A LIQUID-CHROMATOGRAPHIC METHOD FOR THE DETERMINATION OF THE ENANTIOMERIC PURITY OF THE ANTICANCER DRUG, 9-AMINO-20(S)-CAMPTOTHECIN

A method was developed for determining the enantiomeric purity of 9-am ino-20(S)-camptothecin (9-A-20(S)-CAM). The chiral derivatizing reagen t, 1-(1-naphthyl)ethyl isocyanate (NEI) was used to derivatize the ena ntiomers of 9-A-CAM, and H-1-NMR, LC-MS, and LC-UV were used to identi fy and quantitate the two diastereomers produced. During the first 24 h, derivatization was exclusively at the 9-amino nitrogen. The much sl ower reaction involving reaction of NEI with the 20-hydroxy oxygen cou ld be prevented by quenching the reaction within the first 24 h with m ethanol. NMR analysis provided useful information about the site of de rivatization; however. the partial separation of the signals was insuf ficient for quantitative analysis of the two diastereomers. Whereas ba seline resolution of the two diastereomers was achieved by reversed-ph ase LC, the reproducibilities of the resolution and the peak area rati os were dependent on the nature and composition of the mobile phase, t he flow rate, the column temperature. sample concentration and sample preparation.