Influence of diffusion on excitation energy transfer in solutions by gigahertz harmonic content frequency-domain fluorometry

We report the effects of diffusion on fluorescence resonance energy transfer, as observed by using the frequency-domain technique. Energy transfer between indole (donor) and dansylamide (acceptor) was studied in low and high viscosity solutions. At the current level of resolution, the donor fluorescence decays were satisfactorily analyzed in terms of the theoretical function including translational diffusion proposed by Gosele et al. (Chem. Phys. Lett. 1975, 24, 519). The Forster equation, which does not consider diffusion, was found to significantly overestimate the critical transfer distance (R₀) in fluid solutions and to provide an inadequate representation of the data. These results demonstrate the ability to determine mutual diffusion coefficients from the frequency response of the donor emission.