DETERMINATION OF IN-VIVO CYTOPLASMIC ORTHOPHOSPHATE CONCENTRATION IN YEAST

To investigate in-vivo concentrations of cytoplasmic phosphate, especi ally during dynamic conditions, a method has been developed that enabl es reproducible determination of cytoplasmic phosphate from 5 mu M up to 30 mu M. The method involves fast sampling. spontaneous inactivatio n of cell metabolism and differential extraction procedure to gain por osity of the outer cell membrane exclusively. To determine very low cy toplasmic phosphate concentrations, an enzymatic assay was linked to a sensitive spectrophotometric cycling method to increase the detection limit.