Delayed fluorescence from Rhodopseudomonas viridis following single flashes

Delayed fluorescence from Rhodopseudomonas viridis membrane fragments has been studies using a phosphoroscope employing single, short actinic flashes, under conditions of controlled redox potential and temperature. The emission spectrum shows that delayed fluorescence is emitted by the bulk, antenna bacteriochlorophyll. The energy for delayed fluorescence, however, must be stored in a reaction-center complex including the photooxidized form (P+) of the primary electron-donor (P) and the photoreduced form (X MINUS) of the primary electron-acceptor. This is shown by the following observations: (1) Delayed luminescence is quenched (a) at low redox potentials which allow cytochromes to reduce P+ rapidly after the flash, (b) at higher redox potentials which, by oxidizing P chemically, prevent the photochemical formation of P+X minus, and (c) upon transfer of an electron from X minus to a secondary acceptor, Y. (2) Under conditions that prevent the reduction of P+ by cytochromes and the oxidation of X minus by Y, the decay kinetics of delayed fluorescence are identical with those of P+X minus, as measured from optical absorbance changes. The main decay route for P+X minus under these conditions has a rate-constant of approximately 10-3-s-minus 1. In contrase, a comparison of the intensities of delayed and prompt fluorescence indicates that the process in which P+X minus returns energy to the bulk bacteriochlorophyll has a rate-constant of 3.7 s-minus 1, at 295 degrees K and pH 7.8. The decay kinetics of P+X minus and delayed fluorescence change little with temperature, whereas the intensity of delayed fluorescence increases with increasing temperature, having an activation energy of 12.5 kcal mol-mol- minus 1. We conclude that the main decay route involves tunneling of an electron from X minus to P+, without the promotion of P to an excited state. Delayed fluorescence requires such a promotion, followed by transfer of energy to the bulk bacteriochlorophyll, and this combination of events is rare. The activation energy, taken with potentiometric data, indicates that the photochemical conversion of PX to P+X minus results in increases of both the energy and the entropy of the system, by 16.6 kcal-mol- minus 1 and 8.8 cal-mol- minus 1-deg- minus 1. The intensity of delayed fluorescence depends strongly on the pH; the origin of this effect remains unclear.