Abstract: A series of 4(3H)-quinazolinone derivatives containing a Schiff base moiety were synthesized and their antiviral activity against tobacco mosaic virus (TMV) was evaluated using halfleaf method in vivo. Bioactivity results exhibited that the target compounds 3a-3x under 500 μg/mL displayed moderate to good anti-TMV activities with the curative activity, protection activity, and inactivation activity values of 40.4%-53.1%, 51.2%-72.6%, and 46.0%-90.7%, respectively. Especially, compound 3d showed excellent protection activity against TMV, with a half-maximal effective concentration (EC50) value of 203.89 μg/mL, which was even superior to that of ningnanmycin (214.90 μg/mL). SAR analysis showed that compounds with electron-withdrawing groups at R1 and R2 positions displayed better anti-TMV activity. These results indicated that the series of 4(3H)-quinazolinone derivatives containing a Schiff base moiety can effectively control TMV.