Dioxygen uptake by isolated thylakoids from lettuce (Lactuca sativa L.): simultaneous measurements of dioxygen uptake, pH change of the medium and chlorophyll fluorescence parameters

The setup has been elaborated for the simultaneous measurements of dioxygen uptake, pH changes, and chlorophyll a fluorescence parameters of an isolated thylakoid suspension. Using this equipment we have found at least three kinetically distinguishable components in the response of dioxygen uptake and pH increase to light intensity in the range of 0–1600 μE m−2 s−1. The pH changes were not observed in the presence of uncouplers (2 μM valinomycin plus 2 μM nigericin) while O2 uptake increased by about 10% and Fv/Fm ratio appeared to be unaffected by this treatment. Treatment with DNP-INT, an inhibitor of plastoquinol oxidation, led to a significant reduction of pH increase and O2 consumption whereas Fv/Fm was impaired only to 71% of the control. Incubation with catalase (580 U/ml) caused a total inhibition of oxygen uptake, while the pH increased and the Fv/Fm ratio decreased to about 60% and 85% of the control, respectively. The addition of catalase after the irradiation period led to an evolution of the same amount of dioxygen as was consumed during the light period. These results show that hydrogen peroxide was formed in the investigated system and accumulated during illumination. On the basis of the obtained data, three sites of dioxygen reduction within isolated thylakoid membranes and the dependence of dioxygen uptake on the photosystem activities were discussed.