Water-soluble metabolites extracted from 20 astrocytic tumors (11 glioblastomas, 3 anaplastic astrocytomas, and 6 low-grade astrocytomas) and four normal brains were measured qualitatively and quantitatively using in vitro high-resolution proton magnetic resonance (1H-MR) spectroscopy. MR spectra from tumors exhibited characteristic patterns according to malignancy, presumably reflecting the metabolism of gliomas. Concentrations of choline-containing compounds, inositol, alanine, and glycine increased according to the malignancy, while that of total creatine decreased. In particular, glycine concentration was very high in glioblastoma, and an immunohistochemical study using anti-glycine antibody demonstrated that glycine was mainly distributed in glioma cells, not in proliferative endothelial cells. The ratios of choline-containing compounds and glycine to total creatine are useful parameters for grading gliomas, and the ratio of glycine to total creatine is useful for the differential diagnosis of glioblastoma from metastatic tumor. Such indications appearing in in vivo 1H-MR spectroscopy might provide clinically useful information on tumor metabolism and malignancy, and help assess the effects of radiation therapy and chemotherapy on gliomas.