Chemical Constituents And Bioactivities Of The Fruit Of Schisandra Glaucescens&Apoptotic Mechanism Of HepG2Cells Induced By2-methoxyjuglone

This dissertation is composed of three parts. Chapter1presents the chemical constituents and bioactivities of the fruit of Schisandra glaucescens Diels. Chapter2discusses the mechanism of2-methoxyjuglone-induced apoptosis in human hepatocellular carcinoma HepG2cell line. The third part is a review about the outstanding papers awarded by Arthur E. Schwarting and Jack L. Beal Awards in the Journal of Natural Products from2001to2012.1. The chemical constituents and bioactivities of the fruit of Schisandra glaucescens DielsSchisandra glaucescens Diels is a vine plant mainly distributed in the western Hubei and southeastern Sichuan Provinces in China. In the previous work from our laboratory,41compounds including seven new ones were isolated from the stems of S. glaucescens, some of which processing significant neuroprotective effects on SHSY5Y cells compared with H2O2-or CoCl2-treated group. However, to the best of our knowledge, there is no research work on the fruit of S. glaucescens. As a continuation, phytochemical studies along with bioactive assays were carried out in this work.Investigation on the chemical constituents of the fruit of S. glaucescens finally led to the isolation of36compounds (including ten new ones), and their structures were respectively confirmed as (7’R,8’S)-3,4-dimethoxy-3’,4’-methylenedioxy-7,8-seco-7,7’-epoxylignan-7,8-dione (1),(7’R,8’S)-3,4-methylenedioxy-3’,4’-dimethoxy-7,8-seco-7,7’-epoxylignan-7,8-dione (2),(8R,8’R)-9-O-(6’-O-α-L-arabinofuranosyl)-β-D-glucopyranosyl dihydrocubebin (3),(8R,8’R)-9-O-β-D-glucopyranosyl piperphilippinin VI (4a),(8R,8’R)-9’-O-β-D-glucopyranosyl piperphilippinin Ⅵ (4b), tetrecentronside B (5),1-(3,4-dimethoxyphenyl)-4-(3,4-methylenedioxyphenyl)-2,3-dimethylbutane (6), meso-dyhydroguaiavetic acid (7),(±)-anwulignan (8), (-)-galcatin (9),(-)-isootobaphenol (10),(±)-4’,5-O-didemethylcyclogalgravinby (11),(+)-chicanine (12),(+)-zuihonin C (13),(+)-zuihonin A (14), schiglausin P-T (15-19), schisanlactone A (20), schisanlactone B (21), schisanlactone C (22), kadsuphilactone B (23), propinic lactone (24), schisanlactone H (25), heteroclitalactone F (26), schisanlactone G (27), schisanlactone D (28), kadsulactone (29), schisanlactone E (30), micranoic acid B (31),(+)-y-cuparenol (32), chamigrenol (33), schisansphenin A (34),2-monomethyl citrate (35) and n-octacosanol (36), by extensive spectroscopic methods, electronic circular dichroism (ECD) calculations, hydrolysis reactions and comparing with the corresponding literature data. Among them, compounds1-3,4a and4b were new lignans, while compounds15-19were new triterpenoids.The antioxidant activities of all the isolated compounds were tested using2,2-diphenyl-l-picrylhydrazyl and ferric reducing antioxidant power assays. Compounds4,7,8,10,11, and12exhibited antioxidant activities of different potential in both assays. Of these compounds,7showed the strongest DPPH radical scavenging capacity with IC50values of15.7(150μM DPPH) and34.6μM (300μM DPPH) respectively, while4,12, and7displayed higher total antioxidant activities than Trolox in FRAP assay. The neuroprotective effects of these compounds against Aβ25-35-induced cell death in SH-SY5Y cells were also investigated. The results showed that compounds1,2,6,7,8,11, and12exhibited statistically significant neuroprotective effects against Aβ25-35-induced SH-SY5Y cell death compared with the Aβ25-35-treated group. The cytotoxicity of all the isolated triterpenoids (15-31) against B16mouse melanoma cells were tested, and compounds22,25,28and29exhibited strong cytotoxicity in this assay, especially, the IC50values of compounds22and28were less than10μM.2. The apoptotic mechanism of HepG2cells induced by2-methoxyjugloneJuglans cathayensis Dode (Juglandaceae), also called Chinese walnut, is a tree distributed widely in mainland China and Taiwan.2-Methoxyjuglone is one kind of naphthoquinones isolated from the root bark of J. cathayensis Dode in our previous work, and this compound was found to exhibit cytotoxicity against HepG2human hepatocellular carcinoma cells in vitro and anti-tumor effect against H22mouse hepatocellular carcinoma cells in vivo. As a continuation, investigation on the apoptotic mechanism of HepG2cells induced by2-methoxyjuglone was carried out.In this work, cell cycle analysis with propidium iodide staining showed that2-methoxyjuglone induced cell cycle arrest at the S phase in HepG2cells. Flow cytometric analysis with annexin V and propidium iodide staining demonstrated that2-methoxyjuglone induced HepG2cell apoptotic events in a dose-dependent manner. Western blot analysis of apoptosis-related proteins revealed that2-methoxyjuglone induces HepG2cell apoptosis through mitochondrial cytochrome c dependent activation of the caspase-9and caspase-3cascade pathway (mitochondrial pathway).