chlorophyll fluorescence; single-turnover saturating flashes; PSII core complexes; P680

Recent chlorophyll-a fluorescence yield measurements, using single-turnover saturating flashes (STSFs), have revealed the involvement of a rate-limiting step in the reactions following the charge separation induced by the first flash. As also shown here, in diuron-inhibited PSII core complexes isolated from Thermosynechococcus vulcanus the fluorescence maximum could only be reached by a train of STSFs. In order to elucidate the origin of the fluorescence yield increments in STSF series, we performed transient absorption measurements at 819nm, reflecting the photooxidation and re-reduction kinetics of the primary electron donor P680. Upon single flash excitation of the dark-adapted sample, the decay kinetics could be described with lifetimes of 17ns (approximate to 50%) and 167ns (approximate to 30%), and a longer-lived component (approximate to 20%). This kinetics are attributed to re-reduction of P680(center dot+) by the donor side of PSII. In contrast, upon second-flash (with t between 5s and 100ms) or repetitive excitation, the 819nm absorption changes decayed with lifetimes of about 2ns (approximate to 60%) and 10ns (approximate to 30%), attributed to recombination of the primary radical pair P680(center dot+)Pheo(center dot-), and a small longer-lived component (approximate to 10%). These data confirm that only the first STSF is capable of generating stable charge separation-leading to the reduction of Q(A); and thus, the fluorescence yield increments elicited by the consecutive flashes must have a different physical origin. Our double-flash experiments indicate that the rate-limiting steps, detected by chlorophyll-a fluorescence, are not correlated with the turnover of P680.