A simple colorimetric determination of the manganese content in photosynthetic membranes

The functional Mn content of intact photosystem II membrane fragments was measured as 4.06 +/- A 0.13 Mn/reaction center when determined using a simple, sensitive colorimetric assay that will also work with thylakoids and core complexes. This procedure requires minimal sample material, does not need expensive assay equipment, requires four simple steps, and only takes 20-30 min to perform. These include (a) removal of the adventitious Mn ions by CaCl2 treatment of the membranes, (b) extraction of the Mn from the O-2-evolving complex with hydrochloric acid, (c) purification of the extract by centrifugation followed by filtration of the supernatant through an Acrodisc syringe filter (0.2 mu m nylon membrane), and (d) colorimetric determination of Mn in the extract using the reaction of the chromogenic agent, 3,3',5,5'-tetramethylbenzidine, with previously oxidized Mn(II) cations carried out at high pH. The colorimetric assay itself has been used previously by Serrat (Mikrochim Acta 129:77-80, 1998) for assaying Mn concentrations in sea water and drinking water.