Phytolacca americana L. (Phytolaccaceae) is a perennial plant originating from North America. Its rhizome has been used as a traditional crude diuretic drug in spite of having strong toxicity. A number of triterpenes, their glycosides, and neolignanes have been isolated from the roots of P. americana and identified.1-7) As a part of our search for neurotrophic substances in natural products, we reported the isolation and structure of americanol A (8) and isoamericanol A (9) from the seeds of P. americana.8,9) Moreover, we accomplished the convenient syntheses of americanol A (8) and isoamericanol A (9) by horseradish peroxidase (HRP) catalyzed oxidative coupling of caffeic acid. 10) In this paper, we describe our next attempt aiming at production of neurotrophic substances 8 and 9 from the cultures of P. americana. First of all, we tried to establish the cell cultures of P. americana independently of the previously known procedure. 11)A cell culture of P. americana was cultured in the Murashige & Skoog (MS) liquid medium 12) (pH 5.6) containing 10 mM 1-naphthaleneacetic acid (1-NAA) and 3% of glucose, on a rotary shaker (90 rpm) at 25°C. The cell cultures were subcultured at intervals of 4 weeks. The filtered cells were freeze-dried and extracted with methanol. The methanol extract was subjected to silica gel, Sephadex LH-20 and HPLC (octadecyl silica gel (ODS) column) to afford a new triterpene glycoside 1 and the five known triterpene glycosides, esculentoside B (2), 4,13,14) phytolaccoside E (3), 3,4,13) esculentoside S (4), 13,15) esculentoside L1 (5), 13,15) and esculentoside G (6). 3,13,14) The spectral data of 2-6 were in full agreement with those described in the literature.Compound 1 for the anomeric protons of xylose and glucose in 1 indicated that the anomeric configurations of D-xylose and D-glucose are b. These results indicated that 1 was the triterpene glycoside linked with D-xylose and D-glucose. In the 13 C-NMR spectrum, the aglycone part of 1 was similar to phytolaccinic acid (7) 13) except for C-3 which was shifted down-field by 8.1 ppm in comparison with that of 7, thus indicating that the sugar part was attached at the C-3 position. In order to confirm the linkages of the two sugar parts and the structure of the aglycone, heteronuclear multi-bond correlation (HMBC) experiments (Fig. 1) A new triterpene glycoside 1 was isolated together with the five known triterpene glycosides 2-6 from the cultures of Phytolacca americana. The structure of 1 was elucidated by analysis of spectroscopic data and comparison of its NMR data with those of 2-7 and chemical degradation.

We have visualized redox and structural changes in the mitochondria of yeast Saccharomyces cerevisiae as a eukaryotic cell model using a genetically encoded yellow fluorescent protein (Y1-Yellow) and conventional fluorescence microscopy. Y1-Yellow originating from a yellow emitting luminous bacterium Aliivibrio sifiae Y1 was fused with a mitochondria-targeted sequence (mt-sequence). Y1-Yellow fluorescence arising only from the mitochondrial site and the color of yellow fluorescence could be easily differentiated from cellular autofluorescence and from that of conventional probes. Y1-Yellow expressing S. cerevisiae made the yellow fluorescence conspicuous at the mitochondrial site in response to reactive oxygen species (ROS) transiently derived in the wake of pretreatment with hydrogen peroxide. Based on our observation with Y1-Yellow fluorescence, we also showed that mitochondria rearrange to form a cluster structure surrounding chromosomal DNA via respiratory inhibition by cyanide, followed by the generation of ROS. In contrast, uptake of an uncoupler of oxidative phosphorylation is not responsible for mitochondrial rearrangement. These results indicate the utility of Y1-Yellow for visualization of mitochondrial vitality and morphology in living cells.

The relationship between heat treatment and mechanical properties of Al-Si-Mg alloy was investigated. Solution treatment at 808K for 6 hours, preliminary aging at room temperature for 60 hours and artificial aging at 473K for 1 hours were conducted. Not preliminary aged samples were prepared for comparison. The strength improved and the elongation decreased with increasing the amount of Mg. This trend was seen both preliminary aged and not preliminary aged samples. However, preliminary aged samples showed relatively high tensile strength and elongation comparing with not preliminary aged samples. EPMA analysis revealed that the precipitation of Mg2Si in preliminary aged samples was larger than that of not preliminary aged samples, and which possibly contributed to the improvement of strength of the preliminary aged samples. Thermodynamic calculation showed that Mg2Si phase was not undissolved at the solution temperature, 808K, and these undissolved Mg2Si particles might influenced on mechanical properties negatively.