Preparative Separation of Indole Alkaloids from the Rind of Picralima nitida (Stapf) T. Durand & H. Durand by pH‐Zone‐Refining Countercurrent Chromatography

Abstract

Picralima nitida alkaloids have been shown to possess in vitro antimalarial activity comparable to the clinical antimalarial chloroquine and quinine. The in vitro IC₅₀ values for these alkaloids ranged from 0.01 to 0.9 µg/mL against chloroquine‐sensitive (D6) and chloroquine‐resistant (W2) strains of Plasmodium falciparum. As part of our continuing research aimed at identifying new antimalarial leads from African rain forest plants, we have been highly successful in employing pH‐zone‐refining counter‐current chromatography (CCC) to separate and isolate the Picralima alkaloids. The separation was performed in two steps with a two‐phase solvent system composed of tert‐butyl methyl ether (MTBE)/acetonitrile/water (2∶2∶3, v/v), in which the lower phase was used as the mobile phase at a flow‐rate of 2.0 mL min⁻¹ in the head‐to‐tail elution mode pH‐zone‐refining counter‐current chromatography. The resulting fractions contained the alkaloids alstonine, akuammigine, akuammine, picraline, akuammine, akuammicine, picratidine were identified using thermospray liquid chromatography‐mass spectrometry (LC‐MS) and by TLC co‐elution experiments with authentic samples. The application of pH‐zone‐refining CCC to perform successful separations at the multigram level will be discussed. The Picralima alkaloids may represent an entirely new antimalarial chemotype with possible pharmacokinetics advantages over the existing drugs.

Keywords:

• Preparative separation
• pH‐zone‐refining counter‐current chromatography
• indole alkaloids
• Picralima nitida
• Anti‐malarial activity
• alstonine

Acknowledgments

We gratefully acknowledge financial support from the ICBG “Drug Development and Conservation in West and Central Africa” Grant No TW03004 from the Fogarty Center at NIH.