Energetic Basis for the Host-Guest Complexation Between Alendronate and β-Cyclodextrin

Bisphosphonates are widely used for the treatment of bone-related disorders; however, their low bioavailability limits therapeutic efficacy and increases long-term adverse effects. This study investigated the thermal behavior and binding energetics of the alendronate (ALEN)–β-cyclodextrin (β-CD) inclusion complex (IC) at different protonation states. Molecular dynamics simulations identified β-CD as the most suitable host for ALEN, promoting the formation of a stable 1:1 IC. Solid-state characterization by X-ray diffraction, scanning electron microscopy, and thermogravimetric analysis confirmed the successful formation of IC. Thermal analysis revealed distinct behavior compared to ALEN, β-CD, and their physical mixture, including the appearance of a single, well-defined degradation stage. Kinetic studies further supported these results, showing that the IC exhibited the highest activation energy. The ALEN–β-CD interaction in aqueous solution was evaluated by isothermal titration calorimetry, which demonstrated the spontaneous formation of a 1:1 complex over the physiologically relevant pH range (4.0–8.0). Binding was entropically driven at pH 5.3, 7.4, and 8.0, whereas at pH 4.0 the enthalpic contribution dominated. To the best of our knowledge, this work presents for the first time the microscopic binding constants ( K _b ) for the ALEN–β‑CD inclusion complex. MTT assays demonstrated that complexation significantly reduces the cytotoxic profile of ALEN. Overall, these findings confirm the formation of the IC in both the solid and solution states and provide valuable insights for the development of pharmaceutical formulations to improve the safety of long-term bisphosphonate therapy.