Abstract

Bioanalytical techniques are being widely applied for quantitative estimation of xenobiotics and biotics in biological matrices such as blood, serum, plasma, proteins or urine. They are crucial for supporting new drug applications or biologics license applications. Liquid chromatography-tandem mass spectrometry has become an important tool in pharmaceutical industry as it offers reduced analysis times, improved selectivity and increased throughput in drug bioanalysis. In the present work attempt has been made to develop a novel bioanalytical method for estimation of antihypertensive drug Ramipril and its metabolite Ramiprilat in the plasma samples, by a hyphenated technique which includes liquid chromatography combined with mass spectrometry, using Enalapril and Enalaprilat as internal standards. The developed method was validated as per international council for harmonization guidelines for selectivity, specificity, matrix effect, calibration curve, range, accuracy, precision, dilution integrity and stability etc. It exhibited limit of quantification of 1.09 ng/ml for Ramipril and 1.08 ng/ml for Ramiprilat. The analytes, Ramipril and Ramiprilat were extracted from plasma by liquid chromatography-tandem mass spectrometry using solvent mixtures comprising of acetonitrile, methanol and 0.2 % trifluoro acetic acid as mobile phase and Chromolith speed rod RP 18e gold (50×4.6) column as stationary phase. The validated parameters were within the acceptance criteria as per the regulatory guidelines and the validated calibration curve exhibited r² value greater than or equal to 0.98 with high recovery. Hence it can be concluded that the developed method was specific, accurate, sensitive, and reliable to quantify Ramipril and its metabolite Ramiprilat in biological samples and can be potentially applied for Pharmacokinetic and bioequivalence studies.