Light-induced changes of NADPH fluorescence in isolated chloroplasts: a spectral and fluorescence lifetime study

Isolated chloroplasts show a light-induced reversible increase in blue-gree n fluorescence (BGF), which is only dependent on NADPH changes. In the pres ent communication, we report a time-resolved and spectral analysis of this BGF in reconstituted chloroplasts and intact isolated chloroplasts, in the dark and under actinic illumination. From these measurements we deduced the contribution of the different forms of NADPH (free and bound to proteins) to the light-induced variation of BGF and conclude that this variation is d ue only to the redox change of the NADP pool. A simple model estimating the distribution of NADPH between the free and bound form was designed, that e xplains the differences measured for the BGF of reconstituted chloroplasts and intact chloroplasts. From the decay-associated spectra of the chloropla st BGF, we also deduced the participation of flavins to the green peak of c hloroplast fluorescence emission spectrum, and the existence of excitation energy transfer from proteins to bound NADPH in chloroplasts. In addition, we reexamined the use of chloroplast BGF as a quantitative measure of NADPH concentration, and confirmed that chloroplast BGF can be used for non-dest ructive, continuous and probably quantitative monitoring of light-induced c hanges in NADP redox state. (C) 2000 Elsevier Science B.V. All rights reser ved.