Drug delivery from water to lipid bilayer membranes is crucial as a primary step of bioactivities in the cell. To gain insight into molecular mechanisms of drug deliveries, we have developed the method to monitor dynamic properties of drugs and lipid components in membranes, by applying high-resolution solution NMR combined with the pulsedfield- gradient (PFG) technique in a noninvasive manner. The PFG method unveiled the bound component after the preferential decay of the free component at the high field gradient, where the chemical shift difference between these components was not enough to distinguish from each other. Using phospholipid vesicles as model cell membranes, we quantified the diffusivity, the kinetics of membrane binding, and thermodynamic stability of small-sized drugs in relation to the temperature. Cell membrane permeability was also discussed by real-time in-cell NMR spectroscopy in combination with isothermal titration calorimetry of the model system. Finally, the dynamical features in lipid membranes, as platform of drug transport, were characterized by temperature dependence of NMR nuclear overhauser effect (NOE) in cell-sized giant vesicles, to demonstrate large fluctuation of lipids in the vertical direction to the membrane surface related to drug delivery phenomena.
Keywords:NMR, drug delivery, thermodynamic stability, cell-penetrating peptide, ITC.