JP3574495B2

JP3574495B2 - Hair charge

Info

Publication number: JP3574495B2
Application number: JP07384995A
Authority: JP
Inventors: 隆裕三枝
Original assignee: Sansho Pharmaceutical Co Ltd
Current assignee: Sansho Pharmaceutical Co Ltd
Priority date: 1994-03-31
Filing date: 1995-03-30
Publication date: 2004-10-06
Anticipated expiration: 2019-10-06
Also published as: JPH07316026A

Description

【０００１】
【産業上の利用分野】
本発明は、担子菌の培養液の抽出液又は菌体の抽出液の頭髪料への応用に関するものであって、より詳しくは、担子菌の培養物から菌体を除去した培養液から水及び／又は有機溶媒で抽出した液又は培養菌体から水及び／又は有機溶媒で抽出した液のメラニン生成亢進成分の１種又は２種以上を配合することによって白髪防止効果を発揮すると同時に育毛効果を併せ持った頭髪料に関する。
【０００２】
【従来の技術】
担子菌類（キノコ）は、従来食用または漢方薬として利用されてきた。最近では、医薬品または化粧品への応用も見られる。特に担子菌類の頭髪料への応用術を開示したものとしては、特開昭６０−２５５７１５号公報、特公平４−７７７２５号公報などがある。
特開昭６０−２５５７１５号公報には、子実体からの抽出液を利用した発毛促進養毛化粧料の製法、特公平４−７７７２５号公報には、シロキクラゲの液体培養で得られた粘性物を配合した化粧料が開示されている。
【０００３】
他にも、従来の担子菌の利用方法として、制癌剤、抗腫瘍性物質、免疫調節物質などの医薬品への利用は数多く見られるが、化粧品への利用は前述の通り粘性物質を得るものが幾つがあるにすぎず、育毛促進物質を得るものとしては、例えば、特開昭６０−２５５７１５号公報があるが、白髪防止効果と育毛効果を併せ持つような素材は無い。また同公報には、キノコの子実体からの抽出液を利用した化粧料が開示されているが、子実体は、その生育に時間がかかるため安価に大量に入手しにくく、また入手した後も、子実体の個体差による有用性の変動や抽出率の悪さのため量産化が困難であるなどの、実用的な面での問題点が多い。
【０００４】
【発明が解決しようとする課題】
本発明の目的は、頭髪料に求められる有用性成分を培養液又は菌体中に変動無くかつ高濃度に生産させることで量産化を図り、さらに各種処理を施しその濃度をさらに高めることにより、白髪防止効果並びに育毛効果を併せ持った、安全性の高い頭髪料を提供することにある。
【０００５】
【課題を解決するための手段】
本発明者らは、上記課題を解決するため鋭意研究を行った結果、特定の担子菌類を液体培養した培養液の水及び／又は有機溶媒による抽出液、又は菌体の水及び／又は有機溶媒による抽出液に白髪防止効果があることを見出したほか、育毛効果も有するということを確認し、本発明を完成した。
すなわち、本発明によれば、特定のシメジ科、ハリタケ科、カンゾウタケ科、キコブタケ科、モエギタケ科およびハラタケ科からなる群より選択される担子菌の培養液又は菌体の抽出液群のメラニン生成亢進成分の１種又は２種以上を配合することを特徴とする頭髪料が提供される。
これらの担子菌類の培養液の抽出液又は菌体の抽出液は、メラニン生成亢進成分による白髪防止効果並びに育毛効果に優れている。
【０００６】
【発明の具体的説明】
本発明者は、担子菌類数十種類について、白髪防止効果及び育毛効果を得るべく鋭意研究を行った結果、特定のシメジ科、ハリタケ科、カンゾウタケ科、キコブタケ科、モエギタケ科およびハラタケ科の担子菌類を糖類を炭素源とした通気攪拌液体培養することにより得られた培養液の水及び／又は有機溶媒による抽出液又は菌体の水及び／又は有機溶媒による抽出液に、マウスメラノーマＢ１６細胞のメラニン生成を強く亢進する成分があることを初めて見出したものであり、これらには、さらに育毛効果を有することも見いだして本発明を完成した。
【０００７】
本発明における担子菌類としては、ムキタケ、マツオオジ、ブナハリタケ、のカンゾウタケ、メシマコブ、カバノアナタケ、ヌメリスギタケおよびツクリタケに上記特性において高い有効性が認められた。
【０００８】
本発明における培養液の抽出液としては、炭素源、窒素源、無機塩類などを含む液体培地に担子菌の種菌を接種し、１５ないし３５℃、好ましくは２０ないし３０℃の温度条件で、５ないし４５日間、好ましくは１０ないし３０日間、通気攪拌培養した後、培養液から菌体を遠心分離又はろ別などにより除去した溶液又はその溶液を減圧濃縮機などで濃縮したものを水及び／又は有機溶媒で抽出したものが好適なものとして開示できる。
【０００９】
有機溶媒としては、メタノール、エタノール、イソプロピルアルコール、ブタノール、グリセリン、エチレングリコール、１，３−ブチレングリコール、アセトン、クロロホルム、酢酸エチル、ヘキサン、エーテルなどが好ましく、その中でもエタノール、イソプロピルアルコール、ブタノール、グリセリン、エチレングリコールおよび１，３−ブチレングリコールが好ましく使用される。また、有効性、安全性をさらに高めるため抽出液を希釈又は濃縮した後、限外ろ過または逆浸透膜処理したもの若しくはそれらを活性炭または各種樹脂、例えば、セパビーズＳＰ−２０５（三菱化学（株））などで処理したもの、またはそれらの処理液を希釈又は濃縮したものも含めて言う。
【００１０】
本発明における菌体抽出液としては、上記の培養液から遠心分離又はろ別して得られた菌体をそのまま若しくは細断後、水及び／又は有機溶媒にて充分抽出したもの、又はその濃縮液が好適なものとして開示できる。有機溶媒としては、メタノール、エタノール、イソプロピルアルコール、ブタノール、グリセリン、エチレングリコール、１，３−ブチレングリコール、アセトン、クロロホルム、酢酸エチル、ヘキサンおよびエーテルなどが挙げられ、その中でもエタノール、ブタノール、グリセリン、エチレングリコールおよび１，３−ブチレングリコールが好ましく使用される。また、有効性、安全性をさらに高めるため、抽出液を希釈又は濃縮した後、限外ろ過または逆浸透膜処理したもの若しくはそれらを活性炭または各種樹脂、例えば、セパビーズＳＰ−２０５（三菱化学（株））などで処理したもの、またはそれらの処理液を希釈又は濃縮したものも含めて言う。以上のようにして得られた本発明による担子菌類の培養液の抽出液若しくは菌体抽出液は、メラニン生成亢進作用による白髪防止効果に優れていると同時に育毛効果を併せ持っており、しかも皮膚に対し何ら損傷を与えるものではなく安全性にも優れている。
【００１１】
本発明の頭髪料は、前述の有効性成分を頭髪施用上許容し得る公知の剤型に配合して製造するものであり、その配合量は、培養方法、処理方法、濃縮度合いおよび配合する製剤の形態によって多少異なるが、通常、培養液または菌体抽出液またはそれらの濃縮液を製剤中に０．０５ないし５０重量％、好ましくは０．１ないし１０重量％程度配合するのが好ましい。
【００１２】
さらに、本発明の有効成分の他に通常に用いられる種々の公知の有効成分、例えば塩化カルプロニウム、セファランチン、ビタミンＥ、ビタミンＥニコチネート、ニコチン酸、ニコチン酸アミド、ニコチン酸ベンジル、ショウキョウチンキおよびトウガラシチンキなどの末梢血管拡張剤、カンフルおよびメントールなどの清涼剤、ヒノキチオール、塩化ベンザルコニウムおよびウンデシレン酸などの抗菌剤、塩化リゾチーム、グリチルリチンおよびアラントインなどの消炎剤、センブリエキス、ニンニクエキス、ニンジンエキス、オウゴンエキス、ローズマリーエキス、アロエエキス、ヘチマ抽出物、イチョウ抽出物、ニワトコ抽出物、胎盤抽出液および肝臓抽出物および乳酸菌培養抽出物などの動物・植物・微生物由来の各種抽出物などを自由に添加して使用することができる。
【００１３】
先にも述べた如く、本発明の外用剤の公知の剤型とは、外用可能なあらゆる剤型を意味し、例えばヘアクリーム、ヘアエッセンス、シャンプー、リンス、ヘアトニック、ヘアリキッド、ヘアスプレーおよびヘアフォームなどの頭髪適用剤が例示できる。また、前述の外用剤には公知の有効成分の他に、界面活性剤、油脂類などの基剤成分や、必要に応じて公知の保湿剤、増粘剤、防腐剤、酸化防止剤、紫外線吸収剤・散乱剤、キレート剤、ｐＨ調整剤、香料および着色剤など種々の添加剤を適宜使用できる。
【００１４】
【実施例】
以下に、本発明の製造例、その効果を説明するための試験例並びに処方例を挙げるが、これらは本発明を何ら限定するものではない。
【００１５】
＜製造例１＞
グルコース４５ｇ、ペプトン３ｇ、酵母エキス３ｇ、ＫＨ_２ＰＯ_４０．７５ｇを精製水１５００ｍｌに溶解後、ｐＨ６．０に調整し、１２０℃で１５分間殺菌し２８℃に冷却後、ムキタケの種菌をこれに接種し、２８℃で通気攪拌培養をグルコース残量が０．３％以下になるまで２２日間行った。培養物から遠心分離により菌体を集め水洗後、ナイロンメッシュろ布に取り水分を切り、菌体４８ｇを得た。本菌体に７０％エタノール４００ｍｌを添加後、ミキサーで粉砕抽出し遠心分離により上澄液を得た。上澄液を０．４５μｍメンブレンろ過し、０．３ｋｇの生成物を得た。
【００１６】
＜製造例２＞
グルコース６０ｇ、ペプトン４ｇ、酵母エキス４ｇ、ＫＨ_２ＰＯ_４１．０ｇを精製水２０００ｍｌに溶解後、ｐＨ６．０に調整し、１２０℃で１５分間殺菌し３０℃に冷却後、マツオオジの種菌をこれに接種し、３０℃で通気攪拌培養をグルコース残量が０．５％以下になるまで２８日間行った。培養物から遠心分離により菌体を除いた後、培養液を５倍濃縮し、９９％エタノールを濃縮液と等量添加した。本液を５℃に一晩放置後沈殿物を除去し、活性炭を０．２％添加し、セライト（粘土質ろ過助剤）ろ過を行った。本ろ液を０．４５μｍメンブレンろ過し、０．７ｋｇの生成物を得た。
【００１７】
＜製造例３＞
グルコース１５ｇ、酵母エキス１．５ｇを馬鈴薯抽出液（馬鈴薯２００ｇに水１５００ｍｌを加え煮沸後、ろ布ろ過し１５００ｍｌにフィルアップしたもの）に溶解後、ｐＨ６．０に調整し、１２０℃で１５分間殺菌し３０℃に冷却後、ブナハリタケの種菌をこれに接種し、３０℃で通気攪拌培養をグルコース残量が０．５％以下になるまで１８日間行った。培養物から遠心分離により菌体を集め水洗後、再度遠心分離を行い菌体７８ｇを得た。本菌体に精製水３００ｍｌを加えミキサーで粉砕後分液ロートに移し、ブタノール３００ｍｌを添加し振盪抽出を行った。ブタノール層を分取後、減圧濃縮機で乾固し、８０％エタノール５００ｍｌに溶解した。溶解液にセパビーズＳＰ−２０５（三菱化学（株））５％添加しセライトろ過を行った。本ろ液を０．４５μｍメンブレンろ過し、０．５ｋｇの生成物を得た。
【００１９】
＜製造例４＞
グルコース１５ｇ、エビオス錠（田辺製薬（株））１．５ｇを馬鈴薯抽出液（馬鈴薯２００ｇに水１５００ｍｌを加え煮沸後、ろ布ろ過し１５００ｍｌにフィルアップしたもの）に溶解後、ｐＨ４．５に調整し、１２０℃で１５分間殺菌した。２７℃に冷却後、カンゾウタケの種菌をこれに接種し、通気攪拌培養をグルコース残量が０．２％以下になるまで２７℃で２５日間行った。培養物をナイロンメッシュろ布でろ過し菌体３８ｇを集め水洗した後、菌体に５０％エチレングリコール４００ｍｌを添加し、超音波粉砕機に１５分間かけ抽出を行った。その抽出液をセライトろ過後、ろ液に活性炭０．５％を添加し撹拌後、セライトろ過を行い、さらにセパビーズＳＰ−２０５（三菱化学（株））５％添加し攪拌後、セライトろ過を行った。本ろ液を０．４５μｍメンブレンろ過し、０．４ｋｇの生成物を得た。
【００２０】
＜製造例５＞
グルコース４５ｇ、ペプトン３ｇ、酵母エキス３ｇ、ＫＨ₂ ＰＯ₄ ０．７５ｇを精製水１５００ｍｌに溶解後、ｐＨ５．５に調整し、１２０℃で１５分間殺菌した。３０℃に冷却後、メシマコブの種菌をこれに接種し、通気攪拌培養をグルコース残量が０．２％以下になるまで３０℃で２２日間行った。培養物から遠心分離により菌体を除いた後、その培養液を５倍濃縮し、９９％エタノールを濃縮液と等量添加した。本液を５℃に一晩放置後沈殿物を除去し、活性炭を０．５％添加し攪拌後、セライトろ過を行った。本ろ液を０．４５μｍメンブレンろ過し、０．５ｋｇの生成物を得た。
【００２１】
＜製造例６＞
製造例５の培養物から遠心分離により集めた菌体を水洗後、ナイロンメッシュろ布に取り水分を切り、菌体４５ｇを得た。本菌体に４０％１，３−ブチレングリコール４００ｍｌを添加後、ミキサーで粉砕抽出し遠心分離により上澄液を得た。上澄液にアンバーライトＩＲＣ−５０（オルガノ（株））を５％添加しよく攪拌後、ろ紙ろ過を行った。本ろ液をさらに０．４５μｍメンブレンろ過し、０．４ｋｇの生成物を得た。
【００２４】
＜製造例７＞
グルコース４５ｇ、ペプトン３ｇ、酵母エキス３ｇ、ＫＨ₂ ＰＯ₄ ０．７５ｇ、ＭｇＳＯ₄ ０．５ｇを精製水１５００ｍｌに溶解後、ｐＨ６．０に調整し、１２０℃で１５分間殺菌した。３０℃に冷却後、ヌメリスギタケの種菌をこれに接種し、通気攪拌培養をグルコース残量が０．２％以下になるまで３０℃で２０日間行った。培養物を遠心分離して菌体を除いた後、その培養液を１０倍濃縮し、９９％エタノールを濃縮液と等量添加した。本液を５℃に一晩放置後沈殿物をろ別除去し、活性炭を１％添加し、撹拌後、セライトろ過を行った。本ろ液のＲＯ膜（分子量１０００カット）透過液を集め、本発明品０．２ｋｇを得た。
【００２５】
＜製造例８＞
グルコース４５ｇ、ペプトン３ｇ、酵母エキス３ｇ、ＫＨ₂ ＰＯ₄ ０．７５ｇ、ＭｇＳＯ₄ ０．５ｇを精製水１５００ｍｌに溶解後、ｐＨ６．０に調整し、１２０℃で１５分間殺菌した。３０℃に冷却後、ツクリタケの種菌をこれに接種し、通気攪拌培養をグルコース残量が０．２％以下になるまで３０℃で１７日間行った。培養物を遠心分離して菌体を除いた後、その培養液に活性炭１．５％を添加し、撹拌後、セライト重層ろ過を行った。そのろ液を８倍濃縮し、９９％エタノールとプロピレングリコールの４：１混液を濃縮液と等量添加した。本液を５℃に一晩放置後沈殿物をろ別除去し、セライトろ過を行った。
本ろ液のＲＯ膜（分子量１０００カット）透過液を集め、本発明品０．３ｋｇを得た。
【００２６】
＜試験例１＞マウスメラノーマＢ１６細胞のメラニン生成亢進試験
試験方法
試料をＭＥＭ（Ｅａｇｌｅ’ｓＭｉｎｉｍｕｍＥｓｓｅｎｔｉａｌＭｅｄｉｕｍ）に最終濃度が表１に示す濃度になるように調製、溶解し、孔径０．４５μｍの除菌フィルターでろ過した。ＭＥＭに不溶性の試料は、１００μｌのエタノールに溶解後、ＭＥＭに添加した。
２枚のプラスチックシャーレ（Ｆａｌｃｏｎ製、内径９ｃｍ）にそれぞれ、本発明の試料を溶解・ろ過除菌したＭＥＭを８ｍｌ、ＦＢＳ（ウシ胎児血清）１ｍｌおよびＭＥＭ１ｍｌに懸濁した１×１０^５個／ｍｌのＢ１６細胞を添加し、培養開始３日後に培地交換を行い計５日間、５％ＣＯ_２、９５％空気条件下、３７℃で培養した。培養終了後、シャーレの底に増殖した細胞を集めＰｈｏｓｐｈａｔｅｂｕｆｆｅｒｅｄｓａｌｉｎｅ（ＰＢＳ）に懸濁させ、２，０００ｒｐｍで３分間遠心分離を行い、得られた細胞ペレットの黒化度を肉眼的に評価した。
また、培養終了後の細胞数をカウントし、細胞増殖率を算出した。
表１において、肉眼的色調における＋−は、下記の評価を示す。

【００２７】
試験結果
表１に示すごとく、サンプル添加濃度に依存してＢ１６細胞のメラニン生成亢進効果が認められた。
表１担子菌培養液の抽出液又は菌体抽出液の
マウスメラノーマＢ１６細胞に及ぼす効果
【表１】
【００３０】
考察
表１に示した結果は、製造例毎に有効添加濃度が異なるため、予備試験を行い細胞数の減少が少なくかつ有効性の高い濃度を求めた後、再試験した値である。担子菌の培養液の抽出液及び菌体抽出液には添加量に応じてマウスメラノーマＢ１６細胞のメラニン生成亢進作用が認められたことから白髪防止剤として有用であると考えられる。
【００３１】
＜試験例２＞マウスによる発毛試験
試験方法
試験は、Ｍ．ＳｅｉｊｉおよびＩ．Ａ．Ｂｅｒｎｓｔｅｉｎらが編集の「ノーマルアンドアブノーマルエピダーマルディファレンシェーション（ＮｏｒｍａｌＡｎｄＡｂｎｏｒｍａｌＥｐｉｄｅｒｍａｌＤｉｆｆｅｒｅｔｉａｔｉｏｎ）」第１５９ないし１７０頁（１９８２年、東大出版）に記載されている小川らの方法により行った。
即ち、Ｃ３Ｈ／ＨｅＮＣｒＪマウスを一群２０匹、無処置群、基剤群、実施例（本発明）群の３群に分け、背部の毛を剃り取り、それぞれのサンプルを１日１回、０．１ｍｌ塗布した。４週間後、各群の発毛部分を測定し、剃毛した面積に対する毛再生の認められた面積の割合の変化から効果を比較した。
【００３２】
判定基準
▲１▼発毛の長さ ▲２▼発毛の面積
完全に成長：４＋９０ないし１００％：４＋
ほぼ完全に成長：３＋７０ないし９０％：３＋
半分以上成長：２＋５０ないし７０％：２＋
半分以下の成長：１＋２０ないし５０％：１＋
殆ど成長なし：− ２０％以下：−
【００３３】
供試試料を下記の処方に適用し、ヘアエッセンスを製造した。

【００３４】
試験結果
下記の表２のとおりであった。
【表２】
以上の結果から、本発明の外用剤は優れた発毛効果を有することが明らかである。
【００３８】
＜試験例３＞ヒトによる育毛効果試験
男性型脱毛症患者である被試験者１００名により、育毛効果試験を行った。患者１００名をランダムに２群に分け、第１群には試験剤（本発明）を、第２群には基剤を、１日朝夕２回、患者の頭部毛根部に塗擦し、連続６ケ月間使用した後の効果を次の判定基準で評価した。
判定基準
著効：使用前と比較してかなり増毛したもの。
有効：使用前と比較して増毛したもの。
やや有効：使用前と比較して幾分増毛したもの。
無効：使用前と比較して何ら症状が改善されなかったもの。
副作用：上記塗擦方法による６ケ月後の頭部の皮膚異常の有無。
供試試料を下記の処方に適用し、ヘアートニックを製造した。

［供試試料］
No. １上記処方の培養液として製造例２の培養液を用いたもの
No. ２上記処方の培養液として製造例７の培養液を用いたもの
No. ３基剤（上記処方から培養液を除いたもの）
【００３９】
試験結果
結果を表３に示す。
【表３】
【００４０】
【処方例】
以下に本発明の処方例を挙げる。
処方例中、適量とは、処方全体で１００重量％になる割合を意味する。
【００４１】

１ないし４を均一に攪拌し容器に充填して製品とする。
【００４４】

【００４６】

【００４７】

【００４８】

【００５１】
上記の処方例１ないし５は、いずれも表１−１ないし表２に開示したとおりの発毛効果を有する製剤であることが確認された。
【００５２】
【発明の効果】
本発明によれば、特定のシメジ科、ハリタケ科、カンゾウタケ科、キコブタケ科、モエギタケ科およびハラタケ科からなる群より選択される担子菌の培養液の抽出液または菌体の抽出液群のメラニン生成亢進成分の１種又は２種以上を配合した頭髪料が提供され、この頭髪料は、白髪防止効果と育毛効果にすぐれているばかりでなく、副作用がないという優れた特性を有するものである。[0001]
[Industrial applications]
The present invention relates to the application of an extract of a basidiomycete culture solution or an extract of a bacterial cell to hair material, and more specifically, water and water from a culture solution obtained by removing cells from a culture of a basidiomycete. By combining one or more of the melanin production-enhancing components in the liquid extracted with an organic solvent or the liquid extracted from cultured bacterial cells with water and / or an organic solvent, the hair growth-inhibiting effect is exhibited while the hair gray-inhibiting effect is exhibited. It also relates to the hair charge that you have.
[0002]
[Prior art]
Basidiomycetes (mushrooms) have been conventionally used as edible or herbal medicines. More recently, it has found application in medicine or cosmetics. In particular, Japanese Patent Application Laid-Open Nos. 60-255715 and 4-77725 disclose techniques for applying basidiomycetes to hair.
JP-A-60-255715 discloses a method for producing a hair growth-promoting hair nourishing cosmetic using an extract from fruiting bodies, and JP-B-4-77725 discloses a viscous material obtained by liquid culture of white jellyfish. Which discloses a cosmetic.
[0003]
There are many other conventional basidiomycetes that can be used for pharmaceuticals such as anticancer drugs, antitumor substances, and immunomodulators. There is only a method for obtaining a hair growth-promoting substance, for example, there is JP-A-60-255715, but there is no material having both a gray hair prevention effect and a hair growth effect. In addition, the publication discloses a cosmetic using an extract from the fruit body of a mushroom, but the fruit body is difficult to obtain in large quantities at low cost because it takes time to grow, and even after it is obtained. However, there are many practical problems, such as difficulty in mass production due to variation in usefulness due to individual differences in fruiting bodies and poor extraction rate.
[0004]
[Problems to be solved by the invention]
An object of the present invention is to achieve mass production by producing useful components required for hair materials in a culture solution or bacterial cells without fluctuation and at a high concentration, and further increasing the concentration by further performing various treatments. An object of the present invention is to provide a highly safe hair material having both a gray hair prevention effect and a hair growth effect.
[0005]
[Means for Solving the Problems]
The present inventors have conducted intensive studies in order to solve the above-mentioned problems, and as a result, as a result of extraction of a culture solution obtained by liquid culturing a specific basidiomycete with water and / or an organic solvent, or water and / or an organic solvent of cells. It has been found that the extract obtained by the method has an effect of preventing gray hair, and that the extract has a hair-growth effect. Thus, the present invention has been completed.
That is, according to the present invention, certain lyophyllaceae, Haritake family, mosquitoes Nzoutake family, Phellinus igniarius family, model Egitake family and melanin extract group of the culture broth or bacterial cells of Basidiomycetes selected from the group consisting of agaricaceae There is provided a hair preparation characterized by blending one or more kinds of production enhancing components.
The extract of the culture solution or the extract of the cells of these basidiomycetes is excellent in the effect of preventing melanin and the effect of growing hair by the melanin production-enhancing component.
[0006]
DETAILED DESCRIPTION OF THE INVENTION
The present inventors, for basidiomycetes dozens, gray hair prevention effect and the results so intensively studied to obtain a hair growth effect, certain lyophyllaceae, Haritake family, mosquitoes Nzoutake family, Phellinus igniarius family, the motor Egitake family and agaricaceae Mouse melanoma B16 cells were added to an extract of the culture obtained by basidiomycetes by aeration and stirring liquid culture using a saccharide as a carbon source, and an extract of the culture with water and / or an organic solvent or an extract of the cells with water and / or an organic solvent. It has been found for the first time that there is a component that strongly enhances the production of melanin, and it has also been found that these have a hair-growth effect, thereby completing the present invention.
[0007]
The Basidiomycetes of the present invention, Panellus Serotinus, Matsuooji, Bunaharitake, the beefsteak fungus, Phellinus linteus, Kabanoanatake, highly effective in the characteristics Numerisugitake and Agaricus bisporus was observed.
[0008]
As the extract of the culture solution in the present invention, a basidiomycete inoculum is inoculated into a liquid medium containing a carbon source, a nitrogen source, inorganic salts, and the like, and heated at a temperature of 15 to 35 ° C, preferably 20 to 30 ° C. After aeration and agitation cultivation for from 45 to 45 days, preferably from 10 to 30 days, a solution obtained by removing cells by centrifugation or filtration from the culture solution or a solution obtained by concentrating the solution by a vacuum concentrator or the like is treated with water and / or Those extracted with an organic solvent can be disclosed as suitable ones.
[0009]
As the organic solvent, methanol, ethanol, isopropyl alcohol, butanol, glycerin, ethylene glycol, 1,3-butylene glycol, acetone, chloroform, ethyl acetate, hexane, ether and the like are preferable, and among them, ethanol, isopropyl alcohol, butanol, glycerin , Ethylene glycol and 1,3-butylene glycol are preferably used. After further diluting or concentrating the extract to further enhance the efficacy and safety, the extract is subjected to ultrafiltration or reverse osmosis membrane treatment or activated carbon or various resins, for example, Sepabeads SP-205 (Mitsubishi Chemical Corporation) ) Etc., or those obtained by diluting or concentrating these treatment solutions.
[0010]
As the cell extract in the present invention, a cell obtained by centrifugation or filtration from the above culture solution as it is or after chopping, and sufficiently extracted with water and / or an organic solvent, or a concentrated solution thereof is used. It can be disclosed as suitable. Examples of the organic solvent include methanol, ethanol, isopropyl alcohol, butanol, glycerin, ethylene glycol, 1,3-butylene glycol, acetone, chloroform, ethyl acetate, hexane and ether. Among them, ethanol, butanol, glycerin, ethylene Glycol and 1,3-butylene glycol are preferably used. In order to further enhance the efficacy and safety, the extract is diluted or concentrated and then subjected to ultrafiltration or reverse osmosis membrane treatment or activated carbon or various resins such as Sepabeads SP-205 (Mitsubishi Chemical Corp. )) And the like, or those obtained by diluting or concentrating the treated liquids. The extract of the basidiomycete culture solution or the bacterial cell extract according to the present invention obtained as described above is excellent in the effect of preventing gray hair due to the action of enhancing melanin production, and at the same time, has a hair-growth effect, and furthermore, it has an effect on the skin. It does not cause any damage and has excellent safety.
[0011]
The hair preparation of the present invention is produced by blending the above-mentioned active ingredients into a known dosage form acceptable for hair application, and the amount of the preparation is determined by the culturing method, treatment method, degree of concentration, and formulation to be mixed. In general, it is preferable to add about 0.05 to 50% by weight, preferably about 0.1 to 10% by weight of a culture solution, a cell extract, or a concentrate thereof in the preparation.
[0012]
Furthermore, in addition to the active ingredient of the present invention, various known active ingredients usually used, such as carpronium chloride, cepharanthin, vitamin E, vitamin E nicotinate, nicotinic acid, nicotinamide, benzyl nicotinate, tincture tincture and pepper. Peripheral vasodilators such as tincture, cooling agents such as camphor and menthol, antibacterial agents such as hinokitiol, benzalkonium chloride and undecylenic acid, anti-inflammatory agents such as lysozyme chloride, glycyrrhizin and allantoin, assembly extracts, garlic extract, carrot extract, Various extracts derived from animals, plants, and microorganisms, such as pentagonal extract, rosemary extract, aloe extract, luffa extract, ginkgo extract, elder extract, placenta extract, liver extract, and lactic acid bacteria culture extract It can be used by adding to the reason.
[0013]
As described above, the known dosage form of the external preparation of the present invention means any dosage form that can be applied externally, such as hair cream, hair essence, shampoo, rinse, hair tonic, hair liquid, hair spray, and the like. A hair application agent such as a hair foam can be exemplified. In addition, in addition to known active ingredients, the above-mentioned external preparations include surfactants, base components such as fats and oils, and if necessary, known humectants, thickeners, preservatives, antioxidants, and ultraviolet rays. Various additives such as absorbers / scatterers, chelating agents, pH adjusters, fragrances and coloring agents can be used as appropriate.
[0014]
【Example】
Hereinafter, Production Examples of the present invention, Test Examples and Formulation Examples for explaining the effects thereof will be given, but these do not limit the present invention at all.
[0015]
<Production Example 1>
45 g of glucose, 3 g of peptone, 3 g of yeast extract and 0.75 g of KH ₂ PO ₄ are dissolved in 1500 ml of purified water, adjusted to pH 6.0, sterilized at 120 ° C. for 15 minutes and cooled to 28 ° C. And cultivated with aeration and agitation at 28 ° C. for 22 days until the residual amount of glucose became 0.3% or less. The cells were collected from the culture by centrifugation, washed with water, taken on a nylon mesh filter cloth and drained to obtain 48 g of cells. After adding 400 ml of 70% ethanol to the cells, the mixture was pulverized and extracted with a mixer and centrifuged to obtain a supernatant. The supernatant was filtered through a 0.45 μm membrane to obtain 0.3 kg of the product.
[0016]
<Production Example 2>
After dissolving 60 g of glucose, 4 g of peptone, 4 g of yeast extract, and 1.0 g of KH ₂ PO _{4 in} 2000 ml of purified water, adjusted to pH 6.0, sterilized at 120 ° C. for 15 minutes and cooled to 30 ° C. , And cultivated with aeration and agitation at 30 ° C. for 28 days until the residual amount of glucose became 0.5% or less. After removing the cells from the culture by centrifugation, the culture was concentrated 5-fold, and 99% ethanol was added in the same amount as the concentrate. After leaving this solution at 5 ° C. overnight, the precipitate was removed, and 0.2% of activated carbon was added, followed by filtration through Celite (clay filter aid). The filtrate was filtered through a 0.45 μm membrane to obtain 0.7 kg of a product.
[0017]
<Production Example 3>
15 g of glucose and 1.5 g of yeast extract were dissolved in a potato extract (200 g of potato, 1500 ml of water was added, boiled, filtered through a filter cloth and filled up to 1500 ml), adjusted to pH 6.0, and adjusted to 120 ° C. for 15 minutes. After sterilization and cooling to 30 ° C., a seed of B. agaricus was inoculated into the mixture, and aeration and agitation culture was performed at 30 ° C. for 18 days until the residual amount of glucose became 0.5% or less. The cells were collected from the culture by centrifugation, washed with water, and then centrifuged again to obtain 78 g of cells. 300 ml of purified water was added to the cells, and the mixture was pulverized with a mixer, transferred to a separating funnel, and 300 ml of butanol was added, followed by shaking extraction. After separating the butanol layer, it was dried with a vacuum concentrator and dissolved in 500 ml of 80% ethanol. 5% of Sepabeads SP-205 (Mitsubishi Chemical Corporation) was added to the solution, and celite filtration was performed. The filtrate was filtered through a 0.45 μm membrane to obtain 0.5 kg of a product.
[0019]
<Production Example 4 >
15 g of glucose and 1.5 g of Ebios tablets (Tanabe Seiyaku Co., Ltd.) were dissolved in a potato extract (200 g of potato, added with 1500 ml of water, boiled, filtered through a filter cloth and filled up to 1500 ml), and adjusted to pH 4.5. And sterilized at 120 ° C. for 15 minutes. After cooling to 27 ° C., a seed of liquorice mushroom was inoculated thereto, and aeration and agitation culture was performed at 27 ° C. for 25 days until the residual amount of glucose became 0.2% or less. The culture was filtered through a nylon mesh filter cloth to collect 38 g of bacterial cells, washed with water, added with 400 ml of 50% ethylene glycol, and extracted with an ultrasonic grinder for 15 minutes. The extract was filtered through celite, activated carbon 0.5% was added to the filtrate, stirred, filtered through celite, and 5% of Sepabeads SP-205 (Mitsubishi Chemical Corporation) was added, stirred, and filtered through celite. Was. The filtrate was filtered through a 0.45 μm membrane to obtain 0.4 kg of a product.
[0020]
<Production Example 5 >
45 g of glucose, 3 g of peptone, 3 g of yeast extract, and 0.75 g of KH ₂ PO ₄ were dissolved in 1500 ml of purified water, adjusted to pH 5.5, and sterilized at 120 ° C. for 15 minutes. After cooling to 30 ° C., a seed of Mesimakobu was inoculated into this, and aeration and agitation culture was performed at 30 ° C. for 22 days until the residual amount of glucose became 0.2% or less. After removing the cells from the culture by centrifugation, the culture was concentrated 5-fold, and 99% ethanol was added in the same amount as the concentrate. After leaving this solution at 5 ° C. overnight, the precipitate was removed, activated carbon was added by 0.5%, and the mixture was stirred and filtered through celite. The filtrate was filtered through a 0.45 μm membrane to obtain 0.5 kg of a product.
[0021]
<Production Example 6 >
The cells collected by centrifugation from the culture of Production Example 5 were washed with water, taken on a nylon mesh filter cloth and drained to obtain 45 g of cells. After adding 400 ml of 40% 1,3-butylene glycol to the cells, the mixture was pulverized and extracted with a mixer and centrifuged to obtain a supernatant. 5% of Amberlite IRC-50 (Organo Co., Ltd.) was added to the supernatant, and the mixture was stirred well and filtered through a filter paper. The filtrate was further filtered through a 0.45 μm membrane to obtain 0.4 kg of a product.
[0024]
<Production Example 7 >
45 g of glucose, 3 g of peptone, 3 g of yeast extract, 0.75 g of KH ₂ PO ₄ and 0.5 g of MgSO ₄ were dissolved in 1500 ml of purified water, adjusted to pH 6.0, and sterilized at 120 ° C. for 15 minutes. After cooling to 30 ° C., seeds of Numeris mushroom were inoculated into the inoculated culture, and aeration and agitation culture was performed at 30 ° C. for 20 days until the residual amount of glucose became 0.2% or less. After removing the cells by centrifuging the culture, the culture was concentrated 10-fold, and 99% ethanol was added in the same amount as the concentrate. After leaving this solution at 5 ° C. overnight, the precipitate was removed by filtration, 1% of activated carbon was added, and the mixture was stirred and filtered with celite. The permeated liquid of the filtrate obtained from an RO membrane (molecular weight cut: 1,000) was collected to obtain 0.2 kg of the product of the present invention.
[0025]
<Production Example 8 >
45 g of glucose, 3 g of peptone, 3 g of yeast extract, 0.75 g of KH ₂ PO ₄ and 0.5 g of MgSO ₄ were dissolved in 1500 ml of purified water, adjusted to pH 6.0, and sterilized at 120 ° C. for 15 minutes. After cooling to 30 ° C., an inoculum of Amanita mushroom was inoculated into this, and aeration and agitation culture was performed at 30 ° C. for 17 days until the residual amount of glucose became 0.2% or less. After removing the cells by centrifuging the culture, 1.5% activated carbon was added to the culture, and the mixture was stirred and filtered through Celite double-layer filtration. The filtrate was concentrated 8-fold, and a 4: 1 mixture of 99% ethanol and propylene glycol was added in the same amount as the concentrated solution. After leaving this solution at 5 ° C. overnight, the precipitate was removed by filtration and filtered through celite.
The permeated liquid of the RO membrane of the present filtrate (molecular weight cut: 1,000) was collected to obtain 0.3 kg of the product of the present invention.
[0026]
<Test Example 1> Melanin production enhancement test of mouse melanoma B16 cells
Test method A sample was prepared and dissolved in MEM (Eagle's Minimum Essential Medium) so that the final concentration was as shown in Table 1, and filtered with a sterilizing filter having a pore size of 0.45 µm. The sample insoluble in MEM was dissolved in 100 μl of ethanol and then added to MEM.
A sample of the present invention was dissolved and filtered and sterilized in 8 ml, FBS (fetal bovine serum) 1 ml, and 1 × 10 ⁵ cells / ml suspended in 1 ml of MEM, respectively, in two plastic dishes (Falcon, 9 cm inside diameter). 3 days after the start of the culture, the medium was replaced, and the cells were cultured at 37 ° C. under 5% CO ₂ and 95% air conditions for a total of 5 days. After completion of the culture, the cells grown on the bottom of the petri dish were collected, suspended in Phosphate buffered saline (PBS), and centrifuged at 2,000 rpm for 3 minutes, and the degree of blackening of the obtained cell pellet was visually evaluated. .
Further, the number of cells after the completion of the culture was counted, and the cell growth rate was calculated.
In Table 1, +-in the visual color tone indicates the following evaluation.

[0027]
Test results As shown in Table 1 , the melanin production-enhancing effect of B16 cells was observed depending on the sample addition concentration.
Table 1 Effect of basidiomycete culture extract or cell extract on mouse melanoma B16 cells
[0030]
Discussion The results shown in Table 1 show that since the effective addition concentration differs for each production example, a preliminary test was performed to find a concentration with a small decrease in the number of cells and a high effectiveness, and then the values were retested. is there. The extract of the culture of basidiomycetes and the extract of the bacterial cells showed melanin production-enhancing activity of mouse melanoma B16 cells depending on the amount added, and thus are considered to be useful as graying inhibitors.
[0031]
<Test Example 2> Hair growth test using mouse
Test method The test was performed according to M.E. Seiji and I.S. A. Bernard et al., Edited by Ogawa et al., Described in "Normal And Abnormal Epidermal Differentiation", pp. 159 to 170 (1982, published by The University of Tokyo), "Normal and Abnormal Epidermal Differentiation".
That is, C3H / HeNCrJ mice were divided into three groups: a group of 20 mice, a non-treatment group, a base group, and an example (the present invention) group. The back was shaved, and each sample was taken once a day. 1 ml was applied. Four weeks later, the hair growth portion of each group was measured, and the effect was compared based on the change in the ratio of the area where hair regeneration was observed to the shaved area.
[0032]
Judgment criteria ( 1) Hair growth length (2) Hair growth area Complete growth: 4+ 90 to 100%: 4+
Almost completely grown: 3+ 70-90%: 3+
More than half growth: 2+ 50-70%: 2+
Less than half growth: 1+ 20-50%: 1+
Almost no growth:-20% or less:-
[0033]
The test sample was applied to the following formulation to produce a hair essence .

[0034]
The test results were as shown in Table 2 below.
[Table 2]
From the above results, it is clear that the external preparation of the present invention has an excellent hair growth effect.
[0038]
<Test Example 3 > Human Hair Growth Effect Test A hair growth effect test was conducted by 100 test subjects who were male pattern baldness patients. 100 patients were randomly divided into two groups, the test group (the present invention) was applied to the first group, and the base was applied to the second group twice daily in the morning and evening on the hair roots of the patient's head. The effect after use for 6 months was evaluated according to the following criteria.
Judgment criterion Excellent effect: The hair increased considerably compared to before use.
Effective: Increased hair compared to before use.
Somewhat effective: Somewhat increased hair compared to before use.
Ineffective: No improvement in symptoms as compared to before use.
Side effects: presence or absence of abnormal skin on the head after 6 months by the above-mentioned rubbing method.
The test sample was applied to the following formulation to produce a hair tonic .

[ Test sample ]
No. 1 Using the culture solution of Production Example 2 as the culture solution of the above formulation
No. 2 Using the culture solution of Production Example 7 as the culture solution of the above formulation
No. 3 Base (excluding the culture solution from the above formulation )
[0039]
Test results The results are shown in Table 3 .
[Table 3]
[0040]
[Example of prescription]
Examples of the formulation of the present invention are described below.
In the formulation examples, the appropriate amount means a ratio of 100% by weight in the whole formulation.
[0041]

1 to 4 are uniformly stirred and filled into a container to obtain a product.
[0044]

[0046]

[0047]

[0048]

[0051]
It was confirmed that each of the above Formulation Examples 1 to 5 was a preparation having a hair growth effect as disclosed in Tables 1-1 and 2 .
[0052]
【The invention's effect】
According to the present invention, melanin production of an extract of a culture solution of basidiomycetes or a group of extract of fungi selected from the group consisting of a specific Shimeji family, Agaricaceae family, Agaricaceae family, Asteraceae family, Asteraceae family, Moie mushroom family and Agaricaceae family A hair material containing one or more enhancer components is provided, and the hair material has not only an excellent white hair preventing effect and a hair-growth effect, but also has excellent properties of no side effects.

Claims

Mutaketake, matsuooji, beecharitake, licorice mushroom, meshimakobu, kabanoanatake,
A hair care composition comprising one or more melanin production-enhancing components of a basidiomycete culture solution or a cell extract extract group selected from the group consisting of Numerisugitake and Agaricus .