우슬뿌리 추출물의 Cathepsin B에 대한 저해효과
Inhibition Effect of Achyranthes japonica N. Root Extract on Cathepsin B
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초록
우슬이 민간요법으로 관절염 치료에 우수한 한약재로 알려진 바 우슬 추출물이 cathepsin B 에 대한 저해력을 검토하기 위하여 각종 용매로 우슬 추출물을 순차 분획하고 분획 추출물에 대하여 cathepsin B 에 대한 저해활성을 검토하였으며 우슬의 지표물질인 20-hydroxy ecdysone이 추출분획에 검출되는지 TLC 및 HPLC를 이용하여 분석하였으며 우슬의 지표물질이 cathepsin B 에 대한 저해활성여부를 확인한바 다음과 같은 결과를 얻었다. 우슬 뿌리에 대해 methanol/water(4:1, v/v) 추출물을 ethyl acetate, chloroform, chloroform/methanol(3:1, v/v), methanol의 각 용매로 분획한 결과, 분획 F4(methanol 분획)에서 수율이 가장 높아 8.27%를 나타내었다. 각 분획별에 따라 cathepsin B에 대한 저해활성을 측정한 결과 F4분획물에서 가장 저해활성이 높았으며 F1분획에서도 높은 저해활성을 나타내었다. F4분획물 중 우슬의 지표물질인 20-hydroxy ecdysone이 검출되었으며 우슬에 함유되어있는 함량은 0.33%이었다. 20-hydroxy ecdysone에 대하여 cathepsin B저해활성을 보기 위하여 cathepsin B저해활성제로 알려진 leupeptin과 활성을 비교해본 결과 기질을 BANA로 사용하였을 경우 leupeptin은 저해율이 92%인데 비하여 20- hydroxy ecdysone은 88%이었으며 F4분획물은 97%를 나타내었고 기질을 CLN으로 사용하였을 경우 leupeptin은 저해율이 62%인데 비하여 20-hydroxy ecdysone은 36%이었으며 F4추출물은 67%를 나타내었다.
Abstract
This study was carried out to investigate the cathepsin B inhibition effect by Achyranthes japonica N. root extract in vitro. The methanol/H2O(4:1,v/v) extract was fractionated by ethyl acetate(F1), chloroform(F2), chloroform/methanol(3:1, v/v)(F3) and methanol(F4). The yield of F4 in Achyranthes japonica N. root was 8.27%. As an index material of Achyranthes japonica N. root, 20-hydroxy ecdysone was detected by TLC, and HPLC and it's content was 0.33%. Three isolates(F1, F3, F4) showed the cathepsin B inhibition activity, and F4 showed the highest inhibition activity among them. In the inhibition activity on cathepsin B, leupeptin, 20-hydroxy ecdysone and F4(at the same concentration of 20-hydroxy ecdysone.) were 92, 88 and 97% on BANA(Nα-benzoyl-DL-arginine β-naphthylamide) substrate, and 62, 36 and 67% on CLN(Nα-CBZ(carbobenzlyoxy)-L-lysine p-nitrophenyl ester HCl) substrate, respectively.
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