A composition for improving dementia containing buckwheat starch as an active ingredient, and a beverage, food, or pharmaceutical for improving dementia containing the composition.

Abstract

To provide a composition having a dementia remedying action, derived from a safe food.SOLUTION: The inventors have confirmed that buckwheat starch extracted from buckwheat flour has a dementia remedying action, providing a composition containing buckwheat starch, and a beverage, a food, and a medicine each including the composition.SELECTED DRAWING: None

Description

【Technical Field】 【0001】 The present invention relates to a composition for improving dementia containing buckwheat starch as an active ingredient, and a beverage, food or medicine for improving dementia containing the composition. 【Background Art】 【0002】 The number of dementia patients is estimated to be approximately 55 million worldwide and is still increasing. In Japan, the number of dementia patients is also estimated to be approximately 6.3 million, accounting for 18% of the elderly in our country. Therefore, countermeasures against dementia are a global issue and an urgent issue. Thus, the development of medicines for preventing and treating dementia is underway, and food components having an effect of preventing dementia and an effect of maintaining memory have been identified, and some foods containing these components have been registered as foods with functional claims. So far, GABA, EPA, DHA, β-lactolin, curcumin, etc. are known as food components having an effect of preventing dementia and an effect of maintaining memory. 【0003】 Buckwheat is one of the common foods commonly used in our country and is rich in essential amino acids such as lysine, vitamins B1, B2, minerals, rutin, etc., and various effects such as strengthening capillaries, recovering fatigue, and lowering blood pressure and blood sugar have been reported. 【0004】 Patent Document 1 (Japanese Patent Publication No. 4309496) describes a brain function improving composition containing components of buckwheat seeds as an active ingredient. However, Patent Document 1 does not specify the active ingredient in buckwheat seeds, and only refers to it as "components of buckwheat seeds," "buckwheat seeds, buckwheat seed extract or fraction thereof," or "fraction of buckwheat seed extract having a molecular weight of 10,000 or less." Furthermore, Patent Document 1 states that "in order to efficiently separate polyphenol compounds and other useful components contained in buckwheat seeds and utilize them as a composition for solving the problems of the present invention" (paragraph 0008), and states that it may contain "an extract obtained by extracting buckwheat seeds with water, alcohol or other solvents, or a specific fraction obtained by further fractionating and purifying this extract by appropriate means" (paragraph 0010). Here, when buckwheat seeds are extracted with water or alcohol, water-soluble proteins and polyphenols dissolve in the solvent and are extracted, but buckwheat starch does not dissolve in the solvent and remains as a residue. Therefore, although Patent Document 1 does not specify the active ingredients derived from buckwheat seeds, judging from its description, buckwheat starch is not included among the active ingredients derived from buckwheat seeds. 【0005】 Patent Document 2 (Japanese Patent Publication No. 7-236440) describes a "solid composition derived from buckwheat that moves from the buckwheat flour to the liquid layer and separates in a mixture of buckwheat flour and water" as having the effect of lowering blood cholesterol. In this document as well, the active ingredients contained in the solid composition derived from buckwheat are not identified, but since the solid composition derived from buckwheat has the property of moving to the liquid layer when mixed with water, it is clear that such a composition does not contain buckwheat starch. Furthermore, Patent Document 2 does not describe or suggest at all any effect of improving dementia as a characteristic of the solid composition derived from buckwheat. 【0006】 Patent Document 3 (Japanese Patent Publication No. 3977889) describes the cholesterol-lowering, triglyceride-lowering, or hyperlipidemia-improving effects of components obtained by extracting buckwheat hulls. However, since buckwheat hulls do not contain buckwheat starch, it is clear that Patent Document 3 does not suggest buckwheat starch. Furthermore, Patent Document 3 does not describe or suggest at all any dementia-improving effect as a characteristic of components obtained by extracting buckwheat hulls. 【0007】 Regarding buckwheat starch as an ingredient in buckwheat, Patent Document 4 (Japanese Patent Publication No. 11-75747) describes a flour for Japanese soba noodles consisting of buckwheat flour and processed buckwheat starch, Patent Document 5 (Japanese Patent Publication No. 60-133835) describes an edible water-in-oil emulsion spread using buckwheat starch, and Patent Document 6 (Japanese Patent Publication No. 10-174577) describes beer or sparkling alcoholic beverage manufactured using buckwheat-derived starch as a raw material. However, none of these patent documents describe or suggest the pharmacological effects of buckwheat starch. As described above, various studies have been conducted on the pharmacological effects of buckwheat and buckwheat starch, but it was not known that buckwheat starch has an effect on improving dementia. [Prior art documents] [Patent Documents] 【0008】 [Patent Document 1] Japanese Patent Publication No. 4309496 (Composition for the Prevention and / or Treatment of Aging) [Patent Document 2] Japanese Patent Publication No. 7-236440 (Composition derived from buckwheat flour, method of producing the same, and uses thereof) [Patent Document 3] Japanese Patent Publication No. 3977889 (A drug containing buckwheat hull extract as an active ingredient) [Patent Document 4] Japanese Patent Publication No. 11-75747 (Flour for Japanese buckwheat noodles and Japanese buckwheat noodles) [Patent Document 5] Japanese Patent Publication No. 60-133835 (Water-in-Edible Oil Emulsion Spread) [Patent Document 6] Japanese Patent Publication No. 10-174577 (Novel Beer-type Sparkling Wine and Method for Producing the Same) [Overview of the project] [Problems that the invention aims to solve] 【0009】 The present invention aims to provide a composition for improving dementia containing buckwheat starch as an active ingredient, and a beverage, food, or pharmaceutical product for improving dementia containing the composition. [Means for solving the problem] 【0010】 The inventors diligently researched the components and effects of buckwheat and discovered that buckwheat starch, as a component of buckwheat, has an effect on improving dementia. Specifically, they found that dementia can be improved by a dementia-improving composition containing buckwheat starch as an active ingredient, thus completing the present invention. Furthermore, they found that the composition of the present invention can be applied to beverages, foods, or pharmaceuticals for improving dementia, thus completing the present invention. The present invention is defined by the following specific features. 【0011】 [1] A composition for improving dementia containing buckwheat starch as an active ingredient. [2] A beverage for improving dementia containing the dementia-improving composition described in [1]. [3] A food for improving dementia containing the dementia-improving composition described in [1]. [4] A pharmaceutical product for improving dementia containing the dementia-improving composition described in [1]. [Effects of the Invention] 【0012】 The buckwheat starch of the present invention is obtained from buckwheat, a common food commonly consumed in Japan, and is highly safe and possesses properties that improve dementia, making it suitable for use as a dementia-improving composition. Furthermore, by utilizing the dementia-improving properties of the buckwheat starch of the present invention, it is possible to provide a dementia-improving beverage, food, or pharmaceutical product. [Brief explanation of the drawing] 【0013】 [Figure 1] Shows the results of the Barnes maze test. [Figure 2] Shows the results of the passive avoidance test. [Figure 3] Shows the results of gene expression analysis of BDNF by real-time PCR method. [Figure 4] Shows the results of expression analysis of BDNF by Western blotting method. 【Mode for Carrying Out the Invention】 【0014】 The first aspect of the present invention is a composition for improving dementia containing buckwheat starch as an active ingredient. Hereinafter, specific forms for implementing the aspect will be described. 【0015】 Regarding the buckwheat flour used as a raw material for buckwheat starch in the present invention, there are no particular limitations on the raw material variety, raw material origin, quality, milling method, and particle size composition. For example, as the raw material variety, Shinano No. 1, Kitawase buckwheat, etc. can be used. Examples of the raw material origin include Nagano Prefecture, Hokkaido, the United States, Russia, China, etc., and those from any origin can be used. Regarding the quality, for example, when using inner-layer flour with a low protein content and a high starch content, buckwheat starch can be extracted more efficiently, but it is not necessarily limited to inner-layer flour. Examples of the milling method include stone mortar milling, roll milling, etc., but it is not limited thereto, and the particle size composition is also not particularly limited. In addition to buckwheat flour, even those in which buckwheat seeds are slightly crushed can be used as raw materials for buckwheat starch extraction without problems. 【0016】 Buckwheat starch is extracted and separated from buckwheat flour. The method for extracting buckwheat starch is not particularly limited, and for example, the following methods can be applied. Add a 0.2% sodium hydroxide solution to buckwheat flour at about 10 times the amount of the buckwheat flour, stir well, suspend it, and then let it stand at 4°C for 12 hours. Then, remove the supernatant. By repeating this operation several times, the protein is removed from the buckwheat flour. Then, by repeating the same operation several times using distilled water instead of the sodium hydroxide solution, buckwheat starch can be obtained from the buckwheat flour. The operation using the sodium hydroxide solution is preferably repeated 3 to 7 times, more preferably 5 to 7 times, and even more preferably 6 to 7 times, and the operation using distilled water is also preferably repeated the same number of times. 【0017】 The content of buckwheat starch as an active ingredient in the composition for improving dementia of the present invention is not particularly limited as long as it is within the range that exhibits the effect of improving dementia. 【0018】 Another aspect of the present invention is a beverage, food, and medicine for improving dementia containing a composition for improving dementia containing buckwheat starch as an active ingredient. Hereinafter, specific forms for implementing this aspect will be described. 【0019】 The beverage for improving dementia is not particularly limited as long as the composition for improving dementia of the present invention can be uniformly dissolved, and tea such as green tea, black tea, oolong tea, and barley tea, coffee, cocoa, fruit juice beverages, carbonated beverages, sports beverages, and other soft drinks can be used. Regarding the content of the composition for improving dementia in the beverage for improving dementia, there is no particular limitation as long as it is within the range that exhibits the effect of improving dementia. 【0020】 The food for improving dementia is not particularly limited as long as it can contain the composition for improving dementia of the present invention, and any of staple foods, side dishes, and confectioneries excluding buckwheat can be used. Regarding the content of the composition for improving dementia in the food for improving dementia, there is no particular limitation as long as it is within the range that exhibits the effect of improving dementia. 【0021】 There are no particular restrictions on the dosage form of a drug used to improve dementia; it may be an oral preparation such as a tablet, capsule, granule, powder, or syrup, or an injectable preparation. There are also no particular restrictions on the amount of cognitive-improving composition contained in a drug used to improve dementia, as long as it is within a range that produces the desired effect on dementia, and it can be used in combination with other drugs. 【0022】 The dementia-improving effect of the dementia-improving composition of the present invention can be evaluated and confirmed by behavioral tests (Burnes maze test, passive avoidance test) and physiological analysis (BDNF expression analysis: real-time PCR method, Western blotting method) using mice. 【0023】 The Burns maze test, a behavioral test using mice, involves having mice learn a maze space, then placing them back in the maze after a certain period of time. The test evaluates the mice's spatial learning and memory based on their behavior. A shorter time to escape the maze indicates higher learning and memory ability. On the other hand, the passive avoidance test uses the avoidance behavior that mice exhibit in response to previously experienced aversive stimuli as an indicator of memory, and evaluates the degree of learning and memory in mice. A longer duration of avoidance behavior indicates higher learning and memory ability. 【0024】 BDNF expression analysis as a physiological analysis involves evaluating the expression status of BDNF in the mouse brain by performing gene expression analysis of BDNF using real-time PCR and BDNF expression analysis using Western blotting. BDNF stands for brain-derived neurotrophic factor, and it is an essential substance for increasing brain cells, which regulates the growth of nerve cells, such as nerve cell survival, growth, and enhanced synaptic function. Increased BDNF expression is valued as an indicator of improved brain function and improvement of dementia. 【0025】 The following describes specific embodiments of the present invention, including the best mode for carrying it out, with reference to examples and comparative examples. However, the present invention is not limited in any way by these examples. Unless otherwise specified, % refers to weight percent. [Examples] 【0026】 <Extraction of buckwheat starch> Buckwheat (from Nagano Prefecture) was milled using a roller mill (manufactured by Bühler), and the resulting mixture was sieved through a 100 μm mesh sieve. The portion that did not pass through the sieve was milled again and sieved. This process was repeated three times. From the resulting fractions, only those with a protein content of 5% or less were mixed to obtain buckwheat flour for buckwheat starch extraction (buckwheat flour A). The analytical values ​​of buckwheat flour A are shown in Table 1. 【0027】 [Table 1] 【0028】 To 100g of buckwheat flour A, 1,000ml of 0.2% sodium hydroxide solution was added, the mixture was thoroughly stirred, and the suspension was allowed to stand at 4°C for 12 hours. After standing, the supernatant was removed. This procedure was repeated five times to obtain buckwheat starch. To lower the pH of the buckwheat starch, it was washed five times with 1,000ml of distilled water, dehydrated using a centrifuge (6,000rpm, 10 minutes), and the pH was adjusted to 7.0. Wheat starch and rice starch were procured commercially and used for comparison. The compositions of the buckwheat starch, wheat starch, and rice starch used in this experiment are shown in Table 2. 【0029】 [Table 2] 【0030】 Sixteen-week-old aging-accelerated mice (SAMP8, memory impairment, short lifespan) and sixteen-week-old normally aging mice (SAMR1) were divided into five test groups as shown in Table 3. They were fed ad libitum until 28 weeks of age under conditions of room temperature 22±2℃, humidity 50±10%, and a 12-hour light-dark cycle (light period 7:00-19:00). Details of the feed used are shown in Table 4. The proportions of buckwheat starch, wheat starch, and rice starch were each 4%, and the rest of the feed consisted of a general feed composition mainly of corn starch and casein. For the control group, the casein content was increased to 100%. 【0031】 [Table 3] 【0032】 [Table 4] 【0033】 <Barnes Maze Test> The Barnes maze test was conducted using the MBM-020 (Muromachi) Barnes maze for mice. The Barnes maze has 20 holes on the circumference of a disc. A fixed avoidance box was assigned to each mouse within these holes. Three types of external stimuli were placed on a screen as memory cues. The training period consisted of two trials per day, spaced 30 minutes apart, for four consecutive days. Each trial lasted a maximum of 3 minutes, and the time taken to reach the avoidance box was measured. After arrival, the mouse was required to wait in the avoidance box for 1 minute. However, if the mouse did not reach the avoidance box within 3 minutes, it was guided to the box and given another 1 minute to memorize the location. On the day following the completion of the four consecutive days of training, a probe test was conducted with a maximum trial time of 3 minutes, and the time taken to reach the avoidance box was measured. A shorter arrival time to the avoidance box was considered to indicate higher learning ability. The results of the Barnes maze test are shown in Figure 1. Mice in test group 3, fed a diet containing buckwheat starch, showed a reduction in the time it took to escape the maze and reach the escape box compared to mice in test group 2, which were fed a control diet lacking the specific starch. No significant changes were observed in the time it took for mice in test group 4, fed a diet containing wheat starch, or in test group 5, fed a diet containing rice starch, to reach the escape box. 【0034】 <Passive Avoidance Test> The passive avoidance test was conducted using the step-through test cage SCT-001M (Muromachi). The step-through test cage consists of two rooms, a dark room and a light room, separated by a partition door. The test consisted of an acquisition trial on day 1 and a recall trial on day 2. In the acquisition trial, the mouse was placed in the light room, the partition door was opened, and after moving to the dark room, the partition door was closed, and an electrical stimulation of 1.0 mA for 3 seconds was applied. After the electrical stimulation, the mouse was made to wait in the dark room for 10 seconds. The trial time from moving from the light room to the dark room was recorded, with a maximum of 3 minutes. In the recall trial, the mouse was placed in the light room, the partition door was opened, and the trial time from moving to the dark room was recorded, with a maximum of 7 minutes. The longer the time spent in the light room, the higher the learning ability was evaluated. The results of the passive avoidance test are shown in Figure 2. Mice in test group 3, fed a diet containing buckwheat starch, showed a significantly longer duration of stay in the brightly lit room as an avoidance behavior during the regeneration trial compared to mice in test group 2, fed a control diet that did not contain the specific starch. No significant changes were observed in mice in test group 4, fed a diet containing wheat starch, or in test group 5, fed a diet containing rice starch. 【0035】 <Real-time PCR method for gene expression analysis of BDNF> The gene expression analysis of BDNF using real-time PCR was performed as follows. 1.0 mL of RNAiso Plus (Takara Bio) was added to the entire excised hippocampus, and the sample was ground on ice using a biomassher (Nippi). Total RNA was extracted according to the RNAiso Plus protocol. The purity and concentration of total RNA were measured using Nano Drop Lite (Thermo Fisher Scientific). OD 280 OD 260 Ratio (OD 260 / OD 280RNA with a gDNA ratio in the range of 1.8 to 2.1 was used for cDNA synthesis. ReverTra Ace qPCR RT Master Mix with gDNA Remover (TOYOBO) was used for cDNA synthesis. 1.5 μg of RNA template was mixed with 6 μL of 4×DN Master Mix containing gDNA Remover and Nuclease-free Water to a total volume of 24 μL. The mixture was thoroughly mixed to ensure homogeneity, and the DNase reaction was carried out by incubation at 37°C for 5 minutes. The reaction solution was then transferred back to ice, 6 μL of 5×RT Master Mix II was added, and the mixture was thoroughly mixed to ensure homogeneity. Reverse transcription was then performed using a 2720 Thermal Cycler (Applied Biosystems) by incubation at 37°C for 15 minutes, 50°C for 5 minutes, and 98°C for 5 minutes. BDNF gene expression levels were measured by real-time PCR using a Thermal Cycler Dice(R) Real Time System Single (Takara Bio). THUNDERBIRD Next SYBR qPCR Mix (TOYOBO) was used for amplification. The reaction solution was composed of 1 μL of 10-fold diluted synthesized cDNA, 0.2 μL each of primer solution (10 μmol / μL), 10 μL of THUNDERBIRD Next SYBR qPCR Mix, and 3.8 μL of SP water, mixed to a total volume of 10 μL. The reaction conditions were as follows: pre-incubation at 95°C for 1 minute, followed by 40 cycles of 2-step PCR at 95°C for 5 seconds and 60°C for 30 seconds. The expression level of BDNF mRNA was analyzed using the comparative Ct method (ΔΔCt method) with GAPDH mRNA, a housekeeping gene, as the internal standard gene. The results are shown in Figure 3. 【0036】 <Analysis of BDNF expression using Western blotting method> The expression analysis of BDNF using the Western blotting method was performed as follows. For SDS-PAGE, a separation gel containing 12% acrylamide and a concentration gel containing 5% acrylamide were used, and electrophoresis was performed under conditions of 100V, 100mA, and 120min. After electrophoresis, the gels were immersed in Transfer Buffer (39mM Glycine, 42mM Tris, 20% (v / v)MeOH) for 10 minutes, and then transferred to PVDF membranes (Merck Millipore) using a semi-dry transfer apparatus WES-4020 (ATTO) under conditions of 30V, 90mA / mesh, and 90 minutes. The PVDF membranes used were shaken in MeOH for 30 seconds, MilliQ for 1 minute, and Transfer Buffer for 5 minutes before use. After transfer, the PVDF membranes were immersed in Blocking One (Nacalai tesque) and shaken at room temperature for 20 minutes to block. Subsequently, they were washed three times with PBST or TBST for 5 minutes each. As the primary antibody, anti-BDNF antibody (5,000-fold dilution, Abcam) was diluted with 5% Blocking One in PBST and shaken overnight at 4°C. After washing three times with PBST for 5 minutes each, as the secondary antibody, Goat Anti-Rabbit IgG(H+L)-HRP Conjugate (10,000-fold dilution, Bio-Rad, California, USA) was diluted with 5% Blocking One in PBST and reacted by shaking at room temperature for 1 hour. After washing three times with PBST for 5 minutes each, the PVDF membrane, which had undergone a luminescence reaction using Ez WestLumi plus WES-7120L (ATTO), was imaged using a chemiluminescence imaging system EZ-capture MG (ATTO). As the loading control antibody, Goat Anti-Mouse IgG (H+L)-HRP Conjugate (10,000-fold dilution, Bio-Rad) diluted with 5% Blocking One in PBST was used. The mixture was reacted by shaking at room temperature for 1 hour, then washed three times with PBST for 5 minutes each, and finally subjected to imaging. The target protein was quantified using the ImageJ analysis software. The results are shown in Figure 4. 【0037】 As shown in Figures 3 and 4, the results of the physiological analysis of mice (BDNF expression analysis: real-time PCR method, Western blotting method) regarding the effect on improving dementia showed a significant increase in BDNF expression in the hippocampus at both the gene and protein levels in mice fed a diet containing buckwheat starch. On the other hand, there was no change in BDNF expression levels in mice fed a diet containing wheat starch or a diet containing rice starch. [Industrial applicability] 【0038】 The present invention provides a composition for improving dementia, which contains buckwheat starch as an active ingredient, derived from buckwheat, a common food commonly consumed in Japan. This composition can be provided in beverages, foods, and pharmaceuticals.

Claims

[Claim 1] A composition for improving dementia, containing buckwheat starch as an active ingredient. [Claim 2] A beverage for improving dementia containing the dementia-improving composition described in claim 1. [Claim 3] A food for improving dementia containing the dementia-improving composition described in claim 1. [Claim 4] A pharmaceutical agent for improving dementia containing the dementia-improving composition described in claim 1.

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