Differentiation of absolute metabolite concentrations between gray and white matter in the occipital region of normal human brain was performed by in vivo localized single-voxel 1H magnetic resonance spectroscopy at 1.5 Tesla with long echo time (136 ms). With the combination of image segmentation between white and gray matter and cerebrospinal fluid, signal compensation of T1 and T2 effects, tissue water signal as the internal concentration reference, as well as compensation by different water contents in gray and white matters, it was determined that the levels of N-acetylaspartate (NAA), creatine and/or phosphocreatine (Cr), and choline-containing compounds (Cho) in gray matter were significantly higher than in white matter. The averaged NAA, Cr, and Cho concentrations in gray matter were 11.0, 9.7, and 1.9 mM/liter, respectively, in comparison with 7.5, 5.2, and 1.6 mM/liter in white matter. These results suggest that precise composition of white and gray matter and cerebrospinal fluid is necessary to avoid partial voluming effect in a single voxel and to accurately quantify the metabolite concentrations.