Excitation transfer and quenching in photosystem II, enlightened by carotenoid triplet state in leaves

Accumulation of carotenoid (Car) triplet states was investigated by singlet-triplet annihilation, measured as chlorophyll (Chl) fluorescence quenching in sunflower and lettuce leaves. The leaves were illuminated by Xe flashes of 4 μs length at half-height and 525-565 or 410-490 nm spectral band, maximum intensity 2 mol quanta m^-2 s^-1, flash photon dose up to 10 μmol m^-2 or 4-10 PSII excitations. Superimposed upon the non-photochemically unquenched F_md state, fluorescence was strongly quenched near the flash maximum (minimum yield F_e), but returned to the F_md level after 30-50 μs. The fraction of PSII containing a ³Car in equilibrium with singlet excitation was calculated as T_e = (F_md-F_e)/F_md. Light dependence of T_e was a rectangular hyperbola, whose initial slope and plateau were determined by the quantum yields of triplet formation and annihilation and by the triplet lifetime. The intrinsic lifetime was 9 μs, but it was strongly shortened by the presence of O₂. The triplet yield was 0.66 without nonphotochemical quenching (NPQ) but approached zero when NP-Quenched fluorescence approached 0.2 F_md. The results show that in the F_md state a light-adapted charge-separated PSII_L state is formed (Sipka et al., The Plant Cell 33:1286-1302, 2021) in which Pheo^-P680⁺ radical pair formation is hindered, and excitation is terminated in the antenna by ³Car formation. The results confirm that there is no excitonic connectivity between PSII units. In the PSII_L state each PSII is individually turned into the NPQ state, where excess excitation is quenched in the antenna without ³Car formation.