Development and validation of an LC–MS/MS method for pharmacokinetic study of methoxyamine in phase I clinical trial

Methoxyamine (MX) is the first DNA base-excision-repair (BER) inhibitor evaluated in humans. This work described the development and validation of an LC–MS/MS method for quantitative determination of MX in human plasma. In this method, MX and its stable isotope methoxyl-d3-amine (MX-d3 as internal standard) were directly derivatized in human plasma with 4-(N,N-diethylamino)benzaldehyde. The derivatized MX and IS were extracted by methyl-tert-butyl ether, and separated isocratically on a Xterra C18 column (2.1 mm × 100 mm) using an aqueous mobile phase containing 45% acetonitrile and 0.4% formic acid at a flow rate of 0.200 ml/min. Quantitation of MX was carried out by multiple-reaction-monitoring (MRM) mode of positive turbo-ion-spray tandem mass spectrometry. This method has been validated according to FDA guidelines for bioanalytical method. The linear calibration range for MX was 1.25–500 ng/ml in human plasma with a correlation coefficient ≥ 0.9993. The intra- and inter-assay precision (%CV) at three concentration levels (3.50, 45.0 and 450 ng/ml) ranged 0.9–1% and 0.8–3%, respectively. The stability studies showed that MX met the acceptable criteria under all tested conditions. The method developed had been applied to the determination of plasma MX concentrations in the first-in-human phase I clinical trial, and PK data were presented.

► An LC–MS/MS method for quantitative determination of methoxyamine in human plasma has been developed and fully validated. ► This method has been applied to pharmacokinetic study of methoxyamine in a phase I clinical trial. ► The concentration–time profiles and pharmacokinetic parameters of methoxyamine in patients were presented.