Docosahexaenoic acid (DHA), an abundant fatty acid in the brain, is susceptible to auto-oxidation in situ and releases metabolites such as F4 -neuroprostane (4-F4t -NeuroP). The presence of 4-F4t -NeuroP in the brain is not well explored. In this study, 4-F4t -NeuroP was introduced into neuroblastoma cells (SH-SY5Y) and, by in vivo infusion, into rodents. Targeted lipidomic analysis of liver and brain tissues shows significant elevation of anti-inflammatory hydroxylated DHA metabolites and an isomer of neuroprotectin D1, suggesting potential beneficial bioactivities of 4-F4t -NeuroP. Additionally, 4-F4t -NeuroP treatment in SH-SY5Y cells and primary neuronal culture consistently upregulates the transcriptional level of the antioxidant enzyme heme oxygenase-1, but the effect is reduced when 4-F4t -NeuroP is further oxidized. Our data suggest that 4-F4t -NeuroP could be neuroprotective in the native state but may have disadvantageous bioactivity when oxidized extensively.