Phyllanthus urinaria Linnea (Euphorbiaceae), is a traditional anti-hepatitis herb used in Taiwan. In continuation of our search for potent natural anti-inflammatory agents, from the ethanolic extract of this plant, nine compounds including phyllanthin (1), phyltetralin (2), trimethyl-3,4-dehydrochebulate (3), methylgallate (4), and rhamnocitrin (5), methyl brevifolincarboxylate (6), beta-sitosterol-3-O-beta-d-glucopyranoside (7), quercitrin (8), and rutin (9) were isolated. The structures of compounds 3 and 6 were established based on NMR and mass spectral studies. The isolates 1-9 were investigated for their antioxidant, and anti-inflammatory activities in vitro. In the antioxidant assay, the isolates 3, 4 and 6 exhibited significant DPPH radical scavenging activity with an IC(50) value of 9.4, 9.8 and 8.9 microM, respectively. On the other hand, in the inflammatory mediators growth inhibitory assay from LPS/interferon (IFN)-gamma-activated peritoneal macrophages, all the isolates except 7, significantly and dose-dependently inhibited the enhanced production of NO radicals, and such modulation was closely associated with the inhibition of tumor necrosis factor (TNF)-alpha and interleukin (IL)-6. In addition, 30 microM of isolates 3 and 6, and 50 microM of 4, significantly arrest the mitogen-stimulated spleen cells in G0/G1 stage. This is the first report on Phyllanthus urinaria isolates for their growth inhibitory activities against inflammatory mediators, in addition to spleen cell cycle arrest in G0/G1 stage. Therefore, these isolates from Phyllanthus urinaria may be useful for the treatment of cell-mediated immune diseases.