LC–MS/MS assay of quinapril and its metabolite quinaprilat for drug bioequivalence evaluation: prospective, concurrential and retrospective method validation

Abstract

Background: The bioequivalence of two pharmaceutical formulations containing 10 mg quinapril was assessed by assaying the untransformed drug and its active metabolite quinaprilat from plasma samples. Results: A gradient elution liquid chromatographic separation coupled to positive atmospheric pressure electrospray ionization and tandem mass spectrometry detection was used and validated. Sample preparation is simple and uses protein precipitation through addition of an acetonitrile:methanol (8:2 v/v) mixture. The method has a run time of 6.3 min. Carvedilol was used as an internal standard. The multiple reactions monitoring mode was used for both quantitation and structural confirmation of target compounds. Linear 1/x2-weighted regressions characterize detector response function up to concentrations of 1000 ng/ml for quinapril and 2000 ng/ml for quinaprilat. Low limits of quantitation of 5 ng/ml for quinapril and 10 ng/ml for quinaprilat were found. Intra- and inter-day variability of the results were found below 15%. Long- (-20°C/6 months) and short-term (25°C/48 h) stability of analytes in plasma, as well as freeze and thaw stability (six cycles) were demonstrated. Conclusion: The method was found to be selective, precise, accurate and robust when applied to a large number of unknown samples.