A HPLC-MS/MS METHOD DEVELOPMENT AND VALIDATION FOR THE SIMULTANEOUS DETERMINATION OF NIFEDIPINE AND ENALAPRIL IN HUMAN PLASMA

Abstract

Objective: The main purpose of this study was to develop a simple, precise, rapid and accurate method for the simultaneous quantification of nifedipine and enalapril in human plasma.Methods: Chromatography was achieved on Discovery C18, 50 × 2.1 mm, 5 μm column. Samples were chromatographed in a gradient mode (eluent A (acetonitrile-water–formic acid, 5: 95: 0.1 v/v), eluent B (acetonitrile–formic acid, 100: 0.1 v/v)). The initial content of the eluent B is 0%, which increases linearly by 1.0 min to 100% and to 1.01 min returns to the initial 0%. The mobile phase was delivered at a flow rate of 0.400 ml/min into the mass spectrometer ESI chamber. The sample volume was 5 μl.Results: The total chromatographic run time was 2.5 min and the elution of nifedipine, enalapril and IS (verapamil) occurred at ~1.83, 1.57 and 1.61 min, respectively. A linear response function was established at 1-100 ng/ml for nifedipine and 2-200 ng/ml for enalapril maleate in human plasma. The % mean recovery for enalapril in LQC, MQC and HQC was 114.0 %, 112.9 % and 113.2 %, for nifedipine in LQC, MQC and HQC was 104.1 %, 105.0 % and 108.7 % respectively. The lowest concentration with the RSD<20% was taken as LLOQ and was found to be 2.16 ng/ml for enalapril, 1.01 ng/ml for nifedipine. The % accuracy of LLOQ samples prepared with the different biological matrix lots was found 108.2 % for enalapril and 100.5 % for nifedipine, which were found within the range of 80.00-120.00 % for the seven different plasma lots. % CV for LLOQ samples was observed as 3.2 % and 7.4 % respectively, which are within 20.00% of the acceptance criteria.Conclusion: A rapid method was developed for simultaneous determination of nifedipine and enalapril in human plasma. The method was strictly validated according to the ICH guidelines. Acquired results demonstrate that proposed strategy can be effortlessly and advantageously applied for routine examination of nifedipine and enalapril in human plasma.