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 (D1-R-D2) 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 reorienting motions, the interfluorophore D1-D2 distance, as well as the mutual orientations of the donors. For this, the analysis is partly performed in 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. It is found that this theory 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. To exemplify the developed quantitative approach the depolarisation of the fluorescence in the presence of electronic energy migration within a bis-(9-anthrylmethylphosphonate) bisteroid molecule has been studied by time-resolved two-photon excited fluorescence depolarisation experiments. To solely obtain information about local reorientations of the 9-anthrylmethyl group, also the mono-(9-anthrylmethylphosphonate) bisteroid was studied, which enabled modelling of the ordering potential of the donor. Values of the two-photon absorption tensor components were obtained. To describe the discrepancy between the measured values of the initial anisotropy and fundamental anisotropy predicted by theory the distribution of absorption tensor caused by fast processes have been introduced. An angular parameter of absorption tensor was determined. Reasonable values of the distance between the 9-anthrylmethyl groups, as well as for their mutual orientation were obtained.

An extended Förster theory (EFT), previously derived (L. B.-Å. Johansson et al. J. Chem. Phys., 1996,105) has theoretically been adapted and used in simulations of donor-acceptor energy transfer (DAET), which is a process often referred to as FRET. It was shown that the classical Förster theory is only valid in the initial part of the fluorescence decay. In this thesis an EFT is derived and outlined for electronic energy transport between two fluorescent molecules which are chemically identical, but photophysically non-identical. The energy migration within such asymmetric pairs is partially reversible and therefore referred to as partial donor-donor energy migration (PDDEM). The previously derived model of PDDEM (S. V. Kalinin et al. Spectrochim Acta Part A, 2002,58) is an approximation of the EFT. In particular, the EFT accounts for the time-dependent reorientations as well as the distance that influence the rate of electronic energy migration. The reorientation of the fluorophores transition dipole moments has been simulated using Brownian dynamics. As a result, the related “k2-problem” has been solved. The EFT of PDDEM has also been studied regarding the effect of PDDEM on experimental observables e.g. quantum yield of fluorescence and steady-state anisotropies

<p>We have studied fundamental aspects of time-resolved two-photon excited fluorescence depolarisation. The thesis presents experimental as well as theoretical progress. We show that a multi-photon induced instrumental response function obtained from a suspension of gold nanoparticles is appropriate for the analysis of two-photon excited fluorescence decays obtained using time-correlated single photon counting detection. Theoretical expressions have been derived for the fluorescence anisotropy decay obtained upon two-photon excitation of various molecular systems in liquid solutions: a) an anisotropic rigid rotor that undergoes rotational diffusion in the presence of ultrafast unresolved restricted reorientations, e.g. librations. b) a molecular group covalently attached to a stationary macromolecule, and undergoing local reorientation in a uniaxial ordering potential. A new approach to the analysis of two-photon excited fluorescence depolarisation experiments was developed, which combines data obtained by using linearly and circularly polarised excitation light, in a global manner. In the analysis, knowledge about unresolved reorientations was obtained from one-photon excitation studies of the corresponding systems. By means of this procedure it has been possible to obtain quantitative information about the molecular two-photon absorption tensor for perylene and two of its derivatives. Thereby the symmetry of the final excited and intermediate vibronic states could be assigned. The analysis reveals that the two-photon transition studied with the 800 nm laser exhibits mixed character. An important finding from the experiments was that the two-photon absorption tensor appears to be solvent dependent. Furthermore, the thesis presents the first theoretical treatment of two-photon excited donor-donor energy migration in the presence of molecular reorientation and which applies the extended Förster theory. Explicit expressions for molecules that belong to the point groups D_2h, D₂ and C_2v are given. Preliminary experiments are finally also reported on a two-photon excited donor-donor energy migration system consisting of a bisanthryl-bisteroid. </p>

We have studied fundamental aspects of time-resolved two-photon excited fluorescence depolarisation. The thesis presents experimental as well as theoretical progress. We show that a multi-photon induced instrumental response function obtained from a suspension of gold nanoparticles is appropriate for the analysis of two-photon excited fluorescence decays obtained using time-correlated single photon counting detection. Theoretical expressions have been derived for the fluorescence anisotropy decay obtained upon two-photon excitation of various molecular systems in liquid solutions: a) an anisotropic rigid rotor that undergoes rotational diffusion in the presence of ultrafast unresolved restricted reorientations, e.g. librations. b) a molecular group covalently attached to a stationary macromolecule, and undergoing local reorientation in a uniaxial ordering potential. A new approach to the analysis of two-photon excited fluorescence depolarisation experiments was developed, which combines data obtained by using linearly and circularly polarised excitation light, in a global manner. In the analysis, knowledge about unresolved reorientations was obtained from one-photon excitation studies of the corresponding systems. By means of this procedure it has been possible to obtain quantitative information about the molecular two-photon absorption tensor for perylene and two of its derivatives. Thereby the symmetry of the final excited and intermediate vibronic states could be assigned. The analysis reveals that the two-photon transition studied with the 800 nm laser exhibits mixed character. An important finding from the experiments was that the two-photon absorption tensor appears to be solvent dependent. Furthermore, the thesis presents the first theoretical treatment of two-photon excited donor-donor energy migration in the presence of molecular reorientation and which applies the extended Förster theory. Explicit expressions for molecules that belong to the point groups D2h, D2 and C2v are given. Preliminary experiments are finally also reported on a two-photon excited donor-donor energy migration system consisting of a bisanthryl-bisteroid.