丁香酚的分离及其杀多子小瓜虫活性研究

Isolation of Active Compounds fromEugenia caryophyllata against Ichthyophthirius multifiliis in Vitro

利用不同极性大小的溶剂石油醚、乙酸乙酯、氯仿、甲醇回流提取丁香制备粗提物,进行杀灭多子小瓜虫幼虫和包囊的药效活性追踪,确定丁香杀虫的活性部位为石油醚提取部分,进而利用硅胶柱层析和中压制备液相等分离技术结合活性追逐试验追踪分离丁香石油醚提取物（对多子小瓜虫幼虫和包囊的100%致死质量浓度分别为50mg/L和80mg/L）,最终分离获得一种杀虫活性物质,经质谱、核磁氢谱、核磁碳谱等多种波谱分析技术确定该活性物质为丁香酚。体外杀虫结果表明,丁香酚对多子小瓜虫具有较强的杀灭作用,其对幼虫15min、1h、2h、3h和4h的半数致死质量浓度分别为6.21（5.85~6.65）mg/L、3.50（3.24~3.84）mg/L、2.90（2.62~3.22）mg/L、2.33（2.10~2.50）mg/L和2.15（1.98~2.32）mg/L,对幼虫4h的100%杀灭的质量浓度为3.5mg/L,对包囊6h的100%杀灭质量浓度为3.5mg/L。结果表明,丁香酚是一种具有较好开发前景的杀多子小瓜虫药物。

Ichthyophthirius multifiliis is a holotrichous protozoan that invades the gills and skin surfaces of fish and can cause morbidity and high mortality in most species of freshwater fish worldwide. The pres- ent study was undertaken to investigate the antiparasitic activity of crude extracts and pure compounds from Eugenia caryophyllata. The dried E. caryophyllata was extracted successively in a separating funnel with petroleum ether, ethyl acetate, chloroform and methanol, in which only the petroleum ether extract showed promising activity （100% effectiveness against I. multifiliis theronts and encysted tomonts at the concentration of 50.0 mg/L and 80.0 mg/L respectively） and therefore, was fractionated on silica gel column chromatography in a bioactivity-guided isolation affording a compound showing potent activity. The structure of the compound was elucidated as clove oil by hydrogen and carbon-13 nuclear magnetic reso- nance spectrum and electron ionization mass spectrometry. The in vitro tests revealed that clove oil was found to be 100% effective against encysted tomonts at the concentration of 16. 0 mg/L. The results showed that clove oil has potent anti-parasitic efficacy with the 15 min-, 1 h-, 2 h-, 3 h- and 4 h-LC50 （95% Confidence Intervals） of 6.21 （5.85-6.65） mg/L, 3.50 （3.24 -3.84） mg/L, 2.90 （2.62-3.22） mg/L, 2.33 （2.10-2.50） mg/L and 2.15 （1.98-2.32） mg/L against theronts of I. rnultifiliis, respectively. The findings provide important information for the potential application of clove oil in the therapy of serious infection caused by I. multifiliis.