O-β-D-glucopyranoside and (+)-epicatechin 7-O-β-D-glucopyranoside and has the ability to inhibit bile acid micelle solubility and form bile acid-bound insoluble products.

[0032] In a method for producing a food material derived from adzuki beans, adzuki beans (Vigna angularis) are prepared as the raw material. Dried, raw adzuki beans with the skin attached are used. These dried, raw adzuki beans are those that have been harvested, washed, and then naturally dried or air-dried to reduce the moisture content to 10 to 20% without softening them by hydration. Note that raw adzuki beans also include those whose surfaces have been sterilized by steam or roasting. It is preferable to use whole adzuki beans without removing the skin or crushing them (non-crushed).

[0033] In the method for producing a food material derived from adzuki beans of the present invention, an adzuki bean soaking step is carried out in which adzuki beans are soaked in ethanol-containing water. The ethanol-containing water preferably has an ethanol concentration of 30 to 50%. In particular, it is preferable to use ethanol-containing water with an ethanol concentration of 35 to 45%. Furthermore, in the azuki bean soaking step, it is preferable to soak the azuki beans in the ethanol-containing water for 12 hours or more. The soaking time in the azuki bean soaking step is preferably 12 to 48 hours. In particular, soaking for 20 hours or more is preferable. Furthermore, it is preferable that the amount of ethanol-containing water used to soak the azuki beans in the azuki bean soaking step is about 10 times the weight of the azuki beans.

[0034] Next, a liquid collection step is carried out in which the adzuki beans are removed from the soaking water (ethanol-containing water) and a liquid (ethanol-containing water) is collected.

[0035] The soaking liquid produced by soaking the adzuki beans in water (ethanol-containing water) is light reddish-brown in color. The soaking liquid and the hydrated adzuki beans are separated (drained) and the soaking liquid (liquid) is collected.

[0036] Subsequently, a drying step is carried out in which the liquid obtained in the liquid collection step is dried to obtain a dried product. The drying step may be vacuum freeze drying or heat drying, with vacuum freeze drying being preferred. Heat drying may include hot air drying, fluidized bed drying, spray drying, drum drying, etc. [Example]

[0037] (Manufacturing food ingredients derived from adzuki beans) The raw material used was adzuki beans (Vigna angularis) from Hokkaido (variety: Erimo Shouzu). Five grams of adzuki beans were lightly washed with water and then soaked in ethanol-containing water (ethanol concentration 40%) in an amount 10 times the amount of the adzuki beans, and allowed to stand for 24 hours. After soaking, the liquid (supernatant) was separated from the hydrated adzuki beans. The liquid (supernatant) was vacuum freeze-dried to obtain 441.1 mg (yield: 8.8%) of the adzuki bean-derived food material of the present invention (Example 1).

[0038] (HPLC analysis) (+)-Catechin 7-O-β-D-glucopyranoside (C7G), (+)-epicatechin 7-O-β-D-glucopyranoside (E7G), and (+)-catechin were prepared.

[0039] Example 1, C7G, and E7G were analyzed by HPLC at a flow rate of 3 ml / min using an ODS column (Nacalai Tesque, COSMOSIL 5C18-AR, 4.6 x 150 mm) and an intelligent HPLC pump (JASCO PU-1580 Intelligent HPLC Pump, JASCO Corporation) equipped with an ultraviolet detector (JASCO UV-1570, JASCO Corporation) operating at 210 nm. Purified C7G and E7G were also analyzed by HPLC after each separation. The ODS column was balanced by passing methanol through it prior to analysis.

[0040] In the HPLC analysis of Example 1, the mobile phase was ultrapure water and methanol (0-15 min: 100% ultrapure water, 15-45 min: 100% ultrapure water → 100% methanol), and the flow rate was 1 ml / min. (+)-Catechin (Nacalai Tesque) was used as the analytical standard. The total polyphenol content was measured using Folin-Ciocalteu reagent (Nacalai Tesque). 1 is an HPLC peak chart for Example 1. The vertical axis represents the detection potential (mV), and the horizontal axis represents the retention time (min).

[0041] (Cholesterol micellization) Two types of cholesterol micelles were prepared as follows. 2 mM cholesterol (Sigma-Aldrich, St. Louis, MO, USA), 1 mM oleic acid (Nacalai Tesque), and 2.4 mM phosphatidylcholine were dissolved in methanol. Two types of phosphatidylcholine were used: soybean lecithin (Nacalai Tesque) and synthetic lecithin (Fujifilm Wako Pure Chemical Industries, Ltd.).

[0042] After drying under nitrogen, 15 mM phosphate-buffered saline (PBS) containing 6.6 mM taurocholate (Nacalai Tesque) was added, and the solution was sonicated twice for 30 minutes. This solution was incubated overnight at 37°C and used in the experiment. Each compound was added to the micellar solution, mixed, and incubated at 37°C for 2 hours. The amount of cholesterol in the supernatant was measured using a cholesterol measurement kit (Fujifilm Wako Pure Chemical Corporation).

[0043] (bile acid conjugate adjustment) Bile acid conjugates were prepared as follows: 100 μl of sample solution (10 mg / ml) was added to a microtube containing 900 μl of 2 mM taurocholate (Nacalai Tesque, Inc.) in 0.1 M phosphate buffer, pH 7.0. After 2 hours of incubation at 37°C, the sample was centrifuged at 12,000 × g for 15 minutes, and the supernatant was transferred to a black microwell plate. The free bile acid concentration in the supernatant was measured using a Total Bile Acid Assay Kit (Cosmo Bio Co., Ltd.) according to the manufacturer's protocol. Fluorescence was measured using a CytoFluor Series 4000 Multi-Well Plate Reader (PerSeptive Biosystems, Framingham, MA, USA) at an excitation wavelength of 530 nm and an emission wavelength of 580 nm. Experimental values ​​were calculated from a standard curve. Cholestyramine resin (Sigma-Aldrich), a bile acid-binding drug, was used as a positive control.

[0044] (Cell culture and reagents) Caco-2 cells (RIKEN BRC Cell Bank, Tsukuba, Japan; passages 6–20) were cultured in Dulbecco's modified Eagle's medium (Nissui) (Nissui Pharmaceutical Co., Ltd.) supplemented with 10% fetal bovine serum (Hyclone, Logan, UT, USA) at 5% CO2 and 37°C. Cells were harvested using trypsin / EDTA (0.25%; Nacalai Tesque). Before use, cells were cultured until 80% confluent (adherent cells completely covered the adhesive surface of the culture vessel). The medium was changed every 3 days.

[0045] (Cell viability measurement) Caco-2 cells were cultured in a 96-well plate (1.0 × 10 4 The cells were plated on 1000 x g (1000 x g) plates (cells / well) and cultured for 7 days as described above. Subsequently, the cells were treated with Example 1, C7G, E7G, or (+)-catechin at final concentrations of 1 to 100 μg / ml for 24 hours. Then, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide solution (MTT; Nacalai Tesque) (5 mg / ml) was added to each well, and the plate was incubated at 37°C for 4 hours. Formazan crystals were dissolved in 100 μl of dimethyl sulfoxide. Absorbance at 600 nm was quantified using a CHROMATE 4300 Microplate Reader (Practical Co., Ltd.) and expressed as a percentage of the control.

[0046] (NBD-cholesterol uptake activity) Cholesterol uptake activity in Caco-2 cells was measured as follows. In this study, a fluorescent analogue of cholesterol, 22-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-23,24-bisnor-5-cholen-3β-ol (NBD-cholesterol; ThermoFisher Scientific, Waltham, MA, USA), was used.

[0047] Caco-2 cells were seeded in a 96-well plate (1.0 × 10 4Cells were cultured in a 1000-well plate (1000 x g / well) for 7 days. Cells were then incubated with 20 μg / ml NBD-cholesterol at 37°C for 20 minutes or 2 hours, with or without test compounds. After washing twice with PBS to remove unbound NBD-cholesterol, fluorescence was measured using a CytoFluor Series 4000 Multi-Well Plate Reader (PerSeptive Biosystems) at an excitation wavelength of 485 nm and an emission wavelength of 535 nm.

[0048] (RNA isolation and real-time PCR) NPC1L1 mRNA expression was measured by the following method. Caco-2 cells were seeded in a 12-well plate (1.0 × 10 5 The cells were cultured for 7 days. Total RNA was isolated using Trizol Reagent (Invitrogen, Carlsbad, CA, USA) and converted to cDNA using ReverTra Ace® qPCR RT Master Mix (Toyobo Co., Ltd.). Quantitative real-time PCR was performed on the cDNA using a LightCycler® Nano System (Roche Diagnostics, Mannheim, Germany). The thermal cycler program parameters were as follows: 95°C for 60 seconds, 95°C for 15 seconds, 64°C for 60 seconds (40 cycles) (NPC1L1), 95°C for 60 seconds (40 cycles) (NPC1L1). This was followed by 40 cycles of 95°C for 15 seconds and 60°C for 60 seconds (β-actin).

[0049] The following primers were used: NPC1L1, 5′-GTCCCTCATCAACCTGGTCTCG-3′ (forward), 5′-AAACACCGCACTTCCCATAGAG-3′ (reverse); β-actin, 5′-CCTGGCACCCAGCACAAT-3′ (forward), 5′-GCCGATCCACACGGAGTACT-3′ (reverse). Values ​​for each sample were normalized to β-actin and calculated using the 2 −ΔΔCT method.

[0050] (statistical analysis) Data were expressed as mean ± standard deviation (SD) for triplicate wells confirmed in three independent experiments, or mean ± standard error (SE) for three independent experiments. Statistical significance of differences in data was assessed by one-way analysis of variance (ANOVA) followed by Dunnett's test. A value of p < 0.05 was considered to indicate statistical significance.

[0051] (Preparation and HPLC analysis) As a result of 40% ethanol extraction, 441.1 mg of dried food material derived from adzuki beans (Example 1) was obtained from 5 g of adzuki beans (yield: 8.8%). HPLC analysis showed that C7G (Peak I, Figure 1) accounted for 24.5% of the total polyphenols in Example 1. E7G (Peak III, Figure 1) accounted for 1.1% of the total polyphenols in Example 1. (+)-Catechin (Peak II, Figure 1) in Example 1 accounted for 2.2% of the total polyphenols in Example 1. In subsequent experiments, the cholesterol-lowering activity of Example 1 and each polyphenol was investigated.

[0052] (Effect on micellization of cholesterol) The inhibitory effect of adzuki bean polyphenols on the formation of micelles in cholesterol was investigated. Considering the affinity of the lecithin derivatives, both natural lecithin derived from soybeans and synthetic lecithin were used. IC of micelles formed by soybean lecithin 50 value (50% inhibitory concentration (IC 50 [mg / ml] ± SE) were as follows:

[0053] Example 1: 6.15±0.38 mg / ml C7G: 0.23±0.03mg / ml, E7G: 0.22±0.02mg / ml (+)-Catechin: 0.26±0.11mg / ml

[0054] IC of micellization by synthetic lecithin 50 value (50% inhibitory concentration (IC 50 [mg / ml] ± SE) were as follows:

[0055] Example 1: 3.71±1.50 mg / ml C7G: 0.13±0.04mg / ml E7G: 0.08±0.01mg / ml (+)-Catechin: 0.03±0.01mg / ml

[0056] Regardless of the lecithin derivative used (soybean lecithin or synthetic lecithin), Example 1, C7G, E7G, and (+)-catechin showed a dose-dependent inhibitory effect on micellization.

[0057] (bile acid binding activity) The bile acid binding activity of C7G, E7G, and (+)-catechin was evaluated in Example 1. When each sample was compared at the same concentration, the taurocholic acid binding rates were as follows:

[0058] Example 1: 7.95±1.47% C7G: 11.18±2.65% E7G: 13.03±0.66% (+)-Catechin: 16.18±1.36% When cholestyramine, a positive bile acid secretion inhibitor, was administered at the same concentration, the rate was 39.25±3.40%.

[0059] (NBD-cholesterol uptake) Caco-2 cells were treated with Example 1, C7G, and E7G for a short period (20 minutes) or a long period (2 hours), and the amount of NBD-cholesterol taken up was measured to evaluate whether Example 1, C7G, and E7G exhibit cholesterol-lowering effects in Caco-2 cells.

[0060] During 20 minutes of treatment, Example 1 dose-dependently reduced NBD-cholesterol uptake, with a significant difference observed at 100 μg / ml compared to the control. Similarly, C7G, E7G, and (+)-catechin dose-dependently inhibited cholesterol uptake, with a significant difference observed at 100 μg / ml compared to the control. After 2 hours of treatment, Example 1 also dose-dependently inhibited NBD-cholesterol uptake, with a significant difference observed at 100 μg / ml compared to the control. Furthermore, C7G, E7G, and (+)-catechin dose-dependently inhibited cholesterol uptake, with a significant difference observed at 10 μg / ml and 100 μg / ml for C7G. Significant differences were observed for C7G at 10 μg / ml and 100 μg / ml, for E7G at 1 μg / ml, 10 μg / ml, and 100 μg / ml for (+)-catechin.

[0061] Thus, during 20 minutes or 2 hours of treatment, Example 1, C7G, and E7G significantly reduced NBD-cholesterol uptake in Caco-2 cells.

[0062] (Test on the inhibitory effect of adzuki bean extract on bile acid micelle dissolution) Two types of cholesterol micelles were prepared as follows. 2 mM cholesterol (Sigma-Aldrich, St. Louis, MO, USA), 1 mM oleic acid (Nacalai Tesque), and 2.4 mM phosphatidylcholine were dissolved in methanol. Two types of phosphatidylcholine were used: soybean lecithin (Nacalai Tesque) and synthetic lecithin (Fujifilm Wako Pure Chemical Industries, Ltd.).

[0063] After drying under nitrogen, 15 mM phosphate-buffered saline (PBS) containing 6.6 mM taurocholate (Nacalai Tesque) was added, and the solution was sonicated twice for 30 minutes. This solution was incubated overnight at 37°C and used in the experiment. Each compound was added to the micellar solution, mixed, and incubated at 37°C for 2 hours. The amount of cholesterol in the supernatant was measured using a cholesterol measurement kit (Fujifilm Wako Pure Chemical Corporation).

[0064] For Example 1, the inhibition rate of bile acid micelle dissolution was measured and the 50% inhibitory concentration (IC50) was calculated. The results are shown in Figure 2, and Example 1 exhibited a concentration-dependent inhibitory effect on bile acid micelle dissolution. C7G and E7G also exhibited a concentration-dependent inhibitory effect on bile acid micelle dissolution.

Claims

1. The food material is derived from adzuki beans, and is characterized by containing (+)-catechin 7-O-β-D-glucopyranoside, (+)-epicatechin 7-O-β-D-glucopyranoside, and (+)-catechin, and having an inhibitory effect on bile acid micelle solubility and an effect on forming bile acid-bound insolubilized products.

2. 2. The food material derived from adzuki beans according to claim 1, which has an inhibitory effect on the uptake of cholesterol in the body.

3. 3. The food material derived from adzuki beans according to claim 1, wherein the food material is derived from alcohol-containing soaking water for adzuki beans.

4. 3. The food material derived from adzuki beans according to claim 1, wherein the food material is a freeze-dried product.

5. A method for producing a food material derived from adzuki beans, comprising the steps of: an adzuki bean soaking step in which adzuki beans are soaked in ethanol-containing water; a liquid collection step in which, after the adzuki bean soaking step, the adzuki beans are removed from the soaking water and a liquid is collected; and a drying step in which the liquid collected in the liquid collection step is dried to obtain a dried product, thereby obtaining a food material containing (+)-catechin 7-O-β-D-glucopyranoside or (+)-catechin 7-O-β-D-glucopyranoside and (+)-epicatechin 7-O-β-D-glucopyranoside, and having the ability to inhibit bile acid micelle solubility and form bile acid-bound insolubilized products.

6. 6. The method for producing a food material derived from adzuki beans according to claim 5, wherein the food material derived from adzuki beans contains (+)-catechin.

7. 7. The method for producing a food material derived from adzuki beans according to claim 5 or 6, wherein the adzuki bean soaking step uses ethanol-containing water having an ethanol concentration of 30 to 50%.

8. 7. The method for producing a food material derived from adzuki beans according to claim 5 or 6, wherein the adzuki bean soaking step comprises soaking the adzuki beans in the ethanol-containing water for 12 hours or more.

9. 7. The method for producing a food material derived from adzuki beans according to claim 5, wherein the drying step is a heat drying step or a vacuum freeze drying step.

10. 7. The method for producing a food material derived from adzuki beans according to claim 5, wherein the soaking time in the adzuki bean soaking step is 12 to 48 hours.

11. 7. The method for producing a food material derived from adzuki beans according to claim 5, wherein the amount of ethanol-containing water used for soaking the adzuki beans in the adzuki bean soaking step is from the same amount to twice the weight of the adzuki beans.

12. The method for producing a food material derived from adzuki beans comprises the steps of: an adzuki bean soaking step in which dried adzuki beans with husks are soaked in ethanol-containing water having an ethanol concentration of 30 to 50% for 12 to 48 hours; a liquid collection step in which, after the adzuki bean soaking step, the adzuki beans are removed from the soaking water and a liquid is collected; and a drying step in which the liquid collected in the liquid collection step is dried to obtain a dried product, thereby obtaining a food material containing (+)-catechin 7-O-β-D-glucopyranoside or (+)-catechin 7-O-β-D-glucopyranoside, (+)-epicatechin 7-O-β-D-glucopyranoside, and (+)-catechin, and having the ability to inhibit bile acid micelle solubility and form bile acid-bound insolubilized products.

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