Phosphorus and carbon metabolism in Microlunatus phosphovorus was investigated by using a batch reactor to study the kinetics of uptake and release of extracellular compounds, in combination with (31)P and (13)C nuclear magnetic resonance (NMR) to characterize intracellular pools and to trace the fate of carbon substrates through the anaerobic and aerobic cycles. The organism was subjected to repetitive anaerobic and aerobic cycles to induce phosphorus release and uptake in a sequential batch reactor; an ultrafiltration membrane module was required since cell suspensions did not sediment. M. phosphovorus fermented glucose to acetate via an Embden-Meyerhof pathway but was unable to grow under anaerobic conditions. A remarkable time shift was observed between the uptake of glucose and excretion of acetate, resulting in an intracellular accumulation of acetate. The acetate produced was oxidized in the subsequent aerobic stage. Very high phosphorus release and uptake rates were measured, 3.34 mmol g of cell(-1) h(-1) and 1.56 mmol g of cell(-1) h(-1), respectively, values only comparable with those determined in activated sludge. In the aerobic period, growth was strictly dependent on the availability of external phosphate. Natural abundance (13)C NMR showed the presence of reserves of glutamate and trehalose in cell suspensions. Unexpectedly, [1-(13)C]glucose was not significantly channeled to the synthesis of internal reserves in the anaerobic phase, and acetate was not during the aerobic stage, although the glutamate pool became labeled via the exchange with intermediates of the tricarboxylic acid cycle at the level of glutamate dehydrogenase. The intracellular pool of glutamate increased under anaerobic conditions and decreased during the aerobic period. The contribution of M. phosphovorus for phosphorus removal in wastewater treatment plants is discussed on the basis of the metabolic features disclosed by this study.