Bisphosphonate protonation states, conformations, and dynamics on bone mineral probed by solid-state NMR without isotope enrichment.

Recognition of bone mineral by bisphosphonates is crucial to their targeting, efficacy, therapeutic and diagnostic applications, and pharmacokinetics. In a search for rapid and simple NMR approaches to assessing the bone recognition characteristics of bisphosphonates, we have studied alendronate, pamidronate, neridronate, zoledronate and tiludronate, in crystalline form and bound to the surface of pure bone mineral stripped of its organic matrix by a simple chemical process. (31)P NMR chemical shift anisotropies and asymmetries in the crystalline compounds cluster strongly into groupings corresponding to fully protonated, monoprotonated, and deprotonated phosphonate states. All the mineral-bound bisphosphonates cluster in the same anisotropy-asymmetry space as the deprotonated phosphonates. In (13)C{(31)P} rotational echo double resonance (REDOR) experiments, which are sensitive to carbon-phosphorus interatomic distances, the strongly mineral-bound alendronate displays very similar conformational and side chain dynamics to its crystalline state. Pamidronate and neridronate, with shorter and longer sidechains, respectively, and generally weaker mineral binding, display more dynamical sidechains in the mineral-bound state. The REDOR experiment provides a simple rationalization of bisphosphonate-mineral affinity in terms of molecular structure and dynamics, consistent with findings from much more labour- and time-intensive isotope labelling approaches.