Isolation and Purification of Three Ecdysteroids from the Stems of Diploclisia glaucescens by High-Speed Countercurrent Chromatography and Their Anti-Inflammatory Activities In Vitro.

High-speed counter-current chromatography was used to separate and purify ecdysteroids for the first time from the stems of Diploclisia glaucescens using a two-phase solvent system composed of ethyl acetate-n-butanol-ethanol-water (3:0.2:0.8:3, v/v). Three ecdysteroids were obtained from 260 mg of ethyl acetate extract of the residue obtained after evaporation of the crude ethanolicextractof D. glaucescens in one-step separation, which were identified as paristerone (I, 30.5 mg), ecdysterone (II, 7.2 mg), and capitasterone (III, 8.1 mg) by electrospray ionization mass spectrometry (ESI-MS) and nuclear magnetic resonance (NMR). Their anti-inflammatory activities were evaluated by measuring the inhibitory ratios of β-glucuronidase release in rat polymorphonuclear leukocytes (PMNs) induced by platelet-activating factor. Compounds I-III showed significant anti-inflammatory activities with IC50-values ranging from 1.51 to 11.68 μM, respectively.