GRADIENT HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC ASSAY FOR THE DETERMINATION OF THE NOVEL INDOLOQUINONE ANTITUMOR AGENT-E09 IN BIOLOGICAL SPECIMENS

A high-performance liquid chromatographic method for the determination of the novel indoloquinone antitumour agent E09, 3-hydroxymethyl-5-aziridinyl-1-methyl-2-(1H-indole-4,7-dione)prop-β-enα-ol, in mouse plasma and urine is described. Following protein precipitation by means of methanol (2 volumes), separation and quantification of parent drug, metabolites and internal standard E012 (5-morpholine substituted analogue) were achieved on a 5-μm Resolve C18 Rad-Pak with a 15-min linear gradient of 10-30% acetonitrile in a 0.02 M pH 7.4 sodium phosphate buffer with UV detection at 280 and 310 nm. The utility of the assay is also demonstrated for the aziridine ring-opened analogue E05A, 3-hydroxymethyl-5-β-hydroxyethylamino-2-(1H-indole-4,7-dione)prop-β-enα-ol. Plots of area ratios of analytes versus internal standard were linear in the range 50-15 000 ng/ml. The detection limit for indoloquinones in plasma was ca. 30 ng/ml. The within-assay and day-to-day variation were consistently lower than 12.5%. The assay was applied in preliminary pharmacokinetic investigations. One minor metabolite of E09 could be identified; further metabolites were characterized by ultraviolet-visible spectra. © 1990.