JP2004168732A - Active oxygen-scavenging agent, inhibitor for synthesis promotion/decomposition of hyaluronic acid, and aquaporin synthesis promotor - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To obtain an active oxygen scavenging agent causing excellent moisture retention to the skin, chapped skin-improving, and having aging-preventing and bleaching effects on the skin by scavenging active oxygen, inhibiting the production of lipoperoxides in the skin, promoting hyaluronic acid producing ability of a skin cell and inhibiting the decomposition ability and further promoting aquaporin synthesis of a skin cell, and to obtain an inhibitor for synthesis promotion/decomposition of hyaluronic acid and an aquaporin synthesis promotor. <P>SOLUTION: The active oxygen scavenging agent, the inhibitor for synthesis promotor and decomposition of the hyaluronic acid and the aquaporin synthesis promotor comprise each an extract obtained from a plant of the family Tropaeolaceae. <P>COPYRIGHT: (C)2004,JPO

Description

【００２８】

【００２９】
上記より明らかなように、本発明に係るノウゼンハレン科植物抽出物は極めて低濃度でスーパーオキシドを消去することが認められた。
【００３０】
ヒアルロン酸合成促進・分解抑制効果を明らかにするための実施例を示す。
（１）ヒアルロン酸合成促進効果（実施例２）
・ＭＥＭ培地の調製法
Ｍｉｎｉｍｕｍ Ｅｓｓｅｎｔｉａｌ Ｍｅｄｉｕｍ （大日本製薬社製、１０−１０１） １０．６ｇにそれぞれ終濃度として、１％（Ｖ／Ｖ）Ｎｏｎ Ｅｓｓｅｎｔｉａｌ Ａｍｉｎｏ Ａｃｉｄ（大日本製薬社製、１６−８１０）、１ｍｍｏｌ／Ｌピルビン酸ナトリウム（大日本製薬社製、１６−８２０）、１．２％（Ｗ／Ｖ）炭酸水素ナトリウム、蒸留水を加えて１Ｌとした後、炭酸ガスを吹き込んでｐＨを約７にした（以下、ＭＥＭ培地と略記する）。
【００３１】
・ウシ胎仔血清（ＦＢＳ）の非働化
ＦＢＳ（Ｉｒｖｉｎｅ Ｓｃｉｅｎｔｉｆｉｃ社製）を５６℃で３０分間加熱処理した。
【００３２】
・プロナーゼ溶液
２００μｇ／ｍＬプロナーゼ（カルビオケム−ベーリング・コーポレイション社製、Ｓｔｒｅｐｔｏｍｙｃｅｓ ｇｒｉｓｅｕｓ由来）、０．１５ｍｏｌ／Ｌ塩化ナトリウムおよび０．０２％アジ化ナトリウム含有０．５ｍｏｌ／Ｌトリス塩酸緩衝液（ｐＨ８．０）。
【００３３】
・細胞培養
正常ヒト線維芽細胞株〔デトロイト５５１株（ＡＴＣＣ ＣＣＬ １１０）〕を０．６×１０^５個／ウェルの密度で２４ウェルプレートに播種し、１０％（Ｖ／Ｖ）の非働化ＦＢＳを含むＭＥＭ培地にて９５％（Ｖ／Ｖ）空気−５％（Ｖ／Ｖ）炭酸ガスの雰囲気下、３７℃で３日間静置培養し、さらに無血清ＭＥＭ培地にて１日間培養した。次いで、試料を添加した培地にて３日間培養後、培養上清を回収し上清中のヒアルロン酸量を求めた。
【００３４】
・ヒアルロン酸産生量の測定
培養上清０．６３ｍＬにプロナーゼ溶液０．０７ｍＬを加え、３７℃で１８時間静置した後、１００℃で１０分間加熱処理し、プロナーゼを失活させた。次に、反応液０．０１ｍＬからヒアルロン酸プレート「中外」（中外診断科学社製）を用い、添付説明書に従いヒアルロン酸量を測定した。尚、試料添加は各群ｎ＝３で試験を行い、結果はそれぞれの平均値を用いた。
【００３５】
得られた結果を下記に示す。

【００３６】

【００３７】
上記より明らかなように、本発明に係るノウゼンハレン科植物抽出物は極めて低濃度でヒアルロン酸合成促進効果を有することが認められた。
【００３８】
（２）ヒアルロン酸分解抑制効果（実施例３）
・細胞添加用高分子トリチウムヒアルロン酸の調製方法
正常ヒト線維芽細胞株〔デトロイト５５１株（ＡＴＣＣ ＣＣＬ １１０）〕の細胞数を１０％（Ｖ／Ｖ）の非働化ＦＢＳを含むＭＥＭ培地にて２×１０^５個／ｍＬに調整し、２２５ｃｍ^２のフラスコに５０ｍＬ入れ、３日間培養しコンフルエント状態にした。その後ヒアルロン酸の前駆体であるトリチウムグルコサミン（Ａｍｅｒｉｃａｎ Ｒａｄｉｏｌａｂｅｌｅｄ Ｃｈｅｍｉｃａｌｓ Ｉｎｃ．社製）を培養系に添加し（１０μＣｉ／ｍＬ）、さらに３日間培養したのち、培養液からトリチウムラベルされたヒアルロン酸をＵｎｄｅｒｈｉｌｌらの方法（Ｊ．Ｃｅｌｌ Ｂｉｏｌｏｇｙ，８２巻，４７５頁，１９７９年）によって精製し、さらにゲルろ過カラムにより分子量１００万以上の高分子トリチウムヒアルロン酸（比放射活性，０．１μＣｉ／μｇ）を調製した。これを細胞培養系への添加用高分子トリチウムヒアルロン酸とした。
【００３９】
・正常ヒト線維芽細胞への高分子トリチウムヒアルロン酸の添加培養
正常ヒト線維芽細胞株〔デトロイト５５１株（ＡＴＣＣ ＣＣＬ １１０）〕の細胞数を１０％（Ｖ／Ｖ）の非働化ＦＢＳを含むＭＥＭ培地にて１．５×１０^５個／ｍＬに調整し、１２穴プレート（ファルコン社製）に０．８ｍＬずつ播種し、９５％（Ｖ／Ｖ）空気−５％（Ｖ／Ｖ）炭酸ガスの雰囲気下、３７℃で３日間静置培養し、さらに、ＭＥＭ培地のみに培地交換し、１日間培養した。その後、高分子トリチウムヒアルロン酸を含む（１μＣｉ／ｍＬ；３７ＭＢｑ／ｍＬ，１０μｇ／ｍＬ）ＭＥＭ培地を調製し、培地交換をし、３日間培養を行った。
【００４０】
・細胞による高分子トリチウムヒアルロン酸の分解評価
培養終了後、培養液を回収し、１００℃で５分間加熱処理を行った後、培地１ｍＬをセファロースＣＬ−２Ｂカラム（内径１ｃｍ，長さ６０ｃｍ）にアプライし以下の条件でゲルろ過を行った。流速：０．６ｍＬ／ｍｉｎ、分画：４ｍＬ／１Ｆｒａｃｔｉｏｎ、分画総数：２５、更に分子量１０万以下のヒアルロン酸が溶出するフラクション１０〜２５の１６本を集め、［^３Ｈ］放射活性を測定し分解したヒアルロン酸の量を求めた。分解抑制率は以下の式を用いて算出した。
分解抑制率＝（試料無添加時の放射活性−試料添加時の放射活性）×１００
／試料無添加時の放射活性
【００４１】
得られた結果を下記に示す。尚、試料添加は各群ｎ＝３で試験を行い、結果はそれぞれの平均値を用いた。

【００４２】

【００４３】
上記より明らかなように、本発明に係るノウゼンハレン科植物抽出物は極めて低濃度でヒアルロン酸分解抑制効果を有することが認められた。
【００４４】
アクアポリン合成促進効果を明らかにするための実施例等を示す（比較例１、実施例４〜６）。
・培養表皮細胞
ヒト正常表皮角化細胞は市販品（Ｅｐｉｄｅｒｃｅｌｌ／新生児包皮：クラボウ社製）を用いた。
【００４５】
・表皮細胞培養用培地
ＭＣＤＢ１５３ＨＡＡ（和光純薬社より購入）をベースとし、ハイドロコーチゾン（０．５μｍｏｌ／Ｌ）、エタノールアミン（０．１ｍｍｏｌ／Ｌ）、ホスホエタノールアミン（０．１ｍｍｏｌ／Ｌ）、インシュリン（５μｇ／ｍＬ）、及びＥＧＦ（上皮細胞成長因子：１０ｎｇ／ｍＬ）を加えたＫ−ＧＭ培地を調製した。培養には、これにＢＰＥ（牛脳下垂体抽出液、クラボウ社より購入）を培地１ｍＬ当たり４μＬ添加し、表皮細胞培養用培地とした。
【００４６】
・試料添加と表皮細胞抽出物の調製
正常ヒト表皮細胞の細胞数を表皮細胞培養用培地にて２．０×１０^４個／ｍＬに調整し、２４穴プレート（ファルコン製）に１．０ｍＬずつ播種した。培養は、９５％（Ｖ／Ｖ）空気−５％（Ｖ／Ｖ）炭酸ガスの雰囲気下３７℃で行い、培養５日目に試料を添加し、さらに１日間培養した。培養後、培地を除き２５０μＬの抽出バッファー（１００ｍｍｏｌ／Ｌ Ｔｒｉｓ−Ｃｌ ｐＨ９．０、１％ ＳＤＳ、９ｍｏｌ／Ｌ尿素）を添加し細胞を溶解させ、さらに５００μＬの水を加え、これを細胞抽出液とした。
【００４７】
・ウェスタンブロッティング
・ＳＤＳポリアクリルアミドゲル電気泳動；
細胞抽出液８μＬ、４倍濃縮ＳＤＳ Ｓａｍｐｌｅ Ｂｕｆｆｅｒ（インビトロジェン社）４μＬ、１０倍濃縮 Ｓａｍｐｌｅ Ｒｅｄｕｃｉｎｇ Ａｇｅｎｔ（インビトロジェン社） １．６μＬ、水 ２．４μＬを混合し、７０℃１０分加熱後、電気泳動に供した。電気泳動は４−１２％ Ｂｉｓ−Ｔｒｉｓ Ｇｅｌ（インビトロジェン社）を用い、ＮｕＰＡＧＥ ＧｅｌＳｙｓｔｅｍ（インビトロジェン社）の添付説明書に従った。
【００４８】
・タンパク質のＰＶＤＦ膜への転写；
転写は、Ｘｃｅｌｌ ＩＩ Ｂｌｏｔ Ｍｏｄｕｌｅ（インビトロジェン社）を使用し、添付説明書に従った。ＰＶＤＦ膜は Ｉｍｍｏｂｉｌｏｎ（ミリポア社）を使用した。
【００４９】
・免疫学的検出；
転写後のＰＶＤＦ膜は、ブロッキング試薬（５％濃度にスキムミルクを０．０５％Ｔｗｅｅｎ−２０含有ＰＢＳに溶解した試薬）に室温で３０分浸した後、ブロッキング試薬で３０００倍に希釈したヒトアクアポリン−３抗体と４℃で２４ｈｒ反応させた。なお、ヒトアクアポリン−３抗体は、ヒトアクアポリン−３アミノ酸配列のＣ末端側ペプチドを合成し、これを免疫したウサギの血清より作製し、希釈倍率は適切な検出シグナルを得るために事前に設定した。次に、ＰＶＤＦ膜を０．０５％Ｔｗｅｅｎ−２０含有ＰＢＳ中で振盪洗浄し、０．０５％Ｔｗｅｅｎ−２０含有ＰＢＳで２０００倍に希釈した抗ウサギＩｇＧ抗体（ＤＡＫＯ社製）と室温で４５分反応させた。最後に、ＰＶＤＦ膜を０．０５％Ｔｗｅｅｎ−２０含有ＰＢＳ中で振盪洗浄後、ＳｕｐｅｒＳｉｇｎａｌ（ＰＩＥＲＣＥ社製）を用い添付説明書に従いシグナルを検出した。結果を図１に示す。尚、Ｔｗｅｅｎ−２０は、Ｕｎｉｑｅｍａ Ａｍｅｒｉｃａｓ社商品名である。
【００５０】
試料無添加群（比較例１）と比べ、ノウゼンハレン抽出物凍結乾燥品０．００１％（実施例４）、０．０１％添加群（実施例５）、ノウゼンハレン５０％エタノール抽出溶液０．１％添加群（実施例６）では顕著な検出シグナル強度の増加が認められた。これより明らかなように、本発明に係るノウゼンハレン科植物抽出物は極めて低濃度でアクアポリン合成促進効果を有することが認められた。
【００５１】
以下、本発明の活性酸素消去剤、ヒアルロン酸合成促進・分解抑制剤、およびアクアポリン合成促進剤の処方例を示す。
【００５２】
処方例１〜３（スキンクリーム）
試料１又は２の活性酸素消去剤、ヒアルロン酸合成促進・分解抑制剤、およびアクアポリン合成促進剤を表１の組成でそれぞれを配合しスキンクリームを調製した（処方例１〜３）。
【００５３】
（１）組成
【表１】

【００５４】
（２）調製法
（Ａ）成分および（Ｂ）成分を各々８０℃に加熱溶解した後混合して、攪拌しつつ冷却し、３０℃まで冷却して、スキンクリームを調製した。
【００５５】
処方例４〜６（ローション）
試料１又は２の活性酸素消去剤、ヒアルロン酸合成促進・分解抑制剤、およびアクアポリン合成促進剤を表２の組成で配合し、ローションを調製した（処方例４〜６）。
【００５６】
（１）組成
【表２】

【００５７】
（２）調製法
各成分をそれぞれ混合溶解し、攪拌して、ローションを調製した。
【００５８】
処方例７〜９（ジェル）
試料１又は２の活性酸素消去剤、ヒアルロン酸合成促進・分解抑制剤、およびアクアポリン合成促進剤を表３の組成でそれぞれを配合しジェルを調製した（処方例７〜９）。
【００５９】
（１）組成
【表３】

【００６０】
（２）調製法
（Ａ）成分および（Ｂ）成分を各々６０℃に加熱溶解した後混合して、攪拌しつつ冷却し、３０℃まで冷却して、ジェルを調製した。
【００６１】
処方例１０〜１２（ジェル）
試料１又は２の活性酸素消去剤、ヒアルロン酸合成促進・分解抑制剤、およびアクアポリン合成促進剤を表４の組成でそれぞれを配合しジェルを調製した（処方例１０〜１２）。
【００６２】
（１）組成
【表４】

【００６３】
（２）調製法
（Ａ）成分および（Ｂ）成分を各々６０℃に加熱溶解した後混合して、攪拌しつつ冷却し、３０℃まで冷却して、ジェルを調製した。
【００６４】
処方例１３〜１５（親油クリーム）
試料１又は２の活性酸素消去剤、ヒアルロン酸合成促進・分解抑制剤、およびアクアポリン合成促進剤を表５の組成で配合し、親油クリームを調製した（処方例１３〜１５）。
【００６５】
（１）組成
【表５】

【００６６】
（２）調製法
Ａ）成分および（Ｂ）成分を各々６０℃に加熱溶解した後混合して、攪拌しつつ冷却し、３０℃まで冷却して、親油クリームを調製した。
【００６７】
処方例１６〜１８（サンスクリーン）
試料１又は２の活性酸素消去剤、ヒアルロン酸合成促進・分解抑制剤、およびアクアポリン合成促進剤を表６の組成でそれぞれを配合しサンスクリーンを調製した（処方例１６〜１８）。
【００６８】
（１）組成
【表６】

【００６９】
（２）調製法
（Ａ）成分および（Ｂ）成分を各々８０℃に加熱溶解した後混合して、攪拌しつつ冷却し、３０℃まで冷却して、サンスクリーンを調製した。
【００７０】
処方例１９〜２１（入浴剤）
試料１又は２の活性酸素消去剤、ヒアルロン酸合成促進・分解抑制剤、およびアクアポリン合成促進剤を表７の組成でそれぞれを配合し入浴剤を調製した（処方例１９〜２１）。
【００７１】
（１）組成
【表７】

【００７２】
（２）調製法
（Ａ）成分および（Ｂ）成分を各々６０℃に加熱溶解した後混合して、攪拌しつつ冷却し、３０℃まで冷却して、入浴剤を調製した。
【００７３】
次に、効果を示すための試験例を示す。
【００７４】
下記記載の組成のスキンローション及び入浴剤を、それぞれの調製法に従い調製した。
【００７５】
試験例１〜３及び比較例２：スキンローション
配合成分 配合量（％）
−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−
（アルコール相）
９５％エチルアルコール １５．０
ポリオキシエチレン（６０モル）硬化ヒマシ油 ２．０
パラオキシ安息香酸メチル ０．０５
香料 適 量
試料（表８記載） 表８記載の量
（水相）
グリセリン ５．０
クエン酸ナトリウム 適 量
イオン交換水 残 余
（合計） １００．０
【００７６】

【００７７】
・スキンローションの調製法
水相、アルコール相を各々均一に溶解し、そして水相とアルコール相とを混合攪拌分散し可溶化を行い、次いで容器に充填する。使用時には内容物を均一に振盪分散して使用する。
【００７８】
上記で調製したスキンローションを用いて使用試験を行い、皮膚老化防止効果及び美白効果を調べた。試験方法は下記に示す通りである。
【００７９】
・試験方法
８０名の女性被験者の顔面を左右に分け、一方に上記の試験例１〜３、他方に比較例２のスキンローションを毎日２回以上塗布してもらい、２ヵ月後比較例２を基準として下記の判定基準により各評価項目について評点を出してもらい、評点の合計値により評価した。被験者を１群２０名にわけて３群とし、試験例１〜３の各スキンローションについて試験を行った。その結果を表９に示す。
【００８０】

【００８１】
［表９］
評価項目 試験例１ 試験例２ 試験例３
−−−−−−−−−−−−−−−−−−−−−−−−
肌荒れ防止 ２５ ２０ ４１
肌のつや ２７ ２１ ４３
肌のはり ３１ ２８ ４５
肌の明るさ ２９ ２２ ３９
しわ改善 ２４ １９ ３７
【００８２】
表９より明らかなように、本発明に係るノウゼンハレン抽出物凍結乾燥品を有効成分として含有するスキンローションは、肌荒れを防止し、肌のつや、はり、明るさを保ち、しわを改善する効果に優れ、皮膚老化防止用及び美白用の皮膚化粧料として用いることができることが明らかとなった。
【００８３】
試験例４：入浴剤
配合成分 配合量（％）
−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−
硫酸ナトリウム ８５．０
香料及び界面活性剤 適 量
有機色素 適 量
ノウゼンハレン抽出物凍結乾燥品 １０．０
炭酸水素ナトリウム 残 量
（合計） １００．０
【００８４】
・入浴剤の調製法
各成分を混合し入浴剤を調製した。尚、この入浴剤は使用時に約３０００倍に希釈される。
【００８５】
上記入浴剤を被験者２０名に三週間毎日の入浴時に使用させた。結果、肌荒れ防止、肌のつや、はり、明るさの保持、しわの改善において効果が認められた。尚、本発明に係るスキンローション及び入浴剤による発赤や乾燥等の異常は認められなかった。
【００８６】
【発明の効果】
以上説明したように、皮膚の保湿性改善、肌荒れ防止、老化防止に有効な活性酸素消去剤、ヒアルロン酸合成促進・分解抑制剤及びアクアポリン合成促進剤を提供できることは明らかである。
【図面の簡単な説明】
【図１】ヒト正常表皮角化細胞におけるノウゼンハレン抽出物のアクアポリン合成促進効果を示す図である。[0001]
TECHNICAL FIELD OF THE INVENTION
The present invention relates to an active oxygen scavenger, a hyaluronic acid synthesis promoter, a hyaluronic acid decomposition inhibitor, and an aquaporin synthesis promoter, and more particularly, eliminates active oxygen to suppress the production of lipid peroxide in the skin, Active oxygen scavenging that promotes hyaluronic acid-producing ability of cells, suppresses resolution, and promotes aquaporin synthesis of skin cells, which is expected to have excellent moisturizing properties on skin, improve rough skin, prevent aging and whitening effect The present invention relates to an agent, a hyaluronic acid synthesis promoter and a decomposition inhibitor, and an aquaporin synthesis promoter.
[0002]
[Prior art]
Aged skin has increased lipid peroxide, loss of flexibility and elasticity, increased skin wrinkling, and dry, non-smooth rough skin symptoms. As one of the substances causing these skin symptoms, so-called active oxygen, which is generated by oxygen in the atmosphere under the influence of ultraviolet rays and enzymes, is considered. Such active oxygen includes superoxide, hydroxy radical, singlet oxygen, hydrogen peroxide and the like. This active oxygen oxidizes the fatty acids to form peroxides. In addition, the generated peroxide and active oxygen have adverse effects on the living body, such as collagen fiber cross-linking, hyaluronic acid fragmentation, DNA helix partial cleavage, and tissue damage due to the generation of chain radicals. As a result, it is said to promote aging of the living body such as skin wrinkles, loss of elasticity, and hair loss.
[0003]
On the other hand, pigmentation on the skin such as darkness, spots, and freckles due to sunburn is considered to be caused by the production of melanin, a black-brown amorphous pigment. Melanin is produced by tyrosinase converting tyrosine to dopa and dopa to dopaquinone in melanosomes in pigment cells called melanocytes present in the basal layer of the epidermis, and then undergoing oxidative polymerization via various melanin intermediate metabolites. It is said that active oxygen generated by ultraviolet rays activates the oxidation reaction of this system, so that melanin is excessively generated and pigmentation on skin is promoted.
[0004]
Therefore, elimination of active oxygen is very important for skin in terms of improving or preventing skin aging and pigmentation, and is an important factor required for anti-aging skin cosmetics.
[0005]
Therefore, conventionally, a search for a substance having an action of eliminating active oxygen generated in a living body has been widely performed. Substances having such an action which have been conventionally used include those derived from natural products, such as fat-soluble tocopherol (vitamin E) and water-soluble ascorbic acid (vitamin C). Examples include BHT (butylhydroxytoluene) and BHA (butylhydroxyanisole). However, these substances are not sufficient in scavenging active oxygen, and BHA is suspected of having carcinogenicity in a synthetic compound, and none of them is practically usable as an active oxygen scavenger. Therefore, there is a strong demand for an active oxygen scavenger that has a sufficient action of eliminating active oxygen generated in a living body, is highly safe, and has good applicability to skin and the like.
[0006]
From the aspect of preventing skin aging, it is also important to increase the production of hyaluron in the skin. Hyaluronic acid retains water in the intercellular space, retains cells based on the formation of a jelly-like matrix in the tissue, maintains the lubricity and flexibility of the tissue, resists external forces such as mechanical damage, and It has many functions such as prevention of bacterial infection (for example, Non-Patent Document 1).
[0007]
It is said that hyaluronic acid in the skin decreases with age, resulting in aging such as fine wrinkles and dryness. Numerous cosmetics containing collagen and hyaluronic acid have been proposed as such aging skin improving agents, but only those that improve the moisturizing effect on the surface and do not essentially improve aging skin. Absent. Therefore, it is effective to prevent skin aging by enhancing the hyaluronic acid production performance of skin cells and suppressing the resolution.
[0008]
In addition to the above-mentioned effects of eliminating active oxygen and increasing hyaluronic acid on the skin, which have been shown to have excellent effects on the skin, recent studies have shown that aquaporins present in skin cells play an important role in skin physiology. It became clear.
[0009]
Aquaporin (hereinafter abbreviated as AQP) is also called a water channel and is known as a membrane protein that selectively allows water to permeate. To date, 11 types of AQP genes (AQP0 to AQP10) have been discovered in humans, and among them, AQP3, AQP7, and AQP9 transmit not only water but also nonionic small molecules such as glycerol and urea. Also, looking at the distribution of AQP expression in human tissues, it is clear that most organs have a plurality of AQPs.
[0010]
Regarding the expression of AQP in the epidermis, recent studies by the present inventors have reported the expression of AQP3, AQP5, and AQP9 genes in cultured human keratinocytes and model skin epidermis. Among them, AQP3 expression in cultured human keratinocytes is increased by hyperosmotic stress, and thus AQP3 is considered to be one important factor that controls the water environment of keratinocytes (see Non-Patent Document 2). Also, in the skin of AQP3 gene-deficient mice, decreased horny layer water content, skin viscoelasticity, and barrier recovery were observed, indicating that AQP3 is deeply involved in skin physical properties and skin physiology (Non-patent Literature) 3). Therefore, promotion of aquaporin production can be expected to suppress skin dryness, adjust the beam, and enhance the barrier function. However, there are no known aquaporin synthesis promoters and skin cosmetics that have sufficient aquaporin synthesis promoting action, are highly safe, and have good applicability to skin and the like.
[0011]
[Non-patent document 1]
"BIO INDUSTRY" Vol. 8, 1991, p. 346
[Non-patent document 2]
"Biochimica et Biophysica Acta", Vol. 1522, 2001, pp. 82-88)
[Non-Patent Document 3]
"Journal of Dermatological Science", Vol. 29, p. 143
[0012]
[Problems to be solved by the invention]
Under such circumstances, an object of the present invention is to eliminate active oxygen, suppress the production of lipid peroxide in the skin, promote the ability of skin cells to produce hyaluronic acid, suppress the resolution, and furthermore, It is an object of the present invention to provide an active oxygen scavenger, a hyaluronic acid synthesis acceleration / decomposition inhibitor, and an aquaporin synthesis accelerator having excellent moisturizing properties on skin, improvement of rough skin, prevention of aging and whitening by promoting aquaporin synthesis.
[0013]
[Means for Solving the Problems]
In view of the above circumstances, the present inventors have conducted intensive studies, and as a result, the extract obtained from the Nosenharenaceae plant has an excellent active oxygen scavenging effect, a hyaluronic acid synthesis promotion / decomposition suppression effect, and an aquaporin synthesis promotion effect, and The present inventors have found that the dryness, rough skin, aging and pigmentation can be effectively improved or prevented, and the present invention has been completed.
[0014]
That is, the present invention resides in an active oxygen scavenger, a hyaluronic acid synthesis promotion / decomposition inhibitor, and an aquaporin synthesis promoter, characterized by containing an extract obtained from a plant of the family Nosenharenaceae.
[0015]
BEST MODE FOR CARRYING OUT THE INVENTION
Hereinafter, embodiments of the present invention will be described in detail.
[0016]
The Rhododendronaceae plant used in the present invention is a succulent herb with prominent stems, and its place of origin is limited to central and southern South Africa from Mexico to Patagonia. The most famous ornamental one is Tropaeolum majus, which is also called nasturtium (gold lotus flower) and is cultivated in flower beds and potted plants. Also, immature seeds are sometimes pickled and eaten instead of capers.
[0017]
Examples of the extraction solvent for obtaining the Russula plant extract of the present invention include water, alcohols such as methanol, ethanol and isopropyl alcohol, polyhydric alcohols such as ethylene glycol, propylene glycol and 1,3-butylene glycol, and acetone. Or a mixture of one or more of ketones such as ethyl acetate, esters such as ethyl acetate, ethers such as diethyl ether, and aromatic compounds such as benzene.
[0018]
The extract of Rusticaceae may be used as an extraction solvent solution, or may be dried and concentrated by a conventional method. Further, filtration, re-extraction, and the like may be performed.
[0019]
The active oxygen scavenger, hyaluronic acid synthesis accelerator / decomposition inhibitor and aquaporin synthesis accelerator of the present invention are, for example, ointments, lotions, emulsions, creams, packs, powders, granules, etc. Can be used in any dosage form. Examples of the form of use include skin cosmetics. The base of the skin cosmetic may be a known external base, and is not particularly limited. It can be expected to be applied to all skin on the human body including the scalp as the skin to be applied.
[0020]
The amount of the active oxygen scavenger, the hyaluronic acid synthesis accelerating / degrading inhibitor and the aquaporin synthesis accelerating agent of the extract of Rusticaceae plant of the present invention varies depending on the system used, and cannot be said unconditionally. When the amount is 0.00001 to 10.0% by mass in terms of dry matter (hereinafter referred to as% unless otherwise specified), the active oxygen scavenging effect, the hyaluronic acid synthesis promotion / decomposition suppression effect, and the aquaporin synthesis of the present invention are obtained. It is preferable to obtain an accelerating effect, and has a good feel at the time of use, and can stably maintain individual dosage forms. If the amount is less than 0.00001%, the effect may not be sufficiently exhibited, while if the amount exceeds 10.0%, the effect corresponding to the amount may not be obtained.
[0021]
The active oxygen scavenger, the hyaluronic acid synthesis accelerating / degrading inhibitor and the aquaporin synthesis accelerating agent of the present invention, if necessary, are usually used in pharmaceuticals, quasi-drugs, cosmetics and other external skin preparations. -Based pigments, coloring pigments such as iron oxide, preservatives such as paraben and phenoxyethanol, silicone oils such as dimethylpolysiloxane, methylphenylpolysiloxane and cyclic silicone, hydrocarbons such as paraffin and petrolatum, olive squalane, rice squalane, rice Germ oil, jojoba oil, castor oil, safflower oil, olive oil, macadamia nut oil, sunflower oil, and other vegetable oils, beeswax, mocro, carnauba wax and other waxes, octyl dodecyl myristate, cetyl palmitate, isostearyl isostearate, isopropyl myristate Such as ester oil, ethanol, etc. Higher alcohols such as lower alcohols, cetanol, behenyl alcohol, stearyl alcohol, and long-chain branched aliphatic alcohols; sterols and derivatives such as cholesterol, phytosterol, branched fatty acid cholesterol ester, and macadamia nut fatty acid phytosteryl ester; and processing oils such as hardened oil. Kind,
[0022]
Higher fatty acids such as stearic acid, myristic acid, isostearic acid, oleic acid, iso long chain fatty acids and anteiso long chain fatty acids, terpenes such as limonene, hydrogenated bisabolol, glyceryl tricapryl caprate, glyceryl 2-ethylhexanoate , Triglycerides such as triiso long chain fatty acid glyceryl, glyceryl tripalmitate, anionic surfactants such as sodium cetyl sulfate, N-stearoyl-L-glutamate, polyoxyethylene alkyl ether, polyoxyethylene fatty acid ester, polyoxy Nonionic surfactants such as ethylene polyhydric alcohol fatty acid ester, polyoxyethylene hydrogenated castor oil, polyhydric alcohol fatty acid ester, polyglycerin fatty acid ester, modified silicone, sucrose ester, etc. Cationic surfactants such as ammonium salts, amphoteric surfactants such as betaine type, sulfobetaine type and sulfoamino acid type, natural surfactants such as lecithin, lysophosphatidylcholine, ceramide, cerebroside, titanium oxide, and oxidation Pigments such as zinc, antioxidants such as dibutylhydroxytoluene, inorganic salts such as sodium chloride, magnesium chloride, sodium sulfate, potassium nitrate, sodium sulfate, sodium metasilicate, calcium chloride, sodium citrate, potassium acetate, sodium succinate, Organic acids and salts thereof such as sodium aspartate, sodium lactate, dichloroacetic acid, mevalonic acid, and glycyrrhizic acid; organic amines such as ethanolamine hydrochloride, ammonium nitrate, arginine hydrochloride, diisopropylamine salt, urea, and decarboxycarnosine; Salts, chelating agents such as edetic acid, xanthan gum, carboxyvinyl polymer, carrageenan, pectin, alkyl-modified carboxyvinyl polymer, thickeners such as agar, potassium hydroxide, diisopropanolamine, neutralizing agents such as triethanolamine, UV absorbers such as hydroxymethoxybenzophenone sulfonate, dipropylene glycol, 1,3-butylene glycol, glycerin, propylene glycol, sorbitol, malbitol, diglycerin, polyhydric alcohols such as raffinose, various amino acids, ascorbic acid, biotin, Vitamins such as tocopherol and ascorbic acid sulfates, ascorbic acid phosphates, vitamin derivatives such as tocopherol nicotinate and the like are appropriately blended within a range that achieves the object of the present invention. Can be
[0023]
【Example】
Examples for clarifying the active oxygen elimination effect, the hyaluronic acid synthesis promotion / decomposition suppression effect, and the aquaporin synthesis promotion effect according to the present invention will be described below. Note that the present invention is not limited to these.
[0024]
Sample: Nosenhalen extract
As the Nosenhallen extract, a freeze-dried Nosenhallen extract (Sample 1; Maruzen Pharmaceutical Co., Ltd.) and a 50% ethanol extract of Nosenhallen (Sample 2; Maruzen Pharmaceutical Co., Ltd.) were used.
[0025]
・ Active oxygen scavenging effect test (Example 1)
There are various methods for measuring the effect of eliminating active oxygen, but this time, superoxide (O₂ ⁻) Was measured. The method is as follows. That is, the superoxide (O 2) which is one of the active oxygens by the xanthine-xanthin oxidase system₂ ⁻) Was generated and its erasure rate by the sample solution was determined. Superoxide (O₂ ⁻) Was reacted with nitroblue tetrazolium to form diformazan and measured by absorbance at 560 nm (NBT method).
[0026]
2.4 mL of 50 mmol / L sodium carbonate buffer (pH 10.2), 0.1 mL of 3.0 mmol / L xanthine, 0.1 mL of 3.0 mmol / L disodium ethylenediaminetetraacetate, 0.1 mL of 0.15% bovine serum albumin 0.1 mL of a sample solution is added to 2.9 mL of a color developing solution containing 0.1 mL of 0.75 mmol / L nitro blue tetrazolium, and the temperature is kept at 25 ° C. to give an enzyme containing xanthine oxidase (0.5 units / mL). After 0.1 mL of the liquid was added and stirred, the mixture was left at 25 ° C. for 20 minutes. 0.1 mL of a reaction stop solution containing 6.0 mmol / L cupric chloride was added, and the absorbance at 560 nm was measured with a spectrophotometer. For the reagent blank, take 2.9 mL of the color reagent solution and 0.1 mL of the sample solution, keep the temperature at 25 ° C., add 0.1 mL of phosphate buffer in place of the enzyme solution, and repeat the same procedure with a spectrophotometer at 560 nm. The absorbance was measured, and the value was defined as S '. As a control, 2.9 mL of the color reagent solution and 0.1 mL of each diluted solution were used in place of the sample solution. Absorbance at 560 nm was measured using a spectrophotometer in the same manner as above, and the value was designated as C. For the control reagent blank, use 2.9 mL of the color reagent and 0.1 mL of each diluted solution in place of the sample solution, add 0.1 mL of sodium carbonate buffer in place of the enzyme solution, and apply the same procedure to the spectrophotometer in the same manner. The absorbance at 560 nm was measured and the value was taken as C ′. The superoxide elimination rate at each sample solution concentration was calculated by the following equation. The horizontal axis of the semilogarithmic graph was the sample solution concentration (log), and the vertical axis was the superoxide elimination rate. From this graph, the superoxide 50% erasure concentration ( IC50) was determined. In addition, S, S ', C, and C' were measured three times, and the average value of each was used.
[formula]
Superoxide erasure rate (%) = 100 × [1- (S−S ′) / (C−C ′)]
[0027]
The results obtained are shown below.

[0028]

[0029]
As is clear from the above, it was confirmed that the extract of the family Rusticaceae according to the present invention eliminated superoxide at an extremely low concentration.
[0030]
An example for clarifying the effect of promoting hyaluronic acid synthesis and inhibiting decomposition will be described.
(1) Hyaluronic acid synthesis promoting effect (Example 2)
・ Preparation method of MEM medium
Minimum Essential Medium (Dainippon Pharmaceutical Co., Ltd., 10-101) As a final concentration in 10.6 g, 1% (V / V) Non Essential Amino Acid (Dainippon Pharmaceutical Co., Ltd., 16-810), 1 mmol / L pyruvine Sodium acid (Dainihon Pharmaceutical Co., Ltd., 16-820), 1.2% (W / V) sodium bicarbonate, and distilled water were added to make up to 1 L, and then carbon dioxide was blown to adjust the pH to about 7 (hereinafter, referred to as “pH”). , MEM medium).
[0031]
・ Inactivation of fetal bovine serum (FBS)
FBS (Irvine Scientific) was heat-treated at 56 ° C. for 30 minutes.
[0032]
・ Pronase solution
200 μg / mL pronase (from Streptomyces griseus, manufactured by Calbiochem-Bering Corporation), 0.5 mol / L Tris-HCl buffer (pH 8.0) containing 0.15 mol / L sodium chloride and 0.02% sodium azide.
[0033]
・ Cell culture
A normal human fibroblast cell line [Detroit 551 (ATCC CCL 110)] was 0.6 × 10⁵The cells were seeded in a 24-well plate at a density of cells / well, and an atmosphere of 95% (V / V) air-5% (V / V) carbon dioxide gas in MEM medium containing 10% (V / V) inactivated FBS. The cells were cultured at 37 ° C. for 3 days, and further cultured in a serum-free MEM medium for 1 day. Next, after culturing for 3 days in the medium to which the sample was added, the culture supernatant was recovered, and the amount of hyaluronic acid in the supernatant was determined.
[0034]
・ Measurement of hyaluronic acid production
0.07 mL of the pronase solution was added to 0.63 mL of the culture supernatant, allowed to stand at 37 ° C. for 18 hours, and then heat-treated at 100 ° C. for 10 minutes to inactivate pronase. Next, the amount of hyaluronic acid was measured from 0.01 mL of the reaction solution using a hyaluronic acid plate “Chugai” (manufactured by Chugai Diagnostics Science) according to the attached instructions. In addition, the sample was added and the test was performed in each group n = 3, and the average value was used for the results.
[0035]
The results obtained are shown below.

[0036]

[0037]
As is clear from the above, it was confirmed that the extract of the family Rusticaceae according to the present invention has a hyaluronic acid synthesis promoting effect at an extremely low concentration.
[0038]
(2) Hyaluronic acid decomposition inhibitory effect (Example 3)
・ Preparation method of polymer tritium hyaluronic acid for cell addition
The number of cells of a normal human fibroblast cell line [Detroit 551 (ATCC CCL 110)] was increased to 2 × 10 4 in a MEM medium containing 10% (V / V) of inactivated FBS.⁵Adjust to pcs / mL, 225cm²Was placed in a flask and cultured for 3 days to obtain a confluent state. Thereafter, tritium glucosamine (manufactured by American Radioactive Chemicals Inc.), which is a precursor of hyaluronic acid, was added to the culture system (10 μCi / mL), and after further culturing for 3 days, tritium-labeled hyaluronic acid from the culture solution was extracted from Underhill et al. (J. Cell Biology, Vol. 82, p. 475, 1979), and a high molecular weight tritium hyaluronic acid (specific radioactivity, 0.1 μCi / μg) having a molecular weight of 1,000,000 or more was prepared using a gel filtration column. . This was used as a polymer tritium hyaluronic acid for addition to a cell culture system.
[0039]
・ Addition culture of high molecular weight tritium hyaluronic acid to normal human fibroblasts
The number of cells of a normal human fibroblast cell line [Detroit 551 (ATCC CCL 110)] was increased to 1.5 × 10 5 in MEM medium containing 10% (V / V) of inactivated FBS.⁵0.8 mL per 12-well plate (manufactured by Falcon) and seeded at 37 ° C. for 3 days in an atmosphere of 95% (V / V) air-5% (V / V) carbon dioxide gas. The culture was allowed to stand, and the medium was changed to MEM medium only, followed by culturing for one day. Thereafter, a MEM medium containing high molecular tritium hyaluronic acid (1 μCi / mL; 37 MBq / mL, 10 μg / mL) was prepared, the medium was replaced, and the cells were cultured for 3 days.
[0040]
・ Evaluation of degradation of polymer tritium hyaluronic acid by cells
After completion of the culture, the culture solution was collected and subjected to a heat treatment at 100 ° C. for 5 minutes. Then, 1 mL of the medium was applied to a Sepharose CL-2B column (1 cm inside diameter, 60 cm length), and gel filtration was performed under the following conditions. . Flow rate: 0.6 mL / min, fractionation: 4 mL / 1 fraction, total number of fractions: 25, and 16 fractions of 10 to 25 fractions from which hyaluronic acid having a molecular weight of 100,000 or less eluted were collected.³H] The radioactivity was measured to determine the amount of decomposed hyaluronic acid. The decomposition inhibition rate was calculated using the following equation.
Degradation inhibition rate = (radioactivity when no sample is added−radioactivity when sample is added) × 100
/ Radioactivity without sample
[0041]
The results obtained are shown below. In addition, the sample was added and the test was performed with n = 3 in each group, and the average value was used for the results.

[0042]

[0043]
As is clear from the above, it was confirmed that the extract of the family Rusticaceae according to the present invention has a hyaluronic acid decomposition inhibitory effect at an extremely low concentration.
[0044]
Examples and the like for clarifying the effect of accelerating aquaporin synthesis are shown (Comparative Example 1, Examples 4 to 6).
・ Cultivated epidermal cells
As human normal epidermal keratinocytes, a commercially available product (Epidercell / neonatal foreskin: manufactured by Kurabo Industries) was used.
[0045]
・ Epidermal cell culture medium
Based on MCDB153HAA (purchased from Wako Pure Chemical Industries), hydrocortisone (0.5 μmol / L), ethanolamine (0.1 mmol / L), phosphoethanolamine (0.1 mmol / L), insulin (5 μg / mL) , And K-GM medium to which EGF (epithelial cell growth factor: 10 ng / mL) was added. For culture, BPE (bovine pituitary extract, purchased from Kurabo Industries) was added to the culture in an amount of 4 μL per 1 mL of the culture medium to obtain a culture medium for epidermal cells.
[0046]
・ Sample addition and preparation of epidermal cell extract
The number of normal human epidermal cells was determined to be 2.0 × 10⁴And adjusted to 1.0 / mL and inoculated in a 24-well plate (Falcon) in an amount of 1.0 mL each. Culturing was performed at 37 ° C. in an atmosphere of 95% (V / V) air-5% (V / V) carbon dioxide, the sample was added on the 5th day of the culture, and the cells were further cultured for 1 day. After the culture, the medium was removed, 250 μL of an extraction buffer (100 mmol / L Tris-Cl pH 9.0, 1% SDS, 9 mol / L urea) was added to dissolve the cells, and 500 μL of water was further added. And
[0047]
・ Western blotting
-SDS polyacrylamide gel electrophoresis;
8 μL of cell extract, 4 μL of 4 × concentrated SDS Sample Buffer (Invitrogen), 4 μL of 10 × concentrated Sample Reducing Agent (Invitrogen), and 2.4 μL of water are mixed, heated at 70 ° C. for 10 minutes, and then subjected to electrophoresis. did. The electrophoresis was performed using 4-12% Bis-Tris Gel (Invitrogen) according to the instructions attached to NuPAGE GelSystem (Invitrogen).
[0048]
Transfer of proteins to PVDF membranes;
Transcription was performed using Xcell II Blot Module (Invitrogen) according to the attached instructions. The PVDF membrane used was Immobilon (Millipore).
[0049]
-Immunological detection;
After the transfer, the PVDF membrane was immersed in a blocking reagent (a reagent prepared by dissolving skim milk at a concentration of 5% in PBS containing 0.05% Tween-20) for 30 minutes at room temperature, and then diluted with a blocking reagent to 3000 times human aquaporin-. The mixture was reacted with 3 antibodies at 4 ° C for 24 hours. The human aquaporin-3 antibody was prepared by synthesizing a C-terminal peptide of the human aquaporin-3 amino acid sequence and preparing it from the serum of a rabbit immunized with the peptide, and the dilution factor was set in advance to obtain an appropriate detection signal. . Next, the PVDF membrane was washed by shaking in PBS containing 0.05% Tween-20, and then diluted with 2,000-fold anti-rabbit IgG antibody (manufactured by DAKO) in PBS containing 0.05% Tween-20 at room temperature for 45 minutes. Reacted. Finally, the PVDF membrane was washed by shaking in PBS containing 0.05% Tween-20, and then a signal was detected using SuperSignal (manufactured by PIERCE) according to the attached instructions. The results are shown in FIG. Tween-20 is a product name of Uniqema Americas.
[0050]
Compared with the sample-free group (Comparative Example 1), the freeze-dried Nosenhallen extract 0.001% (Example 4), the 0.01% -added group (Example 5), and the Nosenhallen 50% ethanol extraction solution 0.1% In the addition group (Example 6), a remarkable increase in detected signal intensity was observed. As is evident from the above, it was confirmed that the extract of the family Rubenaceae according to the present invention has an aquaporin synthesis promoting effect at an extremely low concentration.
[0051]
Hereinafter, formulation examples of the active oxygen scavenger, the hyaluronic acid synthesis promotion / decomposition inhibitor, and the aquaporin synthesis promoter of the present invention are shown.
[0052]
Formulation Examples 1-3 (skin cream)
A skin cream was prepared by blending the active oxygen scavenger, hyaluronic acid synthesis accelerating / degradation inhibitor, and aquaporin synthesis accelerating agent of Sample 1 or 2 with the compositions shown in Table 1 (Formulation Examples 1 to 3).
[0053]
(1) Composition
[Table 1]

[0054]
(2) Preparation method
The components (A) and (B) were each heated and dissolved at 80 ° C., mixed, cooled with stirring, and cooled to 30 ° C. to prepare a skin cream.
[0055]
Formulation Examples 4-6 (Lotion)
The active oxygen scavenger, hyaluronic acid synthesis promotion / decomposition inhibitor, and aquaporin synthesis promoter of Sample 1 or 2 were blended in the composition shown in Table 2 to prepare lotions (Formulation Examples 4 to 6).
[0056]
(1) Composition
[Table 2]

[0057]
(2) Preparation method
Each component was mixed and dissolved, and stirred to prepare a lotion.
[0058]
Formulation Examples 7-9 (Gel)
Gels were prepared by mixing the active oxygen scavenger, hyaluronic acid synthesis accelerator / decomposition inhibitor, and aquaporin synthesis accelerator of Sample 1 or 2 with the compositions shown in Table 3 (Formulation Examples 7 to 9).
[0059]
(1) Composition
[Table 3]

[0060]
(2) Preparation method
The components (A) and (B) were each dissolved by heating at 60 ° C., mixed, cooled with stirring, and cooled to 30 ° C. to prepare a gel.
[0061]
Formulation Examples 10-12 (Gel)
The active oxygen scavenger, hyaluronic acid synthesis accelerating / degradation inhibitor, and aquaporin synthesis accelerating agent of Sample 1 or 2 were each blended with the compositions shown in Table 4 to prepare gels (Formulation Examples 10 to 12).
[0062]
(1) Composition
[Table 4]

[0063]
(2) Preparation method
The components (A) and (B) were each dissolved by heating at 60 ° C., mixed, cooled with stirring, and cooled to 30 ° C. to prepare a gel.
[0064]
Formulation Examples 13 to 15 (lipophilic cream)
The active oxygen scavenger, hyaluronic acid synthesis accelerating / degradation inhibitor, and aquaporin synthesis accelerating agent of Sample 1 or 2 were blended in the composition shown in Table 5 to prepare lipophilic creams (Formulation Examples 13 to 15).
[0065]
(1) Composition
[Table 5]

[0066]
(2) Preparation method
The components (A) and (B) were each heated and dissolved at 60 ° C., mixed, cooled with stirring, and cooled to 30 ° C. to prepare a lipophilic cream.
[0067]
Formulation Examples 16-18 (Sunscreen)
Sunscreens were prepared by blending the active oxygen scavenger, hyaluronic acid synthesis promotion / degradation inhibitor, and aquaporin synthesis promoter of Sample 1 or 2 with the compositions shown in Table 6 (Formulation Examples 16 to 18).
[0068]
(1) Composition
[Table 6]

[0069]
(2) Preparation method
The components (A) and (B) were each heated and dissolved at 80 ° C., mixed, cooled with stirring, and cooled to 30 ° C. to prepare a sunscreen.
[0070]
Formulation Examples 19 to 21 (bath additives)
The active oxygen scavenger, hyaluronic acid synthesis accelerating / degradation inhibitor, and aquaporin synthesis accelerating agent of Sample 1 or 2 were blended with the compositions shown in Table 7 to prepare bathing agents (Formulation Examples 19 to 21).
[0071]
(1) Composition
[Table 7]

[0072]
(2) Preparation method
The components (A) and (B) were each dissolved by heating at 60 ° C., mixed, cooled with stirring, and cooled to 30 ° C. to prepare a bath agent.
[0073]
Next, test examples for showing the effect will be described.
[0074]
Skin lotions and bath agents having the following compositions were prepared according to the respective preparation methods.
[0075]
Test Examples 1-3 and Comparative Example 2: Skin lotion
Ingredients Ingredients (%)
−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−
(Alcohol phase)
95% ethyl alcohol 15.0
Polyoxyethylene (60 mol) hydrogenated castor oil 2.0
Methyl paraoxybenzoate 0.05
Appropriate amount of fragrance
Sample (described in Table 8) Amount described in Table 8
(Aqueous phase)
Glycerin 5.0
Sodium citrate qs
Ion exchange water residue
(Total) 100.0
[0076]

[0077]
・ Skin lotion preparation method
The aqueous phase and the alcohol phase are each uniformly dissolved, and the aqueous phase and the alcohol phase are mixed, dispersed by stirring, solubilized, and then filled in a container. When used, the contents are shaken and dispersed uniformly.
[0078]
A use test was performed using the skin lotion prepared above, and the effect of preventing skin aging and the effect of whitening were examined. The test method is as shown below.
[0079]
·Test method
The faces of 80 female subjects were divided into left and right sides, and one of the test examples 1 to 3 described above was applied to one side, and the skin lotion of Comparative Example 2 was applied twice or more daily to the other. A rating was given for each evaluation item according to the evaluation criteria described above, and the evaluation was made based on the total value of the ratings. The test subjects were divided into 20 groups and divided into three groups, and the skin lotions of Test Examples 1 to 3 were tested. Table 9 shows the results.
[0080]

[0081]
[Table 9]
Evaluation item Test example 1 Test example 2 Test example 3
−−−−−−−−−−−−−−−−−−−−−−−−−−−
Preventing rough skin 25 20 41
Skin luster 27 21 43
Skin beam 31 28 45
Skin Brightness 29 22 39
Wrinkle improvement 24 19 37
[0082]
As is evident from Table 9, the skin lotion containing the freeze-dried Nosenharen extract according to the present invention as an active ingredient is effective in preventing rough skin, maintaining luster, firmness, brightness, and improving wrinkles. It became clear that it can be used as a skin cosmetic for preventing skin aging and for whitening.
[0083]
Test Example 4: Bath agent
Ingredients Ingredients (%)
−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−
Sodium sulfate 85.0
Flavors and surfactants
Organic dye qs
Nosenhalen extract freeze-dried product 10.0
Sodium bicarbonate balance
(Total) 100.0
[0084]
・ Bath preparation method
Each component was mixed to prepare a bath agent. In addition, this bath agent is diluted about 3000 times at the time of use.
[0085]
The bath preparation was used by 20 subjects for bathing every day for three weeks. As a result, an effect was recognized in preventing rough skin, maintaining gloss, abrasion, brightness of the skin, and improving wrinkles. No abnormalities such as redness and drying caused by the skin lotion and bath agent according to the present invention were observed.
[0086]
【The invention's effect】
As described above, it is clear that an active oxygen scavenger, a hyaluronic acid synthesis promotion / decomposition inhibitor, and an aquaporin synthesis promoter that are effective for improving skin moisture retention, preventing skin roughness, and preventing aging can be provided.
[Brief description of the drawings]
BRIEF DESCRIPTION OF DRAWINGS FIG. 1 is a view showing the aquaporin synthesis-promoting effect of an extract of Nosenhalen on human normal epidermal keratinocytes.

Claims (6)

An active oxygen scavenger, comprising an extract obtained from a Nosenharenaceae plant. A hyaluronic acid synthesis promoter comprising an extract obtained from a plant of the family Nosenharenaceae. A hyaluronic acid decomposition inhibitor comprising an extract obtained from a Nosenharenaceae plant. An aquaporin synthesis promoter comprising an extract obtained from a plant of the family Nosenharenaceae. 2. The active oxygen scavenger according to claim 1, wherein the Russula plant is of the genus Russula. The active oxygen scavenger according to claim 1, wherein the Nosenharenaceae plant is Nosenharen (Nasturtium).

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