Simultaneous Quantification of Some Pharmaceutical Catharanthus roseus Leaf and Root Terpenoid Indole Alkaloids and Their Precursors in Single Runs by Reversed-Phase Liquid Chromatography

A new, rapid, sensitive, and reproducible reversed-phase liquid chromatographic (LC) method with photodiode array detection is described. It allows, in a single run of 30 min, simultaneous separation of 6 pharmaceutically and biologically important Catharanthus roseus leaf and root terpenoid indole alkaloids (TIAs) and 3 of their precursors: TIA precursors tryptophan, tryptamine, and loganine; and TIAs serpentine, catharanthine, ajmalicine, vincristine, vinblastine, and vindoline. The method involves the use of a Phenomenex Luna 5 μm, C18 column (250 mm × 4.6 mm id) and a linear binary gradient mobile phase profile. Detection is performed at 220 and 254 nm, which provided good absorptivity for all of the roseus compounds listed above and gave a minimum detection limit of 0.02 μg/mL. The extraction efficiency, peak purity, and homogeneity parameters of the profiles could be validated using a photodiode array detector. The method was successfully used to quantify major components of leaf and root extracts of C. roseus accessions. The new method thus provides a reliable tool for rapid screening of C. roseus samples in large numbers, which is needed in breeding/genetic engineering and genetic mapping experiments and for monitoring the reaction products, in the in vitro/in vivo conversions of precursors into products, and vice versa.