A method for the extraction and quantification of carvedilol, enalaprilat, and perindoprilat in 50 µL human plasma, using high-performance liquid chromatography with tandem mass spectrometry (LC–MS/MS) detection was developed and validated. Samples were prepared via protein precipitation with chromatographic separation on a Restek Ultra II Biphenyl column using gradient elution at a corresponding flowrate of 300 µL/min. Electrospray ionisation with mass detection at unit resolution in the multiple reaction monitoring (MRM) mode on an AB Sciex API 5500 mass spectrometer was used. Accuracy, precision, selectivity, sensitivity, matrix effects, recovery, process efficiency, and stability were assessed over the validation period. The assay was validated over the calibration range 0.2–200 ng/mL for all three analytes. The inter- and intra-day precision expressed as the coefficient of variation (CV) and accuracy (%Nom) all fell within acceptable limits. The overall recovery was calculated as 72.9%, 77.1%, and 77.0% for carvedilol, enalaprilat, and perindoprilat respectively, with the recovery being shown to be reproducible at the low, medium and high end of the calibration range for all three analytes. The method proved to be specific for all three analytes with no significant matrix effects observed. The validated method facilitated the analysis of carvedilol, enalaprilat, and perindoprilat in human plasma collected from adults as part of a pilot pharmacokinetic study. This validated analytical method lays the foundation for determining adherence in heart failure patients prescribed with carvedilol, enalapril and perindopril.

No abstract