The diffusion through an agarose hydrogel of the ionophoric antibiotic valinomycin dissolved in a mixture of water and ethanol is characterized by electrochemically monitoring the conductivity of a lipid bilayer membrane buried within the gel. Classical Fickian diffusion is observed, with no evidence for traps, and with a best-fit diffusion constant through the gel of 26 mum(2) s(-1) for ethanol and 2.0 mum(2) s(-1) for valinomycin. The valinomycin appears to be present in solution as large aggregates. The time response is consistent with boundary conditions in which the valinomycin enters the membrane but does not re-emerge on the other side. (C) 2003 Elsevier Science B.V. All rights reserved.