Electron spin echo envelope modulation spectroscopy of radical pairs in photosynthetic bacteria
Electron spin echo envelope modulation (ESEEM) spectroscopy is widely used to study the radical pairs created during the primary steps of photosynthesis. In this thesis the analysis of ESEEM spectra is improved, and some new applications and variations of this experiment suggested. Experimental spectra from species such as P+Q-A, the secondary radical pair formed in the reaction centre of the bacterium Rhodobacter sphaeroides, give information about the exchange and dipolar couplings between the radicals. The model used to analyse the data affects the results; this thesis suggests two improvements. First, the effect of anisotropic hyperfine couplings in the radicals is considered by the addition of a single spin-1/2 nucleus to the model. This approach suggests that previous models neglecting the effect of nuclei may have been slightly in error. Secondly, several model fittings are performed in the time domain. This approach avoids the Fourier transformation to the frequency domain so that experimental dead-time does not corrupt the data. An excellent fit to experimental data is found with a model containing one spin-1/2 nucleus on each radical. The hyperfine coupling parameters resulting from the fit are consistent with independent experimental results. Use is made of the method of Cramér-Rao lower bounds to assess the precision to which experimental parameters are determined from a time domain curve fitting. It is shown that the lower bounds may also be used to determine the optimum sampling strategy for the experiment. An example is given of the novel use of ESEEM to determine the distance between the radicals in the strongly coupled, uncorrelated radical pair Q-AQ-B ESEEM has not yet been used for this purpose, and the simulated spectra produced here indicate that the experiment could be used to evaluate the dipolar coupling and hence the inter-radical distance. This thesis considers the possibility of performing ESEEM at higher frequencies than are usually considered. Calculations show that the increased resolution of the g-tensors allow an experiment performed at the W-band frequency of 95 GHz to make a correlation between the relative orientations of the radicals and the dipolar axis, information which has previously been unavailable from a single experiment.