JP4809162B2 - Topical skin preparation - Google Patents

Description

  The present invention relates to a skin external preparation that can be applied to various skin external preparations such as pharmaceuticals, quasi drugs, and cosmetics (preparations used for humans and other animals).

The skin is covered with a stratum corneum, which is a thin biological protective film, and it is possible to live in a dry atmosphere without losing moisture, because the stratum corneum exists on the skin in contact with the outside world, that is, the epidermis. It is. The stratum corneum keeps the water from the body from losing, and at the same time, makes various adjustments so that the skin remains normal.
However, the skin is damaged and hardened due to trauma (burn, inflammation caused by various stimulating factors such as ultraviolet rays, etc.), the stratum corneum is cracked and peeled off, and a large amount of water is lost compared to normal skin. Will happen.

  On the other hand, a decrease in skin water content is often caused or involved in one of the causes for losing the flexibility, extensibility, and strength necessary for the stratum corneum. A functionally defective stratum corneum does not have enough water binding (retention) force to keep the stratum corneum flexible, and conversely allows a large amount of moisture to pass through and lose skin. In general, when the moisture content decreases to 10% or less of the dry weight of the stratum corneum, it loses the softness and softness of the skin that has been maintained until now, and it becomes hard and brittle, so-called crisp, moist feeling It can be said that the moisture content of the skin stratum corneum is very important because it results in no dry skin (skin).

  Therefore, conventionally, in order to make up for this point, cosmetics have been ideally reproduced on the skin with substances similar to those on the skin surface. Many of them are used. In recent years, the stratum corneum component called NMF (Natural Moisturizing Factor) has attracted attention as a third cosmetic component, and research and development of amino acids and their derivatives, which are NMF components, have been actively conducted. (Moisturizing / cleaning / ultraviolet absorption / antibacterial action, etc.).

On the other hand, various cell activators have been proposed in order to keep the skin healthy. For example, it is known that the above-mentioned amino acids have a cell activation action, and various saccharides are provided (Patent Documents 1 and 2).
However, when the additive is, for example, an amino acid, there are many kinds of amino acids, and there remains a problem in the optimum combination combination considering the penetration into the skin. In addition, saccharides tend to produce colored components by Maillard reaction during storage, and amino acids and saccharides are prone to spoilage, leaving problems for maintaining product stability and stabilizing bacterial flora on the epidermis.

JP-A 61-289016 JP-A-6-279227

As described above, amino acids have been known for various uses in the cosmetics field, but there is still room for improvement in terms of formulation. In addition, there remains a problem in using both useful components in consideration of permeability to the skin, storage stability of the product, and stability of the epidermal flora.
Therefore, the present inventors have intensively studied the additive substances that have a cell activation effect and are excellent in moisture retention, useful for pharmaceuticals, quasi drugs, and cosmetics, and stable.
The present invention provides a stable skin external preparation that has a moisturizing action and a cell activation action, is highly permeable and has little deterioration such as discoloration.

The present invention relates to the following [1] to [7], but other matters (for example, items 1 to 8 described later) are also described for reference.
[1]
A skin external preparation containing at least arginine, aspartic acid, isoleucine, leucine, lysine and threonine, or salts thereof among amino acids, and containing xylitol, wherein the amount of amino acids or salts thereof added is arginine 20-300 μg / ml, aspartic acid addition amount 5-60 μg / ml, isoleucine addition amount 30-600 μg / ml, leucine addition amount 30-600 μg / ml, lysine addition amount 30 An external preparation for skin having an addition amount of ˜600 μg / ml, an addition amount of threonine of 30 to 200 μg / ml, and an addition amount of xylitol of 50 to 3000 μg / ml.
[2]
Furthermore, the skin external preparation as described in [1] above, which contains at least one of glycine, histidine, serine, valine, tyrosine, cysteine, phenylalanine, or salts thereof among amino acids.
[3]
The external preparation for skin according to [1] or [2] above, which contains vitamin B.
[4]
The skin external preparation as described in [3] above, wherein the vitamin B is vitamin B1 or vitamin B6.
[5]
The external preparation for skin according to any one of the above [1] to [4], comprising hyaluronic acid or a salt thereof.
[6]
The skin external preparation according to any one of the above [1] to [5], wherein the skin external preparation is a cosmetic or a quasi-drug.
[7]
The external preparation for skin according to [6] above, wherein the external preparation for skin is lotion, emulsion, cream, hair nourishing agent, pack.
The above object was achieved by the following items 1 to 8.
1. A skin external preparation containing at least arginine, aspartic acid, isoleucine, leucine, lysine, threonine or salts thereof among amino acids, and containing xylitol.
2. Regarding the addition amount of amino acids or salts thereof, the addition amount of arginine is 20 to 400 microweight percent, the addition amount of aspartic acid is 3 to 60 microweight percent, the addition amount of isoleucine is 30 to 600 microweight percent, and the addition of leucine 1. The amount is 30-600 micro weight percent, the addition amount of lysine is 30-600 micro weight percent, and the addition amount of threonine is 25-500 micro weight percent. Topical skin preparation.
3. 1. The amount of xylitol added is 30 to 2000 micro weight percent. Topical skin preparation.
4). 1. Among amino acids, at least one of glycine, histidine, serine, valine, tyrosine, cysteine, phenylalanine or salts thereof is included. ~ 3. Topical skin preparation.
5. Contains vitamin Bs ~ 4. Topical skin preparation.
6). 4. B vitamins are vitamin B1 or vitamin B6 Topical skin preparation.
7). 1. Containing hyaluronic acid or a salt thereof ~ 6. Topical skin preparation.
8). 6. The topical skin preparation is a cosmetic or quasi-drug. Topical skin preparation.

  ADVANTAGE OF THE INVENTION According to this invention, it has a moisturizing effect | action and a cell activation effect | action, and can provide a stable skin external preparation with high permeability and little deterioration such as discoloration.

The amino acids used as essential in the present invention are arginine, aspartic acid, isoleucine, leucine, lysine and threonine, which may be optically active or racemic, but all L are particularly preferred.
The amount of amino acids added as essential in the present invention will be described in detail.
The amount of arginine added is preferably 20 to 400 microweight percent, more preferably 40 to 200 microweight percent, and most preferably 60 to 120 microweight percent.
The amount of aspartic acid added is preferably 3 to 60 micro weight percent, more preferably 8 to 30 micro weight percent, and most preferably 12 to 20 micro weight percent.
The amount of isoleucine added is preferably 30 to 600 microweight percent, more preferably 50 to 300 microweight percent, and most preferably 80 to 200 microweight percent.

The amount of leucine added is preferably 30 to 600 microweight percent, more preferably 50 to 300 microweight percent, and most preferably 80 to 200 microweight percent.
The amount of lysine added is preferably 30 to 600 microweight percent, more preferably 50 to 300 microweight percent, and most preferably 80 to 200 microweight percent.
The amount of threonine added is preferably 25 to 250 micro weight percent, more preferably 40 to 150 micro weight percent, and most preferably 60 to 120 micro weight percent.

  In the case of using an amino acid composition as an external preparation for skin, a method of adding trehalose to avoid Maillard reaction is known (Japanese Patent Laid-Open No. 6-279227). However, trehalose has a problem that bacteria easily reproduce in the container and on the skin, and the external preparation is stably maintained, and a large amount of antibacterial agent is required to keep the skin surface normal. As a result of various investigations on additives that improve the skin permeability of amino acid compositions and provide a moisturizing effect, it was found that xylitol among polyhydric alcohols satisfies all of the skin permeability, moisture retention and stability of amino acids. all right.

  The xylitol used in the present invention is usually obtained by reducing α-D-xylose, but the xylose source and its reduction method are not particularly limited. The amount of xylitol added is preferably 0.1 to 10 milliweight percent, more preferably 0.3 to 6 milliweight percent, and most preferably 0.5 to 3 milliweight percent.

  Next, amino acids used in addition to the essential amino acids in the present invention will be described. Further, the amino acid used by addition is glycine, histidine, serine, valine, tyrosine, cysteine, phenylalanine, which may be optically active or racemic, but all L are particularly preferable. At least one or more of these amino acids are preferably added, but more preferably three or more are added, and most preferably all eight are added.

Next, the amount of amino acid added and used in the present invention will be described.
The amount of glycine added is preferably 6 to 120 microweight percent, more preferably 16 to 60 microweight percent, and most preferably 20 to 40 microweight percent.
The amount of histidine added is preferably 8 to 160 micro weight percent, more preferably 20 to 90 micro weight percent, and most preferably 30 to 60 micro weight percent.
The amount of serine added is preferably 8 to 160 micro weight percent, more preferably 20 to 90 micro weight percent, and most preferably 30 to 60 micro weight percent.
The amount of threonine added is preferably 30 to 500 microweight percent, more preferably 50 to 200 microweight percent, and most preferably 70 to 150 microweight percent.
The amount of valine added is preferably 30 to 500 micro weight percent, more preferably 50 to 200 micro weight percent, and most preferably 70 to 150 micro weight percent.
The amount of tyrosine added is preferably 0.08 to 1.6 microweight percent, more preferably 0.2 to 0.9 microweight percent, and most preferably 0.3 to 0.6 microweight percent.
The amount of cysteine added is preferably 15 to 300 micro weight percent, more preferably 30 to 150 micro weight percent, and most preferably 40 to 80 micro weight percent.
The amount of phenylalanine added is preferably 0.12-2.4 microweight percent, more preferably 0.2-1.5 microweight percent, and most preferably 0.3-1.0 microweight percent.

Vitamins have been used in various skin external preparations due to their coenzyme activity, but vitamins belonging to the vitamin B group are particularly preferred when combined with the skin external preparation of the present invention. The vitamin B used in the present invention is specifically vitamin B1, vitamin B2, vitamin B3, vitamin B4, vitamin B5, vitamin B6, vitamin B12, vitamin B13, vitamin B15, vitamin B17, vitamin BT, vitamin Bx, vitamin Bc, vitamin B1 involved in sugar metabolism, vitamin B2 involved in lipid metabolism, and vitamin B6 involved in protein metabolism are preferred, and vitamin B1 and vitamin B6 are more preferred.
The amount of vitamin B to be used in the present invention is preferably 0.8 to 16 microweight percent, more preferably 2 to 9 microweight percent, and most preferably 3 to 6 microweight percent.

In the present invention, it is particularly preferable to add hyaluronic acid in order to maintain skin elasticity. The hyaluronic acid may be a sodium salt or a potassium salt, or may be a free acid. When the skin external preparation of the present invention is a cosmetic, addition of hyaluronic acid is particularly preferable. The amount of hyaluronic acid used in the external preparation for skin is preferably 150 to 3000 microweight percent, more preferably 300 to 1500 microweight percent, and most preferably 400 to 800 microweight percent.
The skin external preparation of the present invention can be used in combination with various medicinal agents. Such medicinal agents are, for example, active oxygen scavengers, antioxidants, cell activators, anti-inflammatory agents, tyrosinase activity inhibitors, UV absorbers and humectants. Specific examples of the medicinal agent include those shown below, but of course not limited thereto.

The active oxygen scavenger or antioxidant that can be used in the present invention is not particularly limited, but is preferably extracted or purified from natural products rather than those obtained by chemical synthesis.
Examples of the active oxygen scavenger include β-carotene, α-carotene, lycopene, lutein, astaxanthin, capsanthin, fucoxanthin, heteroxanthine, loroxanthin, luteoxanthin, lycophy, lycoxanthin, neochrome, neoxanthine, rhodopine, rhodopinal. , Rhodopinol, rhodovibrin, trolixanthine, xanthophyll, zeaxanthin and other carotenoids, superoxide dismutase, mannitol, rutin and derivatives thereof, bilirubin, cholesterol, tryptophan, histidine, quercetin, quercitrin, catechin, catechin derivatives, gallic acid and derivatives thereof , Glutathione and derivatives thereof, and salts thereof, hornon extract, ginkgo biloba extract, keiketto extract, hawthorn extract , Plant extracts containing flavonoids, such as Mikaika extract, Yukinoshita extract, Melissa extract, Gennoshoco extract, Botpi extract, Parsley extract, Tormentilla extract, Rakan fruit extract, Yashajitsu extract and Zicopi extract Can be mentioned.

  Antioxidants include, for example, retinol and its derivatives such as retinol palmitate and retinol acetate, retinal and its derivatives, vitamin A such as dehydroretinal; thiamines such as thiamine hydrochloride and thiamine sulfate, riboflavin, and pyridoxine hydrochloride , Pyridoxines such as pyridoxine dioctanoate, flavin adenine nucleotide, cyanocobalamin, folic acid, nicotinic acid amide, nicotinic acid such as benzyl nicotinate, calcium pantothenate, D-pantothenyl alcohol, pantothenyl ethyl ether, acetyl pantothenyl Pantothenic acids such as ethyl ether, vitamin B such as choline and inositol; L-ascorbic acid, L-ascorbyl palmitate, L-ascorbyl dipalmitate, isopalmitic acid L Ascorbyl, L-ascorbyl diisopalmitate, L-ascorbyl tetraisopalmitate, L-ascorbyl stearate, L-ascorbyl distearate, L-ascorbyl isostearate, L-ascorbyl diisostearate, L-ascorbyl myristate, dimyristin L-ascorbyl acid, L-ascorbyl isomyristate, L-ascorbyl diisomyristate, L-ascorbyl oleate, L-ascorbyl dioleate, L-ascorbyl 2-ethylhexanoate, sodium L-ascorbate phosphate, L-ascorbic acid potassium phosphate, L-ascorbic acid phosphate magnesium, L-ascorbic acid phosphate calcium, L-ascorbic acid phosphate aluminum , Sodium L-ascorbate sulfate, potassium L-ascorbate sulfate, magnesium L-ascorbate sulfate, calcium L-ascorbate sulfate, aluminum L-ascorbate sulfate, sodium L-ascorbate, L- Vitamin C such as potassium ascorbate, magnesium L-ascorbate, calcium L-ascorbate, aluminum L-ascorbate, and derivatives thereof, and salts thereof; ergocalciferol, cholecalciferol, dihydroxystanal, etc. Dl-α (β, γ) -tocopherol, dl-α-tocopherol acetate, nicotinic acid-dl-α-tocopherol, linoleic acid-dl-α-tocopherol, succinic acid d Examples include tocopherols such as l-α-tocopherol and derivatives thereof, salts thereof, vitamin E such as ubiquinones, dibutylhydroxytoluene and butylhydroxyanisole.

  Among the above active oxygen scavengers and antioxidants, mannitol, beta carotene, astaxanthin, rutin and its derivatives, ginkgo biloba extract, urgonum extract, quette extract, yukinoshita extract, melissa extract, gennoshoco Extracts, Ages extract, vitamins C and their derivatives and salts thereof, vitamin E and its derivatives and salts thereof, and more preferable are beta carotene, astaxanthin, ginkgo extract, vitamins C and Their derivatives and their salts, vitamin E and its derivatives and their salts.

  The concentration of the active oxygen removing agent or antioxidant in the external preparation for skin of the present invention is preferably 0.00001 to 50% by weight, more preferably 0.0001 to 30% by weight. However, when using a plant extract, there is no problem as long as the dry solid content is within the above range. Further, the active oxygen scavenger or the antioxidant can be used alone or in combination of two or more.

(Cell activator)
Examples of the cell activator include adenylate derivatives such as deoxyribonucleic acid and salts thereof, adenosine triphosphate and adenosine monophosphate and salts thereof, ribonucleic acid and salts thereof, guanine, xanthine and derivatives thereof and salts thereof. Nucleic acid-related substances such as serum deproteinized extract, spleen extract, placenta extract, chicken crown extract, royal jelly extract, etc .; yeast extract, lactic acid fermentation extract, bifidobacteria extract, ganoderma extract Extracts derived from microorganisms such as foods; extracts derived from plants such as carrot extract, assembly extract, rosemary extract, agaric extract, garlic extract, hinokitiol, cephalanthin; α- or γ-linolenic acid, Eicosapentaenoic acid and their derivatives, succinic acid and its derivatives and their salts, estradiol and its derivatives And α-hydroxy acids such as lactic acid, glycolic acid, citric acid, malic acid, salicylic acid, and derivatives thereof, and salts thereof.

  Among these cell activators, particularly preferred are deoxyribonucleic acid and its salt, adenosine triphosphate and its salt, serum deproteinized extract, placental extract, yeast extract, lactic acid fermentation extract, carrot extract, Hinokitiol, succinic acid and its derivatives and their salts.

  The concentration of the cell activator in the external preparation for skin of the present invention is preferably 0.0001 to 5% by weight, more preferably 0.001 to 3% by weight. However, when using a plant extract, there is no problem as long as the dry solid content is within the above range. Moreover, a cell activator can be used 1 type or in combination of 2 or more types.

(Anti-inflammatory agent)
Anti-inflammatory agents include glycyrrhizic acid, glycyrrhetinic acid, mefenamic acid, phenylbutazone, indomethacin, ibuprofen, ketoprofen, allantoin, guaiazulene and their derivatives and their salts, ε-aminocaproic acid, zinc oxide, diclofenac sodium, aloe extraction Products, salvia extract, arnica extract, chamomile extract, birch extract, hypericum extract, eucalyptus extract, mukuroji extract and the like.

  Among these anti-inflammatory agents, particularly preferred are glycyrrhizic acid, glycyrrhetinic acid, guaiazulene and derivatives thereof, and salts thereof, ε-aminocaproic acid, aloe extract, chamomile extract.

  The concentration of the anti-inflammatory agent is generally 0.0001 to 1%, preferably 0.01 to 0.5% in the composition. However, when using a plant extract, there is no problem as long as the dry solid content is within the above range. The anti-inflammatory agents can be used alone or in combination of two or more.

(Tyrosinase activity inhibitor)
Examples of tyrosinase activity inhibitors include cysteine and derivatives thereof (for example, N, N′-diacetylcystine dimethyl, etc.) and salts thereof, Sempukuka extract, quette extract, sunpens extract, Sohakuhi extract, Toki extract, Ibukitorano extract Products, Clara extract, hawthorn extract, white lily extract, hop extract, Neubara extract, Yokuinin extract and the like.
The concentration of the tyrosinase activity inhibitor is preferably 0.0001 to 2%, particularly preferably 0.001 to 0.5%. However, when using a plant extract, there is no problem as long as the dry solid content is within the above range. In addition, these can be used 1 type or in combination of 2 or more types.

(Humectant)
As the humectant, not only synthetic compounds such as urea, but also low molecular weight compounds such as amino acids known as natural moisturizing factors, pyrrolidone carboxylic acid and lactate can be used. In addition, mucopolysaccharides and / or proteins that are constituents of the skin and are conventionally blended in cosmetics can be used.

Among these, amino acids include those described in “Development of Newly Useful Cosmetics”, Masato Suzuki / Supervision, CMC Publishing, September 2004, pages 16-19.
Examples of the mucopolysaccharide include hyaluronic acid, chondroitin sulfate, dermatan sulfate, heparan sulfate, heparin and keratan sulfate, and salts thereof. Hyaluronic acid, chondroitin sulfate, and salts thereof can be preferably used. .
Examples of the protein include collagen, elastin, keratin and derivatives thereof, and salts thereof, and collagen is particularly preferable. There is no restriction | limiting in particular about the origin of each of these components, Any of animal origin, microorganism origin, and a synthetic product may be sufficient. There are no particular restrictions on the extraction method and the purification treatment method in the case of natural origin.

These moisturizing components may be of one type, or two or more types can be added and used as appropriate.
Furthermore, the amount of the moisturizing agent varies depending on the combination of the components, but is generally preferably 0.0001 to 5%, more preferably 0.001 to 3%.

  The amino acid, vitamin B and xylitol compositions comprising the amino acids of the present invention or salts thereof can be prepared by blending with various forms of bases known as ordinary external compositions according to conventional methods. That is, oil agents (natural animal and vegetable oils, semi-synthetic oils and fats, hydrocarbon oils, higher fatty acids, ester oils, silicone oils, fluorine-based oil agents, etc.), gelling agents, metal soaps, surfactants (anionic, cationic, amphoteric, Powders (inorganic powders, organic powders, pigments, etc.), alcohols (higher alcohols, polyhydric alcohols, sterols, etc.), water-soluble polymers (animal and vegetable systems, microbial systems, synthetic systems), film forming agents, resins, Preservatives, antibacterial agents, fragrances, essential oils, salts, water (purified water, hot spring water and deep water), pH adjusters, refreshing agents, skin roughness improvers, blood circulation promoters, skin astringents, antiseborrheic agents, amino acids , Nucleic acid-related substances, enzymes, hormones, inclusion compounds, plant extracts, animal and microbial extracts, UV scattering agents and the like can be added.

Hereinafter, the present invention will be described in detail based on examples, but it goes without saying that the present invention is not limited thereto.
In addition, Examples 1-3 shall be read as Reference Examples 1-3, respectively. In addition, the present compositions 1 to 3, the present compositions 2-1 and 2-2, and the test liquid 1 (the present invention) are referred to as reference compositions 1 to 3, reference compositions 2-1 and 2-2, and It shall be read as Test Solution 1 (Reference Example).

Example 1 (color tone stability test)
The compositions shown in Tables 1 and 2 were prepared, packed in glass bottles and stored in a 50 ° C. constant temperature bath, and the temporal stability was examined with the naked eye.

The evaluation method was performed in the following three stages.
-: Coloring is not seen.
+-: Slightly colored.
+: Coloring.

  As a result, as shown in Table 3, it was confirmed that the amino acid mixture and the xylitol composition of the present invention showed no coloration even at a high concentration and for a long period of time. This is because xylitol does not have an aldehyde equivalent partial structure, and thus does not cause coloring due to the Maillard reaction and is stable. On the other hand, it was confirmed that the comparative product (amino acid mixture and glucose or trehalose) showed coloring.

Example 2 (antibacterial test)
First, methylparaben (Midori Kagaku) was added to the composition of Example 1 Table 1 to prepare the composition of Table 4.

E. coli (IFO 3301) and Pseudomonas aeruginosa (IID P-1) were used as test bacteria, and these test strains were inoculated into SCDLP medium (Nippon Pharmaceutical Co., Ltd.) and cultured at 37 ° C. for 18 hours to pre-culture the bacteria. A liquid was prepared. Moreover, the composition of Table 4 was diluted 5 times with the SCDLP medium to prepare a test solution.
49.5 ml of the test solution prepared as described above and 0.5 ml of the preculture solution were added to a 200 ml Erlenmeyer flask, shaken at 25 ° C., and the viable cell count after 24 hours was measured by a plate smearing method. The sterilization rate was calculated from the measurement result by the following formula.
Sterilization rate (%) = 100-(number of viable bacteria after 48 hours / number of initial bacteria * 100) (the larger the value, the stronger the antibacterial properties)
The sterilization results are shown in Table 5.

  As shown by the composition 2-1 of the present invention, no antibacterial property is exhibited in the absence of paraben. That is, xylitol itself does not show a remarkable antibacterial property, but when paraben was added, xylitol showed an antibacterial property superior to glucose and trehalose as additives.

Example 3 (skin permeability test)
Permeation test skin
The abdominal skin of a 10-20 week old male male hairless mouse (Kudo Co., Ltd.) was removed and used after removing subcutaneous fat.
Skin Permeation Test Skin samples were mounted in a Keshany-Chien type diffusion cell. 1 ml of the test solution (described later) was added to the donor phase, and the upper part was sealed with parafilm. 0.01 mol / l phosphate buffered saline (Wako Pure Chemical Industries) was added to the receptor phase, stirred and incubated at 32 ° C. After 24 hours, the amount of amino acids was quantified by HPLC, and the permeability was calculated. .
Amino acid quantification method The HPLC amino acid analysis system (Prominence) of Shimadzu Corporation was used and quantified by the post-column fluorescence detection method using OPA (orthophthalaldehyde) as a reaction reagent.

Test solution
20 amino acids (Wako Pure Chemicals: valine, leucine, isoleucine, alanine, arginine, glutamine, lysine, aspartic acid, glutamic acid, proline, cysteine, threonine, methionine, histidine, phenylalanine, tyrosine, tryptophan, asparagine, glycine, serine) 0.1% by weight, 1,3-butylene glycol (Wako Pure Chemical) 1% by weight, xylitol 1% by weight as additive (Wako Pure Chemical) and 0.01 mol / l phosphorus Acid buffered saline (Wako Pure Chemical Industries, Ltd.) was mixed to prepare Test Solution 1.
In place of xylitol, glucose, trehalose, glycerin, sorbitol, and mannitol were mixed in the same manner to prepare test solutions 2, 3, 4, 5, and 6, respectively.
Table 6 shows the results obtained by testing the test solution by the above-described skin permeation test method and measuring the transmittance of all permeated amino acids.

The additive xylitol of the present invention showed a greater permeation promoting effect than other polysaccharides such as glycerin, sorbitol and mannitol, and a permeation promoting effect superior to that of glucose and trehalose.
Table 7 shows the transmittance of each amino acid when the additive of the present invention is xylitol.

  Arginine, aspartic acid, isoleucine, leucine, lysine, and threonine show excellent skin permeability in the presence of xylitol under the test conditions, which may allow these amino acids to penetrate into the cell through a synergistic effect with xylitol. Suggests sex.

Example 4 (Human normal keratinocyte proliferation test)
Preparation of Purified Type IV Collagen-Acetic Acid Solution According to the following protocols (1) to (16), 8 mg of purified high molecular type collagen IV was obtained from the raw pig eyeball. The following protocol was performed at 4 ° C.

(1) Remove the cornea from the eyeball using scissors and remove the lens from the inside of the eyeball.
(2) Remove insoluble parts such as vitreous adhering to the lens as much as possible with scissors.
(3) Add 1 tablet of complete protease inhibitor cocktail (Roche) to 50 ml of cold PBS (phosphate buffered saline), add the lens capsule, and stir for 2 hours.
(4) Centrifugation (2000 g, 10 minutes, 4 ° C.) to remove unnecessary sites present in the supernatant.
(5) The precipitate is suspended in 25 ml of 0.5 M acetic acid in which a complete protease inhibitor cocktail half tablet (Roche) is dissolved.
(6) Crush finely using a homogenizer (IKA).
(7) The finely crushed lens capsule is stirred for 3 days to extract type IV collagen.
(8) Centrifugation (2000 g, 10 minutes, 4 ° C.) to separate the supernatant (acetate-soluble collagen) and the precipitate.
(9) Repeat this agitation extraction and centrifugation once more.
(10) To the supernatant obtained by centrifugation, add NaCl with ground crystals as much as possible in a mortar to a final concentration of 1.7M.
(11) Stir overnight to precipitate collagen.
(12) Centrifugation (5000 g, 30 minutes, 4 ° C.) to collect the precipitate.
(13) Add 0.5M acetic acid to the precipitate and dissolve it sufficiently.
(14) The collagen acidic aqueous solution is put into a dialysis tube (Sanko Junyaku) and dialyzed using 0.5M acetic acid.
(15) Further, dialyze with 2 mM hydrochloric acid.
(16) Collect the collagen solution after dialysis to obtain a purified type IV collagen solution.

Collagen film culture procedure
1. Dilute the aforementioned type IV collagen-acetic acid solution (1 mg / ml) 10-fold with sterilized ultrapure water and pour 50-100 μl per square centimeter into a 48-well culture dish or plate.
2. Extend thinly to spread over the entire culture surface.
3. Open the lid of the culture dish or plate in a clean bench and dry at 25 ° C or lower.
4. After drying, wash with medium 3 times to remove acid from type IV collagen film.
Five. Seed 3500 normal keratinocytes (Sanko Junyaku CC-2503-NZ) per square centimeter.
6. Place IV collagen plate in CO ₂ incubator and start culturing.
7. Change medium every 2 days and incubate for 8 days.

Skin cell proliferation test The proliferation test was performed using the MTT Cell Growth Assay Kit.
Reagent A: MTT, (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyl tetrasodium bromide), 50 mg / vial.
Reagent B: PBS pH 7.4, 15 mL
Reagent C: Color development solution (isopropanol with 0.04 N HCl), 100 mL
Add 10 ml of reagent B to reagent A and mix well.
Leave in a cool dark place overnight.
Sterilize the AB reagent with a 0.22 μm filter.
Add 30 μl of AB reagent per cm ^{2 and} mix gently.
Incubate for 4 hours in a CO ₂ incubator.
After culture, the culture supernatant is transferred to a tube.
Add 300 μl of Reagent C per cm ² .
After mixing, transfer to a tube containing the culture supernatant.
Measure the absorbance (570 nm) within 1 hour.

Test medium Each additive (Wako Pure Chemical Industries) was added to Bullet Kit KGM-2 Medium (Growth Medium for Epidermal Keratinocytes) Sanko Pure Chemical CC-1307-NZ so as to have the concentrations shown in Tables 8-21. The test medium was used.
The test medium shown in Table 19 has a composition similar to that shown in the known examples of JP-A 61-289016.

Skin cell proliferation test results Absorbance at the time of no addition was defined as 100, and relative concentrations thereof were shown in Tables 22 and 23.

The composition 3-1 of the present invention promotes the proliferation of human epidermal keratinocytes on the type IV collagen plate, and its effect is enhanced by vitamin B6 and vitamin B1 (the composition 3-2 of the present invention). (The composition 3-3 of the present invention).
The influence of the concentration of each amino acid is shown in Compositions 3-4 to 3-13 of the present invention. The cell growth effect was substantially saturated at the upper limit indicated for each amino acid. When the amount of xylitol is 50 μg / ml, there is a proliferation effect (composition 3-14 of the present invention), but when the amount is reduced to 10 μg / ml, the effect is lost (comparative composition 3-1), and vitamin Bs and amino acids are added. Even when added, there is no proliferation effect (Comparative composition 3-1).
When the amino acid of the present invention was not added, no cell proliferation effect was obtained (Comparative Examples 3-3 to 3-8).
A human epidermal cell proliferation effect was not recognized only by a combination of known amino acids (Comparative Examples 3-9 to 3-11).

(Cosmetics test)
Production Example 1
A skin care lotion was made with the following composition.
Each component was mixed at 70 ° C. so as to have the concentrations shown in Table 24 below to form an aqueous solution, and then cooled to room temperature.

Production Example 2
A skin care emulsion was prepared with the following composition.
A liquid Each component was mixed at 70 degreeC so that it might become a composition of Table 25, and it was set as aqueous solution.

B liquid Each component was mixed at 70 degreeC so that it might become a composition ratio of Table 26. FIG.

Manufacturing method
65 ml of liquid A and 15 g of liquid B were mixed at 70 ° C., 20 ml of xanthan gum (2% aqueous solution) was added, and the mixture was mixed at 70 ° C. until uniform. Then, it cooled to room temperature.

Production Example 3
A skin care cream was prototyped with the following composition.
A liquid Each component was mixed at 70 degreeC so that it might become a composition of Table 27, and it was set as aqueous solution.

B liquid Each component was mixed at 70 degreeC so that it might become the composition ratio of Table 28. FIG.

Manufacturing method
51 ml of liquid A and 40 g of liquid B were mixed at 70 ° C., 1.0 g of triethanolamine was added, and the mixture was mixed at 70 ° C. until uniform emulsification. Then, it cooled to room temperature.

Example 4
Skin Care Lotion Evaluation Method Test Panel 10 healthy women aged 27-35 years, average age 31.4 years Location Room with a temperature of about 24 ° C and humidity of about 55 percent.
Evaluation method It apply | coated to the random position inside the forearm after washing | cleaning, and test | use feeling (sensory feeling) was scored on the basis of the following sensory test results.

4 Feels very well
3 Well felt well
2 I don't feel much
1 I can't feel

Then, the average evaluation score of 10 panelists was obtained and ranked as follows.
A 3.2 or higher
B 2.7 to less than 3.2
C 2.2 to less than 2.7
D 1.7 or more and less than 2.2
E Less than 1.7

Skin Care Lotion According to the present invention, the composition shown in Table 24 is Comparative Example 4-1, excluding xylitol, Comparative Example 4-2 replacing xylitol with glucose, and Comparative Example 4-3 replacing trehalose. .

  The result of inquiring about the impression after about 5 minutes after applying the coating thoroughly is shown below.

As is apparent from the table, the amino acid lotion combined with the xylitol of the present invention has a strong penetrating feeling and is excellent in overall moist feeling and elongation, and is excellent overall.
Example 5 (moisturizing effect)
The evaluation panel is the same as before, with an average age of 33.6 years.
The moisturizing effect was evaluated by measuring the amount of horny moisture by the high frequency impedance method.
(Asahi BioMed's high-sensitivity horny layer moisture meter ASA-MX, double frequency phase difference amplitude detection method)
Emulsion for skin care The composition shown in Table 25 is the present invention, except that xylitol is excluded from comparative example 5-1, in which xylitol is replaced by glucose, comparative example 5-2, and in which trehalose is replaced by comparative example 5-3 did.

 After application to apply thoroughly, the skin conductivity was measured about 18 hours later and the increase rate after application was determined.

 As apparent from the table, the composition of the present invention showed excellent moisture retention.

Example 6
Evaluation method The evaluation panel is the same as before, with an average age of 33.4 years.
Each day, twice in the morning and noon, after washing both hands, it was applied to the back of the hand and used for 2 weeks.
The test results were scored according to the following criteria.
Skin activation effect

4 I feel a lot of improvement in skin tension and gloss
3 Well felt
2 I don't feel much
1 I can't feel

 Then, the average evaluation score of 10 panelists was obtained and ranked as follows.

A 3.2 or higher
B 2.7 to less than 3.2
C 2.2 to less than 2.7
D 1.7 or more and less than 2.2
E Less than 1.7

Rough skin suppression effect

4 I feel the improvement of rough skin and rough skin
3 Well felt
2 I don't feel much
1 I can't feel

Then, the average evaluation score of 10 panelists was obtained and ranked as follows.
A 3.2 or higher
B 2.7 to less than 3.2
C 2.2 to less than 2.7
D 1.7 or more and less than 2.2
E Less than 1.7

  The composition shown in Table 27 was defined as the present invention, and the xylitol was excluded as Comparative Example 6-1; the xylitol was replaced with glucose as Comparative Example 6-2; and the trehalose was replaced with Comparative Example 6-3.

  As is apparent from the table, the composition of the present invention showed excellent skin activation and rough skin suppression effect.

Claims (7)

A skin external preparation containing at least arginine, aspartic acid, isoleucine, leucine, lysine and threonine, or salts thereof among amino acids, and containing xylitol, wherein the amount of amino acids or salts thereof added is arginine 20-300 μg / ml, aspartic acid addition amount 5-60 μg / ml, isoleucine addition amount 30-600 μg / ml, leucine addition amount 30-600 μg / ml, lysine addition amount 30 An external preparation for skin having an addition amount of ˜600 μg / ml, an addition amount of threonine of 30 to 200 μg / ml, and an addition amount of xylitol of 50 to 3000 μg / ml. Furthermore, the skin external preparation of Claim 1 containing at least 1 sort (s) of glycine, histidine, serine, valine, tyrosine, cysteine or phenylalanine, or those salts among amino acids. The skin external preparation of Claim 1 or 2 containing vitamin B. The external preparation for skin according to claim 3, wherein the vitamin B is vitamin B1 or vitamin B6. The skin external preparation as described in any one of Claims 1-4 containing a hyaluronic acid or its salt. The skin external preparation according to any one of claims 1 to 5 , wherein the skin external preparation is a cosmetic or a quasi-drug. The skin external preparation according to claim 6, wherein the skin external preparation is a lotion, an emulsion, a cream, a hair nourishing agent, or a pack.