Informatics Publishing Limited

Srinivas Reddy ¹, Jibin V. Joseph ¹, Aravind Kumar ¹, Arindam Mukhopadhyay ¹, Saral Thangam ¹

Affiliations
1 Norwich Clinical Services Pvt. Ltd., 147/F, 8th Main, 3rd Block, Koramangala, Bangalore– 560034

Abstract

After extraction from plasma by liquid - liquid extraction method, Felodipine and FelodipineD5, (IS), were separated on a Reverse phase chromatography using the mobile phase mixture of ammonium acetate and methanol at a flow rate of 1.0 ml/min. The analytes were detected in API 4000 Mass spectrometer in the positive atmospheric pressure chemical Ionization (APCI) mode with multiple reactions monitoring (MRM). The MRM transitions were monitored by following m/z for parent ion 384.0&daughter ion 338.0 (Felodipine), and m/z 389.1&daughter 338.1 (Felodipine D5, IS). A linear calibration plot of Felodipine was achieved in the concentration ranges of 0.105 ng/ml to 20.000 ng/ml. Mean recovery was 69.4%.This method was fully validated for specificity, precision, accuracy, reproducibility and other criteria as per regulations.

Keywords

Felodipine, Liquid liquid extraction, LCMS/MS, Human plasma, Validation

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Srinivasa Reddy ¹, Nirmala Nayak ¹, Imran Ahmed ¹, Licto Thomas ¹, Arindam Mukhopadhyay ¹, Saral Thangam ¹

Affiliations
1 Norwich Clinical Services Pvt Ltd, 147/F, 8th Main, 3rd Block, Koramangala, Bangalore 560034, IN

Abstract

Oseltamivir phosphate is licensed for the treatment of patients with influenza virus infection. Two LCMS/MS methods for simultaneous quantification of Oseltamivir and its active metabolite, oseltamivir carboxylate in human plasma were described here. After solid phase extraction sample was separated either on a reversed phase C18 column with a stepwise gradient using 0.05% formic acid and methanol or a cation-exchange column using an isocratic mobile phase (7 mM Ammonium formate, pH 3.5 ± 0.2: Methanol: 50:50, v/v). Flow rate of 1 mL/min was maintained in both cases. A triple quadrupole mass spectrometer operating in the positive ionization mode was used for detection and drug quantification. Both methods were validated over a range of 0.52ng/ml to 207.00 ng/ml for Oseltamivir and 4.08 ng/ml to 1200.00 ng/ml for Oseltamivir Carboxylate. Deuterated Oseltamivir and Oseltamivir carboxylate were used as internal standards. The accuracies and precisions for Oseltamivir were between 91-102% and 0.9 - 13.7% for all concentration levels. The accuracies and precisions for Oseltamivir carboxylate were between 88-109% and 0.5 - 8.2% at all levels. Furthermore, Oseltamivir and its metabolite were stable in plasma ex vivo for at least 191 days when stored at -20°C or below.

Keywords

Oseltamivir, Oseltamivir Carboxylate, LCMS/MS, Human Plasma, Bioequivalence Study.

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Anil Kumar Meher ¹, P. Anandan ¹, Arindam Mukhopadhyay ¹

Affiliations
1 Bioanalytical Laboratories, Lotus Clinical Research Academy Pvt. Ltd., 582, KCA Enclave, Koramangala 8th Block, Bangalore – 560 095, IN

Abstract

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.

Keywords

Isradipine, Nimodipine, LCMS/MS, Validation, Human Plasma.

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