Electrospray–Mass Spectrometry-Guided Targeted Isolation of Indole Alkaloids from Leaves of Catharanthus roseus by Using High-Performance Countercurrent Chromatography

Electrospray mass spectrometry <i>off-line</i> profiling monitored the recovery of targeted indole alkaloids from a fortified crude extract of <i>Catharanthus roseus</i> (790 mg) using semi-preparative <i>high-performance countercurrent chromatography</i> (HPCCC) fractionation. Visualization of selected single-ion traces projected the HPCCC molecular weight elution profile. Experimental <i>partition-ratio</i> values <i>K_D</i> and peak widths for detected metabolites were determined. Structural characterization of metabolites and co-elution effects were monitored in the scan range <i>m</i>/<i>z</i> 100-2000. In this study, the biphasic solvent system containing <i>n</i>-hexane-<i>n</i>-butanol-water with 0.5% ion-pair reagent trifluoro-acetic acid [1:1:2, <i>v</i>/<i>v</i>/<i>v</i>] was used based on <i>partition ratio K_D</i>-value <i>liquid chromatography-electrospray ionization-mass spectrometry</i> (LC-ESI-MS) analysis prediction. The monitoring of target ions resulted in the isolation of six major concentrated indole alkaloids (akuammicine, catharanthine, perivine, vindoline, vindorosine, and <i>19R</i>-vindolinine), which were fully elucidated by 1D and 2D <i>nuclear magnetic resonance</i> (NMR) spectroscopy.