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

Abstract

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-amino-acridine fluorescence quenching method or the [¹⁴C]methylamine uptake method. The ΔpH values obtained with the two methods are compared and a linear relationship is demonstrated in the ΔpH 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 ΔpH determination after calibration with other methods.