A Scientific, General Approach to Robustness Validation for an Ion-Pair LC Assay Method Through a Case Study

Abstract

This paper describes a scientifically sound, systematic approach to the robustness validation of an ion-pair LC assay method. Robustness validation was carried out on all chromatographic parameters, such as ion-pair reagent concentration, buffer concentration and pH, organic to buffer ratio, column oven temperature, and column age. Small, deliberate variations to each parameter were implemented according to reasonably expected laboratory variabilities. For instance, a typical measuring variability with a 2,000-mL graduated cylinder is 20 mL, and this translates into variations of ±1.3% for a volume of 1,560 mL of an aqueous phase. Details on the experimental design of robustness validation are described, such as the parameters considered, the variations chosen for each parameter along with rationales, the probes that were used to assess robustness, and corroboration of method robustness with intermediate precision validation. This robustness validation approach eliminates guess work, ensures scientifically appropriate and customized variations for each method parameter, and has won regulatory acceptance.