JP2018528971A - 高麗人参由来のエクソソーム様ベジクルを含む美白用組成物 - Google Patents
高麗人参由来のエクソソーム様ベジクルを含む美白用組成物 Download PDFInfo
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- JP2018528971A JP2018528971A JP2018515230A JP2018515230A JP2018528971A JP 2018528971 A JP2018528971 A JP 2018528971A JP 2018515230 A JP2018515230 A JP 2018515230A JP 2018515230 A JP2018515230 A JP 2018515230A JP 2018528971 A JP2018528971 A JP 2018528971A
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Abstract
Description
(段階1)先ず、4年根の高麗人参の根を準備し、これを搾汁して高麗人参の根の細胞外液を含む搾汁液を製造した。
前記実施例1で収得した高麗人参由来のエクソソーム様ベジクルに対し、透過電子顕微鏡(transmission electron microscope、TEM)でその大きさや形態を分析した。
ヒト黒色腫細胞株のMNT−1細胞を、ウシ胎児血清が20%入っている最小必須培地(Minimum Essential Medium、MEM)に入れ、37℃、5%のCO2条件下で育てた後、細胞数が各皿当たり1×105になるように60mm培養皿に敷いて、一晩中細胞が器壁にくっ付くことを待った後、前記実施例1で収得した高麗人参由来のエクソソーム様ベジクルを1μg/ml添加した同一の培地で交換した。このとき、比較のために、陰性対照群としては何も処理していないものを使用した。試料が入っている新しい培地は4日に1回ずつ交換し、細胞が皿に一杯になるまで総8日間培養した。細胞が完全に成長すると、溶解バッファー(1%のNP−40、50mMのTris−HCl;pH7.5、150mMのNaCl)を300μl処理してから細胞スクレーパーを使用してマイクロチューブに集めた後、15,000rpmで10分間遠心分離し、次いで、沈殿物を1NのNaOHに溶解させて450nmで吸光度を測定して、メラニン含有量を比較した。
ヒト初代色素形成細胞(human primary melanocyte、Life Technologies、CA、USA)をヒト色素形成細胞成長補充物(human melanocyte growth supplement(HMGS)、Gibco BRL、NY、USA)が添加されたM−254培地(Gibco BRL、NY、USA)に入れ、37℃、5%のCO2条件下で育てた後、細胞数が2×105になるように100mm培養皿に敷いて、一晩中細胞が器壁にくっ付くことを待った後、20mJ/cm2の強さのUVBを24時間置きに2回照射した。次いで、前記実施例1で収得した高麗人参由来のエクソソーム様ベジクルがそれぞれ0.1μg/ml、1μg/ml、5μg/ml、10μg/mlの濃度で含まれた培地で4日に1回ずつ該培地を交換しながら2週間培養した。このとき、比較のために陽性対照群として、30μMのmelasolv、1μMのcompound K、2ppmの発酵サポニンを同じ方法で2週間処理してメラニン生成程度を測定した。陰性対照群としては、UVBを処理していない色素形成細胞とUVBを処理するが物質を処理していない色素形成細胞を同じ方法で培養した。2週間後、細胞を培養皿から引き剥がしてその数を数え、次いで、同数の細胞をマイクロチューブに集めた後、RIPA溶解バッファー(Millipore、20−188)を入れて細胞を壊し、13,500rpmで30分間遠心分離した。沈殿物を1NのNaOHに溶解させ、撮像してその色を比較し、450nmで吸光度を測定してメラニン含有量を比較した。
ヒト初代色素形成細胞(human primary melanocyte、Life Technologies、CA、USA)をヒト色素形成細胞成長補充物(human melanocyte growth supplement(HMGS)、Gibco BRL、NY、USA)が添加されたM−254培地(Gibco BRL、NY、USA)に入れ、37℃、5%のCO2条件下で育てた後、細胞数が0.7×105になるように24ウェルプレートに敷いて、一晩中細胞が器壁にくっ付くことを待った後、前記実施例1で収得した高麗人参由来のエクソソーム様ベジクル(GEV)と高麗人参の搾汁液をそれぞれ1μg/ml、10μg/mlの濃度で含む培地と含まない培地(対照群)で交換し、48時間培養した。48時間が経過した後の細胞の生存率をwst−1(Roche Applied Science、Germany)溶液を用いて測定した。
ヒト初代色素形成細胞(human primary melanocyte、Life Technologies、CA、USA)をヒト色素形成細胞成長補充物(human melanocyte growth supplement(HMGS)、Gibco BRL、NY、USA)が添加されたM−254培地(Gibco BRL、NY、USA)に入れ、37℃、5%のCO2条件下で育てた後、細胞数が2×105になるように100mm培養皿に敷いて、一晩中細胞が器壁にくっ付くことを待った後、20mJ/cm2の強さのUVBを24時間置きに2回照射した。次いで、前記実施例1で収得した高麗人参由来のエクソソーム様ベジクル(GEV)と高麗人参の搾汁液をそれぞれ1μg/ml、10μg/mlの濃度で含まれた培地で4日に1回ずつ該培地を交換しながら2週間培養した。対照群としてUVBを処理していない色素形成細胞とUVBを処理するが物質を処理していない色素形成細胞を同じ方法で培養した。2週後、細胞を培養皿から引き剥がしてその数を数え、次いで、同数の細胞をマイクロチューブに集めた後、RIPA溶解バッファー(Millipore、20−188)を入れて細胞を壊し、13,500rpmで30分間遠心分離した。次いで、沈殿物を1NのNaOHに溶解させて撮像してその色を比較し、450nmで吸光度を測定してメラニン含有量を比較した。
高麗人参由来のエクソソーム様ベジクル50mg、L−カルニチン80〜140mg、大豆油180mg、パーム油2mg、植物性硬化油8mg、黄蝋4mg、及びレシチン6mgを混合し、通常の方法に従って、1カプセル当たり400mgずつ充填して軟質カプセル剤を製造した。
高麗人参由来のエクソソーム様ベジクル50mg、ガラクトオリゴ糖200mg、乳糖60mg、及び麦芽糖140mgを混合し、流動層乾燥機を利用して顆粒化した後、糖エステル(sugar ester)6mgを添加して、打錠機で打錠して錠剤を製造した。
高麗人参由来のエクソソーム様ベジクル50mg、無水結晶ブドウ糖250mg、及び澱粉550mgを混合し、流動層造粒機を使用して顆粒に成形した後、砲に充填して顆粒剤を製造した。
高麗人参由来のエクソソーム様ベジクル50mg、ブドウ糖10g、クエン酸0.6g、及び液状オリゴ糖25gを混合した後、精製水300mlを加えて各瓶に200mlずつ充填した。瓶に充填した後、130℃で4〜5秒間殺菌して、ドリンク剤飲料を製造した。
下記の表3に記載された組成にて通常の方法によりローションを製造した。
下記の表4に記載された組成にて通常の方法によりクリームを製造した。
下記の表5に記載された組成にて通常の方法によりパックを製造した。
Claims (12)
- 高麗人参由来のエクソソーム様ベジクル(exosome−like vesicle)を有効成分として含む、美白用組成物。
- 前記エクソソーム様ベジクルは、高麗人参の根から分離したものである、請求項1に記載の美白用組成物。
- 前記エクソソーム様ベジクルは、20〜500nmの直径を有するものである、請求項1に記載の美白用組成物。
- 前記エクソソーム様ベジクルは、高麗人参の細胞外液の100,000×g以上の超遠心分離で沈降するものである、請求項1に記載の美白用組成物。
- 前記エクソソーム様ベジクルは、イオジキサノール(iodixanol)での浮遊密度が1.00〜1.20g/mlのものである、請求項1に記載の美白用組成物。
- 前記有効成分はメラニン生成を抑制する、請求項1に記載の美白用組成物。
- 前記有効成分は、しみ、そばかす、黒色斑点、母斑、黒色腫、薬物による色素沈着、炎症後色素沈着、及び皮膚炎で発生する色素沈着からなる群から選択される1以上の皮膚色素沈着疾患を予防、改善又は治療する、請求項1に記載の美白用組成物。
- 請求項1〜7のいずれか一項に記載の高麗人参由来のエクソソーム様ベジクル(exosome−like vesicle)を製造する方法であって、
(1)高麗人参を搾汁して搾汁液を収得する段階;
(2)前記搾汁液を遠心分離して残滓を除去し上層液を収得する段階;及び
(3)前記上層液を超遠心分離してエクソソーム様ベジクル(exosome−like vesicle)を収得する段階を含む、高麗人参由来のエクソソーム様ベジクルの製造方法。 - 前記段階(2)における遠心分離は、500〜3,000×gで10〜30分間実施する、請求項8に記載の高麗人参由来のエクソソーム様ベジクルの製造方法。
- 前記段階(3)における超遠心分離は、スクロースクッション密度勾配及びイオジキサノール(iodixanol)密度勾配を用いて超遠心分離するものである、請求項8に記載の高麗人参由来のエクソソーム様ベジクルの製造方法。
- 前記段階(3)における超遠心分離は、100,000〜200,000×gで1〜6時間実施する、請求項8に記載の高麗人参由来のエクソソーム様ベジクルの製造方法。
- 前記段階(3)においてエクソソーム様ベジクルは、上層液を超遠心分離した後、イオジキサノール(iodixanol)で1.00〜1.20g/mlの浮遊密度を有するフラクションを分離する段階を含んで収得する、請求項8に記載の高麗人参由来のエクソソーム様ベジクルの製造方法。
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KR20120037179A (ko) * | 2010-10-11 | 2012-04-19 | 대구한의대학교산학협력단 | 나노리포좀으로 안정화시킨 감 추출물과 인삼 추출물을 함유하는 항아토피 항노화 화장료 조성물 |
CN103271891A (zh) * | 2013-04-28 | 2013-09-04 | 福建南方制药股份有限公司 | 人参皂苷纳米胶束及其制备方法、应用和药物组合物 |
CN103462846A (zh) * | 2013-09-17 | 2013-12-25 | 吉林大学 | 人参美白抗衰纳米乳精华液及其制备方法 |
CN103519178A (zh) * | 2013-10-25 | 2014-01-22 | 天津商业大学 | 人参纳米微胶囊的制备方法 |
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JP2022539228A (ja) * | 2019-07-02 | 2022-09-07 | インダストリー-ユニバーシティー コーペレイション ファンデーション ハンヤン ユニバーシティー エリカ キャンパス | 植物エクソソームの大量生産方法 |
JP7274712B2 (ja) | 2019-07-02 | 2023-05-17 | インダストリー-ユニバーシティー コーペレイション ファンデーション ハンヤン ユニバーシティー エリカ キャンパス | 植物エクソソームの大量生産方法 |
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EP3354272A4 (en) | 2019-05-22 |
EP3354272A1 (en) | 2018-08-01 |
ES2888802T3 (es) | 2022-01-07 |
US20180263871A1 (en) | 2018-09-20 |
TWI699213B (zh) | 2020-07-21 |
HK1249414A1 (zh) | 2018-11-02 |
KR102598793B1 (ko) | 2023-11-07 |
CN108135831B (zh) | 2021-01-05 |
EP3354272B1 (en) | 2021-06-30 |
US10610467B2 (en) | 2020-04-07 |
CN108135831A (zh) | 2018-06-08 |
KR20170035771A (ko) | 2017-03-31 |
JP6890119B2 (ja) | 2021-06-18 |
TW201717891A (zh) | 2017-06-01 |
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