JP6901711B2 - α-Glucosidase inhibitor and its use - Google Patents

Description

The present invention relates to a composition having an α-glucosidase inhibitory activity, and a food or medicine containing the composition.

α-Glucosidase is a disaccharide-degrading enzyme (maltase, sucrase, isomaltase) present at the brush edge of small intestinal mucosal epithelial cells. By inhibiting its action, it suppresses the decomposition of disaccharides into monosaccharides and blood glucose. It can suppress the rise in the value and improve postprandial hyperglycemia. Since an α-glucosidase inhibitor lowers postprandial hyperglycemia, it is one of the means for the prevention and treatment of diabetes (see Non-Patent Document 1).

Currently, there are only a few types of α-glucosidase inhibitors widely used in clinical practice, such as acarbose (see Patent Document 1), voglibose (see Patent Document 2), and miglitol (see Non-Patent Document 2). It has been reported that α-glucosidase inhibitors have serious side effects such as hepatotoxicity, abdominal pain, flatulence, diarrhea, and hypoglycemia (see Non-Patent Document 3), and safety problems have been pointed out. Therefore, a safe and effective α-glucosidase inhibitor is strongly desired.

To date, many α-glucosidase inhibitors have been developed from natural plants. For example, Senkasou (see Patent Document 3), Amla (see Patent Document 4), Salacia reticulata (see Patent Document 5) and the like are used. However, there are only a few types of α-glucosidase inhibitors sold as foods for specified health use, such as Sosoureicha, Glucosedesign, and Touchi extract.

On the other hand, Yanbargoma ( Helicteres angustifolia L.) is a small shrub of the genus Sterculiaceae native to southwestern China and Southeast Asia, and is distributed in the mountains. There is no report that the extract of Yanbar sesame has an α-glucosidase activity inhibitory action.

Special Publication No. 54-39474 Japanese Unexamined Patent Publication No. 09-67271 Japanese Unexamined Patent Publication No. 2006-16388 Japanese Unexamined Patent Publication No. 2006-104094 Japanese Unexamined Patent Publication No. 2008-137925

Jo SH.et al., International Journal of Molecular Science. 12, p1359-1370, 2011 Ana T et al., Journal of Medicinal Chemistry, 55, p10345-10346, 2012 Charpentier G et al., Diabetic Medicine, 26, p73-85, 2000

An object of the present invention is to provide a safe and effective composition having a novel α-glucosidase inhibitory activity, and a food or pharmaceutical product containing the composition.

As a result of diligent studies to solve the above problems, the present inventors have found that the extract of Yanbar sesame has an α-glucosidase inhibitory activity and an effect of suppressing an increase in blood glucose level. It came to be completed.

That is, the present invention is as follows.
(1) An α-glucosidase inhibitor characterized by containing Yanbargoma (Helicteres angustifolia L.) as an active ingredient.
(2) The α-glucosidase inhibitor according to (1), wherein the active ingredient is contained as an extract of yanbal sesame.
(3) The α-glucosidase inhibitor according to (2), wherein the extract is an extract of water or an organic solvent.
(4) The α-glucosidase inhibitor according to (3), wherein the extract is a 30 to 90% ethanol extract.
(5) A drug comprising the α-glucosidase inhibitor according to any one of (1) to (4).
(6) The medicament according to (5), which is in the form of capsules, tablets, granules or powder.
(7) A food containing the α-glucosidase inhibitor according to any one of (1) to (4).
(8) The food according to (7), which is in the form of a beverage.

Since the α-glucosidase inhibitor of the present invention is derived from a natural product, it has low toxicity and few side effects. In addition, the α-glucosidase inhibitor of the present invention is effective in preventing or ameliorating diabetes because it exhibits an inhibitory effect on blood glucose elevation due to its inhibition of the action of disaccharide-degrading enzyme.

Hereinafter, the present invention will be described in detail.
(Α-Glucosidase inhibitor of the present invention)
The α-glucosidase inhibitor of the present invention is characterized by containing Yanbar sesame as an active ingredient.
Helicteres angustifolia L. is a small shrub of the genus Sterculiaceae native to southwestern China and Southeast Asia. The site of yanbalgoma contained as an active ingredient in the α-glucosidase inhibitor of the present invention is not particularly limited, and examples thereof include roots, leaves, stems, flowers and fruits, and it is preferable to use yanbalgoma root as an active ingredient. ..
From the viewpoint of formulation and efficacy, the α-glucosidase inhibitor of the present invention preferably contains an extract of Yanbargoma as an active ingredient. The extract of Yanbar sesame is not particularly limited, and examples thereof include extracts of roots, leaves, stems, flowers and fruits, and it is preferable to use an extract of Yanbar sesame root as an active ingredient.
The α-glucosidase inhibitor of the present invention has a small intestine-derived maltase and / or sucrase inhibition rate of preferably 10% or more, 15% or more, and 20% or more.

(Method for producing α-glucosidase inhibitor of the present invention)
When the root, leaf, stem, flower and fruit are used, the α-glucosidase inhibitor of the present invention is obtained by drying these parts and then cutting or powdering the root, leaves, stems, flowers and fruits, which is used as it is or, if necessary, pharmacology. A pharmaceutically acceptable carrier can be added to obtain the α-glucosidase inhibitor of the present invention. Pharmacologically acceptable carriers include, for example, excipients, lubricants, binders and disintegrants in solid formulations.

When the α-glucosidase inhibitor of the present invention contains an extract of yanbal sesame as an active ingredient, the roots, leaves, stems, flowers and fruits of yanbal sesame can be mixed with water, ethanol, methanol, ethyl acetate, acetone, acetonitrile, butanol and the like. It can be obtained by extracting with the aqueous solvent of No. 1 alone or in combination thereof. The active ingredient of Yanbar sesame is efficiently extracted when water or hydrous ethanol is used as an extraction solvent. Further, when hydrous ethanol is used as the extraction solvent, it is preferable to use 30 to 90% ethanol. By distilling off the solvent of the obtained ethanol extract under reduced pressure, a concentrated extract of yanbal sesame can be obtained, and it is preferable to use this as an extract of yanbal sesame.
By fractionating the obtained ethanol extract with ethyl acetate and n-butanol, a fraction having a high α-glucosidase inhibitory activity can be obtained. In addition, the obtained ethanol extract can be fractionated with a fraction having an α-glucosidase inhibitory activity by using resin beads and / or by chromatography. In the present invention, a fraction having a high α-glucosidase inhibitory activity by fractionating with Diaion (registered trademark) HP20 resin (Mitsubishi Chemical) and / or using column chromatography using HP20 resin. It is preferable to use HP20 resin for fractionation.
The α-glucosidase inhibitor of the present invention can be obtained from the above extract or its concentrate as it is, or by adding a pharmacologically acceptable carrier as needed to obtain the α-glucosidase inhibitor of the present invention. Can be done. Pharmacologically acceptable carriers include, for example, excipients, lubricants, binders and disintegrants in solid formulations, solvents in liquid formulations, solubilizers, buffers and the like.

(Pharmaceutical of the present invention)
The medicament of the present invention is characterized by containing the above-mentioned α-glucosidase inhibitor.
In the formulation of the pharmaceutical product of the present invention, various components usually used in the formulation are optionally used, and examples thereof include starch, dextrin, lactose, cornstarch, and inorganic salts.
Since the α-glucosidase inhibitor of the present invention has extremely low toxicity and no serious side effects have been reported, it can be taken directly. Therefore, the α-glucosidase inhibitor can be ingested as granules, tablets, capsules, powders and the like.
The method for administering the drug of the present invention is not particularly limited, but oral administration is preferable. As a preferred dose, when the Yanbar sesame plant is administered, for example, to a patient (assuming a body weight of 60 kg), the dry weight is 1 g to 500 g, preferably 5 g to 300 g per day. On the other hand, when the extract of Yanbar sesame is administered, the concentration of the extract in the medicine of the present invention is preferably, for example, 10 to 500 μg / ml by dry weight and 20 to 150 μg / ml.
When using the Yanbargoma plant itself instead of the extract, in consideration of the dry weight of the extract in the plant body, administer it in an amount or concentration 9 to 11 times that of the extract, and in the Yanbargoma plant, administer it in an amount or concentration of 9 to 11 times. The reverse is true when the extract is administered without.
The medicament of the present invention is effective in preventing or ameliorating diabetes because it exhibits an inhibitory effect on blood glucose elevation in addition to being an α-glucosidase inhibitor.

(Food of the present invention)
The food product of the present invention is characterized by containing the above-mentioned α-glucosidase inhibitor.
Since the α-glucosidase inhibitor of the present invention has extremely low toxicity and no serious side effects have been reported, it can be taken directly.
The α-glucosidase inhibitor of the present invention is a functional food, a health food, a dietary supplement having an effect of suppressing an increase in blood glucose by blending with a raw material generally used as a food, for example, protein, lipid, carbohydrate, vitamins and the like. , Can also be used as a dietary supplement and a nutritional composition. The food product of the present invention can be produced by blending the above-mentioned α-glucosidase inhibitor with a food material. Preferred examples of the food product of the present invention include dairy products, seasonings, confectionery, noodles, soups, beverages and the like, and beverages such as tea bags and tea are particularly preferable.
The amount of the α-glucosidase inhibitor contained in the food of the present invention is not particularly limited and may be appropriately selected. However, when a Yanbar sesame plant is used, for example, 1 g to 500 g, preferably 5 g to 300 g, per 1 kg of the food. When the Yanbar sesame extract is used, for example, it is preferably 10 to 500 μg / ml, preferably 20 to 150 μg / ml in the beverage.
When using the Yanbargoma plant itself instead of the extract, in consideration of the dry weight of the extract in the plant body, administer it in an amount or concentration 9 to 11 times that of the extract, and in the Yanbargoma plant, administer it in an amount or concentration of 9 to 11 times. The reverse is true when the extract is administered without.
Further, the food of the present invention may be a health food having a preventive or therapeutic effect on the above-mentioned diseases, a food for specified health use, a nutritionally functional food, or the like. The food of the present invention can also be sold with a label such as "a food having an effect of suppressing an increase in blood glucose" or "a food containing a component having an effect of suppressing an increase in blood glucose". In addition, it may have a label such as "food having a diabetic therapeutic effect" or "food containing an ingredient having a diabetic therapeutic effect".

Next, the present invention will be described in more detail with reference to Examples, but the present invention is not limited to these Examples.
In addition, the results in the table for each example are shown by the average value or average value ± standard error of each group. For comparison between each group, a one-way analysis of variance (ANOVA) was performed and then a multiple comparison test was performed by Duncan's multiple range test (DMRT). (* P <0.05, ** p <0.01, *** p <0.001).

Example 1: α-Glucosidase inhibitory action of Yanbargoma extract Example 1-1: Preparation of Yanbargoma root extract and fractions Dry Yanbargoma root (obtained from Laos) is crushed and 500 mL of distilled water is added to 50 g. In addition, 100
Hot water extraction was performed at ° C for 2 hours. After extraction, solid-liquid separation was performed, the extract was concentrated under reduced pressure, and further freeze-dried to obtain a dried product (Extract 1).
Next, dried roots of Yanbar sesame were crushed, 70% ethanol was added, and extraction was performed at room temperature for 3 days. The extracts were combined and the solvent was distilled off at 40 ° C. under reduced pressure to obtain a 70% ethanol extract (extract 2).
Next, Extract 2 was sequentially extracted using petroleum ether, ethyl acetate and butanol, and freeze-dried to obtain Fraction 1, Fraction 2 and Fraction 3. Finally, the residual aqueous layer was concentrated under reduced pressure, and further freeze-dried to obtain a dried product (fraction 4). Fraction 4 was subjected to Diaion HP 20 column chromatography (Mitsubishi Chemical) and fractionated with 10% methanol, 50% methanol and 100% methanol to fraction 5, fraction 6, fraction 6. I got 7 minutes.
Finally, after passing the extract 2 through the synthetic adsorption resin Diaion HP-20 (Mitsubishi Chemical), it was first washed with water to obtain sugars and the like (fraction 8). Then elute with 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99.5% ethanol in sequence, concentrate and dry the eluate, and fraction from fraction 9. Fractions up to 18 minutes were obtained.

試料：試料、マルトース、マルターゼ酵素液
試料盲検：試料、マルトース、リン酸緩衝液
対照：リン酸緩衝液、マルトース、マルターゼ酵素液
対照盲検：リン酸緩衝液、マルトース、リン酸緩衝液

本実験では各試料溶液１種類につき3検体（n＝3）で酵素反応を行った。ヤンバルゴマ
の根の各抽出物の小腸由来マルターゼ阻害活性(1 mg/mL)を表１に記載した。

表１から明らかなように、HP20樹脂により精製した画分 9は1 mg/mLの濃度で46.3％の
マルターゼ阻害率を示し、糖尿病に対する予防又は改善作用を有することが明らかになった。なお、画分７が阻害率０％であったが、これは1 mg/mLの濃度では阻害活性が弱いこ
とが原因と考えられた。 Example 1-2: Measurement of maltase inhibitory effect Preparation of maltase enzyme solution derived from rat small intestine: rat intestinal acetone powder (manufactured by SIGMA) 1 g
Was suspended in 10 mL of 0.1 M phosphate buffer (pH 7.0) and homogenized while cooling with ice. After centrifugation (3,000 rpm, 10 minutes, 4 ° C.), the supernatant was used as a crude enzyme solution (100 mg / mL). The crude enzyme solution diluted 20-fold with 0.1 M phosphate buffer (pH 7.0) was used as the maltase enzyme solution.
0.02 mL of the extract and fraction prepared in Example 1-1 at different concentrations were added to a 96-well plate, 0.02 mL maltose (2%) was added as a substrate, and then preheated at 37 ° C. for 5 minutes. Then, 0.02 mL of maltase enzyme solution derived from rat small intestine was added and the enzyme reaction was carried out accurately for 60 minutes at 37 ° C. Then, 0.01 mL of the reaction solution was placed in another 96-well plate, and the color-developing reagent solution (glucose CII-test, Wako Pharmaceutical) 0.2 mL was added, mixed well, and then heated at 37 ° C for 5 minutes. Then, the absorbance of the sample at 492 nm was measured with a spectrophotometer (Biotrak II plate reader, Biochrom, Cambridge, UK) using reagent blinding as a control, and the maltase inhibitory activity of the sample was calculated.
The inhibitory activity of each sample solution was calculated using the following formula and expressed as the inhibition rate (%).

Sample: Sample, Maltose, Maltase Enzyme Solution Sample Blind: Sample, Maltose, Phosphate Buffer Control: Phosphate Buffer, Maltase, Maltase Enzyme Solution Control Blind: Phosphate Buffer, Maltose, Phosphate Buffer

In this experiment, the enzyme reaction was carried out with 3 samples (n = 3) for each sample solution. The small intestine-derived maltase inhibitory activity (1 mg / mL) of each extract of Yanbar sesame root is shown in Table 1.

As is clear from Table 1, the fraction 9 purified with HP20 resin showed a maltase inhibition rate of 46.3% at a concentration of 1 mg / mL, and it was clarified that it has a preventive or ameliorating effect on diabetes. The inhibition rate of fraction 7 was 0%, which was considered to be due to the weak inhibitory activity at a concentration of 1 mg / mL.

表２から明らかなように、HP20樹脂による精製した画分 9は1 mg/mLの濃度で51.4％の
スクラーゼ阻害率を示し、糖尿病に対する予防又は改善作用を有することが明らかになった。なお、画分４，７，８が阻害率０％であったが、これは1 mg/mLの濃度では阻害活性
が弱いことが原因と考えられた。 Example 1-3: Measurement of sucrase inhibitory action The crude enzyme solution prepared in Example 1-1 diluted 2-fold with 0.1 M phosphate buffer (pH 7.0) was used as the sucrase enzyme solution. The measurement of the sucrase inhibitory effect was carried out in the same manner as in Example 1-2 by changing maltose to sucrose. The small intestine-derived sucrase inhibitory activity (1 mg / mL) of each extract of Yanbar sesame root is shown in Table 2.

As is clear from Table 2, the fraction 9 purified with HP20 resin showed a sucrase inhibition rate of 51.4% at a concentration of 1 mg / mL, and it was clarified that it has a preventive or ameliorating effect on diabetes. The inhibition rate of fractions 4, 7 and 8 was 0%, which was considered to be due to the weak inhibitory activity at a concentration of 1 mg / mL.

表３から明らかなように、ヤンバルゴマ投与群は、対照群と比較して、マルトース投与後30、60分の血糖値を有意に抑制し、糖尿病の予防又は改善作用を有することが明らかになった。 Example 2: Blood glucose elevation inhibitory effect of Yanbargoma (root) methanol extract Example 2-1: Maltose stress test in normal rats
13-week-old SD rats (Nippon SLC) are at room temperature 23 ± 1 ° C, humidity 55 ± 8%, 12-hour light-dark cycle (7: 00-19: 00 light period) under illumination, at the Animal Resource Center, University of Tsukuba. They were bred and free intake of solid feed (MF. Oriental Yeast Industry) and water (tap water). After pre-rearing the rats for 1 week, the control group (6 animals) was orally administered with distilled water, and the administration group (6 animals) was orally administered with the extract 2 (300 mg / kg) prepared in Example 1-1. .. Thirty minutes after administration, maltose (2 g / kg) was orally administered to each group by gastric sonde (10 mL / kg body weight). Immediately before administration of maltose (0 minutes) and 30, 60, 120 minutes after administration of maltose, blood is collected from the tail vein and the blood glucose level is measured using a simple blood glucose meter (Glutest Neoa Alpha, Sanwa Kagaku Kenkyusho, Nagoya). Was measured.
Table 3 shows the changes in blood glucose level 120 minutes after administration of maltose to each group.

As is clear from Table 3, it was clarified that the Yanbargoma-administered group significantly suppressed the blood glucose level 30 to 60 minutes after the maltose administration and had a preventive or ameliorating effect on diabetes as compared with the control group. ..

表４から明らかなように、抽出物２はスクロース摂取30分後の血糖上昇を有意に抑制し、糖尿病の予防又は改善作用を有することが明らかになった。
以上により、ヤンバルゴマの抽出物は血糖値の上昇を抑制できる効果を有する。 Example 2-2: Sucrose load test in normal rats The sucrose load test in normal rats was carried out in the same manner as in Example 2-1 with the substrate changed to sucrose. Table 4 shows the transition of blood glucose level for 120 minutes after administration of sucrose to each group.

As is clear from Table 4, Extract 2 significantly suppresses the increase in blood glucose 30 minutes after ingestion of sucrose, and has a preventive or ameliorating effect on diabetes.
From the above, the extract of Yanbar sesame has an effect of suppressing an increase in blood glucose level.

The α-glucosidase inhibitor of the present invention can be used in the fields of foods, health foods, foods for specified health uses, nutritionally functional foods, pharmaceuticals and the like.

Claims (3)

An α-glucosidase inhibitor characterized by containing a water extract of Yanbargoma ( Helicteres angustifolia L.) as an active ingredient. A food for inhibiting α-glucosidase, which comprises the α-glucosidase inhibitor according to claim 1. The food product according to claim 2 , which is in the form of a beverage.