Implementation of the QbD Approach to the Analytical Method Development and Validation for the Estimation of the...

Quality by design (QbD) is a predictable quality tool that ensures that products meet predefined specifications. QbD involves understanding dependent variables, key factors, and their interactions, assessed through designed experiments analyzing specific responses. The central composite experimental design describes the interrelationships between the flow rate, buffer, and column temperature at different levels, with observed responses including retention time, theoretical plates, and tailing factor analyzed using Design-Expert software. A stability-indicating reverse-phase high-performance chromatography (RP-HPLC) technique was developed and validated for the quantitative estimation of treprostinil, assessing its stability under various forced degradation conditions. Using the optimized method, successful separation of treprostinil was achieved with an Agilent HPLC system with a photodiode array detector and an Express C₁₈ column (5 μm particle size, L × I.D. = 15 cm × 4.6 mm), maintained at 31.4 °C. The mobile phase, a buffer (0.01N KH₂PO₄) and diluent in a ratio of 36.35:63.35 (v/v), was used at a flow rate of 1.04 mL/min. Detection was performed at 276.0 nm, with treprostinil eluting at 2.579 min within a run time of 6.0 min. Treprostinil peak purity values across all degradation conditions confirmed homogeneity, establishing this method as stability-indicating. The proposed method was accurate, precise (relative standard deviation (% RSD) = 0.4%), linear, and robust (% RSD < 2%). The green analytical procedure index method yielded a total score of 83, classifying it as environmentally friendly, with a similar score obtained using the BAGI and GAPI tools. The developed AQbD-based RP-HPLC method is suitable for the assay of treprostinil in regular production batches and stability samples of treprostinil injection.