Development and validation of an ultrasensitive LC-MS/MS method for the quantification of cetagliptin in human plasma and its application in a microdose clinical trial.

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was established and validated for the ultrasensitive determination of cetagliptin in human plasma. Sample pretreatment was achieved by liquid-liquid extraction with ethyl acetate, and chromatographic separation was performed on an XB-C18 (50×2.1 mm, 5 μm) analytical column with gradient elution (0.1% formic acid in acetonitrile and 0.1% formic acid) at a flow rate of 1.0 mL/min. For mass spectrometric detection, the multiple reaction monitoring was used and the ion transitions monitored were m/z 421.2-86.0 for cetagliptin and m/z 424.2-88.0 for cetagliptin-d3. Method validation was performed according to the US Food and Drug Administration Bioanalytical Method Validation Guidance, for which the calibration curve was linear in the range of 50.0-2000 pg/mL. All of the other results, such as selectivity, lower limit of quantitation (LLOQ), precision, accuracy, matrix effect, recovery and stability, could meet the acceptance criteria. The validated method was successfully applied in a microdose clinical trial to systematically investigate the pharmacokinetic profile of cetagliptin in healthy subjects. Both rapid absorption and prolonged duration demonstrate the potential value of cetagliptin for diabetes treatment.