A new sesquiterpene glucoside from Nicotiana rustica L.

Article abstract of DOI:10.1080/14786419.2012.725398

A new compound, rel-2R,5S,9S,10R,11R,12-trihydroxy-6(7)-spirovetiven-8-one- 9- O -β-D-glucopyranoside (1), along with a known spirovetiven glucoside (2), was isolated from the leaves of Nicotiana rustica L. The structure of compound (1) was elucidated on the basis of spectroscopic data.

Full text of DOI:10.1080/14786419.2012.725398

Natural Product Research
Formerly Natural Product Letters
ISSN: 1478-6419 (Print) 1478-6427 (Online) Journal homepage: http://www.tandfonline.com/loi/gnpl20
A new sesquiterpene glucoside from Nicotiana
rustica L.
Rong Zhou , Dong-Liang Li , Guang-Lin Feng & Guo-You Li
To cite this article: Rong Zhou , Dong-Liang Li , Guang-Lin Feng & Guo-You Li (2013) A new
sesquiterpene glucoside from Nicotiana rustica L., Natural Product Research, 27:14, 1261-1264,
DOI: 10.1080/14786419.2012.725398
To link to this article: http://dx.doi.org/10.1080/14786419.2012.725398
Published online: 14 Sep 2012.
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Date: 06 November 2015, At: 06:41

Natural Product Research, 2013
Vol. 27, No. 14, 1261–1264, http://dx.doi.org/10.1080/14786419.2012.725398
A new sesquiterpene glucoside from Nicotiana rustica L.
a
a
a
b
Rong Zhou , Dong-Liang Li , Guang-Lin Feng and Guo-You Li *
a
Technology R&D Center, China Tobacco Chuanyu Industrial Co. Ltd, Chengdu 610066, China;
Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu 610041, China
b
(
Received 16 April 2012; final version received 7 August 2012)
A new compound, rel-2R,5S,9S,10R,11R,12-trihydroxy-6(7)-spirovetiven-8-one-
9-O-�-D-glucopyranoside (1), along with a known spirovetiven glucoside (2), was
isolated from the leaves of Nicotiana rustica L. The structure of compound (1)
was elucidated on the basis of spectroscopic data.
Keywords: Nicotiana rustica; sesquiterpene; glucoside; spirovetiven
1
. Introduction
Nicotiana rustica L. is an important economic crop originating from South America
The Editorial Committee of the Administration Bureau of Flora of China, 2005). Its
(
leaves are mainly used as raw material for the tobacco industry. A few sesquiterpene
glucosides were isolated from the leaves of Nicotiana tabacum L. (Feng, Wang, Luo, &
Kong, 2009, 2010; Kodama, Fujimori, & Kato, 1981, 1982, 1984), which may play an
important role in the quality of tobacco as potential aroma precursors. In the course of
investigation on the potential aroma precursors, a new sesquiterpene glucoside, rel-
2
R,5S,9S,10R,11R,12-trihydroxy-6(7)-spirovetiven-8-one-9-O-�-D-glucopyranoside
along with a known spirovetiven glucoside (2) (Engstrom, 1998), was isolated from the
leaves of N. rustica L. The structure of compound 1 was elucidated on the basis of 1D- and
D-NMR spectra analyses (Figure 1). Here, the isolation, structure elucidation and
antimicrobial activity of compounds 1 and 2 are described.
(1),
¨
2
2
. Results and discussion
Compound 1 was obtained as colourless gum. HR–ESI–MS (high resolution electrospray
þ
ionization mass spectrum, 453.2080 [M þ Na] ) analysis of 1 suggested a molecular
formula of C H O , indicative of five degrees of unsaturation. The IR absorption bands
2
1
34
9
�
1
at 1661 and 1620 cm and C-NMR signals at ꢀ 197.6, 170.5 and 124.6 are characteristic
13
C
1
of an ꢁ,�-unsaturated ketone. The H-NMR spectrum showed two singlet methyls (ꢀ 1.52
H
1
3
and 1.88) and one doublet methyl (ꢀ 1.14, d, J ¼ 6.9 Hz). The C-NMR spectrum of 1
H
1
revealed 21 C-atoms. A moiety of D-glucose group was evident in view of the C-NMR
3
signals at ꢀ 103.5, 78.9, 78.5, 75.1, 71.8 and 62.9 (Liang et al., 2011). Besides the signals
C
1
for glucose, there were 15 C-NMR signals left for a sesquiterpene aglycone containing a
3
*
Corresponding author. Email: lidongliang163@yahoo.com.cn; ligy@cib.ac.cn
©
2013 Taylor & Francis

1
262
R. Zhou et al.
Figure 1. Compounds 1 and 2 from the leaves of N. rustica L.
Figure 2. Key HMBC and NOESY correlations of 1.
ketone group and a trisubstituted double bond. The five unsaturation degrees suggested
that compound 1 should be a bicyclic compound. The HMBC correlations of the methyl at
�H 1.88 (H-14) with the C-atoms at � 51.8 (C-5), 170.2 (C-6) and 124.6 (C-7) and the
C
proton at � 4.49 (d, J ¼ 8.0 Hz, H-9) with the C-atoms at � 51.8 (C-5), 124.6 (C-7), 197.6
H
C
(
C-8), 47.3 (C-10) and 21.7 (C-14) suggested substructure of ring A. The HMBC
correlations of H-1 (� 2.25) with C-2 (� 43.5), C-3 (� 28.1), C-4 (� 33.7), C-5 (� 51.8)
H
C
C
C
C
and C-11 (� 73.6) supported the presence of ring B. H-13 (� 1.52) correlating with the C-
C
H
atoms at � 73.6 (C-11), 70.3 (C-12) and 43.5 (C-2) suggested the presence of the
C
oxygenated isopropyl group. The ꢀ configuration for glucose was inferred by the coupling
constant value (7.8 Hz) of the anomeric proton at � 5.11 (Liang et al., 2011), which was
H
0
connected to C-9 from the HMBC correlation of H-9 with C-1 (� 103.5; Figure 2). Acid
C
�
hydrolysis of compound 1 by 2 N HCl at 80 C yield D-glucose with the optical rotation of
2
0
D
½
ꢁꢀ þ 52.5 (c 0.10, H O), which was also identified by TLC comparison with an authentic
3 3 2 f 3 3 2
2
sample (CHCl –CH OH–H O ¼ 8 : 4 : 1, R ¼ 0.22, n-BuOH–CH COCH –H O ¼ 4 : 1 : 1,
R ¼ 0.50) (Zhang, Ye, Li, Qi, & Zhang, 2005). Detailed analysis of the HSQC and HMBC
f
spectra led to the elucidation of the structure of compound 1. In the NOESY spectrum of
1
, the signals of H-9/H-15, H-15/H-1a and H-14/H-1b suggested the relative stereochem-
istry of C-5, C-9 and C-10 to be S, S and R, respectively. The relative stereochemistry of C-
was inferred from the large J-value (12.0 Hz) between H-1b and H-2. The cross peak of
2
H-12 with H-1a suggested the relative configuration of C-10 should be R. Finally, the
relative stereochemistry of compound 1 was determined to be rel-2R,5S,9S,10R,11R,12-
trihydroxy-6(7)-spirovetiven-8-one-9-O-ꢀ-D-glucopyranoside on the basis of NOESY
experiment in combination with comparing the NMR data with those of the known
compound (Feng et al., 2009; Figure 2).
Compounds 1 and 2 exhibited no inhibitory activity against Escherichia coli,
Staphylococcus aureus, Aspergillus niger or Candida albicans at a concentration
ꢁ
1
.
of 100 mg mL

Natural Product Research
1263
3
3
. Experimental
.1. General details
Optical rotations were measured on a Perkin Elmer 341 polarimeter. UV and IR spectra
were carried out on a Perkin Elmer Lambda 35 UV/VIS spectrometer and a Perkin Elmer
Spectrum One FT-IR spectrometer, respectively. NMR spectra were performed on a
Bruker Avance 600 spectrometer. ESI–MS and HR–ESI–MS were obtained on a BioTOF-
Q mass spectrometer. Silica gel (200–300 mesh) for column chromatography and silica gel
GF254 (10–40 mm) for TLC were obtained from Qingdao Haiyang Chemical Company,
China. Semi-preparative HPLC was performed on a Water 2605 liquid chromatographic
2
ꢀ1
apparatus with a Water RP-18 column (20 � 250 mm , 3 mL min ). All solvents including
ꢁ
petroleum ether (60–90 C) were distilled prior to use.
3
.2. Plant and microorganism material
The leaves of N. rustica L. were collected in Xichang region, Sichuan Province, China, in
September 2010, and were identified by Prof. Yu-Lan Peng, Chengdu Institute of Biology,
Chinese Academy of Sciences. A voucher specimen (zr2011081001) was deposited at the
Herbarium of Chengdu Institute of Biology, CAS.
The micro-organisms used were obtained from Chengdu Institute of Biology, the
Chinese Academy of Sciences. They include bacteria viz. E. coli (ATCC25923) and S.
aureus (ATCC25923) and fungi viz. C. albicans (ATCC10231) and A. niger (ATCC16404).
The bacteria and fungi were maintained on LB dextrose agar slant and potato dextrose
ꢁ
agar slant at 4 C, respectively. The diameter of the Oxford plate is 9.0 mm.
3
.3. Extraction and isolation
The air-dried powdered leaves of N. rustica L. (5.0 kg) were extracted with 95% ethanol
(
residue (800 g). The residue was suspended in H O (5 L) and partitioned successively with
40 L � 2, 3 days) at RT and filtered. The filtrate was evaporated in vacuo to afford a
2
petroleum ether, ethyl acetate and n-butanol. The n-butanol extract (400 g) was subjected
to macroporous resin D101 column, eluted with water, 20% ethanol, 40% ethanol, 60%
ethanol and ethanol. The 20% ethanol fraction (Fraction A, 20 g) was separated over silica
gel eluted gradiently with CHCl –MeOH (7 : 1 to 1 : 1) to give six subfractions AA, AB,
3
AC, AD, AE, AF and AH. Compounds 1 (5 mg) and 2 (40 mg) were obtained by
separation of AD with preparative HPLC (35% MeOH–H O).
2
Rel-2R,5S,9S,10R,11R,12-trihydroxy-6(7)-spirovetiven-8-one-9-O-�-D-glucopyranoside
2
D
0
ꢁ
(
(
6
4
4
1) Colourless oil; ½ꢀꢂ þ 5.0 (c 0.10, MeOH); UV (MeOH) ꢁ
(log "): 202 (4.22), 232
4.41) nm; IR (KBr) ꢂ : 3399, 2928, 1661, 1620, 1464, 1382, 1264, 1162, 1076, 1044,
max
max
ꢀ
1
1
0
19 cm . H-NMR (C D N, 600 MHz) ꢃ 5.80 (1H, s, H-7), 5.11 (1H, d, J ¼ 7.8 Hz, H-1 ),
5
5
0
.50 (1H, brd, J ¼ 12.1 Hz, H-9), 4.49 (1H, dd, J ¼ 11.8, 3.0 Hz, H-6 ), 4.32 (1H, dd, J ¼ 12.1,
0
0
0
.8 Hz, H-6 ), 4.23 (1H, t, J ¼ 9.0 Hz, H-3 ), 4.17 (1H, t, J ¼ 9.0 Hz, H-4 ), 4.00 (1H, t,
0
0
J ¼ 8.4 Hz, H-2 ), 3.96 (1H, m, H-5 ), 3.85 (2H, brs, H-12), 2.53 (1H, m, H-10), 2.49 (1H, m,
H-1a), 2.44 (1H, m, H-2), 2.25 (1H, t, J ¼ 12.0 Hz, H-1b), 1.93 (1H, m, H-3), 1.88 (3H, s, H-
1
4), 1.84 (1H, m, H-4), 1.80 (1H, m, H-3), 1.60 (1H, m, H-4), 1.52 (3H, s, H-13), 1.14 (3H, d,
3
1
J ¼ 6.9 Hz, H-15); C-NMR (C D N, 150 MHz) ꢃ 197.6 (C-8), 170.2 (C-6), 124.6 (C-7),
5
5
0
0
0
0
0
1
1
1
03.5 (C-1 ), 80.9 (C-9), 79.0 (C-5 ), 78.5 (C-3 ), 75.1 (C-2 ), 73.6 (C-10), 71.8 (C-4 ), 70.3 (C-
0
2), 62.9 (C-6 ), 51.8 (C-5), 46.3 (C-11), 43.5 (C-2), 38.2 (C-1), 33.7 (C-4), 28.1 (C-3), 24.6 (C-
þ
3), 21.8 (C-14) and 14.1 (C-15); ESI–MS m/z 453.2 [M þ Na] ; HR–ESI–MS m/z 453.2080
(
Calcd C H O Na [M þ Na], 453.2095).
2
1
34
9

1
264
R. Zhou et al.
3
.4. Acid hydrolysis
�
A solution of compound 1 (5 mg) was added to 5 mL of 2 N HCl at 80 C for 12 h, and then
the mixture was filtrated and extracted with chloroform (10 mL ꢀ 3). The aqueous fraction
was evaporated in vacuo to yield D-glucose, which was identified by with the optical
2
D
0
�
rotation of ½�ꢁ þ 52.5 (c 0.10, H O), which was identified by comparing with an
2
authentic sample by on TLC analysis (CHCl –CH OH–H O ¼ 8:4:1, R ¼ 0.22, n-BuOH–
3
3
2
f
CH COCH –H O ¼ 4:1:1, R ¼ 0.50).
3
3
2
f
3
.5. Antimicrobial assays
Compounds 1 and 2 were tested for in vitro activity against the fungal strains C. albicans
and A. niger, and bacterial strains E. coli and S. aureus by the cup-plate method (Shen,
Fan, & Li, 1999). The antibacterial and antifungal activities were indicated by the presence
of a clear zone around the Oxford plate where the growth of the tested bacteria and fungi
had been inhibited. The degree of inhibition was determined by measuring the diameters of
the inhibition zones in millimetres.
A suspension of each bacterium and fungus tested was prepared from the slides and
�
mixed with melting agar medium at 40–45 C. The compounds were dissolved in dimethyl
ꢂ1
sulphoxide (DMSO) at a concentration of 0.1 mg mL . The solution (0.2 mL) of each
compound was aseptically introduced into Oxford plate. DMSO was used as a negative
�
�
control. The plates of bacteria were incubated at 37 C for 24 h and those of fungi at 30 C
for 48 h. The diameters of the inhibition zones were measured. All assays were performed
in triplicate. Mean values were selected.
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