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Frontier in Structure Elucidation of Natural Products

天然产物结构解析前沿

基本信息

批准号：
12045243
负责人：
MURATA Michio
金额：
$ 10.75万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research on Priority Areas
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2002
项目状态：
已结题

项目摘要

Chemotaxis of sperm toward eggs during fertilization is a crucial event for species conservation, particularly for animals living in aquatic environments. Relevant chemical attractants have been found from a few marine organisms such as sea urchins and corals. We previously showed that sperm of the ascidian Ciona intestinalis is activated and then attracted toward the egg by a factor released from the egg, which led to the purification of sperm-activating and attracting factor (SAAF) from the egg-conditioning medium of Ciona intestinalis.The structure of SAAF was proposed to be a novel polyhydroxysterol sulfate on the basis of 2D-^1H NMR and FAB-MS/MS analysis using no more than approximately 4 microgram of a specimen. However, there still remained small but serious ambiguity that the biological activity might be attributed to a minor constituent. Synthesis is, therefore, essential for the unequivocal identification of SAAF as the active principle, and for the complete structure elucidation including the stereochemistry at C25. SAAF and its C-25 epimer were synthesized from chenodeoxycholic acid in 19 steps via the pivotal intermediate and both the sperm-activating and attracting activity of them were bioassayed. Synthetic SAAF activated the sperms of ascidian Ciona intestinalis at 3.7 nM and concurrently exhibited the attracting activity at<10 nM, which led to the unambiguous structure determination of SAAF and the verification of its dual activity. It is noteworthy that 25-epi-SAAF turned out to possess the comparative activities as those of SAAF.

精子在受精过程中对卵子的趋化作用是物种保护的重要环节，特别是对于生活在水生环境中的动物。从海胆和珊瑚等少数海洋生物中发现了相关的化学引诱剂。我们之前已经证明，海鞘玻璃海鞘的精子被激活，然后被卵子释放的因子吸引到卵子上，从玻璃海鞘卵培养液中分离纯化了精子激活和吸引因子（SAAF），经2D-^1HNMR和FAB-MS确证其结构为一种新的多羟基甾醇硫酸酯。使用不超过约4微克的样品进行MS分析。然而，关于生物活性是否可归因于一种微量成分，仍然存在着微小但严重的模糊性。因此，合成对于明确鉴定SAAF作为活性成分以及完整的结构解析（包括C25处的立体化学）至关重要。以鹅去氧胆酸为起始原料，经19步反应合成了SAAF及其C-25差向异构体，并对它们的精子激活活性和吸引活性进行了生物测定。合成的SAAF在3.7 nM时激活海鞘Ciona spermatitis的精子，同时在<10 nM时表现出吸引活性，这导致了SAAF的明确结构确定和其双重活性的验证。值得注意的是，25-epi-SAAF具有与SAAF相当的活性。