Development of HPLC Method for Simultaneous Quantification of Doxorubicin and Baicalein Encapsulated in Unilamellar Liposomes

ABSTRACT Dual drug delivery systems have transformed breast cancer therapy by enhancing efficacy and reducing side effects. While Doxorubicin (DOX) is highly effective against breast cancer, its clinical utility is limited by substantial adverse effects. Co‐delivery with Baicalein (BCL) in a liposomal formulation offers a promising strategy to mitigate these effects while improving therapeutic outcomes. However, a reliable analytical method is essential for accurate quantification of both drugs in complex formulations. A robust and validated high‐performance liquid chromatography (HPLC) method was developed to simultaneously facilitate DOX and BCL quantification. The method exhibited high sensitivity, with a limit of quantification below 0.3 µg/mL and excellent linearity over a range of 0.0195 to 10 µg/mL. Liposomal formulation co‐loaded with DOX and BCL was prepared using the thin‐film hydration method to validate the method's applicability. The developed HPLC method successfully estimated the entrapment efficiency of both drugs. In addition, in‐vitro drug release studies confirmed the stability of liposomes in acidic conditions and demonstrated sustained drug release in the intestine. This validated method, compliant with International Council for Harmonization guidelines, provides a reliable tool for analyzing and optimizing dual drug delivery systems, paving the way for advanced therapies in breast cancer treatment.