Archive \ Volume.2 2011 Issue 4

DEVELOPMENT AND VALIDATION OF LC-MS/MS METHOD FOR THE ESTIMATION OF ACYCLOVIR IN PHARMACEUTICAL DOSAGE FORM

P. Susantakumar , Ajay Gaur , Piush Sharma
Abstract

The objective of this study was to validate a simple, specific, accurate and precise solid phase high performance liquid chromatographic method with Tandem Mass Spectrometry-Waters Quattro Premier XE method for the determination of acyclovir in human plasma using Ganciclovir as Internal Standard (IS). The precision and accuracy data have to fulfill the requirements for quantification of the analytes in biological matrices to generate data for bioequivalence, bioavailability, pharmacokinetic or toxicology investigations. A Hypersil GOLD C18, 5μ column having 4.6 x 50 mm internal diameter in binary gradient mode with flow rate was 0.5 mL/min of mobile phase containing ammonium acetate and acetonitrile were used. The chromatographic separation was achieved by using elution solution consisting of acetonitrile and waters (80:20%, v/v)], diluent solution of methanol and water (50:50%, v/v)] were monitored on a triple quadrupole mass spectrometer, operating in the multiple reaction monitoring (MRM) mode. The method was validated over the concentration range 5.0-5000.0 ng/mL, by using 500
μL plasma samples. Limit of detection and limit of quantification were found 5.0 ng and 30.0 ng respectively. The retention time for Acyclovir and Internal Standard were 1.24 min and 1.65 min respectively and overall chromatography run time was 2.24 minutes. The mean recovery of Acyclovir (89.09%) and IS (98.84%) from spiked plasma samples was consistent and reproducible. The method was validated for linearity, accuracy, precision, specificity, limit of detection, limit of quantification and robustness. The intra- and inter-day precision and accuracy values were found to be within the assay variability limits as per the FDA guidelines. The developed assay method was applied to a clinical pharmacokinetic study in human volunteers.


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