Determination of O-(chloroacetylcarbamoyl)fumagillol (TNP-470;AGM-1470) and two metabolites in plasma by high-performance liquid chromatography mass spectrometry with atmospheric pressure chemical ionization

High performance liquid chromatography/mass spectrometry using a heat nebulizer atmospheric pressure chemical ionization interface was investigated as a sensitive and selective technique for the chromatographic separation and quantitation of the anti-angiogenic drug TNP-470 and its polar metabolites MII and MIV. Upon extraction from plasma, the analytes were measured using selected-ion monitoring in the positive ionization mode and single quadrapole mass spectrometric detection. A trideuterated C-13-labeled analog of TNP-470 was used as an internal standard, Two plasma extraction procedures were evaluated, An acetonitrile precipitation method resulted in a recovery of 65.9% and 44.4% for MIV and TNP-470, respectively, and a limit of quantitation of 2.5 ng ml(-1) with a limit of detection of 0.63 ng ml(-1) for both analytes. In contrast, a solid-phase extraction method led to a higher recovery of 92.9% and 111.0% for MIV and TNP-470, respectively, and a limit of quantitation of 0.63 ng ml(-1) with a limit of detection of 0.16 ng ml(-1) for both analytes, Intra-day and inter-day precisions were < 10% and < 18% for both analytes, The solid-phase extraction, by decreasing plasma interferences present within the column capacity, also allowed the detection and quantitation of MII, a highly polar metabolite of TNP-470. The recovery of MII in plasma was 77.4% with a limit of quantitation and detection of 2.5 and 1.25 ng ml(-1), respectively, Intra-day and inter-day precisions were 6.19% and 2.99%, Quantitative analysis of plasma samples from a patient receiving 10 mg m(-2) of TNP-470 by a 1 h intravenous infusion is reported.