JP4730642B2 - Active oxygen scavenger and composition thereof - Google Patents
Active oxygen scavenger and composition thereof Download PDFInfo
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- JP4730642B2 JP4730642B2 JP2003151401A JP2003151401A JP4730642B2 JP 4730642 B2 JP4730642 B2 JP 4730642B2 JP 2003151401 A JP2003151401 A JP 2003151401A JP 2003151401 A JP2003151401 A JP 2003151401A JP 4730642 B2 JP4730642 B2 JP 4730642B2
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Description
【0001】
【発明の属する分野】
本発明は活性酸素消去剤を特定の植物樹皮から得るもので、より詳しくは特定植物の樹皮の微粉末あるいは水/低級アルコールを溶剤として抽出して得られる抽出物を有効成分とする食品、飼料、化粧品、医薬部外品、医薬品あるいは抗酸化剤等に配合して用いられる活性酸素消去剤に関するものである。
【0002】
【従来の技術】
一般的に大気中において、酸素は三重項酸素と呼ばれる安定な物質である。その酸素の一部が、生体内でスーパーオキシドやヒドロキシラジカル、一重項酸素、過酸化水素等の活性酸素と呼ばれる反応性の高い物質に変化すると、その一部が好中球やマクロファージで生産され異物に対する生体防御の役割を担う一方で、活性酸素が過剰に生体内に存在すると、脂質、タンパク質、酵素、核酸等を攻撃し、生体膜や組織などを損傷し、動脈硬化、脳心血管障害、肺気腫、癌、リウマチ性関節炎、白内障、高血圧、老人性痴呆、アルツハイマー病あるいはシミ・ソバカスや老化等の発症の原因にもなることが明らかにされている。
【0003】
従って、従来からこれらの過剰な活性酸素による酸化ストレスが引き起こす上述のような各種疾病を予防するために、出発物質であるスーパーオキシド(02 -)を不均化して過酸化水素と酸素分子を生じる反応を触媒する酵素であるスーパーオキシドディスムターゼ(SOD)と類似する働きをするSOD様物質や、活性酸素を捕捉する物質(活性酸素消去活性物質)を食品へ添加する抗酸化物質の探索が広く行われてきた。
【0004】
例えば、この様な物質としては、天然物由来の物質では脂溶性のα-トコフェロール(ビタミンE)や水溶性のアスコルビン酸(ビタミンC)が挙げられ、合成物質ではBHT(ブチルヒドロキシトルエン)、BHA(ブチルヒドロキシアニソール)、TBHQ(第三級ブチルヒドロキノン)等のフェノール性物質がある。
しかしながら、これらの物質は食品の抗酸化物質として一般的には使用されているものの、生体への活性酸素消去作用が十分ではなく、特に合成物質のBHAでは発癌性の疑いが生じる等の問題もでてきている。
【0005】
そこで、近年、動植物等の天然物から安全性の高い活性酸素消去活性物質を探索する研究開発が盛んに行われており、例えば、特許文献1、2、3、4、5等がある。また、実際に製品化まですすんだものとしては、特に、茶葉、カカオ豆エキスから抽出されるカテキン類、ブルーベリー果皮に含有されるアントシアニン系化合物、発酵大豆中から得られるイソフラボン類等、ポリフェノール系化合物が非常に高い活性酸素消去活性があることで注目を集めている。
【0006】
これらの活性酸素消去活性が高いことが明らかになってきたポリフェノール系化合物の中でも特に茶葉に含まれる(−)エピガロカテキン、(−)エピガロカテキン−3−ガレート、(−)エピカテキン、(−)エピカテキン−3−ガレート、(+)カテキンなど、フラバン−3−オールを基本骨格とするフラバノールであるカテキン類が、他のポリフェノール類よりも高い活性酸素消去活性があることがわかってきた。
【0007】
一方で、アカシア属の一種である学名:Acacia mearnsii De Wild. の樹皮から水で抽出されるタンニンは古くから大量に南アフリカやブラジルで生産されており、一般的にはワトルタンニンと呼ばれ、主に皮なめし剤や木材用接着剤として非常に安価で利用されている。また、Acacia mangium Willd.(アカシアマンギュウム)は生長が非常に早いことからも、近年、インドネシア、マレーシアを始めとする亜熱帯〜熱帯で植林が盛んになってきている。このワルトタンニンは、縮合型タンニンに分類されるものである。
【0008】
しかしながら、上記の学名:Acacia mearnsii De Wild.樹皮から熱水等で抽出して得られるタンニンはもとより、Acacia mangium Willd.の様なアカシア属の樹皮を粉砕して得られる樹皮微粉末について、活性酸素を消去する作用は全く知られていなかった。更に、これらを食品、飼料、化粧品、医薬部外品、医薬品あるいは抗酸化剤等へ添加、配合する等して、上述した疾患の予防や治療、老化の防止や改善のために用いる試みはされていなかった。
【0009】
【特許文献1】
特開平01−25726号公報
【特許文献2】
特開平6−65074号公報
【特許文献3】
特開平7−300422号公報
【特許文献4】
特開平11−5975号公報
【特許文献5】
特開2001−98264号公報
【0010】
【発明が解決しようとする課題】
本発明は、上記観点に着眼し、人体及び動物に対して安全性が高く、生体内で発生する過剰の活性酸素を十分に消去する作用を有する活性酸素消去剤及びこれを含有する食品、飼料、化粧品、医薬部外品、医薬品あるいは抗酸化剤等を安価に提供することにある。
【0011】
【課題を解決するための手段】
本発明者らは、上記観点に着眼し、活性酸素消去活性(SOSA;Super Oxide Scavenging Activities)を指標に鋭意研究を重ねていたところ、アカシア属の樹皮に高い活性酸素消去活性があることを見出して本発明を完成した。
アカシア属樹皮の高い活性酸素消去活性は、該樹皮中に含まれる或る一成分によるものか、種々の成分に基づく複合効果によるものか特定できないが、アカシア属樹皮によって得ることができる。
上記活性酸素消去活性は、アカシア属樹皮に共通して示されるが、アカシア属の樹木のうち特に、学名:Acacia mearnsii De Wild.(一般名ブラックワトル)、学名:Acacia mangium Willd.(一般名アカシアマンギュウム)、学名:Acacia dealbata Link.、学名:Acacia decurrens Willd.、及び学名:Acacia pycnantha Benth.からなる群より選ばれるものの樹皮が好ましい。
【0012】
【発明の実施の形態】
アカシア属樹皮による活性酸素消去作用は、樹皮自体を用いても得ることができ、また、樹皮から溶媒によって抽出された抽出物によっても得ることができる。
アカシア属樹皮のうちでも、特に学名:Acacia mearnsii De Wild.(一般名ブラックワトル)、学名:Acacia mangium Willd.(一般名アカシアマンギュウム)、学名:Acacia dealbata Link.、学名:Acacia decurrens Willd.、及び学名:Acacia pycnantha Benth.からなる群より選ばれる樹皮並びにこれら樹皮からの抽出物に非常に高い活性酸素消去作用があることを見い出したことに重要な意義がある。したがって本発明は、これら学名のアカシア樹皮を含むアカシア属樹皮に基づく活性酸素消去剤及びこれを含有する食品、飼料、化粧品、医薬部外品、医薬品あるいは抗酸化剤等の組成物からなるものである。
【0013】
活性酸素消去活性を目的にアカシア属の樹皮を用いるときは、目的に応じて適当な大きさに破砕または粉砕して用いるとよく、特に微粉砕して用いるとよい。
樹皮は、通常、樹木として伐採したのち乾燥し、剥がして樹皮だけ採取し、その後好ましくは更に天日で乾燥するとよい。
アカシア属の樹木(以下、単にアカシアという)の樹皮は、外皮とやや繊維質の内皮とからなり、含水率20%程度以下に乾燥するとハンマーミル等の粉砕機で容易に微粉化する。アカシア樹皮は、内皮、外皮ともに活性酸素消去成分を含み、この内皮と外皮を分離して用いてもよい。
活性酸素消去剤として、この樹皮微粉末をそのまま用いる場合には、分級して好ましくは100μm以下、より好ましくは有効成分(活性酸素消去成分)を80〜99%含有する50ないし70μm以下のものを用いるのがよい。
活性成分を多く含む微粉末の分画は、含水率20%以下に乾燥した樹皮を適当な大きさ、例えば粒径1.6mm以下程度に粉砕し、生じた粉末を振動ふるい機等で分級して所要の微粉末を得る。
上記とは別に、アカシア樹皮を乳牛等の飼料などに用いる場合には、適当な大きさに粉砕した樹皮をそのまま用いることもできる。
【0014】
本発明のアカシア樹皮からの抽出物としての活性酸素消去成分を得るためには、低級アルコールや極性溶媒で抽出するのが好ましい。低級アルコールとしてはメタノール、エタノール、プロパノール、イソプロピルアルコール等、極性溶媒としては水、アセトン、酢酸エチル等が使用され、必要に応じて単独あるいは2種以上を併せて使用してもよい。水/アルコールの混合溶媒と使用してもよく、水又は熱水抽出物を更にアルコール等の溶媒で抽出してもよい。このように、異なる溶媒によって多段回に抽出することによって、より高い活性の成分を得ることができる。
したがって更に別の本発明は、アカシア樹皮からの水又は熱水による抽出物を更にアルコール等の溶媒で抽出した抽出物に関するものである。
【0015】
抽出は、アカシア樹皮に溶媒を加えて必要に応じて攪拌し抽出するが、温度や時間あるいは固液比については特に限定されない。溶媒に水を用いる場合には、熱水で抽出してもよい。抽出して得られた活性酸素消去成分を含有する抽出液は、そのまま凍結乾燥あるいは噴霧乾燥してもよいし、あるいは減圧濃縮してから凍結乾燥あるいは噴霧乾燥してもよい。得られる抽出物は、抽出液、溶液、粉末、濃縮液、ペースト状物等の種々の形態とすることができ、広く必要に応じて使用できる。さらに、これらの形態で得られた本発明のアカシア樹皮抽出物はそのまま活性酸素消去成分として使用できるほか、さらに必要に応じて精製し、その精製物も活性酸素消去成分として使用することができる。
【0016】
また、本発明の活性酸素消去成分はそのまま活性酸素消去剤として用いられるほか、食品、飼料、化粧品、医薬部外品、医薬品あるいは酸化物質捕捉剤等に適宜、目的に応じて任意の形態で配合して用いることができる。
本発明のアカシアの樹皮は、適当な大きさに粉砕して、好ましくは上記のように微粉砕してそのまま飼料又は食品組成物に配合して用いられる。また、活性成分はアカシア樹皮粉砕物から熱水で抽出できることから、ティーパック状として供給し日常の健康飲料としてたしなむこともできる。
【0017】
先に具体的に示したアカシア樹皮の他に、同様に高い活性を示すものとして、アカシア属のうち、学名:Acacia storyi、学名:Acacia glaucocarpa、学名:Acacia neriifolia、学名:Acacia trachyphloiaの樹皮を挙げることができ、また、アカシア樹皮と同様に活性酸素消去活性を示すものとしてユーカリ属のうち学名:Eucalyptus diversicolor(一般名カリ)の材及び樹皮やケプラコ属のケプラコ材を示すことができる。
【0018】
【実 施 例】
以下、製造例、試験例、処方例を挙げて本発明を更に詳しく具体的に説明するが、本発明はこれらに限定されるものではない。特に、ここでは本発明のアカシア樹皮を外皮と内皮に分けないで実施例を示しているが、外皮を内皮から分離してそれぞれ使用することもできる。
以下の製造例、試験例等において、本発明の各アカシアをそれぞれの学名の後の括弧内に示した番号で示す。例えば、学名:Acacia mearnsii De Wild.のアカシアをアカシアNo.1と記す。
学名:Acacia mearnsii De Wild.(No.1)、学名:Acacia mangium Willd.(No.2)、学名:Acacia dealbata Link.(No.3)、学名: Acacia decurrens Willd.(No.4)、学名:Acacia pycnantha Benth.(No.5)。
【0019】
製造例1
アカシアNo.1の樹皮を含水率20%以下まで乾燥し、その乾燥樹皮をハンマーミル(Raymond社製)で1.6mm以下(10メッシュ篩(タイラー:Tyler)通過)の粉末に粉砕した後、更に振動ふるい機で分級し、63μm以下(250メッシュ篩下)の微粉末を得た。
同様にして、残り4種のアカシアNo.2〜5の樹皮を粉砕してそれぞれ63μm以下の微粉末を得た。種類によって250メッシュ篩通過の微粉末の収率に多少の差はあるが、目的とする微粉末が得られた。
【0020】
製造例2
本発明の各アカシアNo.1〜5の樹皮をそれぞれ含水率20%以下まで乾燥し、その乾燥樹皮をハンマーミルで1.6mm以下の粉末に粉砕した後、この乾燥粉砕樹皮100gに対して5倍量の熱水を加え、沸騰してから15分間抽出し、10〜20μmのフィルターを用いて濾過した。得られた濾液をスプレードライヤで噴霧乾燥し、樹皮抽出物各30gを得た。
以下、各樹皮抽出物はアカシアNo.1〜5熱水抽出物と記す。
【0021】
製造例3
本発明の各アカシアの樹皮をそれぞれ含水率20%以下まで乾燥し、その乾燥樹皮をハンマーミルで1.6mm以下の粉末に粉砕した後、この乾燥粉砕樹皮100gに対して3倍量のメタノールを加え、沸騰させて還流させながら15分間抽出し、10〜20μmのフィルターを用いて濾過した。得られた濾液からメタノールを蒸発させた後、濃縮液をクローズドスプレードライヤで噴霧乾燥し、樹皮抽出物45gを得た。
以下、各樹皮抽出物をアカシアNo.1メタノール抽出物の如く記す。
アカシアNo.1〜5のメタノール抽出物を得た。
【0022】
製造例4
本発明のアカシアの樹皮を含水率20%以下まで乾燥し、その乾燥樹皮をハンマーミルで1.6mm以下の粉末に粉砕した後、この乾燥粉砕樹皮100gに対して3倍量のエタノールを加え、沸騰させて還流させながら15分間抽出し、10〜20μmのフィルターを用いて濾過した。得られた濾液からエタノールを蒸発させた後、濃縮液をクローズドスプレードライヤで噴霧乾燥し、樹皮抽出物(以下、アカシアNo.1エタノール抽出物の如く記す)45gを得た。
アカシアNo.1〜5のエタノール抽出物を得た。
【0023】
製造例5
製造例2で得られたアカシア熱水抽出物10gに3倍量のメタノールを加え、沸騰させて還流させながら15分間抽出し、10〜20μmのフィルターを用いて濾過した。得られた濾液からメタノールを蒸発させて、それに水を加えてから凍結乾燥させて9gの抽出物(以下、アカシアNo.1熱水抽出物メタノール画分の如く記す)を得た。
アカシアNo.1〜5の熱水抽出物メタノール画分を得た。
【0024】
製造例6
製造例2で得られたアカシア熱水抽出物10gに3倍量のエタノールを加え、沸騰させて還流させながら15分間抽出し、10〜20μmのフィルターを用いて濾過した。得られた濾液からエタノールを蒸発させて、それに水を加えてから凍結乾燥させて9gの抽出物(以下、アカシアNo.1熱水抽出物エタノール画分の如く記す)を得た。
アカシアNo.1〜5の熱水抽出物エタノール画分を得た。
【0025】
試験例 1 活性酸素消去活性試験
上記製造例で調製したアカシア抽出物の活性酸素消去活性を下記図式で示す反応式に基づき、ヒポキサンチン−キサンチンオキシダーゼ(XOD)系により活性酸素の一つであるスーパーオキシドアニオン(O2ー)を発生させ、発生したO2ーをスピントラップ剤(DMPO:5,5-Dimethyl-1-pyrroline-N-oxide)で捕捉し、ESR(電子スピン共鳴)法を用いて測定した。
【化1】
【0026】
試料の調製
上記製造例1〜6で得られた粉末から5gを秤りとり、0.1Mリン酸緩衝液50mLを加え、攪拌子を入れ、氷冷しながら60分間攪拌抽出する。遠心分離(3,000rpm、5分間)又はろ過(フィルターNo.1)した液を試料溶液とする。
【0027】
測定操作
あらかじめ、小試験管に9.2M DMPO(5,5-Dimethyl-1-pyrroline-N-oxide、ラボテック、冷凍保存品)を15μL、5.5mM DETAPAC(ジエチレントリアミンペンタ酢酸、シグマ、冷凍保存品)を35μL、2mMヒポキサンチン(シグマ、冷凍保存品)を50μL加え混合しておく。この試験管にSOD標準溶液(SOD、東洋紡、3,000単位/mgを10mg秤り取り、0.1Mリン酸緩衝液に溶かし200mLとしたもの)または試料溶液50μLを加え、次いでキサンチンオキシダーゼ溶液(キサンチンオキシダーゼ、ベーリンガー・マンハイム、20単位/mLを20μL取り、0.1Mリン酸緩衝液980μLを加えたもの)50μLを加える時にフットスイッチを押す。試験管をボルテックス等で攪拌し、特殊偏平セル(日本電子、約130μL)に吸い取り、ESR装置にセットする。フットスイッチを押した40秒後に掃引を開始する。
【0028】
活性の評価
スペクトルの読み取りは、DMPO−OOHシグナルの線幅が一定であるので、このシグナルの高さを、内標準として用いているMn2+のシグナルの高さと比較する(比をとる)方法で行う。
【0029】
SOD標準溶液は、0〜20単位/mL濃度で6点程度測定し、検量線を作成する。SODが0の時の比(DMPO-OOHシグナル/Mn2+シグナル=I0)、及びSOD各濃度での比(In)を求める。検量線の縦軸は(I0/In)−1とし、横軸にSOD濃度をとる。
【0030】
試料溶液の濃度は再現性を高めるため、約10単位/mLとなるように0.1Mリン酸緩衝液で希釈してから測定する。検量線から試料溶液中のSOD濃度(SOSA濃度)を求める。
ESRはJES−FR80(日本電子)を用い、マイクロ波出力8mW、磁場掃引幅335.4mT±5mT、掃引時間2分、磁場変調100kHz、0.79×0.1mT、増幅率2×100、応答時間0.1秒の条件で測定する。
Dを希釈率、Wを試料採取量(g)とし、SOSA(単位/g)を試料溶液の活性(単位/mL)×D×(50/W)で求める。
【0031】
結果を、比較品の測定結果と併せて表1に示す。
【表1】
【0032】
表1から本発明の製造例1〜6で得た樹皮粉末及び抽出物の中では、熱水抽出物エタノール画分が最も活性酸素消去作用が高く、アスコルビン酸(ビタミンC)及び(+)カテキンの約7倍の効果を有することがわかる。
【0033】
試験例2 急性毒性試験(経口投与)
OECD(Guidelines for the Testing of Chemicals 401、1987)に準拠し、マウスを用いた急性経口毒性試験を実施した。
ICR系雌雄マウスについて、雄では7,000mg/kg、雌では6,500mg/kgを上限として、公差500mg/kgで各9用量の検体(アカシアNo.1(学名:Acacia mearnsii De Wild.)熱水抽出物)を雌雄マウスに単回経口投与した。検体は、本発明品を純水に懸濁して用いた。その結果、LD50値は雄では4,468mg/kg、雌では3,594mg/kgであり、いずれも3,000mg/kgの投与量で、死亡例を認めなかった。
上記試験をアカシアNo.2〜5の熱水抽出物について行って同様の結果を得た。これにより、本発明の活性酸素消去剤は、安全性に優れていることがわかる。
【0034】
試験例3 突然変異誘起性試験
労働省告示第77号(昭和63年9月1日)に準拠し、突然変異誘起性試験を実施した。
Escherichia coli WP2 uvrA株及びSalmonella typhimurium TA株系4菌株を用いて代謝活性化を含む復帰突然変異試験を156〜5,000μg/プレートの用量の検体アカシアNo.1(学名:Acacia mearnsii De Wild.)熱水抽出物で行った。その結果、いずれの菌株についても復帰変異コロニー数の増加が見られなかったことから、本発明品の突然変異誘起性は陰性であると結論づけられ、安全性に優れていることがわかる。
【0035】
処方例1 内服剤の調製
製造例6のアカシア樹皮熱水抽出物エタノール画分を用い、下記に示す組成にて内服剤を調製した。
【0036】
処方例2 ペットフードの調製
製造例2のアカシア樹皮熱水抽出物を用い、下記に示す組成にてペットフードを調製した。
【0037】
処方例3 錠剤(菓子)の調製
製造例6のアカシア樹皮熱水抽出物エタノール画分を用い、下記に示す組成にて錠剤(菓子)を調製した。
【0038】
処方例4 化粧水の調製
製造例4のアカシア樹皮エタノール抽出物を用い、下記に示す組成にて均一に攪拌、溶解し、化粧水を調製した。
【0039】
【発明の効果】
本発明は、アカシアの樹皮が活性酸素消去作用を示すことを見出して完成されたもので、樹皮を微粉砕し分級することでより活性の強い画分を得ることができ、天然植物由来のものであるため安全で安価に大量に供給できるため、健康食品や飼料等に使用できる。また、アカシアの樹皮を熱水やエタノール等の溶媒で抽出することにより高活性の成分を容易に得ることができるため、健康食品や、飼料、工業製品のみならず、医薬、化粧品等種々の分野で利用することができる。[0001]
[Field of the Invention]
The present invention obtains an active oxygen scavenger from a specific plant bark, and more specifically, a food or feed comprising as an active ingredient an extract obtained by extracting fine powder of bark of a specific plant or water / lower alcohol as a solvent. The present invention relates to an active oxygen scavenger used in cosmetics, quasi drugs, pharmaceuticals, antioxidants and the like.
[0002]
[Prior art]
In general, in the atmosphere, oxygen is a stable substance called triplet oxygen. When some of the oxygen changes into highly reactive substances called active oxygen such as superoxide, hydroxy radicals, singlet oxygen, and hydrogen peroxide in vivo, some of the oxygen is produced by neutrophils and macrophages. While acting as a body defense against foreign bodies, if excessive active oxygen exists in the body, it attacks lipids, proteins, enzymes, nucleic acids, etc., damages biological membranes and tissues, etc., arteriosclerosis, cerebrocardiovascular disorders It has been clarified that it can also cause the onset of emphysema, cancer, rheumatoid arthritis, cataract, hypertension, senile dementia, Alzheimer's disease, blemishes and aging.
[0003]
Therefore, in order to prevent the above-mentioned various diseases caused by oxidative stress caused by these excessive active oxygen, the starting material superoxide (0 2 − ) is disproportionated to produce hydrogen peroxide and oxygen molecules. The search for antioxidants that add SOD-like substances that act similar to superoxide dismutase (SOD), an enzyme that catalyzes the reaction that occurs, and substances that capture active oxygen (active oxygen scavenging active substances) to foods are widespread. Has been done.
[0004]
Examples of such substances include fat-soluble α-tocopherol (vitamin E) and water-soluble ascorbic acid (vitamin C) for substances derived from natural products, and BHT (butylhydroxytoluene) and BHA for synthetic substances. There are phenolic substances such as (butylhydroxyanisole) and TBHQ (tertiary butylhydroquinone).
However, although these substances are generally used as food antioxidants, they do not have a sufficient active oxygen scavenging action on the living body, and in particular, the synthetic substance BHA also causes problems such as suspected carcinogenicity. Is coming out.
[0005]
In recent years, research and development for searching active oxygen scavenging active substances having high safety from natural products such as animals and plants has been actively conducted. For example, there are Patent Documents 1, 2, 3, 4, 5, and the like. In addition, products that are actually commercialized include polyphenolic compounds such as tea leaves, catechins extracted from cacao bean extract, anthocyanin compounds contained in blueberry peels, isoflavones obtained from fermented soybeans, etc. Has attracted attention due to its very high active oxygen scavenging activity.
[0006]
Among these polyphenolic compounds that have been shown to have high active oxygen scavenging activity, (−) epigallocatechin, (−) epigallocatechin-3-gallate, (−) epicatechin, It has been found that catechins that are flavanols based on flavan-3-ol, such as-) epicatechin-3-gallate and (+) catechin, have higher active oxygen scavenging activity than other polyphenols. .
[0007]
On the other hand, tannin extracted with water from the bark of Acacia mearnsii De Wild., A kind of Acacia genus, has been produced in large quantities in South Africa and Brazil since ancient times. It is used very cheaply as a leather tanning agent and wood adhesive. In addition, Acacia mangium Willd. (Acacia mangium) grows very quickly, and in recent years, afforestation has become popular in the subtropics and the tropics including Indonesia and Malaysia. This walt tannin is classified as condensed tannin.
[0008]
However, the above scientific name: Acacia mearnsii De Wild. Not only tannins extracted from bark with hot water, but also bark fine powder obtained by crushing Acacia bark such as Acacia mangium Willd. The effect of erasing was not known at all. Furthermore, attempts have been made to use these for the prevention or treatment of the above-mentioned diseases, prevention or improvement of aging by adding or blending them into food, feed, cosmetics, quasi drugs, pharmaceuticals or antioxidants, etc. It wasn't.
[0009]
[Patent Document 1]
Japanese Patent Laid-Open No. 01-25726 [Patent Document 2]
JP-A-6-65074 [Patent Document 3]
JP 7-300422 A [Patent Document 4]
Japanese Patent Laid-Open No. 11-5975 [Patent Document 5]
Japanese Patent Laid-Open No. 2001-98264
[Problems to be solved by the invention]
The present invention focuses on the above-mentioned viewpoint, is highly safe for human bodies and animals, and has a function of sufficiently eliminating excess active oxygen generated in vivo, and a food and feed containing the same It is to provide cosmetics, quasi drugs, pharmaceuticals or antioxidants at low cost.
[0011]
[Means for Solving the Problems]
The present inventors have focused on the above-mentioned viewpoints, and have conducted extensive research using the active oxygen scavenging activity (SOSA) as an index. As a result, they have found that acacia bark has high active oxygen scavenging activity. The present invention has been completed.
The high active oxygen scavenging activity of Acacia bark cannot be specified by a certain component contained in the bark or by a combined effect based on various components, but can be obtained by Acacia bark.
The above-mentioned active oxygen scavenging activity is commonly shown in Acacia bark, but among Acacia trees, scientific name: Acacia mearnsii De Wild. (Generic name: black wattle), scientific name: Acacia mangium Willd. Mangum), scientific name: Acacia dealbata Link., Scientific name: Acacia decurrens Willd., And scientific name: Acacia pycnantha Benth.
[0012]
DETAILED DESCRIPTION OF THE INVENTION
The active oxygen scavenging action by Acacia bark can be obtained by using the bark itself or by an extract extracted from the bark with a solvent.
Among Acacia bark, scientific name: Acacia mearnsii De Wild. (Generic name: black wattle), scientific name: Acacia mangium Willd. (Generic name: Acacia mangium), scientific name: Acacia dealbata Link., Scientific name: Acacia decurrens Willd. It is important to find that the bark selected from the group consisting of Acacia pycnantha Benth. And extracts from these bark have a very high active oxygen scavenging action. Therefore, the present invention comprises a composition for active oxygen scavengers based on acacia barks including these scientific names of acacia bark and foods, feeds, cosmetics, quasi drugs, pharmaceuticals or antioxidants containing the same. is there.
[0013]
When acacia bark is used for the purpose of scavenging active oxygen, it may be used by pulverizing or pulverizing it to an appropriate size according to the purpose, and in particular finely pulverizing it.
The bark is usually dried as a tree, then dried, peeled off, and only the bark is collected, and then preferably dried further in the sun.
The bark of an Acacia tree (hereinafter simply referred to as acacia) consists of an outer skin and a slightly fibrous endothelium, and when dried to a moisture content of about 20% or less, it is easily pulverized with a pulverizer such as a hammer mill. The acacia bark contains an active oxygen scavenging component for both the inner skin and the outer skin, and the inner skin and the outer skin may be used separately.
When this bark fine powder is used as it is as an active oxygen scavenger, it is preferably classified to 100 μm or less, more preferably 50 to 70 μm or less containing 80 to 99% of an active ingredient (active oxygen scavenging component). It is good to use.
Fractionation of fine powder containing a large amount of active ingredients is obtained by pulverizing bark dried to a moisture content of 20% or less to an appropriate size, for example, a particle size of 1.6 mm or less, and classifying the resulting powder with a vibration sieve or the like. To obtain the required fine powder.
In addition to the above, when the acacia bark is used for feed such as dairy cows, the bark pulverized to an appropriate size can be used as it is.
[0014]
In order to obtain an active oxygen eliminating component as an extract from the acacia bark of the present invention, it is preferable to extract with a lower alcohol or a polar solvent. As the lower alcohol, methanol, ethanol, propanol, isopropyl alcohol, and the like are used. As the polar solvent, water, acetone, ethyl acetate, and the like are used. If necessary, one or two or more of them may be used in combination. It may be used with a mixed solvent of water / alcohol, and water or hot water extract may be further extracted with a solvent such as alcohol. Thus, a higher active component can be obtained by extracting in multiple stages with different solvents.
Accordingly, still another aspect of the present invention relates to an extract obtained by further extracting an extract from water or hot water from acacia bark with a solvent such as alcohol.
[0015]
The extraction is performed by adding a solvent to the acacia bark and stirring as necessary to extract, but the temperature, time, or solid-liquid ratio is not particularly limited. When water is used as the solvent, it may be extracted with hot water. The extract containing the active oxygen eliminating component obtained by extraction may be lyophilized or spray-dried as it is, or may be freeze-dried or spray-dried after concentration under reduced pressure. The obtained extract can be in various forms such as an extract, a solution, a powder, a concentrated liquid, a paste, and can be widely used as necessary. Further, the acacia bark extract of the present invention obtained in these forms can be used as it is as an active oxygen scavenging component, and further purified as necessary, and the purified product can also be used as an active oxygen scavenging component.
[0016]
Further, the active oxygen scavenging component of the present invention is used as it is as an active oxygen scavenger, and it is blended in foods, feeds, cosmetics, quasi drugs, pharmaceuticals or oxidant scavengers, etc., in any form depending on the purpose. Can be used.
The acacia bark of the present invention is pulverized to an appropriate size, preferably finely pulverized as described above, and used as it is in a feed or food composition. Further, since the active ingredient can be extracted from the pulverized Acacia bark with hot water, it can be supplied as a tea pack and enjoyed as a daily health drink.
[0017]
In addition to the acacia bark specifically shown above, the bark of Acacia storyi, scientific name: Acacia glaucocarpa, scientific name: Acacia neriifolia, scientific name: Acacia trachyphloia is listed as having high activity in the same way as the acacia bark specifically shown above. Moreover, the material of Eucalyptus genus: Eucalyptus diversicolor (generic name: Cali) and the bark and Kepraco material of the genus Kepraco can be shown as those exhibiting active oxygen scavenging activity in the same manner as Acacia bark.
[0018]
【Example】
Hereinafter, the present invention will be described in more detail with reference to production examples, test examples, and formulation examples, but the present invention is not limited thereto. In particular, the embodiment shows the acacia bark of the present invention without dividing it into an outer skin and an inner skin, but the outer skin can be used separately from the inner skin.
In the following production examples and test examples, each acacia of the present invention is indicated by a number shown in parentheses after each scientific name. For example, the acacia with the scientific name: Acacia mearnsii De Wild.
Scientific name: Acacia mearnsii De Wild. (No. 1), Scientific name: Acacia mangium Willd. (No. 2), Scientific name: Acacia dealbata Link. (No. 3), Scientific name: Acacia decurrens Willd. (No. 4), Scientific name : Acacia pycnantha Benth. (No. 5).
[0019]
Production Example 1
After drying the acacia No. 1 bark to a moisture content of 20% or less, the dried bark was pulverized to a powder of 1.6 mm or less (passed through a 10 mesh sieve (Tyler)) with a hammer mill (manufactured by Raymond). Furthermore, it classified with the vibration sieve machine and obtained the fine powder of 63 micrometers or less (under 250 mesh sieve).
Similarly, the remaining 4 types of Acacia No. 2-5 bark were pulverized to obtain fine powders of 63 μm or less, respectively. Although there were some differences in the yield of the fine powder that passed through the 250 mesh sieve depending on the type, the desired fine powder was obtained.
[0020]
Production Example 2
The bark of each Acacia No. 1 to 5 of the present invention is dried to a moisture content of 20% or less, and the dried bark is pulverized to a powder of 1.6 mm or less with a hammer mill. Double the amount of hot water was added, boiled, extracted for 15 minutes, and filtered using a 10-20 μm filter. The obtained filtrate was spray-dried with a spray dryer to obtain 30 g of bark extract.
Hereinafter, each bark extract is referred to as Acacia No. 1-5 hot water extract.
[0021]
Production Example 3
Each acacia bark of the present invention is dried to a moisture content of 20% or less, and the dried bark is pulverized to a powder of 1.6 mm or less with a hammer mill, and then 3 times the amount of methanol is added to 100 g of the dry crushed bark. In addition, the mixture was extracted with boiling for 15 minutes while being refluxed, and filtered using a 10 to 20 μm filter. After evaporating methanol from the obtained filtrate, the concentrated solution was spray-dried with a closed spray dryer to obtain 45 g of bark extract.
Hereinafter, each bark extract is described as an Acacia No. 1 methanol extract.
The methanol extract of Acacia No. 1-5 was obtained.
[0022]
Production Example 4
The acacia bark of the present invention was dried to a moisture content of 20% or less, and the dried bark was pulverized to a powder of 1.6 mm or less with a hammer mill, and then 3 times the amount of ethanol was added to 100 g of the dry crushed bark, Extraction was performed for 15 minutes while boiling and refluxing, followed by filtration using a 10-20 μm filter. After evaporating ethanol from the obtained filtrate, the concentrate was spray-dried with a closed spray dryer to obtain 45 g of a bark extract (hereinafter referred to as Acacia No. 1 ethanol extract).
Ethanol extracts of Acacia No. 1-5 were obtained.
[0023]
Production Example 5
Three times the amount of methanol was added to 10 g of the acacia hot water extract obtained in Production Example 2, and the mixture was boiled and refluxed for 15 minutes, and filtered using a 10 to 20 μm filter. Methanol was evaporated from the obtained filtrate, and water was added to the filtrate, followed by lyophilization to obtain 9 g of an extract (hereinafter referred to as an Acacia No. 1 hot water extract methanol fraction).
A hot water extract methanol fraction of Acacia No. 1-5 was obtained.
[0024]
Production Example 6
Three times the amount of ethanol was added to 10 g of the acacia hot water extract obtained in Production Example 2, and the mixture was boiled and refluxed for 15 minutes, and filtered using a 10 to 20 μm filter. Ethanol was evaporated from the obtained filtrate, and water was added thereto, followed by freeze-drying to obtain 9 g of an extract (hereinafter referred to as an Acacia No. 1 hot water extract ethanol fraction).
The hot water extract ethanol fraction of Acacia No. 1-5 was obtained.
[0025]
Test example 1 Active oxygen scavenging activity test The active oxygen scavenging activity of the acacia extract prepared in the above production example was determined based on the reaction formula shown in the following scheme, and the active oxygen scavenging activity was determined using a hypoxanthine-xanthine oxidase (XOD) system. One superoxide anion (O 2− ) is generated, and the generated O 2− is captured by a spin trap agent (DMPO: 5,5-Dimethyl-1-pyrroline-N-oxide), and ESR (Electron Spin Resonance) ) Method.
[Chemical 1]
[0026]
Preparation of sample Weigh 5 g from the powders obtained in the above Production Examples 1 to 6, add 50 mL of 0.1 M phosphate buffer, add a stir bar, and extract with stirring for 60 minutes while cooling with ice. A solution obtained by centrifugation (3,000 rpm, 5 minutes) or filtration (filter No. 1) is used as a sample solution.
[0027]
Measurement operation 9.2M DMPO (5,5-Dimethyl-1-pyrroline-N-oxide, Labotech, frozen storage product) 15μL, 5.5mM DETAPAC (Diethylenetriaminepentaacetic acid, Sigma, frozen storage product) ) 35 μL, 2 mM hypoxanthine (Sigma, frozen storage product) 50 μL and mixed. To this test tube, add SOD standard solution (SOD, Toyobo, 10 mg of 3,000 units / mg, dissolve in 0.1 M phosphate buffer to make 200 mL) or 50 μL of sample solution, and then add xanthine oxidase solution (xanthine oxidase). Boehringer Mannheim, 20 μL of 20 units / mL, added with 980 μL of 0.1 M phosphate buffer) When adding 50 μL, press the foot switch. Stir the test tube with a vortex or the like, suck it into a special flat cell (JEOL, approximately 130 μL), and set it in the ESR device. The sweep starts 40 seconds after the foot switch is pressed.
[0028]
The reading of the activity evaluation spectrum is a method in which the line width of the DMPO-OOH signal is constant, and the height of this signal is compared with the height of the signal of Mn 2+ used as the internal standard. To do.
[0029]
For the SOD standard solution, measure about 6 points at a concentration of 0 to 20 units / mL, and create a calibration curve. The ratio when the SOD is 0 (DMPO-OOH signal / Mn 2+ signal = I0) and the ratio (In) at each SOD concentration are determined. The vertical axis of the calibration curve is (I0 / In) -1, and the horizontal axis is the SOD concentration.
[0030]
In order to improve reproducibility, the concentration of the sample solution is measured after being diluted with 0.1 M phosphate buffer so as to be about 10 units / mL. The SOD concentration (SOSA concentration) in the sample solution is obtained from the calibration curve.
ESR uses JES-FR80 (JEOL), microwave output 8mW, magnetic field sweep width 335.4mT ± 5mT, sweep time 2 minutes, magnetic field modulation 100kHz, 0.79 × 0.1mT, amplification factor 2 × 100, response time 0.1 second Measure under conditions.
D is the dilution rate, W is the sample collection amount (g), and SOSA (unit / g) is determined by the activity of the sample solution (unit / mL) × D × (50 / W).
[0031]
The results are shown in Table 1 together with the measurement results of the comparative products.
[Table 1]
[0032]
Among the bark powders and extracts obtained in Production Examples 1 to 6 of the present invention from Table 1, the hot water extract ethanol fraction has the highest active oxygen scavenging action, and ascorbic acid (vitamin C) and (+) catechin It can be seen that the effect is about seven times that of
[0033]
Test Example 2 Acute toxicity test (oral administration)
In accordance with OECD (Guidelines for the Testing of Chemicals 401, 1987), an acute oral toxicity test using mice was performed.
For ICR male and female mice, each sample (Acacia mearnsii De Wild.) By hot water extraction with a tolerance of 500 mg / kg and 9 doses each up to 7,000 mg / kg for males and 6,500 mg / kg for females Were administered orally to male and female mice once. As the specimen, the product of the present invention was suspended in pure water. As a result, the LD 50 value was 4,468 mg / kg for males and 3,594 mg / kg for females, and no deaths were observed at a dose of 3,000 mg / kg.
The above test was performed on hot water extracts of Acacia Nos. 2 to 5, and similar results were obtained. Thereby, it turns out that the active oxygen scavenger of this invention is excellent in safety | security.
[0034]
Test Example 3 Mutagenicity test A mutagenicity test was carried out in accordance with Ministry of Labor Notification No. 77 (September 1, 1988).
Using the Escherichia coli WP2 uvrA strain and Salmonella typhimurium TA strain 4 strains, a reverse mutation test including metabolic activation was performed at 156 to 5,000 μg / plate dose of specimen Acacia No. 1 (scientific name: Acacia mearnsii De Wild.) Fever Performed with water extract. As a result, since no increase in the number of revertant colonies was observed for any of the strains, it was concluded that the mutagenicity of the product of the present invention was negative, indicating that it was excellent in safety.
[0035]
Formulation Example 1 Preparation of internal preparation An internal preparation was prepared using the ethanol fraction of the acacia bark hot water extract of Production Example 6 with the composition shown below.
[0036]
Formulation example 2 Preparation of pet food Using the acacia bark hot water extract of Production Example 2, a pet food was prepared with the composition shown below.
[0037]
Formulation example 3 Preparation of tablet (confectionery) Using the ethanol fraction of the acacia bark hot water extract of Production Example 6, a tablet (confectionery) was prepared with the composition shown below.
[0038]
Formulation example 4 preparation of skin lotion Using the acacia bark ethanol extract of production example 4 and stirring and dissolving uniformly with the composition shown below, a skin lotion was prepared.
[0039]
【The invention's effect】
The present invention was completed by finding that the acacia bark exhibits an active oxygen scavenging action, and a more active fraction can be obtained by finely pulverizing and classifying the bark. Therefore, it is safe and can be supplied in large quantities at a low cost, so it can be used for health foods and feeds. In addition, a highly active ingredient can be easily obtained by extracting the acacia bark with a solvent such as hot water or ethanol, so that it can be used not only for health food, feed, industrial products, but also for various fields such as pharmaceuticals and cosmetics. Can be used.
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CN101505775B (en) | 2006-08-10 | 2012-09-26 | 株式会社mimozax | Antioxidant composition containing component originating in the bark of tree belonging to the genus acacia |
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WO2008018139A1 (en) | 2006-08-10 | 2008-02-14 | Wood One Co., Ltd. | Antiobesity composition containing component originating in the bark of tree belonging to the genus acacia |
US8124138B2 (en) | 2006-08-10 | 2012-02-28 | Mimozax Co., Ltd. | Composition for prevention and/or treatment of pruritus containing acacia bark derivative |
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CN104352410A (en) * | 2014-11-14 | 2015-02-18 | 陈爱华 | Refrigerant after bath and preparation method thereof |
KR20180058998A (en) | 2016-11-25 | 2018-06-04 | 강동원 | Pharmaceutical composition and food composition for preventing, improving, or treating obesity, and a process for preparing them |
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JP3222672B2 (en) * | 1993-12-16 | 2001-10-29 | 三井農林株式会社 | Protein synthesis inhibitor |
JPH1025238A (en) * | 1996-07-10 | 1998-01-27 | Mikimoto Pharmaceut Co Ltd | Preparation for external use for skin |
JP2001048766A (en) * | 1999-08-02 | 2001-02-20 | Mikimoto Pharmaceut Co Ltd | Histamine liberation suppressing agent |
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