JP2013095708A - Medusa collagen peptide mixture - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a medusa collagen peptide mixture having a repair improvement effect for a horny barrier function.SOLUTION: The medusa collagen peptide mixture contains a medusa collagen peptide containing any of amino acid sequences of (1) Gly-Pro-Ala-Gly, (2) Pro-Gly-Val-Lys-Gly-Glu-Val-Gly-Glu-Ala-Gly-Lys-Arg-Gly, (3) Val-Gly-Pro-Val-Thr-Glu-Glu-Val-Ser-Pro-Ala-Lys, (4) Ser-Ala-Gly-Gln-Asn-Pro-Ala-Asp-Asp-Val, (5) Gly-Thr-Pro(OH)-Gly-Ala-Gly-Gly-Ser-Arg, and (6) Gln-Gly-Pro-Gln-Gly-Glu-Leu, where Pro(OH) represents hydroxyproline.

Description

  The present invention relates to a jellyfish collagen peptide mixture comprising a specific amino acid sequence. The present invention also relates to an external preparation containing the jellyfish collagen peptide, and particularly to an external preparation as a keratinocyte activator, an involucrin production promoter, a transglutaminase production promoter and / or an anti-inflammatory agent.

  In recent years, the mass outbreak of jellyfish in the seas near Japan has become a social problem, and the large amount of echidella jellyfish has damaged the fishing nets and has caused serious damage to the fishery. There are also damages such as water jellyfish and Echizen jellyfish clogging water intakes in factories, thermal power plants, and nuclear power plants. Although a large amount of jellyfish are mainly captured and exterminated, it is costly to dispose of the captured jellyfish. Therefore, an effective solution to capture and extinguishment has been desired from various directions.

  As one of the countermeasures, use of jellyfish as a collagen raw material has been proposed.

  Collagen, which is an important item of basic cosmetics, is mainly derived from mammals such as cows and pigs. In mammals, 1/3 of protein is collagen. On the other hand, jellyfish is composed of 95 to 98% by weight of water and 2 to 5% by weight of solids, 20 to 30% by weight of these solids being protein, and 80% by weight being collagen. Yes.

  In general, mammal-derived collagen is an extract from hard tissues such as animal skin and bone, whereas jellyfish collagen composed of soft tissues containing a large amount of water has other animal properties. Expected to have functions and activities not found in collagen.

  As a study on jellyfish extract, it is reported that the liquid retention obtained from moon jellyfish and Echizen jellyfish can be used in skin preparations to renew the moisture retention capacity of the skin and normalize the turnover of the stratum corneum. (Patent Document 1). However, the liquid substance obtained from jellyfish, that is, the epidermal cell activation effect by the protein containing jellyfish collagen was not fully satisfactory.

  The skin protects internal cells from physical stimuli such as dryness and ultraviolet rays, and prevents the loss of internal components. This function is called a skin barrier function or horny barrier function, and the stratum corneum plays an important role in this function.

  The stratum corneum is formed from a stratum corneum formed by differentiation of epidermal keratinocytes and intercellular lipids filling the cells, and the stratum corneum is composed of keratin fibers and a conformed envelope (CE). Is formed by crosslinking involucrin, loricrin, or the like with an enzyme such as transglutaminase 1. It is known that the keratin barrier function is formed by covalently binding a part of CE and ceramide, which is an intercellular lipid. To improve the keratin barrier function, the proliferation of epidermal keratinocytes, Increases in involucrin and transglutaminase are considered important.

As for the involucrin production promoting effect and the transglutaminase production promoting effect, the effects of natural products such as hypericaceae Ceylon tetsuboku, birch, royal jelly, and extracts thereof have been reported (Patent Documents 2, 3, and 4). However, there has been no report on the activating effect of epidermis cells of jellyfish collagen peptide and the effect of promoting production of involucrin and transglutaminase.

JP 2011-116672 A JP 2005-213187 A JP 2007-217326 A JP 2009-184955 A

  An object of the present invention is to provide a jellyfish collagen peptide mixture having an effect of improving the repair of the keratin barrier function.

  As a result of intensive studies to solve the above problems, the present inventors have found that a mixture of jellyfish collagen peptides containing a specific amino acid sequence has an excellent epidermal keratinocyte activation effect, involucrin production promoting effect, transglutaminase production The present inventors have found a promoting effect and an anti-inflammatory effect, and have completed the present invention based on this finding. These effects make it possible to repair and improve the keratin barrier function.

That is, the present invention relates to the following (1) to (13).
(1) A jellyfish collagen peptide mixture containing a jellyfish collagen peptide containing any of the following amino acid sequences.

<1> Gly-Pro-Ala-Gly (SEQ ID NO: 1)
<2> Pro-Gly-Val-Lys-Gly-Glu-Val-Gly-Glu-Ala-Gly-Lys-Arg-Gly (SEQ ID NO: 2)
<3> Val-Gly-Pro-Val-Thr-Glu-Glu-Val-Ser-Pro-Ala-Lys (SEQ ID NO: 3)
<4> Ser-Ala-Gly-Gln-Asn-Pro-Ala-Asp-Asp-Val (SEQ ID NO: 4)
<5> Gly-Thr-Pro (OH) -Gly-Ala-Gly-Gly-Ser-Arg (SEQ ID NO: 5)
<6> Gln-Gly-Pro-Gln-Gly-Glu-Leu (SEQ ID NO: 6)
Pro (OH) represents hydroxyproline.

(2) The jellyfish collagen peptide mixture according to item 1, obtained by hydrolyzing jellyfish collagen.

(3) The jellyfish collagen peptide mixture according to item 2, wherein the hydrolysis is performed using at least one peptide-degrading enzyme selected from the group consisting of trypsin, pepsin, and papain.

(4) The jellyfish collagen peptide mixture according to item 2, wherein hydrolysis is performed using a solid acid catalyst.

(5) The jellyfish collagen peptide mixture according to item 4, wherein the solid acid catalyst is at least one solid acid catalyst selected from the group consisting of a cation exchanger, zeolite, and diatomaceous earth.

(6) The jellyfish collagen peptide mixture according to any one of Items 1 to 5, wherein the jellyfish collagen is a moon jellyfish collagen or an Echizen jellyfish collagen.

(7) A jellyfish collagen peptide mixture obtained by hydrolyzing jellyfish collagen using a peptide degrading enzyme or a solid acid catalyst.

(8) A peptide comprising any of the following amino acid sequences.

<1> Gly-Pro-Ala-Gly (SEQ ID NO: 1)
<2> Pro-Gly-Val-Lys-Gly-Glu-Val-Gly-Glu-Ala-Gly-Lys-Arg-Gly (SEQ ID NO: 2)
<3> Val-Gly-Pro-Val-Thr-Glu-Glu-Val-Ser-Pro-Ala-Lys (SEQ ID NO: 3)
<4> Ser-Ala-Gly-Gln-Asn-Pro-Ala-Asp-Asp-Val (SEQ ID NO: 4)
<5> Gly-Thr-Pro (OH) -Gly-Ala-Gly-Gly-Ser-Arg (SEQ ID NO: 5)
<6> Gln-Gly-Pro-Gln-Gly-Glu-Leu (SEQ ID NO: 6)
Pro (OH) represents hydroxyproline.

(9) An external preparation containing the jellyfish collagen peptide mixture according to any one of Items 1 to 7 and / or the peptide according to Item 8.

(10) A keratinocyte activator comprising the jellyfish collagen peptide mixture according to any one of Items 1 to 7 and / or the peptide according to Item 8.

(11) An involucrin production promoter containing the jellyfish collagen peptide mixture according to any one of Items 1 to 7 and / or the peptide according to Item 8.

(12) A transglutaminase production promoter containing the jellyfish collagen peptide mixture according to any one of items 1 to 7 and / or the peptide according to item 8.

(13) An anti-inflammatory agent comprising the jellyfish collagen peptide mixture according to any one of items 1 to 7 and / or the peptide according to item 8.

  According to the present invention, an unused resource jellyfish can be effectively used, and a mixture of jellyfish collagen peptides can be provided relatively inexpensively. In addition, keratinocyte activation effect, involculin production promotion effect, transglutaminase production promotion effect of the present invention, jellyfish collagen peptide excellent in anti-inflammatory effect is transdermally administered as an external preparation, the formation of the stratum corneum is promoted, The keratin barrier function can be repaired and improved.

FIG. 1 shows the results of mass spectrometry of the jellyfish collagen peptide mixture produced in Example 1. FIG. 2 shows the results of mass spectrometry of the jellyfish collagen peptide mixture produced in Example 9. FIG. 3 shows the molecular weight distribution of the jellyfish collagen peptide mixture produced in Examples 12-14. In the figure, 1-3 show the molecular weight distribution of the jellyfish collagen peptide mixture produced in Examples 12-14, respectively, 4 shows the molecular weight of the molecular weight marker (the numerical value at the bottom of the table indicates the molecular weight (kDa)). Show.

As used herein, the term “jellyfish collagen peptide” refers to a peptide comprising an amino acid sequence derived from jellyfish collagen. In the present specification, a mixture of jellyfish collagen peptides (jellyfish collagen peptide mixture) may be referred to as jellyfish collagen peptide, and a peptide obtained by synthesis based on amino acid sequence information derived from jellyfish collagen may be referred to as jellyfish collagen peptide.

  The mixture of jellyfish collagen peptides of the present invention can be obtained by hydrolysis of jellyfish collagen. In one embodiment, the mixture of jellyfish collagen peptides contains jellyfish collagen peptides comprising any of the following amino acid sequences: In addition, a peptide containing the following amino acid sequence obtained by synthesis may be used, or a mixture of a peptide containing the following amino acid sequence obtained by synthesis and jellyfish collagen peptide obtained by hydrolyzing jellyfish collagen may be used. You can also.

<1> Gly-Pro-Ala-Gly (SEQ ID NO: 1)
<2> Pro-Gly-Val-Lys-Gly-Glu-Val-Gly-Glu-Ala-Gly-Lys-Arg-Gly (SEQ ID NO: 2)
<3> Val-Gly-Pro-Val-Thr-Glu-Glu-Val-Ser-Pro-Ala-Lys (SEQ ID NO: 3)
<4> Ser-Ala-Gly-Gln-Asn-Pro-Ala-Asp-Asp-Val (SEQ ID NO: 4)
<5> Gly-Thr-Pro (OH) -Gly-Ala-Gly-Gly-Ser-Arg (SEQ ID NO: 5)
<6> Gln-Gly-Pro-Gln-Gly-Glu-Leu (SEQ ID NO: 6)

  Peptide degrading enzymes or solid acid catalysts can be used for hydrolysis of the jellyfish collagen of the present invention, and jellyfish collagen can be obtained from jellyfish protein.

  There are no particular limitations on the type of jellyfish used in the present invention, and for example, jellyfish belonging to the Scyphozoa, Boxbo (Cubozoa), Cross jellyfish (Staurozoa), Hydrozoa, etc. can be used. Specific examples include moon jellyfish, red jellyfish, bizen jellyfish, Echizen jellyfish, Efila jellyfish, kingfisher jellyfish, black camphor jellyfish, Amakusa jellyfish, willow jellyfish, yellow jellyfish, Amaga jellyfish, samurai jellyfish, shrimp jellyfish, scorpion jellyfish, purple jellyfish, etc. Can be used. Among these, from the viewpoint of abundant resources, moon jellyfish and ethene jellyfish can be preferably used, and moon jellyfish can be particularly preferably used. For example, jellyfish collagen containing any one of the amino acid sequences <1> to <6> can be preferably used in the present invention.

First, a method for extracting protein from jellyfish will be described.
When extracting protein from a jellyfish, what was crushed using a cutting machine, a mincing machine, a cutter, etc. can be used, and it can also be used without crushing a jellyfish. Place jellyfish or crushed jellyfish in a low temperature of about 4 ° C. for 1 to 36 hours, or in a phosphate buffer solution of about 4 ° C. or a sodium chloride solution of about 0.2%, or water. By soaking for 1-36 hours, an aqueous solution containing protein is obtained.

  The aqueous solution containing the protein can be filtered, and further impurities can be removed by salting out using an inorganic salt such as sodium chloride or ammonium sulfate, or by precipitation with a lower alcohol such as ethanol.

  The method for obtaining jellyfish collagen from the crude protein thus obtained is not particularly limited. For example, a solid obtained by centrifuging an aqueous solution containing the protein is dispersed in a lower alcohol such as ethanol, and further centrifuged. By this operation, jellyfish collagen can be obtained as a solid. A product obtained by dispersing this jellyfish collagen with water and desalting it with a dialysis membrane or the like can also be used.

Water can be added to the solid material obtained by centrifugation to disperse it, followed by further centrifugation, and the jellyfish collagen in the supernatant and the precipitated jellyfish collagen can be used individually. In the present specification, the precipitated jellyfish collagen is referred to as “acid-soluble collagen” because it is transparently dissolved in an acidic solution having a pH of 1 to 4. This acidic soluble collagen is insoluble in water in the range of pH 4 to 10, but can be dispersed in an alkaline region of pH 10 or higher. Moreover, in this specification, the jellyfish collagen in a supernatant is called "neutral soluble collagen" with respect to acidic soluble collagen. The neutral soluble collagen is soluble in water regardless of the pH range.

  In producing a jellyfish collagen peptide, neutral soluble collagen can be preferably used rather than acidic soluble collagen. In addition, the ratio of acidic soluble collagen can be reduced by removing a metal component beforehand with a chelating agent.

(Hydrolysis of jellyfish collagen by peptide-degrading enzyme)
As used herein, the term “peptide degrading enzyme” refers to an enzyme that degrades a protein to produce a peptide. The molecular weight of the peptide obtained by decomposition is preferably 150 to 35,000 Da, and more preferably 250 to 30,000 Da.

  The peptide degrading enzyme used for hydrolysis of jellyfish collagen is not particularly limited, but is preferably at least one selected from the group consisting of trypsin, pepsin, and papain.

Among them, trypsin is a protease having a molecular weight of about 23,000, secreted from the pancreas as an inactive precursor trypsinogen, and changed to an active form by self-limited degradation. In biological experiments, it is used not only for structural analysis of proteins by specifically hydrolyzing the C-terminal side of arginine and lysine, but also for passage of adherent cells. The amount of trypsin used is preferably 0.0001 to 5% by mass, more preferably 0.001 to 2% by mass, and 0.01 to 1% by mass of the jellyfish collagen as a substrate. Further preferred. Although there is no restriction | limiting in particular in the processing conditions by a trypsin, It is preferable that they are pH 7-9 and 30-40 degreeC.

  Pepsin is a kind of protease having a molecular weight of about 35,000, and the precursor pepsinogen secreted in the vertebrate gastric fluid is converted into pepsin by the action of hydrochloric acid or existing pepsin. The amount of pepsin used is preferably 0.0001 to 5% by mass, more preferably 0.001 to 2% by mass, and 0.01 to 1% by mass of the jellyfish collagen as a substrate. Further preferred. Although there is no restriction | limiting in particular in the processing conditions by pepsin, It is preferable that they are pH 1-4 and 30-40 degreeC.

  Papain is a protease having a molecular weight of about 23,000 contained in papaya pulp, and is an enzyme used for digestives, beer clarifiers, and the like. The amount of papain used is preferably 0.0001 to 5% by mass, more preferably 0.001 to 2% by mass, and 0.01 to 1% by mass of jellyfish collagen as a substrate. Further preferred. Although there is no restriction | limiting in particular in the process conditions by a papain, It is preferable that it is pH 4-10, 20-80 degreeC.

  These peptide degrading enzymes (trypsin, pepsin, papain) can be used alone or in combination of two or more, but it is preferable to use two or more peptidases.

When jellyfish collagen is hydrolyzed with two or more types of peptide degrading enzymes, if jellyfish collagen is acidic soluble collagen or a mixture of acidic soluble collagen and neutral soluble collagen, first hydrolyze with pepsin. Is preferred. By hydrolyzing with pepsin that is active under acidic conditions, a collagen peptide that is soluble even in a neutral pH range can be obtained, and further hydrolyzed with trypsin and / or papain. After the hydrolysis with pepsin, the method of further hydrolyzing with trypsin and / or papain can also be used for the hydrolysis of neutral soluble collagen. For example, a combination of pepsin and trypsin, pepsin, papain and trypsin can be preferably used in the present invention.

(Jellyfish collagen hydrolysis by solid acid catalyst)
The mixture of jellyfish collagen peptides can also be obtained by hydrolyzing jellyfish collagen with a solid acid catalyst. The hydrolysis with a solid acid catalyst is a method of hydrolyzing jellyfish collagen by bringing the solid acid catalyst into contact with jellyfish collagen in the presence of water and under heating. From the viewpoint of hydrolysis efficiency, the jellyfish collagen is preferably neutral soluble collagen.

  In the present invention, a solid catalyst that can function as an acid can be used as a solid acid catalyst without any particular limitation, but from the viewpoint of hydrolysis efficiency, it is at least one selected from the group consisting of a cation exchanger, zeolite, and diatomaceous earth. Preferably, it is a cation exchanger. These solid acid catalysts can be used alone or in combination of two or more.

  The cation exchanger is preferably a resin having at least one of a sulfone group and a carboxyl group, and the sulfone group and the carboxyl group may be a sulfopropyl group and a carboxymethyl group, respectively. When the counter ion of the cation exchanger is other than proton, it is preferable to use it after substitution with the proton type. A particularly preferred cation exchanger is a sulfonic acid type cation exchanger having a strongly acidic cation exchange group such as a sulfopropyl group, and a cation exchange resin and a cation exchange membrane can be mentioned as preferred forms. Specific examples of the sulfonic acid type cation exchange resin include those containing a hydrophilic vinyl polymer as a base material such as TOYOPEARL SP-650C and SP-550C manufactured by Tosoh Corporation, and Nafion (registered trademark). A polytetrafluoroethylene copolymer containing perfluorosulfonic acid can be mentioned.

As zeolite, zeolite which is generally used as a zeolite catalyst can be used, and among them, Zeorum (registered trademark) manufactured by Tosoh Corporation can be preferably used.
Examples of diatomaceous earth include diatomaceous earth (granular) manufactured by Wako Pure Chemical Industries, Ltd.

  There is no restriction | limiting in the shape of the said solid acid catalyst, Arbitrary forms, such as a granular form and a powder form, can be taken. If it is granular or powdery, it can be used in a batch method in which a solid acid catalyst is mixed in an aqueous solution containing jellyfish collagen to be hydrolyzed, and the jellyfish collagen aqueous solution is continuously fed to a column packed with the solid acid catalyst. It can also be used in a continuous process. Further, the jellyfish collagen aqueous solution can be passed through a membrane made of a solid acid catalyst or a membrane on which a granular or powdery solid acid catalyst is immobilized.

  As the solid acid catalyst, for example, one having an average particle diameter of about 2 μm to 2 mm and an ion exchange capacity of about 0.01 to 1 eq / L can be used. The hydrolysis of jellyfish collagen in the present invention is considered to proceed by the action of acid sites on the solid acid catalyst. Therefore, it is preferable to use a solid acid catalyst having a high jellyfish collagen adsorption capacity, and it is preferable to use a solid acid catalyst having a jellyfish collagen adsorption amount of 5 to 150 g / L. For example, the above-mentioned TOYOPEARL SP-650C manufactured by Tosoh Corporation has a jellyfish collagen adsorption amount of about 35 to 55 g / L, and the TOYOPEARL SP-550C has a jellyfish collagen adsorption amount of about 80 to 120 g / L.

  The solid acid catalyst is more preferably a porous body. From the viewpoint of improving the adsorptivity as the specific surface area increases, it is preferable that the porous body has a small pore size, but hydrolysis of jellyfish collagen larger than the pore size in the pores becomes difficult. It is preferable to select a solid acid catalyst having pores of a size suitable for hydrolysis of jellyfish collagen. The preferable pore diameter of the porous body used as the solid acid catalyst is 0.01 to 0.75 μm, and more preferably 0.05 to 0.6 μm.

  Hydrolysis of jellyfish collagen with a solid acid catalyst is carried out by contacting the solid acid catalyst with jellyfish collagen in the presence of water and under heating. Without heating, it is difficult to proceed the hydrolysis reaction well. The heating temperature is preferably about 80 to 160 ° C., and the treatment time is preferably about 5 minutes to 40 hours. It is important that the degradation of jellyfish collagen is limited to the degradation of jellyfish collagen peptide, but when treated at high temperature for a long time, jellyfish collagen may be hydrolyzed to amino acids. The processing time is preferably short. More preferable treatment conditions are 80 to 120 ° C. for 5 minutes to 20 hours, more preferably 80 to 120 ° C. for 5 minutes to 5 hours, more preferably 90 to 110 ° C. for 5 minutes to 2 hours. Yes (see also for example WO 2010/058590 for solid acid catalysts).

  These jellyfish collagen peptide mixtures obtained by hydrolysis using a peptide-degrading enzyme or a solid acid catalyst have increased keratinocyte activation activity, involucrin production promoting activity, transglutaminase compared to jellyfish collagen not hydrolyzed It can exhibit production promoting activity and / or anti-inflammatory activity.

  In the present invention, a peptide containing any one of the amino acid sequences <1> to <6> can be synthesized. There is no particular limitation on the method for synthesizing the peptide containing the amino acid sequence. For example, the amino group can be extended by fixing the C-terminal amino acid to a solid phase made of polystyrene and bonding a protected amino acid thereto, thereby protecting the amino acid. For example, an Fmoc (Fluorenyl-MethOxy-Carbonyl) group or a Boc (tert-ButOxy-Carbonyl) group can be used. After completion of the reaction between the amino group of the solid phase and the protected amino acid, the solid phase is preferably washed to remove the unreacted protected amino acid. Then, the amino group used as the next reaction point is obtained by removing (deprotecting) the protective group at the terminal of the amino acid bonded to the solid phase. A peptide containing the target amino acid sequence can be obtained by sequentially changing the protected amino acid to be used. The peptide of the present invention has a keratinocyte increased by, for example, 1.2 times, preferably 1.5 times, more preferably 1.8 times, and even more preferably 2 times compared to non-hydrolyzed jellyfish collagen. It exhibits activation activity, involucrin production promoting activity, transglutaminase production promoting activity and / or anti-inflammatory activity. The peptide comprising any one of the amino acid sequences <1> to <6> may be a peptide consisting of any one of the amino acid sequences <1> to <6>.

  The jellyfish collagen peptide excellent in cell activation effect, involucrin production promotion effect, transglutaminase production promotion effect and anti-inflammatory effect of the present invention can be transdermally administered in the form of an external preparation such as an external preparation for skin, cosmetics or bath preparation. it can. From the viewpoint of transdermal administration, the molecular weight of the jellyfish collagen peptide is preferably 150 to 35,000 Da, and more preferably 250 to 30,000 Da.

There is no particular limitation on the dosage form when the mixture of the jellyfish collagen peptide of the present invention is formulated as an external preparation into an external preparation for skin or cosmetics. Paste, cream, gel, ointment, lotion, emulsion, pack, powder, papp It can also be used as an agent, and can also be used by impregnating paper products such as wet tissue. Although there is no restriction | limiting in particular in the quantity of the jellyfish collagen peptide mix | blended with a skin external preparation and cosmetics, It is preferable that it is 0.0005-50 mass% in skin external preparations and cosmetics. Although there is no restriction | limiting in particular in the usage-amount of the skin external preparation and cosmetics which mix | blended the jellyfish collagen peptide of this invention, The quantity which becomes 0.1-1000 mg per day for an adult in a jellyfish collagen peptide is divided into 1 to several times. It is preferable to use it.

  There is no limitation on the dosage form when the jellyfish collagen peptide mixture of the present invention is formulated into a bath preparation as an external preparation, and it can be used as a solid preparation such as powder or granule, a liquid preparation such as emulsion or paste. . Although there is no restriction | limiting in particular in the quantity of the jellyfish collagen peptide which can be mix | blended with a bath agent, It is preferable that it is 0.0005-50 mass% in a bath agent.

  In addition to the jellyfish collagen peptide of the present invention, the external preparations such as skin external preparations, cosmetics and bath preparations of the present invention include known functional ingredients used in skin external preparations, cosmetics or bath preparations, such as glycerin, Moisturizers such as butylene glycol, urea and amino acids; emollients such as squalane, macadamia nut oil and jojoba oil; blood circulation promoters such as vitamin E and pepper tincture; dibutylhydroxytoluene (BHT), dibutylhydroxyanisole (BHA), Anti-inflammatory agents such as tocopherol acetate and ascorbic acid; anti-inflammatory agents such as glycyrrhizin and allantoin; antibacterial agents such as hinokitiol, benzalkonium chloride, chlorhexidine salt and parahydroxybenzoate; whitening agents such as ascorbic acid and arbutin; super Oxide Sumutaze (SOD) peroxide inhibitors such as, various known materials such as such can be blended. Moreover, various extracts derived from plants, animals, and microorganisms such as ginkgo biloba extract, placenta extract, and lactic acid bacteria culture extract can be added.

  In addition, in the external preparations such as the above-mentioned external preparations for skin, cosmetics, bath preparations, base components such as surfactants and fats and oils can be blended for the formulation, and thickeners are added as necessary. Various additives such as preservatives, tonicity agents, antioxidants, ultraviolet absorbers, chelating agents, fragrances, and coloring agents can be used in combination.

  There is no limitation on the surfactant that can be used, alkylene oxide adduct of higher alkylamine, alkylene oxide adduct of higher fatty acid amide, fatty acid ester of polyhydric alcohol, alkylene oxide adduct of hardened castor oil, polyethylene glycol sorbitan alkyl ester, Nonionic surfactants such as alkylene oxide adducts such as sterols, anionic surfactants such as sodium alkyl sulfate, sodium alkyloxyl taurate, sodium α-olefin sulfonate, sodium polyoxyalkylene alkyl ether sulfate, alkylpyridinium chloride , Cationic surfactants such as distearyldimethylammonium chloride, amphoteric surfactants such as sodium alkylaminopropionate and alkylpolyaminoethylglycine It can be used. These surfactants can be used alone or in combination of two or more.

  There is no restriction | limiting in particular in the component used as the said base, Oils and fats, such as olive oil, camellia oil, jojoba oil, avocado oil, macadamia nut oil, apricot oil, squalane, squalene, horse oil, can be used.

  The component used as the thickener is not particularly limited, but a water-soluble polymer is preferable, for example, polyvinyl alcohol, polyacrylamide, polyethylene glycol, and derivatives thereof; celluloses such as hydroxyalkyl cellulose and derivatives thereof; dextran, Gums such as gelatin, gum arabic, and tragacanth; carboxyvinyl polymer and the like can be used.

  There is no restriction | limiting in particular in the component used as said preservative, For example, parahydroxybenzoic acid ester, an alkyl pyridinium chloride, benzalkonium chloride, a chlorhexidine salt, hinokitiol etc. can be used.

  There is no restriction | limiting in particular in the component used as said tonicity agent, For example, inorganic salts, such as sodium chloride, potassium chloride, and calcium chloride, can be used.

  There is no restriction | limiting in particular in the component used as the said ultraviolet absorber, For example, a paraamino benzoic acid, a benzophenone series ultraviolet absorber, a benzotriazole type ultraviolet absorber etc. can be used.

  There is no restriction | limiting in particular in the chelating agent which can be used, For example, ethylenediaminetetraacetic acid, phytic acid, a citric acid, and these water-soluble salts can be mentioned.

Hereinafter, the present invention will be described more specifically with reference to examples, but the present invention is not limited thereto.
1. Preparation of Jellyfish Collagen A 1 Kg population of moon jellyfish (collected from: Tokyo Bay) washed with distilled water was cut into 5 mm to 1 cm square pieces. This was stirred at 4 ° C. for 24 hours to obtain a protein-containing solution. This was filtered with gauze, and an equal volume of local ethanol (A) was added to the filtrate and stirred to obtain a precipitate. The same amount of local ethanol as local ethanol (A) was again added to the precipitate, followed by centrifugation (10000 × G). The obtained solid was dispersed in distilled water and subjected to desalting using a dialysis membrane having a molecular weight cut-off (MWCO) of 14,000. This was centrifuged (10000 × G) and separated into a supernatant layer and a precipitate layer to obtain neutral soluble collagen (supernatant) and acidic soluble collagen (precipitate).

2. Amino acid analysis of extracted neutral soluble collagen In the amino acid analysis, 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) was used as a fluorescent derivatization reagent,
10 μg of lyophilized neutral soluble collagen was dissolved in 10 μL of water and dried in a glass tube (6 × 32 mm, CHROMACOL 03-CVG).

  In a glass vial (WHEATON 225289), 200 μL of 6N hydrochloric acid added with a piece of phenol crystal and the glass tube were put, and the glass vial was sealed with a rotary vacuum pump under reduced pressure. This glass vial was set in an aluminum block heater and heated at 110 ° C. for 20 hours to hydrolyze neutral soluble collagen into amino acids.

  To the hydrolyzate (amino acid) of neutral soluble collagen, 10 μL of 20 mM hydrochloric acid, 30 μL of 0.2 M borate buffer (pH 8.8) and 10 μL of AQC solution (AQC 3 mg / ml acetonitrile solution) were added, and the mixture was heated to 55 ° C. The mixture was heated for 10 minutes, and the collagen hydrolyzate (amino acid) was converted to an amino acid fluorescent derivative and separated by high performance liquid chromatography (HPLC).

a) HPLC apparatus: Agilent 1200 series (including vacuum degasser G1379B, binary pump G1312B, autosampler G1329B, column thermostat G1316B, diode array detector G1315C, and fluorescence detector G1321A)
b) Column: Inertsustain C18HP (3.0 × 250 mm, 3 μm), manufactured by GL Sciences Inc. c) Mobile phase A solution: 95% 5 mM tetrabutylammonium bromide-30 mM phosphate buffer (pH 7.3) / 5% acetonitrile B Solution: 50% 30 mM phosphate buffer solution (pH 7.3) / 50% acetonitrile Starting from solution A 98%, solution B 2%, solution B after 3.75 minutes to 7.3%, and further 50 minutes A concentration gradient was provided so that the later B liquid was 72.3%.
d) Flow rate: 0.4 ml / min e) Temperature: 50 ° C
f) Fluorescence excitation wavelength (Ex): 250 nm
g) Monitor wavelength (Em): 395 nm
Amino acid was quantified using Pierce's Amino Acid Standard H (2.5 μmol / ml) in 10 pmol / μL as a standard amino acid solution. The results are shown in the following table.

Jellyfish collagen contained 30% or more Gly.
* Lys (OH), 3Pro (OH), and 4Pro (OH) each represent a hydroxyl group-containing amino acid.

3. Measurement of metal content of extracted jellyfish collagen 0.1 g of lyophilized neutral soluble collagen was precisely weighed, 10 ml of nitric acid was added, and this was wet-decomposed with microwaves (made by MARS5 CEM). The decomposition solution was made up to 50 ml, and the metal content was measured with an emission spectrophotometer (Optima 5300 DV manufactured by PerkinElmer). The same operation was performed on acidic soluble collagen.

4). Preparation of Jellyfish Collagen Peptide, Molecular Weight Distribution, Mass Spectrometry, and Analysis of Amino Acid Sequence The molecular weight distribution of the jellyfish collagen peptide mixture in the following examples was measured using gel filtration chromatography (column: TSKgel Super SW2000, Tosoh).
Solvent: 6M guanidine hydrochloride / 10 mM phosphate buffer (pH 6)
Flow rate: 0.15 ml / min Measurement wavelength: 215 nm

  Moreover, the obtained jellyfish collagen peptide mixture was isolate | separated by the reverse phase chromatography, and mass spectrometry (Ultraflex TOF / TOF, BRUKER DALTONICS) and amino acid sequence analysis (PROCISE 494cLC protein sequencer, Applied Biosystems) were performed.

Separation conditions by reverse phase chromatography Column: Inertsil ODS-3 (manufactured by GL Sciences)
Measurement wavelength: 215 nm
Flow rate 20 μl / min Mobile phase A solution: 0.085% trifluoroacetic acid aqueous solution B solution: 0.075% trifluoroacetic acid / 80% acetonitrile solution Starting from solution A 100%, solution B was 12.5% in 10 minutes. The concentration gradient was set to 50% at 70 minutes and 100% at 75 minutes.

  For specific peaks such as the peak with a molecular weight of 1112.498 in Example 1, a concentration gradient was set so that the liquid B started at 100% and the liquid B reached 100% in 20 minutes.

4-1) Preparation of jellyfish collagen peptide by peptide degrading enzyme Example 1)
0.01 g of lyophilized neutral soluble collagen was dissolved in 10 mL of water and adjusted to pH 2 with 1N HCl. 0.5% by mass of pepsin (reagent, manufactured by Nacalai, Pepsin (1: 10,000)) was added to collagen and reacted at 37 ° C. for 3 hours. Thereafter, the pH was adjusted to 7 with 1N NaOH to inactivate pepsin. Next, 0.5% by mass of papain (reagent manufactured by Wako Pure Chemical Industries, Ltd.) is added to collagen and reacted at 60 ° C. for 3 hours. Reagent) was added and reacted at 37 ° C. for 3 hours. Thereafter, the enzyme was inactivated by maintaining at 95 ° C. for 30 minutes. This was freeze-dried to obtain a mixture of collagen peptides having a molecular weight of 500 to 6000 Da.

As a result of mass spectrometry and amino acid sequence analysis, the collagen peptide of Example 1 contains a collagen peptide having a molecular weight of 1112.498. This collagen peptide has an amino acid sequence <1> Gly-Pro-Ala-Gly (SEQ ID NO: 1).
Confirmed to contain.

  The results of mass spectrometry of Example 1 are shown in FIG. 1, and the relationship between the amino acid sequence having a molecular weight of 1112.498 and the predicted fragment mass of tandem mass spectrometry is shown in Table 3 below.

Example 2)
Except that the neutral soluble collagen of Example 1 was changed to acidic soluble collagen, the same operation as in Example 1 was performed to obtain a mixture of collagen peptides having a molecular weight of 500 to 6000 Da.

Example 3)
0.01 g of lyophilized neutral soluble collagen was dissolved in 10 ml of water and adjusted to pH 2 with 1N HCl. 0.5% by mass of pepsin (reagent, manufactured by Nacalai, Pepsin (1: 10,000)) was added to collagen and reacted at 37 ° C. for 3 hours. Thereafter, the pH was adjusted to 8 with 1N NaOH to inactivate pepsin. This was freeze-dried to obtain a mixture of collagen peptides having a molecular weight of 2 kDa to 30 kDa.

Example 4)
A mixture of collagen peptides having a molecular weight of 2 kDa to 30 kDa was obtained in the same manner as in Example 3 except that the neutral soluble collagen of Example 3 was changed to acidic soluble collagen.

Example 5)
In the operation of Example 3, after inactivating pepsin, 0.5% by mass of trypsin (a reagent manufactured by Wako Pure Chemical Industries, Ltd.) was added to collagen and reacted at 37 ° C. for 3 hours. Subsequently, trypsin was inactivated by treatment at 95 ° C. for 30 minutes. This was freeze-dried to obtain a mixture of collagen peptides having a molecular weight of 500 to 6000 Da.

Example 6)
A mixture of collagen peptides having a molecular weight of 500 to 6000 Da was obtained by performing the same operation as in Example 5 except that the neutral soluble collagen of Example 5 was changed to acidic soluble collagen.

Example 7)
In the operation of Example 3, after reacting at 37 ° C. for 3 hours, pH was adjusted to 7 with 1N NaOH to inactivate pepsin, and then 0.5% by mass of papain (manufactured by Wako Pure Chemical Industries, Ltd.) with respect to collagen. Reagent) and reacted at 60 ° C. for 3 hours. Thereafter, papain was inactivated at 95 ° C. for 30 minutes, and a mixture of collagen peptides having a molecular weight of 1 kDa to 20 kDa was obtained by lyophilization.

Example 8)
A mixture of collagen peptides having a molecular weight of 1 kDa to 20 kDa was obtained in the same manner as in Example 7 except that the neutral soluble collagen of Example 7 was changed to acidic soluble collagen.

Preparation of jellyfish collagen peptide by solid acid catalyst Example 9)
After mixing 0.5 g of cation exchange resin (TOYOPEARL SP-550C) and 0.5 mL of 1% by mass neutral soluble collagen aqueous solution to adsorb neutral soluble collagen to the cation exchange resin, 90 ° C. For 30 minutes to hydrolyze neutral soluble collagen. After cooling, the decomposition product was extracted from the resin with 5% triethylamine / 10% acetonitrile solution. This was freeze-dried to obtain a mixture of collagen peptides having a molecular weight of 2 kDa to 20 kDa.

Moreover, as a result of mass spectrometry and amino acid sequence analysis of the collagen peptide of Example 9, the collagen peptide having a molecular weight of 1950.915 was included, and the collagen peptide having a molecular weight of 1950.915 was
Amino acid sequence <2> Pro-Gly-Val-Lys-Gly-Glu-Val-Gly-Glu-Ala-Gly-Lys-Arg-Gly (SEQ ID NO: 2)
Confirmed to contain.

  The results of mass spectrometry of Example 9 are shown in FIG. 2, and the relationship between the amino acid sequence having a molecular weight of 1950.915 and the predicted fragment mass of tandem mass spectrometry is shown in Table 4 below.

Example 10)
A mixture of collagen peptides having a molecular weight of 6 kDa to 100 kDa was obtained in the same manner as in Example 9, except that the heating time in Example 9 was changed to 15 minutes.

Example 11)
A mixture of collagen peptides having a molecular weight of 1 kDa to 10 kDa was obtained by performing the same operation as in Example 9, except that the heating time of Example 9 was changed to 60 minutes.

Example 12)
A mixture of collagen peptides having a molecular weight of 1 kDa to 10 kDa was obtained in the same manner as in Example 9, except that the heating temperature of Example 9 was 110 ° C. and the heating time was 5 minutes.

Example 13)
Except that the heating time of Example 12 was 30 minutes, the same operation as in Example 12 was performed to obtain a mixture of collagen peptides having a molecular weight of 1 kDa to 5 kDa.

Example 14)
Except that the heating time of Example 12 was 60 minutes, the same operation as in Example 12 was performed to obtain a mixture of collagen peptides having a molecular weight of 200 to 2000 Da.

Example 15)
Except that the heating time of Example 12 was 2 hours, the same operation as in Example 12 was performed to obtain a mixture of collagen peptides having a molecular weight of 200 to 1000 Da.

Example 16)
Except that the heating time of Example 12 was changed to 4 hours, the same operation as in Example 12 was performed to obtain a mixture of collagen peptides having a molecular weight of 200 to 1000 Da.

  The molecular weight distributions of Examples 12 to 14 are shown in FIG.

5. Analysis of cyanogen bromide-degrading collagen peptide 10% by mass of cyanogen bromide was added to lyophilized neutral soluble collagen and reacted at 37 ° C for 20 hours to hydrolyze neutral soluble collagen. A decomposition product was obtained. Hydrolysis with cyanogen bromide that cleaves at the C-terminal side of methionine is a method that has high specificity and is generally used for analysis of amino acid sequences. The neutral soluble collagen hydrolyzate was concentrated and centrifuged to volatilize cyanogen bromide, then redissolved in water, and SDS-polyacrylamide using an acrylamide gel (manufactured by Biocraft) with a gel concentration of 5 to 20% by mass. Separation was performed by gel electrophoresis. The obtained gel information was blot transferred onto a PVDF membrane at a voltage of 60 V for 4 hours, and the transferred band was analyzed with a protein sequencer (PROCISE 494HT, Applied Biosystems) to obtain the following amino acid sequence information.

Amino acid sequence <3> Val-Gly-Pro-Val-Thr-Glu-Glu-Val-Ser-Pro-Ala-Lys (SEQ ID NO: 3)
Amino acid sequence <4> Ser-Ala-Gly-Gln-Asn-Pro-Ala-Asp-Asp-Val (SEQ ID NO: 4)
Amino acid sequence <5> Gly-Thr-Pro (OH) -Gly-Ala-Gly-Gly-Ser-Arg (SEQ ID NO: 5)
Amino acid sequence <6> Gln-Gly-Pro-Gln-Gly-Glu-Leu (SEQ ID NO: 6)
Pro (OH) represents hydroxyproline.

The amino acid sequence <1> (degraded product by peptide degrading enzyme), amino acid sequence <2> (degraded product by solid acid catalyst), and the above amino acid sequences <3> to <6> are all contained in jellyfish collagen. It is an amino acid sequence, and jellyfish collagen before hydrolysis is amino acid sequence <
It has all of 1> to <6>. Therefore, the jellyfish collagen peptide mixture obtained by hydrolyzing the jellyfish collagen contains a collagen peptide containing amino acid sequences <1> to <6> regardless of the means of degradation.

  Among these six kinds of amino acid sequences, a synthetic product of peptides consisting of amino acid sequences <3> to <6> (consignment of production to Hokkaido System Science Co., Ltd.) was prepared. did.

Example 17) Val-Gly-Pro-Val-Thr-Glu-Glu-Val-Ser-Pro-Ala-Lys (SEQ ID NO: 3)
Example 18) Ser-Ala-Gly-Gln-Asn-Pro-Ala-Asp-Asp-Val (SEQ ID NO: 4)
Example 19) Gly-Thr-Pro (OH) -Gly-Ala-Gly-Gly-Ser-Arg (SEQ ID NO: 5)
Example 20) Gln-Gly-Pro-Gln-Gly-Glu-Leu (SEQ ID NO: 6)

6). Evaluation test 1 of the effect of jellyfish collagen peptide (activation test using keratinocytes)
The cell activation effect was evaluated by the cell proliferation effect using activated cultured cells and the cell proliferation reagent WST-1 (Roche).

2 × 10 ⁴ human skin-derived immortalized keratinocytes PHK16-0b distributed by the Human Science Promotion Foundation were seeded on a 48-well multiplate, and 300 μL of KGM medium was added and cultured for 2 days. Thereafter, the medium was replaced with 300 μL of KGM medium having a jellyfish collagen concentration of 10 μg / mL, and the cells were cultured in an incubator at 37 ° C. for 7 days. The medium was replaced with 270 μL of new KGM medium again, 30 μL of cell growth reagent WST-1 was added, and the mixture was reacted in a 37 ° C. incubator for 3 hours, and then the absorbance at 440 nm of the medium was measured. The cell growth rate of the control cultured under the same conditions without adding the collagen peptide was taken as 100%, and the cell growth rate of the collagen peptide was evaluated (n = 3). The results are shown below.

  Jellyfish collagen peptide obtained under any hydrolysis conditions was confirmed to have a cell growth effect. Peptide degrading enzymes are particularly useful for collagen peptides obtained by hydrolysis with pepsin and then with papain and trypsin. Excellent effect was observed.

  Based on the amino acid sequence information of jellyfish collagen, the jellyfish collagen peptide obtained with the solid acid catalyst has a good cell growth effect, and with 110 ° C treatment, good results were obtained with the jellyfish collagen peptide with a treatment time of 30 minutes or less. The cell proliferation effect of the peptide produced in Example 1 was also good, and in particular, the excellent effect was confirmed in Example 17. On the other hand, for the jellyfish collagen itself, the cell growth effect could not be confirmed for both neutral soluble collagen (Comparative Example 1) and acidic soluble collagen (Comparative Example 2).

Evaluation test 2 (Involucrin expression level test using keratinocytes)
Adult human skin-derived normal keratinocytes (manufactured by Kojin Bio) were seeded on a 48-well multiplate at 1 × 10 ⁵ cells, and 300 μL of KGM medium was added thereto and cultured at 37 ° C. for 24 hours. Jellyfish collagen peptide concentration 10 μg The medium was replaced with 300 μL / mL KGM medium and cultured at 37 ° C. for 48 hours. Cells were collected, and involucrin production was measured by Western blotting. In addition, anti-human involucrin (Cosmo Bio, sc-28557) was used as the primary antibody, and anti-rabbit IgG-HRP (Cosmo Bio, sc-2004) was used as the secondary antibody. The results were analyzed by ImageJ, and the involucrin expression rate relative to the additive-free group (control) was evaluated. The results are shown below.

A good involucrin expression effect was confirmed in a mixture of jellyfish collagen peptides obtained by hydrolyzing jellyfish collagen with a peptide degrading enzyme and a solid acid catalyst. A peptide obtained by synthesis based on amino acid sequence information was also found to have an involucrin production effect.
Evaluation test 3 (Transglutaminase 1 expression level test using keratinocytes)
20 × 10 ⁴ human skin-derived immortalized keratinocytes PHK16-0b were seeded on a 48-well multiplate, and 300 μL of KGM medium was added thereto and cultured at 37 ° C. for 24 hours. After replacing with 300 μL of KGM medium having a jellyfish collagen peptide concentration of 10 μg / mL and culturing at 37 ° C. for 48 hours, the cells were collected, and the amount of transglutaminase 1 produced was measured by Western blotting. Note that anti-transglutaminase 1 (Cosmo Bio, PAB15545) was used as the primary antibody, and anti-rabbit IgG-HRP (Cosmo Bio, sc-2004) was used as the secondary antibody. The results were analyzed by ImageJ, and the transglutaminase 1 expression rate relative to the non-added group (control) was evaluated. The results are shown below.

  A favorable transglutaminase 1 expression effect could be confirmed in a mixture of jellyfish collagen peptides obtained by hydrolyzing jellyfish collagen with a peptide degrading enzyme. The effect of producing transglutaminase 1 was also confirmed in a jellyfish collagen peptide mixture obtained by hydrolysis with a solid acid catalyst and a peptide obtained by synthesis based on amino acid sequence information.

Evaluation test 4 (COX-2 inhibitory effect test using keratinocytes)
The anti-inflammatory effect was evaluated by the amount of PGE ₂ produced by COX-2 of keratinocytes that had received ultraviolet irradiation. The smaller the amount of PGE ₂ produced, the more the jellyfish collagen peptide has anti-inflammatory properties.

20 × 10 ⁴ human skin-derived immortalized keratinocytes PHK16-0b were seeded on a 48-well multiplate, and 300 μL of KGM medium was added and cultured overnight. The medium was replaced with 300 μL of KGM medium having an aspirin concentration of 500 μM and cultured for 4 hours to inactivate COX-2 that had already been expressed. Washing with PBS (−) was repeated three times, and the medium was replaced with a DMEM medium with a jellyfish collagen peptide concentration of 10 μg / mL. After irradiation with 50 mJ of UVB, the cells were cultured for 48 hours. Thereafter, the amount of PGE ₂ in the culture supernatant was measured with a prostaglandin E ₂ EIA kit (manufactured by Cayman Chemical) (n = 3).

A good anti-inflammatory effect can be confirmed in the jellyfish collagen peptide mixture, and in particular, an excellent anti-inflammatory effect should be confirmed in the jellyfish collagen peptide mixture obtained by degrading jellyfish collagen with pepsin and then further digesting with trypsin. I was able to.
7). Formulation Example for Cosmetics Each of the A component and the B component was heated to 80 ° C. to dissolve, and the B component was gradually added to the A component and emulsified with stirring. Thereafter, the mixture was cooled with stirring, and a C component adjusted to a 0.2 mass% aqueous solution at 40 ° C. was added to prepare a lotion for whole body.
(A) component
Grape seed oil 0.1 (g)
Polyoxyethylene (60 mol) hydrogenated castor oil 0.7
Diglyceryl monoisostearate0.3
(B) component
Citric acid 0.08
Na citrate 0.3
1,3-butanediol 5
Glycerin 3
Ethanol 5
Xanthan gum 0.1
Methylparaben 0.2
Purified water Remaining (C) component
Jellyfish collagen peptide (0.2% by mass) 0.5
Total 100

Evaluation A lotion for whole body having the above composition is used twice a day (morning and night) as needed on the face and forearm, and cosmetics containing the jellyfish collagen peptide of Examples 1 to 20 on one side of the face and forearm Or the cosmetics which mix | blended acidic soluble collagen (comparative example 2) and the cosmetics which mix | blended neutral soluble collagen (comparative example 1) were used for the other. This was continued for one month and judged according to the following evaluation criteria. The results are shown in Table 9.
No one complained of skin abnormalities during this period of use.

Score value +3: felt moisturizing stronger than the cosmetics formulated with Comparative Example 1 +2: felt moisturized significantly more than the cosmetics formulated with Comparative Example 1 +1: slightly higher than the cosmetics formulated with Comparative Example 1 Moisture was felt 0: No difference -1: Cosmetics formulated with Comparative Example 1 felt slightly moisturized-2: Cosmetics formulated with Comparative Example 1 felt significantly moisturized -3: The cosmetic containing Comparative Example 1 felt moist and strong

  In any mixture of jellyfish collagen peptides, it was possible to obtain a skin moisturizing effect that exceeded that of cosmetics containing neutral soluble collagen.

  Peptide degrading enzyme hydrolyzate is a jellyfish collagen peptide mixture obtained by hydrolyzing neutral or acidic soluble collagen with pepsin and then hydrolyzing with papain and trypsin. It was.

  In the hydrolyzate by a solid acid catalyst, a good moistening effect was observed in the jellyfish collagen peptide mixture obtained by the treatment at a treatment temperature of 90 ° C. and a treatment time of 30 minutes.

  Even in cosmetics containing a peptide synthesized based on amino acid sequence information of jellyfish collagen, the skin moisturizing effect was good.

  By using a collagen peptide derived from jellyfish collagen, keratinocyte activation effect, involucrin production promotion effect, transglutaminase production promotion effect and anti-inflammatory effect can be obtained. The barrier function is repaired and improved, and the function of the epidermal keratinocytes can be normalized.

Claims (13)

A jellyfish collagen peptide mixture containing a jellyfish collagen peptide comprising any of the following amino acid sequences.
<1> Gly-Pro-Ala-Gly (SEQ ID NO: 1)
<2> Pro-Gly-Val-Lys-Gly-Glu-Val-Gly-Glu-Ala-Gly-Lys-Arg-Gly (SEQ ID NO: 2)
<3> Val-Gly-Pro-Val-Thr-Glu-Glu-Val-Ser-Pro-Ala-Lys (SEQ ID NO: 3)
<4> Ser-Ala-Gly-Gln-Asn-Pro-Ala-Asp-Asp-Val (SEQ ID NO: 4)
<5> Gly-Thr-Pro (OH) -Gly-Ala-Gly-Gly-Ser-Arg (SEQ ID NO: 5)
<6> Gln-Gly-Pro-Gln-Gly-Glu-Leu (SEQ ID NO: 6)
Pro (OH) represents hydroxyproline.   The jellyfish collagen peptide mixture according to claim 1 obtained by hydrolyzing jellyfish collagen.   The jellyfish collagen peptide mixture according to claim 2, wherein the hydrolysis is performed using at least one peptide degrading enzyme selected from the group consisting of trypsin, pepsin, and papain.   The jellyfish collagen peptide mixture according to claim 2, wherein the hydrolysis is performed using a solid acid catalyst.   The jellyfish collagen peptide mixture according to claim 4, wherein the solid acid catalyst is at least one solid acid catalyst selected from the group consisting of a cation exchanger, zeolite, and diatomaceous earth. The jellyfish collagen peptide mixture according to any one of claims 1 to 5, wherein the jellyfish collagen is a jellyfish collagen or an Echizen jellyfish collagen.   A jellyfish collagen peptide mixture obtained by hydrolyzing jellyfish collagen using a peptide degrading enzyme or a solid acid catalyst. A peptide comprising any of the following amino acid sequences.
<1> Gly-Pro-Ala-Gly (SEQ ID NO: 1)
<2> Pro-Gly-Val-Lys-Gly-Glu-Val-Gly-Glu-Ala-Gly-Lys-Arg-Gly (SEQ ID NO: 2)
<3> Val-Gly-Pro-Val-Thr-Glu-Glu-Val-Ser-Pro-Ala-Lys (SEQ ID NO: 3)
<4> Ser-Ala-Gly-Gln-Asn-Pro-Ala-Asp-Asp-Val (SEQ ID NO: 4)
<5> Gly-Thr-Pro (OH) -Gly-Ala-Gly-Gly-Ser-Arg (SEQ ID NO: 5)
<6> Gln-Gly-Pro-Gln-Gly-Glu-Leu (SEQ ID NO: 6)
Pro (OH) represents hydroxyproline.   The external preparation containing the jellyfish collagen peptide mixture of any one of Claims 1-7, and / or the peptide of Claim 8.   A keratinocyte activator containing the jellyfish collagen peptide mixture according to any one of claims 1 to 7, and / or the peptide according to claim 8.   The involucrin production promoter containing the jellyfish collagen peptide mixture of any one of Claims 1-7, and / or the peptide of Claim 8. The transglutaminase production promoter containing the jellyfish collagen peptide mixture of any one of Claims 1-7, and / or the peptide of Claim 8.   The anti-inflammatory agent containing the jellyfish collagen peptide mixture of any one of Claims 1-7, and / or the peptide of Claim 8.

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