Nicotiana tabacum, tobacco, is a stout herbaceous plant in the Solanaceae (nightshade family) that originated in the tropical Americas (South America, Mexico, and the West Indies) and now cultivated worldwide as the primary commercial source of tobacco, which is smoked or chewed as a drug for its mild stimulant effects [1] and [2]. N. tabacum is a kind of plant containing most complex secondary metabolites in nature. It was reported that more than 2549 kinds of chemical compositions have been identified in N. tabacum, while up to 8700 kinds of chemical compositions were found in tobacco and tobacco substitutes and cigarette smoke (including combustion products) [3]. Our previous investigation of this species led to the discovery of a number of new compounds that showed various bioactivities, such as anti-HIV-1, anti-TMV, and cytotoxicity by our groups [4], [5], [6], [7], [8] and [9]. In continuing efforts to utilize N. tabacum and identify bioactive natural products, the phytochemistry investigation of the leaves of Yunyan 201 (a variety of N. tabacum) led to the isolation of three new (1–3) and three known (4–6) sesquiterpenes, of which possessed two unprecedented skeletons (one skeleton for compounds 1 and 2 while the other for compound 3). This paper deals with the isolation, structural elucidation, and anti-TMV activity of these compounds.