固相萃取法纯化制备水飞蓟宾和异水飞蓟宾

Purification and preparation of silybin and isosilybin by solid phase extraction

水飞蓟宾和异水飞蓟宾是水飞蓟素中的主要有效成分,其纯化制备主要借助柱色谱法,制备量大,纯化效果好,但过程非常费时。该研究的主要目的是利用更为快速高效的固相萃取(SPE)法从水飞蓟粗提物中分离纯化水飞蓟宾和异水飞蓟宾。建立了用于分析水飞蓟宾和异水飞蓟宾的高相液相色谱法,通过优化洗脱梯度,实现了水飞蓟宾、异水飞蓟宾与其他组分的分离。试用了3种保留机理不同的SPE填料,包括亲水亲脂(HLB)填料、亲水色谱(HILIC)填料及反相C18硅胶填料。通过对比发现C18硅胶对目标化合物的选择性最佳。进一步控制SPE的淋洗及洗脱条件,收集相应的洗脱液,经氮吹干燥后得到纯化的样品。提纯后的水飞蓟宾和异水飞蓟宾混合物的纯度可达94%以上。水飞蓟宾和异水飞蓟宾的平均回收率分别为70.5%~81.7%和66.7%~81.8%,相对标准偏差分别为2.7%~9.4%和1.5%~6.1%。该方法简单、高效,免去传统分离纯化过程中长时间的柱色谱分离过程,适合制备纯度较高的水飞蓟宾和异水飞蓟宾的二元混合标准样品。

Silybin and isosilybin are mainly effective elements of silymarin. To improve the performance of treatment and decrease the side effects from the unknown components,purified silybin and isosilybin are in request. The main purpose of this research was to develop a solid phase extraction（SPE） method for the purification of silybin and isosilybin from silymarin extract. For the analysis of the samples,a liquid chromatography method with ultraviolet detector（LC-UV）was developed to separate the silybin and isosilybin from their impurities.The separation was achieved in 8 min and a total analytical time of 11 min with column washing and equilibration. In the SPE,three different SPE packings were tested,including hydrophiliclipophilic balance（HLB）,hydrophilic interaction chromatography（HILIC）and reverse phase C18 silica microspheres. Based on the selectivity against silybin and isosilybin, the C18 packings were chosen. After the optimization of the elution of SPE,the purity over 94% could be achieved for the silybin and isosilybin mixture with the recoveries of 70. 5%- 81. 7% and66. 7%-81. 8%,and the relative standard deviations（RSDs）of 2. 7%- 9. 4% and 1. 5%- 6. 1%,respectively. Though silybin and isosilybin could not be separated from each other by using SPE,this is a useful method for the purification of silybin and isosilybin mixture from the crude extract of their plant seeds due to its simplicity and efficiency.