Preparative separation of phenylpropanoid glycoside from scrophularia ningpoensis hemsley by high speed countercurrent chromatography and ESI-MSn analysis

The characteristic constituent angoroside C of the famous traditional Chinese medicine Scrophularia ningpoensis hemsley was separated from the crude extract by high speed countercurrent chromatography using reverse phase elution combined with silica gel column chromatography. Meanwhile, harpagoside was also obtained using the normal phase elution method. A two phase solvent system containing n-butanol:acetic acid:ethyl acetate:water (8:1:1:10) was selected according to the partition coefficient of angoroside C. A 311mg quantity of the crude extract was loaded onto a 250mL HSCCC column and yielded 49mg angoroside C at over 89.1% purity, and 14mg harpagoside at over 96.5% purity. Angoroside C was further purified by silica gel column chromatography and the purity reached 98.5% with high recovery. The chemical structures were determined by ESI/MS n and 1H-NMR. The characteristic ESI/MSn fragmentation pattern of angoroside C and the proposed fragmentation mechanism are proposed.