Oxygen evolution by Photosystem II: The contribution of backward transitions to the anomalous behaviour of double-hits revealed by a new analysis method.

Backward transitions in the analysis of oxygen production under flashing light were introduced by Packham et al., 1988, Photosynth. Res. 15: 221-232. In order to take backward transitions into account, a new method of analysis is presented: the 'eigenvalue method'. This method is based on the recurrence relation of oxygen production with four coefficients (also known as the four 'sigma' coefficients). It shows less susceptibility to round-off errors than other methods and permits the computation of double-hits directly from the coefficients, which was not possible before. With it we discovered that the inconsistent behaviour of double-hits observed previously under low flash intensities or low flash frequencies was mainly due to the inclusion of the backward transitions into the double-hit probability. In these conditions backward transitions seemed to be due either to the combination of an S-state deactivation and a miss, or to two S-state deactivations and a single-hit.In the presence of 3-(3, 4-Dichlorophenyl)-1, 1-dimethylurea (DCMU), the previous methods of 'sigma' analysis failed. In contrast, the new method resolved all four S-state probabilities; thus it has the further advantage of being more 'robust' (robustness being defined as the ability to yield a meaningful answer under difficult conditions).