Simultaneous determination of amlodipine, valsartan and hydrochlorothiazide by LC-ESI-MS/MS and its application to pharmacokinetics in rats

Abstract

Polypill is a fixed-dose combination that contains three or more active ingredients used as a single daily pill to achieve a large effect in preventing cardiovascular disease with minimal adverse effects. A novel and accurate liquid chromatography tandem mass spectrometry method using electrospray ionization mode has been developed and validated for the simultaneous determination of amlodipine (AMD), valsartan (VAL) using losartan (LOS) as an internal standard (IS), and hydrochlorothiazide (HCT) using furosemide (FSD) as an IS. The separation was carried on Aquasil C₁₈ (50 mm×2.1 mm, 5 µm) reversed phase column using acetonitrile and water containing 0.1% formic acid (50:50, v/v) as the mobile phase. The method was validated in terms of linearity, accuracy and precision over the concentration range of 1-1000 ng/mL. The intra and inter-day precision and accuracy, stability and extraction recoveries of all the analytes were in the acceptable range. This method can be successfully applied to the pharmacokinetic study of AMD, VAL and HCT when given as a polypill.

Keywords: Amlodipine; Exforge HCT; Hydrochlorothiazide; Polypill; Valsartan.

Fig. 1

Chemical structures of analytes.

Fig. 2

LC–ESI-MS/MS spectra of (A) amlodipine, (B) valsartan, (C) losartan, (D) hydrochlorothiazide and (E) furosemide.

Fig. 3

LC–ESI-MRM chromatograms of (A) blank plasma, (B) spiked plasma at 1 ng/mL, and (C) plasma extracted from rats after 1 h of oral administration.

Fig. 4

Mean plasma concentration–time profile of (A) amlodipine, (B) valsartan and (C) hydrochlorothiazide in rats. Each point represents the mean±SD (n=6).