The kinetics of ampicillin complexation by γ-cyclodextrins. A single molecule approach

Efficient measuring of binding thermodynamics of interactions between cyclodextrins (CDs) and antibiotics remains a challenge in the fields of physical chemistry and pharmaceutics. Here we report on a single-molecule investigation of pH- and voltage-dependent reversible interactions between ampicillin and γ-CDs, through monitoring of ionic current signatures across an α-hemolysin (α-HL) protein entrapping a γ-CD molecule. Our data reveal that electric and electro-osmotic driving forces alter the reversible reaction rates of ampicillin interaction with γ-CDs, as well as free energy changes accompanying the interaction. We found that close to neutral pH values facilitate more unstable γ-CD-ampicillin complexes, as well as a decreased affinity of the γ-CD-ampicillin reversible interaction, as compared to acidic pH. We posit that a pH-dependent partial electric charge on the ampicillin molecule and anionic selectivity of the α-HL·γCD complex account for the antibiotic and γ-CD intracavity manifestations. This approach may provide unique alternatives for the characterization of CD-guest interactions, useful for pharmaceutical formulations and tunable drug delivery systems.