Elution-extrusion counter-current chromatography for the separation of two pairs of isomeric monoterpenes from Paeoniae Alba Radix

In this work, a simple and efficient protocol for the rapid separation of two pairs of isomeric monoterpenes from Paeoniae Alba Radix was developed by combining macroporous resin and elution-extrusion counter-current chromatography. The crude extract was firstly subjected to a D101 macroporous resin column eluted with water and a series of different concentrations of ethanol. Then, effluents of 30 and 95% ethanol were collected as sample 1 and sample 2 for further counter-current chromatography purification. Finally, a pair of isomers, 96 mg of compound 1 and 48 mg of compound 2 with purities of 91.1 and 96.2%, respectively, was isolated from 200 mg of sample 1. The other pair of isomers, 14 mg of compound 3 and 8 mg of compound 4 with purities of 93.6 and 88.9%, respectively, was isolated from 48 mg of sample 2. Their purities were analyzed by high-performance liquid chromatography, and their chemical structures were identified by mass spectrometry and H-1 NMR spectroscopy. Compared to a normal counter-current chromatography separation, the separation time and solvent consumption of elution-extrusion counter-current chromatography were reduced while the resolutions were still good. The established protocol is promising for the separation of natural products with great disparity of content in herbal medicines.