Podolactones are a class of structural diverse diterpenoid lactones, mainly isolated from the Podocarpus species. Several bioactivities have been disclosed for podolactones, including cytotoxicity and anti-atherosclerosis. In this study, the seeds of P. nagi were isolated by comprehensive chromatographic methods to obtain three new podolatones, named nagilactone B 1-O-beta-d-glucoside (1), nagilactone N3 3-O-beta-d-glucoside (2), and 2-epinagilactone B (3), as well as a known compound, nagilactone B (4). Their structures were determined by analyses of NMR and HRESIMS data. Compounds 1 and 2 significantly inhibited nitric oxide (NO) production on LPS-stimulated RAW264.7 macrophages, with IC50 values of 0.18 +/- 0.04 and 0.53 +/- 0.03 mu M, respectively. Indomethacin (IC50 4.21 +/- 0.32 mu M) was used as a positive control. Compound 1 suppressed the expression of inducible NO synthase (iNOS) in a concentration-dependent manner, mediating through inhibiting nuclear factor-kappa B (NF-kappa B) activity. This is the first report regarding the anti-inflammatory effect of podolactones, which could be potential anti-inflammatory agents.