Two new quinoline alkaloids, 4-acetyl-7-isopropyl-6-methylquinolin-2(1H)-one (1) and 4-acetyl-7-isopropyl-5-methylquinolin-2(1H)-one (2), were isolated from the stems of Nicotiana tabacum. Their structures were determined by means of HR-ESI-MS and extensive 1D and 2D NMR spectroscopic studies. Compounds 1 and 2 were tested for their anti-tobacco mosaic virus (anti-TMV) activity. The results showed that compounds 1 and 2 exhibited high anti-TMV activity with inhibition rates of 38.5 and 42.2% at the concentration of 20 mu M, respectively. These rates are higher than that of positive control.