The simultaneous UPLC–MS/MS determination of emerging drug combination; candesartan and chlorthalidone in human plasma and its application

A novel, precise, sensitive and accurate ultra‐performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) method has been developed for the simultaneous determination of a novel drug combination, candesartan (CAN) and chlorthalidone (CHL), in human plasma. Chromatographic separation was achieved on Waters Acquity UPLC BEH C18 (50 × 2.1 mm, 1.7 μm). Mobile phase consisting of 1 mm ammonium acetate in water–acetonitrile (20:80 v/v) was used. The total chromatographic runtime was 1.9 min with retention times for CAN and CHL at 0.7 and 1.1 min respectively. Ionization and detection of analytes and internal standards was performed on a triple quadrupole mass spectrometer, operating in the multiple reaction monitoring and negative ionization mode. Quantitation was done to monitor protonated precursor → product ion transition of m/z 439.2 → 309.0 for CAN, 337.0 → 189.8 for CHL and 443.2 → 312.1 for candesartan D4 and 341.0 → 189.8 for chlorthalidone D4. The method was validated over a wide dynamic concentration range of 2.0–540.0 ng/mL for candesartan and 1.0–180.0 ng/mL for chlorthalidone. The validated method was successfully applied for the assay of CAN and CHL in healthy volunteers.