Abstract In this study, a fast and sensitive liquid chromatography/tandem mass spectrometry method for determination of hydrochlorothiazide in human plasma was developed and validated. The analyte and irbesartan, used as the internal standard, were precipitated and extracted from plasma using methanol. Analysis was performed on a Phenomenex Kromasil C 8 column with water and methanol (27:73, v/v) as the mobile phase. Linearity was assessed from 0.78 to 200 ng/mL in plasma. The analytical method proved to be applicable in a pharmacokinetic study after oral administration of 12 mg hydrochlorothiazide tablets to 20 healthy volunteers.