Peptides, and compositions containing the peptides as active ingredients.

Abstract

The present invention provides: a peptide that can be readily consumed on a daily basis and has various effects but few side effects; and a composition that contains the peptide as an active ingredient.　Provided is a peptide that comprises the amino acid sequence of (a) or (b). (a) The amino acid sequence of SEQ ID NO:1 (Asn-Leu-Pro-Pro-Leu-Thr). (b) An amino acid sequence produced from the amino acid sequence of SEQ ID NO:1 by adding, deleting, or replacing one or more amino acids. Also provided is a composition that contains the peptide as an active ingredient.

Description

[Technical Field]

[0001] The present invention relates to peptides and compositions containing the peptide as an active ingredient. [Background technology]

[0002] In recent years, due to various stressors stemming from diversified living and working environments and increasingly complex interpersonal relationships, many people living in modern society are exposed to a great deal of anxiety and mental stress. Prolonged exposure to such anxiety and mental stress can lead to physical health problems, mental disorders, sleep disorders, learning disabilities, and memory impairments. If such disorders develop, long-term medication with side effects may be necessary; therefore, it is crucial to appropriately alleviate anxiety and mental stress.

[0003] However, using chemically synthesized drugs such as tranquilizers, sleep aids, sleep enhancers, and antidepressants to manage stress on a daily basis is not always appropriate from the perspective of side effects. Therefore, there is a need for ingredients that can be easily taken on a daily basis and have few side effects.

[0004] Here, Patent Document 1 discloses a peptide having an anxiolytic effect, in which tyrosine, phenylalanine, tryptophan, or histidine is adjacent to a hydrophobic amino acid. Furthermore, Patent Document 2 discloses a peptide that improves or treats decreased motivation, depression, or depressive mood disorders or symptoms based thereon, consisting of (i) amino acid sequence LSSTQAQQSY, (ii) amino acid sequence LSSTQAQQSW, or (iii) amino acid sequence LSSTQAQQSF.

[0005] However, because the demand for materials with anti-anxiety, sleep-improving, antidepressant, or learning and / or memory impairment-improving effects is diverse, there is still a demand for new materials. [Prior art documents] [Patent Documents]

[0006] [Patent Document 1] International Publication No. 2010 / 087480 Brochure [Patent Document 2] International Publication No. 2016 / 140277 Brochure [Overview of the project] [Problems that the invention aims to solve]

[0007] Therefore, the main objective of this technology is to provide peptides with various effects that can be easily taken on a daily basis and have few side effects, as well as compositions containing such peptides as active ingredients. [Means for solving the problem]

[0008] As a result of diligent research to solve the aforementioned problems, the present inventors have discovered a peptide (SEQ ID NO: 1, hereinafter also referred to as "peptide NLPPLT") consisting of the amino acid sequence Asn-Leu-Pro-Pro-Leu-Thr, which has an anxiolytic effect, a sleep-improving effect, an antidepressant effect, and an effect of improving learning disabilities and / or memory impairments, and have completed this technology.

[0009] In other words, this technology first provides a peptide consisting of the amino acid sequence of (a) or (b) below. (a) Amino acid sequence described in Sequence ID No. 1 (b) Amino acid sequences in which one or more amino acids are substituted, deleted, or added in the amino acid sequence described in Sequence ID No. 1 The peptides relating to this technology may have an anxiolytic effect, a sleep-improving effect, an antidepressant effect, or an effect that improves learning disabilities and / or memory impairments.

[0010] Furthermore, this technology also provides a composition containing the peptide as an active ingredient. The composition relating to this technology may be food or beverage. The composition relating to this technology may be a pharmaceutical product. In this case, the composition relating to this technology may be used for the treatment, prevention and / or improvement of depression, bipolar disorder, schizophrenia, anxiety disorders, or sleep disorders. [Effects of the Invention]

[0011] This technology provides peptides with various effects that can be easily taken on a daily basis with few side effects, and compositions containing such peptides as active ingredients, because peptide NLPPLT has excellent anti-anxiety, sleep-improving, antidepressant, and learning disability and / or memory impairment-improving effects. The effects described herein are not necessarily limited to those described herein and may include any of the effects described herein. [Modes for carrying out the invention]

[0012] The following describes preferred embodiments for implementing this technology. The embodiments described below are merely examples of typical embodiments of this technology, and this should not be interpreted as narrowing the scope of this technology.

[0013] 1. Peptides The peptide related to this technology has an amino acid sequence represented as Asn-Leu-Pro-Pro-Leu-Thr (SEQ ID NO: 1). In this technology, Asn(N) represents an L-asparagine residue, Leu(L) represents an L-leucine residue, Pro(P) represents an L-proline residue, and Thr(T) represents an L-threonine residue. Furthermore, the peptide relating to this technology has an amino acid sequence in which one or more amino acids are substituted, deleted, or added in the amino acid sequence described in Sequence ID No. 1. In this case, it is preferable that the number of substituted, deleted, or added amino acids is two or less.

[0014] In addition, the peptide according to the present technology may have one or more amino acids substituted, deleted, or added within the range of having any one or more effects selected from an anti-anxiety effect, a sleep improvement effect, an antidepressant effect, and an improvement effect on learning disorder and / or memory disorder. Furthermore, the peptide according to the present technology may be salts of the peptide NLPPLT. Examples of such salts include alkali metals such as potassium and sodium; alkaline earth metals such as calcium and magnesium, etc. Among these, one or more kinds can be appropriately used.

[0015] Regarding the method for producing the peptide NLPPLT, for example, the following methods are exemplified, but it is not particularly limited thereto. Specifically, for example, a method of decomposing a protein or peptide containing an amino acid sequence (SEQ ID NO: 1) represented by Asn-Leu-Pro-Pro-Leu-Thr by hydrolysis or the like and separating and purifying it from the obtained decomposition product; a method of synthesizing the peptide NLPPLT by a chemical synthesis method of a peptide and then separating and purifying the peptide NLPPLT from the obtained synthesis product; a method of extracting from plants, animals, or microorganisms that produce the peptide NLPPLT and peptides containing the same and separating and purifying it from the obtained extract, etc. can be mentioned.

[0016] The peptide according to the present technology can be produced, for example, by appropriately hydrolyzing a protein such as whey protein with an acid, an alkali, an enzyme, or the like. Hereinafter, a method of hydrolyzing a raw material protein with a hydrolytic enzyme to obtain the peptide NLPPLT will be exemplified.

[0017] First, before hydrolyzing the raw material protein with an enzyme, the protein is dissolved, dispersed, or suspended in water. The raw material protein is a protein that structurally contains the peptide according to the present technology, and is not particularly limited as long as the peptide according to the present technology can be generated when appropriately digested with a hydrolytic enzyme. Examples of the protein include those derived from animals, plants, or microorganisms, etc. In the present technology, among these, whey protein that can be obtained in large quantities is preferable.

[0018] The whey protein may be a commercially available product, or it may be a whey protein concentrate or whey protein isolate obtained by separating and purifying whey protein using conventional methods (e.g., ultrafiltration, ion exchange, etc.), or a mixture obtained by mixing these whey proteins with whey, etc. (e.g., whey powder, desalted whey powder, etc.) to adjust the purity of the whey protein.

[0019] In this case, the processing method differs depending on the properties of the raw material protein. If the raw material protein is soluble, it can be dispersed in water or warm water and dissolved. If it is sparingly soluble, the protein can be homogenized by mixing and stirring it in hot water.

[0020] Furthermore, an alkaline agent or an acidic agent may be added to the protein-containing solution to adjust the pH. It is preferable to adjust this pH to the optimal pH of the hydrolytic enzyme used or to a similar level. The alkaline agent or acidic agent is not particularly limited, and any agent acceptable for pharmaceuticals or food and beverages may be used. Examples of alkaline agents include hydroxides such as sodium hydroxide and calcium hydroxide; carbonates such as potassium carbonate, and these may also be alkali metal salts or alkaline earth metal salts. Examples of acidic agents include inorganic acids such as hydrochloric acid and phosphoric acid; organic acids such as citric acid, acetic acid, and formic acid. In this technology, one or more of these may be used as appropriate. Furthermore, it is preferable to heat-sterilize the protein-containing solution at 70-90°C for about 15 seconds to 10 minutes to prevent spoilage due to bacterial contamination.

[0021] Next, a predetermined amount of hydrolytic enzyme is added to the protein-containing solution, and the reaction is carried out at a temperature of approximately 10-85°C for 0.1-48 hours to obtain a hydrolyzed product. At this time, after adding the hydrolytic enzyme, the solution is maintained at an appropriate temperature depending on the type of enzyme, for example, 30 to 60°C, preferably 45 to 55°C, to initiate the hydrolysis of the protein. Furthermore, the hydrolysis reaction time can be adjusted by monitoring the rate of decomposition of the enzymatic reaction and continuing the reaction until a desirable rate of decomposition is reached. The aforementioned hydrolytic enzyme reaction can be stopped, for example, by inactivating the enzyme in the hydrolysis solution, which can be done by conventional heat inactivation treatment. The heating temperature and holding time for the heat inactivation treatment can be appropriately set to conditions that allow for sufficient inactivation, taking into account the thermal stability of the enzyme used, but for example, it can be carried out at a temperature range of 80 to 130°C with a holding time of 30 minutes to 2 seconds.

[0022] The hydrolytic enzyme is not particularly limited, but it is preferably an enzyme capable of hydrolyzing the raw material protein to produce the peptide related to this technology. The hydrolytic enzyme may be used alone or in combination of two or more. When two or more enzymes are used, the enzymatic reactions of each enzyme may be carried out simultaneously or separately. Specifically, as the enzymes that can produce the peptide, it is preferable to use endopeptidase, trypsin, and papain in combination, and it is more preferable to use a mixture of these three enzymes.

[0023] Examples of the endopeptidases include those derived from microorganisms or animals, but specifically, examples include proteases derived from Bacillus subtilus and proteases derived from animal pancreas. Commercially available proteases can be used. Examples of commercially available proteases include proteases derived from Bacillus bacteria such as Protease N Amano (manufactured by Amano Enzyme Co., Ltd.), Bioplase SP-20 (manufactured by Nagase & Co., Ltd.), and Nutrase (manufactured by Novozymes Japan Co., Ltd.); and proteases derived from animal pancreas such as PTN6.0S (manufactured by Novozymes Japan Co., Ltd.).

[0024] Examples of trypsin include PTN6.0S (manufactured by Novozymes Japan), and examples of papain include papain W-40 (manufactured by Amano Enzyme Co., Ltd.) and purified papain (manufactured by Mitsubishi Chemical Corporation).

[0025] The method for calculating the degradation rate of the raw material protein involves measuring the total nitrogen content of the sample using the Kjeldahl method, measuring the formal nitrogen content of the sample using the formal titration method, and then calculating the degradation rate from these measured values ​​using the following formula (1).

[0026]

number

[0027] In this technology, it is preferable to isolate or purify the peptide related to this technology from the hydrolysis solution. The purification of peptides related to this technology can be carried out by appropriately combining methods similar to those normally used for the purification of oligopeptides, such as various chromatography techniques including ion exchange chromatography, adsorption chromatography, reversed-phase chromatography, partition chromatography, and gel filtration chromatography, as well as solvent precipitation, salting out, and partitioning between two liquid phases.

[0028] During the isolation or purification of peptides related to this technology, the fraction containing the target substance can be determined using the various effects described above as indicators. Furthermore, the active components of these fractions can be identified by mass spectrometry.

[0029] Furthermore, the peptides related to this technology can also be produced by chemical synthesis. The chemical synthesis of peptides related to this technology can be carried out by liquid-phase or solid-phase methods commonly used for the synthesis of oligopeptides. The synthesized peptides can be deprotected as needed, and unreacted reagents and by-products can be removed to isolate the peptides related to this technology. Such peptides can be synthesized, for example, using commercially available peptide synthesizers. The acquisition of the target peptide can be confirmed by one or more of the following effects selected from anxiolytic effects, sleep-improving effects, antidepressant effects, and effects that improve learning disabilities and / or memory impairments.

[0030] Peptide NLPPLT has an anxiolytic effect, as shown in Test Example 1 below, and also has an anti-anxiety effect, as shown in Test Example 2 below, as shown in serotonin 5-HT 1A Because it acts through receptor activation, it also has a sleep-improving effect. Furthermore, as shown in Test Example 3 below, it also has an antidepressant effect. Serotonin 5-HT 1A This receptor is the most widely expressed of all serotonin receptors and is involved in behaviors such as sleep, eating, thermoregulation, and anxiety. Therefore, this serotonin 5-HT receptor is important. 1A It is commonly known that activating receptors can produce one or more effects selected from anxiolytic, sleep-improving, and antidepressant effects. Therefore, the peptides related to this technology can be used for anxiolytic, sleep-improving, or antidepressant purposes.

[0031] In addition, the peptide related to this technology enables serotonin 5-HT 1A It is believed that this technology can prevent, improve, or treat various diseases and symptoms related to receptors. Therefore, the peptide related to this technology can be ingested or administered to animals, including humans, to stimulate serotonin 5-HT 1A It can be used as an active ingredient in methods for preventing, improving, and / or treating diseases and symptoms related to receptors. In this specification, "improvement" means a change in the condition; prevention or delay of the worsening of the condition; or reversal, prevention, or delay of the progression of the condition; and "prevention" means preventing or delaying the onset of the condition or symptoms in the area of ​​application, or reducing the risk of the condition or symptoms in the area of ​​application, or maintaining health.

[0032] The aforementioned serotonin 5-HT 1A Examples of diseases and symptoms related to these receptors include depression, bipolar disorder, schizophrenia, anxiety disorders, and sleep disorders.

[0033] Furthermore, as shown in Test Example 3 described later, the peptide NLPPLT has an effect mediated by the activation of AMPA receptors. Therefore, it is considered possible to prevent, improve, or treat various diseases and symptoms related to AMPA receptors using the peptide related to this technology. AMPA receptors are a type of glutamate receptor, widely distributed in the central nervous system, and are known to be significantly involved in memory and learning. Therefore, it is conventionally known that activating these AMPA receptors produces not only antidepressant effects but also improvements in learning disabilities and / or memory impairments. For this reason, the peptide related to this technology can be used for anti-anxiety, sleep improvement, or antidepressant purposes, as well as for improving learning disabilities and / or memory impairments.

[0034] Furthermore, since the peptides related to this technology are highly safe even when ingested over a long period of time, they can also be used as ingredients in non-therapeutic foods and beverages. In this specification, "non-therapeutic purpose" is a concept that does not include medical procedures, i.e., treatments performed on the human body, and includes, for example, cosmetic procedures and health promotion.

[0035] 2. Composition This technology also provides compositions containing the peptide described above as an active ingredient. These compositions can be used as pharmaceuticals such as formulations, food and beverages, animal feed, etc. Furthermore, the peptide can be used in the manufacture of these various compositions.

[0036] The compositions relating to this technology may use any combination of components in addition to the peptide relating to this technology, as needed. These optional components may include any components permitted in pharmaceuticals, food and beverages, or animal feed.

[0037] When using a composition related to this technology in a pharmaceutical product, it can be prepared by adding the peptide related to this technology to a known pharmaceutical product, or by mixing the peptide into the raw materials of a pharmaceutical product to manufacture a new pharmaceutical product. Furthermore, when using the peptides related to this technology in pharmaceuticals, they may be used as is, concentrated, or processed into solid, liquid, granular, or powder form.

[0038] The aforementioned pharmaceutical product can be formulated into any desired dosage form as appropriate, depending on the method of administration, such as oral or parenteral administration. The dosage form is not particularly limited, but in the case of oral administration, it can be formulated into solid preparations such as powders, granules, tablets, lozenges, and capsules; or liquid preparations such as solutions, syrups, suspensions, and emulsions. In the case of parenteral administration, it can be formulated into suppositories, sprays, inhalants, ointments, patches, injections, etc. In this technology, formulation into an oral dosage form is preferred. The formulation can be carried out using known methods as appropriate, depending on the dosage form.

[0039] During formulation, a formulation carrier may be added as appropriate. In addition to the aforementioned pharmaceutical product, ingredients such as excipients, pH adjusters, colorants, and flavorings commonly used in formulation may be used. Furthermore, ingredients that have therapeutic, preventive, and / or ameliorative effects on known or future diseases or symptoms may be used in combination as appropriate.

[0040] Depending on the dosage form, various organic or inorganic carriers can be used as the formulation carrier. Examples of carriers in the case of solid dosage forms include excipients, binders, disintegrants, lubricants, stabilizers, and flavoring / odorizing agents.

[0041] Examples of excipients include sugar derivatives such as lactose, sucrose, glucose, mannitol, and sorbitol; starch derivatives such as corn starch, potato starch, α-starch, dextrin, and carboxymethyl starch; cellulose derivatives such as crystalline cellulose, hydroxypropyl cellulose, hydroxypropyl methylcellulose, carboxymethylcellulose, and carboxymethylcellulose calcium; gum arabic; dextran; pullulan; silicate derivatives such as light anhydrous silicic acid, synthetic aluminum silicate, and magnesium aluminometasilicate; phosphate derivatives such as calcium phosphate; carbonate derivatives such as calcium carbonate; and sulfate derivatives such as calcium sulfate.

[0042] Examples of binders include, in addition to the above-mentioned excipients, gelatin, polyvinylpyrrolidone, macrogol, and the like.

[0043] Examples of disintegrants include, in addition to the above-mentioned excipients, chemically modified starches or cellulose derivatives such as croscarmellose sodium, carboxymethyl starch sodium, and cross-linked polyvinylpyrrolidone.

[0044] Examples of lubricants include talc; stearic acid; metal stearate salts such as calcium stearate and magnesium stearate; colloidal silica; waxes such as pea gum and gayl wax; boric acid; glycol; carboxylic acids such as fumaric acid and adipic acid; sodium carboxylate salts such as sodium benzoate; sulfates such as sodium sulfate; leucine; lauryl sulfates such as sodium lauryl sulfate and magnesium lauryl sulfate; silicic acids such as anhydrous silicic acid and silicic acid hydrate; and starch derivatives.

[0045] Examples of stabilizers include para-hydroxybenzoic acid esters such as methylparaben and propylparaben; alcohols such as chlorobutanol, benzyl alcohol, and phenylethyl alcohol; benzalkonium chloride; acetic anhydride; and sorbic acid.

[0046] Examples of flavoring and odor-modifying agents include sweeteners, acidulants, and flavorings. In the case of liquid formulations for oral administration, examples of carriers used include solvents such as water, and flavoring and odor-masking agents.

[0047] When using the composition according to this technology in food and beverages, it can be prepared by adding the peptide according to this technology to known food and beverages, or by mixing the peptide into the raw materials of food and beverages to produce new food and beverages. Furthermore, when using the peptides related to this technology in food and beverages, they may be used as is, concentrated, or processed into solid, liquid, granular, or powder form.

[0048] Food and beverages include, regardless of form (liquid, paste, solid, powder, etc.), tablets, liquid foods, animal feed (including pet feed), as well as, for example, wheat flour products, instant foods, processed agricultural products, processed marine products, processed livestock products, milk and dairy products, oils and fats, basic seasonings, compound seasonings and foods, frozen foods, confectionery, beverages, and other commercially available foods.

[0049] Examples of dairy products include fermented milk, milk beverages, lactic acid bacteria beverages, sweetened condensed milk, skim milk powder, sweetened milk powder, modified milk powder, cream, cheese, butter, and ice cream. Examples of wheat flour products include bread, macaroni, spaghetti, noodles, cake mix, fried chicken batter, and breadcrumbs. Examples of instant foods include instant noodles, cup noodles, retort / prepared foods, canned prepared foods, microwaveable foods, instant soups / stews, instant miso soup / clear soup, canned soups, freeze-dried foods, and other instant foods. Examples of processed agricultural products include canned agricultural products, canned fruits, jams and marmalades, pickles, boiled beans, dried agricultural products, and cereals (processed grain products). Examples of processed seafood products include canned seafood, fish ham and sausage, processed seafood products, seafood delicacies, and preserved seafood. Examples of processed livestock products include canned and pasteurized livestock products, and meat products such as ham and sausages. Examples of fats and oils include butter, margarine, and vegetable oils. Basic seasonings include, for example, soy sauce, miso, sauces, tomato-based seasonings, mirin, and vinegars, while compound seasonings and foods include cooking mixes, curry bases, sauces, dressings, noodle soup bases, spices, and other compound seasonings. Examples of frozen foods include frozen raw ingredients, partially cooked frozen foods, and pre-cooked frozen foods. Examples of confectionery include caramel, candy, chewing gum, chocolate, cookies, biscuits, cakes, pies, snacks, crackers, Japanese sweets, rice crackers, bean snacks, dessert sweets, and other confectionery. Examples of beverages include carbonated drinks, natural fruit juices, fruit juice drinks, fruit juice-containing soft drinks, fruit pulp drinks, fruit drinks with fruit pulp, vegetable drinks, soy milk, soy milk drinks, coffee drinks, tea drinks, powdered drinks, concentrated drinks, sports drinks, nutritional drinks, alcoholic beverages, and other beverages for enjoyment. Other commercially available foods besides those mentioned above include, for example, baby food, furikake (rice seasoning), and ochazuke nori (rice seasoning).

[0050] Furthermore, the food and beverages defined in this technology can be provided and sold as food and beverages labeled with their health benefits (e.g., for anti-anxiety, sleep improvement, anti-depressant, learning disability and / or memory impairment improvement, etc.).

[0051] In this technology, the act of "displaying" includes all acts that inform consumers of the aforementioned use, and any expression that can evoke or infer the aforementioned use, regardless of the purpose of the display, the content of the display, or the object or medium to which it is displayed, falls under the category of "displaying" in this technology.

[0052] Furthermore, the "display" is preferably made in a way that allows consumers to directly recognize the aforementioned use. Specifically, this includes acts such as transferring, delivering, displaying for transfer or delivery, or importing food and beverage products or product packaging on which the aforementioned use is described; displaying or distributing advertisements, price lists, or transaction documents related to products that describe the aforementioned use; or providing information containing such information by electromagnetic means (e.g., the Internet).

[0053] On the other hand, it is preferable that the displayed content be a display approved by the government or other relevant authority (for example, a display approved based on various systems established by the government and carried out in accordance with such approval). Furthermore, it is preferable to attach such display content to packaging, containers, catalogs, brochures, point-of-purchase (POP) advertising and other promotional materials used at sales sites, and other documents.

[0054] Furthermore, "labeling" also includes labels for health foods, functional foods, foods for the sick, enteral nutrition foods, foods for special dietary uses, health functional foods, foods for specified health uses, foods with functional claims, nutrient functional foods, quasi-drugs, etc. Among these, labels approved by the Consumer Affairs Agency are particularly noteworthy, such as those approved under the Foods for Specified Health Uses system, the Foods with Functional Claims system, and similar systems. More specifically, these include labels for Foods for Specified Health Uses, labels for Foods for Specified Health Uses with Conditions, labels for Foods with Functional Claims, labels indicating an effect on the structure or function of the body, and labels indicating a reduction in disease risk. Typical examples of these include labels for Foods for Specified Health Uses as defined in the Health Promotion Act Enforcement Regulations (Ministry of Health, Labour and Welfare Ordinance No. 86 of April 30, 2003) (especially labels indicating health uses), labels for Foods with Functional Claims as defined in the Food Labeling Act (Act No. 70 of 2013), and similar labels.

[0055] Furthermore, the wording used to make the aforementioned indications is not limited to phrases such as "for anti-anxiety," "for improving sleep," "for antidepressant," or "for improving learning disabilities and / or memory impairment." It goes without saying that any other wording that expresses an anti-anxiety effect, a sleep-improving effect, an antidepressant effect, or an improvement in learning disabilities and / or memory impairment is also included within the scope of this technology. For example, such wording can be used for indications based on various purposes that make consumers aware of an anti-anxiety effect, a sleep-improving effect, an antidepressant effect, or an improvement in learning disabilities and / or memory impairment.

[0056] When using the composition related to this technology in animal feed, it can be prepared by adding the peptide related to this technology to a known feed, or a new feed can be manufactured by mixing the peptide into the raw materials of the feed. Furthermore, when using the peptides related to this technology in animal feed, they may be used as is, concentrated, or processed into solid, liquid, granular, or powder form.

[0057] Examples of raw materials for the aforementioned feed include grains such as corn, wheat, barley, and rye; brans such as wheat bran, wheat bran, rice bran, and defatted rice bran; manufacturing residues such as corn gluten meal and corn jam meal; animal feeds such as skim milk powder, whey, fish meal, and bone meal; yeasts such as brewer's yeast; mineral feeds such as calcium phosphate and calcium carbonate; oils and fats; amino acids; and sugars. Examples of the forms of the aforementioned feed include pet food (e.g., pet food), livestock feed, and fish feed.

[0058] The content or amount of the peptide related to this technology in the composition related to this technology is not particularly limited, but it is preferable to include it in an amount that can be easily taken while efficiently obtaining an anti-anxiety effect or a sleep-improving effect. Furthermore, the administration or ingestion interval and duration of administration or ingestion of the composition relating to this technology are not particularly limited.

[0059] As mentioned above, the peptide related to this technology is serotonin 5-HT 1ABecause it is believed that various diseases and symptoms related to receptors can be prevented, improved, or treated, compositions relating to this technology containing this as an active ingredient can be used, for example, to treat, prevent, and / or improve depression, bipolar disorder, schizophrenia, anxiety disorders, or sleep disorders.

[0060] This technology can also employ the following configurations. [1] Use of the peptide (a) or (b) below in an anxiolytic composition. (a) Amino acid sequence described in Sequence ID No. 1 (b) Amino acid sequences in which one or more amino acids are substituted, deleted, or added in the amino acid sequence described in Sequence ID No. 1 [2] Use of the peptides described in (a) or (b) above for the manufacture of an anxiolytic composition. [3] Use of the peptides described in (a) or (b) above for the treatment, prevention and / or improvement of depression, bipolar disorder, schizophrenia, anxiety disorders, or sleep disorders. [4] A method for treating, preventing and / or improving depression, bipolar disorder, schizophrenia, anxiety neurosis, or sleep disorder, comprising the peptide described in (a) or (b) above as an active ingredient. [5] Use of the peptide described in (a) or (b) above in a sleep-improving composition. [6] Use of the peptides described in (a) or (b) above for the manufacture of a sleep-improving composition. [7] Use of the peptides described in (a) or (b) above in an antidepressant composition. [8] Use of the peptides described in (a) or (b) above for the manufacture of an antidepressant composition. [9] Use of the peptides described in (a) or (b) above in compositions for improving learning disabilities and / or memory impairments.

[10] Use of the peptides of (a) or (b) above for the manufacture of compositions for improving learning disabilities and / or memory impairments.

[11] A method for treating, preventing and / or improving one or more conditions selected from the group consisting of anxiety, sleep disorders, depression, learning disabilities and memory impairment, using the peptides described in (a) or (b) below. (a) Amino acid sequence described in Sequence ID No. 1 (b) Amino acid sequences in which one or more amino acids are substituted, deleted, or added in the amino acid sequence described in Sequence ID No. 1

[12] A method for treating, preventing and / or improving one or more diseases selected from the group consisting of depression, bipolar disorder, schizophrenia, and anxiety disorders, using the peptide described in (a) or (b) below. (a) Amino acid sequence described in Sequence ID No. 1 (b) Amino acid sequences in which one or more amino acids are substituted, deleted, or added in the amino acid sequence described in Sequence ID No. 1 [Examples]

[0061] The technology will be described in more detail below based on the examples provided. The embodiments described below are merely examples of typical embodiments of this technology, and should not be interpreted as narrowing the scope of this technology.

[0062] Manufacturing Example 1: Production of NLPPLT peptide by enzymatic hydrolysis of whey protein One kilogram of commercially available whey protein concentrate was dissolved in nine kilograms of purified water and sterilized in a plate-type sterilizer at 75°C for 15 seconds. Sodium hydroxide was then added to adjust the pH of the solution to 9.0, followed by the addition of a proteolytic enzyme to hydrolyze the solution and obtain a solution containing NLPPLT. The enzyme was then inactivated, the solution was concentrated by conventional methods, dried, and approximately one kilogram of powdered whey protein hydrolysate (hereinafter referred to as "WPH") containing the peptide NLPPLT was obtained.

[0063] Manufacturing Example 2: Chemical Synthesis of Peptide NLPPLT NLPPLT was chemically synthesized by the F-moc method. After deprotection, it was purified by reverse-phase HPLC (Waters 1525) and lyophilized. The resulting peptides were confirmed to have matching molecular weights by LC-MS. I confirmed it.

[0064] Test Example 1: Elevated Cross Maze Test A raised cross maze consisting of two intersecting 25cm x 5cm flat plates (arms) was installed at a height of 50cm from the floor. One arm was a closed arm with a 15cm high transparent enclosure, allowing mice to walk safely without fear of falling. The other arm was an open arm without such an enclosure, and because the surroundings were open, mice that were afraid of heights usually did not like the open arm. However, anxiolytic treatment of mice tended to increase the time they spent in the open arm. Therefore, in the test to confirm the anxiolytic effect, the longer the time mice spent in the open arm, or the more times they entered it, the higher the anxiolytic activity.

[0065] <1> WPH administration trial The inventors of this application first conducted tests on the WPH prepared in Production Example 1.

[0066] <1-1> Test Method Male ddy mice were administered WPH prepared in Production Example 1 at doses of 10 mg, 30 mg, or 100 mg per kg of body weight, respectively, 30 minutes prior to the test. 30 minutes later, the mice were placed on a central platform with intersecting open and closed arms for the test (WPH administration group, n=6). For the first 5 minutes after the start, the cumulative time spent on the open arms, the number of visits to the open arms, and the total number of visits to either arm were recorded. In addition, the same test was conducted on male ddy mice that were not administered the test substance WPH as a control group (control group, n=6).

[0067] Based on the results obtained, the percentage of time spent on the open arm and the percentage of times the open arm was entered were calculated as indicators of anxiolytic activity, and statistical significance was verified using a t-test.

[0068] <1-2> Test Results The test results are shown in Table 1 below.

[0069] [Table 1]

[0070] Administration of WPH increased the time spent on the open arm and the number of times the open arm was entered compared to the control group, confirming that WPH has anxiolytic activity.

[0071] <2> Tests using peptide NLPPLT The present inventors, <1> Based on the results, we focused on the peptide NLPPLT contained in the aforementioned WPH and conducted tests on this peptide as well.

[0072] <2-1> Test Method Male ddy mice were administered the peptide NLPPLT synthesized in Production Example 2 at a dose of 3 μg or 10 μg per kg of body weight 30 minutes prior to the test. 30 minutes later, the mice were placed on a central platform where open and closed arms intersected, and the test was conducted (NLPPLT peptide administration group, n=6). For the first 5 minutes after the start, the cumulative time spent on the open arms, the number of visits to the open arms, and the total number of visits to either arm were recorded. In addition, the same test was conducted on male ddy mice that were not administered the peptide, which is the test substance, as a control (control group, n=6).

[0073] Furthermore, in order to investigate the mechanism of action, in addition to the peptide NLPPLT synthesized in Production Example 2 at 10 μg / kg body weight, serotonin 5-HT was added. 1A A similar test was conducted in male ddy mice that were orally administered 10 mg / kg of the receptor antagonist WAY100135 (Tocris Bioscience) (peptide NLPPLT + serotonin antagonist group, n=5).

[0074] From the obtained results, the percentage of the residence time on the open arm (%) and the percentage of the number of entries into the open arm (%) were calculated as indices of anxiolytic activity, and statistical significance was verified by the Tukey-Kramer method.

[0075] <2-2>Test Results The test results are shown in Tables 2 and 3 below.

[0076]

Table 2

[0077]

Table 3

[0078] Administration of the peptide NLPPLT increased the residence time on the open arm compared to the control, and thus it was confirmed that the peptide NLPPLT has anxiolytic activity.

[0079] On the other hand, when the peptide NLPPLT and an antagonist of the serotonin 5-HT 1A receptor were administered simultaneously, the residence time on the open arm decreased compared to the administration of the peptide alone and reached the same level as the control. Therefore, it was suggested that the action of the peptide NLPPLT is based on the action mediated by the activation of the serotonin 5-HT 1A receptor.

[0080] In addition, since the peptide NLPPLT has an action mediated by the activation of the serotonin 5-HT 1A receptor, it was also suggested that the peptide NLPPLT not only has an anxiolytic action but also may have a sleep-improving action, which is one of the actions based on the activation of the serotonin 5-HT 1A receptor.

[0081] Test Example 2: Open Field Test An open-field test was conducted as a behavioral test used for screening anxiolytics. This test utilizes the fact that administering anxiolytics reduces the proportion of time mice spend in the center of a field (hereinafter referred to as "stay time"). Behavior was observed for 5 minutes 30 minutes after administration. In this open-field test, a circular field consisting of multiple concentric circles with the same center but different radii was used. The circle closest to the center of the field was designated as the center of the field, and the time that a mouse placed in the center of this field spent in the center of the field was measured.

[0082] <1> Tests using peptide NLPPLT The inventors of this application focused on the peptide NLPPLT and conducted tests.

[0083] <2> Test method Male ddy mice were administered the peptide NLPPLT synthesized in Production Example 2 at doses of 10 μg, 30 μg, or 100 μg per kg of body weight, respectively, 30 minutes prior to the experiment. 30 minutes later, the mice were placed in the center of the field for the experiment (NLPPLT peptide administration group, n=5). The duration of stay was recorded for the first 5 minutes after the start of the experiment. In addition, the same test was conducted on male ddy mice that were not administered the peptide test substance as a control (control group, n=5-6).

[0084] Based on the results obtained, the percentage of time spent in the environment was calculated as an indicator of anti-anxiety activity.

[0085] <3> Test results The test results are shown in Table 4 below.

[0086] [Table 4]

[0087] Administration of peptide NLPPLT increased the duration of stay compared to the control group, confirming that peptide NLPPLT has anxiolytic activity.

[0088] Test Example 3: Tail Suspension Test A tail suspension test, used for screening antidepressants, was conducted. Mice were suspended by their tails, creating an inescapable condition. Initially, they would attempt to escape and move around, but over time, they would become desperate and stop moving. In contrast, mice treated with antidepressants showed increased escape behavior and decreased immobility. Therefore, a decrease in immobility was considered indicative of an antidepressant effect. Behavior was observed for 6 minutes 30 minutes after administration.

[0089] <1> WPH administration trial The inventors of this application first conducted tests on the WPH prepared in Manufacturing Example 1.

[0090] <1-1> Test Method Male ddy mice were administered WPH synthesized in Production Example 1 at doses of 1 mg, 10 mg, or 100 mg per kg of body weight, respectively, 30 minutes prior to the test. 30 minutes later, the mice were suspended by their tails, and the test was conducted (WPH administration group, n=9-11). The duration of immobility was recorded for the first 6 minutes after the start of the experiment. In addition, the same test was conducted on male ddy mice that were not administered the peptide test substance as a control (control group, n=9-11).

[0091] Based on the results obtained, the duration (in seconds) of immobility behavior was calculated as an indicator of antidepressant activity.

[0092] <1-2> Test Results The test results are shown in Table 5 below.

[0093] [Table 5]

[0094] The administration of WPH reduced the duration of immobility compared to the control group, confirming that WPH has antidepressant properties.

[0095] <2> Tests using peptide NLPPLT The inventors of this application focused on the peptide NLPPLT and conducted tests.

[0096] <2-1> Test Method Male ddy mice were administered the peptide NLPPLT synthesized in Production Example 2 at doses of 10 μg or 30 μg per kg of body weight, respectively, 30 minutes prior to the experiment. 30 minutes later, the mice were suspended by their tails, and the experiment was conducted (NLPPLT peptide administration group, n=5-6). The duration of immobility was recorded for the first 6 minutes after the start of the experiment. In addition, the same test was conducted on male ddy mice that were not administered the peptide test substance as a control (control group, n=5-6).

[0097] Furthermore, to investigate the mechanism of action, a similar test was conducted in male ddy mice that were orally administered 10 mg / kg of the AMPA receptor antagonist NBQX (Cayman Chemical) in addition to 10 μg / kg body weight of the peptide NLPPLT synthesized in Production Example 2 (peptide NLPPLT + AMPA receptor antagonist administration group, n=10-12).

[0098] Based on the results obtained, the duration (in seconds) of immobility behavior was calculated as an indicator of antidepressant activity.

[0099] <2-2> Test Results The test results are shown in Table 6 below.

[0100] [Table 6]

[0101] Administration of peptide NLPPLT reduced the duration of immobility compared to the control group, confirming that peptide NLPPLT has antidepressant properties.

[0102] Furthermore, the results of the trials using AMPA receptor antagonists are shown in Table 7 below.

[0103] [Table 7]

[0104] When the peptide NLPPLT and an AMPA receptor antagonist were administered simultaneously, the duration of immobility was significantly increased compared to when the peptide was administered alone. Therefore, it was suggested that the effect of peptide NLPPLT is based on its action mediated through the activation of AMPA receptors. AMPA receptors are known to be involved in antidepressant effects, which is consistent with the antidepressant effect of this peptide. Furthermore, since AMPA receptors are known to be involved in memory impairment and learning disabilities, it was also suggested that this peptide may exhibit effects that improve memory impairment and learning disabilities. [Industrial applicability]

[0105] The peptides related to this technology can be used for anti-anxiety, sleep improvement, antidepressant, and improvement of learning disabilities and / or memory impairments. Furthermore, compositions containing these peptides as active ingredients can be used in a wide range of fields, such as pharmaceuticals or food products, for the treatment, prevention, and / or improvement of anxiety symptoms, mental stress, depression, bipolar disorder, schizophrenia, anxiety neurosis, or sleep disorders.

Claims

1. A peptide comprising the amino acid sequence described in Sequence ID No.

1.

2. The peptide according to claim 1, which has an anxiolytic effect.

3. The peptide according to claim 1, which has a sleep-improving effect.

4. The peptide according to claim 1, which has an antidepressant effect.

5. The peptide according to claim 1, which has an effect of improving learning disabilities and / or memory impairments.

6. A composition containing the peptide described in any one of claims 1 to 5 as an active ingredient.

7. The composition according to claim 6, wherein the composition is a food or beverage.

8. The composition according to claim 6, wherein the composition is a pharmaceutical product.

9. The composition according to claim 8, used for the treatment, prevention and / or improvement of depression, bipolar disorder, schizophrenia, anxiety neurosis, or sleep disorder.

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