Determination of metolazone in human blood by liquid chromatography with electrospray ionization tandem mass spectrometry

A rapid, sensitive and accurate liquid chromatographic–tandem mass spectrometric method is described for the determination of metolazone in human blood. Metolazone was extracted from blood using ethyl acetate and separated on a C18 column interfaced with a triple quadrupole tandem mass spectrometer. The mobile phase consisting of a mixture of acetonitrile, 10 mmol/l ammonium acetate and formic acid (60:40:0.1, v/v/v) was delivered at a flow rate of 0.5 ml/min. Electrospray ionization (ESI) source was operated in positive ion mode. Selected reaction monitoring (SRM) mode using the transitions of m/z 366 → m/z 259 and m/z 321 → m/z 275 were used to quantify metolazone and the lorazepam (internal standard), respectively. The linearity was obtained over the concentration range of 0.5–500 ng/ml for metolazone and the lower limit of quantitation (LLOQ) was 0.5 ng/ml. For each level of QC samples, inter- and intra-run precision was less than 8.07 and 3.56% (relative standard deviation (RSD)), respectively, and the bias was within ±4.0%. This method was successfully applied to the pharmacokinetic study of metolazone formulation after oral administration to humans.