Using inductively coupled plasma-mass spectrometry after samples microwave-assisted acid digestion, zinc (Zn), copper (Cu), and manganese (Mn) levels were measured in 14 different areas of the human brain of adult individuals (n = 42; 71 ± 12, range 50-101 years old) without a known history of neurodegenerative, neurological, or psychiatric disorder. The main goals of the work were to establish the "normal" (reference) values for those elements in the human brain and to evaluate the age-related changes, a prior and indispensable step in order to enlighten the role of trace element (TE) in human brain physiology and their involvement in aging and neurodegenerative processes. Considering the mean values for the 14 regions, Zn (mean ± sd; range 53 ± 5; 43-61 μg/g) was found at higher levels, followed by Cu (22 ± 5; 10-37 μg/g) and Mn (1.3 ± 0.3; 0.5-2.7 μg/g). The TE distribution across the brain tissue showed to be quite heterogeneous: the highest levels of Zn were found in the hippocampus (70 ± 10; 49-95 μg/g) and superior temporal gyrus (68 ± 10; 44-88 μg/g) and the lowest in the pons (33 ± 8; 19-51 μg/g); the highest levels of Cu and Mn were found in the putamen (36 ± 13; 21-76 μg/g and 2.5 ± 0.8; 0.7-4.5 μg/g, respectively) and the lowest in the medulla (11 ± 6; 2-30 μg/g and 0.8 ± 0.3; 0.2-1.8 μg/g, respectively). A tendency for an age-related increase in Zn and Mn levels was observed in most brain regions while Cu levels showed to be negatively correlated with age.