익지(益智)의 간 세포 보호 성분

Abstract

Alpinia oxyphylla Miq. (Zingiberaceae) is traditional oriental herbal medicine widely used as tonic, aphrodisiac, anti-salivation, anti-polyuria and anti-diarrhea. Crude methanolic extract of A. oxyphylla increased antioxidant response element (ARE) luciferase activity 6 fold at 30 g/ml. Thus, methanolic extract was fractionated successively to obtain n-hexane, EtOAc, n-BuOH and water fractions. Among these fractions, n-hexane and EtOAc fractions induced ARE activity. n-Hexane fraction was divided into 17 sub-fractions by centrifugal partition chromatography (CPC) with two different biphasic solvent systems, consisting of n-hexane-EtOAc-methanol-water (7:3:7:3, v/v). In active four sub-fractions (C, D, H and I), a new diaryheptanoid, one diarylheptanoid and four sesquiterpenoids were isolated by repeated open column chromatography and preparative HPLC. In EtOAc fraction, six sesquiterpenoids were purified by Diaion HP-20 and silica gel open column chromatography and preparative HPLC. Chemical structures of isolated compounds were identified as nootkatone (1), eudesma-3,11-dien-2-one (2), yakuchinone A (3), 5'-hydroxyl-yakuchinone A (4), alinenone (5), 6-hydroxy-cyperone (6), (4S, 5E, 10R)-7-oxo-tri-nor-eudesm-5-en-4-ol (7), teuhetenone A (8), 7-epi-teucrenone B (9), 11-hydroxyvalenc-1(10)-en-2-one (10), oxyphyllenodiol A (11), oxyphyllendiol B (12) on the basis of physicochemical evidence and spectroscopic analysis such as 1D, 2D-NMR and ESI-MS. Among isolated compounds, eudesma-3, 11-dien-2-one (2) enhanced ARE luciferase activity in HepG2 cells transiently transfected with the ARE-luciferase, and yakuchinone A (3) and 5'-hydroxyl-yakuchinone A (4) exerted radical scavenging activities on ABTS and DPPH assays. Crude extract, eudesma-3, 11-dien-2-one (2), yakuchinone A (3) and 5'-hydroxyl-yakuchinone A (4) showed protective effects via inhibiting reactive oxygen species generation against oxidative cellular injury by t-BHP. Moreover, crude extract and eudesma-3, 11-dien-2-one (2) promoted the nuclear accumulation of the nuclear factor, E2-related factor 2 (Nrf2), and increased the promoter property of the antioxidant response element (ARE). In conclusion, A. oxyphylla extract showed protective effect on oxidative stress induced by t-BHP and enhancement of cellular Nrf2, HO-1 proteins in HepG2 cell. Among purified compounds, eudesma-3, 11-dien-2-one (2) was inducing ARE-Nrf2 pathway and yakuchinone A (3) and 5'-hydroxyl-yakuchinone A (4) showed radical scavenging effects on DPPH, ABTS assay. Taken together, this investigation provides evidence supporting that A. oxyphylla is hepatoprotective against oxidative stress that causes abnormal liver damage. All these considered, A. oxyphylla is worthy of further investigation for its potential as a hepatoprotective agent.