On the analyses of fluorescence depolarisation data in the presence of electronic energy migration. Part I: Theory and general description.

A new and general procedure is described for a detailed analysis of time-resolved fluorescence depolarisation data in the presence of electronic energy migration. An isotropic ensemble of bifluorophoric molecules (D(1)-R-D(2)) has been studied to demonstrate its utility. Intramolecular donor-donor energy migration occurs between the two donor groups (D), which are covalently connected to a rigid linker group (R). These groups undergo restricted reorientational motions with respect to the R group. The analysis of depolarisation data basically involves the search for best-fit parameters which describe the local reorientational motions, the intermolecular D(1)-D(2) distance, as well as the mutual orientations of the donors. For this, the analysis is partly performed on the Fourier domain and the best-fit parameters are determined by using an approach based on a Genetic Algorithm. The energy migration process has been described by using Monte Carlo simulations and an extended Förster theory (EFT). It is found that the EFT provides the least time-consuming computational method. Since one-photon and two-photon excited fluorescence experiments can be applied for energy migration studies, a general and unified theoretical formulation is given.