Studies on Oryzalide A, a novel antibacterial diterpene, isolated from healthy leaves of rice plant

1. 10种Oryzalides及其相关化合物的分离与鉴定从健康水稻叶片中分离得到一种新的抗菌二萜类物质Oryzalide A及9种结构相关物质，并对其结构进行了鉴定。从鲜重约20 kg的大白菜剑叶中分离出乙酸乙酯提取的酸性和中性粗提物。Norin 27，经硅胶柱层析、SephadexLH-20柱层析、高效液相色谱、分配薄层层析等分离纯化，通过核磁共振氢谱、核磁共振碳<13>谱、高分辨质谱、红外光谱等方法确定了它们的结构，结果新发现了4种酸性物质Oryzalides A、B和Oryzalic acids A、B，以及6种中性物质化合物A、B、C、D、E和F，并对上述10种化合物的结构特征、产率和抑菌活性进行了研究.稻瘟内酯和稻瘟酸微量分析方法的建立及内源水平的测定 ...更多信息 在稻叶中。转化为甲酯衍生物后，通过配备毛细管柱的GC-MS-SIM分析稻苷（A和B）和稻酸（A和B）。加入氘代标记的[17-D_2]Oryzalide B作为内标物测定了内源性Oryzalide和Oryzalic酸的量。在m/z 305碎片离子（4种物质中的特征离子和共有离子）处检测，水稻鲜叶中的主要成分为Oryzalide A和Oryzalic acid A，初步测定结果表明，在8个抗白叶枯病不同的水稻品种的健康叶片中均检测到Oryzalide A和Oryzalic acid A，但在量和抗性程度之间没有观察到相关性。4种内源物质在水稻叶片中几乎都能检测到，但在根和穗中检测不到，且含量随植株生长而增加。针刺伤叶后，其含量比未伤健叶（对照）增加3-4倍。在接种亲和或不亲和菌株的叶片中，其数量分别增加到对照的7.2倍和9.1倍。少

1. Isolation and identification of ten kinds of Oryzalides and related compounds.A novel antibacterial diterpene substances, Oryzalide A and structurally related 9 substances, were isolated from healthy rice leaves and the structures were determined. An ethyl acetate-extractable acidic and neutral crude extracts were isolated from about 20 kg fresh weight of flag leaves of cv. Norin 27, and they were purified by silica gel and sephadex LH-20 column chromatography, HPLC, partition TLC, etc., and finally their structures were determined by ^1H-NMR, ^<13>C-NMR, HR-MS, IR, etc. As the results, four kinds of acidic substances of Oryzalides A, B, and Oryzalic acids A, B, and six kinds of neutral substances of Compounds A, B, C, D, E and F were newly found. The structural characteristics, yields and antibacterial activities of the above mentioned ten substances were clarified.2. Establishment of a microanalytical method of Oryzalides and Oryzalic acids, and measurement of an endogenous level … More in rice leaves. Oryzalides(A and B)and Oryzalic acids(A and B)were analyzed by means of GC-MS-SIM equipped with a capillary column after conversions to the Methyl ester derivative. Amounts of endogenous Oiyzalides and Oryzalic acids were determined by addition of deuterated labeled[17-D_2]Oryzalide B as an internal standard. Major components were found to be Oryzalide A and Oryzalic acid A in fresh leaves of rice plant when monitoring at m/z 305 fragment ion, which was a characteristic and common ion among the four substances.As the results of preliminary measurement using the above microanalytical method, all Oryzalides and Oryzalic acids were detected from healthy leaves of 8 rice cultivars different in their resistance to bacterial leaf blight, but no correlation was observed between the amount and the degree of resistance. An endogenous four substances were almost detected from leaves but not from roots and ears, and their amount increased as a growth of rice plant proceeds. When the leaves were wounded by a needle bundle-prick, the amount increased to 3-4 times compared with that of not wounding healthy leaves(control). In the leaves inoculated with a compatible or an incompatible strain of pathogenic bacterium, the amount increased to 7.2 or 9.1 times of control, respectively. Less