Two new sesquiterpenes from the leaves of Nicotiana tabacum and their anti-tobacco mosaic virus activities

Abstract Two new sesquiterpenes, nicotianasesterpenes A and B (1 and 2), together with five known sesquiterpenes (3–7) were isolated from the leaves of Nicotiana tabacum. Their structures were determined mainly by spectroscopic methods, including extensive 1D- and 2D-NMR techniques. The anti-tobacco mosaic virus (anti-TMV) activities of compounds 1–7 were evaluated. The results revealed that compound 1 exhibited high anti-TMV activities with inhibition rates of 33.6%. This rate is high than that of positive control. The other compounds also showed potential activities with inhibition rates in the range of 18.8–28.4%, respectively. Graphical abstract The structures of compounds 1–2.

Nicotianasesterpene A (1) was isolated as a yellow gum. Its molecular formula was determined to be C 16 h 20 O 2 based on hReSIMS with seven degrees of unsaturation. The 1 h NMR spectrum of 1 displayed two aromatic protons [δ h 6.92 (d, J = 8.2 hz), h-6 and 7.33 (d, J = 8.2 hz), h-7], two olefinic protons [δ h 6.55 (s), h-3 and 6.36 (d, J = 10.0 hz), h-10], one  Figure S5) of h 3 -12,13/ h-11/h-10 indicated that the existence of a methylpropylidene moiety (=Ch-Ch(Ch 3 ) 2 ). In addition, the 13 C and DePT NMR spectroscopic data revealed that compound 1 presence of 16 carbon signals, including one 1,2,3,4-tetrasubstituted aromatic ring (C-1 and C-5-C-9), two pair of double bonds (C-2-C-4, C-10), three methyl (C-12, C-13, and C-15), one oxidated methylene (C-14), one methane (C-11) and one methoxy group (δ C 61.0). The hMBC correlations ( Figure S5) of h-3 with C-1, C-2, C-4 and C-5, of h-6 with C-1, C-4 and C-5 indicate the existence of an indene ring (C-1-C-9) in 1. The remainder signals, a methylpropylidene moiety, an oxidated methylene group, a methyl group and a methoxy group are considered as the substitutents on indene ring. The methylpropylidene moiety located at C-2 was supported by the hMBC correlations of h-10 with C-1, C-2 and C-3, of h-11 with C-2, and of h-3 with C-10. The oxidated methylene group located C-4 was supported by the hMBC corrections of h-14 with C-3, C-4 and C-5, and of h-3 with C-14. The methyl group located C-8 was supported by the hMBC corrections of h-15 with C-7, C-8 and C-9, and of h-7 with C-15. The hMBC correlation of methoxy proton signal (δ h 3.80) with C-9 suggested the methoxy group located at C-9. In addition, the E-configuration of the double bond between C-2 and C-10 was determined by the comparison of NMR data of 1 with these of known compound, anmindenol A (Lee et al. 2014), and a clear ROeSY correlation observed from h-11 and h-3. Thus, the structure of 1 was established.
Compound 2 was also obtained as a yellow gum, and assigned the molecular formula C 16 h 20 O 3 by hReSIMS at m/z 283.1310 [M + Na] + , which indicated seven degrees of unsaturation. The 1 h NMR and 13 C NMR spectra of 2 (Table 1) display a 1,2,3,4-tetrasubstituted aromatic ring (C-1 and C-5-C-9), two pair of double bonds (C-2-C-4, C-10), two methyl (C-12, C-13), two oxidated methylene (C-14 and C-15), one methane (C-11) and one methoxy group (δ C 61.0), and these data were very similar to those of compound 1. The obvious chemical shift differences resulted from the substituent group variation on the benzene ring. The appearance of one oxidated methylene (δ C 58.3 t, δ h 4.82, 4.85 s) signals and the disappearance of one methyl signals were observed in compound 2. These changes indicated that the methyl group in 1 was replaced by a hydroxymethyl group in 2. Further analysis of its hMBC correlations can confirm the methylpropylidene moiety located at C-2, the methoxy group located at C-9 and two hydroxymethyl groups located at C-4 and C-8, respectively. The structure of 2 is therefore determined, and gives the trivial name of nicotianasesterpene B.
Compounds 1-7 were tested for their anti-TMV activities. The inhibitory activities of compounds 1-6 against TMV replication were tested using the half-leaf method (hu, ). Ningnanmycin (a commercial product for plant disease in China), with inhibition rate of 31.5%, was used as a positive control. The results showed that compound 1 exhibited high anti-TMV activities with inhibition rates of 33.6%. This rate is higher than that of positive control. The other compounds also showed potential activities with inhibition rates in the range of 18.8-28.4%, respectively (Table 1).

General
uV spectra were obtained using a Shimadzu uV-2401A spectrophotometer. eCD spectra were measured on a JASCO J-810 spectropolarimeter. A Tenor 27 spectrophotometer was used for scanning IR spectroscopy with KBr pellets. 1D and 2D NMR spectra were recorded on a DRX-500 NMR spectrometer with TMS as internal standard. Chemical shifts (

Plant material
The

Anti-TMV assay
The anti-TMV activities were tested using the half-leaf method (hu, , and ningnanmycin, a commercial product for plant disease in China, was used as a positive control.

Disclosure statement
No potential conflict of interest was reported by the authors.