On the estimation of proton gradient and osmotic volume in chloroplast membranes

The dependence of thylakoid osmotic volume on NH4Cl uncoupling and on phosphorylation substrates is determined by the centrifuge filtration method. The values obtained are used to evaluate the transmembrane proton gradient in conjunction with either the 9-aminoacridine fluorescence quenching method or the [14C]methylamine uptake method. The DeltapH values obtained with the two methods are compared and a linear relationship is demonstrated in the DeltapH range from 1.4 to 2.7 ([14C]methylamine values). Different linear relationships are obtained depending on the presence or absence of electron acceptor. We conclude that the 9-aminoacridine method can be used for DeltapH determination after calibration with other methods.