Doxorubicin, a potent antineoplastic agent, is widely utilizedin the treatment of various malignancies, including breast can-cer, malignant lymphomas, and soft tissue sarcomas. This studyinvestigates the interaction between doxorubicin and humanserum albumin (HSA) under physiological conditions using acombination of biophysical techniques, including circular dichro-ism (CD), Fourier-transform infrared spectroscopy (FT-IR), equilib-rium dialysis, molecular docking, and UV–visible (vis) absorptionspectroscopy. The results indicate that doxorubicin quenchesthe intrinsic fluorescence of HSA via a static quenching mecha-nism. Binding constants (Kb), Stern–Volmer quenching constants(Ksv), the number of binding sites (n), and thermodynamicparameters were determined at multiple temperatures. Dataanalysis revealed cooperative binding, wherein the binding ofone doxorubicin molecule enhances the affinity for subsequentmolecules. Molecular docking studies further demonstrated thatdoxorubicin interacts primarily with the IIA, IIIA, and IIB sub-domains of HSA, inducing subtle conformational changes. Bothfluorescence spectroscopy and molecular docking analyses sug-gest that hydrophobic interactions are a dominant force in thedoxorubicin-HSA binding process
