Advantages of automation in plasma sample preparation prior to HPLC/MS/MS quantification: application to the determination of cilazapril and cilazaprilat in a bioequivalence study

• Published in: Journal of AOAC International 2011-05, Vol.94 (3), p.758-764
• Main Authors: Kolocouri, Filomila, Dotsikas, Yannis, Apostolou, Constantinos, Kousoulos, Constantinos, Soumelas, Georgios-Stefanos, Loukas, Yannis L
• Format: Article
• Language: English
• Subjects: Angiotensin-Converting Enzyme Inhibitors - chemistry; Angiotensin-Converting Enzyme Inhibitors - pharmacokinetics; Automation; Chemical properties; Chromatography, High Pressure Liquid - methods; Cilazapril; Cilazapril - administration & dosage; Cilazapril - analogs & derivatives; Cilazapril - chemistry; Cilazapril - pharmacokinetics; Drug Stability; High performance liquid chromatography; Humans; Identification and classification; Mass spectrometry; Mechanization; Methods; Molecular Structure; Sensitivity and Specificity; Specimen Handling - methods; Tandem Mass Spectrometry - methods; Technology application; Therapeutic Equivalency
• ISSN: 1060-3271; 1944-7922
• DOI: 10.1093/jaoac/94.3.758

An HPLC/MS/MS method characterized by complete automation and high throughput was developed for the determination of cilazapril and its active metabolite cilazaprilat in human plasma. All sample preparation and analysis steps were performed by using 2.2 mL 96 deep-well plates, while robotic liquid handling workstations were utilized for all liquid transfer steps, including liquid-liquid extraction. The whole procedure was very fast compared to a manual procedure with vials and no automation. The method also had a very short chromatographic run time of 1.5 min. Sample analysis was performed by RP-HPLC/MS/MS with positive electrospray ionization using multiple reaction monitoring. The calibration curve was linear in the range of 0.500-300 and 0.250-150 ng/mL for cilazapril and cilazaprilat, respectively. The proposed method was fully validated and proved to be selective, accurate, precise, reproducible, and suitable for the determination of cilazapril and cilazaprilat in human plasma. Therefore, it was applied to a bioequivalence study after per os administration of 2.5 mg tablet formulations of cilazapril.