Proton MR spectroscopy ((1)H MRS)-visible total choline-containing compounds (tCho-compounds) are derivatives of membrane phospholipids and, in part, may act as a long-term second-messenger system for cellular proliferation. Experimental evidence suggests increasing concentrations of tCho-compounds during cellular proliferation. The present study was conducted in order to test the hypothesis that in vivo measurements of tCho-concentrations using (1)H MRS allow assessment of the proliferative activity of neuroepithelial brain tumors presurgically. Single-voxel (1)H MRS (PRESS, TR 1500 ms, TE 135 ms) was performed in 101 patients with neuroepithelial brain tumors prior to surgery and 19 healthy volunteers. Histological diagnoses were confirmed postsurgically according to the WHO classification. Measured tCho-compound signal intensities were corrected for coil loading, numbers of acquisitions and voxel size, and tCho concentrations calculated as institutional arbitrary units. They were matched with the mean immunohistochemical marker of cell proliferation, the Ki-67 (MIB.1) labeling index, using correlation analysis according to Spearman. Compared with low-grade tumors (i.e. WHO grade I/II) and normal white brain matter, high-grade tumors (i.e. WHO grade III/IV) revealed significantly (p < 0.05) elevated labeling indices paralleled by increasingly elevated tCho-concentrations. In contrast tCho-concentrations in low-grade tumor did not differ significantly from physiological values. A highly significant positive correlation (p < 0.0001, r(2) = 0.81) was found between the tCho-concentration and the labeling index. It was concluded that the determination of tCho-concentrations using in vivo (1)H MRS could provide a novel and noninvasive assessment of the proliferative activity of neuroepithelial brain tumors, pointing at (1)H MRS as a useful method for differentiating proliferating from non-proliferating tissues. Hence, potential indications for the clinical application of (1)H MRS are grading tumors presurgically, early detection of anaplastic transformation, and monitoring treatment.