Decreasing nitrogen (N) supply affected the normal growth of Oryza sativa (O. sativa) seedlings, reducing CO2 assimilation, stomatal conductance (gs), the contents of chlorophylls (Chl) and the ratio of Chl a/Chl b, but increasing the intercellular CO2 concentration. Polyphasic chlorophyll a fluorescence transient and relative fluorescence parameters (JIP test) results indicated that N deficiency increased Fo, but decreased the maximum quantum yield of primary photochemistry (Fv/Fm) and the maximum of the IPphase, implying that N-limiting condition impaired the whole photo electron transport chain from the donor side of photosystem II (PSII) to the end acceptor side of PSI in O. sativa. N deficiency enhanced the activities of the antioxidant enzymes, such as ascorbate peroxidase (APX), guaiacol peroxidase (GuPX), dehydro-ascorbate reductase (DHAR), superoxide dismutase (SOD), glutathione peroxidase (GlPX), glutathione reductase (GR), glutathione S-transferase (GST) and O-acetylserine (thiol) lyase (OASTL), and the contents of antioxidant compounds including reduced glutathione (GSH), total glutathione (GSH+GSSG) and non-protein thiol compounds in O. sativa leaves. In contrast, the enhanced activities of catalase (CAT), DHAR, GR, GST and OASTL, the enhanced ASC-GSH cycle and content of sulfur-containing compounds might provide protective roles against oxidative stress in O. sativa roots under N-limiting conditions. Quantitative real-time PCR (qRT-PCR) analysis indicated that 70% of the enzymes have a consistence between the gene expression pattern and the dynamic of enzyme activity in O. sativa leaves under different N supplies, whereas only 60% of the enzymes have a consistence in O. sativa roots. Our results suggested that the antioxidant system and sulfur metabolism take part in the response of N limiting condition in O. sativa, and this response was different between leaves and roots. Future work should focus on the responsive mechanisms underlying the metabolism of sulfur-containing compounds in O. sativa under nutrient deficient especially N-limiting conditions.