Simple method for simultaneous quantification of two new derivatives of betulinic and ursolic acids with antimalarial activity by RP-HPLC-UV-CAD

Leonardo Trevisan Secretti, Andrea Pereira, Simone Cristina Baggio Gnoatto, Lívia Maronesi Bueno, Pâmela Lukasewics Ferreira, Pedro Eduardo Froehlich


Endemic in more than 90 countries and territories, malaria is the most widely, populational, and geographically, parasitic disease in the world. Plasmodium sp. resistance to available drugs is one of the biggest problems for malaria eradication. In this study, we develop a method for the simultaneous determination of two new derivatives of betulinic and ursolic acids with antimalarial activity designated 3-O-butanoylbetulinic and 3-O-butanoylursolic acids. An analytical method was developed by high-performance liquid chromatography coupled, in series, to ultraviolet (UV) and charged aerosol (CAD) detectors. The chromatographic system, operated isocratically by reversed-phase, consisted in a mobile phase composed of acetonitrile: water pH 3.0 adjusted with formic acid (85:15, v/v), flow rate of 1.2 mL/min and a PhenoSphere Next octadecylsilane column (250 mm x 4.6 mm, 5 μm particle size). Chromatograms were recorded simultaneously in UV and CAD, at a concentration of 50 µg mL-1, an injection volume of 20 μL at a controlled temperature of 50 °C. The method was found to be selective, linear (r > 0.99), precise (RSD < 2.0%), accurate, and robust for both analytes, and considered statistically validated, and can be applied to the identification and quantification of these new drug candidates.

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