Determination of N-(3-nitro-4-quinoline)morpholino-4-carboxamidine in plasma high-performance liquid chromatography

A high-performance liquid chromatographic (HPLC) method was developed for the determination of the radiosensitizing agent N-(3-nitro-4-quinoline)morpholino-4-carboxamidine (EGIS-4136) in plasma using an internal standard. HPLC separation was achieved on a LiChrosorb C18 column using acetonitrile-sodium acetate buffer (pH 7.2) (40:60) as the mobile phase and UV detection at 330 nm. The plasma samples were prepared for measurement by protein precipitation with methanol and centrifugation. The assay was validated with respect to linearity, sensitivity, accuracy, precision, stability and recovery in plasma. The limit of detection for EGIS-4136 in plasma was 0.05 microgram/ml. A straight line was obtained on plotting the peak-area ratio of EGIS-4136 to the internal standard against concentration in plasma in the range 0.1-20 microgram/ml. The method was applied to study the pharmacokinetics of EGIS-4136 in six male rats after a single oral dose of 50 mg/kg, and allowed the compound to be monitored in the concentration range 5-10 micrograms/ml occurring in rats 24 h post-administration.