Short communicationDetermination of metolazone in human blood by liquid chromatography with electrospray ionization tandem mass spectrometry
Introduction
Metolazone (Fig. 1), (7-chloro-1,2,3,4-tetrahydro-2-methyl-4-oxo-3-o-tolyl-6-quinazoline-sulfonamide), is a diuretic agent. It has been clinically used for the treatment of hypertension and edema [1], [2]. For the pharmacokinetic study of metolazone formulation in humans, an analytical method with simplicity and high sensitivity was required in our laboratory. Various methods have been reported for the determination of metolazone in biological samples, which involved high-performance liquid chromatographic method [3], [4], liquid chromatography with fluorescence detection [5] and liquid chromatography–tandem mass spectrometry (LC–MS–MS) [6]. Taking into consideration the low levels of metolazone in human blood, LC–MS–MS method is the first choice for our purpose. But the published LC–MS–MS method [6] addressed the qualitative analysis of a metolazone metabolite, not the quantitative determination of metolazone. This paper describes a simple, specific and highly sensitive LC–MS–MS method with an electrospray ionization (ESI) source in selected reaction monitoring (SRM) mode for the determination of metolazone in human blood. The described method was validated in terms of matrix effect, selectivity, sensitivity, linearity, accuracy, precision and stability of analyte in human blood, and successfully applied to the pharmacokinetic studies of metolazone formulation in humans.
Section snippets
Chemicals and reagents
Metolazone (purity, 99.0%) and tablets (2.5 mg, batch no. 20051124) were supplied by Tianjin Institute of Pharmaceutical Research (Tianjin, China). Lorazepam (purity, 99.5%, internal standard) was also supplied by Tianjin Institute of Pharmaceutical Research. HPLC-grade acetonitrile and formic acid were purchased from Tianjin Concord Tech Reagent Company (Tianjin, China). Ammonium acetate, analytical-grade, was purchased from Tianjin Chemical Reagent Co. Ltd. (Tianjin, China). Water was purified
Method development
Liquid–liquid extraction (LLE) was used for the sample preparation in this work. LLE can be helpful in producing a clean sample and avoiding the introduction of non-volatile materials onto the column and MS system. Clean samples are essential for minimizing ion suppression and matrix effect in LC–MS–MS analyses. Two organic solvents, ethyl acetate and chloroform were evaluated. Finally, ethyl acetate was found to be optimal, which can produce a clean chromatogram for a blank blood sample and
Conclusions
A rapid, sensitive and accurate liquid chromatography with electrospray ionization tandem mass spectrometry was developed for the determination of metolazone in human blood. The method was successfully applied to the pharmacokinetic studies of metolazone formulation in humans.
Acknowledgement
This work was supported by the National High Technology Program (863 program) of China (grant number 2003AA2Z347D).
References (6)
- et al.
J Chromatogr. B
(1994) - et al.
J Chromatogr.
(1981) - et al.
J Chromatogr.
(1990)