JP6782956B2 - Absorption enhancer - Google Patents

Description

The present invention comprises an absorption enhancer, an endocytosis promoter, a receptor-independent endocytosis promoter, and a pharmaceutical composition containing the absorption promoter, which comprises an extract of Atemoya, cosmetics, and foods. It is about.

The cell absorbs nutrients into the cell through the cell membrane, which is a lipid bilayer structure that separates the inside and the outside, and maintains vital activity.

Endocytosis is a cell membrane function in which cells take up extracellular substances, and is classified in detail according to the type and size of the substances taken up, the differences in biomolecules involved, and the like. For example, clathrin-dependent endocytosis, caveolae-dependent endocytosis, clathrin-independent endocytosis, phagocytosis (phagocytosis), pinocytosis (pinocytosis), macropinocytosis, etc. are known. ing.

When these are classified according to whether or not stimulation of the receptors present on the cell membrane is necessary, they can be broadly classified into receptor-dependent endocytosis and receptor-independent endocytosis. Receptor-dependent endocytosis is a mechanism that includes clathrin-dependent endocytosis, caveolae-dependent endocytosis, etc., forms a structure called coated vesicles, and selectively takes up a specific recognized substance. .. On the other hand, receptor-independent endocytosis includes clathrin-independent endocytosis, phagocytosis (phagocytosis), pinocytosis (pinocytosis), macropinocytosis, etc., and forms coated vesicles. It is a mechanism for non-selective uptake of extracellular substances (Patent Document 1, Non-Patent Document 1).

The mechanism for incorporating the medicinal ingredient into the cell by utilizing this receptor-independent endocytosis is a composition for topical treatment of the eye containing a selective anticancer agent (Patent Document 2), triamcinolone acetonide and hyaluronic acid. (Patent Document 3), a polynucleotide having a bioreversible group (Patent Document 4), and the like are already known. In addition, a mechanism for more selective delivery of medicinal ingredients to cells by devising nanocarriers has been attempted for some time, and an anticancer drug delivery system of cationic polysaccharide copolymer (Patent Document 5) is given as an example. Be done.

In addition to these, there are attempts to enhance the receptor-independent endocytosis activity of cells to further increase the uptake of medicinal ingredients. It has been reported that the induction of macropinocytosis by gene transfer promoted the uptake of anticancer agents and promoted the death of cancer cells (Non-Patent Document 2). Therefore, in order to provide a drug having a higher medicinal effect, it is desired to develop a substance having an effect of promoting receptor-independent endocytosis of cells.

Atemoya (scientific name: Annona atemoya) is a plant belonging to the family Annonaceae and the genus Annona. It is known that it has a saccharification reaction inhibitory effect (Patent Document 8). However, nothing is known about the receptor-independent endocytosis-promoting effect.

JP 2013-139391 JP-A-6-157346 JP 2012-102141 Special table 2015-52973 JP 2014-001198 JP 2011-335494 JP-A-2009-269842 JP 2015-160843

Yoichiro Fujioka, Biochemistry Vol. 87, No. 1, pp. 91-100 (2015) I. Nakase et. al. , Sci. Rep. , Jun 3, 5, (2015)

The present invention is to provide an absorption enhancer capable of enhancing the uptake of drugs and nutritional components into cells.

As a result of diligent studies aimed at solving the above problems, the present inventors have found that the extract of Atemoya has an excellent absorption promoting action, and have completed the present invention.

That is, the present invention includes the following inventions.
(1) An absorption enhancer containing an extract of Atemoya.
(2) An endocytosis promoter characterized by containing an extract of Atemoya.
(3) A receptor-independent endocytosis promoter containing an extract of Atemoya.
(4) A pharmaceutical composition containing the agent according to any one of (1) to (3).
(5) Cosmetics containing the agent according to any one of (1) to (3).
(6) A food containing the agent according to any one of (1) to (3).

The absorption-promoting agent of the present invention contains an extract of Atemoya as an active ingredient, and when administered or applied in combination with other drugs or nutritional components, cells of the same drug or nutritional component through the cell membrane. It has the effect of increasing the absorption efficiency into the inside. Therefore, this absorption enhancer enhances the uptake of the functioning drug through the cell membrane into cells and enhances its effect. In addition, the absorption promoter of the present invention is also expected to have an effect of improving the nutritional status of non-lesioned tissues including skin by increasing the uptake of nutritional components of cells. Since the absorption promoter of the present invention contains a plant extract having a mild action as an active ingredient, it has no side effects and is highly safe. Therefore, it can be safely used in pharmaceuticals, cosmetics or foods.

Atemoya (scientific name: Annona atemoya) used in the present invention is a plant belonging to the genus Annona genus of the Annonaceae family, and is an artificially improved variety of Annona squamosa and Cherimoya (Annona squamosa). It is a fruit tree.

The Atemoya extract used in the present invention is an extract from a part or whole plant of a plant such as leaves, stems, bark, flowers, fruits and roots of Atemoya. Preferably, it is extracted from fruits, flesh, peels and seeds. The extraction method is not particularly limited, and for example, it may be extracted by heating or may be extracted at room temperature. In addition, raw Atemoya may be used, or dried one may be used.

The method for extracting Atemoya of the present invention is not particularly limited, and may be, for example, heat-extracted or extracted at room temperature or low temperature. Examples of the solvent to be extracted include water, lower alcohols (methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, etc.) and liquid polyhydric alcohols (1,3-butylene glycol, propylene). Glycol, glycerin, etc.), ketones (acetone, methyl ethyl ketone, etc.), acetonitrile, esters (ethyl acetate, butyl acetate, etc.), hydrocarbons (hexane, heptane, liquid paraffin, etc.), ethers (ethyl ether, tetrahydrofuran, propyl, etc.) Ether, etc.). Polar solvents such as water, lower alcohols and liquid polyhydric alcohols are preferred, and water, ethanol, 1,3-butylene glycol and propylene glycol are particularly preferred. These solvents may be used alone or in admixture of two or more.

The above extract may be used as it is in the extracted solution, or may be used after being concentrated, diluted, filtered, decolorized with activated carbon or the like, deodorized, or the like, if necessary. Further, the extracted solution may be subjected to treatments such as concentrated drying, spray drying, freeze-drying and used as a dried product, or may be used after column purification and the like to concentrate or isolate the active ingredient. Is also good. Atemoya used in the present invention is a naturally occurring plant, and the component extracted from Atemoya is a mixture in which a large number of compounds having various structures are present at the same time. Therefore, it is difficult to clarify all the structures or properties of the contained components, and it is preferable to treat them as an extract.

As the pharmaceutical composition, cosmetics, and foods of the present invention, the extract of Atemoya may be used as it is, and fats and oils, waxes, which are components used in cosmetics, pharmaceuticals, foods, etc., within a range that does not impair these effects. Kinds, hydrocarbons, fatty acids, alcohols, esters, surfactants, metal soaps, pH adjusters, preservatives, fragrances, moisturizers, powders, UV absorbers, thickeners, pigments, antioxidants , Whitening agents, chelating agents, excipients, film agents, sweeteners and the like can also be contained.

The present invention can be used in pharmaceuticals, cosmetics, foods, etc., and the dosage forms thereof include, for example, powders, rounds, tablets, granules, capsules, troches, liquids (tinki agents, drink extracts, alcoholic beverages). Includes agents, suspending agents, limonade agents, etc.), ointments, poultices, pastes, plasters, essences, injections, suppositories, emulsions, lotions, creams, massage creams, emulsions, gels, aerosols, Packs, cleaning agents, bathing agents, foundations, dusting, lipsticks, breads, noodles, confectionery, dairy products, processed marine and livestock foods, oils and fats processed foods, seasonings, various beverages (soft beverages, carbonated beverages, beauty beverages) Drinks, nutritional beverages, fruit beverages, dairy beverages, etc.) and the like.

The content of the Atemoya extract used in the present invention varies depending on the age, body weight, symptoms, therapeutic effect, administration method, treatment time, etc. for internal use, but is usually the daily amount per adult. In terms of dry matter, 5 mg or more is preferable, and 10 mg to 5 g is more preferable. Further, 100 mg to 1 g is most preferable. On the other hand, in the case of external use, 0.0001% by weight or more is preferable, and 0.001 to 10% by weight is more preferable in terms of solid matter with respect to the total amount. Further, 0.01 to 5% by weight is most preferable. If it is less than 0.0001% by weight, it is difficult to expect a sufficient effect. If it is contained in an amount exceeding 10% by weight, the enhancement of the effect is hardly recognized and it is uneconomical.

Next, in order to explain the present invention in detail, specific examples will be given. These examples specifically explain the effect and do not limit the scope of the invention. The content in the examples is% by weight.

An extract of Atemoya was produced as follows.

Production Example 1 Hot water extract of Atemoya peel Add 400 mL of purified water to 20 g of Atemoya (dried peel), extract at 90-100 ° C for 2 hours, filter, concentrate the filtrate, freeze-dry and Atemoya. 5.4 g of a hot water extract of the peel was obtained.

Production Example 2 50% ethanol extract of Atemoya peel Add 400 mL of 50% ethanol aqueous solution to 20 g of Atemoya (dried peel), extract at room temperature for 1 week, filter, concentrate the filtrate under reduced pressure, freeze-dry. 5.5 g of a 50% ethanol extract of Atemoya peel was obtained.

Production Example 3 Ethanol extract of Atemoya peel Add 400 mL of ethanol to 20 g of Atemoya (dried peel), extract at room temperature for 1 week, filter, concentrate the filtrate under reduced pressure, freeze-dry and extract ethanol from Atemoya peel. 0.6 g of the product was obtained.

Production Example 4 Hot water extract of Atemoya fruit Add 400 mL of purified water to 20 g of Atemoya (dried fruit), extract at 90-100 ° C for 2 hours, filter, concentrate the filtrate, freeze-dry and dry Atemoya. 9.2 g of hot water extract of fruit was obtained.

Production Example 5 Hot water extract of Atemoya seeds Add 400 mL of purified water to 20 g of Atemoya (seed), extract at 90-100 ° C. for 2 hours, filter, concentrate the filtrate, freeze-dry and heat the Atemoya seeds. 1.5 g of water extract was obtained.

An experiment to evaluate the effect of Atemoya extract was conducted as follows.

Experimental Example 1 Evaluation of absorption-promoting effect on fibroblasts The effect of the Atemoya extract produced in Example 1 (Production Examples 1 to 5) on the absorption of drugs and nutrients through the cell membrane was examined intracellularly in Neutral Red. The amount of uptake was used as an index for evaluation. Neutral red is one of the staining pigments taken up by living cells via the cell membrane. The specific method will be described below.

Human skin fibroblasts (NB1RGB) were seeded 2 × 10 ³ per 1well in 96well plates, in DMEM culture medium containing 10% FBS, 37 ° C., and cultured for 4 days under 5% CO _2. Next, the cells were cultured in DMEM culture medium supplemented with a sample (final concentration 5, 20 μg / mL) for 24 hours, and then cultured in DMEM culture medium supplemented with 50 μg / mL neutral red for another 2 hours. After washing the excess neutral red with PBS, 100 μL of 50% ethanol containing 1% acetic acid was added per well, and the neutral red taken up by the cells was extracted for 15 minutes. Using the blank as a control, the absorbance at 540 nm of the obtained extract was measured and used as a neutral red value. On the other hand, the cells after extracting the neutral red were fixed, stained with methylene blue, and then extracted with 0.1N hydrochloric acid. Using the blank as a control, the absorbance at 650 nm of the methylene blue extract was measured and used as the methylene blue value. The methylene blue value was used to correct the data as a value reflecting the number of cells. That is, the neutral red value was determined as a ratio to the methylene blue value, and was used as the amount of neutral red uptake per fixed number of cells. The neutral red uptake amount was calculated as the ratio of the neutral red uptake amount of the sample addition group to the neutral red uptake amount of the control.

The results of these tests are shown in Table 1. As a result, the extract of Atemoya (Production Examples 1 to 5) was found to have a remarkable absorption promoting effect.

A prescription example of the product containing the extract of Atemoya produced in Production Examples 1 to 5 is shown below.

Prescription example 1 Tablet prescription Content (% by weight)
1. 1. Atemoya extract 1.0
2. Dried cornstarch 25.0
3. 3. Carboxymethyl Cellulose Calcium 24.0
4. Microcrystalline Cellulose 40.0
5. Polyvinylpyrrolidone 7.0
6. Talc 3.0
[Manufacturing method] Ingredients 1 to 5 are mixed, then 10% water is added as a binder, and the mixture is extruded and granulated and then dried. Ingredient 6 is added to the molded granules, mixed and locked. One tablet weighs 0.52 g.

Prescription example 2 Capsules Prescription content (% by weight)
1. 1. Atemoya extract 5.0
2. Microcrystalline Cellulose 60.0
3. 3. Corn starch 15.0
4. Lactose 18.0
5. Polyvinylpyrrolidone 2.0
[Manufacturing method] Ingredients 1 to 5 are mixed and granulated, and then 250 mg is filled in a No. 2 hard capsule to obtain a capsule.

Prescription example 3 Powder prescription content (% by weight)
1. 1. Atemoya extract 5.0
2. Microcrystalline Cellulose 40.0
3. 3. Corn starch 55.0
[Manufacturing method] Ingredients 1 to 3 are mixed to obtain a powder by a conventional method.

Prescription example 4 Cream prescription content (% by weight)
1. 1. Atemoya extract 0.1
2. Squalene 5.5
3. 3. Olive oil 3.0
4. Stearic acid 2.0
5. Beeswax 2.0
6. Octyldodecyl myristate 3.5
7. Polyoxyethylene cetyl ether (20EO) 3.0
8. Behenyl alcohol 1.5
9. Glycerin monostearate 2.5
10. Fragrance 0.1
11. Methyl paraoxybenzoate 0.2
12. Ethyl paraoxybenzoate 0.05
13.1,3-butylene glycol 8.5
14. Remaining amount of purified water [Manufacturing method] Components 2 to 9 are heated and dissolved and mixed, and kept at 70 ° C. to prepare an oil phase. Ingredients 1 and 11 to 14 are heated and dissolved and mixed, and kept at 75 ° C. to prepare an aqueous phase. Next, an aqueous phase is added to the oil phase to emulsify, and the mixture is cooled while stirring, component 10 is added at 45 ° C., and the mixture is further cooled to 30 ° C. to obtain a product.

Prescription example 5 Ointment prescription content (% by weight)
1. 1. Atemoya extract 2.0
2. Polyoxyethylene cetyl ether (30EO) 2.0
3. 3. Glycerin monostearate 10.0
4. Liquid paraffin 5.0
5. Cetanol 6.0
6. Methyl paraoxybenzoate 0.1
7. Propylene glycol 10.0
8. Remaining amount of purified water [Manufacturing method] Components 2 to 5 are heated and dissolved and mixed, and kept at 70 ° C. to prepare an oil phase. Ingredients 1 and 6 to 8 are heated, dissolved and mixed, and kept at 75 ° C. to prepare an aqueous phase. An aqueous phase is added to the oil phase to emulsify, and the product is cooled to 30 ° C. with stirring.

Prescription example 6 Tablet confectionery Prescription content (% by weight)
1. 1. Atemoya extract 1.0
2. Dried cornstarch 50.0
3. 3. Erythritol 40.0
4. Citric acid 5.0
5. Sucrose fatty acid ester 3.0
6. Fragrance 1.0
[Manufacturing method] Purified water is added to components 1 to 4 in an appropriate amount, kneaded, extruded and granulated, and then dried to obtain granules. Ingredients 5 and 6 are added to the granules and tableted. 1 tablet is 1 g.

Prescription example 7 Beverage prescription content (% by weight)
1. 1. Atemoya extract 0.1
2. Stevia 0.05
3. 3. Malic acid 5.0
4. Sodium ascorbate 1.0
5. Fragrance 0.1
6. Remaining amount of purified water [Manufacturing method] Components 1 to 5 are stirred and dissolved in a part of purified water of component 6. The remaining purified water of component 6 is then added and mixed, heated to 90 ° C. and filled in a 50 mL glass bottle.

The absorption-promoting agent according to the present invention, which comprises an extract of Atemoya, enhances the effect of the drug which is taken up by cells through the cell membrane and acts. Therefore, it can be used to enhance the effect of a pharmaceutical composition utilizing receptor-independent endocytosis, for example, a gene therapy agent using an anticancer agent or siRNA, and to enhance the effect more safely.

Claims (2)

An endocytosis promoter characterized by containing a hot water extract of Atemoya (excluding application to multidrug-resistant tumor cells) . A receptor-independent endocytosis promoter characterized by containing a hot water extract of Atemoya (excluding applications to multidrug-resistant tumor cells) .