The critical period of human cerebral myelination is characterized by rapid production of cellular membranes. We hypothesize that this period is subject to the "physiological" generation of free radicals resulting in lipid peroxidation (LPO). In this study, oxidative markers were examined in developing human parietal white matter using 4-hydroxy-2-nonenal (HNE) protein adducts as an indicator of LPO. Immunocytochemistry showed an increase in HNE-positive glia from 40 gestational weeks to 1.5 postnatal years encompassing the peak period of myelin sheath synthesis at this site. Western blots showed a distinct pattern of HNE-modified proteins at fetal/term ages 26 to 42 gestational weeks and a second, different pattern at 45 gestational weeks to 2.5 postnatal years. Proteins modified by HNE in the latter period, corresponding to active myelination, were identified using mass spectrometry. The most prominent category of HNE modification included cytoskeletal proteins such as tubulins and neurofilaments. Other categories included cell type-specific proteins for mature oligodendrocytes and astrocytes and proteins involved in cell cycle and energy metabolism. We conclude that human brain development involves basal levels of oxidative stress and resulting LPO and that these processes target different proteins in an age-specific manner, thereby likely playing distinct roles during different periods of brain maturation.