New triterpenoid sapoin from Ardisia gigantifolia Stapf.

Article abstract of DOI:10.1016/j.cclet.2009.12.029

One new triterpenoid sapoin with two known triterpenoid sapoins: 3β-O-{α-l-rhamnopyranosyl-(1 → 3)-[β-d-xylopyranose-(1 → 2)]-β-d-glucopyranosyl-(1 → 4)-α-l-arabinopyranosyl}-3β-hydroxy-13β,28-epoxy-oleanan-16-oxo-30-al (1), 3β-O-{α-l-rhamnopyranosyl-(1 →

Full text of DOI:10.1016/j.cclet.2009.12.029

Available online at www.sciencedirect.com
Chinese Chemical Letters 21 (2010) 449–452
www.elsevier.com/locate/cclet
New triterpenoid sapoin from Ardisia gigantifolia Stapf.
b
Qiang Qiang Gong a,b, Li Hua Mu a,1, Ping Liu a,1, , Shi Lin Yang ,
*
Bo Wang a, Yu Lin Feng b
a Department of Clinical Pharmacology, General Hospital of PLA, Beijing 100853, China
b JiangXi University of Traditional Chinese Medicine, Nanchang 330006, China
Received 17 August 2009
Abstract
One new triterpenoid sapoin with two known triterpenoid sapoins: 3b-O-{a-L-rhamnopyranosyl-(1 ! 3)-[b-D-xylopyranose-
(1 ! 2)]-b-D-glucopyranosyl-(1 ! 4)-a-L-arabinopyranosyl}-3b-hydroxy-13b,28-epoxy-oleanan-16-oxo-30-al (1), 3b-O-{a-L-
rhamnopyranosyl-(1 ! 3)-[(b-D-xylopyranosyl-(1 ! 2)]-b-D-galactopyranosyl-(1 ! 4)-[(b-D-glucopyranosyl-(1 ! 2)]-a-L-ara-
binopyranoside}-16a-hydroxy-13,28-epoxy-oleanane (2) and cyclamiretin A 3b-O-a-L-rhamnopyranosyl-(1 ! 3)-[b-D-xylopyr-
anosyl-(1 ! 2)]-b-D-glucopyranosyl-(1 ! 4)-[b-D-glucopyranosyl-(1 ! 2)]-a-L-arabinopyranoside (3) were isolated from
Ardisia gigantifolia Stapf. Their structures were elucidated by means of 1H and 13C NMR spectroscopic studies, including
2D-NMR technique.
# 2009 Ping Liu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Keywords: Ardisia gigantifolia Stapf.; Myrsinaceae; Triterpenoid sapoin
The dried rhizome of Ardisia gigantifolia Stapf. is mainly used as Chinese folk medicine in south of China for the
treatment of rheumatism, pain of muscles and bones and traumatic injury. It distributes in the provinces of Guangxi,
Jiangxi and Fujian in China, which is a kind of evergreen dwarf shrub [1]. Previous chemical studies showed that
triterpenoid saponins were the main components from this genus. Thirty-two triterpenoid saponins have been isolated
from the rhizomes of A. gigantifolia Stapf. over the last 20 years [2]. In this paper, the isolation and characterization of
one new triterpenoid sapoin and two known triterpenoid sapoins from 60% ethanol percolation of the dried rhizome of
the plant were reported.
1. Experimental
The dry rhizome of A. gigantifolia Stapf. was obtained from Guangdong, China in 2007 and was authenticated by
Prof. Ping Liu. The voucher specimen has been deposited in traditional Chinese medicine pharmacy of General
Hospital of PLA.
* Corresponding author.
E-mail address: liuping126@126.com (P. Liu).
1
Both these authors contributed equally to this work.
1001-8417/$ – see front matter # 2009 Ping Liu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
doi:10.1016/j.cclet.2009.12.029

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Q.Q. Gong et al. / Chinese Chemical Letters 21 (2010) 449–452
The dried and cut rhizome parts of A. gigantifolia (13.5 kg) was percolated with 60% ethanol. The 60% ethanol
extract (1.54 kg) was partitioned successively between water and petroleum, ethyl acetate, n-butanol, respectively.
After removing the solvent, the n-butanol layer (600 g) was separated with macroporous resin D101 eluted with EtOH/
H2O (0:100, 30:70, 50:50, 70:30, and 95:5, v/v) gradient to afford five fractions. Fraction 4 (50 g) was subjected to
column chromatography on silica gel several times, which gave rise to 2 (0.7450 g) and 3 (10.0 mg). At last,
preparative HPLC [MeOH/H2O (70:30, v/v), flow rate 15.2 mL/min, 200 nm] to give 1 (0.3950 g).
Compound 1 (10 mg) was heated in an ampule with 7 mL of 2 mol/L HCL at 90 8C for 2 h. The aglycone was
extracted with EtOAC and H2O. The H2O layer was evaporated under reduced pressure. Pyridine (1.5 mL) and 3 mg of
NH2OHꢀHCL were added to residue, and the mixtures were agitated at room temperature for 24 h. Then, Ac2O (2 mL)
was added and mixtures were agitated at room temperature for 24 h. The reaction mixtures were evaporated under
reduced pressure, and the resulting aldononitrile peracetates were analyzed by GC-MS using standard aldononitrile
peracetates as reference samples.
The GC–MS experiment was carried out on a Shimadzu GC–MS chromatograph using a DB-17 MS capillary
column (30 m ꢁ 0.25 mm i.d., 0.25 mm film), ion source temperature: 250 8C, interface temperature: 300 8C, and split
ratio: 10.0. Helium (1.16 mL/min) was used as carrier gas. The initial column oven temperature was 100 8C for 1 min;
then the temperature was increased by 20 8C/min to a value of 200 8C for 1 min; then the temperature was increased by
8 8C/min to a final value of 280 8C. Compound 1 gave L-rhamaose, L-arabinose, D-xylose and D-glucose at tR 8.693,
8.983, 8.713 and 11.737 min.
2. Results and discussion
The dried and cut rhizome parts of A. gigantifolia were percolated with 60% ethanol and the ethanol extract was
partitioned between water and petroleum, ethyl acetate and n-butanol, respectively. Chromatography of the n-butanol
extract on silica gel, macroporous resin D101 and Sephadex LH-20, followed by repeated HPLC purification over
ODS, afforded compounds 1–3. Compound 1, obtained as a white powder, showed positive Libermann–Burchard and
Molish reactions, suggesting that 1 might be a triterpenoid sapoin or steroidal sapoin. HR-ESI-MS of 1 showed the
quasi-molecular ion at m/z 1065.5246 [M+Na]+ and 1041.5270 [MꢂH]ꢂ, establishing the molecular formula of
Fig. 1. Structures of compounds 1–3 and key HMBC correlations of compound 1.

Q.Q. Gong et al. / Chinese Chemical Letters 21 (2010) 449–452
451
C52H82O21. The 13C NMR spectral data of the sapogenin part of 1 were similar to those of 3, which is the known
oleanane type triterpene cyclamiretin A. In 3, the 13b,28-epoxy bridge and 16a-OH is evident from the 13C NMR
resonances at d 86.0 (C-13, C by DEPT), 77.3 (C-28, CH2) and 76.5 (C-16, C-H), respectively. However, in compound
1, no resonance was observed for C-16 at d 76.5; instead, a signal was seen at d 212.6 (C by DEPT), indicating that the –
OH at C-16 of compound 3 is oxidized to a carbonyl group. This assignment was confirmed low-field shifts at C-17
(+11.7), C-15 (+9.2) and C-14 (+5.9), and the high-field shifts at C-28 (ꢂ3.0). Furthermore, long-range coupling of H-
28 with C-16 in HMBC also supported the same conclusion. Further, in the 1H NMR spectrum, one carbinylic proton
signal assignable to H-3 of the aglycon were observed at d 3.25(dd, J = 12.0, 4.0 Hz), suggesting the carbinylic protons
could be placed at 3a [3]. Based on these findings, the aglycone was identified as 3b-hydrox-13b,28-epoxy-oleanan-
16-oxo-30-al (cyclamigenin B).
The monosaccharides obtained after acid hydrolysis of 1 were derivatized into aldononitrile peracetatederivative
and analyzed by GC–MS using authentic samples as References L-rhamaose, L-arabinose, D-xylose, and D-glucose in
the relative proportions of 1:1:1:1 were detected. The 1H NMR spectrum showed four anomeric proton signals at d 4.90
(d, 1H, J = 7.8 Hz, Glc-H-1), 4.91 (d, 1H, J = 8.0 Hz, Xyl-H-1), 4.66 (d, 1H, J = 5.2 Hz, Ara-H-1) and 6.00 (brs, 1H,
Rha-H-1). All proton signals due to sugars were assigned by careful analysis of the TOCSY, and HMQC spectra, and
the carbon signals were assigned by HMQC and DEPT spectra. The b-anomeric configurations for the glucopyranose
and xylopyranose units were determined from their 3JH-1, H-2 coupling constants (6.9–7.8 Hz). The small H-1 coupling
constant of arabinose, which indicated the arabinose should have an a-configuration at its anomeric carbon. The broad
singlet of the anomeric proton of the Rha unit indicated an a-orientation [4]. The sugar signals of 1 were similar with
known 3 [5], 1 was one glucopyranosyl unit less than 3. The arabinose was connected to C-3 of the aglycon, which was
Table 1
The NMR spectral data of compound 1 (400 MHz, pyridine-d5).a,b
.
No.
dC (DEPT)
dH (JHZ
)
No.
dC (DEPT)
dH (JHZ)
1
2
39.1 (CH2)
26.7 (CH2)
88.6 (CH)
39.7 (C)
0.78, 1.62
Arabinose (A)
A-1
1.84, 2.08
107.4 (CH)
73.8 (CH)
74.3 (CH)
81.5 (CH)
65.9 (CH2)
4.66 d (5.2)
3
3.25 dd (4.0, 12.0)
A-2
4.30
4
–
A-3
4.01
5
55.5 (CH)
17.7 (CH2)
33.8 (CH2)
43.0 (C)
0.69 d (9.2)
A-4
4.14
6
1.42
A-5
3.72 d (11.8), 4.64 dd (3.2, 11.8)
7
0.94
Glucose (G)
G-1
8
–
105.2 (CH)
81.8 (CH)
86.0 (CH)
70.6 (CH)
78.0 (CH)
62.1 (CH2)
4.90 d (7.8)
4.08
9
50.0 (CH)
36.7 (C)
1.06
G-2
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
–
G-3
4.12
18.8 (CH2)
31.5 (CH2)
86.1(C)
1.37, 1.75
G-4
4.13
2.05
G-5
3.69
–
G-6
4.32, 4.42
50.1(C)
–
Xylose(X)
X-1
45.7 (CH2)
212.6 (C)
55.3 (C)
1.27, 1.95
106.0 (CH)
75.3 (CH)
78.2 CH)
4.91 d (8.0)
–
X-2
4.03
–
X-3
3.99
55.7 (CH)
33.8 (CH2)
47.9 (C)
1.88
X-4
69.6 (CH)
66.8 (CH2)
4.10
1.36, 2.77
X-5
3.45 t-like (10.4) 4.20 m
–
Rhamnose(R)
R-1
29.6 (CH2)
33.7 (CH2)
28.0 (Me)
16.7 (Me)
16.1 (Me)
18.8 (Me)
21.8 (Me)
74.3 (CH2)
23.8 (Me)
206.1 (CH)
1.95
103.7 (CH)
72.3 (CH)
72.6 (CH)
73.7 (CH)
70.5 (CH)
18.5 (Me)
6.00 br s
5.04 br s
4.52 dd (9.0, 3.0)
4.32
0.97
R-2
1.26 s
R-3
1.00 s
R-4
0.82 s
R-5
4.85
1.28 s
R-6
1.65 d (6.4)
1.10 s
3.35 d (8.0), 3.87(8.0)
0.89 s
9.63 s
a
Assignments based on TOCSY, HMQC and HMBC experiments.
Overlapped signals are reported without designating multiplicity.
b

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Q.Q. Gong et al. / Chinese Chemical Letters 21 (2010) 449–452
deduced from the HMBC correlation between dH 4.66 (d, 1H, 5.2 Hz, Ara-H-1) and dC 88.6 (C-3). The sequence of the
sugar chain at C-3 was further determined by analysis of the HMBC and HMQC NMR spectra. Thus, HMBC
correlations were observed between dH 4.90 (Glc-H-1) and dC 81.5 (Ara-C-4), dH 4.91 (Xyl-H-1) and dC 81.8 (Glc-C-
2), dH 6.00 (Rha-H-1) and dC 86.0 (Glc-C-3). On the basis of the above results, the structure of compound 1 was
concluded to be 3b-O-{a-L-rhamnopyranosyl-(1 ! 3)-[b-D-xylopyranose-(1 ! 2)]-b-D-glucopyranosyl-(1 ! 4)-a-
L-arabinopyranosyl}-3b-hydroxy-13b,28-epoxy-oleanan-16-oxo-30-al (Fig. 1 and Table 1).
The structures of the known compounds were identified by spectral data comparison with 3b-O-{a-L-
rhamnopyranosyl-(1 ! 3)-[(b-D-xylopyranosyl-(1 ! 2)]-b-D-galactopyranosyl-(1 ! 4)-[(b-D-glucopyranosyl-
(1 ! 2)]-a-L-arabinopyranoside}-16a-hydroxy-13,28-epoxy-oleanane (2) [6] and cyclamiretin A 3b-O-a-L-
rhamnopyranosyl-(1 ! 3)-[b-D-xylopyranosyl-(1 ! 2)]-b-D-glucopyranosyl-(1 ! 4)-[b-D-glucopyranosyl-
(1 ! 2)]-a-L-arabinopyranoside (3) [5].
Acknowledgments
The authors are grateful to Prof. Ping Liu, LiHua Mu and Yuan Hu. This project was supported by National Natural
Science Foundation of China (No. 30701079).
References
[1] Jiangsu New Medicinal College, Dictionary of Chinese Drugs, Shanghai Scientific and Technological Press, Shanghai, 2001, p. 1097.
[2] D.L. Liu, X.M. Zhang, N.L. Wang, et al. Shenyang Pharm. Univ. 21 (2004) 394.
[3] R.O. Dorchai, B.J. Thomson, Tetrahedron 24 (3) (1968) 1377.
[4] Y. Mimaki, A. Yokosuka, M. Hamanaka, et al. J. Nat. Prod. 67 (2004) 1511.
[5] P. Wen, X.M. Zhang, Z. Yang, et al. J. Asian Nat. Prod. Res. 10 (9) (2008) 873.
[6] P. Wen, X.M. Zhang, Z. Yang, et al. Chinese Patent Application for Invention. CN 193187.