Palladium(II) complexes with tris(2-carboxyethyl)phosphine (PdTCEP) show promise for biomedical applications due to their distinct chemical characteristics. This study explored the toxicity of PdTCEP towards normal human cells and examined its interactions with model cell membranes. Two cell types were used to evaluate cytotoxicity: human microvascular endothelial cells (HMEC-1) and red blood cells (RBCs). In HMEC-1 cells, PdTCEP reduced survival to about 80% at 15 µM, with the most significant drop-down to 40%-occurring at 40 µM. The production of reactive oxygen species (ROS) increased in a manner dependent on both dose and time, especially after 72 h of incubation. Despite these effects, PdTCEP caused only minor hemolysis in RBCs, with hemolysis levels staying below 10% even at higher concentrations. Fluorescence anisotropy measurements showed that PdTCEP minimally affects the hydrophobic core of the lipid bilayer, with slight changes observed at concentrations above 40 µM. Generalized polarization (GP) analysis indicated a slight decrease in lipid polar head packing with increasing PdTCEP concentration. Complementary FTIR analysis supported these findings by providing detailed insights into PdTCEP-membrane interactions. This research underscores PdTCEP's selective cytotoxicity and structural effects on membranes, suggesting its promise for more in-depth biological and pharmacological studies.