Picosecond time-gated fluorescence imaging of Photosystem II particles and spinach leaves

Abstract

The distribution of chlorophylls and other molecular components in whole leaf tissue and the aggregation state of Photosystem II (PS II) particles were studied using picosecond time-gated fluorescence microscopy of PS II core complexes, BBY particles and whole spinach leaves. In addition, the effects of UV-B on these and other properties of these samples where characterized for precisely the same regions of these samples. Typically, spatial resolutions ~ 2 mm and temporal resolutions of 100 ps for 5 ns scans were used. For leaf material, the initial fluorescence (0 ps) shows localized and well-defined cellular structure in the case of the control but relatively diffuse and non-specific structures in the UV-irradiated case indicating significant UV-induced rupture of the cellular structure. The total initial fluorescence amplitude integated over the whole image is similar for the control and irradiated cases because, although the fluorescence decay times differ, the net chlorophyll emission cross-section is not substantially altered by this UV treatment. Structural changes on UV irradiation are most evident in the images for later times (> 1 ns) and in the fluorescence decay times and amplitudes which indicate faster fluorescence decay in the UV-irradiated case. A slow risetime component, which we assign to carotenoid, appears in images and decay curves for leaf samples. The changes in the chlorophyll fluorescence decay kinetics indicate UV-induced damage by uncoupling of chlorophyll molecules in the light-harvesting system. This uncoupling adversely effects the energy reorganization and transfer in the antennae, and subsequent exciton transfer to the PS II reaction centre.