Nitric Oxide Diffusion in Membranes Determined by Fluorescence Quenching

Quenching of pyrene derivative fluorescence by nitric oxide was used to evaluate the apparent diffusion coefficients of nitric oxide in artificial and biological membranes. The apparent second-order quenching constants of nitric oxide were obtained from Stern–Volmer plots using methyl- and undecylpyrene derivatives incorporated into liposomes and erythrocyte plasma membranes in order to assess the ability of nitric oxide to interact with molecules located at different positions in the membrane. Diffusion coefficients were estimated from the determined second-order quenching constants and compared to that of oxygen obtained under the same conditions. Oxygen and nitric oxide presented similar diffusional behavior in agreement with their similarity in structures, with the differences observed attributable to the higher lipophilicity of oxygen compared to nitric oxide. In solution, both showed the same quenching efficiency while in liposomes and erythrocyte ghosts oxygen diffusion was twice that of nitric oxide (kO2/kNO= 2). Nitric oxide diffusion coefficients determined at 20°C ranged from 1.3 × 10−5cm2s−1in liposomes to 0.4 × 10−5cm2s−1in surface erythrocyte plasma membranes. Both nitric oxide and oxygen had larger quenching constants for the undecyl derivative compared to the methylpyrene compound incorporated into erythrocyte plasma membranes, indicating an increased solubility of both gases toward the center of the membrane.