Nymphaea mexicana Zucc. is an aquatic plant species which belongs to the family Nymphaea and is commonly known as the yellow water lily. The aim of this work was to study the in vitro anti-inflammatory effects of phenolic compounds isolated from the flowers of Nymphaea mexicana Zucc. Seven phenolic compounds including vanillic acid (1), 4-methoxy-3,5-dihydroxybenzoic acid (2), (2R,3R)-3,7-dihydroxyflavanone (3), naringenin (4), kaempferol 3-O-(3-O-acetyl-α-L-rhamnopyranoside) (5), kaempferol 3-O-(2-O-acetyl-α-L-rhamnopyranoside) (6), and quercetin 3-(3′′-acetylrhamnoside) (7) were isolated from the flowers of Nymphaea mexicana Zucc. These results revealed that compound 4 has the most prominent inhibitory effect on the LPS-stimulated nitric oxide (NO), monocyte chemotactic protein-1 (MCP-1), and tumor necrosis factor-alpha (TNF-α) production in RAW 264.7 macrophages. In addition, compound 4 also inhibited LPS-mediated induction of protein expressions of inducible nitric oxide synthase (iNOS), cyclooxygenase 2 (COX-2), and phospho-ERK in RAW 264.7 macrophages. Thus, compound 4 from the flowers of Nymphaea mexicana Zucc. may provide a potential therapeutic approach for inflammation-associated disorders.