Key words Inula japonica; topoisomerase I; topoisomerase IIInula japonica THUNB. (Asteraceae) is a traditional herbal medicine and is widely distributed in Korea, Japan and China, and the flowers of this plant have been used to relieve phlegm, peptic disorder, inflammation and detumescence.1) Some terpenes 2-7) and flavonoids 8,9) have previously been isolated from this medicinal plant and have been reported to possess diverse biological activities, such as anti-diabetes, 10) antihypolipidemia, 11) hepatoprotective, 12) anti-inflammatory 7) and anti-tumor 1,2,13) effects. We have been isolating compounds that inhibited topoisomerases I and II from the source of medicinal plants. [14][15][16][17][18] DNA topoisomerases are enzymes that control DNA topology in the cell and are targets of anticancer drugs. 19,20) Topoisomerase I cleaves and reseals one DNA strand at a time and does not require ATP for the complementary strand to pass through the enzyme-linked strand break, thereby effecting DNA relaxation. Topoisomerase II cleaves both strands of DNA during catalysis. In a reaction coupled to ATP binding and hydrolysis, these proteins cleave one DNA duplex, transport a second duplex through the break, and then religate the cleaved duplex. 21) Camptothecin (CPT) and etoposide (VP-16) are typical inhibitors of topoisomerases I and II, respectively. 19) In this paper, we report isolation of fourteen compounds from the flowers of I. japonica and their DNA topoisomerases I and II inhibitory activities and cytotoxicities. Results and DiscussionTwo new (1, 2) and twelve known compounds (3-14) were isolated by various chromatographic separations from the EtOAc extract of the flowers of I. japonica (Fig. 1) C-NMR and distortionless enhancement by polarization transfer (DEPT) spectra of 1 showed seventeen peaks due to three methyl, four methylene, five methine and five quaternary carbons. The corresponding proton signals were determined by 1 H-detected heteronuclear multiple quantum coherence (HMQC) spectrum.The

Two rapid and simple HPLC methods with UV detector to determine three main compounds (magnoflorine, spinosin and 6′′′-feruloyl spinosin) and evaporative light scattering detector (ELSD) to determine jujuboside A were developed for the chemical analyses of Zizyphi Semen. Magnoflorine, spinosin, and 6′′′-feruloyl spinosin were separated with an YMC J’sphere ODS-H80 column (250 mm × 4.6 mm, 4 μm) by the gradient elution followed by the isocratic elution using methanol with 0.1 % formic acid and water with 0.1 % formic acid as the mobile phase. The flow rate was 1.0 mL/min. Jujuboside A was separated by HPLC–ELSD with YoungJinBioChrom Aegispak C18-L column (250 mm × 4.6 mm, 5 μm) column in a gradient elution using methanol with 0.1 % formic acid (A) and water with 0.1 % formic acid as the mobile phase. These two methods were fully validated with respect to linearity, precision, accuracy, stability, and robustness. These HPLC methods were applied successfully to quantify four compounds in a Zizyphi Semen extract. The HPLC analytical methods were validated for pattern recognition analysis by repeated analysis of 91 seed samples corresponding to 48 Zizyphus jujuba var. spinosa (J01–J48) and 43 Zizyphus mauritiana (M01–M43). The results indicate that these methods are suitable for a quality evaluation of Zizyphi Semen.