JP7044306B2 - Leucine blood concentration increase promoter and its uses - Google Patents

Description

Published on the website to which Article 30, Paragraph 2 of the Patent Act is applied. Website publication date April 27, 2018 Website address http: // www. jsnfs. or. jp https: // member. jsnfs. or. jp / eishoku_db / index. php

Applicable to Article 30, Paragraph 2 of the Patent Act Published in publications Publisher name Publication name of the Japan Society of Nutrition and Food Science Publication name Proceedings of the 72nd Annual Meeting of the Japan Society of Nutrition and Food Science Publication date April 27, 2018 Day

Patent Law Article 30, Paragraph 2 Applicable Society Open to the public Date May 12, 2018 Meeting name, venue 72nd Japan Society for Nutrition and Food Science Okayama Prefectural University (111 Kuboki, Soja City, Okayama Prefecture)

The present invention relates to a leucine blood concentration increase accelerator containing nanoparticles and its use, and is used, for example, for improving endurance.

Endurance is indispensable for the continuation of whole body exercise and local muscle exercise, and it is no exaggeration to say that endurance determines the outcome, especially in sports competitions where it is necessary to continue exercising for a long time. .. Endurance is indispensable for all daily activities such as working, walking, and standing, in addition to sports. However, in recent years, with the development of transportation means and the development of information and communication technology, exercise tends to be insufficient, and as a result, endurance, muscle weakness, and motor function deterioration occur, and the subsequent QOL (Quality Of Life) There is concern that it will have a serious adverse effect on the quality of life.

One way to improve endurance is to improve muscle endurance. Muscle endurance is the endurance of the muscle, which is how many times the repeated load can be continued. In order to increase muscle endurance, it is important to strengthen skeletal muscle. Skeletal muscle mass is largely due to an increase or decrease in the amount of skeletal muscle protein. If the synthesis of skeletal muscle protein exceeds the decomposition of skeletal muscle protein, the skeletal muscle is enlarged, and conversely, if the decomposition exceeds the synthesis, the skeletal muscle Causes atrophy. Therefore, in order to increase skeletal muscle mass and strengthen skeletal muscle, it is important to promote the synthesis of skeletal muscle protein and suppress its degradation.

Milk-derived whey protein is an ingredient that is ingested for the purpose of increasing skeletal muscle mass. Whey protein contains a lot of branched-chain amino acids such as valine, leucine, and isoleucine, and has a faster rate of digestion and absorption after ingestion than casein, which is a protein derived from the same milk, so it promotes immediate muscle synthesis. It is known that it is effective and can efficiently strengthen muscles while preventing muscle fatigue, especially when taken during exercise.

Among the branched-chain amino acids, leucine has a higher effect of promoting skeletal muscle protein synthesis than other amino acids, but also has a function of suppressing the decomposition of skeletal muscle protein, so that it plays a key role in muscle building. It is known that this is achieved (Non-Patent Document 1). That is, when the blood leucine level decreases, the action of mTOR (mammalian Target Of Rapamycin) present in skeletal muscle cells is suppressed, and protein synthesis is also suppressed accordingly. On the other hand, when the blood leucine concentration increases, the action of mTOR is activated and protein synthesis is switched on, and as a result, muscle synthesis is considered to be promoted (Non-Patent Document 2). Therefore, how to maintain high blood leucine concentration is considered to be one of the keys to strengthen muscles and improve endurance.

For example, Patent Document 1 discloses an agent for promoting an increase in blood amino acid concentration, which comprises a whey protein hydrolyzate containing a specific dipeptide as an active ingredient. However, although the blood amino acid increase-promoting agent has an immediate (immediate) blood amino acid concentration increase-promoting effect, the blood leucine concentration over a long period of time such as 10 hours, 15 hours, and 20 hours after ingestion. The effect of keeping the blood high has not been shown.

Patent Document 2 discloses a casein composition for increasing the blood leucine concentration to the same level as when whey protein is ingested. However, the effect of ingesting whey protein to raise it to a level exceeding the blood leucine concentration has not been shown.

Patent Document 3 shows that an increase in blood amino acid concentration stimulates muscle protein synthesis, and leucine is particularly important in the elderly. However, as shown in FIGS. 1 to 3, administration is performed. Since the blood amino acid concentration returned to the same level as before the administration in about 4 hours, it is considered that the duration of the effect is about 4 hours.

Considering the prior art from the viewpoint of the endurance improving effect, for example, Patent Document 4 discloses an endurance improving agent containing a branched α-glucan mixture as an active ingredient. ing. However, when the endurance enhancer was ingested at a high dose of 25 g to 41.7 g of the branched α-glucan mixture in terms of anhydride, the exercise duration by the bicycle ergometer was 25 minutes and 28 seconds in the control group. It is 9 to 43 seconds longer, and it is hard to say that it has reached the level expected by consumers in terms of dose and effect.

Patent Document 5 discloses an endurance improving agent containing S-allylcysteine as an active ingredient. S-allylcysteine is a water-soluble sulfur compound derived from garlic, and although it has less odor and irritation than garlic-specific stimulants such as allicin, it still has a peculiar odor and has a problem in material stability.

Patent Document 6 describes a method for exerting an endurance-enhancing effect including a step of continuously ingesting a catechin compound to a human or a non-human animal, and a step of ingesting sugar, alanine and proline before exercise. Is disclosed. That is, the combined intake of the composition containing the catechin compound and the pre-exercise intake of the glycogenic amino acid composition containing sugar, alanine and proline is the composition containing the catechin compound or the glycogenic amino acid composition. It has been shown to show a higher endurance-enhancing effect than when each of the substances is ingested alone. Here, Patent Document 6 does not describe leucine, and it is described that a healthy human adult acts for 30 to 180 minutes by ingesting it 10 to 120 minutes before the start of exercise. Therefore, it is considered that the duration of the effect is about 5 hours at the maximum.

Japanese Unexamined Patent Publication No. 2017-212954 Japanese Patent Application Laid-Open No. 2015-509531 Special Table 2013-515718A Japanese Unexamined Patent Publication No. 2018-24619 Japanese Unexamined Patent Publication No. 2017-88549 Japanese Unexamined Patent Publication No. 2016-160216

Nagasawa T et al. , J Nutr Biochem, 13 (2): 121-127 (2002). Wolfson RL et al. , Science, 351 (6268): 43-48 (2016)

The subject of the present invention is not only sports competition, but also a blood leucine concentration increase promoter for improving endurance necessary for sustaining movements such as labor, walking, and standing, and a method for producing the same. It is an object of the present invention to provide a safe and effective endurance-enhancing food and drink composition containing the above-mentioned agent for increasing blood leucine concentration.

As a result of diligent research to solve the above problems, the present inventors have an effect of improving endurance on nanoparticles obtained by heating and drying a preparation obtained by mixing catechin, whey protein, and leucine. It was found that there was a problem, and the present invention was completed.

[1] The first aspect of the present invention contains nanoparticles containing catechin, whey protein and leucine as active ingredients, and is a zeta potential / nanoparticle size measurement system (“DelsaMax PRO” manufactured by Beckman Coulter Co., Ltd.). A blood leucine concentration increase promoter having an average particle size of 40 to 200 nm of the nanoparticles measured in 1 .
[2] The agent for increasing the blood leucine concentration according to the above [1], wherein the nanoparticles further contain a polysaccharide .

[ 3 ] The second aspect of the present invention is an endurance-enhancing food and drink composition containing the blood leucine concentration increase-promoting agent according to the above [1] or [2] as an active ingredient.

[ 4 ] The third aspect of the present invention is a step (A) of mixing catechin, whey protein, and leucine to prepare a preparation, and heat-treating the preparation obtained in the step (A) to prepare nanoparticles. The zeta potential includes a step (B) of obtaining a nanoparticle-containing liquid by producing the nanoparticles, and a step (C) of drying the nanoparticles-containing liquid obtained in the step (B) to obtain a nanoparticle-containing powder. A method for producing a blood leucine concentration increase accelerator having an average particle size of 40 to 200 nm of the nanoparticles-containing powder measured by a nanoparticle size measurement system (“DelsaMax PRO” manufactured by Beckman Coulter Co., Ltd.). ..
[ 5 ] The method for producing the blood leucine concentration increase accelerator according to the above [ 4 ], which prepares a preparation solution further containing a polysaccharide in the step (A).
[ 6 ] The method for producing the blood leucine concentration increase accelerator according to the above [ 4 ] or [ 5 ], wherein the heat treatment is carried out at 70 to 98 ° C. for 2 to 180 minutes in the step (B).

The blood leucine concentration increase promoter of the present invention can maintain a blood leucine concentration at a higher level than before ingestion for at least 24 hours after ingestion by animals including humans. Therefore, a food and drink composition for improving endurance for improving the endurance necessary for spending a healthy life in sports competition or daily life by containing the above-mentioned blood leucine concentration increase promoter as an active ingredient is provided. It will be possible to do.

FIG. 1 is a schematic explanatory view of the manufacturing process of nanoparticles of the present invention in Example 1. FIG. 2 is a diagram showing changes over time in blood leucine concentration after administration of a whey protein-leucine mixture and leucine-supported nanoparticles (leucine-supported whey), respectively. In FIG. 2, "*" represents P <0.05, "**" represents P <0.01, and "***" represents P <0.001, a whey protein-leucine mixture at the same time point. Represents a significant difference from the intake group. FIG. 3 is a diagram showing changes over time in oxygen consumption after administration of a whey protein-leucine mixture and leucine-supported nanoparticles (leucine-supported nanoparticles), respectively. FIG. 4 is a diagram showing changes over time in respiratory quotient after administration of a whey protein-leucine mixture and leucine-supported nanoparticles (leucine-supported nanoparticles), respectively. FIG. 5 is a diagram showing oxygen consumption from 5 hours to 12 hours (7 hours in total) after administration of the whey protein-leucine mixture and leucine-supported nanoparticles (leucine-supported nanoparticles), respectively. In FIG. 5, "*" represents P <0.05. FIG. 6 is a diagram showing the amount of lipid oxidation from 5 hours to 12 hours (7 hours in total) after administration of the whey protein-leucine mixture and the leucine-supported nanoparticles (leucine-supported nanoparticles), respectively. FIG. 7 is a diagram showing the amount of sugar oxidation from 5 hours to 12 hours (7 hours in total) after administration of the whey protein-leucine mixture and the leucine-supported nanoparticles (leucine-supported nanoparticles), respectively. FIG. 8 is a diagram showing the total amount (integrated value) of the travel distance from 5 hours to 12 hours (7 hours in total) after administration of the whey protein-leucine mixture and the leucine-supported nanoparticles (leucine-supported nanoparticles), respectively. FIG. 9 shows the correlation between the oxygen consumption and the integrated value of the travel distance from 5 hours to 12 hours (7 hours in total) after administration of the whey protein-leucine mixture and the leucine-supported nanoparticles (leucine-supported nanoparticles), respectively. Is.

1. 1. Blood leucine concentration increase promoter In the present invention, the "blood leucine concentration increase promoter" refers to an agent having a blood leucine concentration increase promoting effect, and specifically, an animal including humans (for example, a mammal). , Reptiles, birds, etc., hereinafter also referred to as the target) refers to an agent having the effect of increasing the blood leucine concentration of the target as compared with the case of not ingesting it.

In particular, the agent for increasing the blood leucine concentration of the present invention has the effect of maintaining the blood leucine concentration of the subject at a higher level for a long period of time when the subject ingests it, as compared with before the ingestion. Play.

In the present invention, "high level" means that the subject who ingested the blood leucine concentration increase promoter of the present invention has a chemical component composition with catechin, whey protein and leucine contained in the blood leucine concentration increase promoter. The blood leucine concentration is 5% or more, preferably 10% or more, more preferably 20% or more higher than the case of ingesting the same "whey protein-leucine mixture" (non-nanoparticles). Means that.

Further, in the present invention, "long time" means at least 24 hours, preferably about 24-36 hours, more preferably about 24-48 hours.
For example, when the subject is a human, the blood leucine concentration increases about 10 to 120 minutes (preferably 10 to 60 minutes) after the subject ingests the "promoter for increasing blood leucine concentration" of the present invention. It rises and then retains high levels of leucine blood levels for about 24-36 hours compared to when the subject ingested the control "whey protein-leucine mixture". Therefore, according to a preferred embodiment of the present invention, the blood leucine concentration increase-promoting agent of the present invention is also suitable as a "blood leucine concentration high level long-term holding agent".

The blood leucine concentration increase promoter of the present invention contains nanoparticles containing catechin, whey protein and leucine as an active ingredient.

In the present invention, the "active ingredient" means an ingredient required for exerting an effect of promoting an increase in blood leucine concentration.

In the present invention, the "nanoparticle" contains leucine in nanoparticles containing whey protein as a main component (hereinafter, also referred to as leucine-supported nanoparticles).
The nanoparticles are obtained by converting whey protein into nanoparticles by heat-treating a solution containing catechin, whey protein and leucine.
The structure of the nanoparticles is not clear in detail, but it is presumed that leucine is adsorbed on the surface of the nanoparticles of whey protein.

In the present invention, "catechin" is used to make whey protein nanoparticles in a solvent as described below. The catechin used in the present invention may be any polyphenol peculiar to green tea. "Gallate-type catechins" such as Cg), galocatechin gallate (GCg), and epigalocatechins (EGC), epicatechins (EC), catechins (C), galocatechins (GC) without "gallate groups". Examples include "free catechins".

In the present invention, the catechin may be used alone or as a combination of two or more types (catechin mixture), but it is easy to obtain and has a good flavor. A catechin mixture can be preferably used. Examples of such a catechin mixture include a tea extract obtained by drying a tea extract obtained by extracting tea with water. The tea extract may be prepared and used by itself, or a commercially available product may be obtained and used. Commercially available tea extracts include, for example, "Sanphenon 90S" (Taiyo Kagaku Co., Ltd.), "Sanphenon BG-3" (Taiyo Kagaku Co., Ltd.), "Sanphenon BG-5" (Taiyo Kagaku Co., Ltd.), "Sanphenon EGCg". -OP "(Taiyo Kagaku Co., Ltd.)," Camellia Extract 30S "(Taiyo Kagaku Co., Ltd.) and the like can be mentioned.

In the present invention, "whey protein" is a component constituting the main component of nanoparticles. The whey protein used in the present invention is a general term for proteins contained in whey from which casein and milk fat are removed from raw milk of cattle, sheep, humans, etc., and is, for example, whey protein concentrate (whey protein concentrate). Whey Protein Concentrate (also called WPC), whey protein isolate (also called Whey Protein Isolate, WPI), whey protein with specific proteins such as β-lactoglobulin extracted, and other undiluted whey solutions (sweetness). Examples include whey (whey, acid whey, etc.), its dried product (whey powder, etc.), and its frozen product. The whey protein may be prepared and used by itself, or a commercially available product may be obtained and used.

Examples of commercially available whey proteins include "Enlacto HUS" (Nippon Shinyaku Co., Ltd.), a WPC material derived from cheese whey, "Enlacto ALC" (Nippon Shinyaku Co., Ltd.), and "PROGEL 800", a WPC material derived from acid whey. Nippon Shinyaku Co., Ltd.), WPI material "Enlacto SAT" (Nippon Shinyaku Co., Ltd.), "Enlacto YYY" (Nippon Shinyaku Co., Ltd.), "Lact Crystal" (Nippon Shinyaku Co., Ltd.), etc. can be mentioned.

In the present invention, "leucine" means an amino acid, L-leucine or a physiologically acceptable salt thereof. Examples of leucine applicable in the present invention include naturally derived ones, chemically synthesized ones, and those obtained by combining these. Examples of the commercially available L-leucine include L-leucine (Ajinomoto Co., Inc.).

Further, the nanoparticles may further contain polysaccharides, if necessary. In the present invention, "polysaccharide" means a general term for substances in which a plurality of monosaccharide molecules are polymerized by glycosidic bonds. Examples of the polysaccharide used in the present invention include polysaccharides having a "suspension stabilizing effect" that keeps a state in which difficult-to-dissolve components put into a solution are uniformly dispersed for a long period of time. Specific examples of such a polysaccharide include tamarind seed gum, xanthan gum, locust bean gum and the like, but the polysaccharide is particularly limited as long as it can be used as a food or drink and has the suspension stabilizing effect. Not done. Examples of commercially available tamarind seed gum include "Glyroid 3C" (DSP Gokyo Food & Chemical Co., Ltd.).

In the present invention, the average particle size of the nanoparticles is preferably 40 to 200 nm, more preferably 45 to 160 nm, still more preferably 50 to 50, from the viewpoint of dispersion stability of the nanoparticles in a liquid and absorption into the body. It is 120 nm. The average particle size of the nanoparticles can be measured by a zeta potential / nanoparticle size measurement system (“DelsaMax PRO” manufactured by Beckman Coulter, Inc.).

As described above, the agent for increasing the blood leucine concentration of the present invention may be only nanoparticles containing catechin, whey protein and leucine and, if necessary, polysaccharides, or may be necessary while using the nanoparticles as an active ingredient. Depending on the content, components such as a bulking agent, a solubilizer, a dispersant, a suspending agent, an emulsifier, an antioxidant, a bacterial inhibitor, a coloring agent, a flavoring agent, and a flavoring agent may be contained. These components may be the rest of the catechin, whey protein and leucine and, if necessary, polysaccharides, for example, the total amount of these components is based on 100% by weight of the dry weight of the blood leucine concentration increase promoter. It may be 55% by weight or less.

As for the content of each component in the blood leucine concentration increasing accelerator of the present invention, the blood leucine concentration is obtained from the viewpoint that nanoparticles having a desired particle size can be efficiently obtained and the blood leucine concentration increasing effect is excellent. It is preferable that the concentration is adjusted to 0.5 to 5% by weight of catechin, 30 to 98.5% by weight of whey protein, and 0.3 to 10% by weight of leucine with respect to 100% by weight of the dry weight of the ascending accelerator. When the blood leucine concentration increase accelerator of the present invention contains a polysaccharide, the content of the polysaccharide is preferably 0.005 to 0.1% by weight.

The agent for increasing the blood leucine concentration of the present invention is
Step (A) of mixing catechin, whey protein, and leucine to prepare a preparation.
The step (B) of obtaining the nanoparticles-containing liquid by heat-treating the preparation liquid obtained in the step (A) to generate nanoparticles, and the step (B) of drying the nanoparticles-containing liquid obtained in the step (B). , Step of obtaining nanoparticles containing nanoparticles (C)
It can be manufactured by passing through.

Hereinafter, steps (A) to (C) will be described.
[Step (A)]
In this step, three raw materials of catechin, whey protein, and leucine are mixed to prepare a preparation. Specifically, catechin, whey protein, and leucine are dissolved or dispersed in water or a hydrous organic solvent or an organic solvent to prepare a preparation solution uniformly containing them.

Examples of the water used as the solvent for dissolving or dispersing the three raw materials include pure water, distilled water, tap water, and commercially available drinking water, but the water is not particularly limited. When an organic solvent is used as the solvent, it may be miscible with water and is not particularly limited. Further, it is preferable to select a solvent suitable for the intended use of the obtained nanoparticles. For example, examples of the solvent suitable for food use include glycerin, propylene glycol, ethanol and the like, and examples of the solvent are suitable for pharmaceutical use. In addition to the above, methanol, acetone, dimethyl sulfoxide and the like can be mentioned. The water-containing organic solvent used as the solvent means a mixed solvent of the organic solvent and water.

Means for mixing each raw material with the solvent include dissolving or dispersing each raw material in the solvent. For example, when preparing a formulation containing catechin, whey protein, leucine, and polysaccharide, each may be mixed in a powder state and then dissolved or dispersed, or a solution or dispersion of each raw material may be prepared. Then they may be mixed. Further, when dissolving or dispersing, it is preferable to adjust the temperature of the solvent to 20 to 50 ° C. from the viewpoint of improving the solubility of each raw material, but there is no particular limitation as long as it is dissolved or dispersed. ..

From the viewpoint of efficiently producing nanoparticles, the amount of catechin in the preparation solution prepared in step (A) is preferably 0.01 to 0.5% by weight, more preferably 0.02 to 0.4% by weight. %, Most preferably 0.03 to 0.3% by weight.
The amount of whey protein in the preparation solution prepared in the step (A) is preferably 0.2 to 7% by weight, more preferably 0.2 to 6% by weight, from the viewpoint of efficiently producing nanoparticles. , More preferably 0.2 to 5% by weight.
From the viewpoint of efficiently producing nanoparticles, the amount of leucine in the preparation solution prepared in the step (A) is preferably 0.5 to 2% by weight, more preferably 0.5 to 1% by weight of the amount of whey protein. It is preferably prepared in an amount of 5% by weight, most preferably 0.5 to 1.2% by weight.

The pH of the preparation is preferably 1.0 to 2.5 or 5.0 to 8.0, more preferably 1.5 to 2, from the viewpoint of adjusting the particle size of the nanoparticles to a desired range. Adjust with acid or alkali to .2 or 5.5 to 7.0, more preferably 1.7 to 2.0 or 6.0 to 6.8. When the pH is less than 1.0 or around 3.9, the particle size becomes large and the nanoparticles dissolve.

In order to adjust the pH of the preparation, for example, the pH of a catechin-containing solution or dispersion, a whey protein-containing solution or swelling solution, a leucine-containing solution or dispersion, and, if necessary, a polysaccharide-containing solution can be adjusted. Each may be pre-adjusted with an acid or an alkali. By adjusting each raw material solution to a desired pH in advance and then mixing them in this way, the pH of the prepared solution can also be adjusted to the above-mentioned suitable range. Further, it is also possible to put each raw material into a solvent as a powder and mix an acid or an alkali with the obtained preparation liquid to adjust the pH to a desired value.

[Step (B)]
In this step, leucine-supported nanoparticles are produced by heating the preparation solution prepared in the step (A). Further, in this step, from the viewpoint of efficiently obtaining leucine-supported nanoparticles having an average particle diameter of 40 to 200 nm, it is preferably 70 to 98 ° C. for 2 to 120 minutes, more preferably 70 to 90 ° C. for 2 to 160 minutes, and further. It is preferable to treat at 75 to 85 ° C. for 2 to 180 minutes. The heat treatment is preferably performed while stirring the above-mentioned preparation solution. In the present invention, "stirring" means that the contents in the container are mixed. Specifically, stirring may be performed manually using a stirring rod or the like, or may be performed using a stirrer such as a magnetic stirrer or a mixer.

Further, by further diluting the preparation solution that produced the leucine-supported nanoparticles, the aggregation of the obtained leucine-supported nanoparticles was suppressed, and the particles were dried while maintaining the desired average particle size even during the drying described later. It has the advantage of being easy. The dilution method may be any method as long as it can be mixed uniformly, and examples thereof include a method of adding the diluted solution while stirring, a method of adding the diluted solution while homogenizing, and the like, but the method is not particularly limited. Here, as the dilution ratio, for example, a diluted solution in an amount of 2 to 100 times the amount of the obtained leucine-supported nanoparticles-containing compounding solution may be mixed.

[Step (C)]
In this step, the leucine-supported nanoparticles-containing compound obtained in the step (B) is dried to obtain solid leucine-supported nanoparticles. In this step, the leucine-supported nanoparticles formed in the diluted solution obtained in the step (B) can be further dried to produce solid leucine-supported nanoparticles having stable physical properties. can. Examples of the drying method include spray drying, heating and vacuum drying, and the like.

Spray drying is a method of spraying a liquid into a gas and rapidly drying it to produce a dry powder. Heating vacuum drying is a method of promoting drying by depressurizing the inside of a heating device to lower the boiling point, and evaporating and drying with less energy. All of these drying methods may be performed using a known drying device. The temperature conditions at the time of drying may be set to an appropriate temperature range according to each drying method. For example, the outlet temperature is adjusted to 50 to 100 ° C. for spray drying and 20 to 100 ° C. for heated vacuum drying. However, there are no particular restrictions.

The blood leucine concentration increase promoter of the present invention obtained as described above is excellent in the effect of increasing the blood leucine concentration of the subject who ingested it and maintaining the state.

For the measurement of the blood leucine concentration, blood collected from the subject before and after ingestion of the blood leucine concentration increase promoter of the present invention is used. The method for collecting blood is not particularly limited, but a blood anticoagulant may be used to prevent blood coagulation. Examples of the blood anticoagulant include heparin, sodium citrate, EDTA and the like. In addition, the blood may be prepared as serum by a method well known in the art, and may be directly used for measurement or stored. The blood thus collected may be measured by, for example, a high performance liquid chromatography (HPLC) method. In the high performance liquid chromatography method, amino acids are derivatized by a pre-column derivatization method (AQC method, OPA method, FMOC method, etc.) or a post-column derivatization method (ninhydrin method, OPA method, etc.), and the derivatized amino acids are ultraviolet. There is a method of detecting by a visible absorptiometry method (UV method), a fluorescence method, a mass spectrometry method (MS method, MS / MS method), or the like.

The agent for increasing the blood leucine concentration of the present invention may be formulated. The form of this formulation is not particularly limited, and for example, tablets, coated tablets, powders, fine granules, granules, capsules, liquids, pills, suspensions, emulsions, troches, chewable tablets, syrups and the like. Oral preparations and the like can be mentioned. At the time of formulation, carriers, excipients, lubricants, binders, disintegrants, diluents, stabilizers, tonicity agents, pH adjusters, buffers and the like are used.

Examples of carriers and excipients include lactose, sucrose, sodium alginate, glucose, maltose, mannitol, erythritol, xylitol, maltitol, inositol, dextran, sorbitol, albumin, urea, starch, calcium carbonate, kaolin, and crystalline cellulose. , Silica, methylcellulose, glycerin, sodium alginate, gum arabic and mixtures thereof.

Examples of the lubricant include purified talc, stearate, borax, polyethylene glycol and mixtures thereof.
Examples of the binder include simple syrup, glucose solution, starch solution, gelatin solution, polyvinyl alcohol, polyvinyl ether, polyvinyl pyrrolidone, carboxymethyl cellulose, cellac, methyl cellulose, ethyl cellulose, water, ethanol, potassium phosphate and mixtures thereof. Can be mentioned.

Examples of the disintegrant include dried starch, sodium alginate, canten powder, laminaran powder, sodium hydrogencarbonate, calcium carbonate, polyoxyethylene sorbitan fatty acid esters, sodium lauryl sulfate, stearic acid monoglyceride, starch, lactose and mixtures thereof. Can be mentioned.

Examples of the diluent include water, ethyl alcohol, macrogol, propylene glycol, ethoxylated isostearyl alcohol, polyoxylated isostearyl alcohol, polyoxyethylene sorbitan fatty acid esters, and mixtures thereof.
Examples of the stabilizer include sodium metabisulfite, ethylenediaminetetraacetic acid, thioglycolic acid, thiolactic acid and mixtures thereof.

Examples of the tonicity agent include sodium chloride, boric acid, glucose, glycerin and a mixture thereof.
Examples of the pH adjuster and buffer include sodium citrate, citric acid, sodium acetate, sodium phosphate and mixtures thereof.

2. 2. Endurance-enhancing food and drink composition The endurance-enhancing food and drink composition of the present invention contains the blood leucine concentration increasing accelerator as an active ingredient.
Here, the "active ingredient" means an ingredient required for exerting an endurance improving effect. In one preferred embodiment, the "active ingredient" has the effect of retaining high levels of leucine in the blood for extended periods of time for subjects who need to maintain high levels of leucine in the blood for extended periods of time. It means the ingredients required above.

In the present invention, "a subject whose blood leucine concentration needs to be maintained at a high level for a long period of time" means a subject expected to have an endurance improving effect, for example, a marathon, on land / long distance, or a bicycle road. , Triathlon, cross-country, swimming / long-distance, speed skating long-distance, soccer, basketball, rugby and other endurance-requiring athletes and sports enthusiasts (athletes), health promotion, athletic performance improvement , Subjects exercising for the purpose of recovery from fatigue or improvement of endurance.

In the present invention, "endurance" means endurance in exercise or muscle exertion, and means a certain action / function, for example, the ability to continuously perform an exercise state.
The "exercise" means physical activity. For example, marathon, long-distance running / long-distance running, bicycle road, triathlon, cross-country skiing, long-distance swimming / long-distance running, speed skating long-distance running, soccer, basketball, rugby and other endurance-requiring sports, labor, walking, standing, etc. Includes all daily activities of.
The above-mentioned "muscle exertion" means to move a muscle.
Further, the "endurance improving action" means an endurance improving action in exercise / muscle exertion, and an action of enhancing the ability to be continuously performed, for example, specifically, the duration of exercise / muscle exertion. It can be confirmed by extending. For example, "you can continue exercising and playing sports for a long time", "you will not get tired even if you work for a long time", "you will not get tired in your daily life" and so on.
Therefore, the target for ingesting the blood leucine concentration increase promoter or the endurance-enhancing food and drink composition of the present invention is not limited to, for example, athletes and sports enthusiasts (athletes), but is not limited to athletes, sports enthusiasts (athletes), etc. , Subjects in need of recovery from fatigue, or improvement of endurance. Further, the target may be animals other than humans (livestock such as horses and cows, pet animals such as dogs and cats, and ornamental animals bred in zoos and the like).

The endurance improving effect in the present invention can be evaluated by oxygen consumption analysis or measurement of travel distance using mice, for example, as in the examples described later.

In the food and drink composition for improving endurance of the present invention, other food raw materials and / or food additive raw materials may be added in addition to the blood leucine concentration increasing accelerator as long as the endurance improving effect is exhibited. can.
For example, sweeteners (sugars, liquid sugars, fructose, malt sugar, trithermal sugar, etc.), sugar alcohols (erythritol, etc.), high-sweetness sweeteners (stevia, aspartame, sucralose, acesulfame potassium, etc.) for the purpose of imparting palatability. ), Acidulants, thickening polysaccharides, fragrances, fructose, vegetable juice and the like can be added. Further, for the purpose of imparting function, minerals (calcium, iron, manganese, magnesium, zinc, etc.), vitamins, functional materials, probiotic lactic acid bacteria, etc. are added to the food and drink composition for improving endurance of the present invention. It can be a thing.

The form of the food and drink composition for improving endurance of the present invention is other than the pharmaceutical composition and may be a form that can be orally ingested such as a solution, a suspension, an emulsion, a powder, or a solid molded product. It is not particularly limited. Specifically, for example, instant foods such as instant noodles, retort foods, canned foods, microwave foods, instant soups / miso juices, and freeze-dried foods; soft drinks, fruit juice drinks, vegetable drinks, soy milk drinks, coffee drinks, and tea. Beverages, powdered beverages, concentrated beverages, alcoholic beverages and other beverages; bread, pasta, noodles, cake mixes, bread flour and other wheat flour products; candy, caramel, chewing gum, chocolate, cookies, biscuits, cakes, pies, snacks, crackers, Confectionery such as Japanese confectionery and dessert confectionery; seasonings such as sauces, processed tomato seasonings, flavor seasonings, cooking mixes, sauces, dressings, soups, curry and stew ingredients; Oils and fats such as mayonnaise; dairy products such as dairy beverages, yogurts, lactic acid bacteria beverages, ice creams and creams; canned agricultural products, jams and marmalades, processed agricultural products such as cereals; frozen foods and the like. Preferably, the food is a dairy product, more preferably a milk beverage, a lactic acid bacteria beverage.

In addition, foods include health foods, functional foods, nutritional supplements, foods with functional claims, foods for specified health use, foods for the sick, adjusted powdered milk for infants, powdered milk for pregnant or lactating women, or increased blood amino acid concentration. Also included are foods classified as foods labeled as being used for promotion. Further, in the present invention, food is a concept including beverages.

In addition, the food and drink composition for improving endurance of the present invention can be in the form of powder in a container as one aspect for the purpose of providing it on the market.
As the container, known ones such as paper, bags, boxes, cans, and bottles can be used.
The capacity of the container is not particularly limited, and is, for example, 1 to 5000 g, preferably 2 to 4000 g, and more preferably 3 to 3000 g. Among them, in the case of family size, for example, 300 to 5000 g, preferably 300 to 4000 g, more preferably 300 to 3000 g, and in the case of personal size, for example, 1 to 500 g, preferably 2 to 250 g, more preferably. Is 3 to 200 g.

Further, the food and drink composition for improving endurance of the present invention can be in the form of a beverage in a container (a beverage in which powder is dispersed and dissolved) as one aspect for the purpose of providing it on the market.
As the container, a known container such as a paper container, a soft bag, a PET bottle, a can, or a bottle can be used.
The capacity of the container is not particularly limited, and is, for example, 1 to 5000 g, preferably 2 to 4000 g, and more preferably 3 to 3000 g. Among them, in the case of family size, for example, 300 to 5000 g, preferably 300 to 4000 g, more preferably 300 to 3000 g, and in the case of personal size, for example, 10 to 500 g, preferably 20 to 500 g, more preferably. Is 30-500 g.

Further, the food and drink composition for improving endurance of the present invention comprises a unit package form per meal, and the total protein amount including whey protein and leucine in the unit is 1 g or more as a meal intake. The adjusted one is desirable, and specifically, it is prepared so as to be 1 to 40 g, preferably 2 to 20 g, more preferably 3 to 20 g, still more preferably 3 to 15 g, and particularly preferably 3 to 10 g. Here, the term "unit packaging form per meal" means that the amount of intake per meal is predetermined. For example, as a food that can be orally ingested in a specific amount, not only general foods but also beverages. It means forms such as (drinks, etc.), health supplements, foods with health claims, and supplements.
In the "unit packaging form per serving", for example, in the case of liquid beverages, gel-like / paste-like / paste-like jelly, powder-like / granular / capsule-like / block-like solid foods, etc., metal. A form in which a specific amount (dose) can be specified for packaging containers such as cans, glass bottles (bottles, etc.), plastic containers (pet bottles, etc.), packs, pouches, film containers, paper boxes, etc., or intake per serving (usage, dose) ) Can be displayed on the packaging container, homepage, etc. to specify the specific amount. The endurance-enhancing food and drink composition of the present invention is easy to ingest and swallow in any form, such as a bite-sized block or a mini canned beverage that can be ingested in one drink, and is easy for children and the elderly. It is possible to improve the convenience for the person.

As a method for ingesting (administering) the blood leucine concentration increasing promoter or the food and drink composition for improving endurance of the present invention to a subject requiring endurance improvement, oral ingestion, enteral administration, gastric fistula, etc. Therefore, it can be appropriately selected depending on the subject and use, but it is preferably taken orally.

The intake amount of the blood leucine concentration increasing promoter or the food and drink composition for improving endurance of the present invention depends on the sex, age, physiological condition, pharmaceutical form, administration route, administration frequency, active ingredient concentration, etc. of the subject. The content of L-leucine added to the blood leucine concentration increase promoter of the present invention per day for adults is about 20 to 200 mg / kg as a solid content, preferably 30 to 30. It may be about 100 mg / kg. Administration may be divided into, for example, once or several times per day.

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

[Example 1] Production of leucine-supported nanoparticles (leucine-supported nanoparticles) (production of test specimen)
FIG. 1 shows a method for producing a leucine-supported nanowhey.
First, (1) 1.14 g of a green tea extract (trade name: Sanphenon 90S, manufactured by Taiyo Kagaku Co., Ltd.) having a catechin content of 70% was added to 1000 mL of distilled water, and (2) an aqueous catechin solution was prepared by stirring well (2). Catechin concentration 800 mg / L).
Next, (3) 50 g of whey protein (trade name: Enlacto SAT, manufactured by Nippon Shinyaku Co., Ltd.) and 0.5 g of leucine were added, and (4) a well-stirred mixture prepared a leucine-whey protein-catechin aqueous solution.
Further, (5) 0.5 mL of a thickening polysaccharide (trade name: Glyroid 3C, manufactured by DSP Gokyo Food & Chemical Co., Ltd.) prepared in advance with distilled water to a concentration of 1% was added, and (6) An aqueous solution of leucine-whey protein-catechin-thickening polysaccharide was prepared by stirring well [step (A)].

Next, (7) a leucine-supported nanoparticle-containing liquid was obtained by performing nano-izing treatment by heat treatment by holding at 80 ° C. for 30 minutes with stirring [step (B)].

Next, (8) the leusin-supported nanoparticles-containing liquid was spray-dried using a spray dryer (manufactured by Tokyo University of Science) to obtain (9) leucine-supported nanoparticles powder [step (C)]. ..
The conditions for spray drying were an inlet temperature of 160 ° C., an outlet temperature of 60 ° C., a dry air volume of 0.4 m ³ / min, a spray pressure of 50 kPa, and a flow rate of 400 mL / h.
The obtained nanoparticle powder was dispersed in water again, and the average particle size was measured using a zeta potential / nanoparticle size measurement system (“DelsaMax PRO” manufactured by Beckman Coulter Co., Ltd.). The average particle size was 90 nm. Met.

[Comparative Example 1] Production of whey protein-leucine mixture (production of control sample)
The whey protein-leucine mixture is the same as in Example 1 except that (7) no nano-treatment by heat treatment is performed.
That is, first, (1) 1.14 g of a green tea extract (trade name: Sanphenon 90S, manufactured by Taiyo Kagaku Co., Ltd.) having a catechin content of 70% was added to 1000 mL of distilled water, and (2) an aqueous catechin solution was prepared by stirring well. (Catechin concentration 800 mg / L).
Next, (3) 50 g of whey protein (trade name: Enlacto SAT, manufactured by Nippon Shinyaku Co., Ltd.) and 0.5 g of leucine were added, and (4) a well-stirred mixture prepared a leucine-whey protein-catechin aqueous solution.
Next, (5) 0.5 mL of a thickening polysaccharide (trade name: Glyroid 3C, manufactured by DSP Gokyo Food & Chemical Co., Ltd.) prepared in advance with distilled water to a concentration of 1% was added, and (6) Aqueous solution of leucine-whey protein-catechin-thickening polysaccharide was prepared by stirring well.

Then, (8) the leucine-whey protein-catechin-thickening polysaccharide aqueous solution was spray-dried using a spray dryer (manufactured by Tokyo Rika Kikai Co., Ltd.) to obtain a powder of a whey protein-leucine mixture. The conditions for spray drying were an inlet temperature of 160 ° C., an outlet temperature of 60 ° C., a dry air volume of 0.4 m ³ / min, a spray pressure of 50 kPa, and a flow rate of 400 mL / h. The obtained nanoparticle powder was dispersed in water again, and the average particle size was measured using a zeta potential / nanoparticle size measurement system (“DelsaMax PRO” manufactured by Beckman Coulter Co., Ltd.). The average particle size was 350. It was ~ 480 nm.

[Example 2] Measurement of blood leucine concentration after ingestion of leucine-supported nanoparticles (leucine-supported nanowhey) 1.72 g of leucine-supported nanowhey obtained in Example 1 as a test sample, obtained in Comparative Example 1 as a control sample. 1.72 g of a leucine protein-leucine mixture powder (with the same amount of leucine and whey protein as the leucine-supported nanowhey except that it is not nano-sized) was suspended in 21 mL of distilled water, of which 3 mL was used. Was intragastrically administered to 5 SD male rats each 6 weeks old. The dose of each food was 1.06 g / kg in terms of rat body weight. Blood was collected 0.5, 1, 2, 4, 6, 8, and 24 hours after administration, and the plasma leucine concentration was measured.

The results are shown in FIG. In FIG. 2, "whey + leucine" ingested the blood concentration of leucine in rats to which the whey protein-leucine mixture powder was administered as a control sample, and "nano whey + leucine" ingested the leucine-supported nanowhey powder as a test sample. It shows the leucine blood concentration in rats.

From the results shown in FIG. 2, in rats to which the nanowhey + leucine solution was intragastrically administered, the blood leucine concentration was about 2.1 mg / dl before the administration, whereas it was about 4 hours after the administration. After increasing to 9.9 mg / dl, about 2.5 mg / dl 2 hours after administration, about 2.4 mg / dl at 4 hours, about 2.4 mg / dl at 6 hours, and about 2.3 mg at 8 hours. At / dl, at 24 hours, it was maintained at a significantly high level of about 2.5 mg / dl and about 2.3 to 2.5 mg / dl.
On the other hand, in the rat to which the control whey + leucine solution was administered, the blood leucine concentration was about 2.1 mg / dl before the administration, whereas it was about 5.9 mg / dl 0.5 hours after the administration. After increasing to dl, about 2.2 mg / dl 2 hours after administration, about 2.1 mg / dl at 4 hours, about 1.9 mg / dl at 6 hours, about 2.1 mg / dl at 8 hours, At 24 hours, the time was about 1.8 mg / dl, which returned to the same level as before administration or at 4 hours after administration, and remained at a low level thereafter.
In particular, rats to which nanowhey + leucine solution was intragastrically administered had blood leucine concentrations compared to rats to which whey + leucine solution was intragastrically administered at 2, 4, 6, 8, and 24 hours after administration. However, about 13.4% at 2 hours after administration, about 15.0% at 4 hours, about 27.4% at 6 hours, about 9.9% at 8 hours, and about 34.9% at 24 hours. It was kept significantly higher.
From these results, it was found that the leucine-supported nanowhey has an effect of maintaining a high blood leucine concentration for 24 hours after administration, as compared with a whey mixed with leucine.

[Example 3] Measurement of blood leucine concentration after ingestion of leucine-supported nanoparticles (leucine-supported nanowhey) 4-week-old C57BL / 6J male mice were placed in a breeding room in which the dark and light periods were switched every 12 hours. Preliminary breeding for a week. After that, fasting and water cessation were started from 16:00, 3 hours before switching to the dark period, and the chamber with a rotating basket (manufactured by Shin Factory Co., Ltd.) was trained. At 19:00, nanowhey + 1% leucine as a test sample and whey + 1% leucine (8.74 g of each food suspended in 40 mL of distilled water) were used as test samples in mice (7 nanowhey group, 6 whey group). ) Was intragastrically administered so that the dose per body weight was 8.74 g / kg, and energy metabolism and spontaneous momentum (calculated from the cage diameter of 15 cm and the number of rotations) were measured during the dark period of 12 hours. ..

The breath gas analyzer "ARCO-2000" (Arco System Co., Ltd.) was used for the measurement of energy metabolism. The exhaled gas of the mouse filled in the chamber with a rotating cage was aspirated at a rate of 0.3 mL / min, and the oxygen concentration and the carbon dioxide concentration were measured with a connected mass spectrometer. Based on these oxygen concentration and carbon dioxide concentration values, oxygen consumption, respiratory quotient, carbohydrate oxidation amount, and lipid oxidation amount were calculated.

FIG. 3 shows the change over time in oxygen consumption, and FIG. 4 shows the change over time in the respiratory quotient.
Throughout the dark period, nanowhey + 1% leucine intake did not affect oxygen consumption and respiratory quotient compared to whey + 1% leucine, but in the latter half of the dark period (after 300 minutes indicated by the dashed line), the nanowhey group was affected. It was confirmed that the oxygen consumption tended to be high.
The above trend continued for up to 24 hours. Oxygen is used to make chemical energy in the body, but the more energy it consumes, the more energy it makes. Since the larger the amount of energy produced, the longer the body can be moved, it can be evaluated that the increase in oxygen consumption is directly linked to the increase in endurance.

The oxygen consumption from 5 hours to 12 hours (7 hours in total) after administration is shown in FIG. 5, the lipid oxidation amount is shown in FIG. 6, the sugar oxidation amount is shown in FIG. 7, and the total travel distance (integrated value) is shown in FIG. Further, FIG. 9 shows a correlation diagram between the oxygen consumption for 7 hours and the integrated value of the travel distance. Oxygen consumption was significantly higher in nanowhey + 1% leucine intake than in whey + 1% leucine, and the lipid oxidation amount, sugar oxidation amount, and travel distance tended to be higher.
Furthermore, in the nano-whey group, individuals with higher oxygen consumption tended to travel longer distances, whereas in the whey group, such a tendency was not observed (Fig. 9). From these results, it was found that the administration of nanowhey + leucine increased the exercise endurance, and the exercise performance in the latter half was less likely to decrease than the administration of whey + leucine.
The above tendency continued for up to 24 hours.

Based on the above results, leucine-supported nanoparticles (leucine-supported nanoparticles) act as a "promoter for increasing blood leucine concentration" that keeps the blood leucine concentration high for a long period of time, and leucine-supported nanoparticles are an active ingredient. It has been shown that the food and drink composition contained in the above is useful as a "food and drink composition for improving endurance" that enhances endurance.

Claims (6)

Contains nanoparticles containing catechin, whey protein and leucine as active ingredients ,
A blood leucine concentration increase promoter having an average particle size of 40 to 200 nm of the nanoparticles measured by a zeta potential / nanoparticle size measurement system (“DelsaMax PRO” manufactured by Beckman Coulter Co., Ltd.) . The agent for increasing blood leucine concentration according to claim 1, wherein the nanoparticles further contain a polysaccharide . A food or drink composition for improving endurance, which comprises the blood leucine concentration increase promoter according to claim 1 or 2 as an active ingredient. Step (A) of mixing catechin, whey protein, and leucine to prepare a preparation.
The step (B) of obtaining the nanoparticles-containing liquid by heat-treating the preparation liquid obtained in the step (A) to generate nanoparticles, and the step (B) of drying the nanoparticles-containing liquid obtained in the step (B). , Step of obtaining nanoparticles containing nanoparticles (C)
Have,
A method for producing a blood leucine concentration increase accelerator having an average particle size of 40 to 200 nm of the nanoparticles-containing powder measured by a zeta potential / nanoparticle size measurement system (“DelsaMax PRO” manufactured by Beckman Coulter Co., Ltd.). .. The method for producing a blood leucine concentration increase accelerator according to claim 4 , wherein in the step (A), a preparation solution further containing a polysaccharide is prepared. The method for producing a blood leucine concentration increase accelerator according to claim 4 or 5 , wherein the heat treatment is performed at 70 to 98 ° C. for 2 to 180 minutes in the step (B).

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