Estimation of Isradipine in Human Plasma by LCMS/MS

A rapid and sensitive method for quantification of the antihypertensive drug, isradipine in human plasma using LCMS/MS has been developed. Protein was precipitated from the plasma by 20% trichloroacetic acid. It was then made alkaline with 1M sodium hydroxide. Isradipine and the internal standard (IS), Nimodipine, were extracted by a simple liquid -liquid extraction method using extraction solvent mixture (n-Hexane: t-butyl methyl ether-70:30, v/v). The extract was then separated by a reverse phase HPLC on a XTerra MSC18 column (100mm x 3.0 mm, 5μm) using solvent mixture 42:58, v/v ( Mobile phase 1: 0.1%, v/v, Formic Acid; Mobile phase 2: Acetonitrile: Methanol- 75:25, v/v). Flow rate was 0.5 ml/min without splitter and the column oven temperature was 35°C. The analytes were then detected by monitoring the transitions m/z 372.1 → m/z 312.0 for isradipine and 419.0 → 343.0 for nimodipine using API 3000 LCMS/MS system (Applied Biosystems) with Turbo Ion Spray interface. A good linear standard curve was obtained within the range of 0.051ng/ml - 20.448ng/ml (r > 0.9973). The lower limit of quantitation (LLOQ) was 0.051 ng/ml. The overall accuracy and precision were 94.28% and 6.19%, respectively. No significant degradation for isradipine in human plasma was observed at room temperature (7h) or when subjected to freeze thaw procedures (3 cycles). No significant metabolic compounds were found to interfere with the analysis. The results of method validation offer the acceptable performance for the specificity, selectivity, sensitivity, linearity, accuracy, precision and stability. This method can be applied for quantitation of isradipine in dosed human plasma samples.