31P NMR studies on microorganisms have been carried out with the cells embedded in agarose gel. The novel use of the gel for the NMR studies has advantages over the usual liquid suspensions in terms of improved reproducibility of data and cell viability, with no net loss of spectral quality. Polyphosphate formation in Escherichia coli was monitored continuously for up to 24 h and metabolic changes in yeast for 6 h. Changes of the intracellular pH during glycolysis in yeast were determined from the chemical shift of the internal Pi. NMR titration curves of Pi in the presence of Mg2+ indicate uncertainties in internal pH values estimated by this technique.