蟾毒配基磺酸化及基于自制串联固相萃取对磺酸化产物富集的研究

Study of Bufogenins on Sulfonation Reaction and Enrichment of Sulfated Bufogenins via a Homemade RP/HILIC-SPE

关于抗癌活性的蟾蜍二烯内酯的结构修饰化合物已达到100多个。甾体母核3位磺酸化的蟾蜍二烯内酯是从蟾蜍皮中分离得到的一类具有抗癌活性的微量成分,但目前尚无关于蟾蜍二烯内酯磺酸化修饰的报道。该文以沙蟾毒精为例,首次发展了一种高效的3位磺酸化沙蟾毒精的合成方法,对目标产物进行了结构鉴定。基于自制亲水性材料Click TE-Cys,发展了反相/亲水串联固相萃取的反应产物后处理方法,用于磺酸化沙蟾毒精的富集。依此法对总蟾毒配基进行了磺酸化和目标产物的富集及LC-MS定性分析,实现了大量具有潜在抗癌活性的新化合物的制备和富集,为进一步实现高效低毒化合物的筛选提供了物质基础。

Bufadienolide is a kind of natural steroids with significant anti-cancer activities. Hitherto, the number of structural modification on the bufadienolides has reached more than 100. The purification of sulfated, a trace cytotoxic component bufadienolides from toad skin is a tough work due to its low content and unstable structures. However, there is no report on synthesis of bufadienolides sulfonation though they have promising cytotoxic activities. In this paper, the arenobufagin was firstly taken as an example, to develop an effective method for the synthesis of arenobufagin-3-O-sulfonate. In order to enrich the arenobufagin-3-O-sulfonate quickly and mildly, a reversed phase/hydrophilic solid-phase extraction method based on our homemade hydrophilic Click TE - Cys was developed. The structures of arenobufagin-3-O-sulfonate and arenobufagin were both identified by MS,＇ H NMR and 13C NMR spectra. In addition, the total bufogenins were sulfonated by this method and the sulfated bufogenins were also successfully enriched via the reversed phase/hydrophilic solid-phase extraction approach. Finally, a LC - MS system was employed to test the result, and about 20 sulfated bufogenins were primarily identified according to their molecular weight. Based on this, a systemic method was first developed for the synthesis and enrichment of the sulfated total bufogenins and a large number of potential anticancer new compounds were obtained, which provided lots of agents for further screening to achieve high effective and low toxic compounds. The efficient reversed phase/hydrophilic solid-phase extraction method for the enrichment of sulfated bufogenins could be used to enrich sulfated bufogenins in toad skin extract.