Time-resolved chlorophyll fluorescence studies of photosynthetic membranes: Resolution and characterisation of four kinetic components

Room temperature single-photon timing measurements on intact green algae and higher plant thylakoids at low excitation energies have been analysed using a four-exponential kinetic model. Closing the Photosystem II (PS II) reaction centres produced two variable lifetime components (0.25-1.4 ns and 0.45-2.6 ns) and two constant components (50 and 220 ps) between the initial (F₀) and maximal (F_M) fluorescence levels. The yield of the variable components paralleled the changes in their lifetimes, while the yields of the constant emissions remained insensitive to the state of PS II. These observations indicate the presence of well-connected PS II centres favouring energy transfer between each other. Time-resolved emission spectra, at F₀ and F_M, show that the three 'slow' components have fluorescence maxima at 685 nm, while the fast decay has a maximum emission between 695 and 705 nm. Kinetic and spectral analyses of decays from a Chlamydomonas mutant and PS II-enriched membranes lacking Photosystem I (PS I) showed the absence of the fast, red-shifted component and only required a triexponential model to fit the decays. At F_M the three components retrieved from the PS II-enriched thylakoids were identical to the PS II-associated emissions of intact thylakoids. Such observations indicate the requirement for at least four kinetic components to describe the fluorescence decays of intact light-harvesting Chl a b protein complex (LHC) containing photosynthetic material best. Furthermore, these data cannot be interpreted in terms of the α β heterogeneity of PS II.