The sample preparation of samples containing bovine serum albumin (BSA), e.g., as used in transdermal Franz diffusion cell (FDC) solutions, was evaluated using an analytical quality-by-design (QbD) approach. Traditional precipitation of BSA by adding an equal volume of organic solvent, often successfully used with conventional HPLC-PDA, was found insufficiently robust when novel fused-core HPLC and/or UPLC-MS methods were used. In this study, three factors (acetonitrile (%), formic acid (%) and boiling time (min)) were included in the experimental design to determine an optimal and more suitable sample treatment of BSA-containing FDC solutions. Using a QbD and Derringer desirability (D) approach, combining BSA loss, dilution factor and variability, we constructed an optimal working space with the edge of failure defined as D<0.9. The design space is modelled and is confirmed to have an ACN range of 83±3% and FA content of 1±0.25%. Keywords: Bovine serum albumin (BSA) solutions, Franz diffusion cell (FDC), Analytical quality-by-design (QbD), Sample preparation, Design of experiment (DOE), Derringer desirability (D)