Method Development, Validation, and Sample Analysis for Regulated Quantitative Bioanalysis Using LC-MS/MS

Abstract

There are three primary stages of the regulated bioanalysis process using Liquid Chromatography–Tandem Mass Spectrometry (LC-MS/MS): method development, method validation, and sample analysis including incurred sample reanalysis (ISR). Robust and rugged LC-MS/MS methods are essential in support of drug discovery, toxicology studies, and clinical trials. The development of a robust bioanalytical method requires careful consideration of many critical parameters, such as accuracy and precision, linearity, matrix effect, sensitivity, selectivity, stability, throughput, and ruggedness (or reproducibility). Because bioanalytical data is critical for determining the safety and efficacy of a new drug, a bioanalytical method must be validated following the Food and Drug Administration’s (FDA) guidance for the industry, the recommendations from various white papers and Standard Operating Procedures (SOPs). A complete regulated method validation in biological matrix minimally requires three interday precision and accuracy runs, various short- and long-term solution and matrix stability assessments, extraction recovery, dilution capability and linearity, extract stability, reinjection reproducibility, selectivity and specificity, assessment of matrix effects, interference from concomitant medications and prodrug/metabolites, etc. The final evaluation of any high quality bioanalytical method is not complete until it passes the ultimate test of regulated sample analysis and incurred sample reanalysis (ISR) which are also conducted ­following the similar rules as validation.