A simple, rapid and reproducible HPLC method has been developed and validated for the quantification of albendazole sulfoxide (ALBSOX) and praziquantel (PRZ), which can coexist in various dosage forms. Chromatographic separation was performed in gradient mode using a mobile phase consisting of an InertSustain® C18 column (150 × 4.6 mm, 5 µm) and acetonitrile: water (v/v) at a flow rate of 1.0 mL/min. The active substance peaks were well separated and detected by a DAD detector at 217 nm. The HPLC method was linear for ALBSOX and PRZ in the concentration range of 0.1-50 µg/mL. Limit of detection (LOD) was found to be 0.01 µg/mL for ALBSOX and 0.009 µg/mL for PRZ. The limit of quantification (LOQ) was found to be 0.03 µg/mL for ALBSOX and 0.027 µg/mL for PRZ. The developed HPLC method was validated based on ICH guidelines for specificity, linearity, system suitability, precision and accuracy. The method was applied for simultaneous quantification and characterization studies of ALBSOX and PRA in PLGA nanoparticles.