We have studied a series of normal subjects and patients with brain tumors, by using 31P three-dimensional chemical shift imaging to obtain localized 31P spectra of the brain. A significant proportion of brain cytosolic ATP in normal brain is not complexed to Mg2+, as indicated by the chemical shift delta of the beta-P resonance of ATP. The ATP beta-P resonance position in brain thus is sensitive to changes in intracellular free Mg2+ concentration and in the proportion of ATP complexed with Mg because this shift lies on the rising portion of the delta vs. Mg2+ titration curve for ATP. We have measured the ATP beta-P shift and compared intracellular free Mg2+ concentration and fractions of free ATP for normal individuals (n = 6) and a limited series of patients with brain tumors (n = 5). In four of the five spectra obtained from brain tissue containing a substantial proportion of tumor, intracellular free Mg2+ was increased, and the fraction of free ATP was decreased, compared with normal brain.