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Analysis of selected withanolides in plant extract by capillary electrochromatography and microemulsion electrokinetic chromatography
Author(s) -
Cherkaoui Samir,
Cahours Xavier,
Veuthey JeanLuc
Publication year - 2003
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.200390043
Subject(s) - chromatography , chemistry , capillary electrochromatography , micellar electrokinetic chromatography , mass spectrometry , microemulsion , electrochromatography , sodium dodecyl sulfate , capillary electrophoresis , electrospray ionization , electrospray , high performance liquid chromatography , analytical chemistry (journal) , electrokinetic phenomena , pulmonary surfactant , biochemistry
Microemulsion electrokinetic chromatography (MEEKC) coupled with a diode‐array detector was developed for the simultaneous analysis of natural steroidal compounds, withanolides including withaferin A, withacnistin and iochromolide. Optimal resolution was obtained with a microemulsion consisting of 70 m M octane, 800 m M 1‐butanol, 100 m M sodium dodecyl sulfate (SDS), and 10 m M phosphate‐borate buffer (pH 7) using a fused‐silica capillary at 25 kV and 40°C. Since this technique is not compatible with mass spectrometry detection, a capillary electrochromatographic method was developed to separate the investigated withanolides. The effects of mobile phase composition and pH were systematically investigated. Complete separation was obtained with a capillary electrochromatography (CEC) Hypersil C18 bonded silica column (packed length, 25 cm×100 μm ID and 375 μm OD), packed with 3 μm particles. The mobile phase consisted of formic acid‐ammonia, pH 8 / acetonitrile (40/60 v/v); the voltage was set at 25 kV and the temperature at 20°C. Under these conditions, resolution of these closely related compounds, including the critical pair withacnistin and iochromolide, was achieved in less than 5 min. The separations by MEEKC and CEC were compared with that obtained by reversed‐phase liquid chromatography and showed similar retention order, indicating the analogy of the retention mechanism of these techniques. To further improve specificity and sensitivity, the developed CEC method was interfaced with electrospray ionization mass spectrometry using a Teflon connection between the CEC column and a void fused‐silica capillary. Finally, the described methods were applied to the qualitative analysis of withanolides in Iochroma gesnerioides plant extract.