High-speed countercurrent chromatography of Harpagophytum procumbens constituents and their identification by TLC-MS
Harpagophytum procumbens DC. (Pedaliaceae), known as Devil's claw, is native to the arid regions of Southern Africa including the Kalahari desert. The dried secondary root tubers have been used to reduce pain and inflammation especially in rheumatism and arthritis . Iridoid glycosides are considered to be the main pharmacologically active constituents with other constituents such as phenylethanoid glycosides and flavonoids contributing to the effect . Rapid isolation and identification of the constituents was necessary in order to acquire sufficient quantities of the reference compounds for use in further biological studies as well as to develop quantitative calibration models. To achieve these goals, a methanol extract of the secondary root tubers was rapidly filtered over silica gel to remove sugars and other polar compounds. The resulting fraction, which consisted mainly of iridoid and phenylpropanoid glycosides, was subjected to high-speed countercurrent chromatography (HSCCC). This allowed a one-step separation of the major constituents. The minor constituents were obtained either by a second HSCCC operation or by a final column chromatographic step. In order to distinguish close-running compounds in the absence of reference standards, TLC-MS  was performed on the extract and the isolated constituents. This method could be used, for example, to distinguish the close-eluting pair 8-E-p-coumaroylharpagide and 8-E-p-feruloylharpagide .
Acknowledgement: The authors thank the National Research Foundation of South Africa for financing this study.
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