College of Pharmaceutical Science, Zhejiang University of Technology, Hangzhou, 310014, China.
Institute of Engineering Biology and Health, Collaborative Innovation Center of Yangtze River Delta Region Green Pharmaceuticals, College of Pharmaceutical Science, Zhejiang University of Technology, Hangzhou, 310014, China.
J Chromatogr A. 2020 Dec 20;1634:461666. doi: 10.1016/j.chroma.2020.461666. Epub 2020 Nov 2.
Off-line two-dimensional countercurrent chromatography has been widely applied to the isolation of complex samples, but little research on the investigation of orthogonality in the selection of biphasic solvent systems is available. In the present work, the orthogonality in the selection of a biphasic solvent system for liquid-liquid chromatographic separation of aqueous extract and ether extract from the traditional Chinese medicinal plant Polygonum cuspidatum Sieb. et Zucc was evaluated by the correlation coefficient and space occupancy rate. In total, 25 different biphasic solvent systems were tested, and 313 system combinations were analysed. A convex hull methodology was used to determine the separation space and to optimize separation conditions. The correlation coefficient matrix was transformed into dendrograms and a colour map to visualize the dissimilarity between, and orthogonality for, all solvent systems. The aqueous extracts from Polygonum cuspidatum were separated using selected biphasic solvent systems with high orthogonality: ethyl acetate-ethanol-water (70:1:70, v/v) and petroleum ether-ethyl acetate-water (1:5:5, v/v). The ether extracts from Polygonum cuspidatum were also separated using selected biphasic solvent systems with high orthogonality: petroleum-ethyl acetate-methanol-aqueous 0.25 M NH•HO (5:5:5:5, v/v) and petroleum-ethyl acetate-methanol-water (5:5:5:5, v/v). Thirteen compounds were successfully obtained. The experimental results demonstrated that the evaluation of orthogonality provided an alternative strategy to select an applicable solvent system for the separation of complex samples using off-line two-dimensional countercurrent chromatography.
离线二维逆流色谱已广泛应用于复杂样品的分离，但在两相溶剂系统选择中的正交性研究较少。本工作采用相关系数和空间占有率评价了从传统中药虎杖的水提物和醚提物中液液色谱分离两相溶剂系统选择中的正交性。共测试了 25 种不同的两相溶剂系统，分析了 313 种系统组合。采用凸包法确定分离空间并优化分离条件。将相关系数矩阵转化为聚类图和色图，直观地显示所有溶剂系统之间的差异和正交性。选择具有高正交性的两相溶剂系统分离虎杖的水提物：乙酸乙酯-乙醇-水（70:1:70，v/v）和石油醚-乙酸乙酯-水（1:5:5，v/v）。选择具有高正交性的两相溶剂系统分离虎杖的醚提物：石油醚-乙酸乙酯-甲醇-0.25 M NH 4 OH（5:5:5:5，v/v）和石油醚-乙酸乙酯-甲醇-水（5:5:5:5，v/v）。成功获得了 13 种化合物。实验结果表明，正交性评价为使用离线二维逆流色谱分离复杂样品选择适用溶剂系统提供了一种替代策略。
