KINETICS OF THE OXYGEN EVOLUTION STEP IN PLANTS DETERMINED FROM FLASH-INDUCED CHLOROPHYLL-A FLUORESCENCE

Photosystem II (PS II) of plants and cyanobacteria, :which catalyzes t he light-induced splitting of water and the release of oxygen, is the primary source of oxygen in the earth atmosphere. When activated by sh ort light flashes, oxygen release in PS II occurs periodically with ma xima after the third and the seventh flashes. Many other processes, in cluding chlorophyll (Chi) a fluorescence, are also modulated with peri od of four, reflecting their sensitivity to the activity of Photosyste m II. A new approach has teen developed for the analysis of the flash- induced fluorescence of Chl a in plants, which is based on the use of the generalized Stern-Volmer equation for multiple quenchers. When app lied to spinach thylakoids, this analysis reveals the presence of a ne w quencher of fluorescence whose amplitude is characterized by a perio dicity of four with maxima after the third and the seventh flashes, in phase with oxygen release. The quencher appears with a delay of appro ximate to 0.5 ms followed by a rise time of 1.2-2 ms at pH 7, also in agreement with the expected time for oxygen evolution. It is concluded that the quencher is a product of the reaction leading to the oxygen evolution in PS II. The same quenching activity, maximal after the thi rd flash, could be seen in dark adapted leaves, and provides the first fully time-resolved measurement of the kinetics of the oxygen evoluti on step in the leaf. Thus, the non-invasive probe of Chl a fluorescenc e provides a new and sensitive method for measuring the kinetics of ox ygen evolution with potential for use in plants and cyanobacteria in v ivo.