A quality by design (QbD) strategy for developing and validating Droxidopa in API and formulation using an RP-HPLC procedure ensuring regulatory compliance

Background: Droxidopa, a prodrug for neurogenic orthostatic hypotension, requires precise quality analysis. RP-HPLC is valued for its sensitivity and reliability in pharmaceutical quality control, but traditional methods are time-consuming and lack robustness, highlighting the need for systematic approaches. Method: This study innovatively applies the Quality by Design (QbD) systematic approach to establish and validate an RP-HPLC procedure for analyzing Droxidopa in active pharmaceutical ingredient (API) and pharmaceutical dosage form. Emphasizing risk assessment and management, the procedure was systematically designed employing Plackett-Burman design (PBD) for initial screening studies, followed by Central Composite Design (CCD) for optimization. Critical method parameters (CMPs) such as buffer pH, % organic composition, and organic modifier were precisely adjusted. Statistical tools, including ANOVA, evaluated the significance and interactions of factors on responses like theoretical plates, retention time, and tailing factor. Results: Using Design Expert version 13, the procedure was optimised for an Agilent C18 packed column (150 mm x 4.6 mm i.d. and 3.5 mu size particles). The eluent comprising Acetonitrile and 10mM hexane sulphonic acid buffer (pH 3 adjusted with 0.1% orthophosphoric acid) in 35:65 v/v ratio, achieved a 1.0 ml/min flow rate and a 2.66 min retention time. The optimised procedure was validated, and stress studies were conducted per ICHQ2(R1) and ICHQ1A guidelines. The procedure exhibited high sensitivity and precision, with a linearity range of 25-150 mu g/ml (R2 = 0.9999), theoretical plates of 4226, and 1.07 as a tailing factor. Precision was validated within one day and between days with RSD values of 0.36 and 0.77, respectively, and robustness values below 2%. Forced degradation studies confirmed the ability of a procedure to distinguish Droxidopa from degradation products. Conclusion: This research marks a significant advancement in RP-HPLC method development for pharmaceutical analysis, offering a robust, and sensitive, procedure for Droxidopa analysis in bulk medication and formulations.