Influenza A virus subtype H1N1 can cause severe acute respiratory distress syndrome and death in young children and elderly individuals. H1N1 initiates inflammatory responses that aim to contain and eliminate microbial invaders. Various lipid mediators (LMs) are biosynthesized and play a critical role in fighting viruses during inflammation; thus, by profiling the LMs in patients, researchers can obtain mechanistic insights into diseases, such as the pathways disrupted. To date, the relationship between molecular alterations in LMs and the pathogenesis of H1N1 influenza in children is poorly understood. Here, we employed a targeted liquid chromatography coupled with tandem mass spectrometry (LC?MS/MS) to profile LMs in serum from children with H1N1 influenza (H1N1 children) and recovered children. We found that 22 LM species were altered in H1N1 children with mild symptoms. Analysis of the LM profiles of recovered children revealed a decrease in the levels of thromboxane B2 (TxB2) and thromboxane B3 (TxB3) and an increase in the levels of other 8 altered LM species associated with H1N1 influenza, including cytochrome P450 (CYP) enzyme-derived dihydroxyeicosatrienoic acids (DiHETrEs) and hydroxyeicosatetraenoic acids (HETEs) from arachidonic acid (AA), and epoxyoctadecamonoenoic acids (EpOMEs) from linoleic acid (LA). Taken together, the results of this study revealed that serum LMs change dynamically in H1N1 children with mild symptoms. The dramatically altered LMs in H1N1 children could serve as a basis for potential therapeutics or adjuvants against H1N1 influenza.