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On the basis of our previous studies on the antiviral mechanism against tobacco mosaic virus (TMV) and structure-activity relationship of phenanthroindolizidine alkaloids, a series of 9-substituted tylophorine derivatives targeting TMV RNA were designed, synthesized, and assessed for their anti-TMV activities. The bioassay results indicated that most of these compounds showed good in vivo anti-TMV activities, and some of them displayed higher activity than that of commercial ribavirin. Especially, the anti-TMV activities of compound 3b,