JP7213019B2 - Blood uric acid level reducing agent and xanthine oxidase inhibitor - Google Patents

Description

TECHNICAL FIELD The present invention relates to a blood uric acid level-lowering agent and a xanthine oxidase inhibitor containing a plant of the genus Brassica or an extract thereof as an active ingredient.

Uric acid is a metabolite of purines and is excreted through urine and feces after being produced in the body. In living organisms, the amount of purines in the body is kept constant by balancing the production and excretion of uric acid from purines. However, genetic and environmental factors can disrupt this balance and increase the uric acid level in the body. Patent document 1).

Regarding the relationship between blood uric acid levels and diseases, hyperuricemia with blood uric acid levels exceeding 7.0 mg/dL increases the risk of gouty arthritis, and blood uric acid levels and the onset of chronic kidney disease (CKD). It has been reported that the frequency of metabolic syndrome increases as the blood uric acid level increases (Non-Patent Document 1).

As therapeutic agents for hyperuricemia, those having actions such as inhibition of uric acid synthesis, promotion of uric acid excretion, and inhibition of purine absorption are known. Xanthine oxidase inhibitors are known as uric acid synthesis inhibitors.

Xanthine oxidase is an enzyme involved in the biosynthesis of uric acid in the body, and is responsible for the synthesis of uric acid from xanthine. Xanthine oxidase inhibitors such as allopurinol are used as uric acid level-lowering agents for the treatment of hyperuricemia. However, overdosing of allopurinol to patients with renal failure may increase the risk of side effects associated with oxypurinol accumulation (Non-Patent Document 1).

For the purpose of preventing or improving hyperuricemia or gout, a uric acid level-lowering agent that can be easily taken in daily life for a long period of time is preferable. For example, a uric acid level-lowering agent containing an active ingredient derived from a natural product is considered suitable. So far, chitosan (Patent Document 1), water-soluble dietary fiber (Patent Document 2), activated carbon (Patent Document 3), ε-polylysine (Patent Document 4), etc. have been reported as natural products having purine absorption inhibitory action. It is Grape extract (Patent Document 5), ellagic acid (Patent Document 6), epigallocatechin gallate (Patent Document 7), etc. have been reported as natural products having xanthine oxidase inhibitory activity. Furthermore, shikuwasa pericarp (Patent Document 8) has been reported as a natural product having a uric acid excretion promoting effect.

JP-A-2001-163788 Japanese Patent Application Laid-Open No. 2005-047828 Japanese Patent Application Laid-Open No. 2005-187405 Japanese Patent Application Laid-Open No. 2006-001898 Japanese Patent Application Laid-Open No. 2002-121145 Japanese Patent Application Laid-Open No. 2000-290188 JP-A-2002-370980 JP 2016-155825 A

Japanese Society for Gout and Nucleic Acid Metabolism Guideline Revision Committee Edited "Treatment Guidelines for Hyperuricemia and Gout" 2nd Edition (2012 supplement, digest version), published by Medical Review Co., Ltd.

As described above, in order to prevent and improve hyperuricemia and gout, there is a demand for a blood uric acid level-lowering agent that is excellent in safety and can be easily taken in daily life for a long period of time. Accordingly, an object of the present invention is to provide an agent containing a new active ingredient derived from a natural product and useful for preventing or improving hyperuricemia/gout.

The present inventors have studied in detail the physiological effects of plants belonging to the genus Brassica. As a result, the present inventors have found that kale, which is a kind of plant belonging to the genus Brassica, or an extract thereof has a blood uric acid level-lowering effect, and that kale or an extract thereof has a high xanthine oxidase inhibitory activity, thereby completing the present invention. .

One aspect of the present invention is a blood uric acid level-lowering agent containing a plant of the genus Brassica or an extract of a plant of the genus Brassica as an active ingredient.

Preferably, the Brassica plant is kale.

Preferably, the blood uric acid level-lowering agent is orally administered.

Preferably, the blood uric acid level-lowering agent is a food composition.

Another aspect of the present invention is a xanthine oxidase inhibitor containing a Brassica plant or an extract of a Brassica plant as an active ingredient.

Preferably, the Brassica plant is kale.

Preferably, said xanthine oxidase inhibitor is orally administered.

Preferably, said xanthine oxidase inhibitor is a food composition.

Preferably, the xanthine oxidase inhibitor is used to prevent or improve hyperuricemia or gout.

INDUSTRIAL APPLICABILITY According to the present invention, it is possible to provide a blood uric acid level-lowering agent and a xanthine oxidase inhibitor that contain active ingredients derived from natural products and are excellent in safety.

1 is a graph showing changes in serum uric acid levels in rats to which kale was administered. 13 is a graph showing serum uric acid levels in rats to which kale was administered 13 days after administration. 1 is a graph showing the results of examining the xanthine oxidase inhibitory activity of kale extracts.

The blood uric acid level-lowering agent and the xanthine oxidase inhibitor of the present invention (hereinafter sometimes collectively referred to as the agent of the present invention) contain a plant of the genus Brassica or an extract of a plant of the genus Brassica as an active ingredient.

The Brassica plant in the present invention refers to a plant belonging to the Brassicaceae Brassica genus. Specific examples of plants of the genus Brassica include kale, broccoli, broccoli sprouts, cabbage, Chinese cabbage, cauliflower, Brussels sprouts, Taisai, mizuna, sugukina, Japanese mustard spinach, mustard greens, and turnips. These plants of the genus Brassica may be used alone, or two or more of them may be used in combination. Among these, it is preferable to use at least kale as the plant belonging to the genus Brassica in the present invention.

The plant parts of the Brassica plant used in the present invention are not particularly limited, and examples thereof include leaves, stems, roots, seeds, and flower buds. These plant body parts may be used alone or in combination of two or more. In the present invention, leaves and stems are preferably used, and leaves are particularly preferably used.

The form of the Brassica plant contained in the agent of the present invention is not particularly limited. pulverized products can be mentioned. In addition, squeezed juice and squeezed lees obtained by squeezing the plant parts can be mentioned. A Brassica plant having one or more of these forms can serve as an active ingredient of the agent of the present invention.

The extract of the Brassica plant contained in the agent of the present invention is not particularly limited, and can be obtained using a known extraction method. Examples of the object to be extracted include the plant body part itself, as well as the dried and pulverized plant body parts described above. The solvent used for extraction may be an aqueous solvent or a non-aqueous solvent. Extraction methods include immersion, decoction, leaching, and the like. A purified product obtained by further purifying a primary extract extracted from a plant body part or the like may be used as an extract of a plant of the genus Brassica as an active ingredient. Purification operations include desalting, salting out, recrystallization, chromatography, membrane separation, and the like.

Oral administration is typical as the administration route of the agent of the present invention. Other examples of parenteral administration include transdermal administration. Other routes of administration include intravenous, subcutaneous, intramuscular, intraperitoneal, intracerebroventricular, and the like.

Regarding the dosage of the agent of the present invention, the target disease etc. (e.g., hyperuricemia, gout), administration purpose (e.g., prevention or improvement), type of active ingredient, administration route, age and weight of the subject of administration , etc., can be set as appropriate. For the purpose of preventing hyperuricemia and gout, for example, it is preferable to orally ingest the agent of the present invention on a daily and continuous basis. Many plants of the genus Brassica are widely used as food, and have a wide range of food experiences and are excellent in safety. Therefore, the agent of the present invention is suitable for long-term continuous ingestion.

As an example, in the case of an adult male (body weight 60 kg), this product is continuously taken so that 290 g to 830 g of kale leaves, preferably 350 g to 750 g, and more preferably 450 g to 650 g of kale leaves are ingested per day. All you have to do is ingest the agent of the invention. This is expected to prevent hyperuricemia and gout in healthy individuals. In addition, in the case of a person who has developed or has symptoms of hyperuricemia/gout, an increase in the blood uric acid level is suppressed, and an ameliorating effect of hyperuricemia/gout can be expected.
The human equivalent dose (HED) can be calculated from dose data obtained from animal experiments.

Regarding the content of the Brassica plant or Brassica plant extract in the agent of the present invention, the above-mentioned target diseases (e.g., hyperuricemia/gout), purpose of administration (e.g., prevention or improvement), efficacy It can be appropriately set according to the type of component, administration route, age and weight of the subject of administration, and the like. The agent of the present invention may consist of the dried or pulverized plant parts of Brassica plants.

The agent of the present invention can be, for example, a pharmaceutical composition, a food composition, or a cosmetic composition. In a preferred embodiment, the agent of the invention is a food composition.

Examples of food compositions include various processed foods, health foods, functional foods, health supplements, nutritional functional foods, foods for special dietary uses, foods with function claims, foods for specified health uses, and foods for the sick. and other health foods. Others include animal supplements.
The agent of the present invention can also be used as an additive to add functionality to other foods and drinks. That is, by adding the agent of the present invention to any food or drink, the blood uric acid level reducing effect and the xanthine oxidase inhibitory effect can be imparted. In this case, the addition target is not particularly limited, and all food and drink can be targeted.

The agent of the present invention may be in liquid, semi-solid or solid form. Dosage forms for oral administration in the case of pharmaceutical compositions include powders, granules, fine granules, tablets, capsules, syrups, drops, and the like. The same applies to food compositions such as supplements.

The agent of the present invention may further contain other ingredients within the range that does not impair its performance. For example, various carriers and bases that can be added to pharmaceuticals, foods, cosmetics and the like may be further included. Specifically, excipients, stabilizers, binders, disintegrants, solubilizers, tonicity agents, buffers, and the like may be further contained. In the case of food compositions, sugars (sucrose, fructose, lactose, sugar alcohols, etc.), amino acids, acids (citric acid, tartaric acid, etc.), vitamins (vitamin C, etc.), thickeners, pigments , fragrance, etc. may be further included.

EXAMPLES The present invention will be described in more detail with reference to examples below, but the present invention is not limited to these examples.

In this example, changes in blood uric acid levels in rats fed with kale were investigated.

(1) Test method Wister rats (6 weeks old) were divided into two groups, a control group of 5 rats and a kale-administered group of 5 rats. Both groups were preliminarily fed with a standard diet (AIN93G) for 1 week. (registered trademark), Nippon Flour Mills Co., Ltd.).
Table 1 shows the nutritional components of the feed administered to each group. That is, the nutritional components of each feed were almost the same.

Blood was collected 0, 1, 4, 7, 10 and 13 days after administration of each feed, and serum was collected. For deproteinization, 200 μL of 0.3 mol/L perchloric acid was added to 20 μL of serum. After standing on ice for 30 minutes, it was centrifuged at 10000 rpm for 10 minutes and the supernatant was recovered. The same amount of 0.2 mol/L disodium hydrogen phosphate aqueous solution was added to 200 μL of the supernatant, and subjected to HPLC using a C18 reverse phase column.

<HPLC conditions>
・Column: Inertsil ODS-3, particle size 4 μm, inner diameter 4.6 mm, length 250 mm (GL Sciences)
・ Eluent: A: 74 mM phosphate buffer (pH 2.2), B: methanol
A:B=98:2
・Column temperature: 40°C
・Detection wavelength: 284 nm

(2) Results Results are shown in FIGS. FIG. 1 is a graph showing changes in serum uric acid levels, where the vertical axis represents changes in serum uric acid levels (mg/dL), and the horizontal axis represents the number of days of breeding (days). ◯ indicates the kale-administered group, and ▪ indicates the control group. FIG. 2 is a graph showing serum uric acid levels 10 days after administration.
As shown in FIGS. 1 and 2, the kale-administered group (hyperuric acid-inducing diet + kale) showed significantly lower values after 1, 10, and 13 days than the control group (high-uric acid-inducing diet). (Mann-Whitney U test, days 1 and 13: p<0.05, day 10: p<0.01). From the above, it was clarified that the plant parts of kale have the effect of reducing the blood uric acid level.

In this example, the xanthine oxidase inhibitory activity of the kale extract was examined.

(1) Preparation of Kale Extract Kale leaves were freeze-dried to obtain freeze-dried kale powder. 20 mL of water was added to 2 g of freeze-dried kale powder heated at 100° C. for 30 minutes, and the mixture was shaken for 1 hour. After adding 20 mL of hexane and shaking for 1 hour, centrifugation was performed to separate the aqueous layer and the hexane layer. The aqueous layer was collected, ethanol was added to the final concentration of 90%, and ethanol precipitation was performed. After centrifugation, the supernatant was recovered, dried and dissolved in water. This solution was passed through a C18 solid-phase extraction column, and the flow-through liquid that came out without being adsorbed was dried to obtain an aqueous kale extract (kale extract). Samples with three concentrations of 10 mg/mL, 100 mg/mL, and 500 mg/mL were prepared as samples to be subjected to the following tests.

(2) Test method 720 µL of 0.1 M potassium phosphate buffer (pH 7.5), 352 µL of purified water, and 48 µL of sample were mixed. Further, 80 μL of 0.04 unit/mL bovine milk-derived xanthine oxidase dissolved in 0.1 M potassium phosphate buffer (pH 7.5) was added and incubated at 37° C. for 10 minutes. 1200 μL of 0.1 mM xanthine was added and incubated at 37° C. for 30 minutes for enzymatic reaction. 200 μL of 1N hydrochloric acid was added to stop the reaction. The reaction solution was subjected to HPLC under the same conditions as in Example 1, and the uric acid level was measured. The xanthine oxidase inhibitory activity was evaluated by the ratio of uric acid produced when the sample (kale water extract fraction) was added to the uric acid produced when water (control) was added as 100. At the same time, the same procedure was performed using allopurinol (a known xanthine oxidase inhibitor) in place of the aqueous kale extract, and it was confirmed that the reaction system functioned normally.

(3) Results The results are shown in FIG. The vertical axis in FIG. 3 represents xanthine oxidase relative activity (%). That is, xanthine oxidase inhibitory activity was observed in the water-extracted kale fraction, and concentration dependence was also observed. From the above, it was clarified that the plant parts of kale have xanthine oxidase inhibitory activity.

Claims (4)

A method for producing a xanthine oxidase inhibitor containing a kale extract as an active ingredient, comprising:
After heating the freeze-dried powder of kale leaves at 100° C. for 30 minutes, water was added and the mixture was shaken for 1 hour. ,
Ethanol is added to the collected aqueous layer so that the final concentration is 90%, ethanol precipitation is performed, and the supernatant is recovered,
The collected supernatant is passed through a C18 solid-phase extraction column to obtain a non-adsorbed flow-through liquid as a kale water extraction fraction,
using the kale water extract fraction as the kale extract;
A method for producing a xanthine oxidase inhibitor. The method for producing a xanthine oxidase inhibitor according to claim 1, wherein the xanthine oxidase inhibitor is orally administered. The method for producing a xanthine oxidase inhibitor according to claim 1 or 2, wherein the xanthine oxidase inhibitor is a food composition. The method for producing a xanthine oxidase inhibitor according to any one of claims 1 to 3, wherein the xanthine oxidase inhibitor is used for preventing or improving hyperuricemia or gout.

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