Powdered preparation for improving brain function, etc., and method for manufacturing the same.

Abstract

This invention provides a stabilized formulation of alariadiol for use as a pharmaceutical or functional food (a formulation for improving brain function, etc.) and a method for producing the same. [Solution] A powder formulation containing alariadiol, dextrin, and cyclodextrin. Preferably further containing high molecular weight polysaccharides.

Description

Technical Field

[0001] The present invention relates to a stabilized powder formulation containing asiaticadiol for use in humans and the like, and a method for producing the same.

Background Art

[0002] Centella asiatica (also known as Gotu Kola) is a herb native to India, Indonesia, China, and Southeast Asia, and is a plant used in Ayurveda, the traditional medicine of India. It is known to have a wide range of effects such as antipyretic, analgesic, anti-inflammatory, and cognitive function improvement. For example, when a Centella asiatica extract is administered to rats, reduction of oxidative stress and improvement of cognitive function are observed, and it has been reported that it is expected as a therapeutic agent for Alzheimer's disease (see, for example, Non-Patent Document 1).

[0003] By the way, in order to develop such an active ingredient as a pharmaceutical or functional food, it is desirable to be able to be stably stored at room temperature for a certain period or more. For example, in order to stabilize vitamins and coenzyme Q formulated in pharmaceuticals and foods, a technique of encapsulating them with cyclodextrin to form a powder composition has been reported (see, for example, Patent Document 1). 10 To stabilize them, a technique of encapsulating them with cyclodextrin to form a powder composition has been reported (see, for example, Patent Document 1).

Prior Art Documents

Non-Patent Documents

[0004]

Non-Patent Document 1

Patent Documents

[0005] [Patent Document 1] Japanese Patent Publication No. 2005-2005 [Overview of the project] [Problems that the invention aims to solve]

[0006] The inventors have discovered that alariadiol, a single compound purified by HPLC from an ethanol extract of gotu kola, is the main active ingredient for the prevention and / or treatment of neurodegenerative diseases, and have already filed an international patent application (PCT / JP2020 / 039361). However, the storage stability of formulations containing alariadiol is still unknown.

[0007] The problems that this invention aims to solve include providing a stabilized formulation (a formulation for improving brain function, etc.) for use as a pharmaceutical or functional food for human use, and a method for producing such a formulation. [Means for solving the problem]

[0008] To solve the above problems, the present inventors conducted diligent research and found that the above problems can be solved by mixing alariadiol with a specific excipient to form a powder formulation. That is, the present invention includes the following embodiments.

[0009] [1] A powder formulation for improving brain function containing alariadiol, dextrin, and cyclodextrin. [2] The powder formulation according to [1], further comprising high molecular weight polysaccharides. [3] The powder formulation according to [1] or [2], wherein cyclodextrin is contained in an amount of 20% by mass or more of the total amount of the powder formulation. [4] A powder formulation according to any one of the following items [1] to [3], wherein the cyclodextrin contains γ-cyclodextrin. [5] The powder formulation according to any one of the items [2] to [4], wherein the high molecular weight polysaccharide is gum arabic or ghati gum. [6] A powder formulation according to any one of the items [1] to [5] for stabilizing alariadiol as an active ingredient. [7] A method for producing alariadiol, comprising the steps of: extracting the plant body of gotu kola with a mixed solvent of an organic solvent and water; passing the extract obtained in the extraction step through a column packed with a hydrophobic synthetic adsorbent to adsorb alariadiol onto the hydrophobic synthetic adsorbent; and eluting the components adsorbed on the hydrophobic synthetic adsorbent with a mixed solvent of an organic solvent and water to obtain an eluted fraction containing alariadiol. [8] A method for producing a powder formulation for improving brain function, comprising the steps of: mixing alariadiol, dextrin and cyclodextrin to obtain a mixture; homogenizing the mixture to prepare an emulsion; and drying the emulsion. [9] A method for producing the powder formulation according to [8], wherein the cyclodextrin comprises γ-cyclodextrin. [Effects of the Invention]

[0010] According to the present invention, it is possible to provide a stabilized formulation (a formulation for improving brain function) for use as a pharmaceutical or functional food, and a method for producing such a formulation. [Brief explanation of the drawing]

[0011] [Figure 1] Figure 1 is a flow chart showing the manufacturing process of alariadiol in one embodiment. [Figure 2] Figure 2 is a flowchart showing the process for manufacturing a powder formulation containing alariadiol. [Figure 3] Figure 3 shows the flowchart for Test Example 5 (Human Monitor Test). The Stroop test has two purposes: "to perform an activity that fatigues the brain" and "to evaluate brain function (concentration, judgment, reaction speed) because good results were obtained." The 15 minutes of the 100-square calculation are broken down as follows: "the first 5 minutes are multiplication to confuse the brain," and the following 10 minutes are addition to evaluate brain function (concentration, calculation ability, judgment, reaction speed). [Modes for carrying out the invention]

[0012] The present invention relates to a powder formulation containing at least alariadiol, dextrin, and cyclodextrin, and more preferably to a powder formulation further containing a high molecular weight polysaccharide. The following describes the components contained in a typical embodiment of the powder formulation.

[0013] (Alariadiol) The formulation of this embodiment is the following formula (I): [ka] The active ingredient is alariadiol (CAS No. 1354638-93-9), represented by formula (I). The compound represented by formula (I) above contains chiral carbon atoms at positions 3 and 8, and the double bond between carbon atoms at positions 9 and 10 has a cis (Z) conformation. Therefore, the active ingredient may be an optical isomer, a racemic mixture, and / or a geometric isomer with respect to these positions. Naturally occurring alariadiol has also been isolated from ethanol extracts of Aralia cordata leaves and has been reported to have inhibitory activity on the proliferation of human breast cancer cells (Cheng WL et al. (2011) Planta Med; 77: 164-168). Alariadiol can be produced from medicinal plants such as Gotu Kola and Aralia cordata by organic solvent extraction and purification by column chromatography.

[0014] Incidentally, the active ingredient may be in the form of a salt or derivative. When the hydroxy group of the compound represented by the formula (I) has moderate acidity, examples of the salt include metal salts such as aluminum, alkali metal salts such as lithium, sodium or potassium, alkaline earth metal salts such as calcium or magnesium, and ammonium or substituted ammonium salts, for example, salts with lower alkylamines such as triethylamine, hydroxyalkylamines such as 2-hydroxyethylamine, bis-(2-hydroxyethyl)-amine or tri-(2-hydroxyethyl)-amine, cycloalkylamines such as bicyclohexylamine, or salts with pyridine-type bases such as procaine, dibenzylpiperidine, N-benzyl-β-phenethylamine, dehydroabietylamine, N,N'-bisdehydroabietylamine, glucamine, N-methylglucamine, or pyridine, collidine, quinene or quinoline. Further, the hydroxy group of the compound represented by the formula (I) may be an acylated, alkylated, phosphorylated or borated derivative. Accordingly, in the present specification, the term "allaradiol" includes the compound represented by the above formula (I), its isomers or its derivatives or salts thereof.

[0015] The content of allaradiol in the preparation of this embodiment is not particularly limited. For example, as long as an effective amount of allaradiol is contained to exhibit a desired effect (such as the effect of preventing or treating a disease or condition presenting a disorder of brain function), for example, when the total preparation is 100% by mass, · The lower limit of the content of allaradiol is preferably 1 ppm (0.0001% by mass) or more, more preferably 10 ppm or more (0.001% by mass), still more preferably 100 ppm (0.01% by mass) or more, · The upper limit of the content of allaradiol is preferably 50% by mass or less, more preferably 40% by mass, still more preferably 30% by mass or less, is satisfied.

[0016] (Dextrin) Dextrin refers to starch that has been partially hydrolyzed, and there are dextrins with various molecular weights depending on the degree of partial hydrolysis of starch. The DE (Dextrose Equivalent) value is commonly used as an indicator to understand the degree of partial hydrolysis of starch (i.e., the molecular weight distribution of dextrin). A high DE value means that the hydrolysis of starch has progressed, the average molecular weight has decreased, and it has properties similar to glucose, while a low DE value means that the degree of hydrolysis is low and it is closer to starch. DE can take values ​​from 0 to 100. In this specification, "DE" is a value obtained by the Wilstetter-Schudel method, and is calculated using the formula [(mass of direct reducing sugar (indicated as glucose)) / (mass of solids)] × 100.

[0017] In this embodiment, dextrin with a DE of 20 or less is preferably used. Using dextrin with a DE of 20 or less contributes to improved dispersibility and stability when it is made into a powder formulation. More preferably, dextrin with a DE of 5 or less is used in the present invention. Dextrin with a DE of 5 or less used in the present invention is commercially available, and examples of commercially available products include corn-derived dextrin with a DE of 4 (Pinex #100: manufactured by Matsutani Chemical Industry Co., Ltd.) and Sandec #30 (DE 2-5: manufactured by Sanwa Starch Industry Co., Ltd.).

[0018] The dextrin content is not particularly limited, but if the total mass of the formulation of this embodiment is 100% by mass, The lower limit of the dextrin content is preferably 10% by mass or more, more preferably 15% by mass, and even more preferably 20% by mass. The upper limit of the dextrin content is preferably 50% by mass or less, more preferably 45% by mass or less, and even more preferably 40% by mass or less. That is the case.

[0019] (Cyclodextrin) Cyclodextrin is a type of cyclic oligosaccharide in which several D-glucose molecules are linked by α(1→4) glucosidic bonds to form a cyclic structure. This cyclodextrin is not particularly limited, and examples include α-cyclodextrin (αCD, a 6-membered sugar ring molecule), β-cyclodextrin (βCD, a 7-membered sugar ring molecule), and γ-cyclodextrin (γCD, an 8-membered sugar ring molecule), but γCD is particularly preferred. This is because it allows for a higher concentration of alariadiol and enables the production of a highly stable powder formulation. Note that only one type may be included, or two or more types may be used in combination.

[0020] Furthermore, cyclodextrins having branched side chains such as glucose and maltose (hereinafter simply referred to as "branched cyclodextrins"), as well as cyclodextrins having substituents such as methyl groups, hydroxypropyl groups, and acetyl groups (hereinafter referred to as "cyclodextrin derivatives"), may be used as the cyclodextrins mentioned above. This is because including these allows for the acquisition of a water-soluble powder formulation. Additionally, cyclodextrin alone may be used, or a mixture with cyclodextrin derivatives may be used, but it is preferable to use cyclodextrin alone.

[0021] The cyclodextrin in the powder formulation of this embodiment acts as both an excipient and a stabilizer of the active ingredient. In this specification, an excipient is an additive used to improve the handling, molding, and administration of a composition. Although not bound by any particular theory, the cyclodextrin's stabilizing effect is thought to be due to its ability to encapsulate the active ingredient, alariadiol, thereby suppressing oxidation caused by heat and light from the external environment, or making it less susceptible to the influence of coexisting substances.

[0022] From the viewpoint of improving the storage stability of alariadiol, the cyclodextrin content in the powder formulation of this embodiment is 20% by mass or more, more preferably 30% by mass or more, and even more preferably 40% by mass or more. The upper limit of the cyclodextrin content is preferably 80% by mass or less, more preferably 70% by mass or less, and even more preferably 60% by mass or less. This is because if the cyclodextrin content is too high, it is difficult to obtain a powder formulation with excellent fluidity.

[0023] (High molecular weight polysaccharide) As high molecular weight polysaccharides, a wide range of natural and synthetic high molecular weight polysaccharides can be selected, such as guar gum, locust bean gum, tara gum, tamarind gum, glucomannan, xanthan gum, soy fiber, corn fiber, okara fiber, gum arabic, ghati gum, pectin, gellan gum, pullulan, curdlan, agar, sodium alginate, cellulose, and carboxymethylcellulose. One or more of these can be selected and used. In particular, in this embodiment, it is more preferable to select and use one or more from gum arabic, ghati gum, carrageenan, guar gum, tamarind gum, tara gum, glucomannan, xanthan gum, locust bean gum, and corn fiber.

[0024] In one embodiment, gum arabic is a gum-like secretion obtained from the trunks and branches of Acacia senegal Willdenow or other related plants (Leguminosae). Gum arabic is composed of approximately 2% protein, including sugars such as galactose, arabinose, rhamnose, glucuronic acid, and 4-O-methylglucuronic acid, and amino acids such as hydroxyproline, proline, and serine. Gum arabic is commercially available, and examples of commercially available products include Gum arabic HP Powder (manufactured by DSP Gokyo Food & Chemical Co., Ltd.).

[0025] Another embodiment of ghati gum is an amorphous, translucent secretion obtained from Anogeissus latifolia, a Combretaceae tree found in the arid deciduous forest areas of India. It is a water-soluble gum containing approximately 3% protein and is an acidic complex polysaccharide composed of l-arabinose, d-galactose, d-glucuronic acid, and other sugars. Gattigum is commercially available, and examples of commercially available products include Gattikol SS (manufactured by Sanei Pharmaceutical Trading Co., Ltd.).

[0026] The content of high molecular weight polysaccharides is not particularly limited, but if the total mass of the formulation of this embodiment is 100% by mass, The lower limit of the content of high molecular weight polysaccharides is preferably 5% by mass or more, more preferably 6% by mass, more preferably 7% by mass, and even more preferably 8% by mass. The upper limit of the content of high molecular weight polysaccharides is preferably 20% by mass or less, more preferably 15% by mass, and even more preferably 12% by mass or less. That is the case.

[0027] (Other additives) Alariadiol, the active ingredient in this embodiment, is oil-soluble and, as is, has poor dispersibility in water, making it difficult to add to food and beverages. Therefore, a solution of alariadiol dissolved in a water-miscible organic solvent can be mixed with oils, surfactants, and other additives to emulsify it, and then dried to produce a powder formulation.

[0028] (1)Oils and fats Oils and fats include avocado oil, linseed oil, almond oil, fennel oil, perilla oil, olive oil, orange oil, orange raffia oil, cocoa butter, chamomile oil, carrot oil, cucumber oil, apricot kernel oil, kukui nut oil, walnut oil, wheat germ oil, sesame oil, rice oil, rice bran oil, camellia oil, safflower oil, salad oil, shea butter, soybean oil, tea oil, evening primrose oil, camellia oil, corn oil, and rapeseed oil. Examples include oils, peach oil, safflower oil, castor oil, sunflower oil, grape seed oil, hazelnut oil, macadamia nut oil, cottonseed oil, meadowfoam oil, peanut oil, rosehip oil, turtle oil, mink oil, egg yolk oil, cocoa butter, palm oil, palm kernel oil, Japanese wax, coconut oil, fish oil, whale oil, shark oil, liver oil, beef tallow, pork tallow, chicken tallow, rabbit tallow, sheep tallow, horse tallow, or hydrogenated versions of these oils and fats or their derivatives.

[0029] (2) Surfactants The surfactant includes, but is not limited to, at least one of sucrose fatty acid esters and polyglycerol fatty acid esters. Both act as surfactants and can reduce the average particle size of the emulsion particles when the emulsion composition is prepared.

[0030] In this embodiment, the sucrose fatty acid ester used is preferably one with 12 or more carbon atoms in the fatty acid, and more preferably one with 12 to 20 carbon atoms, from the viewpoint of surfactant activity. By having 12 or more carbon atoms, it may be possible to produce emulsion particles with a smaller average particle size. Examples of sucrose fatty acid esters include sucrose dioleate, sucrose distearate, sucrose dipalmitate, sucrose dimyristate, sucrose dilaurate, sucrose monooleate, sucrose monostearate, sucrose monopalmitate, sucrose monomyristate, and sucrose monolaurate. Among these, sucrose monoesters are preferred, and sucrose monolaurate and sucrose monooleate are particularly preferred. In the present invention, these sucrose fatty acid esters can be used individually or in combination.

[0031] Examples of commercially available products include Ryoto Sugar Ester S-070, S-170, S-270, S-370, S-370F, S-570, S-770, S-970, S-1170, S-1170F, S-1570, S-1670, P-070, P-170, P-1570, P-1670, M-1695, O-170, O-1570, and DK Ester manufactured by Daiichi Kogyo Seiyaku Co., Ltd. Examples include SS, F160, F140, F110, F90, F70, F50, F-A50, F-20W, F-10, F-A10E, Cosme-like B-30, S-10, S-50, S-70, S-110, S-160, S-190, SA-10, SA-50, P-10, P-160, M-160, L-10, L-50, L-160, L-150A, L-160A, R-10, R-20, O-10, O-150, etc.

[0032] Examples of polyglycerin fatty acid esters used in this embodiment include esters of polyglycerin having an average degree of polymerization of 2 or more, preferably 6 to 15, more preferably 8 to 10, and fatty acids having 8 to 18 carbon atoms, such as caprylic acid, capric acid, lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid, and linoleic acid. Preferred examples of polyglycerin fatty acid esters include hexaglycerin monooleate, hexaglycerin monostearate, hexaglycerin monopalmitate, hexaglycerin monomyristate, hexaglycerin monolaurate, decaglycerin monooleate, decaglycerin monostearate, decaglycerin monopalmitate, decaglycerin monomyristate, and decaglycerin monolaurate.

[0033] Among these, more preferably are decaglycerin monooleate (HLB=12), decaglycerin monostearate (HLB=12), decaglycerin monopalmitate (HLB=13), decaglycerin monomyristate (HLB=14), and decaglycerin monolaurate (HLB=16). These polyglycerin fatty acid esters can be used individually or in combination.

[0034] Examples of commercially available products include NIKKOL DGMS, NIKKOL DGMO-CV, NIKKOL DGMO-90V, and NIKKOL Decaglyn PR-20 from Nikko Chemicals Co., Ltd., Ryoto Polyglycerides L-7D, L-10D, and M-10D from Mitsubishi Chemical Foods Corporation, Sunsoft Q-17UL, Sunsoft Q-14S, and Sunsoft A-141C from Taiyo Kagaku Co., Ltd., and Poem DO-100 and Poem J-0021 from Riken Vitamin Co., Ltd.

[0035] (Powdered preparations and their uses) The "powder formulation" in the present invention and this embodiment includes not only powder, but also, for example, tablets, granules, powders, etc., which are made by molding powder. The composition containing the powder formulation of this embodiment (including the powder formulation itself, capsules containing the powder formulation, etc.) contains at least three components (alariadiol, dextrin, and cyclodextrin) that are effective in exerting the desired effects of alariadiol (such as cognitive function improvement effects) and have storage stability. In a particular embodiment, for example, the powder formulation is: (a) 0.0001 to 50% by mass of alariadiol; (b) 5-50% by mass of dextrin; (c) 10-80% by mass of cyclodextrin; and (d) Optionally, other additives in an amount of 50% by mass or less, Includes.

[0036] To achieve the desired effect, a composition containing the powder formulation of this embodiment (including the powder formulation itself) can be administered to a human being, for example, at a dose of 0.01 to 10 mg / day, more preferably 0.11 to 10 mg / day, of alariadiol. The administration method may be, for example, oral administration, topical administration (including ophthalmic, vaginal, rectal, nasal, and transdermal administration), or parenteral administration. Parenteral administration methods include intravenous injection or drip infusion, subcutaneous, intraperitoneal, or intramuscular injection, pulmonary administration by aspiration or inhalation, intrathecal administration, and intraventricular administration.

[0037] The powder formulation containing alariadiol of this embodiment and the composition containing said powder formulation are useful, for example, for the prevention or treatment of diseases or conditions exhibiting impaired brain function. Examples of diseases or conditions exhibiting such impaired brain function include dementia (e.g., dementia caused by various diseases such as senile dementia, Alzheimer's disease, vascular dementia, post-traumatic dementia, dementia caused by brain tumors, dementia caused by chronic subdural hematoma, dementia caused by normal pressure hydrops, post-meningitis dementia, and Parkinson's disease), non-dementia-related cognitive impairment (e.g., mild cognitive impairment (MCI)), and memory or learning impairment (e.g., memory and learning impairments associated with brain developmental disorders).

[0038] When a composition containing the powder formulation of this embodiment is used as a food or beverage, it can be used in, for example, foods, functional foods, foods for specified health uses, foods for sick people, or supplements that are based on the concept of preventing, improving, or enhancing cognitive decline and, if necessary, display that fact. When manufacturing a supplement using the powder formulation of this embodiment, the powder formulation can be directly compressed into tablets, but additionally, additives commonly used in formulations, such as starch, lactose, sucrose, mannitol, carboxymethylcellulose, corn starch, and inorganic salts, can be added as needed.

[0039] On the other hand, when a composition containing the powder formulation of this embodiment is used as a pharmaceutical product (including quasi-drugs, etc.), it will be administered in any of the permitted dosage forms for drugs containing a therapeutically effective amount of alariadiol and providing similar utility. The therapeutically effective amount will be determined depending on numerous factors, including the severity of the disease to be treated, the age and relative health status of the subject, the route and form of administration, the indications for which the administration is directed, and the preferences and experience of the physicians involved. A person skilled in the art treating such diseases will be able to determine the therapeutically effective amount of alariadiol for a given disease without conducting unnecessary experiments and relying on personal knowledge and the disclosures of this application.

[0040] (Improved brain function) The "improvement of brain function" in the present invention and this embodiment refers to, for example, the observation of improvements in motor control function, which moves the limbs in accordance with commands output from brain nerve cells; physical balance function, which controls the balance of the body; and / or higher brain function (in the present invention, improvement of function includes the recovery or improvement of impaired function, and the suppression of its decline). The following examples (Test Example 5, Human Monitor Test) demonstrate the effect of improving brain function in areas such as improved concentration, improved calculation ability, improved judgment, improved reaction speed, improved concentration after continuous work, improved reaction speed after continuous work, reduced brain fatigue after continuous work, and / or reduced fatigue in the test subjects.

[0041] "Higher brain functions" is a general term for intellectual functions such as understanding, judgment, memory, calculation / learning, problem-solving, execution, and orientation, which are exerted in response to perceptual information obtained from the five senses (sight, hearing, touch, smell, taste), vestibular sense (sense of balance), and proprioception (sense of feeling the intention of the limbs), based on brain functions that have developed normally after birth. "Higher brain functions" include "cognition," which involves actively collecting external information, storing it in recent memory, and processing it by adding reasoning and judgment; "language," which involves explaining this information in words; "memory," which involves creating new memories; "execution," which involves acting with purpose; and "emotion / personality," which involves controlling reason to behave socially. In other words, higher brain functions are a general term for cognitive processes, actions / behaviors, and emotional / psychological functions. Furthermore, "higher brain functions" are broadly synonymous with "cognitive function," which is commonly used when evaluating the degree of impairment in dementia.

[0042] In the present invention and its embodiments, "higher brain dysfunction" refers to a state in which, for example, comprehension, judgment, memory, language comprehension, problem-solving ability, executive ability, orientation, etc., are reduced due to a decline in brain function (such as regression of brain nerve cells or a decrease in nerve conduction velocity).

[0043] The "core symptoms of higher brain dysfunction" in the present invention and this embodiment are symptoms that appear in conjunction with higher brain dysfunction caused by damage to brain nerve cells, and include memory impairment (inability to remember new things or to forget past actions, words, and knowledge), comprehension and judgment impairment (inability to think logically), problem-solving ability impairment (inability to think deeply to solve problems and becoming confused), executive function impairment (inability to make plans or think of procedures), disorientation (inability to know the time and place), apraxia, agnosia, and aphasia (inability to perform actions that combine movements, inability to know how to use familiar objects, inability to know the names of objects), and left-right recognition impairment (inability to recognize the difference between left and right sides of one's own or others' bodies and difficulty in grasping spatial relationships).

[0044] The "peripheral symptoms of higher brain dysfunction" in this invention and its embodiments are secondary symptoms that occur in conjunction with core symptoms, such as behavioral abnormalities (sleep disorders, verbal and physical aggression, wandering, rejection, overeating / pica, unhygienic behavior) and psychological symptoms (anxiety / restlessness, depression / apathy, hallucinations, delusions). As a symptom of mild cognitive impairment (MIC), sleep may become shallow, and individuals may complain of insomnia as a subjective symptom.

[0045] The "exercise control function" of the present invention and this embodiment is, for example, a function that works to make subtle adjustments to muscle strength or maintain muscle balance during exercise, thereby enabling smooth movement and maintaining a stable posture.

[0046] The "body balance function" in the present invention and this embodiment refers to, for example, the function of integrating sensory information (visual, somatosensory, and vestibular) that contributes to postural stability, controlling the body balance that acts on the limb skeletal muscles by inducing postural reflexes, and maintaining posture.

[0047] (Effects other than improved brain function) The effects of using the powder formulation of the present invention and this embodiment on humans, etc. (effects other than improvement of brain function) include, for example, stress reduction, reduction of eye strain, improvement of eye focusing ability, reduction of leg swelling, suppression of facial skin roughness, and suppression of facial loss of firmness. These effects are shown in the following examples (Test Example 5, Human Monitor Test).

[0048] (Method for producing alariadiol and powder formulations containing the same) In another aspect of the present invention, a method for producing alariadiol is provided. Figure 1 is a flow chart showing the process for producing alariadiol. This method includes an extraction step (S01) in which the Gotu Kola plant body is extracted with a mixed solvent of an organic solvent and / or water; a column adsorption step (S02) in which the obtained extract is passed through a column packed with a hydrophobic synthetic adsorbent to adsorb alariadiol thereon; a column elution step (S03) in which the components adsorbed on the hydrophobic synthetic adsorbent are eluted with a mixed solvent of an organic solvent and water to obtain an eluted fraction containing alariadiol; and a concentration step (S04) in which the obtained extract is concentrated.

[0049] The Gotu Kola plant material used in the extraction step (S01) can be extracted from the flowers, inflorescences, fruits, stems, leaves, petioles, branches, branch leaves, rhizomes, roots, seeds, or the whole plant, but other species of the same genus can also be used. The extraction method can be water or an organic solvent (such as an ethanol solution) or a mixture thereof. For example, a 30-90% aqueous ethanol solution, preferably a 50% aqueous ethanol solution, is used as the extraction solvent. In this embodiment, it is preferable to use an ethanol extract using the whole Gotu Kola plant.

[0050] The adsorbent used in the column adsorption step (S02) is not particularly limited as long as it is a hydrophobic synthetic adsorbent capable of adsorbing highly hydrophobic alariadiol, but it is preferable to use an aromatic synthetic adsorbent. A "hydrophobic synthetic adsorbent" is a hydrophobic adsorbent composed of a crosslinked polymer having a porous structure, and has the property of adsorbing various organic substances in solution through physical interaction between the pores and the substance to be adsorbed. The type of hydrophobic synthetic adsorbent is not particularly limited as long as it can achieve the objective of the present invention, and can be appropriately selected, but it is preferable to use an aromatic synthetic adsorbent because it improves the efficiency of impurity removal. An "aromatic synthetic adsorbent" is a synthetic adsorbent in which the crosslinked polymer constituting the adsorbent has an aromatic ring group such as a benzene ring. An example of an aromatic synthetic adsorbent is a porous body made of styrene-divinylbenzene copolymer. Examples of aromatic synthetic adsorbents include Diaion HP20, Diaion HP21, Sepabeads SP825, Sepabeads SP850, Sepabeads SP700, Sepabeads SP270 (all product names of Mitsubishi Chemical Corporation) and Amberlite XAD4, Amberlite XAD16HP, Amberlite XAD1180, Amberlite XAD2000 (all product names of Rohm & Haas).

[0051] A further aspect of the present invention is a method for preparing a powder formulation containing alariadiol, dextrin, and cyclodextrin. Figure 2 is a flow chart showing the steps for producing the powder formulation by this method. This method includes an excipient addition step (S11) in which dextrin, cyclodextrin, etc. are added to an alariadiol solution to obtain a mixture; a homogenization step (S12) in which the mixture obtained in S11 is homogenized to prepare an emulsion; a drying step (S13) in which the emulsion obtained in S12 is dried; and a recovery step (S14) in which the powder formulation after drying is recovered.

[0052] The excipient addition step (S11) can be achieved by mixing the oil phase, in which alariadiol has been added and dissolved, with the aqueous phase, in which excipients such as dextrin and cyclodextrin have been dissolved.

[0053] In the homogenization step (S12), the above mixture is homogenized to prepare an emulsion. The emulsification method is not particularly limited, and an emulsion with excellent stability can be obtained by adding various emulsifiers that have been conventionally used in food and beverages, such as the high molecular weight polysaccharides and surfactants mentioned above, and then emulsifying the mixture using a homomixer, colloid mill, rotary disc homogenizer, high-pressure homogenizer, etc.

[0054] The emulsion obtained in this way is then subjected to drying in a drying step (S13). Any drying method commonly used for this application can be applied to this manufacturing method, including spray drying, freeze drying, vacuum drying, shelf drying, belt drying, and drum drying. Of these, spray drying and freeze drying are preferred from the viewpoint of handling powder.

[0055] The present invention will now be described in more detail with reference to examples, but the present invention is not limited in any way to these examples. In the following examples, the unit % in the numerical values ​​indicating the amount of each component added means mass %. [Examples]

[0056] [Example 1] Preparation of Gotu Kola Extract 6 g of dried and ground Gotu Kola leaves, stems, or whole plants were each mixed with 75 mL of 30-90% ethanol, and extracted at room temperature for 7 days with occasional stirring. The resulting extracts were filtered and used as test samples. Alariadiol in these test samples was analyzed under the following conditions.

[0057] Column: Mightysil RP-18 GP 250-4.6 (5μm) Elutate: CH3CN:0.1% phosphoric acid = 52:48 Flow rate: 1.0mL / min Analysis temperature: 40℃ Detection: UV210nm

[0058] The alariadiol content in gotu kola extract varied depending on the origin and part of the gotu kola plant, but the 50% ethanol extract showed the highest recovery rate. In addition, other components such as phenylpropanoids, flavonoids, and triterpenoids were also extracted simultaneously from the 50% ethanol extract of gotu kola. Under the above HPLC analysis conditions, alariadiol was found to elute at a retention time of approximately 24 minutes. In the following experiment, samples of gotu kola extract purified and stored under various conditions were analyzed by HPLC under the same conditions as above, and the alariadiol concentration in the samples was quantified by comparing it with the peak area of ​​the standard.

[0059] [Example 2] Purification of Gotu Kola extract (Preparation of alariadiol concentrate) 100 g of Gotu Kola whole plant was mixed with 1500 g of 50% ethanol and extracted at room temperature for 7 days. The extract was then filtered to obtain an extract. This extract was placed on a column packed with approximately 500 mL of an aromatic synthetic adsorbent (e.g., Diaion HP20). Other components were eluted with approximately 1500 mL of 50% ethanol, and then alariadiol was eluted with 55% ethanol. The fraction containing alariadiol was concentrated to obtain 100 mL of concentrate. This concentrate contained approximately 750 μg / mL of alariadiol. The concentrate was evaporated to dryness by evaporating the ethanol and water in a 50°C water bath, and then dissolved again in ethanol. The alariadiol peak was measured by HPLC, and the concentration of alariadiol was found to be 296.2 μg / mL.

[0060] [Test Example 1] Measurement of storage stability of Gotu Kola extract The Gotu Kola extract obtained in Example 1 was further concentrated and dried into a powder, which was then placed in a resealable bag, heat-sealed, and stored. Samples (the sealed powder) stored for 1 month and 3 months at -20°C and 40°C were dissolved in ethanol and analyzed by HPLC under the same conditions as in Example 1. The amount of alariadiol remaining in the samples after 1 month and 3 months from the start of storage is shown in Table 1 below, with the amount of alariadiol in the sample immediately after the start of storage (day 0) set to 100. In Table 1, "-20°C" indicates the group in which the sample was stored at -20°C, and "40°C" indicates the group in which the sample was stored at 40°C.

[0061] [Table 1] As shown in Table 1, the amount of alariadiol in the sample after 3 months of storage at 40°C decreased to 79.3. It is thought that the decomposition of alariadiol is accelerated at higher storage temperatures.

[0062] [Test Example 2] Accelerated Degradation Test of Alariadiol Concentrate, Part 1 An accelerated degradation test at 40°C was conducted using the following compositions (Powder 1, Powder 2). The test results are shown in Table 2 below.

[0063] Powder 1: Powder 1 is obtained by mixing 0.1% by mass of the alaria diol concentrate obtained in Example 2 with 99.9% by mass of highly branched chain dextrin (product name: cluster dextrin, manufactured by Nippon Shokuhin Kako Co., Ltd.). Powder 2: Powder 2 is obtained by mixing 0.1% by mass of the alaria diol concentrate obtained in Example 2 with 99.9% by mass of starch hydrolysate (Pinex #100 (DE3.8), manufactured by Matsutani Chemical Industry Co., Ltd.).

[0064] In Table 2, the amount of alariadiol in the sample immediately after the start of storage (day 0) was set to 100. [Table 2] As shown in Table 2 (values ​​of 17.2 and 19.3 after one month from the start of storage), simply adding common excipients using conventional technology, such as highly branched cyclic dextrin or starch hydrolysates, was not enough to solve the problem of improving the storage stability of alariadiol.

[0065] [Test Example 3] Accelerated degradation test of purified alariadiol, part 2 The following Table 3 shows the results of accelerated degradation tests conducted at 60°C using the compositions listed below (Composition 1, Composition 2, and Composition 3). In Table 3, the amount of alariadiol in the sample immediately after the start of storage (day 0) is set to 100 (100%).

[0066] • Composition 1: A mixture of the alariadiol concentrate (0.1% by mass) obtained in Example 2 and Dexypearl SD-20 (99.9% by mass). This Dexypearl SD-20 (Shiosuiko Sugar Refining Co., Ltd.) is a product containing cyclodextrin and dextrin. • Composition 2: A mixture of alariadiol concentrate (0.1% by mass) obtained in Example 2 and cyclodextrin-β (99.9% by mass). The cyclodextrin-β is "Product name: Celldex B-100, manufactured by Nippon Shokuhin Kako Co., Ltd." • Composition 3: A mixture of alaria diol concentrate (0.1% by mass) obtained in Example 2 and olive oil (99.9% by mass). The olive oil is manufactured by Kanto Chemical Co., Ltd.

[0067] This 60°C accelerated degradation test is based on the Arrhenius equation and assumes the following (Shigeyuki Sakagami, Akito Kawase: Storage Stability Test of Pharmaceuticals, SCAS NEWS, 2000-1, pp. 7-11). • 14 days after the start of storage: In the accelerated degradation test at 40°C, assuming 2 years from the start of storage.

[0068] [Table 3]

[0069] As shown in Table 3, when a mixture of cyclodextrin and dextrin was used as an excipient, the amount of alariadiol remaining was 90% or more in the group that had been stored for 14 days compared to the group that had been stored for 0 days (90% or more after 2 years at 40°C).

[0070] [Example 3] Preparation of Formulation 1 A powder formulation was prepared using the alariadiol concentrate obtained in Example 2 and various excipients. First, predetermined amounts of the aqueous components of polyglycerin fatty acid ester, sucrose fatty acid ester, cyclodextrin, gum arabic, and dextrin shown in Table 4 were mixed and dissolved in purified water heated to 85°C. Next, predetermined amounts of the alariadiol concentrate, rice oil, and tocopherol shown in Table 4 were mixed and dissolved by heating to 50°C. The aqueous and oily component mixtures prepared in this way were mixed and diluted with water to approximately 100 mL, and then homogenized using a precision emulsification and dispersion machine, Creamix (manufactured by M-Technique Co., Ltd.). The supernatant obtained by centrifugation at 13,000 rpm for 5 minutes was concentrated until water was removed, and then freeze-dried using a freeze-dryer (FDU-2100, Tokyo Rikakikai Co., Ltd.). The following day, the freeze-dried product was pulverized using a lab mill and stored in a resealable bag under predetermined conditions.

[0071] [Table 4]

[0072] [Example 4] Preparation of Formulation 2 Formulation 2, with the composition shown in Table 5 below, was prepared in the same manner as in Example 3, except that ghatti gum was used instead of gum arabic.

[0073] [Table 5]

[0074] [Example 5] Preparation of Formulation 3 Formulation 3, with the composition shown in Table 6 below, was prepared using the same method as in Example 3.

[0075] [Table 6]

[0076] The product names and manufacturers of the various excipient components used in Examples 3 to 5 are shown in Table 7 below. [Table 7]

[0077] [Test Example 4] The results of accelerated degradation tests conducted using formulations 1-3, manufactured in Examples 3-5, in the same manner as in Test Example 3 are shown in Table 8 below.

[0078] This 60°C accelerated degradation test was conducted based on the Arrhenius equation, under the following assumptions (Shigeyuki Sakagami, Akito Kawase: Storage Stability Tests for Pharmaceuticals, SCAS NEWS 2000-1 p.7-11). Storage was started at day 0 (indicated as day 0 in Table 8). • 7 days from the start of storage: In the accelerated degradation test at 40°C, it is assumed that the storage period is 1 year from the start of storage. In Table 8, it is indicated as 7 days. • 14 days from the start of storage: In the accelerated degradation test at 40°C, it is assumed to be 2 years from the start of storage. In Table 8, it is indicated as 14 days.

[0079] In Table 8, the amount of alariadiol in the sample immediately after the start of storage (day 0) was set to 100. In Table 8, the group using Formulation 1 as the sample was designated "Formulation 1," the group using Formulation 2 as the sample was designated "Formulation 2," and the group using Formulation 3 as the sample was designated "Formulation 3."

[0080] [Table 8]

[0081] As shown in Table 8, in all groups, more than 90% of the alariadiol remained on day 14 compared to day 0.

[0082] [Example Test 5]: Human Monitor Test A human monitoring study was conducted using Formulation 3, which was manufactured in Example 5. The flowchart of the human monitoring study is shown in Figure 3. The study participants were 29 healthy men and women aged 20 to 50.

[0083] (Information on oral intake by test subjects, etc.) (1) Test food Formulation 3: Composition with the composition described in Table 6 (2) Active ingredient Alariadiol, etc. (3) Examination period Oral intake period: 6 weeks (4) Dosage and Administration The test subjects ingested 100 mg of Formulation 3 (powder) per day. (5) Exam The following tests, as shown in Figure 3, were conducted on the test subjects before taking Formulation 3 (at day 0) and 6 weeks after starting to take Formulation 3 (at day 42). (Test items, etc.) • Item 1 of the test: A 5-minute Stroop task using color words (New Stroop Test 2, Yuji Hakoda, Megumi Watanabe, Toyo Physical Co., Ltd.). This task was performed to put a load on the test subjects (cause brain fatigue), and the number of correct answers performed by the test subjects was measured. • Test item 2: 15-minute 100-square calculation task: In this example test (Test 5), the test subjects performed single-digit multiplication for 5 minutes to induce mental confusion, followed by single-digit addition for 5 minutes and double-digit addition for 5 minutes. The total number of answers and correct answers for each task performed by the test subjects were measured. • Test item 3: 4 sessions of 5 minutes each (total 20 minutes) of computer typing: Test subjects typed random tasks. The "average number of characters typed, average number of correct typed characters, and the trend of the average number of characters typed over the 1st to 4th sessions" were measured for the tasks performed by the test subjects. • Test item 4: Chaldor Fatigue Scale (CFS, subjective evaluation): This is a subjective evaluation using the method described in the paper (Development of a fatigue scale, Volume 37, Issue 2, February 1993, Pages 147-153) and the paper (Journal of the Japanese Society of Oral Examination, Vol. 6, No. 1, pp. 15 to 22, 2014, Table 3, etc.). The test subjects selected one of four options for each of the following 14 questions related to fatigue. The total score (0-42 points) for the responses to these 14 items indicates the degree of fatigue of the test subject, with a higher total score indicating a higher degree of fatigue. • 14 items related to fatigue: 1. What problems do you experience due to fatigue (feeling tired)? 2. Do you need to rest more? 3. Do you feel sleepy or drowsy? 4. Do you have any problems starting things? 5. Do you ever find yourself starting something without difficulty but losing motivation as you continue? 6. What do you feel you lack in energy? 7. Do you feel a decline in your abilities? 8. Do you feel weak or frail? 9. What do you find difficult to concentrate on? 10. What do you do when you have trouble organizing your thoughts? 11. Do you ever get tongue slips when you try to speak? 12. What do you do when you try to speak but can't find the right words? 13. How is your memory? 14. Have you ever lost interest in the things you used to do? • Fatigue level investigation using VAS for test item 5: This method is based on the literature (Fatigue Sensitivity VAS Test Method published on the website of the Japanese Society for Fatigue Research), and the fatigue level felt after the test is indicated by an "X" on a straight line, referencing the sensations shown at both ends of a 10 cm line. The left end of the line represents the best feeling, where the subject feels no fatigue at all, something they have never experienced before, while the right end represents the worst feeling, where the subject is so exhausted that they are unable to do anything, something they have never experienced before. The subject's fatigue level (length from the left end to the "X" mark) was measured using this method. • Test item 6: A prescribed questionnaire (VAS evaluation) was used to evaluate daily stress, eye strain and focus problems, leg swelling, facial skin problems, and loss of facial firmness using a VAS scale. For each question, participants were asked to indicate their sensations with an "X" on a straight line, referring to the sensations shown at both ends of a 10 cm line. The left end of the line represented the worst sensation ever experienced, and the right end represented the best sensation, no sensation at all, also never experienced. This method was used to measure the sensations (length of the drawn line) of the test subjects.

[0084] (Test results) The test results are shown in Table 9. The results for the Stroop task (Test Item 1) show the average value calculated from the measured values ​​(N=29 test subjects). For all tasks, an increase in values ​​was observed at day 42 of intake compared to before intake (day 0). This suggests improvements in the test subjects' concentration, judgment, and / or reaction speed.

[0085] The results of the 100-square calculation task in Test Item 2 were calculated as follows: Each value for the test subjects (N=29) before intake (day 0) was set to 100%, and each value (relative value) at day 42 of intake was calculated. The values ​​at day 42 of intake shown in Table 9 are the average values ​​calculated from these relative values. An increase in values ​​was observed at day 42 of intake compared to before intake (day 0) for all items. This observation suggests improvements in the test subjects' concentration, calculation ability, judgment, and / or reaction speed.

[0086] The average number of characters typed and the average number of correct typed characters for the first to fourth attempts in the computer typing task (Test Item 3) were calculated as follows. The values ​​for each item before intake (day 0) for the test subjects (N=29) were set to 100%, and the relative values ​​for each item at day 42 were calculated. The values ​​at day 42 shown in Table 9 are the average values ​​calculated from these relative values. For all items, an increase in values ​​was observed at day 42 compared to before intake (day 0). Furthermore, the average number of characters typed increased from the first to the third attempt before intake (day 0), but decreased in the fourth attempt compared to the third attempt. This is thought to be due to fatigue or decreased concentration. On the other hand, at day 42, the average number of characters typed in the fourth attempt increased compared to the third attempt. This suggests an improvement in concentration after continuous work, an improvement in reaction speed after continuous work, and / or a reduction in brain fatigue after continuous work in the test subjects.

[0087] The results of the fatigue assessment using Test Item 4 (CFS) and Test Item 5 (VAS) show the average values ​​calculated from the measured values ​​(N=29 test subjects). For both items, a decrease in values ​​was observed at 42 days after ingestion compared to before ingestion (day 0). This observation suggests a reduction in fatigue among the test subjects.

[0088] In the prescribed questionnaire (VAS evaluation) for test item 6, the change is expressed as the difference between the measured value at day 42 of intake and the measured value at day 0 before intake for each test subject (N=29), and the average value is shown. Participants experienced a reduction in daily stress, a reduction in eye strain and focus problems, a reduction in leg swelling, a reduction in facial skin roughness, and a reduction in facial loss of firmness.

[0089] [Table 9]

[0090] Furthermore, Table 10 shows the results of tests conducted 14 days after discontinuing the intake, following 42 days of intake. In Table 10, the results before intake (day 0) and at 42 days after intake are the same as those in Table 9. In Table 10, "St" is the result of Student's t-test, and "Wi" is the result of Wilcoxon's two-sample test. In the results at 14 days after discontinuation shown in Table 10, "St" is the result of Student's t-test comparing with the results at 42 days after intake, and "Wi" is the result of Wilcoxon's two-sample test comparing with the results at 42 days after intake. Compared to 42 days after intake, in some test items (test items 4 to 6, etc.), at 14 days after discontinuation, for example, test subjects were more likely to feel fatigued (test items 4 and 5), and test subjects were more likely to feel daily stress (test item 6).

[0091] [Table 10]

[0092] Although embodiments of the present invention (including examples) have been described above with reference to the drawings, the specific configuration of the present invention is not limited thereto, and any design changes, etc., that do not depart from the spirit of the present invention are still included. [Industrial applicability]

[0093] Because the powder formulation of the present invention contains stabilized alariadiol (it is presumed to remain stable even after being stored at room temperature for more than one year) and exhibits effects such as improving brain function, it may be possible to use it as a pharmaceutical product containing the powder formulation, or as a functional food product (such as a supplement) containing the powder formulation.

Claims

1. A powder formulation for improving brain function, containing alariadiol, dextrin, and cyclodextrin.

2. The powder formulation according to claim 1, further comprising high molecular weight polysaccharides.

3. The powder formulation according to claim 1 or 2, wherein the cyclodextrin is contained in an amount of 20% by mass or more relative to the total amount of the powder formulation.

4. The powder formulation according to claim 1 or 2, wherein the cyclodextrin comprises γ-cyclodextrin.

5. The powder formulation according to claim 2, wherein the high molecular weight polysaccharide is gum arabic or ghati gum.

6. A powder formulation according to claim 1 or 2 for stabilizing alariadiol as an active ingredient.

7. A step of mixing alariadiol, dextrin, and cyclodextrin to obtain a mixture, A step of homogenizing the aforementioned mixture to prepare an emulsified solution, The process of drying the emulsified liquid, A method for producing a powder formulation for improving brain function, including [the specified ingredient].

8. The method for producing a powder formulation according to claim 8, wherein the cyclodextrin comprises γ-cyclodextrin.

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